Multidrug resistant

Scand J Infect Dis Suppl, 2003 Dec, 35 Suppl 106, 45 - 8
Cellular issues relating to the resistance of HIV to antiretroviral agents; Turriziani O et al.; It has been proposed that the declining efficiency of antiretroviral agents in human immunodeficiency virus (HIV) infection may also depend on cellular factors at their site of action . Two in particular have been proposed: (i) the defective intracellular metabolism of NRTI in target cells and the altered uptake; and (ii) efflux of nucleoside reverse transcriptase inhibitors (NRTI) and protease inhibitors (PI) by cellular transporter molecules . Several studies have shown that: changes in the activities of various purine and pyrimidine biosynthetic enzymes may occur in lymphocytes of HIV-infected patients; HIV-infected patients on prolonged treatment with nucleoside analogues, e.g . zidovudine, show significantly decreased activity of thymidine kinase (TK) compared with untreated HIV-infected people; and NRTI and PI are substrates for the multidrug membrane transporters . With regard to the latter issue, it is known that the ATP-binding cassette transporter proteins such as the P-glycoprotein (MDR), and the newly discovered family of multidrug resistance-associated proteins (MRP1-6), promote the active extracellular efflux of a wide variety of therapeutics drugs and overexpression of some of them lowers intracellular concentration of PI . In the very near future such mechanisms, also called 'cellular drug resistance', might be taken into account, together with other immunological, virological and behavioural factors, to explain the 'drug failure' and/or the variability of response in HIV patients undergoing antiretroviral treatment.

Scand J Infect Dis Suppl, 2003 Dec, 35 Suppl 106, 17 - 20
Epidemiological aspects of transmitted HIV drug resistance; Girardi E; Transmitted human immunodeficiency virus (HIV) resistance to antiretrovirals (i.e . resistance in antiretroviral naive patients) emerged during the 1990s as a potentially relevant public health problem . HIV variants resistant to all classes of approved antiretroviral agents have been identified in significant proportions antiretroviral naive patients, and this phenomenon appears as a potential threat to the effectiveness of highly active antiretroviral therapy . Available data from surveys conducted between 1996 and 2001 show the prevalence of drug resistance among newly HIV-infected individuals to range from 3%, to above 20% in North America, and from 5% to 15% in Europe . Increases in prevalence observed during the late 1990s in some studies are not confirmed by most recent data . Transmission of multidrug resistance still appears to be an uncommon occurrence . However, methodological heterogeneity and problems in study design make it difficult to compare results between different surveys and to draw firm conclusions from the results . There is a clear need to improve surveillance systems aimed at identifying patients at the time of primary infection and to standardize laboratory methods for the identification of genetic markers of resistance to be used for epidemiological purposes.

Expert Rev Mol Diagn, 2004 Mar, 4(2), 209 - 17
Progress in defining multidrug resistance in leukemia; Kappelmayer J et al.; Multidrug resistance (MDR) is a naturally occurring defense phenomenon by which cells battle against chemically foreign substances (xenobiotics), including some cytotoxic drugs . Membrane transporter hyperactivity is a major contributor to MDR and is the primary target of both diagnostic and therapeutic interventions . Multi-xenobiotic resistance can be exploited as several fluorescent indicator probes are extruded by the same drug transporters, making it possible to quantitatively measure MDR activity in cell lines and clinical samples by flow cytometry . The literature on MDR is reported in a number of different formats, making it difficult to compare data from various groups . This article will briefly review the pathomechanism, then focus upon the diagnostic approach, the interpretation of results from clinical samples and correlations with other variables . The authors believe that a standardized MDR assay, as well as a suitable monitoring test, may become a prognostic marker in several types of leukemia . Copyright Future Drugs Ltd.

J Pharm Sci, 2004 Apr, 93(4), 932 - 42
Drug efflux transport properties of 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester (BCECF-AM) and its fluorescent free acid, BCECF; Bachmeier CJ et al.; 2',7'-Bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) is a fluorescent probe used to examine multidrug resistance-associated protein (MRP) transporter activity in cells . BCECF is introduced into the cell as the nonfluorescent membrane permeable acetoxymethyl ester, BCECF-AM, where it is hydrolyzed to the membrane impermeable BCECF . The lipophilic nature of BCECF-AM suggests it may be a substrate for other drug efflux transporters such as P-glycoprotein (P-gp) and the breast cancer resistance protein (BCRP) . To assess the drug efflux transporter interactions of BCECF-AM and BCECF, accumulation studies were examined in various drug efflux-expressing cells . Inhibition of P-gp, BCRP, and/or MRP produced distinct changes in the time-dependent accumulation of BCECF in the cells . Treatment with GF120918 produced an immediate and sustained effect throughout the entire time course examined . Fumitremorgin C only affected BCECF accumulation at the early time points, whereas the impact of indomethacin on BCECF accumulation was observed only at the latter time points . Permeability studies in bovine brain microvessel endothelial cells indicated an increased basolateral-to-apical transport of BCECF, which could be reduced in the presence of either indomethacin or GF120918 . These results indicate that the intracellular accumulation and transcellular permeability of BCECF are sensitive to a variety of drug efflux interactions . These results likely reflect an interaction of the ester form with P-gp and BCRP during the initial accumulation process, and an interaction of the free acid form with MRP after hydrolysis in the cell .

Hepatology, 2004 Mar, 39(3), 779 - 91
BSEP and MDR3 haplotype structure in healthy Caucasians, primary biliary cirrhosis and primary sclerosing cholangitis; Pauli-Magnus C et al.; Primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC) are characterized by a cholestatic pattern of liver damage, also observed in hereditary or acquired dysfunction of the canalicular membrane transporters bile salt export pump (BSEP, ABCB11) and multidrug resistance protein type 3 (MDR3, ABCB4) . Controversy exists whether a genetically determined dysfunction of BSEP and MDR3 plays a pathogenic role in PBC and PSC . Therefore, 149 healthy Caucasian control individuals (control group) were compared to 76 PBC and 46 PSC patients with respect to genetic variations in BSEP and MDR3 . Sequencing spanned approximately 10,000 bp including promoter and coding regions as well as 50-350 bp of flanking intronic regions . In all, 46 and 45 variants were identified in BSEP and MDR3, respectively . No differences between the groups were detected either in the total number of variants (BSEP: control group: 37, PBC: 37, PSC: 31; and MDR3: control group: 35; PBC: 32, PSC: 30), or in the allele frequency of the common variable sites . Furthermore, there were no significant differences in haplotype distribution and linkage disequilibrium . In conclusion, this study provides an analysis of BSEP and MDR3 variant segregation and haplotype structure in a Caucasian population . Although an impact of rare variants on BSEP and MDR3 function cannot be ruled out, our data do not support a strong role of BSEP and MDR3 genetic variations in the pathogenesis of PBC and PSC.

Clin Infect Dis, 2004 Mar 15, 38(6), 851 - 6 Epub 2004 Mar 01.
Multidrug-resistant tuberculous meningitis in KwaZulu-Natal, South Africa; Patel VB et al.; Multidrug-resistant (MDR) pulmonary tuberculosis (TB) is well described in the literature . Reports of MDR TB meningitis (MDR-TBM), however, are limited to case reports and a single case series . During the period of 1999-2002, 350 patients with TBM were identified by cerebrospinal fluid culture for TB . Thirty patients (8.6%) had TB that was resistant to at least isoniazid and rifampicin . All 30 patients were included in this study . We reviewed hospital charts of the patients with MDR-TBM and describe our experience . Seventeen patients with MDR-TBM died, and, of those who were known to be alive, many experienced significant morbidity . Eighteen patients were HIV positive . Twenty-two patients had been treated for TB in the past, 3 patients had received no previous treatment for TB, and the history of TB treatment was unknown for 5 patients . The study highlights the prevalence of MDR-TBM and identifies new challenges in the management of affected patients.

J Med Chem, 2004 Mar 11, 47(6), 1413 - 22
Synthesis and evaluation of dihydropyrroloquinolines that selectively antagonize P-glycoprotein; Lee BD et al.; In a search for improved multiple drug resistance (MDR) modulators, we identified a novel series of substituted pyrroloquinolines that selectively inhibits the function of P-glycoprotein (Pgp) without modulating multidrug resistance-related protein 1 (MRP1) . These compounds were evaluated for their toxicity toward drug-sensitive tumor cells (i.e . MCF-7, T24) and for their ability to antagonize Pgp-mediated drug-resistant cells (i.e . NCI/ADR) and MRP1-mediated resistant cells (i.e . MCF-7/VP) . Cytotoxicity and drug accumulation assays demonstrated that the dihydropyrroloquinolines inhibit Pgp to varying degrees, without any significant inhibition of MRP1 . The compound termed PGP-4008 was the most effective at inhibiting Pgp in vitro and was further evaluated in vivo . PGP-4008 inhibited tumor growth in a murine syngeneic Pgp-mediated MDR solid tumor model when given in combination with doxorubicin . PGP-4008 was rapidly absorbed after intraperitoneal administration, with its plasma concentrations exceeding the in vitro effective dose for more than 2 h . PGP-4008 did not alter the plasma distribution of concomitantly administered anticancer drugs and did not cause systemic toxicity as was observed for cyclosporin A . Because of their enhanced selectivity toward Pgp, these substituted dihydropyrroloquinolines may be effective MDR modulators in a clinical setting.

J Med Chem, 2004 Mar 11, 47(6), 1339 - 50
Design and synthesis of new templates derived from pyrrolopyrimidine as selective multidrug-resistance-associated protein inhibitors in multidrug resistance; Wang S et al.; In our continued effort to identify selective MRP1 modulators, we have developed two novel templates, 3 and 4, through rational drug design by identifying the key pharmacophore interaction at the 7-position of the pyrrolopyrimidine template 1 . Further synthesis and SAR work on these novel templates gave a number of potent MRP1 modulators with great selectivity against Pgp . Additional studies to reduce the CYP3A4 inhibition are also reported . Several compounds of these classes were subjected to in vivo xenograft studies and in vivo efficacies were demonstrated.

J Med Chem, 2004 Mar 11, 47(6), 1329 - 38
Studies on pyrrolopyrimidines as selective inhibitors of multidrug-resistance-associated protein in multidrug resistance; Wang S et al.; Multidrug resistance mediated by P-glycoprotein (Pgp) or multidrug-resistance-associated protein (MRP) remains a major obstacle for successful treatment of cancer . Inhibition of Pgp and MRP transport is important for high efficacy of anticancer drugs . While several Pgp inhibitors have entered clinical trials, the development of specific MRP1 inhibitors is still in its infancy . In our screening program, we have identified a pyrrolopyrimidine (4) as a novel and selective MRP1 inhibitor . Subsequent SAR work on the 4-position of the template revealed the phenethylpiperazine side chain as a potent replacement of the benzylthio group of the lead molecule . Introduction of groups at the 2-position seems to have no detrimental effect on activity . Modifications to the nitrile group at the 7-position resulted in the identification of analogues with groups, such as amides, with superior pharmacokinetic profiles . In vivo efficacy has been demonstrated by xenograft studies on selected compounds.

Blood, 2004 Jun 15, 103(12), 4636 - 43 Epub 2004 Mar 02.
T-cell lymphoma as a model for the use of histone deacetylase inhibitors in cancer therapy: impact of depsipeptide on molecular markers, therapeutic targets, and mechanisms of resistance; Piekarz RL et al.; Depsipeptide (FK228) is a novel histone deacetylase inhibitor currently in clinical trials and the first to demonstrate clinical activity in patients . Responses have been observed in patients with T-cell lymphomas, despite prior treatment with multiple chemotherapeutic agents . To better understand the effects of histone deacetylase inhibitors on T-cell lymphoma, the human T-cell lymphoma cell line HUT78 was tested for sensitivity and molecular response to depsipeptide . Treatment with depsipeptide, as well as other histone deacetylase inhibitors, caused induction of histone acetylation, induction of p21 expression, and substantial apoptosis without significant cell cycle arrest . Treatment with the caspase inhibitor z-VAD-fmk significantly inhibited depsipeptide-induced apoptosis, enabling detection of cell cycle arrest . Treatment with depsipeptide increased expression of the interleukin-2 (IL-2) receptor, and combination with the IL-2 toxin conjugate denileukin diftitox resulted in more than additive toxicity . Cells selected for resistance to depsipeptide overexpressed the multidrug resistance pump, P-glycoprotein (Pgp) . However, cells selected for resistance to depsipeptide in the presence of a Pgp inhibitor had a Pgp-independent mechanism of resistance . These studies confirm the activity of depsipeptide in a T-cell lymphoma model and suggest a general sensitivity of T-cell lymphoma to histone deacetylase inhibitors, an emerging new class of anticancer agents.

Infect Control Hosp Epidemiol, 2004 Feb, 25(2), 162 - 4
Which strategies follow from the surveillance of multidrug-resistant bacteria to strengthen the control of their spread? A French experience; Lepelletier D et al.; Efforts to enhance standard precautions and to isolate patients with positive routine clinical cultures during 3 years were insufficient to decrease multidrug-resistant bacteria infection rates . Routine screening for carriage in high-risk patients may be necessary to halt transmission and control the hospital reservoir.

Clin Ter, 2003 Sep-Oct, 154(5), 325 - 35
{MDR (multidrug resistance) in hepatocarcinoma clinical-therapeutic implications}; Petraccia L et al.; Our purpose was to summarize current knowledge on "multidrug resistance", or MDR, an intrinsic or acquired cross resistance to a variety of structurally and functionally unrelated drugs, still representing one of the major problems in the therapy of cancer and other diseases . MDR depends on various mechanisms, the best known being the activity of ABC transport proteins, mainly Pgp, MDR1 gene product,and MRPs; but also other transporters can cause resistance, for example TAP, a peptide transporter, CFTR, cystic fibrosis transmembrane regulator, ABCG2, or breast cancer resistance protein (BCRP) and LRP, lung resistance protein . MDR has been detected in nearly all types of cancer, because it affects many organs and can occur against a wide number of drugs; it is frequent even in other diseases, such as epilepsy and HIV . We focused on MDR phenomenon in HCC, one of the commonest tumors in the world, and one of the most resistant to pharmacological treatment . This characteristic might be partly determined by a link between MDR and angiogenic phenotypes . The relationship between MDR in hepatocellular carcinoma and the effectiveness of therapeutic treatments has been particularly examined . Finally, the importance to overcome the strong chemoresistance of hepatocellular carcinoma with methods alternative to drugs, namely gene therapy, which makes use of antisense oligonucleotides and anti-MDR1 ribozymes, has been pointed out.

Mol Interv, 2001 Jun, 1(2), 117 - 25
Transcription of the multidrug resistance gene MDR1: a therapeutic target; Scotto KW et al.; Two decades ago, the overexpression of P-glycoprotein (Pgp) was first demonstrated to mediate the energy-dependent efflux of a variety of chemotherapeutic agents from tumor cells, resulting in the development of multidrug resistance (MDR) . Not surprisingly, this discovery triggered an ongoing search for agents that would inhibit Pgp function, with the hope that by doing so the MDR phenotype could be reversed . As our understanding of Pgp function and pharmacokinetics has increased, this quest has become more urgent, as well as more complex.

Yao Xue Xue Bao, 2003 Nov, 38(11), 805 - 8
{Effects of 3-substituted aryl oxindole(PH II-7) on cell cycle of tumor cells}; Tan YH et al.; AIM: To study the antitumor mechanism of 3-substituted aryl oxindole (PH II-7) and determine its effects on cell cycle distribution of tumor cells . METHODS: The cell cycle distributions were determined with FACS . The cell cycle regulation-related proteins of K562 lysates were analyzed with Western Blot . The inhibition of PH II-7 on DNA synthesis of tumor cells were estimated though 3H-thymidine incorporation and the tyrosine kinase activity of EGFR of A431 lysates was measured with ELISA . RESULTS: PH II-7 effected cell cycle distribution of several tumor cells, including multidrug resistant tumor cell lines, and accumulation of cells in the G0-G1 stages was observed . The cell cycle regulation-related proteins CDK2, Rb and c-myc were inhibited by PH II-7 in a dose dependent manner, whereas the expression of CyclinE was increased after exposure to PH II-7 . Furthermore, PH II-7 2.0 mg.L-1 was shown to inhibit the incorporation of 3H-thymidine into DNA, and 21.89%-41.29% of the PTK activity of EGFR in A431 lysates was inhibited by PH II-7 2-8 mg.L-1 in a dose-dependant manner . CONCLUSION: PH II-7, a new anti-tumor agent, blocks the transition of cell cycle of tumor cells from G1 to S phase by inhibition CDK2.

J Pathol, 2004 Mar, 202(3), 305 - 12
Expression of COX-2, mPGE-synthase1, MDR-1 (P-gp), and Bcl-xL: a molecular pathway of H pylori-related gastric carcinogenesis; Nardone G et al.; Helicobacter pylori up-regulates cyclo-oxygenase-2 (COX-2) expression, which in turn is involved in tumourigenesis . Recently, a causal link between COX-2 and multidrug resistance 1 (MDR-1) gene expression, implicated in cancer chemoresistance, has been demonstrated . Thus, the expression of COX-2 and the downstream enzyme involved in PGE2 biosynthesis, microsomal PGE-synthase1 (mPGES1), was correlated with P-gp, the product of MDR-1, and the anti-apoptotic protein, Bcl-xL, in gastric biopsies from patients with H pylori infection and in patients with gastric cancer . In a retrospective analysis of endoscopic and pathology files, 40 H pylori-negative patients (Hp-), 50 H pylori-positive patients who responded to eradication therapy (Hp+R), 84 H pylori-positive patients who did not respond to eradication therapy (Hp+NR), and 30 patients with gastric cancer (18 intestinal and 12 diffuse types) were selected . COX-2, mPGES1, P-gp, and Bcl-xL were detected by immunohistochemistry . COX-2, mPGES1, P-gp, and Bcl-xL expression was undetectable in gastric mucosa from Hp- patients . By contrast, COX-2 and mPGES1 expression was detected in 42% and 44% of Hp+R patients, respectively, and in up to 66% (range 63-66%) of Hp+NR patients (p < 0.05) . The expression of COX-2 and mPGES1 correlated significantly (p < 0.0001) with that of P-gp and Bcl-xL . High levels of COX-2, mPGES1, P-gp, and Bcl-xL expression were found in intestinal-type gastric cancer samples . In conclusion, H pylori-dependent induction of COX-2 and mPGES1 is associated with enhanced production of P-gp and Bcl-xL that may contribute to gastric tumourigenesis and resistance to therapy .

J Virol, 2004 Mar, 78(6), 3123 - 32
Crystal structures of a multidrug-resistant human immunodeficiency virus type 1 protease reveal an expanded active-site cavity; Logsdon BC et al.; The goal of this study was to use X-ray crystallography to investigate the structural basis of resistance to human immunodeficiency virus type 1 (HIV-1) protease inhibitors . We overexpressed, purified, and crystallized a multidrug-resistant (MDR) HIV-1 protease enzyme derived from a patient failing on several protease inhibitor-containing regimens . This HIV-1 variant contained codon mutations at positions 10, 36, 46, 54, 63, 71, 82, 84, and 90 that confer drug resistance to protease inhibitors . The 1.8-angstrom (A) crystal structure of this MDR patient isolate reveals an expanded active-site cavity . The active-site expansion includes position 82 and 84 mutations due to the alterations in the amino acid side chains from longer to shorter (e.g., V82A and I84V) . The MDR isolate 769 protease "flaps" stay open wider, and the difference in the flap tip distances in the MDR 769 variant is 12 A . The MDR 769 protease crystal complexes with lopinavir and DMP450 reveal completely different binding modes . The network of interactions between the ligands and the MDR 769 protease is completely different from that seen with the wild-type protease-ligand complexes . The water molecule-forming hydrogen bonds bridging between the two flaps and either the substrate or the peptide-based inhibitor are lacking in the MDR 769 clinical isolate . The S1, S1', S3, and S3' pockets show expansion and conformational change . Surface plasmon resonance measurements with the MDR 769 protease indicate higher k(off) rates, resulting in a change of binding affinity . Surface plasmon resonance measurements provide k(on) and k(off) data (K(d) = k(off)/k(on)) to measure binding of the multidrug-resistant protease to various ligands . This MDR 769 protease represents a new antiviral target, presenting the possibility of designing novel inhibitors with activity against the open and expanded protease forms.

Zhongguo Shi Yan Xue Ye Xue Za Zhi, 2004 Feb, 12(1), 6 - 10
{Methylation status of multidrug resistance (mdr1) gene and its correlation with expression of mdr1 gene in patients with hematologic malignancies}; Zhu Y et al.; To investigate the correlation between methylation and expression of multidrug resistance (mdr1) gene, restriction endonuclease HpaII combined with competitive PCR technique was used to quantitatively detect the methylation status of two CCGG sites located at -110 and -50 bp (region I and II) up to the transcription start site in mdr1 promoter in 54 AL and 9 MM patients . Semi-quantitative RT-PCR was used to detect the expression level of mdr1 gene . The results showed that inverse correlation between methylation rate of either region or total methylation rate and expression of mdr1 gene was observed . The correlation in the region I (r = -0.64) was closer than that in the region II (r = -0.4) . High expression rate of mdr1 ascended significantly in low methylation group (n = 36) (P < 0.001) . In comparison with chemotherapy sensitive group (n = 8), the methylation rate in refractory AL patients (n = 16) was lower (P = 0.05) in the region I, P < 0.05 in the region II and total regions . Comparing with the untreated patients (n = 36), the methylation rate in the region I and total methylation rate were lower in the patients with chemotherapy (n = 14) (P < 0.05) . The methylation rate in the region II was also decreased after chemotherapy, however, no statistical significance was shown (P > 0.05) . Increased mdr1 expression level accompanying with decreased methylation rate after chemotherapy was found, although no significant difference was shown (P = 0.06) . It is concluded that the expression level of mdr1 gene was associated with the methylation status of CCGG in -110 and -50 bp upstream to the transcription start site, especially the -110 site . In both the patients treated with chemotherapy and the refractory patients, the methylation level of mdr1 gene decreased relatively . The rising expression of mdr1 gene after chemotherapy was associated with the decrease of methylation level.

Eur J Med Chem, 2004 Jan, 39(1), 59 - 67
Structure-activity relationship study of antimalarial indolo {2,1-b}quinazoline-6,12-diones (tryptanthrins) . Three dimensional pharmacophore modeling and identification of new antimalarial candidates; Bhattacharjee AK et al.; A widely applicable three-dimensional QSAR pharmacophore model for antimalarial activity was developed from a set of 17 substituted antimalarial indolo{2,1-b}quinazoline-6,12-diones (tryptanthrins) that exhibited remarkable in vitro activity (below 100 ng/mL) against sensitive and multidrug-resistant Plasmodium falciparum malaria . The pharmacophore, which contains two hydrogen bond acceptors (lipid) and two hydrophobic (aromatic) features, was found to map well onto many well-known antimalarial drug classes including quinolines, chalcones, rhodamine dyes, Pfmrk cyclin dependent kinase inhibitors, malarial FabH inhibitors, and plasmepsin inhibitors . The phamacophore allowed searches for new antimalarial candidates from multiconformer 3D databases and enabled custom designed synthesis of new potent analogues.

Eur J Med Chem, 2004 Feb, 39(2), 161 - 77
Anti-calmodulin acridone derivatives modulate vinblastine resistance in multidrug resistant (MDR) cancer cells; Hegde R et al.; Multidrug resistance (MDR) is one of the main obstacles limiting the efficacy of chemotherapy treatment of tumors . Parent acridones 1A and 1B were prepared by the Ullmann reaction followed by cyclization and N-alkylation . N-(omega-Chloroalkyl) analogues were subjected to iodide catalyzed nucleophilic substitution reaction with secondary amines to get the compounds 3A-13A and 3B-13B, which enhanced the uptake of vinblastine in KBChR-8-5 cells to a greater extent (2.6-13.1-fold relative to control) than verapamil . The study on the structure-activity relationship revealed that substitution of -H at position C-4 in acridone nucleus by -OCH3 increased the cytotoxic and anti-MDR activities . The ability of acridones to inhibit calmodulin dependent cyclic AMP phosphodiesterase has been determined and the results have shown a strong positive correlation between anti-calmodulin activity and cytotoxicity in KBChR-8-5 cells or anti-MDR activity.

J Nat Prod, 2004 Feb, 67(2), 245 - 56
Biological activity and chemistry of taxoids from the Japanese yew, Taxus cuspidata; Shigemori H et al.; Approximately 120 taxoids have been isolated to date from the Japanese yew, Taxus cuspidata . These taxoids possess various skeletons containing 5/7/6-, 6/10/6-, 6/5/5/6-, 6/8/6-, or 6/12-membered ring systems . Among the taxoids, some non-paclitaxel-type compounds have been shown to reduce CaCl(2)-induced depolymerization of microtubules, increase cellular accumulation of vincristine in multidrug-resistant tumor cells, and exhibit potent cytotoxicity . Chemical derivatization of taxoids of T . cuspidata is also reviewed.

Clin Neuropathol, 2004 Jan-Feb, 23(1), 21 - 7
Heterogeneity in the expression of markers for drug resistance in brain tumors; Andersson U et al.; Brain tumors, in general, display a multidrug-resistant phenotype . This study evaluated the immunohistochemical expression and distribution of P-glycoprotein (Pgp), multidrug resistance protein (MRP1), lung resistance protein (LRP) and O6 methylguanine-DNA methyltransferase (MGMT) in low- and high-grade astrocytoma, oligodendroglioma and in different subgroups of meningioma . The results revealed a marked heterogeneity in the expression and distribution among the analyzed tumors . In astrocytoma and oligodendroglioma, Pgp and MRP1 were observed in the capillary endothelium and in scattered tumor cells, whereas LRP occurred only in tumor cells . A pronounced expression of MGMT was found independent of the histopathological grade . An enhanced expression of MRP1 and LRP in astrocytoma and oligodendroglioma were more often evident in older patients (> 50 years) . Survival analysis suggested a markedly decreased overall survival for patients suffering from low-grade glioma overexpressing Pgp . In meningioma, a heterogeneous expression of Pgp, MRP1, LRP and MGMT was seen with the most prominent staining localized to the capillary endothelium . Pgp was significantly more often overexpressed (p < 0.05) in transitional compared to meningothelial meningioma . The marked heterogeneity in the expression suggests that analysis of these factors can be of importance in the selection of individualized chemotherapy, regardless of tumor type.

Biochem Biophys Res Commun, 2004 Mar 19, 315(4), 991 - 7
Effects of tributyltin on barrier functions in human intestinal Caco-2 cells; Tsukazaki M et al.; The effect of tributyltin (TBT) on human intestinal epithelial cell functions was investigated by using human intestinal Caco-2 cell monolayers . We paid particular attention to the effect of TBT on two barrier functions: the tight junction as a physical barrier and MDR1/P-glycoprotein as a biological barrier . A loss of monolayer integrity was apparent from the TBT treatment and the paracellular permeability was increased by TBT . On the other hand, the activity of P-glycoprotein, which was examined by measuring the accumulation of Rhodamine-123 and daunomycin, was increased by prolonged TBT treatment in a concentration-dependent manner (1-100 nM) . Furthermore, it was clarified by Western and Northern blots that this increase was accompanied by the increased expression of MDR1 mRNA and protein . The activation of a multidrug resistance transporter P-glycoprotein by TBT would cause a disorder of the human intestines by changing the drug pharmacokinetics.

Biochem Biophys Res Commun, 2004 Mar 19, 315(4), 942 - 9
Distinct groups of multidrug resistance modulating agents are distinguished by competition of P-glycoprotein-specific antibodies; Nagy H et al.; P-glycoprotein (Pgp) is one of the ABC transporters responsible for the multidrug resistance of cancer cells . The conformational changes of Pgp that occur in the presence of substrates/modulators or ATP depletion are accompanied by the up-shift of UIC2 monoclonal antibody (mAb) binding . In the case of cyclosporin A, vinblastine or valinomycin, this up-shift was found to be concomitant with the near-complete suppression of labeling with other mAbs specific for Pgp epitopes overlapping with UIC2, while pre-treatment with verapamil or Tween 80 brings about a modest suppression . Here we have extended these observations to 44 Pgp interacting agents, and found that only 8 fall into the cyclosporin-like category, inducing a conformational state characterized by the complete UIC2 dominance . The rest of the drugs either did not affect antibody competition or had a modest effect . Thus, Pgp substrates/modulators can be classified into distinct modalities based on the conformational change they elicit.

Pediatr Neurol, 2004 Feb, 30(2), 102 - 6
Multidrug resistance proteins in tuberous sclerosis and refractory epilepsy; Lazarowski A et al.; Tuberous sclerosis is an autosomal dominant syndrome characterized by seizures that are refractory to medication in severely affected individuals . The mechanism involved in drug resistance in tuberous sclerosis is unknown . The proteins MDR-1 (multidrug resistance) and MRP-1 (multidrug resistance-associated protein-1) are linked to chemotherapy resistance in tumor cells . However, the relationship between refractoriness to antiepileptic drugs and MDR-1 or MRP-1 brain expression has been poorly studied . We have previously described a case of tuberous sclerosis with refractory epilepsy that expressed multidrug resistance gene (MDR-1) in tuber cells from epileptogenic brain lesion . In this retrospective study, we describe the expression of MDR-1 and MRP-1 in the epileptogenic cortical tubers of three pediatric patients with tuberous sclerosis and refractory epilepsy surgically treated . Monoclonal antibodies for MDR-1 and MRP-1 proteins were used for immunohistochemistry . In epileptogenic cortical tuber brain specimens, MDR-1 and MRP-1 proteins were strongly immunoreactive in abnormal balloon cells, dysplastic neurons, astrocytes, microglial cells, and some blood-brain vessels . A more extensive MDR-1 immunoreactivity was observed . These data suggest that refractory epilepsy phenotype in tuberous sclerosis can be associated with the expression of both multidrug resistance MDR-1 and MRP-1 transporters in epileptogenic cortical tubers.

Proc Natl Acad Sci U S A, 2004 Feb 24, 101(8), 2470 - 5
Bilirubin protects astrocytes from its own toxicity by inducing up-regulation and translocation of multidrug resistance-associated protein 1 (Mrp1); Gennuso F et al.; Unconjugated bilirubin (UCB) causes encephalopathy in severely jaundiced neonates by damaging astrocytes and neurons . Astrocytes, which help defend the brain against cytotoxic insults, express the ATP-dependent transporter, multidrug resistance-associated protein 1 (Mrp1), which mediates export of organic anions, probably including UCB . We therefore studied whether exposure to UCB affects the expression and intracellular localization of Mrp1 in cultured mouse astroglial cells (>95% astrocytes) . Mrp1 was localized and quantitated by confocal laser scanning microscopy and double immunofluorescence labeling by using specific antibodies against Mrp1 and the astrocyte marker glial fibrillary acidic protein, plus the Golgi marker wheat germ agglutinin (WGA) . In unexposed astrocytes, Mrp1 colocalized with WGA in the Golgi apparatus . Exposure to UCB at a low unbound concentration (Bf) of 40 nM caused rapid redistribution of Mrp1 from the Golgi throughout the cytoplasm to the plasma membrane, with a peak 5-fold increase in Mrp1 immunofluorescence intensity from 30 to 120 min . Bf above aqueous saturation produced a similar but aborted response . Exposure to this higher Bf for 16 h markedly decreased Trypan blue exclusion and methylthiazoletetrazoilum activity and increased apoptosis 5-fold by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling assay . These toxic effects were modestly increased by inhibition of Mrp1 activity with 3-({3-(2-{7-chloro-2-quinolinyl}ethenyl)phenyl-(3-dimethylamino-3-oxopropyl)-thio-methyl}thio)propanoic acid (MK571) . By contrast, Bf=40 nM caused injury only if Mrp1 activity was inhibited by MK571, which also blocked translocation of Mrp1 . Our conclusion is that in astrocytes, UCB up-regulates expression of Mrp1 and promotes its trafficking from the Golgi to the plasma membrane, thus moderating cytotoxicity from UCB, presumably by limiting its intracellular accumulation.

Antimicrob Agents Chemother, 2004 Mar, 48(3), 954 - 60
Plasmodium falciparum-based bioassay for measurement of artemisinin derivatives in plasma or serum; Teja-Isavadharm P et al.; Artemisinin and its derivatives, artesunate and artemether, are rapidly acting antimalarials that are used for the treatment of severe and uncomplicated multidrug-resistant falciparum malaria . To optimize treatment regimens that use this new class of antimalarials, there is a need for readily available and reproducible assays to monitor drug levels closely in patients . A sensitive and reproducible bioassay for the measurement of the concentrations of artemisinin derivatives in plasma and serum is described . By modifying the in vitro drug susceptibility test, it was found that antimalarial activity in plasma or serum containing an unknown concentration of drug could be equated to the known concentrations of dihydroartemisinin (DHA) required to inhibit parasite growth . Dose-response curves for a Plasmodium falciparum clone (clone W2) and DHA were used as a standard for each assay . Assays with plasma or serum spiked with DHA proved to be reproducible (coefficient of variation, <or=10.9%), with a lower limit of quantitation equivalent to 2.5 ng of DHA per ml . For plasma spiked with artesunate or artemether, there was good agreement of the results obtained by the bioassay and the concentrations measured by high-performance liquid chromatography (HPLC) with electrochemical detection . The bioassay for measurement of the antimalarial activities of artemisinin derivatives in body fluids requires a smaller volume of plasma or serum and is more sensitive than the presently available HPLC methods, can provide pharmacodynamic parameters for determination of activity against the parasite, and should enhance the design of more appropriate dosage regimens for artemisinin drugs.

Antimicrob Agents Chemother, 2004 Mar, 48(3), 809 - 14
Possible involvement of the drug transporters P glycoprotein and multidrug resistance-associated protein Mrp2 in disposition of azithromycin; Sugie M et al.; P glycoprotein and multidrug resistance-associated protein 2 (Mrp2), ATP-dependent membrane transporters, exist in a variety of normal tissues and play important roles in the disposition of various drugs . The present study seeks to clarify the contribution of P glycoprotein and/or Mrp2 to the disposition of azithromycin in rats . The disappearance of azithromycin from plasma after intravenous administration was significantly delayed in rats treated with intravenous injection of cyclosporine, a P-glycoprotein inhibitor, but was normal in rats pretreated with intraperitoneal injection erythromycin, a CYP3A4 inhibitor . When rats received an infusion of azithromycin, cyclosporine and probenecid, a validated Mrp2 inhibitor, significantly decreased the steady-state biliary clearance of azithromycin to 5 and 40% of the corresponding control values, respectively . However, both inhibitors did not alter the renal clearance of azithromycin, suggesting the lack of renal tubular secretion of azithromycin . Tissue distribution experiments showed that azithromycin is distributed largely into the liver, kidney, and lung, whereas both inhibitors did not alter the tissue-to-plasma concentration ratio of azithromycin . Significant reduction in the biliary excretion of azithromycin was observed in Eisai hyperbilirubinemic rats, which have a hereditary deficiency in Mrp2 . An in situ closed-loop experiment showed that azithromycin was excreted from the blood into the gut lumen, and the intestinal clearance of azithromycin was significantly decreased by the presence of cyclosporine in the loop . These results suggest that azithromycin is a substrate for both P glycoprotein and Mrp2 and that the biliary and intestinal excretion of azithromycin is mediated via these two drug transporters.

Med Mycol, 2004 Feb, 42(1), 59 - 71
Profiling gene expression in Coccidioides posadasii; Delgado N et al.; Coccidioides posadasii is a dimorphic fungal pathogen which grows as a filamentous saprobe in the soil and multicellular parasitic form in host lung tissue . Studies of gene expression profiles during saprobic and parasitic phase development can provide clues about morphogenetic regulation and may lead to the discovery of molecular targets for novel antifungal drugs . Suppression-subtractive hybridization (SSH) and quantitative real-time polymerase chain reaction (QRT-PCR) were used to identify and quantify differential gene expression during in vitro growth of Coccidioides . DNA fragments obtained from the subtraction of cDNA pools derived from the saprobic and parasitic phase RNA preparations were each cloned into an appropriate vector and subjected to sequence analysis . Semi-quantitative, reverse transcription polymerase chain reaction (RT-PCR) experiments were first conducted to assess whether these inserts represented differentially expressed genes . Nucleotide sequences of the partial and full-length genes selected by RT-PCR were obtained by genome walking and rapid amplification of cDNA ends (RACE) methods . QRT-PCR analysis of the expression of these genes during saprobic and parasitic cell growth was then conducted using DNA standard curves normalized to a constitutively expressed control gene . Four C . posadasii genes whose expression is essentially restricted to the parasitic cycle were discovered using this approach . These genes include homologues of OPS1 (encodes opsin-related protein), MDR1 (multidrug resistance protein), ALDR1 (aldehyde reductase), and PSP1 (hypothetical lipid transporter/flippase protein) . The combined applications of SSH and QRT-PCR permit global analysis of gene expression patterns in C . posadasii.

Anticancer Res, 2003 Nov-Dec, 23(6C), 4781 - 7
Conjugated bilirubin induces multidrug resistance-associated protein 2 mRNA expression and in vivo cisplatin resistance in rat hepatoma AH66 cells; Tamai M et al.; In vivo cisplatin resistance of rat ascites hepatoma AH66 cells is suggested to result from the induction of multidrug resistance-associated protein 2 (MRP2) expression by ascites fluid (ASF) in the peritoneal cavity . The in vitro cisplatin sensitivity of AH66 cells grown in assay medium containing 5% fetal bovine serum in Dulbecco's modified Eagle's medium (5% FBS DMEM) did not change when the cells were treated with probenecid, an inhibitor of anion transporters, while the decreased cisplatin sensitivity of AH66 cells cultured in an assay medium containing 5% ASF (5% ASF DMEM) was restored by probenecid . Furthermore, in an in vivo study, the survival span (%ILS) of AH66-bearing rats was markedly extended by combination therapy with cisplatin and probenecid, compared with either agent alone . The expression of MRP2 mRNA was increased when AH66 cells were cultured in medium containing 5% ASF or 5% bile for 24 h . The induction of MRP2 mRNA expression in AH66 cells was also observed in the presence of heat-denatured ASF . The bilirubin content in ASF was characteristically higher than that in FBS, normal rat serum or AH66-bearing rat serum . Unconjugated bilirubin did not change the expression of MRP2 mRNA, whereas conjugated bilirubin markedly increased it . The cisplatin uptake in AH66 cells after culture in 5% FBS DMEM containing conjugated bilirubin was about half that of the cells cultured in 5% FBS DMEM alone (p < 0.01) . In addition, the cisplatin sensitivity of the cells was significantly lowered by the addition of conjugated bilirubin . The expression of MRP2 mRNA in rat normal hepatocytes was also increased after culture in medium containing 5% ASF or 5% bile . These results indicated that conjugated bilirubin, a component of ASF, induces the mRNA expression of MRP2, which is a determinant of the in vivo cisplatin resistance of AH66 cells.

Anticancer Res, 2003 Nov-Dec, 23(6C), 4737 - 46
Differential control of cholesterol and fatty acid biosynthesis in sensitive and multidrug-resistant LoVo tumor cells; Santini MT et al.; Multidrug resistance (MDR) describes the decrease in sensitivity of tumor cells to a wide variety of cytotoxic compounds . Although a central role has been ascribed to the P-glycoprotein (Pgp) pump in MDR, lipids also appear to be extremely important . However, their precise role in MDR is not yet fully understood . It was the aim of the present paper to gain a deeper understanding of intracellular lipid equilibrium in both sensitive and MDR tumor cells . In particular, intracellular cholesterol biosynthesis and cholesterol esterification were examined in LoVo-sensitive and Pgp-overexpressing resistant cells . The data presented seem to suggest that the higher synthesis of cholesteryl ester and triglyceride observed in resistant with respect to wild-type cells is due to a greater production of fatty acids in these cells . The results are discussed in view of the possible roles of sterol regulatory element-binding proteins and Pgp in these phenomena.

Anticancer Res, 2003 Nov-Dec, 23(6C), 4607 - 11
P-glycoprotein modulation improves in vitro chemosensitivity in malignant pediatric liver tumors; Warmann S et al.; BACKGROUND: Multidrug resistance (MDR) is a major reason for the poor outcome of advanced pediatric liver malignancies . The P-glycoprotein (P-gP), which contributes to this phenomenon, has been potently antagonized in other tumors . Our aim was to investigate the influence of P-gP antagonizers on the chemotherapy of pediatric liver malignancies in vitro . MATERIALS AND METHODS: One hepatocellular carcinoma (HCC) and three hepatoblastoma (HB) cell lines were incubated with doxorubicin or cisplatin . Additional effects of three P-gP-modulators were determined in a cytotoxicity assay . Expression levels of the MDR1 gene were determined using rT-PCR . RESULTS: Modulation of P-gP improved chemotherapy results in all HB cell lines, more effectively in the more highly differentiated tumors . Combined treatment of the HCC cell line was only more efficient using doxorubicin and PSC 833 . CONCLUSION: Modulation of P-gP can overcome MDR in HCC and HB in vitro . Our data encourage further studies analyzing this effect under in vivo conditions.

Leukemia, 2004 Mar, 18(3), 513 - 20
Efficient inhibition of multidrug-resistant human tumors with a recombinant bispecific anti-P-glycoprotein x anti-CD3 diabody; Gao Y et al.; Overexpressing of P-glycoprotein (Pgp) has been shown to be responsible for cancer resistance to multiple chemotherapeutic agents . Immunotherapy with biological agents, such as bispecific antibodies (BsAbs), may represent a promising approach to overcome the emergence of drug resistance . Here we constructed a recombinant BsAb, a diabody, with specificities to both CD3 on human T-lymphocyte and Pgp on cancer cells . The diabody was produced in Escherichia coli in a soluble functional form and purified by an affinity chromatography with a yield of >4 mg/l culture medium in shaker flask . The diabody binds to both CD3 on T-lymphocytes and Pgp on multidrug-resistant (MDR) tumor cells with affinities that are comparable to its respective parental single chain Fv molecules . In the presence of activated human peripheral blood lymphocytes (PBLs), the diabody mediates effectively the lysis of the Pgp-overexpressing human leukemia K562/A02 and epidermoid carcinoma KBv(200) cells, but is much less potent in mediating the lysis of the parent K562 and KB cells . Further, the diabody localized selectively within the K562/A02 xenografts in mice . When combined with activated PBL, the diabody significantly inhibited the growth of K562/A02 and KBv(200), but had no effect on K562 and KB xenografts . In contrast, treatment with doxorubicin, a standard chemotherapeutic agent, only inhibited the growth of K562 and KB, but had no effect on K562/A02 and KBv(200) xenografts . Taken together, our results suggest that the anti-Pgp x anti-CD3 diabody may have a great potential in the treatment of various MDR cancers.

Bioorg Med Chem, 2004 Mar 1, 12(5), 1177 - 82
Synthesis and antimalarial activity of a new series of trioxaquines; Singh C et al.; Trioxanes 8a-b, easily accessible in two steps from allylic alcohol 6a-b, on reductive amination with 4-aminoquinolines 4a-c furnish a new series of trioxaquines 9a-b, 10a-b, 11a-b in 32-77% yields . Dicitrate salts of these trioxaquines have been evaluated for antimalarial activity against multidrug resistant Plasmodium yoelii in mice model.

Zhonghua Gan Zang Bing Za Zhi, 2004 Feb, 12(2), 95 - 8
{Establishment of human hepatocellular carcinoma multidrug-resistance cell line (HepG2/Adm) and study apoptosis induced by low-frequency pulse ultrasound exposure}; Zhai BJ et al.; OBJECTIVE: To establish human hepatocellular carcinoma multidrug-resistance cell line (HepG2/ADM) and to determine the effect of low-frequency pulse ultrasound (US) on MDR cells and investigate its mechanism . METHODS: Using gradual increase of adriamycin (ADM) concentrations in culture, an adriamycin-resistant human hepatocellular carcinoma cell sub line (HepG2/ADM) was established in vitro . HepG2/ADM cells were cultured in vitro and randomly divided into 4 groups: the control group (HepG2/ADM only), the group ADM by 1.0 mug/ml adriamycin for 1 h, the group US by low-frequency pulse ultrasound for 10 min, and the group US by low-frequency pulse ultrasound and treated with adriamycin simultaneously at same time . A sonication at a frequency of 0.8 MHz, was delivered with an intensity level of 0.5W/cm2, with continuous exposure of 10 min was applied . The ability of US to induce the apoptosis of MDR was evaluated by analyses of fluorescence microscopy, DNA fragmentation assay and flow cytometry assay . RESULTS: HepG2/Adm was resistant to many anti-tumor agents, and its IC50 of ADM was 26 times higher than that of parent cell line HepG2 . Significant over expressions of P-gp, MRP, LRP and GSTs were detected . HepG2/ADM cells radiated by US had the typical characteristics of apoptosis . Compared with the control group (HepG2/ADM, 3.47%); the apoptosis rates were higher in US (12.23%) . The therapeutic alliance of US with ADM for MDR cells, have a significant change in the ratio of apoptosis (18.81%, t=1.46 to 5.36, P<0.01) . CONCLUSION: HepG2/ADM could have the biological characteristics of human multidrug-resistance cell line . The US sonication of 0.8 MHz could induce apoptosis of HepG2/ADM cell in vitro, and could act synergistically with Adriamycin.

Zhonghua Gan Zang Bing Za Zhi, 2004 Feb, 12(2), 85 - 7
{Reversal of multidrug resistance gene MDR1 and MRP of drug-resistant human hepatocellular carcinoma cells SMMC-7721/ADM with antisense phosphorothioate oligonucleotides}; Luo HY et al.; OBJECTIVES: To investigate the reversal effect of gene MDR1 and MRP with combinational antisense phosphorothioate oligonucleotide on Drug-resistant human hepatocellular carcinoma cells SMMC-7721/ADM . METHODS: SMMC-7721/ADM was transfected with synthetic antisense phosphorothioate oligonucleotides complementary to gene MDR1 and MRP mediated by Lipofectamine . Drug sensitivity was measured by MTT assay, Fluorescence intensity of cells was determined by flow cytometric analysis, RH123 and DNR retention was assayed by confocal scanning laser microscopy . RESULTS: ASODN of MDR1+MRP increased the sensitivity of SMMC-7721/ADM to chemotherapeutic drug more significantly than that any of MDR1 and MRP did separately . But they did not enhance the inhibition expression of protein of p190 or p170 . CONCLUSION: Drug-resistance could be reversed significantly when antisense phosphorothioate oligonucleotide of MDR1+MRP were transfected into drug-resistant human hepatocellular carcinoma cells SMMC-7721/ADM together.

Biochemistry, 2004 Mar 2, 43(8), 2345 - 52
Glutathione S-transferases (GSTs) inhibit transcriptional activation by the peroxisomal proliferator-activated receptor gamma (PPAR gamma) ligand, 15-deoxy-delta 12,14prostaglandin J2 (15-d-PGJ2); Paumi CM et al.; 15-Deoxy-Delta(12,14)prostaglandin J(2) (15-d-PGJ(2)), a terminal metabolite of the J-series cyclopentenone prostaglandins, influences a variety of cellular processes including gene expression, differentiation, growth, and apoptosis . As a ligand of peroxisomal proliferator-activated receptor gamma (PPAR gamma), 15-d-PGJ(2) can transactivate PPAR gamma-responsive promoters . Previously, we showed that multidrug resistance proteins MRP1 and MRP3 attenuate cytotoxic and transactivating activities of 15-d-PGJ(2) in MCF7 breast cancer cells . Attenuation was glutathione-dependent and was associated with formation of the glutathione conjugate of 15-d-PGJ(2), 15-d-PGJ(2)-SG, and its active efflux by MRP . Here we have investigated whether the glutathione S-transferases (GST) can influence biological activities of 15-d-PGJ(2) . MCF7 cells were stably transduced with human cytosolic GST isozymes M1a, A1, or P1a . These GSTs had no effect on 15-d-PGJ(2) cytotoxicity when expressed either alone or in combination with MRP1 . However, expression of any of the three GSTs significantly inhibited 15-d-PGJ(2)-dependent transactivation of a PPAR gamma-responsive reporter gene . The degree of inhibition correlated with the level of GST expressed . Under physiologic conditions, the nonenzymatic rate of 15-d-PGJ(2) conjugation with glutathione was significant . Of the three GST isozymes, only GSTM1a-1a further stimulated the rate of 15-d-PGJ(2)-SG formation . Moreover, GSTM1a-1a rate enhancement was only a transient burst that was complete within 15 s . Hence, catalysis plays little, if any, role in GST inhibition of 15-d-PGJ(2)-dependent transactivation . In contrast, inhibition of transactivation was associated with strong GST/15-d-PGJ(2) interactions . Potent inhibition by 15-d-PGJ(2) and 15-d-PGJ(2)-SG of GST activity was observed with K(i) in the 0.15-2.0 microM range for the three GST isozymes, results suggesting avid associations between GST and 15-d-PGJ(2) or 15-d-PGJ(2)-SG . Electrospray ionization mass spectrometry (ESI/MS) studies revealed no stable adducts of GST and 15-d-PGJ(2) indicating that GST/15-d-PGJ(2) interactions are primarily noncovalent . These results are consistent with a mechanism of GST-mediated inhibition of transactivation in which GST binds 15-d-PGJ(2) and 15-d-PGJ(2)-SG thereby sequestering the ligands in the cytosol away from their nuclear target, PPAR gamma.

No Shinkei Geka, 2004 Jan, 32(1), 19 - 26
{Report of two cases with germinoma treated by individual adjuvant chemotherapy based on the mRNA expression of drug-resistance gene}; Kunishio K et al.; We reported two cases with germ cell tumor in which new preliminary treatment trials were performed by chemotherapy using anti-cancer drug selected on the basis of multidrug resistance gene mRNA expression, such as MDR1, MRP1, MRP2, MXR1, MGMT, GST pi and TopoII alpha, from RT-PCR assay . A 28-year-old male had gradually developed DI . MR imaging revealed enhanced tumors in the medulla oblongata, the pineal region and the suprasella region . Biopsy of tumor in the medulla oblongata demonstrated germinoma histologically . RT-PCR assay of this tissue revealed overexpression of MRP1, MGMT and GST pi mRNA, but neither MDR1, MRP2 nor MXR1 was observed . The patient was successfully given carboplatin, mitoxanthrone and ifosphamide after irradiation . A 15-year-old male was admitted to our hospital with high intracranial pressure syndrome . MR imaging revealed enhanced tumor in the pineal region . The tumor was diagnosed as a malignant germ cell tumor, histopathologically . RT-PCR assay of this tissue revealed overexpression of MRP1, MRP2, MXR1, MGMT and GST pi mRNA . Only MDR1 was not expressed . The patient was treated by irradiation including radiosurgery combined with chemotherapy, given cisplatin, etoposide and ifosphamide (ICE regimen), but he died because of progressive disease such as CSF dissemination . It seems that preliminary individual adjuvant chemotherapy based on mRNA expression of drug-resistance gene is available for the treatment of germ cell tumors.

Mol Pharmacol, 2004 Mar, 65(3), 675 - 84
The molecular basis of the action of disulfiram as a modulator of the multidrug resistance-linked ATP binding cassette transporters MDR1 (ABCB1) and MRP1 (ABCC1); Sauna ZE et al.; The overexpression of multidrug resistance protein 1 (MDR1) and multidrug resistance protein 1 (MRP1) gene products is a major cause of multidrug resistance in cancer cells . A recent study suggested that disulfiram, a drug used to treat alcoholism, might act as a modulator of P-glycoprotein . In this study, we investigated the molecular and chemical basis of disulfiram as a multidrug resistance modulator . We demonstrate that in intact cells, disulfiram reverses either MDR1- or MRP1-mediated efflux of fluorescent drug substrates . Disulfiram inhibits ATP hydrolysis and the binding of {alpha-32P}8-azidoATP to P-glycoprotein and MRP1, with inhibition curves comparable with those of N-ethylmaleimide, a cysteine-modifying agent . However, if the ATP sites are protected with excess ATP, disulfiram stimulates ATP hydrolysis by both transporters in a concentration-dependent manner . Thus, in addition to modifying cysteines at the ATP sites, disulfiram may interact with the drug-substrate binding site . We demonstrate that disulfiram, but not N-ethylmaleimide, inhibits in a concentration-dependent manner the photoaffinity labeling of the multidrug transporter with 125I-iodoarylazidoprazosin and {3H}azidopine . This suggests that the interaction of disulfiram with the drug-binding site is independent of its role as a cysteine-modifying agent . Finally, we have exploited MRP4 (ABCC4) to demonstrate that disulfiram can inhibit ATP binding by forming disulfide bonds between cysteines located in the vicinity of, although not in, the active site . Taken together, our results suggest that disulfiram has unique molecular interactions with both the ATP and/or drug-substrate binding sites of multiple ATP binding cassette transporters, which are associated with drug resistance, and it is potentially an attractive agent to combat multidrug resistance.

J Pharmacol Exp Ther, 2004 Jun, 309(3), 1029 - 35 Epub 2004 Feb 20.
Cyclosporin A, but not tacrolimus, inhibits the biliary excretion of mycophenolic acid glucuronide possibly mediated by multidrug resistance-associated protein 2 in rats; Kobayashi M et al.; The onset of diarrhea after the administration of mycophenolate mofetil (MMF) is possibly associated with the biliary excretion of its metabolite, mycophenolic acid glucuronide (MPAG) . This study was undertaken to clarify the mechanism underlying the biliary excretion of MPAG . Intravenously administered mycophenolic acid (MPA, 5 mg/kg) rapidly disappeared from plasma and was efficiently excreted as MPAG in the bile of Wistar (26% of dose) and Sprague-Dawley rats (21% of dose) over 1 h . On the other hand, in spite of the rapid disappearance of MPA from plasma, the biliary excretion of MPAG was very limited in Eisai hyperbilirubinemic rats (EHBRs), which display mutations in multidrug resistance-associated protein 2 (Mrp2)/canalicular multispecific organic anion transporter, and constituted only 0.5% of dose . Instead, high levels of MPA were noted in the plasma of EHBRs . Intravenous administration of CsA (5 mg/kg) to Wistar rats significantly lowered the biliary excretion of MPAG . However, intravenously administered tacrolimus (0.1 mg/kg) failed to produce such effect . In conclusion, it is suggested that there is an efficient MPAG transport mediated by Mrp2 on the bile canalicular membrane of rat hepatocytes and that the therapeutic range of CsA potentially interferes with Mrp2 . However, the therapeutic range of tacrolimus does not inhibit the transporter . Thus, it should be noted that MMF coadministered with tacrolimus instead of CsA might increase the occurrence of diarrhea related to the biliary excretion of MPAG in transplant recipients.

Toxicol Sci, 2004 May, 79(1), 189 - 95 Epub 2004 Feb 19.
Biliary secretory function in rats chronically intoxicated with aluminum; Gonzalez MA et al.; The effects of a chronic aluminum (Al) exposure on biliary secretory function, with special emphasis on hepatic handling of non-bile salt organic anions, was investigated . Male Wistar rats received, intraperitoneally, either 27 mg/kg body weight of Al, as Al hydroxide {Al (+) rats}, or the vehicle saline {Al (-) rats} three times a week for 3 months . Serum and hepatic Al levels were increased by the treatment (approximately 9- and 4-fold, respectively) . This was associated with enhanced malondialdehyde formation (+110%) and a reduction in GSH content (-17%) and in the activity of the antioxidant enzymes catalase (-84%) and GSH peroxidase (-46%) . Bile flow (-23%) and the biliary output of bile salts (-39%), cholesterol (-43%), and proteins (-38%) also decreased . Compartmental analysis of the plasma decay of the model organic anion bromosulphophthalein revealed that sinusoidal uptake and canalicular excretion of the dye were significantly decreased in Al (+) rats (-53 and -43%, respectively) . Expression of multidrug resistance-associated protein 2 (Mrp2), the main, multispecific transporter involved in the canalicular excretion of organic anions, was also decreased (-40%), which was associated with a significant decrease in the cumulative biliary excretion of the Mrp2 substrate, dinitrophenyl-S-glutathione (-50%) . These results show that chronic Al exposure leads to oxidative stress, cholestasis, and impairment of the hepatic handling of organic anions by decreasing both sinusoidal uptake and canalicular excretion . The alteration of the latter process seems to be causally related to impairment of Mrp2 expression . We have addressed some possible mechanisms involved in these deleterious effects.

Biochem Biophys Res Commun, 2004 Mar 12, 315(3), 719 - 25
Mapping of the MRPm5 epitope to the cytosolic region between transmembrane helices 13 and 14 in the drug and organic anion transporter, MRP1 (ABCC1); Koike K et al.; Multidrug resistance in human tumour cells is often associated with increased expression of the 190kDa multidrug resistance protein, MRP1, that belongs to the ATP-binding cassette superfamily of transport proteins . MRP1 is also an efficient transporter of many organic anions . In the present study, we have mapped the epitope of the MRP1-specific murine monoclonal antibody (MAb) MRPm5 to the decapeptide (1063)FFERTPSGNL(1072) located in the cytoplasmic loop (CL6) linking transmembrane helices 13 and 14 in the third membrane spanning domain of the protein . Several amino acids in the cytoplasmic loops of MRP1 have been reported to be important for its transport function; nevertheless, MAb MRPm5 does not inhibit vesicular uptake of the high affinity substrate leukotriene C(4) . None of the other MRP1-reactive MAbs described to date map to CL6 of MRP1 which in turn enhances the utility of MAb MRPm5 for both clinical and experimental investigations of this transporter.

Int J Tuberc Lung Dis, 2004 Jan, 8(1), 52 - 8
Identifying early treatment failure on category I therapy for pulmonary tuberculosis in Lima Ciudad, Peru; Chavez Pachas AM et al.; SETTING: Ambulatory, public tuberculosis treatment facilities, central Lima, Peru . OBJECTIVE: To identify risk factors for failure on directly observed Category I therapy . DESIGN: Case-control study . All failures of Category I (2HREZ/4H2R2) therapy in 2000 (2.9% of smear-positive TB patients) were included as cases; two controls per case were matched on health center and approximate time of treatment initiation . RESULTS: The study included 38 cases and 76 controls, all new smear-positive, pulmonary TB patients treated with Category I therapy in central Lima in 2000 . Neither treatment irregularity nor incidence of adverse events predicted failure in the study group . Mean baseline body mass index was lower in cases than in controls (P = 0.06) . Cases gained less weight during therapy (P = 0.01) . Smear positivity at 2 months of therapy was strongly associated with failure (OR 11.7; 95%CI 2.4-57.5) . No controls had positive smears at or after 3 months of therapy (OR {corrected} 144.9; 95%CI 8.4-2500) . Nearly 75% of cases with isolates tested for susceptibility to first-line drugs had multidrug-resistant TB (MDR-TB) . CONCLUSION: A large proportion of failures on Category I therapy can be identified early . As three-quarters of patients with susceptibility results have MDR-TB, early referral for culture and drug susceptibility testing is critical for prompt initiation of appropriate therapy and improved outcomes.

Int J Tuberc Lung Dis, 2004 Jan, 8(1), 31 - 8
Development of acquired drug resistance in recurrent tuberculosis patients with various previous treatment outcomes; Yoshiyama T et al.; SETTING: Chiang Rai province, Northern Thailand . OBJECTIVE: To study the probability of acquiring drug resistance to isoniazid (H) and rifampicin (R) on recurrence after treatment success, default and failure, among sputum smear-positive pulmonary tuberculosis (TB) patients treated with standardised short-course chemotherapy . DESIGN: Retrospective analysis of registration records of TB patients from May 1996 to December 2000 in Chiang Rai, where routine drug susceptibility testing (DST) is conducted for surveillance purposes . Patients registered twice or more were examined . RESULTS: Of 59 cases treated with HRZE/HR who underwent DST at the time of registration, 31 were fully susceptible to H and R at first registration, of whom four acquired drug resistance to H or R . Of 13 cases resistant to H or R at first registration, 11 became multidrug-resistant (MDR) . The remaining 15 patients were original MDR cases . Among 28 MDR or H- or R-resistant cases, six reverted from resistant to susceptible . DISCUSSION: A high proportion of patients with H- or R-resistant TB became MDR after treatment with 2HRZE/HR . Using this regimen, MDR may increase in a population with a high prevalence of H or R resistance . We are unable to explain why some drug-resistant cases became drug-susceptible . Further investigation is necessary.

Int J Tuberc Lung Dis, 2004 Jan, 8(1), 23 - 30
Drug resistance monitoring: combined rates may be the best indicator of programme performance; Van Deun A et al.; SETTING: Greater Mymensingh District, Bangladesh . OBJECTIVES: To determine changes in prevalence of drug resistance of Mycobacterium tuberculosis under DOTS . DESIGN: Drug susceptibility testing of systematic samples of M . tuberculosis isolated from all sputum smear-positive cases newly registered in sentinel centres during 1995 and 2001 . Continuous monitoring of retreatment registrations and resistance of strains from relapse and failure cases . RESULTS: Of 942 strains from the new cases in 2001, 10.8% showed resistance to any drug, 6.2% to isoniazid, 0.4% to rifampicin (all of them multidrug-resistant, MDR), 7.1% to streptomycin, and 1.0% to ethambutol . Corresponding rates for 99 strains from previously treated cases were 32%, 20%, 3%, 20% and 2%, respectively . Although most rates of resistance had decreased since 1995, increased streptomycin resistance was the only significant change when new and previously treated cases were considered separately . However, combined resistance for any drug, isoniazid, rifampicin and MDR had decreased significantly . CONCLUSION: As suggested by monitoring of resistance in failure and relapse cases and by routine programme reports, drug resistance had decreased . Combined resistance demonstrated changes between periodic surveys better than its subgroups, and may be a more reliable and comprehensive indicator . However, continuous monitoring of the pool of resistant retreatment cases is a more efficient strategy.

Int J Tuberc Lung Dis, 2004 Jan, 8(1), 15 - 22
Study of resistance to anti-tuberculosis drugs in five districts of Equatorial Guinea: rates, risk factors, genotyping of gene mutations and molecular epidemiology; Tudo G et al.; SETTING: Five districts in Equatorial Guinea, March 1999 to February 2001 . OBJECTIVES: To determine tuberculosis drug resistance among new and previously treated cases, the risk factors associated with resistance, and the mutations associated with isoniazid and rifampicin (katG, inhA and rpoB genes) resistance, and to genotype resistant strains . RESULTS: A positive culture identified as Mycobacterium tuberculosis complex was obtained in 240/499 patients . Susceptibility testing was performed in 236 strains . The overall resistance rate in new cases was 16.9% compared to 41.6% in previously treated cases . Isoniazid resistance was the most frequent (respectively 12.5% and 16.6%) in the two groups, while multidrug resistance was observed in 1.7% and 25% of new and previously treated cases, respectively . Female sex was statistically associated with resistance in new cases . Of 41 isoniazid-resistant strains, 33 (80.5%) had mutations in the inhA gene; none had mutations in the katG gene and eight had no mutations in either gene . All strains had low-level isoniazid resistance . Of eight strains resistant to rifampicin, six had mutations in the rpoB gene . Genotyping defined seven clusters . CONCLUSIONS: Moderate resistance was found in new cases . Low-level isoniazid resistance predominated among mutations in the inhA gene, with a high percentage of clustering in resistant strains.

Cancer Res, 2004 Feb 15, 64(4), 1242 - 6
Pheophorbide a is a specific probe for ABCG2 function and inhibition; Robey RW et al.; Pheophorbide a (PhA), a chlorophyll catabolite, was shown to be an ABCG2 substrate based on Abcg2(-/-) knockout mouse studies (J . W . Jonker et al., Proc . Natl . Acad . Sci . USA, 99: 15649-15654, 2002) . We developed a functional assay for ABCG2 using PhA and the ABCG2 inhibitor fumitremorgin C . In selected cell lines expressing high levels of P-glycoprotein, multidrug resistance-associated protein 1, or ABCG2, PhA transport was observed only in cells expressing ABCG2 . Fumitremorgin C-inhibitable PhA transport was found to correlate with cell surface ABCG2 expression as measured by the anti-ABCG2 antibody 5D3 . We found that 100 micro M of the cyclin-dependent kinase inhibitor UCN-01 or 1 micro M of the P-glycoprotein inhibitor tariquidar inhibited ABCG2-mediated PhA transport . In 4-day cytotoxicity assays, ABCG2-mediated resistance to SN-38 and topotecan was abrogated in ABCG2-transfected HEK-293 cells treated with 1 micro M tariquidar, and ABCG2-transfected cells were 6-7-fold resistant to UCN-01 . PhA is an ABCG2-specific substrate with potential value in measuring ABCG2 function and expression in clinical samples.

J Bacteriol, 2004 Mar, 186(5), 1423 - 9
Role of histone-like protein H-NS in multidrug resistance of Escherichia coli; Nishino K et al.; The histone-like protein H-NS is a major component of the bacterial nucleoid and plays a crucial role in global gene regulation of enteric bacteria . It is known that the expression of a variety of genes is repressed by H-NS, and mutations in hns result in various phenotypes, but the role of H-NS in the drug resistance of Escherichia coli has not been known . Here we present data showing that H-NS contributes to multidrug resistance by regulating the expression of multidrug exporter genes . Deletion of the hns gene from the DeltaacrAB mutant increased levels of resistance against antibiotics, antiseptics, dyes, and detergents . Decreased accumulation of ethidium bromide and rhodamine 6G in the hns mutant compared to that in the parental strain was observed, suggesting the increased expression of some drug exporter(s) in this mutant . The increased drug resistance and decreased drug accumulation caused by the hns deletion were completely suppressed by deletion of the multifunctional outer membrane channel gene tolC . At least eight drug exporter systems require TolC for their functions . Among these, increased expression of acrEF, mdtEF, and emrKY was observed in the Deltahns strain by quantitative real-time reverse transcription-PCR analysis . The Deltahns-mediated multidrug resistance pattern is quite similar to that caused by overproduction of the AcrEF exporter . Deletion of the acrEF gene greatly suppressed the level of Deltahns-mediated multidrug resistance . However, this strain still retained resistance to some compounds . The remainder of the multidrug resistance pattern was similar to that conferred by overproduction of the MdtEF exporter . Double deletion of the mdtEF and acrEF genes completely suppressed Deltahns-mediated multidrug resistance, indicating that Deltahns-mediated multidrug resistance is due to derepression of the acrEF and mdtEF drug exporter genes.

Mol Cell Biochem, 2004 Jan, 255(1-2), 11 - 8
Toxicokinetic and genomic analysis of chronic arsenic exposure in multidrug-resistance mdr1a/1b(-/-) double knockout mice; Xie Y et al.; Multidrug-resistance gene knockout mdr1a/1b(-/-) mice, which are deficient in P-glycoproteins, are more sensitive than wild-type (WT) mice to acute arsenic toxicity . This study assessed toxic manifestations of chronic oral arsenic in mdr1a/1b(-/-) mice, including oxidative stress and altered gene expression, and investigated altered toxicokinetics as a potential basis of enhanced arsenic toxicity . Thus, mdr1a/1b(-/-) and WT mice were exposed to sodium arsenite (0-80 ppm as arsenic) in the drinking water for 10 weeks at which time hepatic arsenic accumulation, lipid peroxidation (LPO), redox status and change in gene expression level were assessed . All mice survived the arsenic exposure, but body weight gain in the highest dose group was reduced in both mdr1a/1b(-/-) and WT mice . Arsenic induced pathological changes, elevated LPO levels and enhanced glutathione S-transferase (GST) activity, in the liver to a greater extent in mdr1a/1b(-/-) than in WT mice . Arsenic also decreased Cu/Zn superoxide dismutase activity in both mdr1a/1b(-/-) and WT mice . The expressions of certain genes, such as those encoding cell proliferation, GST, acute-phase proteins and metabolic enzymes, were modestly altered in arsenic-exposed mice . The expression of cyclin D1, a potential hepatic oncogene, was enhanced in arsenic-exposed mdr1a/1b(-/-) mice only . At the highest level of exposure, hepatic arsenic content was higher in mdr1a/1b(-/-) than in WT mice, suggesting that enhanced accumulation due to transport deficiency may, in part, account for the enhanced toxicity in these mice . In summary, this study shows that chronic arsenic toxicity, including liver pathology and oxidative stress, is enhanced in mdr1a/1b(-/-) mice, possibly due to enhanced accumulation of arsenic as a result of transport system deficiency.

Proc Natl Acad Sci U S A, 2004 Mar 2, 101(9), 2852 - 7 Epub 2004 Feb 17.
Structure of the multidrug resistance efflux transporter EmrE from Escherichia coli; Ma C et al.; Multidrug resistance efflux transporters threaten to reverse the progress treating infectious disease by extruding a wide range of drug and other cytotoxic compounds . One such drug transporter, EmrE, from the small multidrug resistance family, utilizes proton gradients as an energy source to drive substrate translocation . In an effort to understand the molecular structural basis of this transport mechanism, we have determined the structure of EmrE from Escherichia coli to 3.8 A . EmrE is a tetramer comprised of two conformational heterodimers related by a pseudo two-fold symmetry axis perpendicular to the cell membrane . Based on the structure and biochemical evidence, we propose a mechanism by which EmrE accomplishes multidrug efflux by coupling conformational changes between two heterodimers with proton gradient . Because of its simplicity and compact size, the structure of EmrE can serve as an ideal model for understanding the general structural basis of proton:drug antiport for other drug efflux systems.

Am J Physiol Renal Physiol, 2004 Jul, 287(1), F33 - 8 Epub 2004 Feb 17.
Involvement of guanylyl cyclase and cGMP in the regulation of Mrp2-mediated transport in the proximal tubule; Notenboom S et al.; In killifish renal proximal tubules, endothelin-1 (ET-1), acting through a basolateral ET(B) receptor, nitric oxide synthase (NOS), and PKC, decreases cell-to-lumen organic anion transport mediated by the multidrug resistance protein isoform 2 (Mrp2) . In the present study, we examined the roles of guanylyl cyclase and cGMP in ET signaling to Mrp2 . Using confocal microscopy and quantitative image analysis to measure Mrp2-mediated transport of the fluorescent drug fluorescein methotrexate (FL-MTX), we found that oxadiazole quinoxalin (ODQ), an inhibitor of NO-sensitive guanylyl cyclase, blocked ET-1 signaling . ODQ was also effective when signaling was initiated by nephrotoxicants (gentamicin, amikacin, diatrizoate, HgCl(2), and CdCl(2)), which appear to stimulate ET release from the tubules themselves . ODQ blocked the effects of the NO donor sodium nitroprusside but not of the phorbol ester that activates PKC . Exposing tubules to 8-bromo-cGMP (8-BrcGMP), a cell-permeable cGMP analog, decreased luminal FL-MTX accumulation . This effect was abolished by bisindoylmaleimide (BIM), a PKC inhibitor, but not by N(G)-methyl-l-arginine, a NOS inhibitor . Together, these data indicate that ET regulation of Mrp2 involves activation of guanylyl cyclase and generation of cGMP . Signaling by cGMP follows NO release and precedes PKC activation.

J Lipid Res, 2004 May, 45(5), 933 - 40 Epub 2004 Feb 16.
Oligonucleotides blocking glucosylceramide synthase expression selectively reverse drug resistance in cancer cells; Liu YY et al.; High glucosylceramide synthase (GCS) activity is one factor contributing to multidrug resistance (MDR) in breast cancer . Enforced GCS overexpression has been shown to disrupt ceramide-induced apoptosis and to confer resistance to doxorubicin . To examine whether GCS is a target for cancer therapy, we have designed and tested the effects of antisense oligodeoxyribonucleotides (ODNs) to GCS on gene expression and chemosensitivity in multidrug-resistant cancer cells . Here, we demonstrate that antisense GCS (asGCS) ODN-7 blocked cellular GCS expression and selectively increased the cytotoxicity of anticancer agents . Pretreatment with asGCS ODN-7 increased doxorubicin sensitivity by 17-fold in MCF-7-AdrR (doxorubicin-resistant) breast cancer cells and by 10-fold in A2780-AD (doxorubicin-resistant) ovarian cancer cells . In MCF-7 drug-sensitive breast cancer cells, asGCS ODN-7 only increased doxorubicin sensitivity by 3-fold, and it did not influence doxorubicin cytotoxicity in normal human mammary epithelial cells . asGCS ODN-7 was shown to be more efficient in reversing drug resistance than either the GCS chemical inhibitor d-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol or the P-glycoprotein blocking agents verapamil and cyclosporin A . Experiments defining drug transport and lipid metabolism parameters showed that asGCS ODN-7 overcomes drug resistance mainly by enhancing drug uptake and ceramide-induced apoptosis . This study demonstrates that a 20-mer asGCS oligonucleotide effectively reverses MDR in human cancer cells.

Chem Res Toxicol, 2004 Feb, 17(2), 158 - 64
Multidrug resistance-associated protein 2 (MRP2) enhances 4-hydroxynonenal-induced toxicity in Madin-Darby canine kidney II cells; Ji B et al.; 4-Hydroxy-trans-2,3-nonenal (HNE) is a toxic end product of lipid peroxidation . This multifunctional aldehyde reacts with proteins, phospholipids, and nucleic acids, consequently activating/inactivating enzymes, affecting signal transduction and gene expression . HNE is mainly detoxified by glutathione (GSH) conjugation . In our previous report, we showed that GSH conjugates of 4-hydroxynonenal (HNE-SG) are substrates of multidrug resistance-associated protein 2 (MRP2) . MRP2 has been shown to export HNE-SG conjugates into the extracellular space . In the present study, the role of MRP2 in the detoxification of HNE was studied using Madin-Darby canine kidney II (MDCK II) cells expressing human MRP2 . MRP2 reduced the intracellular accumulation of HNE-SG conjugate but unexpectedly increased the susceptibility of cells to HNE . The viability of cells was reduced to approximately 70% in the presence of 62.5 microM HNE in MDCK II cells expressing MRP2, whereas MDCK II cells remained unaffected . MRP2 accelerated the elimination of intracellular GSH via a conjugation reaction with HNE (half-life of GSH was 30.1 and 12.2 min for MDCK II cells and MDCK II cells expressing MRP2, respectively) . Moreover, the consumption of GSH was unlimited in MDCK II cells expressing MRP2, finally resulting in necrosis . These results indicate that MRP2 has an adverse effect during the detoxification of HNE in MDCK II cells and suggest that expression of MRP2 may enhance the damage caused by oxidative stress.

Neoplasia, 2003 Nov-Dec, 5(6), 475 - 80
Bypass of tumor drug resistance by antivascular therapy; Preise D et al.; Multidrug resistance (MDR) presents a major obstacle for the successful chemotherapy of cancer . Its emergence during chemotherapy is attributed to a selective process, which gives a growth advantage to MDR cells within the genetically unstable neoplastic cell population . The pleiotropic nature of clinical MDR poses a great difficulty for the development of treatment strategies that aim at blocking MDR at the tumor cell level . Targeting treatment to the nonmalignant vascular network-the lifeline of the tumor-is a promising alternative for the treatment of drug-resistant tumors . The present study demonstrates that MDR in cancer can be successfully circumvented by photodynamic therapy (PDT) using an antivascular treatment protocol . We show that, although P-glycoprotein-expressing human HT29/MDR colon carcinoma cells in culture are resistant to PDT with Pd-bacteriopheophorbide (TOOKAD), the same treatment induces tumor necrosis with equal efficacy (88% vs 82%) in HT29/MDR-derived xenografts and their wild type counterparts, respectively . These results are ascribed to the rapid antivascular effects of the treatment, supporting the hypothesis that MDR tumors can be successfully eradicated by indirect approaches that bypass their inherent drug resistance . We suggest that with progress in ongoing clinical trials, TOOKAD-PDT may offer a novel option for local treatment of MDR tumors.

Curr Pharm Des, 2004, 10(6), 647 - 57
Cyclooxygenase-2: potential role in regulation of drug efflux and multidrug resistance phenotype; Sorokin A; Multidrug resistance (MDR) of cancer cells to cytostatic agents is the major obstacle for the succesfull chemotherapy . One of the causes of the development of cellular resistance to a wide variety of drugs is the elevated expression of membrane transporter proteins such as members of ATP binding cassette (ABC) protein superfamily . Expression of the ABC transporter MDR1, also termed P-glycoprotein (P-gp), seems to correlate with drug resistance of tumors to chemotherapy . Cyclooxygenase-2, an inducible isoform of enzyme, responsible for generation of prostaglandins from arachidonic acid, is constitutively expressed in a number of cancer cells . Anti-cancer potency of cyclooxygenase inhibitors is established, but the mechanism of Cox-2-dependent potentiation of tumor growth is a subject of intense discussion . Here we focus on the discussion of potential link between Cox-2 expression and development of multidrug resistance phenotype . Our observation, that enforced expression of Cox-2 causes enhancement in MDR1 expression and functional activity suggests the existence of causal link between Cox-2 activity and MDR1 expression . The use of Cox-2 inhibitors to decrease function of MDR1 may enhance accumulation of chemotherapy agents and decrease resistance of tumors to chemotherapeutic drugs.

Curr Drug Metab, 2004 Feb, 5(1), 21 - 53
The MRP-related and BCRP/ABCG2 multidrug resistance proteins: biology, substrate specificity and regulation; Haimeur A et al.; Several members of different families of the ATP-binding cassette (ABC) superfamily of transport proteins are capable of transporting an extraordinarily structurally diverse array of endo- and xenobiotics and their metabolites across cell membranes . Together, these transporters play an important role in the absorption, disposition and elimination of these chemicals in the body . In tumor cells, increased expression of these drug transporters is associated with resistance to multiple chemotherapeutic agents . In this review, current knowledge of the biochemical, physiological and pharmacological properties of nine members of the multidrug resistance protein (MRP)-related ABCC family (MRP1, MRP2, MRP3, MRP4, MRP5, MRP6, MRP7, ABCC11 and ABCC12) as well as the G family member, ABCG2/BCRP, are summarized . A focus is placed on the structural similarities and differences of these drug transporters as well as the molecular determinants of their substrate specificities and transport activities . Factors that regulate expression of the MRP-related proteins and ABCG2/BCRP are also reviewed.

Curr Drug Metab, 2004 Feb, 5(1), 11 - 9
The role of MDR1 genetic polymorphisms in interindividual variability in P-glycoprotein expression and function; Woodahl EL et al.; The human multidrug resistance gene (MDR1), spanning greater than 200 kb, encodes for the ATP-dependent membrane efflux transporter, P-glycoprotein (Pgp) . Significant progress has been made in the discovery of MDR1 polymorphisms and the assessment of allelic frequencies . The search for key genetic determinants that predispose individuals to drugs that are substrates or inhibitors of Pgp has just begun . Reports in the literature, particularly focusing on the C3435T polymorphism, have provided discordant results with respect to functional modification in vitro, and Pgp expression and disposition of probe drugs in vivo . Due to the large size of the MDR1 gene, genotyping based on individual single nucleotide polymorphism (SNPs) analysis is not sufficient to predict functional consequences . Strong linkage disequilibrium has been detected between several MDR1 polymorphisms, and discrepancies in the literature may be due to the focus on the influence of single nucleotide variations instead of on linked nucleotide variations . Multiple SNPs found on the same chromosome are assigned to a specific haplotype, and some attempts have been made to determine the role of MDR1 haplotypes in Pgp variability . Most of the data for MDR1 haplotype have been predicted based on computational or mathematical models . However, molecular haplotyping techniques, analysis of linkages on the same chromosome directly by biophysical and biochemical means, may be needed to characterize haplotypes in individuals with a highly polymorphic and large gene like MDR1 . Haplotype identification may prove to be vital in identifying the functional significance of MDR1 polymorphisms on disease susceptibility and drug disposition.

Clin Nephrol, 2004 Jan, 61(1), 25 - 9
Mycophenolate mofetil in children with multidrug-resistant nephrotic syndrome; Bayazit AK et al.; AIM: The aim of the present study is to report our clinical experiences with MMF in problematic children with chronic glomerulonephritis resistant to corticosteroids and/or other immunosuppressive drugs . PATIENTS AND METHODS: Ten patients with chronic glomerulonephritis resistant to treatment with corticosteroids and other immunosuppressive drugs were treated with mycophenolate mofetil (MMF) . Causes of chronic glomerulonephritis were mesangial proliferative glomerulonephritis (4), membranoproliferative glomerulonephritis (3), chronic sclerosing glomerulonephritis (1), focal segmental glomerulosclerosis (1), diffuse endo- and extracapillary proliferative glomerulonephritis (1) . MMF 15 mg/kg was used in combination with low-dose corticosteroids and angiotensin-converting enzyme inhibitors . RESULTS: During 24 weeks of MMF therapy, no significant changes were detected in mean serum creatinine, albumin and proteinuria . Severe leukopenia was seen in 1 patient . Additional adverse effects, including nausea and diarrhea, were observed in another patient when the dosage was increased to 20 mg/kg per day . During MMF treatment proteinuria decreased slightly without remission in 6 of 10 patients . CONCLUSION: Further data and clinical trials are needed to evaluate the possible role of MMF in the treatment of chronic glomerulonephritis of similar etiologies in pediatric patients.

Blood, 2004 Jun 1, 103(11), 4276 - 84 Epub 2004 Feb 12.
The peripheral benzodiazepine receptor ligand PK11195 overcomes different resistance mechanisms to sensitize AML cells to gemtuzumab ozogamicin; Walter RB et al.; The antibody-targeted therapeutic, gemtuzumab ozogamicin (GO, Mylotarg), is approved for treatment of relapsed acute myeloid leukemia (AML) . We previously showed that AML blasts from GO refractory patients frequently express the drug transporters P-glycoprotein (Pgp) and/or multidrug resistance protein (MRP) . We also previously reported that inhibition of drug transport by the Pgp modulator, cyclosporine A (CSA), can increase GO sensitivity in Pgp(+) AML cells and that the peripheral benzodiazepine receptor ligand, PK11195, sensitizes AML cells to standard chemotherapeutics both by inhibiting Pgp-mediated efflux and by promoting mitochondrial apoptosis . We now show that PK11195 also can overcome multiple resistance mechanisms to increase GO sensitivity in AML cells, including resistance associated with expression of drug transporters and/or antiapoptotic proteins . PK11195 substantially increases GO cytotoxicity in AML cells from many different cell lines and primary patient samples, often more effectively than CSA . We also show that PK11195 is nontoxic in NOD/SCID mice and can sensitize xenografted human AML cells to GO . Since PK11195 is well tolerated in humans as a single agent, its further study as a multifunctional chemosensitizer for anti-AML therapies, including GO-based therapies, is warranted.

Eur J Cancer, 2004 Mar, 40(4), 606 - 13
Inhibition of multidrug resistance by immunisation with synthetic P-glycoprotein-derived peptides; Pawlak-Roblin C et al.; Overexpression of the membrane glycoprotein (P170) represents the most common multidrug resistance (MDR) mechanism in cancer therapy . Specific auto-antibodies to extracellular loops 1, 2 and 4 of murine P170 were elicited in mice using palmitoylated synthetic peptides reconstituted in liposomes, with or without Lipid A, and resuspended in alum . IgM antibodies were detected 14 days following the first injection and IgG1 became predominant after the third challenge . Animals did not show any auto-immune symptoms or induced toxicity up to 18 months after the immunisation . Previous immunisations of mice using liposomes with MDR1 peptides increases the efficacy of chemotherapy treatments with doxorubicin and vinblastine against P388 R cells with increase of 77% in the survival half time in the immunised group . Sera from the immunised mice were also effective in reducing cellular resistance to vinblastine and doxorubicin in vitro . Taken together, these data suggest that this immunisation approach might have potential clinical applications.

Lancet, 2004 Feb 7, 363(9407), 474 - 81
Programmes and principles in treatment of multidrug-resistant tuberculosis; Mukherjee JS et al.; Multidrug-resistant tuberculosis (MDR-TB) presents an increasing threat to global tuberculosis control . Many crucial management issues in MDR-TB treatment remain unanswered . We reviewed the existing scientific research on MDR-TB treatment, which consists entirely of retrospective cohort studies . Although direct comparisons of these studies are impossible, some insights can be gained: MDR-TB can and should be addressed therapeutically in resource-poor settings; starting of treatment early is crucial; aggressive treatment regimens and high-end dosing are recommended given the lower potency of second-line antituberculosis drugs; and strategies to improve treatment adherence, such as directly observed therapy, should be used . Opportunities to treat MDR-TB in developing countries are now possible through the Global Fund to Fight AIDS, TB, and Malaria, and the Green Light Committee for Access to Second-line Anti-tuberculosis Drugs . As treatment of MDR-TB becomes increasingly available in resource-poor areas, where it is needed most, further clinical and operational research is urgently needed to guide clinicians in the management of this disease.

Eur J Haematol, 2004 Jan, 72(1), 45 - 51
Significant co-expression of WT1 and MDR1 genes in acute myeloid leukemia patients at diagnosis; Galimberti S et al.; A high expression of Wilms' tumor gene (WT1) in acute myeloid leukemia (AML) seems to correlate with a poor outcome and its increased levels can be predictive of an impending relapse . WT1 has been shown in vitro to interact with the promoter of the MDR1, a gene involved in the multidrug resistance phenomenon . The aim of this study was to measure, by real-time polymerase chain reaction, levels of WT1 and MDR1 expression, in order to find a possible association between these genes, in a series of 50 newly diagnosed AML cases . Twenty-five percent of patients carried very high (>75 degrees percentile) MDR1- and 23.3%WT1-mRNA levels . Interestingly, high levels of WT1 were significantly correlated with correspondent high levels of MDR1 gene . Nevertheless, the co-expression of these genes did not significantly influence the complete response rate to the induction therapy . Reported data confirm the existence of a co-expression of WT1 and MDR1 genes even in vivo; this may be relevant because one consequence could be the positive selection by chemotherapeutic regimens of cells with higher MDR1 levels already present before treatment . Thus, the association between these genes could suggest avoiding the use of drugs involved in the multidrug resistance (MDR) phenomenon in patients carrying high levels of WT1 at diagnosis.

Leuk Lymphoma, 2003 Dec, 44(12), 2109 - 16
Physiological oxidants induce apoptosis and cell cycle arrest in a multidrug-resistant natural killer cell line, NK-YS; Than TA et al.; Natural-killer (NK) cell-derived malignant tumors, such as angiocentric lymphoma, is often resistant to various chemotherapeutic agents and follows an aggressive clinical course . We report the effects of physiological oxidants (hydrogen peroxide, H2O2; sodium hypochlorite, NaOCl and monochloramine, NH2Cl) on the cell growth and cell death in a multidrug-resistant NK tumor cell line, NK-YS . Among the oxidants tested, NH2Cl was most cytotoxic, in which more than 90% of the cells died at 150 nmol/1 x 10(6) cells . H2O2 was less cytotoxic, whereas NaOCl showed no significant cell death at this dose . The cell death induced by NH2Cl was accompanied by DNA cleavage and caspase activation, which suggested apoptosis . In addition, lower dose of NH2Cl (70 nmol/1 x 10(6) cells) retarded cell growth and inhibited the cell cycle transition from G1 to S . This cell cycle arrest accompanied a decrease in the phosphorylation of retinoblastoma tumor suppressor protein at serine 795 . These observations suggest that NH2Cl may induce apoptotic cell death and growth arrest in multidrug-resistant NK cell tumors.

J Biol Chem, 2004 Apr 23, 279(17), 17134 - 41 Epub 2004 Feb 09.
RNA helicase A in the MEF1 transcription factor complex up-regulates the MDR1 gene in multidrug-resistant cancer cells; Zhong X et al.; RNA helicase A (RHA) is a member of the DEAD/H family of RNA helicases and unwinds duplex RNA and DNA . Recent studies have shown that RHA regulates the activity of gene promoters . However, little information is available about the in vivo relevance of RHA in the regulation of natural genes . We previously characterized a nuclear protein (MEF1) that binds to the proximal promoter of the multidrug resistance gene (MDR1) and up-regulates the promoter activity . In the present study, we isolated and identified RHA as a component of the MEF1 complex by using DNA-affinity chromatography and mass spectrometry . The antibody against RHA specifically disrupted the complex formation in electrophoretic mobility shift assay, confirming the identity of RHA . Western blotting showed that RHA in drug-resistant cells had a higher molecular weight than that in drug-sensitive cells . Similar results were obtained when FLAG-tagged RHA was overexpressed in these cells . This size difference probably reflects posttranslational modification(s) of RHA in drug-resistant cells . Chromatin immunoprecipitation revealed that RHA occupies the MDR1 promoter in vivo . Overexpression of RHA enhanced expression of the MDR1 promoter/reporter construct and endogenous P-glycoprotein (P-gp), the MDR1 gene product, and increased drug resistance of drug-resistant cells but not the drug-sensitive counterpart . Introduction of short interfering RNA targeting the RHA gene sequence selectively knocked-down RHA expression and concomitantly reduced P-gp level . Thus, our study demonstrates, for the first time, the involvement of RHA in up-regulation of the MDR1 gene . Interactions of RHA with other protein factors in the MEF1 complex bound to the promoter element may contribute to P-gp overexpression and multidrug resistance phenotype in drug-resistant cancer cells.

Oncol Rep, 2004 Mar, 11(3), 641 - 5
Chemotherapeutic potential of plant alkaloids and multidrug resistance mechanisms in malignant fibrous histiocytoma of the heart; Reinecke P et al.; Primary malignant fibrous histiocytoma (MFH) of the heart is a rare and highly malignant soft tissue tumor, which is largely resistant to conventional chemotherapy and radiotherapy . Therefore, we analyzed growth inhibitory effects of different chemotherapeutic agents and mechanisms of drug resistance in the recently established cell line MFH-H derived from a human primary cardiac MFH . The growth inhibitory effects of etoposide, vincristine, and paclitaxel were tested using the MTT assay . The expression and function of multidrug resistance-related proteins, i.e . the P-glycoprotein, the multidrug resistance-associated protein (MRP) and the lung resistance-related protein (LRP) were determined by FACScan and functional assays of cellular drug efflux . The concentration required for a 50% inhibition of growth (IC50) was 0.001 microM for etoposide and 0.035 microM for vincristine . Paclitaxel dissolved in Cremophor EL/ethanol inhibited the cell growth of MFH-H cells more intensively (IC50: 0.27 microM) than paclitaxel dissolved in DMSO (IC50: 11.09 microM) suggesting that Cremophor EL is contributing to the inhibitory effects of paclitaxel . The response of MFH-H to etoposide, vincristine and paclitaxel/Taxol could not be predicted by the expression and function of P-glycoprotein, MRP and LRP . This study demonstrates that etoposide and to a lesser extent vincristine can effectively inhibit the growth of MFH-H cells, irrespective of the multidrug resistance phenotype . MFH-H cells are relatively insensitive to paclitaxel dissolved in DMSO, in contrast to paclitaxel dissolved in Cremophor EL/ethanol indicating that the diluent Cremophor contributes to the antiproliferative effects of the taxane paclitaxel.

J Biol Chem, 2004 Apr 30, 279(18), 18256 - 61 Epub 2004 Feb 06.
Cytoprotective effect of glucosylceramide synthase inhibition against daunorubicin-induced apoptosis in human leukemic cell lines; Grazide S et al.; Several studies have shown that ceramide (CER) glucosylation contributes to drug resistance in multidrug-resistant cells and that inhibition of glucosylceramide synthase sensitizes cells to various drug treatments . However, the role of glucosylceramide synthase has not been studied in drug-sensitive cancer cells . We have demonstrated previously that the anthracycline daunorubicin (DNR) rapidly induces interphasic apoptosis through neutral sphingomyelinase-mediated CER generation in human leukemic cell lines . We now report that inhibition of glucosylceramide synthase using d,l-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP) or 1-phenyl-2-palmitoylamino-3-morpholino-1-propanol (PPMP) protected U937 and HL-60 cells from DNR-induced apoptosis . Moreover, blocking CER glucosylation did not lead to increased CER levels but to increased CER galactosylation . We also observed that pretreating cells with galactosylceramide (GalCER) significantly inhibited DNR-induced apoptosis . Finally, we show that GalCER-enriched lymphoblast cells (Krabbe's disease) were significantly more resistant to DNR- and cytosine arabinoside-induced apoptosis as compared with normal lymphoblasts, whereas glucosylceramide-enriched cells (Gaucher's disease) were more sensitive . In conclusion, this study suggests that sphingomyelin-derived CER in itself is not a second messenger but rather a precursor of both an apoptosis second messenger (GD3) and an apoptosis "protector" (GalCER).

J Clin Microbiol, 2004 Feb, 42(2), 891 - 4
Genotypic and phenotypic characterization of drug-resistant Mycobacterium tuberculosis isolates from rural districts of the Western Cape Province of South Africa; Streicher EM et al.; Genotypic and phenotypic analysis of drug-resistant Mycobacterium tuberculosis isolates from the Western Cape Province of South Africa showed that drug resistance is widespread and recently transmitted . Multidrug-resistant (MDR) isolates comprise 40% of this collection, and a large pool of isoniazid monoresistance may be a future source of MDR tuberculosis.

J Biol Chem, 2004 Apr 23, 279(17), 17090 - 100 Epub 2004 Feb 06.
JNK regulates HIPK3 expression and promotes resistance to Fas-mediated apoptosis in DU 145 prostate carcinoma cells; Curtin JF et al.; Elevated endogenous JNK activity and resistance to Fas receptor-mediated apoptosis have recently been implicated in progression of prostate cancer and can promote resistance to apoptosis in response to chemotherapeutic drugs . In addition, JNK has been demonstrated to promote transformation of epithelial cells by increasing both proliferation and survival . Although numerous studies have reported a role for JNK in promoting Fas receptor-mediated apoptosis, there is a paucity in the literature studying the antiapoptotic function of JNK during Fas receptor-mediated apoptosis . Consequently, we have used the recently described specific JNK inhibitor SP600125 and RNA interference to inhibit endogenous JNK activity in the prostate carcinoma cell line DU 145 . We demonstrated that endogenous JNK activity increased the expression of a kinase, HIPK3, that has previously been implicated in multidrug resistance in a number of tumors . HIPK3 has also been reported to phosphorylate FADD . The interaction between FADD and caspase-8 was inhibited, but abrogation of JNK activity or HIPK3 expression was found to restore this interaction and increased the sensitivity of DU 145 cells to Fas receptor-mediated apoptosis . In conclusion, we present novel evidence that JNK regulates the expression of HIPK3 in prostate cancer cells, and this contributes to increased resistance to Fas receptor-mediated apoptosis by reducing the interaction between FADD and caspase-8.

Natl Med J India, 2003 Nov-Dec, 16(6), 321 - 7
Beyond DOtS: avenues ahead in the management of tuberculosis; Chaudhury RR et al.; India has almost 30% of the global burden of tuberculosis (TB)--one person dies of the disease every minute in our country . India has mounted the second-largest DOTS programme in the world to control this disease . However, DOTS has its limitations and newer approaches have been developed over the years to overcome the global burden of tuberculosis . Problems with health facilities, patients, drugs and the disease itself constitute some of the hurdles in the implementation of the DOTS programme . In an attempt to go beyond DOTS, the WHO launched the 'Stop TB Initiative' in 1988 . Against the background of irrational antituberculosis drug use, which contributes to increasing drug resistance, the effective involvement of private healthcare providers is imperative to achieve better geographical and patient coverage for the implementation of DOTS . The WHO is currently addressing the issue of involving private practitioners in tuberculosis control in a programme called Public-Private Mix DOTS (PPM DOTS) . The Stop TB Initiative is also active in the area of dual infection with HIV and tuberculosis, and the initiatives that have been taken in this area include 'ProTEST', community contribution to tuberculosis care, and development and dissemination of training materials and guidelines . The DOTS-Plus strategy for the management of multidrug resistant (MDR)-TB and the establishment of the Green Light Committee to review project applications in this area are initiatives taken to curb the problem of drug resistance in tuberculosis . Even decades after the introduction of the DOTS strategy, much needs to be done to expand the services to the entire population; it is now essential to develop strategies that go beyond DOTS.

Planta Med, 2004 Jan, 70(1), 81 - 4
Pubescenes, jatrophane diterpenes, from Euphorbia pubescens, with multidrug resistance reversing activity on mouse lymphoma cells; Valente C et al.; The macrocyclic jatrophane diterpene polyesters, pubescenes A - D ( 1 - 4) were isolated from the whole dried plant of Euphorbia pubescens, and evaluated for multidrug resistance (MDR) reversing activity on mouse lymphoma cells . All the compounds displayed very strong activity compared with the positive control verapamil . Pubescene D ( 4) is a new compound, whose structure was established as 3beta,9alpha,-diacetoxy-7beta-benzoyloxy-15beta-hydroxy-14-oxo-2beta H-jatropha-5 E,12 E-diene by spectroscopic methods, including (1)H- (1)H COSY, HMQC, HMBC and NOESY.

Planta Med, 2004 Jan, 70(1), 45 - 9
Rearranged jatrophane-type diterpenes from euphorbia species . Evaluation of their effects on the reversal of multidrug resistance; Madureira AM et al.; The rearranged jatrophane-type diterpenes ( 1 - 3), isolated from the Me (2)CO extracts of Euphorbia portlandica and Euphorbia segetalis, were examined for their effects on multidrug resistance (MDR) in mouse lymphoma cells . Compounds 2 and 3 revealed to be active with the latter being more active than the positive control verapamil, a known resistance modifier . The new compound 1, named portlandicine, was isolated from Euphorbia portlandica and its structure characterised by high-field NMR spectroscopic methods including 2D NMR techniques: COSY, HMQC, HMBC and NOESY . The known diterpene 2, together with aleuritolic acid ( 4), oleanolic acid ( 5), and betulin diacetate ( 6), were also isolated from the same species.

JPEN J Parenter Enteral Nutr, 2004 Jan-Feb, 28(1), 1 - 6
Hepatic P-glycoprotein changes with total parenteral nutrition administration; Tazuke Y et al.; BACKGROUND: The mechanism(s) responsible for the development of parenteral nutrition-associated liver disease (PNALD) is unknown . Recently, a number of bile canalicular transport proteins have been identified that transport bile components out of hepatocytes . One group of these genes, multidrug resistance 1 (mdr1) and mdr2, encode P-glycoproteins . Mice lacking mdr2 expression develop liver disease that appears similar to PNALD . This study investigated the alteration in the expression of these transport proteins during the administration of total parenteral nutrition (TPN) . METHODS: Mice received either physiologic saline and standard chow or TPN . Mice were sacrificed on day 7, and hepatic DNA and RNA content, mRNA expression, and levels of mdr1 and mdr2 proteins were measured . RESULTS: TPN administration led to a significant (p < .05) decline in mdr2 mRNA expression and an increase in mdr1 mRNA expression . Mdr2 protein expression declined by 66% in the TPN-treated group, and mdr1 protein expression significantly increased by 58% . Histology and biochemical parameters showed no evidence of liver injury . Serum bile acid levels were elevated in the TPN group, suggesting the development of early cholestasis . CONCLUSIONS: The decline in mdr2 and rise in mdr1 mRNA and protein expression with TPN administration occurred before the development of liver injury but during an early state of cholestasis . This suggests that alterations in mdr gene expression may be a causative factor in the development of PNALD.

Biosci Rep, 2003 Aug, 23(4), 199 - 212
Mitotracker green is a P-glycoprotein substrate; Marques-Santos LF et al.; P-glycoprotein has a widespread expression on normal tissues . The protein has also been strongly associated with the multidrug resistance phenotype (MDR) on tumor cells . The employment of flow cytometry and confocal microscopy has contributed to the discovery and application of new particular fluorescent dyes . Nevertheless, several studies are being performed in different cellular types neglecting the expression/activity of MDR proteins . Because many fluorochromes have been reported as P-glycoprotein substrates, an especial attention must be given to the properties of new dyes in the presence of MDR proteins . Flow cytometric analyzes of Mitotracker Green (MTG) fluorescence profile were performed in a human erythroleukemic cell line and its resistant counterpart . In this report we demonstrated that MTG, a probe used to evaluate the mitochondrial mass, is a P-glycoprotein substrate and its staining profile is dependent on the activity of this protein . In vitro studies on a human erythroleukemic cell line and its resistant counterpart revealed that MDR modulators (Cyclosporin A, Verapamil, and Trifluoperazine) alter the MTG fluorescence pattern on a resistant cell line . The findings suggest that attention should be given to the expression of P-glycoprotein when performing an evaluation of mitochondria properties with MTG.

Curr Opin Investig Drugs, 2003 Dec, 4(12), 1416 - 21
Multidrug resistance in cancer therapy: role of the microenvironment; Galmarini CM et al.; Despite advances in the design of chemotherapeutic agents, and although many of these effective agents are now available for clinical use, most human cancers are resistant to therapy at presentation or become resistant after an initial response . The effectiveness of chemotherapy is compromised by several microenvironmental factors that affect the bioavailability and efficacy of chemotherapeutic agents . These factors vary from one tumor to another, and from one location to another, within the same tumor . A better comprehension of microenvironmental multidrug resistance mechanisms would lead to a clearer understanding of the reason for chemotherapeutic failure . This improved knowledge will permit a more rational utilization of chemotherapy.

J Pharmacol Exp Ther, 2004 Jun, 309(3), 1221 - 9 Epub 2004 Feb 04.
Multidrug resistance protein (MRP) 4- and MRP 5-mediated efflux of 9-(2-phosphonylmethoxyethyl)adenine by microglia; Dallas S et al.; The pathogenesis of human immunodeficiency virus (HIV)-associated dementia has been linked to microglial responses after infection . We have recently confirmed expression of several ATP-dependent efflux transporters in microglia, namely, multidrug resistance protein 1 (MRP1) and P-glycoprotein (P-gp) . In the present study, we investigated whether cultured rat microglia express two additional MRP family members, rMRP4 and rMRP5 . Using reverse transcriptase-polymerase chain reaction, rMRP4 and rMRP5 mRNA was detected in primary cultures of microglia and in a rat microglia cell line, MLS-9 . Western blot analysis further confirmed protein expression of the two MRP isoforms in MLS-9 cells . Bis(pivaloxymethyl)-9-(2-phosphonylmethoxyethyl)adenine {bis(POM)PMEA}, a lipophilic ester prodrug of the well characterized MRP4 and 5 substrate 9-(2-phosphonylmethoxyethyl)adenine (PMEA), was chosen to examine transport characteristics in MLS-9 . Using thin layer chromatography, we verified that more than 90% of radioactivity recovered in MLS-9 loaded with 1 microM {(3)H}bis(POM)PMEA for 1 h under ATP-depleting conditions was converted to PMEA . Efflux of PMEA by MLS-9 cell monolayers was ATP-dependent, glutathione-independent, and significantly inhibited by several MRP inhibitors (i.e., sulfinpyrazone, genistein, indomethacin, and probenecid) as well as the antiretroviral drug azidothymidine-monophosphate . Similar results were not observed in MRP1- or P-gp-overexpressing cell lines, suggesting that PMEA is not a substrate for either P-gp or MRP1 . These studies provide further evidence that microglia express multiple subfamilies of ATP-binding cassette transporters (i.e., P-gp, MRP1, MRP4, and MRP5) that could restrict permeation of several different classes of antiretroviral drugs in a brain cellular target of HIV-1 infection.

Zhonghua Xue Ye Xue Za Zhi, 2003 Dec, 24(12), 617 - 20
{CIK cells acquired multidrug resistance and maintained cytotoxic activity to tumor cells after mdr1 gene transfection}; Yang YH et al.; OBJECTIVE: To investigate whether the cytokine-induced killer (CIK) cells could acquire multidrug resistance and maintain the original cytotoxic activity after multidrug resistance (mdr1) genes transfection . METHODS: CIK cells were generated from peripheral blood cultured with IFN-gamma, CD(3) monoclonal antibody, IL-2, IL-1 and transfected with a plasmid (pHamdr) containing human mdr1 gene via electroporation . RT-PCR method was used to assay mRNA expression of mdr1 gene in transfected CIK cells, flow cytometry with anti-P-gp monoclonal antibody to detect P-glycoprotein (P-gp) expression on CIK cells membrane, and MTT assay to compare both the multidrug resistance to doxorubicin and colchicines and cytotoxic activity to human mammary cancer cell line MCF7 between transfected and non-transfected CIK cells . RESULTS: mdr1 expression was detected in the transfected CIK cells . There was a strong expression of P-gp on the transfected CIK cells and the percentages of P-gp positive cells were 21% - 37% (average 27%) . The IC(50) of transfected CIK cells to doxorubicin was 22.3 - 45.8 times and 6.7 - 11.35 times to colchicines of those of non-transfected CIK cells . The cytotoxic activity to MCF7 remained unchanged (P > 0.05) . CONCLUSION: It demonstrated that CIK cells transfected with mdr1 gene via electroporation could express multidrug resistance successfully without changes of cytotoxic activity.

Zhonghua Bing Li Xue Za Zhi, 2003 Dec, 32(6), 563 - 6
{Reversal of multidrug resistance property of carcinoma cells by down-regulating transcription of mdr-1}; Gao P et al.; OBJECTIVE: To reverse the multidrug resistance (MDR) property of carcinoma cells by blocking transcription of activating sites of mdr-1 . METHODS: Breast carcinoma cells were transinfected with several antisense oligonucleotide (ASODN) complementary to mdr-1 by lipofectin . RT-PCR was used to detect the production of mdr-1mRNA . The expression of P-glycoprotein (gp) was then detected by immunohistochemistry and the function of P-gp was detected by rhodamine123 retention . RESULTS: Forty-eight hours after transfection, mdr-1 index of cells treated by ASODN complementary to MA zone (major initiation start zone), MI (minor initiation start zone), C zone (CAAT box), G zone (GC box) of mdr-1 gene was 1.4, 1.9, 1.6 and 2.1 respectively . The rate of P-gp protein expression in treated cells was 14%, 43%, 26% and 39% respectively . The intracellular Rh123 retention in treated cells was 125%, 83%, 102% and 77% respectively . There was significant difference between cells treated by ASODN complementary to MA zone and C zone and drug-resistant cells . CONCLUSIONS: The ASODN complementary to MA zone and C zone of mdr-1 gene can reverse MDR of drug-resistant cells to various extent, amongst which the former is more effective . Down-regulating transcription of mdr-1 by blocking transcription activating sites can reduce the expression of mdr-1mRNA and P-gp, and thus reversing MDR of carcinoma cells . The ASODN complementary to MI zone, G zone of mdr-1 however do not significantly reverse the MDR property.

J Med Chem, 2004 Feb 12, 47(4), 988 - 92
Jatrophane diterpenes as modulators of multidrug resistance . Advances of structure-activity relationships and discovery of the potent lead pepluanin A; Corea G et al.; From the whole plant of Euphorbia peplus L., five new diterpenes based on a jatrophane skeleton (pepluanins A-E, 1-5) were isolated, together with two known analogues (6 and 7), which served to divulge in detail the structure-activity relationships within this class of P-glycoprotein inhibitors . The results revealed the importance of substitutions on the medium-sized ring (carbons 8, 9, 14, and 15) . In particular, the activity is collapsed by the presence of a free hydroxyl at C-8, while it increases with a carbonyl at C-14, an acetoxyl at C-9, and a free hydroxyl at C-15 . The most potent compound of the series, pepluanin A, showed a very high activity for a jatrophane diterpene, outperforming cyclosporin A by a factor of at least 2 in the inhibition of Pgp-mediated daunomycin transport.

Eur J Pharm Sci, 2004 Feb, 21(2-3), 283 - 93
Reversal of P-glycoprotein mediated vincristine resistance of L1210/VCR cells by analogues of pentoxifylline . A QSAR study; Kupsakova I et al.; In our previous papers we described the ability of methylxanthine pentoxifylline (PTX) to depress the P-glycoprotein (P-gp) mediated multidrug resistance (MDR) of the mouse leukemic cell line L1210/VCR . Other methylxanthines like caffeine and theophylline were found to be ineffective in this respect . In the present paper we have analysed the capability of 25 methylxanthines to depress MDR of L1210/VCR cells . These methylxanthines structurally differ in substituents located in positions N1, N3, N7 and C8 . The results indicate that for an effective reversal of P-gp mediated MDR of our cells the existence of a longer polar substituent in the position N1 plays a crucial role . The elongation of the substituent in the positions N3 and N7 (from methyl to propyl) increases and in the position C8 (from H to propyl) decreases the efficacy of xanthines to reverse the vincristine resistance of L1210/VCR cells . The multiple linear regression for effectiveness of methylxanthines in reversal of P-gp mediated MDR of L1210/VCR cells (expressed as respective IC(50r) values) has been computed, with molar weight: M(w), molar volume: V(M), molar refractivity: R(M), crystal density: d and partition coefficient n-octanol/water: logP as descriptors . A high intercorrelation of M(W), V(M) and R(M) was found for the tested group of methylxanthines indicating that only one of these parameters is necessary for testing a potential correlation . The best fit in the multiple linear regression was obtained for R(M) applied together with d and logP and resulted in a QSAR model given by the following equation: IC(50r)=-{(32.3+/-7.2)x10(-3)xR(M)}+{(10.1+/-2.3)xd}+{(0.74+/-0.10)xlogP}-{10.5+/-3.2} . Model revealed that: (i) the molar refractivity influences the effectiveness of xanthine positively; (ii) the crystal density and partition coefficient influence the MDR reversal effectiveness of xanthine negatively.

Eur J Pharm Sci, 2004 Feb, 21(2-3), 243 - 50
Membrane effects of the antitumor drugs doxorubicin and thaliblastine: comparison to multidrug resistance modulators verapamil and trans-flupentixol; Pajeva I et al.; The interactions of the antitumor drugs doxorubicin and thaliblastine with model membranes composed of neutral (phosphatidylcholine) and negatively charged (phosphatidylserine) phospholipids were studied by differential scanning calorimetry and nuclear magnetic resonance . The membrane activities of doxorubicin and thaliblastine were compared to those of the powerful multidrug resistance (MDR) modulators trans-flupentixol and verapamil . The results point out to the potential role of the drug-membrane interactions for the effects of doxorubicin and thaliblastine in resistant tumor cells . They direct also to the artificial membranes as a suitable tool for screening of compounds with potential ability to modulate MDR.

Biochim Biophys Acta, 2004 Jan 28, 1660(1-2), 155 - 63
Fluorescent modified phosphatidylcholine floppase activity of reconstituted multidrug resistance-associated protein MRP1; Huang Z et al.; Multidrug resistance-associated protein (MRP1) may function as a floppase in human red blood cells to translocate phosphatidylserine and/or phosphatidylcholine from inner membrane leaflet to outer leaflet . Here we report that the purified and reconstituted MRP1 protein into asolectin proteoliposomes is mainly in an inside-out configuration and possesses the ability to flop a fluorescent labeled phosphatidylcholine (NBD-PC) from outer leaflet (protoplasmic) to inner leaflet (extracytoplasmic) . The reconstituted MRP1 protein retains endogenous ATPase activity . ATP hydrolysis is required for the flopping since removal of ATP and/or Mg2+ inhibits the translocation of NBD-PC . Further evidence to support this conclusion is that the translocation of NBD-PC is inhibited by vanadate, which traps ATP hydrolysis product ADP in the nucleotide binding domains . In addition, the translocation of NBD-PC by proteoliposomes containing MRP1 protein is in a glutathione-dependent manner, similar to the process of translocating anticancer drugs such as daunorubicin . verapamil, vincristine, vinblastine, doxorubicin and oxidized glutathione partially inhibited the translocation of NBD-PC, whereas MK 571, an inhibitor of MRP1 protein, inhibited the translocation almost completely . Taken together, the purified and reconstituted MRP1 protein possesses the ability to flop NBD-PC from outer to inner leaflet of the proteoliposomes.

Biochem J, 2004 Jun 1, 380(Pt 2), 549 - 60
Nucleotides and transported substrates modulate different steps of the ATPase catalytic cycle of MRP1 multidrug transporter; Kern A et al.; The human ABC (ATP-binding cassette) transporter MRP1 (human multidrug-resistance-associated protein 1; ABCC1) is involved in the cellular extrusion of conjugated metabolites and causes multidrug resistance in tumour cells . The transport of substrate molecules by ABC proteins is energized by ATP hydrolysis, performed by two co-operating ABC units . Orthovanadate (Vi), a non-covalent inhibitor of the ABC ATPases, was found to catalyse a photo-oxidative cleavage of various ATP-binding proteins . In the present study, we have identified three Vi-cleavage sites within MRP1, and found that the cleavage reactions were variably modulated by the presence of nucleotides and by transported substrates . We concluded that Vi cleavage of MRP1 at Site I detects conformational changes due to the binding of MgATP . In contrast, Site II could be identified as part of the substrate-modulated catalytic cycle, probably containing an MRP1.MgADP.Vi transition-state-like complex . Cleavage at Site III was modulated by both the binding and hydrolysis of MgATP, in a biphasic pattern, which was also affected by the presence of transported substrates . We detected two different allosteric effects and found that they control two consecutive steps of the MRP1 ATPase catalytic cycle . Nucleotide binding to the low-affinity site accelerated the formation of the pre-hydrolytic intermediate in the other catalytic centre . Interaction of the transporter with its transported substrates stimulated a later reaction of the hydrolytic cycle, the formation of the post-hydrolytic intermediate, which could be detected in both catalytic sites by the experimental strategy used.

SAR QSAR Environ Res, 2003 Oct-Dec, 14(5-6), 447 - 54
QSAR studies on P-glycoprotein-regulated multidrug resistance and on its reversal by phenothiazines; Dearden JC et al.; Multidrug resistance is brought about largely by membrane transport proteins such as P-glycoprotein (P-gp) . We have developed a quantitative structure-activity relationship (QSAR) for P-gp-associated ATPase activity for a diverse set of 22 drugs, and found that such activity is related to substrate molecular size and polarity . We have also developed a QSAR for drug efflux from the blood-brain barrier of another diverse set of 22 drugs, and found that such efflux is a function of drug size and polarisability . Thirdly, we have carried out a QSAR analysis of the ability of 157 phenothiazines and related drugs to reverse multidrug resistance . We were unable to obtain a good QSAR for the whole data-set, but when we divided the data-set into sub-sets of closely related structures, a series of good correlations was obtained, most of which incorporated descriptors that model molecular size and polarity/polarisability . In no instance did we find any evidence that hydrogen bonding or hydrophobicity play a part in multidrug resistance or its reversal, despite that fact that several other workers have reported that these effects appear to be important here.

Lasers Surg Med, 2004, 34(1), 62 - 72
Effect of 5-aminolevulinic acid-mediated photodynamic therapy on MCF-7 and MCF-7/ADR cells; Tsai T et al.; BACKGROUND AND OBJECTIVES: Photodynamic therapy (PDT) has been proposed as an alternative approach in overcoming multidrug resistance (MDR) phenotype . To verify whether 5-aminolevulinic acid (ALA)-mediated PDT is effective in MDR cells, we studied the protoporphyrin IX (PpIX) content, intracellular localization, and phototoxicity in human breast cancer cells MCF-7 and derived MDR subline, MCF-7/ADR . STUDY DESIGN/MATERIALS AND METHODS: The fluorescence kinetics of ALA-induced PpIX was evaluated by spectrofluorometer . The phototoxicity of MCF-7 and MCF-7/ADR cells was determined by tetrazolium (MTT) assays and clonogenic assay . Furthermore, Annexin V and propidium iodide (PI) binding assays were performed to analyze the characteristics of cell death after ALA-PDT . RESULTS: MCF-7/ADR accumulated a lower level of PpIX as compared to parental MCF-7 cells . Significant phototoxicity was observed in MCF-7 and increased in a fluence-dependent manner with LD(50) around 8 J/cm(2) . Compared to its parental counterpart, MCF-7/ADR cells were less sensitive to ALA photodynamic treatment and PDT-induced cytotoxicity did not increase in a dose responsive manner as the concentration of ALA increased or the fluence of light increased . ALA-PDT was less effective for MCF-7/ADR cells than MCF-7 cells even under the condition when these two cell lines contained the similar amounts of PpIX . CONCLUSIONS: These results indicate that, except for the MDR related characteristics, MCF-7/ADR cells might possess intrinsic mechanisms that render them less sensitive to ALA-PDT induced phototoxicity .

Neuroendocrinology, 2004 Jan, 79(1), 1 - 12
Hypothalamic-pituitary-adrenocortical system and mood disorders: highlights from mutant mice; Muller MB et al.; In recent years, refined molecular technologies and the generation of genetically engineered mice have allowed to specifically target individual genes involved in the regulation of the hypothalamic-pituitary-adrenocortical (HPA) system . Given the fundamental role of the corticotropin-releasing hormone (CRH) system in anxiety, stress-associated pathologies, and mood disorders, we describe genetic modifications of the genes that encode proteins integral to the CRH/CRH receptor system with particular emphasis on conditional gene-targeting strategies . The profile of results, consistent with current knowledge of CRH function from more traditional assays, indicates that enhancement of the CRH function is associated with an activation of the HPA system, an anxious phenotype, alterations in cognitive performance, reductions in food intake, and disturbances of autonomic functions . In general, blockade of CRH activity produces the opposite effects, namely an anxiety-reduced phenotype . Molecular genetic strategies for conditional inactivation or overexpression of the glucocorticoid receptor contribute to our understanding of the genetics of endocrine activity and behavior, the most complex form of biological organization . In addition, we introduce mice with a genetic manipulation in the function of the blood-brain barrier as an animal model for the study of neuroendocrine regulation and, in particular, of HPA system activity . By use of mice deficient for abcb1- (also called multidrug resistance gene 1, mdr1-) type P glycoproteins, it was shown most recently that abcb1-type P glycoproteins control the access of endogenous glucocorticoids into the central nervous system . Thus, the ABCB1-type P glycoprotein function exerts a profound influence on activity and regulation of the HPA system under both basal conditions and during stress . Taken together, these genetically engineered mice are valuable tools for increasing our understanding of HPA system dysregulation in anxiety and stress-related pathologies, including human affective disorders . The identification and detailed characterization of these molecular pathways will ultimately lead to the development of novel neuropharmacological intervention strategies .

Proc Natl Acad Sci U S A, 2004 Feb 10, 101(6), 1702 - 7 Epub 2004 Jan 30.
Imaging reversal of multidrug resistance in living mice with bioluminescence: MDR1 P-glycoprotein transports coelenterazine; Pichler A et al.; Coelenterazine is widely distributed among marine organisms, producing bioluminescence by calcium-insensitive oxidation mediated by Renilla luciferase (Rluc) and calcium-dependent oxidation mediated by the photoprotein aequorin . Despite its abundance in nature and wide use of both proteins as reporters of gene expression and signal transduction, little is known about mechanisms of coelenterazine transport and cell permeation . Interestingly, coelenterazine analogues share structural and physiochemical properties of compounds transported by the multidrug resistance MDR1 P-glycoprotein (Pgp) . Herein, we report that living cells stably transfected with a codon-humanized Rluc show coelenterazine-mediated bioluminescence in a highly MDR1 Pgp-modulated manner . In Pgp-expressing Rluc cells, low baseline bioluminescence could be fully enhanced (reversed) to non-Pgp matched control levels with potent and selective Pgp inhibitors . Therefore, using coelenterazine and noninvasive bioluminescence imaging in vivo, we could directly monitor tumor-specific Pgp transport inhibition in living mice . While enabling molecular imaging and high-throughput screening of drug resistance pathways, these data also raise concern for the indiscriminate use of Rluc and aequorin as reporters in intact cells or transgenic animals, wherein Pgp-mediated alterations in coelenterazine permeability may impact results.

Haematologica, 2004 Jan, 89(1), 34 - 41
P-glycoprotein and multidrug resistance associated protein-1 activity in 132 acute myeloid leukemias according to FAB subtypes and cytogenetics risk groups; Legrand O et al.; BACKGROUND AND OBJECTIVES: We studied the function of both Pgp and MRP1 to identify subgroups of patients who could benefit from Pgp reversion, and to clarify in different FAB subtypes and in cytogenetic risk groups their expression and function . DESIGN AND METHODS: We examined 132 adults with de novo acute myeloid leukemia (AML) for Pgp and MRP1 expression and function . We correlated our finding with the FAB subtypes and the cytogenetics, and clinical data of our patients . RESULTS: Among FAB subtypes and cytogenetic subgroups, patients with good risk cytogenetics have a low expression and activity of Pgp and MRP1 except patients with inv(16) who have a higher activity of MRP1 than t(8;21) and t(15;17) (p=0.05) . All other AML patients, except M5, have a high expression and activity of Pgp . In contrast, M5 have a high expression, but a low activity of Pgp . In this subgroup, M5 with MLL gene rearrangement did not express an active Pgp . Others patients with M5 AML did not have a functional Pgp . Monosomy 7, 11q2.3 gene rearrangement and complex cytogenetic have a higher activity of MRP1 than other cytogenetic (p=0.03) . INTERPRETATION AND CONCLUSIONS: Resistance mechanism in M5 was not mediated by Pgp . In contrast, MRP1 may play a role in patients who have a 11q2.3 gene rearrangement, or in M4E with inv(16) . Thus trials that modulate Pgp are likely to achieve limited success in AML with low activity of Pgp, i.e., M5 and, AML with good risk cytogenetics.

Ann Trop Med Parasitol, 2003 Dec, 97(8), 783 - 91
Therapeutic efficacies of antimalarial drugs in the treatment of uncomplicated, Plasmodium falciparum malaria in Assam, north-eastern India; Dev V et al.; In the Indian state of Assam, the current therapeutic efficacies of the drugs commonly used in the area for the treatment of uncomplicated, Plasmodium falciparum malaria were investigated . As is routine in this area, subjects found positive for P . falciparum malaria were initially treated with chloroquine (CQ) . They were given sulfadoxine-pyrimethamine (SP) if this treatment failed, and subsequently quinine if the SP failed . The protocol of the World Health Organization's extended in-vivo test was used to follow parasite clearance and clinical cure . Therapeutic response was assessed by comparing the baseline (day-0) level of parasitaemia with that observed on day 3 . Many (75.7%) of the 144 evaluable subjects were treatment successes after CQ, but six early (4.2%) and 29 (20.1%) late CQ-treatment failures were observed . Of the 34 CQ-treatment failures followed, 31 (91.2%) responded adequately to SP but the other three were early (one) or late (two) SP-treatment failures . Two (66.7%) of the SP-treatment failures responded adequately to parenteral quinine but the other (a late quinine-treatment failure) had to be given an artemisinin derivative to achieve a clinical cure . The foci in which multidrug-resistant cases of malaria are developing in India need to be identified quickly, so that such cases can be cured before the mutant strains of P . falciparum that are resistant to several drugs have a chance to become more widespread.

Curr Top Med Chem, 2004, 4(2), 203 - 17
3D QSAR models of interactions between beta-tubulin and microtubule stabilizing antimitotic agents (MSAA): a survey on taxanes and epothilones; Manetti F et al.; In the last two decades, paclitaxel (Taxol), 1) has dominated the anticancer chemotherapy as one of the most important antimitotic agents . Despite its clinical success, it presents some limitations due to its low aqueous solubility or multidrug-resistance (MDR) susceptibility . Among new compounds sharing paclitaxel's mechanism of action, epothilones have emerged as very promising candidates and are currently under clinical trials . While the electron crystallography (EC) structure of tubulin with embedded paclitaxel is available, only hypotheses about epothilone binding upon the protein may be advanced . This review illustrates our efforts in the minireceptor modeling approach as the most recent advances in the field of three-dimensional quantitative structure-activity relationship (3D QSAR) studies involving taxanes, epothilones and the corresponding protein environment.

Curr Drug Targets Infect Disord, 2003 Dec, 3(4), 329 - 44
Mathematical approaches in the study of viral kinetics and drug resistance in HIV-1 infection; Muller V et al.; We review some crucial aspects of drug therapy and viral resistance that have been investigated within the framework developed for the modelling of virus kinetics . First, we give a general overview on the use of mathematical models in the field of HIV research . We seek to identify the factors that determine the steady state virus load and show that stable reductions during antiviral therapy are difficult to explain within the standard model of virus dynamics . We discuss possible extensions that enable the models to account for the moderately reduced virus loads during non-suppressive treatment and argue that the residual viremia under suppressive treatment can probably be attributed to the survival of long-lived infected cells, rather than to new rounds of replication . Next, we address the emergence of resistance during suppressive therapy and demonstrate that the resistant virus is more likely to be present already at the start of treatment than to be generated during therapy . The appearance of resistance after a prolonged period of initial suppression indicates that drug efficacy is not continuously maintained over time . We investigate the potential risks and benefits of therapy interruptions . Considering the effect of recombination, we argue that it probably decelerates, rather than accelerates the evolution of multidrug-resistant virus . We also review state-of-the-art methods for the estimation of fitness, which is crucial to the understanding of the emergence of resistance during therapy or the re-emergence of wild type upon the cessation of therapy.

Curr Drug Targets Infect Disord, 2003 Dec, 3(4), 273 - 81
A review of HIV-1 resistance to the nucleoside and nucleotide inhibitors; Shulman N et al.; The nucleoside reverse transcriptase inhibitors (NRTIs) were the first class of agents used for the treatment of HIV and remain an important component of combination antiretroviral therapy . Resistance to the NRTIs occurs by the acquisition of mutations in the reverse transcriptase gene that result in a structural change that either decreases the NRTI incorporation into the extending nucleotide chain or enhances removal of the NRTI from the terminated chain, also known as primer unblocking or pyrophosphorylysis . There are several major genetic mutational patterns of resistance and cross-resistance that evolve with the NRTIs including the thymidine analog mutations M41L, D67N, K70R, L210W, T215Y, and K219Q/E/W, the non-thymidine mutations M184V, L74V, and K65R, and the multidrug resistant Q151M complex, as well as others . Increasing knowledge of resistance and cross-resistance patterns that evolve on the NRTIs as well as the other antiretroviral classes will help optimize antiretroviral treatment strategies . Advancing knowledge of the biochemical and structural basis of resistance will aid in the design of newer compounds that are active against HIV resistant to the currently available drugs, ultimately prolonging virologic suppression and life in the millions of people who are infected with HIV.

Curr Med Chem Anti-Canc Agents, 2004 Jan, 4(1), 53 - 62
Structure of multidrug-resistance proteins of the ATP-binding cassette (ABC) superfamily; Altenberg GA; Multidrug resistance of tumors, characterized by resistance against a variety of chemically unrelated anticancer agents, can be caused by overexpression of ATP-binding cassette (ABC) proteins, such as P-glycoprotein and MRP1 . These multidrug-resistance proteins are plasma-membrane proteins that actively extrude chemotherapeutic agents from the cell interior, decreasing drug accumulation and thus, allowing the cells to survive in the presence of toxic levels of anticancer agents . ABC proteins contain multispanning transmembrane domains and nucleotide-binding domains (NBDs) . The NBDs are responsible for the ATP binding/hydrolysis that drives drug transport, and their structure is conserved independently of the degree of primary-sequence homology . The transmembrane domains contain the drug-binding sites that are likely located in a flexible internal chamber that is sufficiently large to accommodate different drugs . It has been recently proposed that dimerization of the NBDs induced by ATP binding is a key step for the coupling of ATP hydrolysis to substrate transport . The power stroke for substrate transport can be the formation or the dissociation of the dimers . Since the NBDs and TMDs are tightly associated, association/dissociation of the NBDs may control the "gate" of the translocation pathway, formed by intracellular loops . In the case of P-glycoprotein it seems that the power stroke for transport is ATP binding (and therefore NBD dimerization), and not hydrolysis, because the major conformational and functional changes seem to occur at this step.

Curr Med Chem Anti-Canc Agents, 2004 Jan, 4(1), 43 - 52
Reversing agents for ATP-binding cassette (ABC) transporters: application in modulating multidrug resistance (MDR); Lee CH; One of the main reasons for the failure in cancer chemotherapy is the existence of multidrug resistance (MDR) mechanisms . One form of MDR phenotype is contributed by a group of plasma membrane proteins that belong to a large superfamily of proteins called the ATP-binding cassette (ABC) transporters . There has been intense search for compounds, which can act at reversing MDR phenotype exhibited by ABC transporters such as P-glycoprotein (P-gp), multidrug-resistance protein (MRP) and breast cancer resistance protein (BCRP) . Reversing agents can be designed to target MDR-associated ABC transporters at three levels - the protein, mRNA or DNA level . This review aims at describing, over-viewing and discussing currently known MDR reversing agents, which have been shown to act at either of the three levels against ABC transporters . Other potential agents and strategies, which can be used to reverse the MDR phenotype, are also discussed.

Curr Med Chem Anti-Canc Agents, 2004 Jan, 4(1), 31 - 42
Multidrug resistance in cancer chemotherapy and xenobiotic protection mediated by the half ATP-binding cassette transporter ABCG2; Han B et al.; ABCG2, also termed BCRP/MXR/ABCP, is a half ATP-binding cassette (ABC) transporter expressed on plasma membranes . ABCG2 was independently cloned from placenta as well as cell lines selected for resistance to mitoxantrone or anthracyclines . ABCG2 consists of a nucleotide-binding domain (NBD) at the amino terminus and a transmembrane domain (TMD) at the carboxyl terminus and it is postulated to form a homodimer to perform its biological functions . Over-expression of ABCG2 in cell lines confers resistance on a wide variety of anticancer drugs including mitoxantrone, daunorubicin, doxorubicin, topotecan and epirubicin . The expression of ABCG2 has been implicated in multidrug resistance (MDR) of acute myeloid leukemia and some solid tumors . In addition, ABCG2 can transport several fluorescent dyes or toxins . ABCG2 is found to be expressed in epithelial cells of intestine and colon, liver canaliculi, and renal tubules, where it serves to eliminate the plasma level of orally administered anticancer drugs as well as ingested toxins . ABCG2 is found to be highly expressed in placenta and the luminal surface of microvessel endothelium blood-brain barrier where it may play a role in limiting the penetration of drugs, such as topotecan from the maternal plasma into the fetus and from blood to brain . A variety of inhibitors for ABCG2 including GF120918 may prove useful for sensitizing cancer cells to chemotherapy or altering the distribution of orally administered drug substrates of ABCG2 . Interestingly, ABCG2 is also expressed highly in hematopoietic stem cells . However, the function of ABCG2 in stem cells is currently unknown, although it may provide protection to stem cells from a variety of xenobiotics.

Curr Med Chem Anti-Canc Agents, 2004 Jan, 4(1), 19 - 30
Drug binding domains of MRP1 (ABCC1) as revealed by photoaffinity labeling; Karwatsky JM et al.; Drug resistance is a major impediment in the treatment of cancer patients receiving single or multiple drug treatment . Efforts to reverse drug resistance of tumor cells have not been successful . In recent years, considerable emphasis has been placed on understanding the underlying mechanisms that confer drug resistance . The expression of the multidrug resistance protein 1 (MRP1 or ABCC1) in cancer cells has been shown to confer resistance to diverse classes of anti-cancer drugs . MRP1 is a member of the ATP-binding cassette (ABC) family whose function, in tumor cells, is to reduce drug accumulation through energized drug efflux . To learn more about the functions of MRP1 in tumor drug resistance, knowledge of the protein binding characteristics and the location of its binding sites are essential . Photoaffinity labeling (PAL) has emerged as a leading technique that can rapidly shed light on a protein's drug binding characteristics and ultimately drug binding domains . Several MRP1-specific photoreactive probes have been developed . PAL of MRP1 was first demonstrated with the quinoline-based drug, IAAQ . Other studies showed that the high affinity endogenous substrate of MRP1, LTC(4), has intrinsic photoreactive properties and binds within both N- and C-terminal domains of MRP1 . LTC(4) is conjugated to glutathione (GSH), a property common to several MRP1 substrates . In addition, several unconjugated drugs have been identified that interact with MRP1: {(3)H}VF-13,159, IAAQ, IACI and IAARh123 . Mapping studies showed that IACI and IAARh123 bind two sites within transmembrane (TM) regions 10-11 and 16-17 of MRP1 . Interestingly, the GSH-dependent PAL of {(125)I}azidoAG-A and {(125)I}LY475776 occurs within, or proximal to TM 16-17 . The PAL with several analogs of GSH, IAAGSH and azidophenacyl-{(35)S}GSH found to interact specifically with MRP1 within TM 10-11 and TM 16-17 in addition to binding two cytoplasmic regions in MRP1, L0 and L1 . This review focuses on the use of PAL for studying MRP1 interactions with various drugs and cell metabolites . Furthermore, knowledge of MRP1 drug binding domains, as identified by PAL with various photoreactive drug analogs, provides an important first step towards more detailed analyses of MRP1 binding domains.

Curr Med Chem Anti-Canc Agents, 2004 Jan, 4(1), 1 - 17
Identification and characterization of the binding sites of P-glycoprotein for multidrug resistance-related drugs and modulators; Safa AR; A major problem in cancer treatment is the development of resistance to multiple chemotherapeutic agents in tumor cells . A major mechanism of this multidrug resistance (MDR) is overexpression of the MDR1 product P-glycoprotein, known to bind to and transport a wide variety of agents . This review concentrates on the progress made toward understanding the role of this protein in MDR, identifying and characterizing the drug binding sites of P-glycoprotein, and modulating MDR by P-glycoprotein-specific inhibitors . Since our initial discovery that P-glycoprotein binds to vinblastine photoaffinity analogs, many P-glycoprotein-specific photoaffinity analogs have been developed and used to identify the particular domains of P-glycoprotein capable of interacting with these analogs and other P-glycoprotein substrates . Furthermore, significant advances have been made in delineating the drug binding sites of this protein by studying mutant P-glycoprotein . Photoaffinity labeling experiments and the use of site-directed antibodies to several domains of this protein have allowed the localization of the general binding domains of some of the cytotoxic agents and MDR modulators on P-glycoprotein . Moreover, site-directed mutagenesis studies have identified the amino acids critical for the binding of some of these agents to P-glycoprotein . Furthermore, equilibrium binding assays using plasma membranes from MDR cells and radioactive drugs have aided our understanding of the modes of drug interactions with P-glycoprotein . Based on the available data, a topological model of P-glycoprotein and the approximate locations of its drug binding sites, as well as a proposed classification of multiple drug binding sites of this protein, is presented in this review.

Hepatology, 2004 Jan, 39(1), 167 - 78
Genipin enhances Mrp2 (Abcc2)-mediated bile formation and organic anion transport in rat liver; Shoda J et al.; Inchin-ko-to (ICKT), an herbal medicine, and its ingredients exert potent choleretic effects by a "bile acid-independent" mechanism . The current study was designed to determine whether ICKT or its ingredients potentiate multidrug resistance-associated protein 2 (Mrp2; Abcc2)-mediated choleresis in vivo . Biliary secretion of Mrp2 substrates and the protein mass, subcellular localization, and messenger RNA (mRNA) level of Mrp2 were assessed in rat liver after infusion of genipin, an intestinal bacterial metabolite of geniposide, a major ingredient of ICKT . The function of Mrp2 was also assessed by the adenosine triphosphate (ATP)-dependent uptake of Mrp2-specific substrates using canalicular membrane vesicles (CMVs) from the liver . Infusion of genipin increased bile flow by 230% . It also increased biliary secretion of bilirubin conjugates and reduced glutathione (GSH) by 513% and 336%, respectively, but did not increase bile acid secretion . The ATP-dependent uptake of estradiol 17-beta-D-glucuronide (E(2)17 beta G; by 265%), leukotriene C4 (LTC(4); by 161%), taurolithocholate-3-sulfate (TLC-3S; by 266%), and methotrexate (MTX; by 234%) was significantly stimulated in the CMVs from the liver . These effects were not observed in Mrp2-deficient rats . Under these conditions, genipin treatment increased the protein mass of Mrp2 in the CMVs but not the mRNA level . In immunoelectron microscopic studies, a marked increase in Mrp2 density in the canalicular membrane (CM) and microvilli was observed in the genipin-treated liver tissue sections when compared with the vehicle-treated liver tissue sections . In conclusion, genipin may enhance the bile acid-independent secretory capacity of hepatocytes, mainly by stimulation of exocytosis and insertion of Mrp2 in the bile canaliculi . ICKT may be a potent therapeutic agent for a number of cholestatic liver diseases.

Dig Dis, 2003, 21(4), 326 - 38
Molecular analysis of therapy resistance in gastric cancer; Lage H; Therapy resistance is the main cause of therapeutic failure and death in patients suffering from gastric carcinoma . Clinical resistance against systemic chemotherapy of gastric cancer is likely to be multifactorial and heterogenous . So far, no significant resistance factor that predicts the clinical outcome of systemic treatment of gastric carcinoma has been identified . In order to gain further understanding of therapy resistance in gastric carcinoma, various in vitro model systems were established . One of these models consists of the parental, drug-sensitive and thermosensitive human gastric carcinoma cell line EPG85-257P, its classical multidrug-resistant variant EPG85-257RDB, its atypical multidrug-resistant subline EPG85-257RNOV and their thermoresistant counterparts EPG85-257P-TR, EPG85-257RDB-TR, and EPG85-257RNOV-TR . This panel of cells was analyzed using morphological, biochemical, cellular and molecular biological methods to identify potential new factors involved in therapy resistance of gastric carcinoma . Cellular alterations that could be identified in these models were evaluated by functional investigations . This review will discuss the current state of knowledge of these new therapy resistance-associated factors, e.g . glypican-3 (GPC3), as well as the impact of well-known drug resistance-associated factors, such as MDR1/P-glycoprotein, on therapy resistance of gastric carcinoma .

Lancet, 2004 Jan 24, 363(9405), 320 - 4
Scaling-up treatment for HIV/AIDS: lessons learned from multidrug-resistant tuberculosis; Gupta R et al.; The UN has launched an initiative to place 3 million people in developing countries on antiretroviral AIDS treatment by end 2005 (the 3 by 5 target) . Lessons for HIV/AIDS treatment scale-up emerge from recent experience with multidrug-resistant tuberculosis . Expansion of treatment for multidrug-resistant tuberculosis through the multipartner mechanism known as the Green Light Committee (GLC) has enabled gains in areas relevant to 3 by 5, including policy development, drug procurement, rational use of drugs, and the strengthening of health systems . The successes of the GLC and the obstacles it has encountered provide insights for building sustainable HIV/AIDS treatment programmes.

Gan To Kagaku Ryoho, 2004 Jan, 31(1), 1 - 6
{Drug resistance mediated by ABC transporters}; Yoshikawa M et al.; Remarkable advances have been made in cancer chemotherapy by developing new anticancer drugs and pharmacogenomics strategies . However, multidrug resistance in human cancers is the major obstacle to long-term, sustained patient response to chemotherapy . Several ATP-binding cassette (ABC) transporters cause multidrug resistance in cancer cells by actively extruding the clinically administered chemotherapeutic drugs . P-glycoprotein (ABCB1/MDR1/P-gp) and MRP1 (ABCC1/GS-X pump) have been well characterized in terms of their molecular structure and function . In addition, ABCG2/breast cancer resistance protein (BCRP) is the most recently identified/ABC transporter, and is also reportedly associated with cellular resistance against chemotherapeutic agents, such as DNA topoisomerase I, II inhibitor . It is important to note that these ABC transporters are expressed not only in cancer cells but also in normal tissues to play a pivotal role in the absorption, distribution, and excretion of endogenous substances as well as xenobiotics . ABC transporters are key factors that can affect the pharmacokinetic profiles of drugs . Recent studies have revealed that many single nucleotide polymorphisms (SNPs) reside in these ABC transporter genes . Functional analysis of the genetic polymorphism of ABC transporters would greatly contribute to our understanding of individual differences in the drug response and also to the development of personalized medicine in the near future.

Int J Cancer, 2004 Mar 20, 109(2), 174 - 81
Dynamic and intracellular trafficking of P-glycoprotein-EGFP fusion protein: Implications in multidrug resistance in cancer; Fu D et al.; In our present study, a P-glycoprotein-EGFP (P-gp-EGFP) fusion plasmid was constructed and functionally expressed in HeLa cells to investigate the intracellular localization and trafficking of P-glycoprotein (P-gp) . Using immunocytochemistry and fluorescent confocal microscopy techniques, colocalization studies showed that after transfection, P-gp-EGFP was progressively transported from the endoplasmic reticulum (ER) to the Golgi and finally to the plasma membrane within 12-48 hr . The degree of intracellular accumulation of daunorubicin was related to the particular localization of P-gp-EGFP . Significant daunorubicin accumulation occurred in transfected cells when P-gp-EGFP was localized predominantly within the ER, and accumulation remained high when P-gp-EGFP was mainly localized in the Golgi . However, there was little or no intracellular accumulation of daunorubicin when P-gp-EGFP was localized predominantly on the plasma membrane . Blocking the intracellular trafficking of P-gp-EGFP with brefeldin A (BFA) and monensin resulted in inhibition of traffic of P-gp-EGFP and retention of P-gp-EGFP intracellularly . Intracellular accumulation of daunorubicin also increased in the presence of BFA or monensin . Our study shows that P-gp-EGFP can be used to define the dynamics of P-gp traffic in a transient expression system, and demonstrates that localization of P-gp on the plasma membrane is associated with the highest level of resistance to daunorubicin accumulation in cells . Modulation of intracellular localization of P-gp with agents designed to selectively modify its traffic may provide a new strategy for overcoming multidrug resistance in cancer cells .

Ther Drug Monit, 2004 Feb, 26(1), 44 - 6
Neuronal MDR-1 gene expression and persistent low levels of anticonvulsants in a child with refractory epilepsy; Lazarowski A et al.; SUMMARY: It is estimated that 20-25% of epileptic patients fail to achieve good control with antiepileptic drug (AED) treatment; thus, refractory epilepsy (RE) has been described in patients who have adequate therapeutic levels of AEDs without control of seizures . Multidrug resistance genes have been reported to be highly expressed in brain of patients with RE . Persistent low plasma levels of AEDs and high brain expression of the multidrug resistance product P-glycoprotein (P-gp) have been previously communicated in a case report of RE secondary to tuberous sclerosis . Here, the authors report a case of an 8-year-old boy diagnosed with partial RE with focal seizures who was admitted to hospital for a severe episode of subintrant crisis . The patient received polytherapy with carbamazepine (CBZ), phenytoin (PHT), and valproic acid (VA); however, habitual doses of these AEDs failed to control the patient's symptoms . AED blood levels were monitored for 25 consecutive days and showed low values in 8/25 (33%) for CBZ, 10/25 (40%) for PHT, and 25/25 (100%) for VA of samples studied . Because the patient developed focal status epilepticus, surgical treatment by callosotomy was done, resulting in a significant improvement in epileptic symptoms . The immunostaining of brain specimens showed significantly increased expression of P-gp not only in vascular endothelial cells and related astrocytes but also in neurons . Overexpression of P-gp in the brain does not explain the low blood levels of AEDs described in these cases . Different mechanisms such as drug-drug interactions and drug transporters can be involved in the results observed . The P-gp overexpression and/or its pharmacologic induction should be considered as a potential mechanism responsible for drug resistance to epilepsy treatment and highly suspected in patients with persistent subtherapeutic AEDs plasma levels.

J Antimicrob Chemother, 2004 Mar, 53(3), 518 - 21 Epub 2004 Jan 28.
Contribution of integrons, and SmeABC and SmeDEF efflux pumps to multidrug resistance in clinical isolates of Stenotrophomonas maltophilia; Chang LL et al.; OBJECTIVES: The contribution of integrons and efflux pumps to multidrug resistance in Stenotrophomonas maltophilia was evaluated . MATERIALS AND METHODS: Ninety-three S . maltophilia clinical isolates were studied . PCR and direct sequencing were used to detect the presence of integrons . Real-time PCR was performed to assess and quantify the expression of the Sme efflux pumps of S . maltophilia . RESULTS: Class 1 integrons were detected in 22% of clinical isolates and carried cassettes conferring resistance mainly to aminoglycosides and trimethoprim . The small multidrug resistance gene, smr, was found on class 1 integrons in six isolates . Thirty-one percent of the isolates overexpressed the smeDEF gene, as compared with a control strain, and 59% overexpressed the smeABC gene . Extrusion of ciprofloxacin and meropenem was specific to the SmeABC and SmeDEF pumps, respectively . CONCLUSION: SmeABC and SmeDEF efflux pumps play important roles in resistance of S . maltophilia to ciprofloxacin and meropenem.

Psychoneuroendocrinology, 2004 May, 29(4), 423 - 47
Do antidepressants regulate how cortisol affects the brain?
Pariante CM, Thomas SA, Lovestone S, Makoff A, Kerwin RW.
Although the effects of antidepressants on glucocorticoid hormones and their receptors are relevant for the therapeutic action of these drugs, the molecular mechanisms underlying these effects are unclear . Studies in depressed patients, animals and cellular models have demonstrated that antidepressants increase glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) expression and function; this, in turn, is associated with enhanced negative feedback by endogenous glucocorticoids, and thus with reduced resting and stimulated hypothalamic-pituitary-adrenal (HPA) axis activity . In a series of studies conducted over the last few years, we have shown that antidepressants modulate GR function in vitro by inhibiting membrane steroid transporters that regulate the intracellular concentration of glucocorticoids . In this paper, we will review the effects of membrane steroid transporters and antidepressants on corticosteroid receptors . We will then present our unpublished data on GR live microscopy in vitro, showing that ligand-induced translocation of the GR starts within 30 seconds and is completed within minutes . Furthermore, we will present our new data using an in situ brain perfusion model in anaesthetised guinea-pigs, showing that entry of cortisol to the brain of these animals is limited at the blood-brain barrier (BBB) . Finally, we will present a comprehensive discussion of our published findings on the effects of chemically unrelated antidepressants on membrane steroid transporters, in mouse fibroblasts and rat cortical neurones . We propose that antidepressants in humans could inhibit steroid transporters localised on the BBB and in neurones, like the multidrug resistance p-glycoprotein, and thus increase the access of cortisol to the brain and the glucocorticoid-mediated negative feedback on the HPA axis . Enhanced cortisol action in the brain might prove to be a successful approach to maximise therapeutic antidepressant effects.

Zhonghua Er Ke Za Zhi, 2003 Jul, 41(7), 525 - 7
{Reversal of multidrug resistance in leukemic cell line K562/AO2 by chlordelazine in vitro}; Chen LJ et al.; OBJECTIVE: Some recent studies revealed that phenthiazine might be able to reverse tumor cell drug-resistance . Chlorderazin belongs to the phenthiazine compounds . The study aimed to investigate the reversing effect and mechanism of chlorderazin on multidrug resistance of leukemic cell line K562/AO2 . METHODS: (1) The cytotoxicities of chlorderazin were assayed with the tetrazolium dye, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method . (2) The reverse effect of chlorderazin on K562/AO2 cells was analyzed with MTT method . The multidrug resistance reversal index (RI) was equal to the ratio of control group IC(50)/test group half inhibition concentration IC(50) . (3) The intracellular daunorubicin (DNR) concentrations were measured by the flow cytometry . (4) Mdr1 mRNA expression was detected by reverse transcription-polymerase chain reaction (RT-PCR) . The ratio of mdr-1/beta-actin density was calculated . RESULTS: (1) Chlorderazin 3 micro g/ml showed little toxicity to K562/AO2 cells and the suppression rate was less than 5%, so the concentration of 3 micro g/ml chlorderazin was selected as the experiment concentration . (2) The cytotoxicities of DNR to K562/AO2 were enhanced by 3 micro g/ml of chlorderazin (P < 0.05) and RI was 1.901 . (3) Chlorderazin of 3 micro g/ml could increase the intracellular DNR accumulation significantly (P < 0.05), and the fluorescence staining by the flow cytometry was higher (250.95 +/- 18.96) than the control group (112.75 +/- 15.78) and shift right in K562/AO2 cells treated with chlorderazin, and the difference was significant (P < 0.05) . (4) Chlorderazin has no significant influence to the expression level of mdr-1 mRNA . Both test group and control group showed a clear mdr-1 mRNA band located at the position of 157 kb . The ratios of mdr-1/beta-actin density were 0.414 +/- 0.012 in the test group and 0.447 +/- 0.027 in the control group, respectively, and the difference was not significant (P > 0.05) . CONCLUSION: Chlorderazin could reverse the multidrug resistance by increasing the intracellular DNR accumulation in K562/AO2 cells . The effects had no correlation to the mdr-1 gene . Further study is needed.

Annu Rev Med, 2004, 55, 519 - 26
Management of infections in the neutropenic patient; Rolston KV; Neutropenic patients continue to be at increased risk for developing serious infections despite substantial advances in supportive care . Epidemiologic shifts occur periodically and need to be detected early because they influence prophylactic, empiric, and specific therapy strategies . Although effective in preventing bacterial and some fungal infections, prophylaxis must be used with caution because it is associated with the emergence of resistance . The choices for empiric therapy include combination regimens and monotherapy . Specific choices depend on local factors (epidemiology, susceptibility/resistance patterns, availability) . Various treatment settings (hospital-based, early discharge, outpatient) are also available, and the choice depends on the patient's risk category . Early diagnosis and treatment of many fungal and viral infections remains suboptimal . Infection control and prevention are important strategies, especially with the emergence of multidrug-resistant organisms.

Cancer Chemother Pharmacol . 2004 Jan 27; {Epub ahead of print}
Broad-spectrum modulation of ATP-binding cassette transport proteins by the taxane derivatives ortataxel (IDN-5109, BAY 59-8862) and tRA96023; Minderman H et al.; PURPOSE . The taxanes paclitaxel and docetaxel are substrates for P-glycoprotein (Pgp), an ATP-binding cassette (ABC) transport protein associated with multidrug resistance (MDR) . In contrast, the synthetic taxane ortataxel (BAY 59-8862, IDN-5109) is effective against Pgp-expressing cells by virtue of modulation of Pgp-mediated transport . The synthetic taxane tRA96023 also modulates Pgp and is noncytotoxic due to removal of the tubulin-binding side chain at the C-13 position of the taxane backbone . We studied the effects of ortataxel and tRA96023 on the other MDR-associated ABC transport proteins, multidrug resistance protein (MRP-1) and breast cancer resistance protein (BCRP, MXR, ABCG2) . METHODS . Modulation of mitoxantrone, daunorubicin and doxorubicin retention and cytotoxicity by ortataxel and tRA96023 was studied in established cell lines overexpressing Pgp, MRP-1 and wild type (BCRP(R482)) and mutant (BCRP(R482T)) BCRP, and was compared with modulation by the established Pgp-, MRP-1- and BCRP-specific modulators PSC-833, probenecid and fumitremorgin C, respectively . RESULTS . Ortataxel effectively modulated drug retention and cytotoxicity in cell lines overexpressing MRP-1 and BCRP(R482), in addition to Pgp . tRA96023 modulated drug retention and cytotoxicity in cell lines overexpressing BCRP(R482) and Pgp, but not those overexpressing MRP-1 . Neither ortataxel nor tRA96023 modulated BCRP(R482T) . CONCLUSIONS . The synthetic taxane derivatives ortataxel and tRA96023 are broad-spectrum ABC protein modulators . Further studies will seek to identify a noncytotoxic synthetic taxane that modulates Pgp, MRP-1 and BCRP.

Eur J Pharmacol, 2004 Jan 26, 484(2-3), 333 - 9
Azithromycin reverses anticancer drug resistance and modifies hepatobiliary excretion of doxorubicin in rats; Asakura E et al.; The present study aims to investigate whether azithromycin reverses P-glycoprotein-dependent anticancer drug resistance in vitro and modifies the hepatobiliary excretion of doxorubicin, a substrate for P-glycoprotein in vivo . Azithromycin increased dose-dependently the intracellular accumulation of doxorubicin in adriamycin-resistant human myelogenous leukemia cells (K562/ADR) with no effect on the expression of P-glycoprotein in the cells . However, the inhibitory effect was much weaker than that of cyclosporin A and was comparable to that of erythromycin . When Sprague-Dawley (SD) rats, which have drug transporting P-glycoprotein and multidrug resistance-associated protein 2 (Mrp2) in the bile canalicular membrane of hepatocytes, received an infusion of doxorubicin, the steady-state biliary clearance of doxorubicin was significantly decreased for 40 min after a single intravenous injection of azithromycin . However, azithromycin did not increase the plasma concentration of doxorubicin . The biliary clearance of doxorubicin in Eisai hyperbilirubinemic rats (EHBRs), which have a hereditary deficiency in Mrp2, was significantly decreased compared with that in Sprague-Dawley rats, suggesting the involvement of Mrp2 in the biliary excretion of doxorubicin . The present findings suggest that azithromycin overcomes P-glycoprotein-dependent anticancer drug resistance of tumors by inhibiting the binding of doxorubicin to P-glycoprotein in K562/ADR cells and inhibits the hepatobiliary excretion of drugs that are substrates for P-glycoprotein and Mrp2.

Am J Respir Crit Care Med, 2004 May 15, 169(10), 1103 - 9 Epub 2004 Jan 23.
Treatment and outcome analysis of 205 patients with multidrug-resistant tuberculosis; Chan ED et al.; Multidrug-resistant tuberculosis, a disease caused by Mycobacterium tuberculosis strains that are resistant at least to rifampin and isoniazid, entails extended treatment, expensive and toxic regimens, and higher rates of treatment failure and death . We retrospectively analyzed the outcomes in 205 patients treated at our center for multidrug-resistant tuberculosis, with strains resistant to a median of six drugs, and compared the results with those of our previous series . Logistic regression and survival analysis were used to evaluate short- and long-term outcomes, respectively . Initial favorable response, defined as at least three consecutive negative sputum cultures over a period of at least 3 months, was 85% compared with 65% in the prior cohort . The current cohort had greater long-term success rates, 75% versus 56%, and lower tuberculosis death rates, 12% versus 22%, than the earlier one . Surgical resection and fluoroquinolone therapy were associated with improved microbiological and clinical outcomes in the 205 patients studied after adjusting for other variables . The improvement was statistically significant for surgery and among older patients for fluoroquinolone therapy.

Xenobiotica, 2003 Dec, 33(12), 1173 - 83
Altered expression of sulfotransferases, glucuronosyltransferases and mrp transporters in FVB/mrp1-/- mice; Bain LJ et al.; 1 . Genetically altered mice increasingly are being used in toxicology and pharmaceutical development . As such, knowledge of the compensatory activity of enzymes is critical when interpreting the results of studies using these animals . 2 . The present study examined alterations in hepatic phase I and II enzyme activity, and alterations in phase III (transporter) RNA expression, between FVB mice and mice lacking the multidrug resistance-associated protein 1 (mrp1) gene (FVB/mrp1-/- mice) . It was hypothesized that other transporters and phase I and II enzymes would be increased in the FVB/mrp1-/- mice, presumably as a compensatory mechanism . 3 . No differences was found in hepatic cytochrome P450 activity between FVB and FVB/mrp1-/- mice, nor were there differences in the amount of total hepatic glutathione or in glutathione S-transferase enzyme activity . 4 . However, sulfotransferase activity towards 2-naphthol was significantly increased by 2.6-fold in the FVB/mrp1-/- mice, whereas glucuronosyltransferase activity towards both 4-nitrophenol and testosterone was significantly reduced 1.5-fold . In addition, mrp2 RNA expression was significantly increased by 3.4-fold and mrp5 expression was significantly increased by 1.6-fold in the FVB/mrp1-/- mice . 5 . Mice lacking mrp1 have significantly increased hepatic transcription of at least two other ATP-binding cassette transporters, as well as increased 2-naphthol sulfotransferase activity, presumably to compensate for the lack of mrp1.

Nucl Med Biol, 2004 Jan, 31(1), 53 - 65
Imaging recognition of multidrug resistance in human breast tumors using 99mTc-labeled monocationic agents and a high-resolution stationary SPECT system; Liu Z et al.; Imaging recognition of multidrug-resistance by 99mTc-labeled sestamibi, tetrofosmin and furifosmin in mice bearing human breast tumors was evaluated using a high-resolution SPECT, FASTSPECT . Imaging results showed that the washout rates in drug-resistant MCF7/D40 tumors were significantly greater than that in drug-sensitive MCF7/S tumors . Furifosmin exhibited greater washout from both MCF7/S and MCF7/D40 than sestamibi, while tetrofosmin washout was greater than sestamibi in MCF7/D40 only . Feasibility of the monocationic agents for characterizing MDR expression was well clarified with FASTSPECT imaging.

J Biomol NMR, 2004 Jan, 28(1), 43 - 57
An evaluation of detergents for NMR structural studies of membrane proteins; Krueger-Koplin RD et al.; Structural information on membrane proteins lags far behind that on soluble proteins, in large part due to difficulties producing homogeneous, stable, structurally relevant samples in a membrane-like environment . In this study 25 membrane mimetics were screened using 2D (1)H-(15)N heteronuclear single quantum correlation NMR experiments to establish sample homogeneity and predict fitness for structure determination . A single detergent, 1-palmitoyl-2-hydroxy-sn-glycero-3-{phospho-RAC-(1-glycerol)} (LPPG), yielded high quality NMR spectra with sample lifetimes greater than one month for the five proteins tested - R . sphaeroides LH1 alpha and beta subunits, E . coli and B . pseudofirmus OF4 ATP synthase c subunits, and S . aureus small multidrug resistance transporter - with 1, 2, or 4 membrane spanning alpha-helices, respectively . Site-specific spin labeling established interhelical distances in the drug transporter and genetically fused dimers of c subunits in LPPG consistent with in vivo distances . Optical spectroscopy showed that LH1 beta subunits form native-like complexes with bacteriochlorophyll a in LPPG . All the protein/micelle complexes were estimated to exceed 100 kDaltons by translational diffusion measurements . However, analysis of (15)N transverse, longitudinal and (15)N{(1)H} nuclear Overhauser effect relaxation measurements yielded overall rotational correlation times of 8 to 12 nsec, similar to a 15-20 kDalton protein tumbling isotropically in solution, and consistent with the high quality NMR data observed.

Oncogene, 2004 Jan 22, 23(3), 753 - 62
MYCN-mediated regulation of the MRP1 promoter in human neuroblastoma; Manohar CF et al.; In the childhood cancer neuroblastoma (NB), the level of expression of the multidrug resistance-associated protein (MRP1) gene is strongly correlated with expression of the MYCN oncogene in primary NB tumors, suggesting that MRP1 may be a target for MYCN-mediated gene regulation . In this study, we show that MYCN induction in human NB cells results in increased MRP1 mRNA and protein levels, which in turn is accompanied by increased drug resistance and enhanced MRP1-mediated drug efflux . Furthermore, luciferase activity from MRP1 promoter/luciferase gene reporter constructs was significantly increased in NB cells with exogenous overexpression of MYCN, whereas activity was decreased in NB cells stably transfected with MYCN-antisense vectors . Decreased luciferase activity was observed with promoter constructs that lacked one or two E-box sequences or had E-box double point mutations, while a truncated MRP1 promoter lacking all three E-boxes exhibited only basal levels of activity . Specific electrophoretic mobility shifts of MRP1 E-box sequences were detected with nuclear extracts from NB cells with MYCN overexpression, and complex formation was inhibited with the addition of antibodies directed against MYCN or MYC . These findings indicate that by interacting with E-box elements within the promoter, MYCN can upregulate MRP1 expression and modulate drug resistance in NB.

J Med Chem, 2004 Jan 29, 47(3), 576 - 87
SAR studies of dihydro-beta-agarofuran sesquiterpenes as inhibitors of the multidrug-resistance phenotype in a Leishmania tropica line overexpressing a P-glycoprotein-like transporter; Cortes-Selva F et al.; A series of dihydro-beta-agarofuran sesquiterpenes isolated from the leaves of Maytenus cuzcoina (1-10) or semisynthetic derivatives (11-30) have been tested on a multidrug-resistant Leishmania tropica line overexpressing a P-glycoprotein-like transporter to determine their ability to revert the resistance phenotype and to modulate intracellular drug accumulation . Almost all natural compounds showed potent reversal activity with different degrees of selectivity . Compounds 2, 7, and 8 are the most effective reversal agents tested against the multidrug resistance phenotype of Leishmania . Three-dimensional quantitative structure-activity relationships using the comparative molecular similarity indices analysis (CoMSIA) were employed to characterize the steric (contribution of 5.4%), electrostatic (58.9%), lipophilic (10.0%), and hydrogen-bond-donor (13.3%) and -acceptor (7.5%) requirements of these sesquiterpenes as modulators at the P-glycoprotein-like transporter . The most salient features of these requirements are the H-bond interaction between the substituents at the C-2 and C-6 positions with the receptor.

Planta Med, 2003 Nov, 69(11), 1009 - 12
Transport of parthenolide across human intestinal cells (Caco-2); Khan SI et al.; This study examined the intestinal epithelial membrane transport of the sesquiterpene lactone parthenolide, a bioactive compound present in the migraine prophylactic herb feverfew . The Caco-2 human colonic cell line was used as an in vitro model of the human intestinal mucosal barrier . The bidirectional transport (apical to basolateral and basolateral to apical) of parthenolide was investigated using Caco-2 monolayers grown on Transwell inserts . Quantitation of parthenolide was performed using high performance liquid chromatography (HPLC) . Apical to basolateral and basolateral to apical permeability coefficients and percent transport were calculated and a potential bioavailability of parthenolide was determined . Sodium fluorescein was used as a marker for paracellular leakage . Parthenolide, at a concentration of 250 microM, demonstrated substantial linear transport across the monolayer . The transport parameters were not affected by the presence of MK-571, an inhibitor of multidrug resistance transporter P-glycoprotein (MRP) . Upon comparison of the transport parameters of parthenolide with atenolol under identical conditions and the reported values for model compounds like mannitol and propranolol, it is concluded that parthenolide is effectively absorbed through the intestinal mucosa via a passive diffusion mechanism.

Eur J Clin Microbiol Infect Dis, 2004 Mar, 23(3), 174 - 9 Epub 2004 Jan 20.
Impact of drug-resistant Mycobacterium tuberculosis on treatment outcome of culture-positive cases of tuberculosis in the Archangel oblast, Russia, in 1999; Toungoussova OS et al.; The objective of this study was to evaluate the outcome of treatment of culture-positive cases of tuberculosis registered in Archangel, Russia, in 1999, and to analyse the influence of Mycobacterium tuberculosis drug resistance on treatment outcome . The outcome of tuberculosis treatment was evaluated for 235 new and 61 previously treated culture-positive cases diagnosed in 1999 . Of the 235 new cases, there were 150 (63.8%) cases of treatment completion, 20 (8.5%) cases of treatment failure, 29 (12.3%) cases of death during treatment, and 29 (12.3%) cases in which the patient failed to pick up medications for at least 2 consecutive months . The outcome in 7 (3%) cases was unknown, as the patients were transferred outside the oblast region . Among the 61 previously treated cases, the rate of treatment completion was low (26.2%), and rates of treatment failure (23%) and failure to pick up medications for at least 2 consecutive months (29.5%) were high . The relation between the susceptibility pattern of the infecting strain as determined by the Bactec method and tuberculosis treatment outcome was analysed for 76 patients . The majority (69%) of patients infected with drug-susceptible strains was cured . A large proportion (58.8%) of patients infected with Mycobacterium tuberculosis resistant to more than two drugs did not respond to treatment, i.e . the treatment failed or the patients died . The high rates of death (16.7%) and failure (66.7%) among patients infected with multidrug-resistant strains illustrate the negative impact of multidrug resistance on the outcome of tuberculosis treatment . Pan-resistance was significantly associated with treatment failure (P<0.001) . The spread of resistant Mycobacterium tuberculosis has a serious negative impact on the outcome of tuberculosis treatment in Archangel, Russia.

Mol Vis, 2003 Dec 31, 9, 756 - 61
Vasospastic persons exhibit differential expression of ABC-transport proteins; Wunderlich K et al.; PURPOSE: To quantify the gene expression levels of the ABC-proteins MDR1 (P-glycoprotein) and MRP (multidrug resistance-associated protein) isoforms in isolated mononuclear cells of vasospastic persons with increased Endothelin-1 plasma levels . METHODS: Quantitative real-time RT-PCR was performed to determine the expression levels of the MDR1 (P-glycoprotein) gene and MRP1 to MRP5 genes as well as the expression of the ETA and ETB receptor in mononuclear cells derived from 11 vasospastic subjects compared to 10 healthy controls . RESULTS: Mononuclear cells of vasospastic subjects showed a significant decrease in the expression of MDR1 (P-glycoprotein) gene (p=0.029), MRP2 gene (p=0.003), and MRP5 gene (p=0.013) when compared to healthy controls . These effects were poorly correlated with ET-1 plasma levels . No significant ETA and ETB receptor expression was observed in both groups . CONCLUSIONS: Vasospastic persons differ in their expression pattern of MDR proteins from healthy controls . This might be an indirect effect of elevated ET-1 levels.

Clin Cancer Res, 2004 Jan 1, 10(1 Pt 1), 272 - 84
Identification of genes with differential expression in acquired drug-resistant gastric cancer cells using high-density oligonucleotide microarrays; Kang HC et al.; PURPOSE: A major obstacle in chemotherapy is treatment failure due to anticancer drug resistance . The emergence of acquired resistance results from host factors and genetic or epigenetic changes in the cancer cells . The purpose of this study was to identify differentially expressed genes associated with acquisition of resistance in human gastric cancer cells . EXPERIMENTAL DESIGN: We performed global gene expression analysis in the acquired drug-resistant gastric cancer cell lines to the commonly used drugs 5-fluorouracil, doxorubicin, and cisplatin using Affymetrix HG-U133A microarray . The gene expression patterns of 10 chemoresistant gastric cancer cell lines were compared with those of four parent cell lines using fold-change and Wilcoxon's test for data analysis . RESULTS: We identified over 250 genes differentially expressed in 5-fluorouracil-, cisplatin-, or doxorubicin-resistant gastric cancer cell lines . Our expression analysis also identified eight multidrug resistance candidate genes that were associated with resistance to two or more of the tested chemotherapeutic agents . Among these, midkine (MDK), a heparin-binding growth factor, was overexpressed in all drug-resistant cell lines, strongly suggesting that MDK might contribute to multidrug resistance in gastric cancer cells . CONCLUSIONS: Our investigation provides comprehensive gene information associated with acquired resistance to anticancer drugs in gastric cancer cells and a basis for additional functional studies.

Clin Cancer Res, 2004 Jan 1, 10(1 Pt 1), 4 - 12
A systematic review of molecular and biological tumor markers in neuroblastoma; Riley RD et al.; PURPOSE: The aim of this study was to conduct a systematic review, and where possible meta-analyses, of molecular and biological tumor markers described in neuroblastoma, and to establish an evidence-based perspective on their clinical value for the screening, diagnosis, prognosis, and monitoring of patients . Experimental Design: A well-defined, reproducible search strategy was used to identify the relevant literature from 1966 to February 2000 . RESULTS: A total of 428 papers studying the use of 195 different tumor markers in neuroblastoma were identified . Small sample sizes, poor statistical reporting, large heterogeneity across studies (e.g., in cutoff levels), and publication bias limited meta-analysis to the area of prognosis only; MYCN, chromosome 1p, DNA index, vanillylmandelic acid:homovanillic acid ratio, CD44, Trk-A, neuron-specific enolase, lactate dehydrogenase, ferritin, and multidrug resistance were all identified as potentially important prognostic tools . CONCLUSIONS: This systematic review forms a knowledge base of the tumor markers studied thus far in neuroblastoma, and has identified some of the most important prognostic markers, which should be considered in future research and treatment strategies . Importantly, the review has also highlighted some general problems across primary tumor marker studies, in particular poor and heterogeneous reporting . These need to be addressed to allow better clinical interpretation and enable more appropriate evidence-based reviews in the future . In particular, collaboration of cancer research groups is needed to enable bigger sample sizes, standardize methods of analysis and reporting, and facilitate the pooling of individual patient data.

Biochem Biophys Res Commun, 2004 Feb 6, 314(2), 565 - 70
Annexin-I expression modulates drug resistance in tumor cells; Wang Y et al.; The use of anti-cancer chemotherapy often leads to the rise of multidrug-resistant (MDR) tumors . We have previously reported the overexpression of a 40kDa protein (P-40) in several MDR tumor cell lines . In this report we describe the cloning of a 1.4kb cDNA with an open reading frame of 344 amino acids that encodes the P-40 protein . Analysis of the P-40 amino acid sequence showed it is identical to the human annexin I (Anx-I) protein . The identity of the isolated P-40 cDNA as Anx-I was confirmed by the specific binding of IPM96 mAb to a 40kDa protein following the in vitro expression of P-40 full-length cDNA . Northern blot analysis of total RNA from drug-sensitive and -resistant cells revealed an increase in P-40 (or Anx-I) mRNA in drug-resistant cells relative to drug-sensitive cells . Transfection of Anx-I cDNA into drug-sensitive MCF-7 cells was carried out without further drug selection and showed 2- to 5-fold increase in resistance of transfected cells to adriamycin, melphalan, and etoposide . Conversely, transfection of reverse Anx-I cDNA into SKOV-3 cells decreased the expression of Anx-I without affecting the expression of other members of the annexin family and showed a 3- to 8-fold increase in sensitivity to these drugs . Of interest was the correlation between the presence of Anx-I and MDR in MDA-MB-231 cells when compared to MCF-7 cells . MDA-MB-231 cells show 3- to 20-fold increase in resistance to adriamycin, melphalan, and etoposide in the absence of detectable levels of P-glycoprotein (P-gp1), the multidrug resistance protein (MRP1) or the breast cancer resistance protein (BCRP) . Taken together, these results provide the first direct evidence for the role of Anx-I in MDR of tumor cells.

Trends Cell Biol, 1992 Mar, 2(3), 81 - 6
New genes in the MHC that encode proteins for antigen processing; DeMars R et al.; Most cells process proteins into short peptides that are displayed on the cell surface bound to class I or class II proteins encoded by the major histocompatibility complex (MHC) . These protein-peptide complexes can then be recognized by the circulating lymphocytes of the immune system . Several genes found recently in the MHC encode proteins with possible roles in the supply of peptides to class I molecules . The results imply that the peptides are produced in the cytoplasm by proteasomes and are translocated into the endoplasmic reticulum by 'peptide transporters' related to the multidrug resistance proteins . While there is little biochemical evidence to validate these ideas, Robert DeMars and Thomas Spies discuss here the arguments supporting this view . New data indicate that there may also be factors for class II peptide-processing hidden in the MHC.

J Gastroenterol Hepatol, 2004 Feb, 19(2), 146 - 53
Increased hepatic and renal expressions of multidrug resistance-associated protein 3 in Eisai hyperbilirubinuria rats; Kuroda M et al.; BACKGROUND AND AIM: Eisai hyperbilirubinuria rats (EHBR) are animal models of Dubin-Johnson syndrome, which suffer from jaundice due to impaired biliary excretion of bilirubin glucuronides . In EHBR, deficiency of multidrug resistance-associated protein 2 (mrp2) causes defective biliary excretion of numerous organic anions . However, little is known about the expression of other organic anion transporters in this mrp2-deficient model . The aim of the present study was to investigate adaptive expressions of mrp1, mrp3, mrp6, organic anion transporting polypeptide 1 (oatp1) and oatp2 in liver and kidney of EHBR . METHODS: For the present study, EHBR (n = 5) were used . Hepatic and renal mRNA expression of the aforementioned transporters was determined by constructed semiquantitative reverse transcription polymerase chain reaction assay . Their protein expression was determined by western blotting . Localization of hepatic and renal mrp3 was confirmed by immunohistochemistry . Sprague-Dawley (SD) rats (n = 5) were used as normal controls . RESULTS: Deficiency of mrp2 protein was confirmed in EHBR . Hepatic and renal expression of mrp3 mRNA was 53.6% (P < 0.001) and 82.9% (P < 0.001), and its protein expression was 298.9% (P < 0.001) and 245.0% (P = 0.001) higher in EHBR than in SD rats, respectively . Hepatic and renal expression of mrp1 and mrp6 mRNA was not significantly different between EHBR and SD rats . The mrp1 and mrp6 proteins were expressed in very low amounts in the liver and kidney of both EHBR and SD rats . In contrast to mrp3, hepatic expression of oatp1 and oatp2 mRNA was 33.9% (P = 0 . 001) and 38.6% (P < 0.001), and their protein expression was 57.4% (P < 0.05) and 51.0% (P < 0.01) lower in EHBR than in SD rats, respectively . Hepatic and renal mrp3 protein was localized at the basolateral membrane . CONCLUSIONS: Mrp3 plays an important role in the compensation of mrp2 deficiency in liver and kidney of EHBR . Hepatic expressions of mrp3, oatp1 and oatp2 changed adaptively in this animal model . This is a compensatory mechanism for reducing injury to hepatocytes from cytotoxic materials that increase in mrp2 deficiency.

Electrophoresis, 2004 Jan, 25(1), 173 - 83
Study of the development of thermoresistance in human pancreatic carcinoma cell lines using proteome analysis; Poland J et al.; In order to find candidate proteins that are potentially associated with the thermoresistant phenotype in combination with drug resistance, we analyzed the differential protein expression in vitro in the human pancreatic cancer cell line EPP85-181-P and classical and atypical multidrug-resistant variants and their thermoresistant counterparts using proteomics . This study identifies sets of proteins that may lead to the development of thermoresistance . These results provide a fundamental basis to elucidate the molecular mechanism of thermoresistance and chemoresistance phenomena that may assist the therapy of inoperable cancers.

J Med Microbiol, 2004 Feb, 53(Pt 2), 107 - 13
Genotypic analysis of multidrug-resistant Mycobacterium tuberculosis isolates from Monterrey, Mexico; Ramaswamy SV et al.; Thirty-seven multidrug-resistant and 13 pan-susceptible isolates of Mycobacterium tuberculosis were analysed for the diversity of genotypes associated with known drug-resistance mechanisms . The isolates were obtained from patients attending a university tuberculosis clinic in Monterrey, Mexico . A total of 25 IS6110-RFLP patterns were obtained from the multidrug-resistant tuberculosis (MDR-TB) isolates . Approximately 65% of the MDR-TB isolates were attributed to secondary resistance . Different drug-susceptibility patterns were seen with the clustered isolates . The percentage of isolates resistant to isoniazid (INH), rifampicin (RIF), ethambutol (EMB) and streptomycin (STR) was 100, 97.3, 48.7 and 67.6, respectively . The most common resistance-associated polymorphisms for the four drugs were as follows: INH, Ser315Thr (67.6%) in katG; RIF, Ser450Leu (41.7%) in rpoB; EMB, Met306Ile/Val/Leu (66.7%) in embB; and STR, Lys43Arg (24%) in rpsL . Drug-resistance-associated mutations were similar to changes occurring in isolates from other areas of the world, but unique, previously unreported, mutations in katG (n=5), rpoB (n=1) and rrs (n=3) were also identified.

Cancer Res, 2004 Jan 1, 64(1), 322 - 8
Multidrug-resistant cancer cells facilitate E1-independent adenoviral replication: impact for cancer gene therapy; Holm PS et al.; Resistance to chemotherapy is responsible for a failure of current treatment regimens in cancer patients . We have reported previously that the Y-box protein YB-1 regulates expression of the P-glycoprotein gene mdr1, which plays a major role in the development of a multidrug resistant-tumor phenotype . YB-1 predicts drug resistance and patient outcome in breast cancer . Thus, YB-1 is a promising target for new therapeutic approaches to defeat multidrug resistance . In drug-resistant cancer cells and in adenovirus-infected cells YB-1 is found in the nucleus . Nuclear accumulation of YB-1 in adenovirus-infected cells is a function of the E1 region, and we have shown that YB-1 facilitates adenovirus replication . Here we report that E1A-deleted or mutant adenovirus vectors, such as Ad312 and Ad520, replicate efficiently in multidrug-resistant (MDR) cancer cells and induce an adenovirus cytopathic effect resulting in host cell lysis . Thus, replication-defective adenoviruses are a previously unrecognized vector system for a selective elimination of MDR cancer cells . Our work forms the basis for the development of novel oncolytic adenovirus vectors for the treatment of MDR malignant diseases in the clinical setting.

Carcinogenesis, 2004 Jun, 25(6), 941 - 9 Epub 2004 Jan 16.
Reversal of P-glycoprotein-mediated multidrug resistance by diallyl sulfide in K562 leukemic cells and in mouse liver; Arora A et al.; Multidrug resistance (MDR) mediated by the overexpression of drug efflux protein P-glycoprotein (P-gp) is one of the major obstacles to successful cancer chemotherapy . P-gp acts as an energy-dependent drug efflux pump, reducing the intracellular concentration of structurally unrelated drugs . Modulators of P-gp function can restore the sensitivity of multidrug-resistant cells to such drugs . In the present study, we evaluated the P-gp modulatory potential of diallyl sulfide (DAS), a volatile organosulfur compound present in garlic, known to possess many medicinal properties, including antimutagenic and anticarcinogenic activities . For in vitro studies, K562 leukemic cells were made resistant (K562/R) to the cytotoxicity of vinblastine (VBL) by progressive adaptation of the sensitive K562 parental cells to VBL . Cross-resistance of K562/R was found between vincristine (VCR), doxorubicin and other antineoplastic agents . A non-toxic concentration of DAS (8.75 x 10(-3) M) enhanced the cytotoxic effects of VBL and another vinca alkaloid, VCR, time dependently in VBL-resistant human leukemia (K562/R10) cells but had no effect on the parent (K562/S) cells . The results show that DAS decreased the induced levels of P-gp in resistant cells back to the normal levels as analyzed both qualitatively and quantitatively by western blotting and immunocytochemistry . Furthermore, in vivo combination studies showed that DAS effectively inhibited vinca alkaloid-induced P-gp overexpression in mouse hepatocytes . Quantitation of immunostained tissue sections with image analysis showed that the reduction in P-gp levels was up to 73% for VBL- and 65% for VCR-induced drug resistance . The above features thus indicate that DAS can serve as a novel, non-toxic modulator of MDR and can be used as a dietary adjuvant.

Toxicol Appl Pharmacol, 2004 Jan 1, 194(1), 1 - 9
Human intestinal absorption of imidacloprid with Caco-2 cells as enterocyte model; Brunet JL et al.; In order to assess the risk to mammals of a chronic exposure to imidacloprid (IMI), we investigated its absorption with the human intestinal Caco-2 cell line . Measurements of transepithelial transport revealed an apparent permeability coefficient of 21.6 x 10(-6) +/- 3.2 x 10(-6) cm/s reflecting a 100% absorption . The comparison of apical to basal (A-B) and basal to apical (B-A) transports showed that the monolayer presents a basal to apical polarized transport . Studies of apical uptake demonstrated that the transport was concentration-dependent and not saturable from 5 to 200 microM . Arrhenius plot analysis revealed two apparent activation energies, Ea(4-12 degrees C) = 63.8 kJ/mol and Ea(12-37 degrees C) = 18.2 kJ/mol, suggesting two temperature-dependent processes . IMI uptake was equivalent when it was performed at pH 6.0 or 7.4 . Depletion of Na+ from the transport buffer did not affect the uptake, indicating that a sodium-dependent transporter was not involved . Decrease of uptake with sodium-azide or after cell surface trypsin (Ti) treatment suggested the involvement of a trypsin-sensitive ATP-dependent transporter . Investigations on apical efflux demonstrated that initial velocities paralleled the increase of loading concentrations . A cell surface trypsin treatment did not affect the apical efflux . The lack of effect when the efflux was performed against an IMI concentration gradient suggested that an energy-dependent transporter was involved . However, the inhibition of P-glycoproteins (P-gp) and multidrug resistance-associated proteins (MRP) by taxol, vincristine, and daunorubicine had no effect on IMI intracellular accumulation suggesting the involvement of transporters distinct from classical ATP binding cassette transport (ABC-transport) systems . All results suggest that IMI is strongly absorbed in vivo by inward and outward active transporters.

Zhonghua Wai Ke Za Zhi, 2003 Dec, 41(12), 893 - 6
{The role of cell adhesion, multidrug resistance and cell proliferation in short-term recurrent cases with T1G3 superficial bladder cancer}; Zhu YY et al.; OBJECTIVE: To evaluate the roles of cell adhesion, multidrug resistance and cell proliferation in short-term recurrent cases with superficial bladder cancer, and the prognostic value of the three indexes . METHODS: Immunohistochemical staining for E-cad, P-gp and Ki-67 was performed on the tumors of 100 patients with stage T0-T1 transitional cell carcinoma of the bladder who had been included in a retrospective research by follow-up . RESULTS: E-cad and P-gp expression was positive in 51 (43.2%)and 17 (14.4%) of the tumors, respectively and mean proliferation index (PI) was 22.1% . The decrease in E-cad expression was accompanied with the increasing recurrent episodes (P < 0.05), while increase of P-gp expression and PI were accompanied with the increasing recurrence episodes (P < 0.05) . There was significant difference according to E-cad, P-gp positivity and between T(1)G(3) patients and no-T(1)G(3) patients (P < 0.05) . There was negative correlation of E-cad expression with P-gp expression and PI . CONCLUSIONS: Minimum adhesion, strong drug resistance and maximum proliferation are the main factors that promote short-term recurrence of superficial bladder cancer and also the inherent reasons for easy recurrence and high malignancy of T(1)G(3) tumors . During this course, the three aspects may interact.

Cancer Biol Ther, 2004 Apr, 3(4), 377 - 81 Epub 2004 Apr 17.
Overexpression of ZNRD1 promotes multidrug-resistant phenotype of gastric cancer cells through upregulation of P-glycoprotein; Shi Y et al.; ZNRD1, a new zinc ribbon gene, has been previously identified as an upregulated gene in a multidrug-resistant gastric cancer cell line SGC7901/VCR comparing to its parental cell SGC7901 by subtractive hybridization and RT-PCR . The antisense nucleic acid for ZNRD1 could enhance adriamycin accumulation in SGC7901/VCR cells and sensitize SGC7901/VCR cells to vincristine . The present study aims to explore the role of ZNRD1 in multidrug resistance in gastric cancer cells . Upregulation of ZNRD1 protein in SGC7901/VCR cells was confirmed by Western blot and immunocytochmical staining . ZNRD1 was genetically overexpressed in SGC7901 cells by gene transfection . It was found that overexpression of ZNRD1 could sensitize SGC7901 cells to P-glycoprotein (P-gp)-related anticancer drugs (vincristine, adriamycin, etoposide) but not to P-gp-nonrelated drugs (5-fluorouracil and cisplatin), which was accompanied with significantly decreased adriamycin accumulation and retention and increased adriamycin releasing in SGC7901 cells . Verapamil, an inhibitor for P-gp, could reverse the effects of ZNRD1 on drug sensitivity and drug accumulation in SGC7901 cells to a great extent . Western blot and Northern blot revealed that overexpression of ZNRD1 could upregulate P-gp at both protein and mRNA levels . Together, these results suggest that overexpression of ZNRD1 could promote multidrug-resistant phenotype of gastric cancer cells through upregulation of P-gp.

Shi Yan Sheng Wu Xue Bao, 2003 Oct, 36(5), 342 - 6
{Development of a K562 multidrug-resistant cell line and study on proteins with altered expression}; Wang Y et al.; In an attempt to study the whole protein expression alterations of tumer cells after becoming multidrug-resistant, which may provide useful information on new drug target identification, an adriamycin-resistant variant of the human leukemia cell line K562 (K562/ADR) was developed in vitro by continuous exposure to adrimycin . MTT assay was used to determine IC50 of K562/ADR cells to adriamycin (ADR), cisplatin (DDP), 5-fluorouracil (5-FU) and vincristin (VCR) . The total proteins of K562 and K562/ADR were separated by two-dimensional gel electrophoresis and visualized by silver staining . Proteins with significant expression alterations were selected and their peptide mass fingerprints (PMFs) were obtained by matrix-assisted laser desorption/ionization time of flying mass spectrometry (MALDI-TOF-MS) . The PMFs were used to search NCBInr database by AutoMS-Fit software . The results showed that K562/ADR cell demonstrated cross-resistance to other antineoplastic drugs . The IC50 of K562/ADR cells to ADR, DDP, 5-FU, VDR were much higher than those of K562 . The proteins differentially expressed in the two cell lines were identified as cell cycle-related proteins, zinc finger protein 165, etc . These proteins are involved in cell cycling and transcription regulation, whose expression alterations may contribute to the multidrug resistant phenotype of K562/ADR cells.

Transplantation, 2004 Jan 15, 77(1), 22 - 7
Expression of MRP2 and MRP3 during liver regeneration after 90% partial hepatectomy in rats; Chang TH et al.; BACKGROUND: Small-for-size grafts often cause persistent conjugated hyperbilirubinemia in the recipient after adult-to-adult living donor liver transplantation, but the cause has not yet been clarified . In physiologic status, bilirubin is excreted from hepatocytes to the bile canaliculus by means of multidrug resistance protein (MRP) 2 and, in particular circumstances, by means of MRP3 to the sinusoidal space . The aim of this study was to research whether there is any change in bilirubin excretion pattern during liver regeneration with reference to expression of MRP2 and MRP3 . METHODS: Sprague-Dawley rats underwent sham operation (n=37), 70% hepatectomy (n=38), or 90% hepatectomy (n=37) . The degree of liver regeneration, total and direct bilirubin, protein synthesis, and interleukin (IL)-6 were serially assessed . Expression of MRP2 and MRP3 were semiquantified by Western blotting . RESULTS: The proliferating cell nuclear antigen labeling index indicated rapid liver regeneration after 70% and 90% hepatectomy . Serum levels of total and direct bilirubin increased significantly (P<0.05), and conjugated hyperbilirubinemia was proved only in the 90% hepatectomy group . Coagulation factor VII dipped but increased as early as 12 to 24 hr postoperatively in both hepatectomy groups . Plasma IL-6 levels were significantly increased in the 90% hepatectomy group (P<0.05) . Expression of MRP2 was decreased and MRP3 was expressed at 36 and 72 hr postoperatively in the 90% hepatectomy group, whereas no change was observed in MRP expression in the 70% hepatectomy group . CONCLUSIONS: During liver regeneration after critical hepatectomy such as 90% hepatectomy, decrease of MRP2 and expression of MRP3 may play an important role in postoperative hyperbilirubinemia.

Lancet, 2004 Jan 3, 363(9402), 18 - 22
Dihydroartemisinin-piperaquine against multidrug-resistant Plasmodium falciparum malaria in Vietnam: randomised clinical trial; Tran TH et al.; BACKGROUND: Southeast Asia has the most resistant malaria parasites in the world, which severely limits treatment options . There is general acceptance that to combat resistance, combinations of antimalarial drugs that include an artemisinin derivative should be used, and, if possible, these should be formulated in a single tablet . METHODS: We did a pilot randomised study in a tertiary referral hospital in Vietnam to compare the efficacy of 3-day regimens of dihydroartemisinin-trimethoprim-piperaquine (DHA-TP total dose 4.8/13.6/48 mg/kg, respectively) with the standard antimalarial regimen in Vietnam, artesunate-mefloquine (A3M total dose 12/25 mg/kg, respectively) in non-immune patients with uncomplicated Plasmodium falciparum malaria . 114 patients were randomised, 76 to DHA-TP and 38 to A3M . The subsequent open randomised trial at a Provincial Health Station compared DHA-TP, dihydroartemisinin-piperaquine, and A3M in 400 patients . In both studies all patients received directly observed therapy and were followed up for 56 days . The primary endpoint was reappearance of P falciparum malaria within 56 days of treatment . Analysis was by intention to treat . FINDINGS: The 56-day cure rate in the pilot study, adjusted for reinfections identified by PCR genotyping, was 97.4% (74/76) in the DHA-TP group and 100% (38/38) in the A3M group . In the second study, cure rates were similar in the three groups; DHA-TP 97.4% (153/157), dihydroartemisinin-piperaquine 98.7% (164/166), and A3M 98.7% (76/77) . The DHA-TP and dihydroartemisinin-piperaquine regimens were well tolerated; fewer than 3% of patients had side-effects that might have been related to treatment, compared with 16% of A3M patients (p<0.001) . No patients were lost to follow-up . INTERPRETATION: Dihydroartemisinin-piperaquine is an inexpensive, safe, highly efficacious fixed-dose antimalarial combination treatment that could make an important contribution to the control of multidrug-resistant falciparum malaria.

Neurol Med Chir (Tokyo), 2003 Dec, 43(12), 573 - 80; discussion 581
Technetium-99m sestamibi single photon emission computed tomography findings correlated with P-glycoprotein expression, encoded by the multidrug resistance gene-1 messenger ribonucleic acid, in intracranial meningiomas; Kunishio K et al.; The present study evaluated whether technetium-99m sestamibi (99mTc-MIBI) single photon emission computed tomography (SPECT) characteristics of intracranial meningioma are correlated with the histological malignancy, proliferative potential, and P-glycoprotein (Pgp) expression, encoded by the multidrug resistance gene-1 (MDR-1) messenger ribonucleic acid (mRNA) . Twenty-one patients with intracranial meningiomas, including 17 benign and four nonbenign meningiomas, underwent 99mTc-MIBI SPECT imaging at 15 minutes (early) and 3 hours (delayed) after injection . The tumor-to-normal pituitary gland ratio was calculated on both early (ER) and delayed (DR) images . Retention index (RI) was calculated using the following formula: (DR - ER)/ER x 100% . Meningioma specimens were examined by immunohistochemistry using anti-Pgp and MIB-1 monoclonal antibody . MDR-1 mRNA expression was also investigated using reverse transcription-polymerase chain reaction assay . 99mTc-MIBI was highly accumulated and retained in the tumors . 99mTc-MIBI SPECT findings were not related to MIB-1 labeling index . 99mTc-MIBI SPECT RI of the Pgp-positive group (-9.12 +/- 22.27%) was significantly lower than that of the Pgp-negative group (28.79 +/- 22.80%) (p = 0.0016) . No significant difference was seen in ER and DR between the positive and negative groups . These results show that 99mTc-MIBI may not be useful for determining proliferative potential and histological malignancy, but could predict anticancer drug resistance related to the expression of MDR-1 mRNA and its gene product Pgp in patients with intracranial meningiomas.

J Pharmacol Exp Ther, 2004 Apr, 309(1), 156 - 64 Epub 2004 Jan 13.
Transport of ethinylestradiol glucuronide and ethinylestradiol sulfate by the multidrug resistance proteins MRP1, MRP2, and MRP3; Chu XY et al.; Ethinylestradiol (EE) is one of the key constituents of oral contraceptives . Major metabolites of EE in humans are the glucuronide and sulfate conjugates, EE-3-O-glucuronide (EE-G) and EE-3-O-sulfate (EE-S) . In the present study, transport of EE-G and EE-S by the human multidrug resistance proteins MRP1, MRP2, and MRP3 was investigated using inside-out membrane vesicles, isolated from Sf9 cells expressing human MRP1, MRP2, or MRP3 . Vesicular uptake studies showed that EE-G was not a substrate for MRP1, whereas an ATP-dependent and saturable transport of {(3)H}EE-G was observed in MRP2 (K(m) of 35.1 +/- 3.5 microM) and MRP3 (K(m) of 9.2 +/- 2.3 microM) containing vesicles . EE-S was not transported by either MRP1, MRP2, or MRP3 . However, low concentrations of EE-S stimulated MRP2-mediated uptake of ethacrynic acid glutathione . EE-S also stimulated MRP2 and MRP3-mediated uptake of 17beta-estradiol-17beta-D-glucuronide . Interestingly, EE-S stimulated strongly MRP2- and MRP3-mediated uptake of EE-G by increasing its apparent transport affinity, whereas no reciprocal stimulation of EE-S uptake by EE-G was observed . These data indicate that EE-S allosterically stimulates MRP2- and MRP3-mediated transport of EE-G and is not cotransported with EE-G . Our studies demonstrate specific active transport of a pharmacologically relevant drug conjugate by human MRP2 and MRP3, involving complex interactions with other organic anions . We also suggest that caution needs to be taken when using only competition studies as screening tools to identify substrates or inhibitors of MRP-mediated transport.

Mol Pharmacol, 2004 Jan, 65(1), 77 - 84
Salvinal, a novel microtubule inhibitor isolated from Salvia miltiorrhizae Bunge (Danshen), with antimitotic activity in multidrug-sensitive and -resistant human tumor cells; Chang JY et al.; Aqueous extracts of Salvia miltiorrhizae Bunge have been extensively used in the treatment of cardiovascular disorders and cancer in Asia . Recently, a compound, 5-(3-hydroxypropyl)-7-methoxy-2-(3'-methoxy-4'-hydroxyphenyl)-3-benzo{b}furancarbaldehyde (salvinal), isolated from this plant showed inhibitory activity against tumor cell growth and induced apoptosis in human cancer cells . In the present study, we investigated the cytotoxic effect and mechanisms of action of salvinal in human cancer cell lines . Salvinal caused inhibition of cell growth (IC50 range, 4-17 microM) in a variety of human cancer cell lines . Flow cytometry analysis showed that salvinal treatment resulted in a concentration-dependent accumulation of cells in the G(2)/M phase . We observed, using Hoechst 33258 dye staining, that salvinal blocked the cell cycle in mitosis . In vitro and in vivo examinations showed that salvinal inhibited tubulin polymerization in a concentration-dependent manner . Immunocytochemical studies demonstrated that salvinal treatment caused the changes of cellular microtubule network, similar to the effect of colchicine . In addition, salvinal treatment resulted in upregulation of cyclin B1 levels, activation of Cdc2 kinase, and Cdc25c phosphorylation . Furthermore, elevation of levels of MPM-2 phosphoepitopes in salvinal-treated cells in a concentration-dependent manner was also observed . Similar to the effect of other antitubulin agent, hyperphosphorylation of Bcl-2, induction of DNA fragmentation and activation of caspase-3 activity occurred in salvinal-treated cells . In particular, salvinal exhibited similar inhibitory activity against parental KB, P-glycoprotein-overexpressing KB vin10 and KB taxol-50 cells, and multidrug resistance-associated protein (MRP)-expressing etoposide-resistant KB 7D cells . Taken together, our data demonstrate that salvinal inhibits tubulin polymerization, arrests cell cycle at mitosis, and induces apoptosis . Notably, Salvinal is a poor substrate for transport by P-glycoprotein and MRP . Salvinal may be useful in the treatment of human cancers, particularly in patients with drug resistance.

Ann Rheum Dis, 2004 Feb, 63(2), 138 - 43
Development of sulfasalazine resistance in human T cells induces expression of the multidrug resistance transporter ABCG2 (BCRP) and augmented production of TNFalpha; van der Heijden J et al.; OBJECTIVE: To determine whether overexpression of cell membrane associated drug efflux pumps belonging to the family of ATP binding cassette (ABC) proteins contributes to a diminished efficacy of sulfasalazine (SSZ) after prolonged cellular exposure to this disease modifying antirheumatic drug (DMARD) . METHODS: A model system of human T cells (CEM) was used to expose cells in vitro to increasing concentrations of SSZ for a period of six months . Cells were then characterised for the expression of drug efflux pumps: P-glycoprotein (Pgp, ABCB1), multidrug resistance protein 1 (MRP1, ABCC1), and breast cancer resistance protein (BCRP, ABCG2) . RESULTS: Prolonged exposure of CEM cells to SSZ provoked resistance to SSZ as manifested by a 6.4-fold diminished antiproliferative effect of SSZ compared with parental CEM cells . CEM cells resistant to SSZ (CEM/SSZ) showed a marked induction of ABCG2/BCRP, Pgp expression was not detectable, while MRP1 expression was even down regulated . A functional role of ABCG2 in SSZ resistance was demonstrated by 60% reversal of SSZ resistance by the ABCG2 blocker Ko143 . Release of the proinflammatory cytokine tumour necrosis factor alpha (TNFalpha) was threefold higher in CEM/SSZ cells than in CEM cells . Moreover, twofold higher concentrations of SSZ were required to inhibit TNFalpha release from CEM/SSZ cells compared with CEM cells . CONCLUSION: Collectively, ABCG2 induction, augmented TNFalpha release, and less efficient inhibition of TNFalpha production by SSZ may contribute to diminished efficacy after prolonged exposure to SSZ . These results warrant further clinical studies to verify whether drug efflux pumps, originally identified for their roles in cytostatic drug resistance, can also be induced by SSZ or other DMARDs.

Ann Rheum Dis, 2004 Feb, 63(2), 131 - 7
Acquired resistance of human T cells to sulfasalazine: stability of the resistant phenotype and sensitivity to non-related DMARDs; van der Heijden J et al.; BACKGROUND: A recent study from our laboratory showed that induction of the multidrug resistance related drug efflux pump ABCG2 contributed to acquired resistance of human T cells to the disease modifying antirheumatic drug (DMARD) sulfasalazine (SSZ) . OBJECTIVES: To investigate the duration of SSZ resistance and ABCG2 expression after withdrawal of SSZ and rechallenging with SSZ, and to assess the impact of SSZ resistance on responsiveness to other DMARDs . METHODS: Human CEM cells (T cell origin) with acquired resistance to SSZ (CEM/SSZ) were characterised for (a) SSZ sensitivity and ABCG2 expression during withdrawal and rechallenge of SSZ, and (b) antiproliferative efficacy of other DMARDs . RESULTS: ABCG2 protein expression was stable for at least 4 weeks when CEM/SSZ cells were grown in the absence of SSZ, but gradually declined, along with SSZ resistance levels, to non-detectable levels after withdrawal of SSZ for 6 months . Rechallenging with SSZ led to a rapid (<2.5 weeks) resumption of SSZ resistance and ABCG2 expression as in the original CEM/SSZ cells . CEM/SSZ cells displayed diminished sensitivity to the DMARDs leflunomide (5.1-fold) and methotrexate (1.8-fold), were moderately more sensitive (1.6-2.0 fold) to cyclosporin A and chloroquine, and markedly more sensitive (13-fold) to the glucocorticoid dexamethasone as compared with parental CEM cells . CONCLUSION: The drug efflux pump ABCG2 has a major role in conferring resistance to SSZ . The collateral sensitivity of SSZ resistant cells for some other (non-related) DMARDs may provide a further rationale for sequential mono- or combination therapies with distinct DMARDs upon decreased efficacy of SSZ.

J Biol Chem, 2004 Mar 26, 279(13), 12325 - 36 Epub 2004 Jan 13.
Identification of proline residues in the core cytoplasmic and transmembrane regions of multidrug resistance protein 1 (MRP1/ABCC1) important for transport function, substrate specificity, and nucleotide interactions; Koike K et al.; Multidrug resistance protein 1 (MRP1/ABCC1) is an ATP-binding cassette transporter that confers resistance to drugs and mediates the transport of organic anions . MRP1 has a core structure of two membrane spanning domains (MSDs) each followed by a nucleotide binding domain . This core structure is preceded by a third MSD with five transmembrane (TM) helices, whereas MSD2 and MSD3 each contain six TM helices . We investigated the consequences of Ala substitution of 18 Pro residues in both the non-membrane and TM regions of MSD2 and MSD3 on MRP1 expression and organic anion transport function . All MRP1-Pro mutants except P1113A were expressed in human embryonic kidney cells at levels comparable with wild-type MRP1 . In addition, five mutants containing substitutions of Pro residues in or proximal to the TM helices of MSD2 (TM6-Pro(343), TM8-Pro(448), TM10-Pro(557), and TM11-Pro(595)) and MSD3 (TM14-Pro(1088)) exhibited significantly reduced transport of five organic anion substrates . In contrast, mutation of Pro(1150) in the cytoplasmic loop (CL7) linking TM15 to TM16 caused a substantial increase in 17beta-estradiol-17-beta-(D-glucuronide) and methotrexate transport, whereas transport of other organic anions was reduced or unchanged . Significant substrate-specific changes in the ATP dependence of transport and binding by the P1150A mutant were also observed . Our findings demonstrate the importance of TM6, TM8, TM10, TM11, and TM14 in MRP1 transport function and suggest that CL7 may play a differential role in coupling the activity of the nucleotide binding domains to the translocation of different substrates across the membrane.

Emerg Infect Dis, 2003 Dec, 9(12), 1571 - 8
Multidrug-resistant Mycobacterium tuberculosis in HIV-infected persons, Peru; Campos PE et al.; During 1999 to 2000, we identified HIV-infected persons with new episodes of tuberculosis (TB) at 10 hospitals in Lima, Peru, and a random sample of other Lima residents with TB . Multidrug-resistant (MDR)-TB was documented in 35 (43%) of 81 HIV-positive patients and 38 (3.9%) of 965 patients who were HIV-negative or of unknown HIV status (p<0.001) . HIV-positive patients with MDR-TB were concentrated at three hospitals that treat the greatest numbers of HIV-infected persons with TB . Of patients with TB, those with HIV infection differed from those without known HIV infection in having more frequent prior exposure to clinical services and more frequent previous TB therapy or prophylaxis . However, MDR-TB in HIV-infected patients was not associated with previous TB therapy or prophylaxis . MDR-TB is an ongoing problem in HIV-infected persons receiving care in public hospitals in Lima and Callao; they represent sentinel cases for a potentially larger epidemic of nosocomial MDR-TB.

Emerg Infect Dis, 2003 Dec, 9(12), 1553 - 7
Mycobacterium tuberculosis Beijing genotype; Lillebaek T et al.; Molecular epidemiologic studies of strains of Mycobacterium tuberculosis are currently conducted worldwide . The genetically distinct Beijing family of strains has been associated with large outbreaks of tuberculosis, increased virulence, and multidrug resistance . However, in this first population-based search for Beijing strains in the Danish DNA fingerprint database, analysis of 97% of all culture-positive tuberculosis patients in 1992 to 2001 showed that 2.5% of 3,844 patients, 1.0% of Danish-born patients, and 3.6% of immigrants (from 85 countries) had Beijing strains . No Beijing strains were found among 201 strains from Danish-born patients sampled in the 1960s, and no evidence of an increase in Beijing strains was found over time . The true prevalence of Beijing strains worldwide is unknown because only a fraction of global strains have been analyzed.

Hinyokika Kiyo, 2003 Nov, 49(11), 649 - 58
Proliferative status is a risk index for recurrence in primary superficial (pTa/T1) low-grade urothelial bladder carcinoma; Su JS et al.; The current clinicopathologic study for evaluation of superficial bladder cancer still has limitations in predicting the true behavior of recurrence . To determine the high-risk recurrence factors, we studied the influence of Ki-67, c-erbB-2, p53 and multidrug resistance-associated protein (MRP) expression . Samples were obtained from 33 pTa and 46pT1 diagnosed bladder cancer patients with a mean follow-up of 48.7 +/- 30.6 months . The contingency table method, Kaplan-Meier curve and multivariate analysis were used to evaluate the association among the immunohistochemical factors expression, clinicopathologic parameters with tumor recurrence . Stage pT1 tumors, sessile tumors and large tumors (> 3 cm) showed a significantly high recurrence rate (p = 0.0158, p = 0.0162, p = 0.0001 respectively) . Tumors with overexpression of Ki-67, c-erbB-2 and p53 were more likely to recur (p = 0.0035, p = 0.0027, p = 0.0076 respectively), MRP expression was not associated with recurrence . Multivariate analysis showed that large tumors and high Ki-67 expression were independent indicators of recurrence . On the other hand, in tumors less than 1 cm, recurrence was significantly correlated with overexpression of Ki-67 and p53 . High Ki-67 expression could discriminate higher recurrence cases in grade 2, pT1 and single tumors . The c-erbB-2 overexpression was more frequently associated with recurrence in sessile tumors, large tumors, multiple and grade 1 tumors . The p53 overexpression also predicted a higher risk of recurrence in pTa tumors . These data demonstrated that the use of proliferative related proteins yields significant prognostic information in addition to clinicopathological factors, high Ki-67 expression is a reliable indicator of recurrence . A combination rather than any factor alone could more accurately predict tumor recurrence.

Int J Oncol, 2004 Feb, 24(2), 365 - 72
Effectiveness of Type I interferons in the treatment of multidrug resistant osteosarcoma cells; Manara MC et al.; P-glycoprotein overexpression is an important adverse prognostic marker for osteosarcoma (OS) patients, which is associated with higher risk for developing metastases as a consequence of the limited responsiveness to standard treatments of P-glycoprotein overexpressing OS cells . The use of cytokines has been advocated as a possible therapeutic approach to overcome multidrug resistance (MDR), being active on cell lines that are resistant to conventional drugs . In this study, we evaluated in vitro effects of interferons (IFNs) on MDR P-glycoprotein overexpressing OS cells . Type I IFNs, but not IFNgamma, showed tangible inhibitory effects on OS cell growth, which were higher in MDR cell lines compared to parental cells . The higher sensitivity of P-glycoprotein overexpressing cells to Type I IFNs correlates with higher expression of the activator of the transcription (STAT)-2 and (STAT)-3, two intracellular mediators of the IFNalpha and IFNbeta signaling pathways, whereas no differences were observed with respect to the expression or activation of the Type I IFN receptor and STAT-1 . Exposure of OS MDR cells to Type I IFN decreased the expression of P-glycoprotein . This effect resulted in a significantly increased chemosensitivity of MDR cells to doxorubicin . Therefore, our data support the use of IFNalpha or IFNbeta in the treatment of osteosarcoma patients who overexpress P-glycoprotein in their primary tumors, and respond insufficiently to current therapeutic regimens.

Br J Pharmacol, 2004 Feb, 141(3), 415 - 22 Epub 2004 Jan 12.
3,5-Dibenzoyl-4-(3-phenoxyphenyl)-1,4-dihydro-2,6-dimethylpyridine (DP7) as a new multidrug resistance reverting agent devoid of effects on vascular smooth muscle contractility; Saponara S et al.; The aim of this study was to investigate the effects of 3,5-diacetyl- (DP1-DP5) and 3,5-dibenzoyl-1,4-dihydropyridines (DP6-DP11) on vascular functions in vitro, by comparing their mechanical and electrophysiological actions in rat aorta rings and single rat tail artery myocytes, respectively, and to quantify their multidrug resistance (MDR)-reversing activity in L5178 Y mouse T-lymphoma cells transfected with MDR1 gene . In rat aorta, the 11 compounds tested, but 3,5-dibenzoyl-4-(3-phenoxyphenyl)-1,4-dihydro-2,6-dimethylpyridine (DP7), 3,5-dibenzoyl-4-(3-chlorophenyl)-1,4-dihydro-2,6-dimethylpyridine (DP9), 3,5-dibenzoyl-4-(4-chlorophenyl)-1,4-dihydro-2,6-dimethylpyridine (DP10) and 3,5-dibenzoyl-4-phenyl-1,4-dihydro-2,6-dimethylpyridine (DP11), antagonized 60 mm K+ (K60)-induced contraction in a concentration-dependent manner, with IC50 (m) values ranging between 5.65 x 10(-7) and 2.23 x 10(-5) . The 11 dihydropyridines tested, but DP7, inhibited L-type Ca2+ current recorded in artery myocytes in a concentration-dependent manner, with IC50 (M) values ranging between 1.12 x 10(-6) and 6.90 x 10(-5) . The K+ -channel opener cromakalim inhibited the Ca2+ -induced contraction in K30 but not that evoked in K60 . On the contrary, DP7 was ineffective in both experimental conditions . When the rings were preincubated with 1 mm Ni2+ plus 1 microm nifedipine, the response to phenylephrine was significantly reduced by 2,5-di-t-butyl-1,4-benzohydroquinone (BHQ), a well-known endoplasmic reticulum Ca2+ -ATPase inhibitor . DP7 had no effects on this model system . In L5178 MDR cell line, the 11 dihydropyridines tested, but 3,5-diacetyl-4-(2-nitrophenyl)-1,4-dihydro-2,6-dimethylpyridine (DP1), 3,5-diacetyl-4-(3-phenoxyphenyl)-1,4-dihydro-2,6-dimethylpyridine (DP2) and 3,5-diacetyl-4-(3-chlorophenyl)-1,4-dihydro-2,6-dimethylpyridine (DP4), exhibited an MDR-reversing activity, with IC50 values ranging between 3.02 x 10(-7) and 4.27 x 10(-5), DP7 being the most potent . In conclusion, DP7 may represent a lead compound for the development of potent dihydropyridine MDR chemosensitizers devoid of vascular effects.British Journal of Pharmacology (2004) 141, 415-422 . doi:10.1038/sj.bjp.0705635

Oncogene, 2004 Feb 26, 23(8), 1586 - 93
Apoptosis signaling triggered by the marine alkaloid ascididemin is routed via caspase-2 and JNK to mitochondria; Dirsch VM et al.; The marine alkaloid ascididemin (ASC) was shown to exert cytotoxicity even against multidrug-resistant cancer cells . Here, we address the signaling pathways utilized by ASC to trigger apoptosis in Jurkat leukemia T cells . We show that ASC (0.5-20 microM) induces a mitochondrial pathway that requires the activation of the initiator caspase-2 upstream of mitochondria . ASC-triggered apoptosis occurred independent of CD95, but required mitochondrial dysfunction . The activation of caspase-2 was shown to precede the processing of caspase-8, -9 and -3 . The specific caspase-2 inhibitor zVDVADfmk abrogated ASC-induced DNA fragmentation almost completely . Overexpression of Bcl-x(L) blocked caspase-8 but not caspase-2 processing . Conversely, caspase-2 inhibition strongly reduced caspase-9 activation . As a possible link between caspase-2 and mitochondrial dysfunction, Bid was found to be cleaved by ASC . In addition, JNK was activated by ASC upstream of mitochondria via reactive oxygen species . The specific JNK inhibitor SP600125 partially inhibited caspase-2 and -9 processing as well as cytochrome c release and DNA fragmentation indicating that JNK contributes to, but is not necessary for ASC-mediated apoptosis . Thus, ASC triggers a pathway in which early activation of caspase-2 provides a possible link between its DNA-damaging activity and the induction of mitochondrial dysfunction . The activation of JNK contributes to this signaling upstream of mitochondria.

J Clin Microbiol, 2004 Jan, 42(1), 212 - 9
IS6110 mediates increased transcription of the phoP virulence gene in a multidrug-resistant clinical isolate responsible for tuberculosis outbreaks; Soto CY et al.; Drug resistance in Mycobacterium tuberculosis complex strains is solely due to chromosomal mutations that could affect bacterial virulence . Molecular epidemiology studies have shown that resistant strains are less likely to be clustered than susceptible strains . However, a few multidrug-resistant (MDR) M . tuberculosis complex strains have been described as causing outbreaks, suggesting that they have restored virulence or increased transmission . One of the biggest MDR tuberculosis outbreaks documented to date was caused by the B strain of M . bovis . Restriction fragment length polymorphism fingerprinting revealed that the B strain contains two copies of IS6110 . Here, we mapped and sequenced the regions flanking the two copies of IS6110 in the B strain . Ligation-mediated PCR showed that one of these IS6110 copies is located within the promoter region of phoP, a transcriptional regulator that is essential for M . tuberculosis virulence . We used PCR to screen 219 MDR M . tuberculosis complex strains (90.4% of all MDR isolates) isolated in Spain between 1998 and 2002 and found that the B strain was the only strain that contained a copy of IS6110 in the phoP promoter . To determine whether IS6110 affects phoP promoter activity in the B strain, we individually cloned the phoP gene and its promoter region (including IS6110 from the B strain and the equivalent region from M . tuberculosis without IS6110 as a control) into a mycobacterial replicative plasmid and transformed M . smegmatis with the resulting plasmid . Primer extension analysis showed that phoP transcription was strongly upregulated when the promoter region contained IS6110, as in the case of the B strain.

Am J Physiol Heart Circ Physiol, 2004 May, 286(5), H2020 - 7 Epub 2004 Jan 08.
Regulation of rat pial arteriolar smooth muscle relaxation in vivo through multidrug resistance protein 5-mediated cGMP efflux; Xu HL et al.; Multidrug resistance protein 5 (MRP5) has been linked to cGMP cellular export in peripheral vascular smooth muscle cells (VSMCs) and is widely expressed in brain vascular tissue . In the present study, we examined whether knockdown of MRP5 in pial arterioles {via antisense oligodeoxynucleotide (ODN) applications} affected nitric oxide (NO)/cGMP-induced dilations . The antisense or (as a control) missense ODN was applied to the cortical surface approximately 24 h before study via closed cranial windows . The efficacy of the antisense vs . missense ODN in eliciting selective reductions in MRP5 expression was confirmed by analysis of MRP5 mRNA in pial tissue . Unexpectedly, in initial studies, a significantly lower maximal pial arteriolar diameter increase in the presence of the NO donor S-nitrosoacetylpenicillamine (SNAP) was seen in the antisense vs . missense ODN-treated rats (35 vs . 48% diameter increase, respectively) . It was suspected that this related to a reduced vascular smooth muscle cell sensitivity to cGMP due to prolonged exposure to increased intracellular cGMP levels elevated by overnight restriction of cGMP efflux . That postulate was supported by a finding of a diminished vasodilating response to the cGMP-dependent protein kinase-activating cGMP analog 8-p-chlorophenylthio-cGMP in antisense vs . missense ODN-treated rats . To prevent desensitization, additional rats were studied in the presence of chronic NOS inhibition via Nomega-nitro-L-arginine . In the NO synthase (NOS)-inhibited rats, the maximal SNAP response was much higher in the antisense (62% increase) vs . the missense ODN (40% increase) group . A similar result was obtained when monitoring responses to the soluble guanylyl cyclase-activating drugs YC-1 and BAY 41-2272 . Moreover, in the presence of NOS inhibition, the normal SNAP-induced rise in periarachnoid cerebrospinal fluid cGMP levels, which reflects cGMP efflux, was absent in the antisense ODN-treated rats, a finding consistent with loss of MRP5 function . In conclusion, if one minimizes the confounding effects of basal cGMP production, a clearer picture emerges, one that indicates an important role for MRP5-mediated cGMP efflux in the regulation of NO-induced cerebral arteriolar relaxation.

J Comb Chem, 2004 Jan-Feb, 6(1), 135 - 41
Synthesis of a library of 3-oxopiperazinium and perhydro-3-oxo-1,4-diazepinium derivatives and identification of bioactive compounds; Masip I et al.; The design and synthesis of a library of novel families of 3-oxopiperazinium and perhydro-3-oxo-1,4-diazepinium derivatives is reported . The library was composed of 44 3-oxopiperazinium derivatives (11 of these compounds had a spiranic skeleton) and 22 perhydro-3-oxo-1,4-diazepinium compounds . The synthetic procedure involved a 6-step sequence carried out in solution, along with the use of solid-phase linked scavengers and microwave activation for the rapid removal of the excess of amine reagents . A final cyclization step performed under mild conditions led to the charged heterocyclic moiety . Screening of this library in two biological assays identified active compounds that inhibit the activity of the vanilloid receptor TRPV1 and modulators of the multidrug resistance phenomenon . Thus, this synthetic sequence represents a facile and convenient entry to unprecedented libraries of this sort of tetraalkylammonium derivatives that may be of use for identification of novel scaffolds of diverse biological activity.

Dig Dis Sci, 2003 Dec, 48(12), 2315 - 22
Unique inhibition of bile salt-induced apoptosis by lecithins and cytoprotective bile salts in immortalized mouse cholangiocytes; Komichi D et al.; Bile duct epithelium is physiologically exposed to high concentrations of bile salts, suggesting the presence of a cytoprotective mechanism(s) . The aim of this study was to clarify whether bile salts cause bile duct cell damage and to elucidate the mechanism(s) providing protection against such an action of bile salts . Immortalized mouse cholangiocytes were incubated with taurocholate, taurochenodeoxycholate, glycochenodeoxycholate (GCDC), taurodeoxycholate, and tauroursodeoxycholate (TUDC), followed by flow-cytometric analysis and caspase activity assay to evaluate the induction of apoptosis . GCDC time-dependently induced caspase 3 (3.4-fold)- and caspase 9 (1.4-fold)-mediated apoptosis of cholangiocytes, but this was inhibited by lecithins and TUDC . Further, expression of cholangiocyte bile salt transporters (apical sodium-dependent bile salt transporter {Asbt} and multidrug resistance protein 3 {Mrp3}) was examined by RT-PCR and western blotting, and cholangiocyte bile salt uptake was determined using radiolabeled bile salts . Expression of cholangiocyte Asbt and Mrp3 was increased by bile salts, whereas lecithins interestingly reduced bile salt uptake to inhibit cholangiocyte apoptosis . In conclusion, bile salts themselves cause cholangiocyte apoptosis when absorbed by and retained inside the cell, but this is inhibited by washing out cytotoxic bile salts according to Mrp3, a rescue exporting molecule . Biliary lecithin is seemingly another cytoprotective player against cytotoxic bile salts, reducing their uptake, and this is associated with a reduced expression of Mrp3.

J Pharm Pharmacol, 2003 Nov, 55(11), 1531 - 7
Persistent reversal of P-glycoprotein-mediated daunorubicin resistance by tetrandrine in multidrug-resistant human T lymphoblastoid leukemia MOLT-4 cells; Liu ZL et al.; Multidrug resistance (MDR) represents a major problem in cancer chemotherapy . P-glycoprotein (P-gp), the drug efflux pump that mediates this resistance, can be inhibited by compounds with a variety of pharmacological functions, thus circumventing the MDR phenotype . The present study was performed to evaluate a unique MDR-reversal feature of a bisbenzylisoquinoline alkaloid tetrandrine (TET) in a P-gp expressing MOLT-4 MDR line (MOLT-4/DNR) established in our laboratory . Cell viability was determined by an MTT assay . P-gp function was characterized by determining the Rh123 accumulation/efflux capacity . P-gp overexpression in resistant MOLT-4/DNR cells was confirmed by flow cytometry analysis after staining with phycoerythrin-conjugated anti-P-gp monoclonal antibody 17F9 . Compared to ciclosporin A (CsA), TET exhibited stronger activity to reverse drug resistance to daunorubicin (DNR), vinblastine (VLB) and doxorubicin (DOX) in MOLT-4/DNR cells . TET showed no cytotoxic effects on parental MOLT-4 cells lacking P-gp expression or on the resistant MOLT-4/DNR cells . TET modulated DNR cytotoxicity even after it was washed with the medium for 24 h, while CsA almost completely lost its reversal capability 24 h after washing . TET and CsA similarly increased the accumulation of Rh123 in resistant MOLT-4/DNR cells . However, TET inhibited Rh123 efflux from resistant cells even after washing with the medium, while CsA rapidly lost its ability to inhibit Rh123 efflux after washing . The current study suggests that TET enhances the cytotoxicity of anticancer drugs in the P-gp expressing MDR cell line by modulating P-gp in a different manner to the well-known P-gp inhibitor CsA.

Leukemia, 2004 Mar, 18(3), 521 - 9
In vitro drug-resistance profile in infant acute lymphoblastic leukemia in relation to age, MLL rearrangements and immunophenotype; Ramakers-van Woerden NL et al.; Acute lymphoblastic leukemia (ALL) in infants under 1 year is strongly associated with translocations involving 11q23 (MLL gene), CD10-negative B-lineage (proB) immunophenotype, and poor outcome . The present study analyses the relationship between age, MLL rearrangements, proB-lineage, and in vitro drug resistance determined using the MTT assay . Compared to 425 children aged over 1 year with common/preB (c/preB) ALL, the 44 infants were highly resistant to steroids (for prednisolone (PRED) more than 580-fold, P=0.001) and L-asparaginase (L-ASP) (12-fold, P=0.001), but more sensitive to cytarabine (AraC) (1.9-fold, P=0.001) and 2-chlorodeoxyadenosine (2-CdA) (1.7-fold, P<0.001) . No differences were found for vincristine, anthracyclines, thiopurines, epipodophyllotoxines, or 4-hydroperoxy (HOO)-ifosfamide . ProB ALL of all ages had a profile similar to infant ALL when compared with the group of c/preB ALL: relatively more resistant to L-ASP and PRED (and in addition thiopurines), and more sensitive to AraC and 2-CdA . Age was not related to cellular drug resistance within the proB ALL group (<1 year, n=32, vs >/=1 year, n=19), nor within the MLL-rearranged ALL (<1 year, n=34, vs >/=1 year, n=8) . The translocation t(4;11)(q21;q23)-positive ALL cases were more resistant to PRED (>7.4-fold, P=0.033) and 4-HOO-ifosfamide (4.4-fold, P=0.006) than those with other 11q23 abnormalities . The expression of P-glycoprotein, multidrug-resistance protein, and lung-resistance protein (LRP) was not higher in infants compared to older c/preB ALL patients, but LRP was higher in proB ALL and MLL-rearranged ALL of all ages . In conclusion, infants with ALL appear to have a distinct in vitro resistance profile with the proB immunophenotype being of importance . The role of MLL cannot be excluded, with the t(4;11) being of special significance, while age appears to play a smaller role.

J Med Chem, 2004 Jan 15, 47(2), 285 - 7
Discovery of a bulky 2-tert-butyl group containing primaquine analogue that exhibits potent blood-schizontocidal antimalarial activities and complete elimination of methemoglobin toxicity; Jain M et al.; To eliminate an unwarranted metabolic pathway of the quinoline ring, a set of two compounds, where C-2 position of the antimalarial drug primaquine is blocked by metabolically stable bulky alkyl group are synthesized . Compound 2 {R = C(CH(3))(3)} of the series has produced excellent antimalarial efficacy against P . berghei and highly virulent multidrug-resistant P . yoelii nigeriensis strain in vivo . Compound 2 was also evaluated for methemoglobin (MetHb) toxicity . This study describes the discovery of a highly potent blood-schizontocidal antimalarial analogue 2, completely devoid of MetHb toxicity.

Drug Metab Dispos, 2004 Jan, 32(1), 35 - 42
The pregnane X receptor binds to response elements in a genomic context-dependent manner, and PXR activator rifampicin selectively alters the binding among target genes; Song X et al.; The pregnane X receptor (PXR) is a key regulator of genes encoding several major types of cytochrome P450 enzymes and transporters (e.g., multidrug resistance-1, MDR1); therefore, PXR contributes significantly to drug-drug interactions . PXR binds to response elements and confers transactivation . Several target genes such as CYP3A4 and 3A7 contain two PXR elements (distant and proximal) that are separated by more than 7000 nucleotides in the genome . Disruption of the distant element causes a 73% decrease of the reporter activity, whereas inactivation of the proximal element decreases by only 53% . This study was undertaken to test the hypothesis that PXR differentially binds to the elements with the distant enhancer being bound to a higher extent . To test this hypothesis, a stable transfected line (hPXR-HRE) was prepared to constitutively express human PXR and harbor a chromatinized CYP3A4-ER6 reporter . This line responded to rifampicin and dexamethasone similarly as hepatocytes based on the relative potency and activation kinetics . Contrary to the hypothesis, chromatin immunoprecipitation experiments showed that the genomic fragment harboring the proximal element was preferably precipitated over the fragment containing the distant element in the CYP3A4 gene, but the opposite was true with the CYP3A7 gene . In addition, the promoters from the MDR1 and CYP2B6 genes were abundantly present in the PXR immunocomplexes from the vehicle-treated cells . However, such abundant interactions were markedly diminished when cells were treated with PXR activator rifampicin . These findings suggest that PXR binding is dependent on the genomic context and PXR activators modulate such bindings.

Drug Metab Dispos, 2004 Jan, 32(1), 20 - 7
Suppression of drug-metabolizing enzymes and efflux transporters in the intestine of endotoxin-treated rats; Kalitsky-Szirtes J et al.; Infection and inflammation impose a suppression in the expression and activity of several drug transporters and drug-metabolizing enzymes in liver . In the intestine, cytochrome P450 3A (CYP3A), P-glycoprotein (PGP/mdr1), and the multidrug resistance-associated protein 2 (MRP2) are important barriers to the absorption of many clinically important drugs; thus, the expression and activity of these proteins were examined in inflammation . Transport and metabolism were determined in jejunum segments isolated at 24 h from endotoxin-treated or control rats (n = 8) mounted in Ussing chambers . Transport and metabolism of (3)H-digoxin, 5-carboxyfluorescein (5-CF), amiodarone (AM), and 7-benzyloxyquinoline (7-BQ) were measured for 90 min in the presence and absence of inhibitors . Reverse transcription-polymerase chain reaction was used to measure mRNA levels . As compared with controls, levels of mdr1a and mrp2 mRNA were significantly decreased by approximately 50% in the jejunum of LPS-treated rats . Corresponding reductions in the basolateral-->apical efflux of digoxin, AM, and 5-CF were observed, resulting in significant increases in the apical-->basolateral absorption of these compounds . Intestinal CYP3A mRNA levels and CYP3A-mediated metabolism of 7-BQ and AM were also decreased by approximately 50 to 70% (p < 0.05) in the LPS group . Mannitol permeability and lactate dehydrogenase release were not altered . These studies indicate that endotoxin-induced inflammation imposes a reduction in the intestinal expression and activity of PGP, mrp2, and CYP3A in rats, which elicits corresponding changes in the intestinal transport and metabolism of their substrates . Hence, infection and inflammatory diseases may impose variability in drug bioavailability through alterations in the intestinal expression and activity of drug transporters and metabolic enzymes.

Eur J Pharmacol, 2004 Jan 1, 483(1), 19 - 28
Induction of proteins involved in multidrug resistance (P-glycoprotein, MRP1, MRP2, LRP) and of CYP 3A4 by rifampicin in LLC-PK1 cells; Magnarin M et al.; P-glycoprotein, multidrug resistance-related proteins (MRPs) and lung resistance-related protein (LRP) are involved in multidrug resistance in tumor cells but are also expressed in normal tissues . In the LLC-PK(1) tubular renal cell line, a 15-day treatment with 25 microM rifampicin significantly increased the mRNA levels of P-glycoprotein, MRP1, MRP2, LRP and cytochrome P450 3A4 (CYP 3A4) . Western blot analysis confirmed a moderate increase in the expression of P-glycoprotein and MRP2, but not MRP1 also at the protein level . The intracellular uptake of doxorubicin was significantly lower in rifampicin pretreated cells . A pretreatment with 6-{82S,4R,6E)-4-methyl-2-(methylamino)-3-oxo-6-octenoic acid}cyclosporin D, valspodar (PSC 833), a specific inhibitor of P-glycoprotein, with (3-(3-(2-(7-chloro-2-quinidinyl)ethenyl-phenyl)((3-diimethyl amino-3oxo propyl)thio)methyl)thio)propanoic acid, sodium salt (MK-571), a specific inhibitor of MRP1, and with verapamil, that inhibits both proteins, significantly increased doxorubicin cell accumulation in rifampicin pretread cells . In rifampicin treated cells cultured on porous membranes, doxorubicin showed a polarized transport, that was reduced by a pretreatment with PSC 833 . A chronic treatment with rifampicin induces the expression of transport proteins and of CYP 3A4 and could therefore alter the renal elimination kinetics of drugs that are their substrates.

Pancreas, 2004 Jan, 28(1), 45 - 52
Expression of multidrug resistance proteins in rat and human chronic pancreatitis; Schaarschmidt T et al.; BACKGROUND AND AIMS: The expression of the ABC-transporters MDR-1, MRP1, and MRP-2 was investigated in healthy pancreas and in chronic pancreatitis tissue samples in rats and humans to evaluate their possible involvement in a multidrug resistance of the pancreas with consequences for the pharmacologic treatment of pancreatic diseases . METHODS: Human pancreatic tissue samples of healthy tissue and chronic pancreatitis were collected during pancreas surgery . In rats, the time-course of the expression of transporter proteins was studied 14, 28, and 56 days after experimental induction of chronic pancreatitis . The expression of MDR-1, MRP-1, MRP-2, and furthermore, LRP and PAP was investigated by RT-PCR, Real Time TaqManPCR, and immunohistochemistry . RESULTS: In rat pancreas, MDR-1 (P-gp) and MRP-1 but in human pancreas MDR-1 (P-gp), MRP-1 and MRP-2 were found to be expressed . Chronic pancreatitis lead to an increased transcription of mRNA of MDR-1 (rat and human) and much lower, MRP-2 (human) . CONCLUSIONS: The expression of P-gp and related transporters could have impact on the metabolism, distribution, and availability of various compounds, including drugs, in the pancreas . The results indicate that this could be more pronounced in chronic pancreatitis.

Mol Cancer Ther, 2003 Dec, 2(12), 1351 - 9
Genetic polymorphism at the 5' regulatory region of multidrug resistance 1 (MDR1) and its association with interindividual variation of expression level in the colon; Taniguchi S et al.; The multidrug resistance 1 (MDR1) is a key molecule in determining not only the resistance of cancer cells to anticancer agents but also the disposition of a variety of drugs in intestinal and other tissues . However, the mechanism underlying interindividual variations in levels of MDR1 activity and expression in various tissues remains unclear . We analyzed the nucleotide sequence polymorphisms in the 5' upstream regulatory region of the gene spanning 4 kb from the transcriptional start site of MDR1 and tried to identify any associations between polymorphisms and MDR1 expression . Within that region, we identified eight single nucleotide polymorphisms (SNPs) in the region in the Japanese population . Of the SNPs identified, -2410T>C, -1910T>C, and 692T>C were in perfect linkage disequilibrium . In normal colorectal mucosa, diplotypes at the region showed more significant association with the expression level of MDR1 mRNA than each SNP did . In an in vitro reporter assay, transcription activity of the minor-type construct carrying haplotypes 2 and 3 was significantly lower than that of the major-type construct carrying haplotype 1 . We next identified two DNA binding proteins: one protein bound to the nucleotide sequence carrying -692T but not to that carrying -692C and another bound to the nucleotide sequence carrying -2352G but three times weaker than that carrying -2352A . This suggested the significance of SNP at -692 and -2352 of MDR1 in variable expression in the colon interindividually . This is the first report connecting SNPs and interindividual variety of MDR1 expression rationally.

Am J Surg, 2004 Jan, 187(1), 134 - 45
A consensus statement on empiric therapy for suspected gram-positive infections in surgical patients; Solomkin JS et al.; BACKGROUND: Multidrug resistance among gram-positive pathogens in tertiary and other care centers is common . A systematic decision pathway to help select empiric antibiotic therapy for suspected gram-positive postsurgical infections is presented . DATA SOURCES: A Medline search with regard to empiric antibiotic therapy was performed and assessed by the 15-member expert panel . Two separate panel meetings were convened and followed by a writing, editorial, and review process . CONCLUSIONS: The main goal of empiric treatment in postsurgical patients with suspected gram-positive infections is to improve clinical status . Empiric therapy should be initiated at the earliest sign of infection in all critically ill patients . The choice of therapy should flow from beta-lactams to vancomycin to parenteral linezolid or quinupristin-dalfopristin . In patients likely to be discharged, oral linezolid is an option . Antibiotic resistance is an important issue, and in developing treatment algorithms for reduction of resistance, the utility of these new antibiotics may be extended and reduce morbidity and mortality.

Zhongguo Shi Yan Xue Ye Xue Za Zhi, 2003 Dec, 11(6), 604 - 8
{Reversal effect of berbamine on multidrug resistance of K562/A02 cells and its mechanism}; Han YQ et al.; This preject is to explore the reversal efficacy of calmodulin antagonist berbamine (BBM) on multidrug resistance (MDR) and its mechanism . Human erythroleukemic cell line K562 and its adriamycin-resistant counterpart K562/A02 were used in the study . The cells were co-cultured with ADR and BBM in different concentrations . MTT assay was used to analyze the effect of BBM on cell growth inhibition . According to the MTT assay, the 50% inhibitory concentration (IC(50)), the multiples of drug resistance and increased sensitivity of ADR were calculated . The concentration of intracellular ADR and expression level of P-gp were detected by flow cytometry (FCM) . The expression level of mdr1 mRNA and survivin mRNA was detected by semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) with beta-actin as internal reference . The results showed that IC(50) of ADR in K562 and K562/A02 cells was 1.16 +/- 0.09 micro mol/L and 37.47 +/- 1.76 micro mol/L, respectively . The resistant multiple of K562/A02 cells to ADR was 32.30 higher than that of K562 cells . BBM increased the chemo-sensitivity of ADR in K562/A02 cells with dose-dependent relationship, i.e . when 5, 10 and 20 micro mol/L BBM was added in the culture the chemo-sensitivity of ADR was increased to 2.01-, 9.68-, and 41.18-fold (P < 0.01), respectively . After treating K562/A02 cells by 5 or 10 micro mol/L BBM for 2 hours the accumulation of intracellular ADR was increased to 1.41- and 1.52-fold (P < 0.01), respectively . Treating by BBM for 72 hours decreased 4.12% (P < 0.05) and 27.09% (P < 0.01) of P-gp expression, respectively, meanwhile down-regulated expression of mdr1 mRNA and survivin mRNA was found . In conclusion, BBM could increase intracellular concentration of ADR in K562/A02 that down-regulated expression level of mdr1 mRNA and P-gp and survivin so that the sensitivity of K562/A02 to ADR was increased significantly.

Drug Metab Rev, 2003 Nov, 35(4), 417 - 54
Clinical relevance of P-glycoprotein in drug therapy; Lin JH et al.; The drug efflux transporter P-glycoprotein (P-gp) is known to confer multidrug resistance in cancer chemotherapy . The P-gp is highly expressed in many types of tumor cells, as well as many normal tissues, including the apical surface of intestinal epithelial cells, and the luminal surface of capillary endothelial cells in the brain . Because of its expression and localization, it has been suggested that P-gp plays an important role in cancer chemotherapy, intestinal absorption, and brain uptake . This review addresses the significance of the role of P-gp in cancer chemotherapy, drug absorption, and brain uptake . Based on the clinical and animal studies with P-gp modulators, it has become apparent that the role of P-gp in multidrug resistance is far less important compared to other biological factors . Although P-gp is highly expressed in both intestinal epithelial cells and endothelial cells of brain capillaries and functions as an efflux transporter in both organs, the magnitude of P-gp's impact on intestinal absorption and brain uptake of drugs is quantitatively very different . From animal and clinical studies, it is evident that P-gp plays a very important role in CNS penetration of drugs, whereas the effect of P-gp on drug absorption is not as important as generally believed.

Drug Metab Rev, 2003 Nov, 35(4), 305 - 17
Regulation of drug and bile salt transporters in liver and intestine; Kullak-Ublick GA et al.; Major determinants of the bioavailability of drugs are the degree of intestinal absorption and the hepatic first-pass effect . Drugs need to overcome several membrane barriers before reaching the systemic circulation, each of which expresses an array of specialized transport proteins for drug uptake or efflux . The P-glycoprotein MDR1 (multidrug resistance gene product, ABCB1) is expressed at the apical surface of enterocytes, where it mediates the efflux of xenobiotics into the intestinal lumen before these can access the portal circulation . Increased expression of MDR1 reduces the bioavailability of MDR1 substrates such as digoxin, cyclosporin, and taxol . Numerous xenobiotics can induce the MDR1 gene through activation of the nuclear pregnane X receptor (PXR) . This explains the risk for drug interactions that is inherent to pharmacotherapy with PXR ligands such as rifampin, phenobarbital, statins, and St . John's wort . Other PXR-regulated genes include cytochrome P450 3A4, the digoxin and bile salt transporter Oatp2 (organic anion transporting polypeptide 2, Slc01a4) of the basolateral hepatocyte membrane, and the xenobiotic efflux pump Mrp2 (multidrug resistance associated protein 2, Abcc2) of the canalicular hepatocyte membrane . A second orphan nuclear receptor that is activated by xenobiotics is the constitutive androstane receptor (CAR), which induces Mrp2 and Mrp3 (Abcc3) . The PXR and CAR are thus important "xenosensors" that mediate drug-induced activation of the detoxifying transport and enzyme systems in liver and intestine.

Med Sci Monit, 2004 Jan, 10(1), RA5 - 14
Multidrug resistance P-glycoprotein: crucial significance in drug disposition and interaction; Sun J et al.; This article reviews recent advances in our understanding of the structure, drug interaction mechanism, and substrate molecular requirements of P-glycoprotein (P-gp) and its emerging crucial role in drug disposition and the modulation of drug interaction . In view of its wide localization in normal tissues, the broad variety of structurally and functionally unrelated substrates of P-gp, and its ATP-dependent outward-oriented transport, P-gp actively participates in intestinal secretion, blood-tissue barriers, and biliary and renal excretions for many exogenous substrates, and also performs a protective role to prevent entry of xenobiotics . Moreover, the importance of P-gp-mediated drug interactions in clinical practice can hardly be underestimated, since it may result in severe side effects, such as digitalis drug interaction . Polymorphism or single nucleotide polymorphism (SNP) associated with P-gp may exert a significant effect on the pharmacokinetic behavior of its substrates, a fact which has major clinical implications and suggests careful dose adjustment for individual treatment . Moreover, dietary components and pharmaceutical excipients may modulate P-gp activity, and as a result affect in vivo drug disposition and therapeutic efficacy; examples include grapefruit juice, Pluronic P85, PEG 300, etc . In summary, it should be emphasized that P-gp is an integral component in the process of drug discovery, development strategy,

Clin Lung Cancer, 2001 Feb, 2(3), 220 - 6
Potential clinical application of antioncogene ribozymes for human lung cancer; Tong AW et al.; Non-small-cell lung cancer frequently contains oncogenetic defects (mutations in ras, retinoblastoma, and p53 genes) that contribute to disease pathophysiology . Recent studies and clinical trials have focused on gene therapy approaches that either replace the function of defective tumor-suppressor genes such as p53 or inactivate mutant oncogenes such as ras . Ribozymes are RNA molecules with highly specific intrinsic enzymatic activity against target RNA sequences, which can discriminate mutant sequences that differ by a single base from their wild-type counterparts . Following binding to the RNA substrate by base-pair complementation, the ribozyme cleaves the target RNA irreversibly, then releases itself for new rounds of subsequent cleavage, resulting in significantly improved target:effector stoichiometry as compared with antisense oligonucleotides of the same specificity . Transcript-specific ribozymes have been used extensively for experimental oncogene inactivation . Ribozymes are effective for targeting mutant ras, p53, or the multidrug-resistant gene product for lung cancer cells in vitro . However, their in vivo effect is not well defined against this malignancy . We recently characterized the antitumor properties of an anti-K-ras ribozyme specific for the K-ras codon 12 mutation (GGT-->GTT) . When delivered as a transgene by an adenoviral vector (ADV), the K-ras ribozyme (KRbz) suppressed growth of lung tumor xenografts expressing the relevant mutation, whereas the corresponding antisense sequence lacking catalytic activity did not . Multiple intratumoral (3-5) injections of KRbz-ADV were effective in producing complete tumor regressions of preexisting tumor xenografts . Clinical trials are under consideration to examine the applicability of this anti-K-ras ribozyme for treatment of non-small-cell lung cancers expressing the relevant mutation.

Gastroenterology, 2004 Jan, 126(1), 322 - 42
Enterohepatic bile salt transporters in normal physiology and liver disease; Kullak-Ublick GA et al.; The vectorial transport of bile salts from blood into bile is essential for the generation of bile flow, solubilization of cholesterol in bile, and emulsification of lipids in the intestine . Major transport proteins involved in the enterohepatic circulation of bile salts include the hepatocellular bile salt export pump (BSEP, ABCB11), the apical sodium-dependent bile salt transporter (ASBT, SLC10A2) in cholangiocytes and enterocytes, the sodium-dependent hepatocyte bile salt uptake system NTCP (SLC10A1), the organic anion transporting polypeptides OATP-C (SLC21A6), OATP8 (SLC21A8) and OATP-A (SLC21A3), and the multidrug resistance protein MRP3 (ABCC3) . Synthesis and transport of bile salts are intricately linked processes that undergo extensive feedback and feed-forward regulation by transcriptional and posttranscriptional mechanisms . A key regulator of hepatocellular bile salt homeostasis is the bile acid receptor/farnesoid X receptor FXR, which activates transcription of the BSEP and OATP8 genes and of the small heterodimer partner 1 (SHP) . SHP is a transcriptional repressor that mediates bile acid-induced repression of the bile salt uptake systems rat Ntcp and human OATP-C . A nuclear receptor that activates rodent Oatp2 (Slc21a5) and human MRP2 (ABCC2) is the pregnane X receptor/steroid X receptor PXR/SXR . Intracellular trafficking and membrane insertion of bile salt transporters is regulated by lipid, protein, and extracellular signal-related kinases in response to physiologic stimuli such as cyclic adenosine monophosphate or taurocholate . Finally, dysfunction of individual bile salt transporters such as BSEP, on account of genetic mutations, steric inhibition, suppression of gene expression, or disturbed signaling, is an important cause of cholestatic liver disease.

Thorax, 2004 Jan, 59(1), 79 - 80
Multidrug resistant tuberculosis following lung transplantation: treatment with pulmonary resection; Shitrit D et al.; Recipients of organ transplants are at increased risk for infection owing to their immunosuppressed state and the possibility of contamination of the donor organ . We report a case of multidrug resistant tuberculosis (MDR) transmission via a donor lung . After medical treatment with four drugs had failed, the patient underwent right upper lobectomy . There were no signs of disease on follow up more than 2 years later . To our knowledge, this is the first report of MDR tuberculosis in a lung transplant recipient . The need for a non-conservative approach, including pulmonary resection, to eradicate the infection is emphasised.

Ai Zheng, 2003 Dec, 22(12), 1339 - 42
{Expression of P-glycoprotein and lung resistance protein in breast carcinoma and its relationship with prognosis}; Yu P et al.; BACKGROUND & OBJECTIVE: The multidrug resistance of tumor cells is the reason to failure of chemotherapy . The overexpression of P-glycoprotein (P-gp) and lung resistance protein (LRP) play important roles in the multidrug resistance of breast carcinoma . The objective of this study was to investigate the expression of P-gp and LRP in breast carcinoma and its relationship with prognosis . METHODS: Immunohistochemical assay with the LSAB immunostaining technique was used to determine protein expression of P-gp and LRP in the specimens of paraffin-embedded tissues from 151 cases of breast carcinoma and 40 normal breast tissues (control) . The results were evaluated on the basis of clinical pathological factors and the survival time of these cases . RESULTS: The expression of P-gp were found in 62 (40.8%) of the breast carcinoma samples and in 1 (2.5%) of control samples . The difference between the two groups was statistically significant (Chi(2)=21.27,P< 0.01) . The expression of LRP were found in 121 (80.1%) of the breast carcinoma samples and in 22 (55.0%) of control samples . The difference between the two groups was statistically significant (Chi(2)=10.62,P< 0.01) . The expression of P-gp were irrelative to age,tumor size,histotype,lymph nodes metastasis,tissue grading,clinical staging, ER condition,and prognosis.The expression of LRP were found in 62 (88.6%) of 70 cases with positive lymph nodes and 59 (72.8%) of 81 cases with negative lymph nodes . The difference between the two groups was statistically significant (Chi(2)=4.891, P< 0.05) . The expression of P-gp were found in 55 (45.5%) of breast carcinomas with positive immunohistochemical reaction of LRP protein; while the expression of P-gp were found in 7(23.3%) of breast carcinomas with negative immunohistochemical reaction of LRP protein . The difference between the two groups was statistically significant (Chi(2)=3.990,P< 0.05).The expression of LRP were positively correlated with the lymph nodes metastases (r=0.197, P< 0.05) and the expression of P-gp (r=0.179,P< 0.05) and irrelative to age tumor size, histotype, lymph nodes metastases, tissue grading,clinical staging, and ER condition . Univariate analysis demonstrated worse disease-free survival in the patients with positive LRP than in those with negative LRP (Chi(2)=6.892,P< 0.01) . Multivariate analysis of clinical and pathological data by Cox regression method did not demonstrated that both expression of P-gp and LRP were independent factors affecting the survival of breast carcinoma patients . CONCLUSION: The expression of P-gp and LRP have their characteristics and dissimilarity clinical significance in breast carcinoma tissues . The expression of LRP may correlate not only with MDR but also the lymph nodes metastases,prognosis and expression of P-gp in breast carcinoma tissues.

Ai Zheng, 2003 Dec, 22(12), 1284 - 8
{Reversal of multidrug resistance of human hepatocarcinoma HepG2/Adm cells by high intensity focused ultrasound}; Zhai BJ et al.; BACKGROUND & OBJECTIVE: The effects of ultrasound have been shown on increasing cell membrane permeability and thereby might promote cellular uptake of cytotoxic drugs and enhance chemotoxicity . The aim of this study was to examine the reversal effects of high intensity focused ultrasound (HIFU) and co-administration with Adriamycin (ADM) on multidrug resistance (MDR) in HepG2/Adm cells, and to investigate its mechanism . METHODS: HepG2 cells and HepG2/Adm cells were divided into 4 groups: HepG2 (control group),HepG2 (treated with HIFU 5s),HepG2/Adm (non HIFU), and HepG2/Adm (treated with HIFU 5s) . MTT assay was used to examine the chemosensitivity to ADM in HIFU-treated HepG2 and HepG2/Adm cell lines . Before and after the treatment with HIFU 5s, MDR1 gene expression was determined by immunocytochemistry assay . Flow cytometry was used to determine intracellular ADM concentration and the expression of P-gp on cell membranes . RESULTS: HIFU (0.8MHz, 460W/cm2) partly overcame the MDR of cell line HepG2/Adm and decreased the drug resistance . The relative reverse efficiencies to ADM, cisplatin (DDP), mytomycin (MMC), 5-fluorouracil (5-FU), and methotrexate (MTX) were 66.4%, 63.4%, 89.4%, 72.4%, and 75.0%, respectively . While, the concentration of ADM in HepG2/Adm cells was increased by 88% . The expression of P-glycoprotein (P-gp) protein in HepG2/Adm was down-regulated by 83.1% . CONCLUSION: HIFU can not only increase the ADM concentration in HepG2/Adm cells and decrease P-gp expression on cellular and nuclear membrane, but also partly overcome MDR of tumor cells.

Addict Biol, 2003 Dec, 8(4), 413 - 8
P-glycoprotein modulation by the designer drugs methylenedioxymethamphetamine, methylenedioxyethylamphetamine and paramethoxyamphetamine; Ketabi-Kiyanvash N et al.; There are increasing numbers of deaths related to taking MDMA, MDE and PMA reported where the deceased typically took several different drugs with these compounds . Hence, mutual modulation of the pharmacokinetics in drug combinations with "ecstasy" might be a risk factor for "ecstasy"-related morbidity . Regarding potential drug - drug interactions, there are no data evaluating a possible contribution of the multidrug resistance transporter P-glycoprotein (Pgp) in contrast to the cytochrome P450 enzyme system . Therefore, individual "ecstasy" compounds have been tested for their ability to interact with Pgp using a fluorometric calcein assay as a model for Pgp inhibition in porcine kidney epithelial cells with overexpression of human Pgp (L-MDR1) . All three compounds increased calcein retention in L-MDR1 cells in a concentration-dependent manner, with MDE being the most potent and MDMA the weakest Pgp inhibitor . The effective concentrations were 1 - 3 orders of magnitude higher than plasma concentrations observed in vivo, suggesting that these compounds are only weak inhibitors of Pgp, which is unlikely to influence the access of other compounds to the brain . However, it cannot be excluded that co-administration of Pgp inhibitors such as ritonavir or paroxetine could increase MDMA, MDE and PMA bioavailability and also enhance brain entry leading to severe side effects.

J Hum Genet, 2004, 49(1), 40 - 5 Epub 2003 Dec 18.
Common single nucleotide polymorphisms of the MDR1 gene have no influence on its mRNA expression level of normal kidney cortex and renal cell carcinoma in Japanese nephrectomized patients; Uwai Y et al.; In this study, we have quantified the mRNA expression levels of multidrug resistance gene 1 (MDR1) in the normal kidney cortex and renal cell carcinoma (RCC) segments from 24 Japanese nephrectomized patients by real-time polymerase chain reaction (PCR) . The mRNA expression level of MDR1 in RCC segments was significantly decreased in comparison with each normal segment (P=0.0042, by Student's paired t-test) . In addition, the ten common single nucleotide polymorphisms (SNPs) of the MDR1 gene in the patients were assessed using the PCR-restriction enzyme fragment length polymorphism method to investigate the influence of these SNPs on its mRNA expression levels . The allele frequencies of these SNPs were comparable with our previous report in the Japanese recipients of living-donor liver transplantation (Goto et al., Pharmacogenetics 12:451-457; 2002) . MDR1 expression levels in the normal kidney cortex were independent on the five SNPs, which were polymorphic in the Japanese population . Furthermore, the effect of the SNPs on expression levels of MDR1 mRNA in RCC segments was not recognized . These findings suggest that the common SNPs in the MDR1 gene have no influence on the expression of its transcript in RCC segments as well as in the normal kidney cortex.

Rev Port Pneumol, 2003 Mar-Apr, 9(2), 99 - 107
{The cost of tuberculosis care: in-patient estimated costs}; Gomes C et al.; OBJECTIVE: To calculate the costs of in hospital tuberculosis treatment and to compare the estimated charges of multidrug-resistant tuberculosis (MDR-TB) and drug susceptible tuberculosis patients (TB) . DESIGN: Descriptive study . SETTING: Tuberculosis Unit (Pulido Valente Hospital), Lisbon, Portugal . METHODS: The records of all TB patients discharged between January and December 2000 were reviewed . The cost analysis was conducted by using the hospital cost accounting system data and the charges approved by the national public health system . The main outcome measures were the following costs: 1) laboratory and ancillary services, 2) medication and 3) other direct and indirect components . RESULTS: The 116 study patients were divided into 3 groups: HIV/TB-48 (41.4%), TB 62 (53.4%) and MDR-TB-6 (5.2%) . The estimated cost of treatment for all patients was PTE 213,732.769, but only 42% was covered by diagnosis-related groups financing system . In the MDR-TB group, the median cost per day (PTE 7,531) and the median cost per episode (PTE 316,593) were significantly higher comparing with the TB group, regarding laboratory services and medication items . CONCLUSIONS: The hospital care budget based on the diagnosis-related groups financing system, accounted for less than half the estimated costs of tuberculosis in-patients . Also, in regarding tuberculosis the diagnosis-related groups system does not account for a major confounder like MDR-TB, which has an huge impact on the length of hospital stay and increasing laboratory and medication costs.

Rev Port Pneumol, 2003 May-Jun, 9(3), 195 - 204
{Immigration and tuberculosis . Five year experience}; Rifes G et al.; Immigrants are a tuberculosis risk group . In Portugal, in 2000, they had an incidence rate 3.6 times higher than the global incidence, and were native, predominantly, from Angola and Cabo Verde . Being the Chest Disease Center of Venda Nova located in a residential area with a great number of immigrants, most of them living in slums, we decided to evaluate the tuberculosis cases in this group, between 1996/2000, comparing the data obtained with some data of the tuberculosis cases in the non-immigrants . Immigrants with tuberculosis corresponded to 24.5% of all cases, 71.4% male, 93.9% black and mostly native from Cabo Verde and Angola . 73% lived in Portugal for more than 5 years, 86.7% were new cases and 13.3% relapses . 174 were pulmonary forms, 70.7% of which were D+ and 81% confirmed (against 75% in the non-immigrants) . Of the 91 drug susceptibility tests done in the pulmonary forms, 9.9% revealed multidrug resistance, against 5% in the non-immigrants . Twenty six point six percent had AIDS against 18% in the non-immigrants . Some conclusions: important percentage of immigrants with tuberculosis in the Chest Disease Center of Venda Nova; immigrants have a higher confirmation rate of pulmonary tuberculosis, more multidrug resistance and AIDS cases.

Plant Physiol, 2004 Jan, 134(1), 528 - 38 Epub 2003 Dec 18.
Binding of sulfonylurea by AtMRP5, an Arabidopsis multidrug resistance-related protein that functions in salt tolerance; Lee EK et al.; Recently, a new member of the ABC transporter superfamily of Arabidopsis, AtMRP5, was identified and characterized . In the present work, we found that AtMRP5 can bind specifically to sulfonurea when it is expressed in HEK293 cells . We also present evidence for a new role of AtMRP5 in the salt stress response of Arabidopsis . We used reverse genetics to identify an Arabidopsis mutant (atmrp5-2) in which the AtMRP5 gene was disrupted by transferred DNA insertion . In root-bending assays using Murashige and Skoog medium supplemented with 100 mm NaCl, root growth of atmrp5-2 was substantially inhibited in contrast to the almost normal growth of wild-type seedlings . This hypersensitive response of the atmrp5-2 mutant was not observed during mannitol treatment . The root growth of the wild-type plant grown in Murashige and Skoog medium supplemented with the MRP inhibitor glibenclamide and NaCl was inhibited to a very similar extent as the root growth of atmrp5-2 grown in NaCl alone . The Na(+)-dependent reduction of root growth of the wild-type plant in the presence of glibenclamide was partially restored by diazoxide, a known K+ channel opener that reverses the inhibitory effects of sulfonylureas in animal cells . Moreover, the atmrp5-2 mutant was defective in 86Rb+ uptake . When seedlings were treated with 100 mm NaCl, atmrp5-2 seedlings accumulated less K+ and more Na+ than those of the wild type . These observations suggest that AtMRP5 is a putative sulfonylurea receptor that is involved in K+ homeostasis and, thus, also participates in the NaCl stress response.

Zhonghua Wai Ke Za Zhi, 2003 Sep, 41(9), 676 - 8
{To set up a mdrl multidrug resistant model of orthotopic transplantation of liver carcinoma in nude mice}; Han Y et al.; OBJECTIVE: To set up a mdr1 multidrug resistant model of orthotopic transplantation of liver carcinoma in nude mice by intermittent abdominal chemotherapy . METHODS: The hepatocellular carcinoma cell line HepG2 was first cultured, then subcutaneous carcinoma was produced to form the tumor donor mice . An orthotopic mdr1 hepatoma was produced by implanting the tumor fragment subserosally to the mice liver, and intermittent chemotherapy with Pharmorubicin given to induce drug resistance . Physical examination, ultrasonography, spiral CT and laparotomy were used to examine the growth of the tumor . RT-PCR and SP method by the monoclonal antibody JSB-1 were adoped to detect the expression of mdr1-mRNA and p-gp protein . RESULTS: There was no mortality . The successful rate of tumor implantation is 88% (22/25), the successful rate of supplementary implantation was 100% (3/3), and the successful rate of induction of drug resistance was 80% (16/20) . The expressions of mdr1-mRNA and p-gp in the Pharmorubicin group were 23 folds and 13 folds of the control group, respectively . CONCLUSIONS: The nude mice mdr1 multidrug resistant model of orthotopic liver carcinoma were set up successfully, with its features similar to clinical cases . It would be helpful for the further study of the diagnosis and reversal strategy of the MDR phenomenon.

Chem Res Toxicol, 2003 Dec, 16(12), 1642 - 51
Interplay between MRP inhibition and metabolism of MRP inhibitors: the case of curcumin; Wortelboer HM et al.; The multidrug resistance proteins MRP1 and MRP2 are efflux transporters with broad substrate specificity, including glutathione, glucuronide, and sulfate conjugates . In the present study, the interaction of the dietary polyphenol curcumin with MRP1 and MRP2 and the interplay between curcumin-dependent MRP inhibition and its glutathione-dependent metabolism were investigated using two transport model systems . In isolated membrane vesicles of MRP1- and MRP2-expressing Sf9 cells, curcumin clearly inhibited both MRP1- and MRP2-mediated transport with IC(50) values of 15 and 5 microM, respectively . In intact monolayers of MRP1 overexpressing Madin-Darby canine kidney (MDCKII-MRP1) cells, curcumin also inhibited MRP1-mediated activity, although with a 3-fold higher IC(50) value than the one observed in the vesicle model . Interestingly, MRP2-mediated activity was hardly inhibited in intact monolayers of MRP2-overexpressing MDCKII (MDCKII-MRP2) cells upon exposure to curcumin, whereas the IC(50) value in the vesicle incubations was 5 microM . The difference in extent of inhibition of the MRPs by curcumin in isolated vesicles as compared to intact cells, observed especially for MRP2, was shown to be due to a swift metabolism of curcumin to two glutathione conjugates in the MDCKII cells . Formation of both glutathione conjugates was about six times higher in the MDCKII-MRP2 cells as compared with the MDCKII-MRP1 cells, a phenomenon that could be ascribed to the significantly lower glutathione levels in the cell line . The efflux of both conjugates, identified in the present study as monoglutathionyl curcumin conjugates, was demonstrated to be mediated by both MRP1 and MRP2 . From dose-response curves with Sf9 membrane vesicles, glutathionylcurcumin conjugates appeared to be less potent inhibitors of MRP1 and MRP2 than their parent compound curcumin . In conclusion, curcumin clearly inhibits both MRP1- and MRP2-mediated transport, but the glutathione-dependent metabolism of curcumin plays a crucial role in the ultimate level of inhibition of MRP-mediated transport that can be achieved in a cellular system . This complex interplay between MRP inhibition and metabolism of MRP inhibitors, the latter affecting the ultimate potential of a compound for cellular MRP inhibition, may exist not only for a compound like curcumin but also for many other MRP inhibitors presently or previously developed on the basis of vesicle studies.

Semin Respir Infect, 2003 Dec, 18(4), 292 - 306
Mycobacterium tuberculosis: the treatment of active disease; Neralla S et al.; Mycobacterium tuberculosis (MTB) remains a worldwide health care challenge despite the relatively recent evolution of effective antituberculous medications and combination drug therapy . In many parts of the world, the continued high prevalence of MTB disease is caused in part by the lack of availability of medications and the growing problem of multidrug-resistant TB (MDR-TB) . In the United States, however, errors in treatment constitute a significant portion of treatment failures and relapses . The eradication of MTB in the United States is an achievable goal through the strict adherence to several treatment principles that have been developed over the latter half of the 20th century . These include (1) the use of multiple drugs to which the organism is sensitive; (2) using these drugs in appropriate combinations for a sufficient period of time; (3) using directly observed therapy whenever possible; (4) using in vitro drug susceptibility and local resistance patterns to guide initial drug choices; and (5) never adding a single drug to a failing regimen . Strict adherence to these principles along with an emphasis on completing the total number of doses for a particular regimen will help to ensure fewer relapses/failures, achieve higher cure rates, and help eradicate MTB disease in this country.

Semin Respir Infect, 2003 Dec, 18(4), 241 - 8
The diagnosis of tuberculosis: what's old, what's new; Schluger NW; The approach to the diagnosis of both active tuberculosis and latent infection has changed very little in the past several decades . For active disease, sputum smears with or without chest radiographs to aid in diagnostic accuracy, form the cornerstone of the diagnostic approach in many high-burden countries . These tests usually are supplemented by cultures when resources permit . The diagnosis of latent infection still relies on the use of the tuberculin skin test using purified protein derivative . The current global tuberculosis epidemic, which features large numbers of patients with human immunodeficiency virus infection and increasing rates of multidrug-resistant tuberculosis, makes accurate and rapid diagnosis of tuberculosis more urgent than ever before . Currently available technologies, most involving techniques of DNA amplification, can substantially improve the accuracy of the diagnosis of tuberculosis, although the use of such assays has been sharply limited because of concerns about cost . However, economic analyses suggest that these assays can be cost effective if they lead to sharp reductions in transmission through earlier treatment of infectious cases.

Semin Respir Infect, 2003 Dec, 18(4), 225 - 40
Epidemiology of tuberculosis; Iademarco MF et al.; In the United States, many people erroneously think that tuberculosis (TB) is a disease of the past--an illness that no longer constitutes a public health threat . In reality, TB is one of the leading global causes of morbidity and mortality . According to the World Health Organization, which compiles annual country profiles of reported TB cases using standardized case definitions, 2.4 million cases were reported in 2001 . However, because of underreporting, the number of new TB cases is estimated to be 8.3 million, including 1.8 million deaths . In the United States, after more than 3 decades of steady downward trends, an unprecedented resurgence of TB occurred between 1985 and 1992 . This increase was associated with deficient infrastructure, the human immunodeficiency virus (HIV) epidemic, immigration, outbreaks in congregate settings, and widespread occurrence of multidrug resistant TB strains . The resultant increased concerns provided the impetus for the development of a national action plan to combat MDR and the mobilization of new resources . Consequently, the incidence of TB cases has decreased from 1992 through 2002 . New challenges are evident and must be addressed to achieve the agreed-on goal of eliminating TB in the United States . This report describes the global epidemiology of TB and the epidemiology in the United States, and outlines future challenges to the elimination of TB in the United States.

Hybrid Hybridomics, 2003 Oct, 22(5), 321 - 7
Mouse myeloma cell line Sp2/0 multidrug-resistant variant as parental cell line for hybridoma construction; Melixetian MB et al.; The use of multidrug-resistant variant of Sp2/0 mouse myeloma cell line SpEBr-5 as a partner for making mouse hybridoma producing monoclonal antibodies is described here . The resulting hybridoma cell line 1F7 was characterized with a high level of monoclonal antibody production and karyotype containing all normal mouse chromosomes . 1F7 cells were separately selected for resistance to ethidium bromide (EBr) and adriamycin (ADR) and different mechanisms of drug resistance were found in these cell variants . The resistance in ADR-selected 1F7 cells was due to amplification and overexpression of mdr genes . In EBr-resistant 1F7 cells, mdr genes were overexpressed without amplification . Substantially decreased level of Topo II activity in both cell lines also suggests the existence of additional mechanisms for MDR phenotype of hybridoma cells . Finally, adriamycin-resistant 1F7 hybridoma cell variant was found to produce higher level of specific immunoglobulins due to the increased level of Iggamma(2b) heavy chain mRNA.

Int J Tuberc Lung Dis, 2003 Dec, 7(12), 1191 - 8
Analysis of multidrug-resistant Mycobacterium bovis from three clinical samples from Scotland; Hughes VM et al.; OBJECTIVE: To determine whether multidrug-resistant (MDR) strains of Mycobacterium bovis isolated from patients in Scotland were genotypically related . DESIGN: Genotypes of MDR strains were determined using three molecular fingerprinting techniques: pulsed-field gel electrophoresis (PFGE), spoligotyping and restriction fragment length polymorphism (RFLP) . PFGE profiles were also obtained for all medical and veterinary isolates occurring in Scotland in 1997-1998 . RESULTS: MDR strains showed individual Dra I PFGE profiles . Case III/98 had a profile represented in both veterinary and medical populations, Case I/94 had a profile observed in medical but not veterinary isolates, and Case II/98 had a profile unique to this study . Afl II PFGE discriminated the resistant strains . Spoligotyping grouped Cases I/94 and II/98 (ST-134) . Case III/98 had a spoligotype ST-140, which is commonly observed in veterinary isolates . Similarly, DRr-RFLP analysis grouped cases I/94 and II/98, whereas Case III/98 had a common veterinary profile . DRX(PGRS) RFLP gave three unique profiles . CONCLUSION: Three resistant strains were discriminated by PFGE and DRX(PGRS) RFLP, indicating that the three strains are not related in an epidemiologically relevant time scale . However, Cases I/94 and II/98 were more closely linked by spoligotyping and DRr-RFLP data . PFGE and DRr-RFLP linked Case III/98 profiles to the most common veterinary isolate.

Int J Tuberc Lung Dis, 2003 Dec, 7(12 Suppl 3), S501 - 9
Contact investigations as a means of detection and timely treatment of persons with infectious multidrug-resistant tuberculosis; Bayona J et al.; SETTING: Two regions of metropolitan Lima, Peru . OBJECTIVE: To determine the outcomes of two contact investigation strategies used in therapy enrollment cohorts of patients with multidrug-resistant tuberculosis (MDR-TB) . DESIGN: From 28 August 1996 to 31 December 1999, 91 index patients received individualized MDR-TB therapy (Group A), and from 1 October 1997 to 31 December 1999, another 101 index patients received a standardized MDR-TB regimen (Group B) . We conducted a retrospective chart review and home visits to identify secondary cases among close contacts of both of these groups . Group A secondary cases with MDR-TB received therapy based on the drug susceptibility profile of their infecting strain, while Group B secondary cases received standard short-course therapy . RESULTS: Among 945 close contacts, 72 secondary TB cases (8%) were found . Of 42 who had drug-susceptibility testing, 35 (84%) were MDR-TB, but only seven (17%) had the same drug susceptibility profile as the index case . Cure exceeded 80% in Group A secondary cases, while only half of Group B secondary cases were cured (RR 1.6, 95%CI 1.1-2.2) . CONCLUSION: Contact investigation protocols coupled with enrollment in MDR-TB therapy are a useful means of detecting and promptly treating persons with infectious MDR-TB . In settings with endemic MDR strains of Mycobacterium tuberculosis, effective therapy of contacts of MDR-TB patients requires knowledge of drug susceptibility for each contact with active disease.

Int J Tuberc Lung Dis, 2003 Dec, 7(12 Suppl 3), S494 - 500
Tuberculosis infection and disease in children living in households of Filipino patients with tuberculosis: a preliminary report; Salazar-Vergara RM et al.; SETTING: DOTS Clinic with a DOTS-Plus pilot project for the management of multidrug-resistant tuberculosis (MDR-TB) in a high burden country . OBJECTIVE: To determine the prevalence of tuberculosis (TB) infection and disease among pediatric household contacts of patients with pulmonary TB (PTB) . DESIGN: Cross-sectional study . METHODOLOGY: One hundred and fifty-three children aged 0-15 years in the households of 62 bacteriologically confirmed PTB patients, including 44 with MDR-TB, were studied . BCG scars were noted, and tuberculin skin test (TST), screening chest radiography, and sputum or gastric aspirate smear and culture for Mycobacterium tuberculosis in those with radiographic findings suggestive of PTB were done . RESULTS: For children in this study, the prevalences of latent TB infection (LTBI), radiographically diagnosed pulmonary TB, and bacillary pulmonary TB were 69.2%, 3.3%, and 0.65%, respectively . Only age > or = 5 years was found to be a significant predictor of LTBI (OR 3.17, 95%CI 1.43-7.01) . CONCLUSION: Contact investigation for active case-finding and early treatment of TB in children from households of patients with active PTB is essential for TB control . Further study on a more precise definition of TB infection and strategies for control in this population will be pursued.

Pharmacogenomics J, 2004, 4(1), 40 - 8
Warfarin sensitivity related to CYP2C9, CYP3A5, ABCB1 (MDR1) and other factors; Wadelius M et al.; The required dose of the oral anticoagulant warfarin varies greatly, and overdosing often leads to bleeding . Warfarin is metabolised by cytochrome P450 enzymes CYP2C9, CYP1A2 and CYP3A . The target cell level of warfarin may be dependent on the efflux pump P-glycoprotein, encoded by the adenosine triphosphate-binding cassette gene ABCB1 (multidrug resistance gene 1) . Genetic variability in CYP2C9, CYP3A5 and ABCB1 was analysed in 201 stable warfarin-treated patients using solid-phase minisequencing, pyrosequencing and SNaPshot . CYP2C9 variants, age, weight, concurrent drug treatment and indication for treatment significantly influenced warfarin dosing in these patients, explaining 29% of the variation in dose . CYP3A5 did not affect warfarin dosing . An ABCB1 haplotype containing the exon 26 3435T variant was over-represented among low-dose patients . Thirty-six patients with serious bleeding complications had higher prothrombin time international normalised ratios than 189 warfarin-treated patients without serious bleeding, but there were no significant differences in CYP2C9, CYP3A5 or ABCB1 genotypes and allelic variants.

Pharmacol Toxicol, 2003 Dec, 93(6), 297 - 304
Spontaneous reversal of p-glycoprotein expression in multidrug resistant cell lines; Green H et al.; Increased expression of P-glycoprotein encoded by the mdr-1 gene is a well-characterised mechanism for resistance to cancer chemotherapeutic drugs in cell lines . However, the P-glycoprotein expression after removal of the selection pressure has not fully been elucidated . The stability of P-glycoprotein expression in the presence (+) and absence (-) of vincristine (30 or 150 nM) was studied in multidrug resistant K562 cell lines (VCR30+, VCR150+, VCR30- and VCR150-) for 11 months . The P-glycoprotein protein and mdr-1 mRNA levels were determined at regular intervals using flow cytometry and real-time PCR, respectively . Chemosensitivity to a panel of antineoplastic drugs was measured using an MTT assay . The presence of vincristine (VCR30+ and VCR150+) resulted in high and stable levels of P-glycoprotein and mdr-1 mRNA during the whole period compared to wild type . As for the VCR30- and VCR150- subcultures, the expressions of P-glycoprotein and mdr-1 mRNA were stable for five months, and then the levels decreased rapidly . Concomitantly, the sensitivity to drugs known as P-glycoprotein substrates was restored . In conclusion, resistant cells growing in the presence of the inducing drug have a stable P-glycoprotein expression and resistance level, but removing the inducing drug may result in a sudden and rapid lowering of P-glycoprotein and mdr-1 mRNA levels as long as five months after drug withdrawal.

Int J Gynecol Cancer, 2003 Nov-Dec, 13(6), 776 - 84
Multivariate analysis for prognostic significance of histologic subtype, GST-pi, MDR-1, and p53 in stages II-IV ovarian cancer; Ikeda K et al.; It has been suggested that histologic subtype of ovarian cancer is a factor that determines the chemoresponsiveness of tumor . In this study, we wanted to clarify the prognostic significance of histologic subtype and its correlation to expression of chemoresistance-related proteins (CRPs) in ovarian cancer . A total of 93 stage II-IV ovarian cancers, where the proportion of serous, endometrioid, mucinous, and clear cell subtype was 61.3%, 14.0%, 7.5%, and 17.2%, respectively, were investigated for glutathione S-transferase-pi (GST-pi), MDR (multidrug resistance)-1, and p53 expression using immunohistochemistry . GST-pi expression was detected in 62.4% of the tumors and was not related to histologic subtype of tumor . MDR-1 expression was observed in 12.9% of the tumors tested and was more frequently detected in clear cell adenocarcinomas than other histologic subtypes of tumor (10/ 16 vs . 2 / 77, P < 0.001) . P53 expression was found in 49.1% of serous, 53.8% of endometrioid, and 50% of mucinous adenocarcinomas . In contrast, none of 16 clear cell adenocarcinomas showed positive p53 staining . In univariate analysis, no direct correlations were found between CRPs and overall survival . Histology of mucinous/clear cell tumors (P = 0.0063), as well as FIGO stage III/IV (P = 0.0091) and residual tumor >or= 2 cm (P = 0.0045), was found to have independent prognostic value in multivariate analysis . In conclusion, histologic subtype proved to be the significant independent prognostic factor in addition to FIGO stage and residual tumor in stage II-IV ovarian cancer . GST-pi, MDR-1, and p53 expression pattern is closely related to histologic subtype of ovarian cancer, although they are not significant predictors of survival.

Clin Pharmacokinet, 2003, 42(15), 1331 - 57
Role of orphan nuclear receptors in the regulation of drug-metabolising enzymes; Wang H et al.; During the past several years, important advances have been made in our understanding of the mechanisms that regulate the expression of genes that determine drug clearance, including phase I and phase II drug-metabolising enzymes and drug transporters . Orphan nuclear receptors have been recognised as key mediators of drug-induced changes in both metabolism and efflux mechanisms . In this review, we summarise recent findings regarding the function of nuclear receptors in regulating drug-metabolising and transport systems, and the relevance of these receptors to clinical drug-drug interactions and the development of new drugs . Emphasis is given to two newly recognised 'orphan' receptors (the pregnane X receptor {PXR} and the constitutive androstane receptor {CAR}) and their regulation of cytochrome P450 enzymes, such as CYP3A4, CYP2Cs and CYP2B6; and transporters, such as P-glycoprotein (MDR1), multidrug resistance-associated proteins (MRPs) and organic anion transporter peptide 2 (OATP2) . Although 'cross-talk' occurs between these two receptors and their target sequences, significant species differences exist between ligand-binding and activation profiles for both receptors, and PXR appears to be the predominant or 'master' regulator of hepatic drug disposition in humans . Several important physiological processes, such as cholesterol synthesis and bile acid metabolism, are also tightly controlled by certain ligand-activated orphan nuclear receptors (farnesoid X receptor {FXR} and liver X receptor {LXR}) . In general, their ability to bind a broad range of ligands and regulate an extensive array of genes that are involved in drug clearance and disposition makes these orphan receptors attractive targets for drug development . Drugs have the capacity to alter nuclear receptor expression (modulators) and/or serve as ligands for the receptors (agonists or antagonists), and thus can have synergistic or antagonistic effects on the expression of drug-metabolising enzymes and transporters . Coadministration of drugs that are nuclear receptor agonists or antagonists can lead to severe toxicity, a loss of therapeutic efficacy or an imbalance in physiological substrates, providing a novel molecular mechanism for drug-drug interactions.

J Infect Dis, 2003 Dec 15, 188(12), 1878 - 84 Epub 2003 Dec 09.
Effect of drug resistance on the generation of secondary cases of tuberculosis; Burgos M et al.; BACKGROUND: The results of animal studies suggest that isoniazid-resistant strains of Mycobacterium tuberculosis are less pathogenic than isoniazid-susceptible strains . Here, we assess the relative pathogenicity of drug-resistant and drug-susceptible strains, in a human population . METHODS: We linked IS6110 genotype patterns of M . tuberculosis strains with drug-susceptibility test results and epidemiologic information for 85% of culture-positive incident cases of tuberculosis (TB) in San Francisco during 1991-1999 . We assumed that drug-susceptible and drug-resistant strains were transmitted to secondary case patients if the drug-resistance and genotype patterns were identical . We calculated the number of secondary cases for each drug-resistance pattern and determined the relative secondary-case rate ratio (SR) of drug-resistant TB to drug-susceptible TB . RESULTS: There were 1800 patients with culture-positive TB, drug-susceptibility test results, and genotyping results . The overall SR of drug-resistant to drug-susceptible TB cases was 0.51 (95% confidence interval {CI}, 0.37-0.69) . The SR was 0.29 (95% CI, 0.15-0.57) for isoniazid-resistant strains, 0.10 (95% CI, 0.02-0.42) for strains resistant to both isoniazid and streptomycin, and 0.88 (95% CI, 0.53-1.47) for streptomycin-resistant strains . There were no secondary cases caused by multidrug-resistant (MDR) TB . The SR for rifampin-resistant cases was 2.33 (95% CI, 1.04-5.25) . Seventy-eight percent (7/9) of the patients with rifampin-resistant secondary cases of TB were seropositive for human immunodeficiency virus . CONCLUSION: In the context of an effective TB program in San Francisco, strains that were resistant to isoniazid either alone or in combination with other drugs were less likely to result in secondary cases than were drug-susceptible strains . In this setting, isoniazid-resistant and MDR TB cases were not likely to produce new, incident drug-resistant TB cases.

Chemotherapy, 2003 Dec, 49(6), 303 - 8
The farnesyl protein transferase inhibitor lonafarnib (SCH66336) is an inhibitor of multidrug resistance proteins 1 and 2; Wang EJ et al.; Clinical studies indicate that the farnesyl protein transferase inhibitor SCH66336 (lonafarnib), an anticancer agent developed to antagonize oncogenic Ras, is generally well tolerated . Lonafarnib has also demonstrated therapeutic synergy with coadministered taxanes, vincristine, cisplatin, cyclophosphamide, 5-fluorouracil (5-FU) and Gleevec . Lonafarnib has recently been shown, in addition, to be a potent inhibitor of the transmembrane efflux transporter P-glycoprotein (P-gp), which confers cellular resistance to the substrates vincristine, taxol and paclitaxel . Treatment with lonafarnib would therefore be predicted to be synergistic with these coadministered cancer therapeutics that are substrates of P-gp . However, cisplatin, 5-FU and cyclophosphamide are not P-gp substrates, yet cisplatin, 5-FU and possibly cyclophosphamide are purported substrates for multidrug resistance proteins (MRPs) 1 and 2 (known to cause chemotherapy resistance) . Lonafarnib is shown here to inhibit the function of MRP1 and MRP2 with a potency similar to that of cyclosporin A and may therefore cause the observed synergy with cisplatin and other agents by inhibiting these MRPs . Coadministration of lonafarnib could thus reduce chemotherapy dosage and hence produce lower exposure to normal cells and less undesired toxicity .

Arch Dis Child, 2003 Dec, 88(12), 1106 - 11
Culture confirmed multidrug resistant tuberculosis: diagnostic delay, clinical features, and outcome; Schaaf HS et al.; AIMS: To determine the delay in diagnosis of multidrug resistant (MDR) tuberculosis (TB), the correlation between drug susceptibility patterns of adult-child contact pairs, the effectiveness of treatment, and the outcome in these children . METHODS: MDR M tuberculosis culture results of children were prospectively collected during a four year period in the Western Cape Province of South Africa, an area with a TB incidence of 589/100 000 population, and a new MDR TB rate of 0.94% . Folder reviews were done to retrieve clinical information . Children not already on treatment at our MDR TB clinic or TB hospital were recalled and appropriate treatment was started . Follow up was done for as long as possible . RESULTS: Thirty nine children, median age 4.5 years at first TB diagnosis and 6.2 years on MDR culture confirmation, were seen . Delay in starting appropriate MDR treatment after TB diagnosis was a median of 2 days if MDR TB source cases were taken into account, but 246 days if the drug susceptibility pattern of the source case was not considered, and 283 days if there was no known tuberculosis source case . Correlation between the drug susceptibility results of the child's and adult source case's isolates was 68% . Seventeen children had smear positive tuberculosis, of whom 13 had cavitatory pulmonary disease . Eight children had central nervous system TB . Thirty six children were treated for MDR tuberculosis, of whom four died . CONCLUSIONS: Obtaining a detailed contact history is essential as a delay in starting appropriate MDR antituberculosis treatment has potentially serious consequences.

Biochem Pharmacol, 2004 Jan 1, 67(1), 31 - 9
Glucuronidation as a mechanism of intrinsic drug resistance in colon cancer cells: contribution of drug transport proteins; Cummings J et al.; We have recently shown that drug conjugation catalysed by UDP-glucuronosyltransferases (UGTs) functions as an intrinsic mechanism of resistance to the topoisomerase I inhibitors 7-ethyl-10-hydroxycamptothecin and NU/ICRF 505 in human colon cancer cells and now report on the role of drug transport in this mechanism . The ability of transport proteins to recognise NU/ICRF 505 as a substrate was evaluated in model systems either transfected with breast cancer-resistance protein 1 (Bcrp1), multidrug-resistance protein 2 (Mrp2) or Mrp3, or overexpressing MRP1 or P-170 glycoprotein . Results from chemosensitivity assays suggested that NU/ICRF 505 was not a substrate for any of the above proteins . In drug accumulation studies in human colon cancer cell lines NU/ICRF 505 was taken up avidly and retained in cells lacking UGTs (HCT116), whereas, following equally rapid uptake, it was cleared rapidly from cells displaying UGT activity (HT29) as glucuronide metabolites . HT29 cells were shown to express MRP1 and 3, but not P-170 glycoprotein, MRP2 or breast cancer-resistance protein . The major glucuronide of NU/ICRF 505 inhibited ATP-dependent transport of estradiol 17-beta-glucuronide in Sf9 insect cell membrane vesicles containing MRP1 or MRP3, while co-incubation of HT29 cells with the MRP antagonist, MK571, significantly restored intracellular concentrations of NU/ICRF 505 . These data lead us to conclude that the presence of a glucuronide transporter is essential for glucuronidation to represent a major de novo resistance mechanism and that UGTs will contribute more as a primary resistance mechanism when the parent drug (e.g . NU/ICRF 505) is not itself recognised by transport proteins.

Matrix Biol, 2003 Nov, 22(6), 491 - 500
Multidrug resistance protein-6 (MRP6) in human dermal fibroblasts . Comparison between cells from normal subjects and from Pseudoxanthoma elasticum patients; Boraldi F et al.; Multidrug resistance protein-6 (MRP6) is a membrane transporter whose deficiency leads to the connective tissue disorder Pseudoxanthoma elasticum (PXE) . In vitro dermal fibroblasts from normal and PXE subjects, homozygous for the R1141X mutation, were compared for their ability to accumulate and to release fluorescent calcein, in the absence and in the presence of inhibitors and competitors of the MDR-multidrug resistance protein (MRP) systems, such as 3-(3-(2-(7-choro-2 quinolinyl) ethenyl)phenyl ((3-dimethyl amino-3-oxo-propyl)thio) methyl) propanoic acid (MK571), verapamil (VPL), vinblastine (VBL), chlorambucil (CHB), benzbromarone (BNZ) and indomethacin (IDM) . In the absence of chemicals, calcein accumulation was significantly higher and the release significantly slower in PXE cells compared to controls . VBL and CHB reduced calcein release in both cell strains, without affecting the differences between PXE and control fibroblasts . VPL, BNZ and IDM consistently delayed calcein release from both control and PXE cells; moreover, they abolished the differences between normal and MRP6-deficient fibroblasts observed in the absence of chemicals . These findings suggest that VPL, BNZ and IDM interfere with MRP6-dependent calcein extrusion in in vitro human normal fibroblasts . Interestingly, MK571 almost completely abolished calcein release from PXE cells, whereas it induced a strong but less complete inhibition in control fibroblasts, suggesting that MRP6 is not inhibited by MK571 . Data show that MRP6 is active in human fibroblasts, and that its sensitivity to inhibitors and competitors of MDR-MRPs' membrane transporters is different from that of other translocators, namely, MRP1 . It could be suggested that MRP1 and MRP6 transport different physiological substances and that MRP6 deficiency cannot be overcome by other membrane transporters, at least in fibroblasts . These data further support the hypothesis that MRP6 deficiency may be relevant for fibroblast metabolism and responsible for the metabolic alterations of these cells at the basis of connective tissue clinical manifestations of PXE.

Anticancer Res, 2003 Sep-Oct, 23(5A), 3607 - 15
Pervilleine F, a new tropane alkaloid aromatic ester that reverses multidrug resistance; Mi Q et al.; P-glycoprotein (Pgp)-mediated drug efflux can yield a multidrug resistance (MDR) phenotype that is associated with poor response to cancer chemotherapy . Pervilleine F, a new tropane alkaloid aromatic ester obtained from a chloroform extract of the roots of Erythroxylum pervillei as the result of bioactivity-guided fractionation, was found to restore the vinblastine sensitivity of cultured multidrug-resistant KB-V1 cells, with an IC50 value of 0.40 microM . Pervilleine F (8 microM) was also able to partially reverse the cross-resistance of KB-V1 cells to the clinically used or experimental anticancer agents actinomycin D (45.1-fold), baccatin III (> 3.4-fold), daunomycin (> 22.5-fold), ellipticine (1.9-fold), mithramycin A (42.5-fold), podophyllotoxin (1.6-fold), paclitaxel (32.2-fold) and vincristine (73.6-fold) . While pervilleine F alone at the concentration of 10 microM had no significant effect on the KB-V1 cell cycle, pervilleine F (at concentrations of 0.2, 1, 2, and 8 microM) combined with vinblastine (1 microgram/ml) induced dose-dependent G2/M phase arrest, ranging from 20.2, 51.0, 63.7, to 79.5%, as an indication of the restoration of vinblastine sensitivity . To confirm this activity with an in vivo animal model, KB-V1 cells were placed in hollow fibers and implanted into NCr nu/nu mice . Cell growth was not significantly inhibited when vinblastine or pervilleine F was administered as single agents, but when these two compounds were used in combination, inhibition of up to 64.1% was observed . Equimolar doses of verapamil were less effective . These data suggest that pervilleine F is an effective inhibitor of Pgp and should be further evaluated for clinical utility.

Cancer Chemother Pharmacol, 2004 Apr, 53(4), 349 - 56 Epub 2003 Dec 10.
Characterization of tetrandrine, a potent inhibitor of P-glycoprotein-mediated multidrug resistance; Fu L et al.; Multidrug resistance (MDR) is one of the main obstacles in tumor chemotherapy . A promising approach to solving this problem is to utilize a nontoxic and potent modulator able to reverse MDR, which in combination with anticancer drugs increases the anticancer effect . Experiments were carried out to examine the potential of tetrandrine (Tet) as a MDR-reversing agent . Survival of cells incubated with Tet at 2.5 micromol/l for 72 h was over 90% . Tet at 2.5 micromol/l almost completely reversed resistance to vincristine (VCR) in KBv200 cells . Tet at a concentration as low as 0.625 micromol/l produced a 7.6-fold reversal of MDR, but showed no effect on the sensitivity of drug-sensitive KB cells in vitro . In the KBv200 cell xenograft model in nude mice, neither Tet nor VCR inhibited tumor growth . However, VCR and Tet combined inhibited tumor growth by 45.7%, 61.2% and 55.7% in three independent experimental settings . In the KB cell xenograft model in nude mice, Tet did not inhibit tumor growth, but VCR and the combination of VCR and Tet inhibited tumor growth by 40.6% and 41.6%, respectively . Mechanism studies showed that Tet inhibited {(3)H}azidopine photoaffinity labeling of P-gp and increased accumulation of VCR in MDR KBv200 cells in a concentration-dependent manner . The results suggest that Tet is a potent MDR-reversing agent in vitro and in vivo . Its mechanism of action is via directly binding to P-gp and increasing intracellular VCR accumulation.

Braz J Med Biol Res, 2003 Dec, 36(12), 1653 - 7 Epub 2003 Nov 17.
Age-related changes of the multidrug resistance P-glycoprotein function in normal human peripheral blood T lymphocytes; Machado CG et al.; The multidrug resistance P-glycoprotein is a transmembrane efflux pump expressed by lymphocytes and is involved in their cytolytic activity . In the present study, we investigated the age-related changes of P-glycoprotein function in normal peripheral blood lymphocytes . Blood samples from 90 normal volunteers (age range, 0 to 86 years) were analyzed . P-glycoprotein function was assessed by the flow cytometric rhodamine 123 assay . P-glycoprotein function was highest in cord blood and progressively declined with age in peripheral blood T CD4+ and CD8+ cells . In contrast, P-glycoprotein function did not vary with age in CD19+ B or CD16+CD56+ natural killer cells . These data suggest that the decline in P-glycoprotein function in T CD4+ and CD8+ lymphocytes as a function of age may contribute to the decrease in T cell cytolytic activity with aging.

J Clin Microbiol, 2003 Dec, 41(12), 5774 - 7
Evaluation of a PCR-based universal heteroduplex generator assay as a tool for rapid detection of multidrug-resistant Mycobacterium tuberculosis in Peru; Mayta H et al.; Multidrug-resistant tuberculosis is an increasing health problem worldwide, especially in developing countries . The PCR-UHG-Rif assay, which detects mutations within the rpoB gene associated with rifampin resistance, was evaluated for its ability and reliability to detect and identify drug-resistant Mycobacterium tuberculosis in a developing country where tuberculosis is highly endemic.

Hepatol Res, 2003 Dec, 27(4), 323 - 326
Novel mutations identified in the human multidrug resistance-associated protein 2 (MRP2/ABCC2) gene in a Japanese patient with Dubin-Johnson syndrome; Shoda J et al.; A 39-year-old Japanese women who has been jaundiced since childhood and indicative of Dubin-Johnson syndrome (DJS) underwent MRP2/ABCC2 mutation analysis . The results of the analysis revealed two novel mutations, C298T and C3928T, and two single nucleotide polymorphisms (SNPs), C3972T and C-24T . One of the two mutations (C298T) is in the transmembrane domain . The other mutation (C3928T) and the SNP (C3972T) are in the cytoplasmaic domain which are concentrated in the second adenosine triphosphate (ATP)-binding cassette . Both of the mutations, C298T and C3928T, are immature stop codons . C298T, C-24T and C3972T (originating from the mother) and C3928T (from the father) were observed to be heterozygous in this patient . Light-microscopy examination of the liver biopsy specimens revealed the accumulation of dark pigment granules in most of the hepatocytes . In the immunohistochemistry using pAb recognizing the C-terminal of the human MRP2, no staining of MRP2 in the canalicular membrane domain was observed in the liver sections from this patient . In this patient, the mutations with immature stop codons presumably resulted in instability of MRP2 mRNA and/or defective synthesis of the truncated protein, which may underlie the loss of the expression of functional MRP2 protein.

Gen Physiol Biophys, 2003 Jun, 22(2), 265 - 73
Hypoxia increases cell death in multidrug-resistant leukemia cells . Differences in viability and ultrastructure between sensitive and multidrug-resistant L1210 mouse leukemic cells under hypoxia; El-Saggan AH et al.; The comparative study of sensitive and multidrug-resistant L1210 cells under 24 hours of hypoxia (2% O2 and 5% CO2 at 37 degrees C) was done to see if differences in energetic metabolism between both cell lines are paralleled by differences in cellular morphology . During the dye exclusion assay the viability of sensitive cells was about 70 to 90%, whereas only 30 to 50% of resistant cells were viable . Electron microscopic study of sensitive and resistant L1210 cells under hypoxia has shown cells of different ultrastructural appearance in both cell lines . Cells with necrotic changes (swollen mitochondria, lysed cells) prevailed in resistant cells . The highest incidence of cells with normal or slightly dense mitochondria was found among the sensitive L1210 cells . Additionally, cells with pyknotic nuclei, shrunken cytoplasm and dense mitochondria, reminiscent of apoptosis, could be found sporadically, especially in the sensitive L1210 cell line . These results are in agreement with flow cytometry measurements: in resistant cells the number of necrotic cells was on the average 2.3 times higher than in sensitive cells . Ultrastructural differences and differences in the numbers of necrotic cells as measured by flow cytometry between sensitive and resistant L1210 cells under hypoxia are consistent with differences in energetic metabolism between these cell lines, as described in earlier studies, and document an increased cell death in the resistant L1210 cell line.

Oncogene, 2004 Jan 29, 23(4), 945 - 55
Glypican-3 is involved in cellular protection against mitoxantrone in gastric carcinoma cells; Wichert A et al.; Elevated expression of the heparan sulphate proteoglycan glypican-3 (GPC3) was found on mRNA and protein levels in the atypical multidrug-resistant gastric carcinoma cell line EPG85-257RNOV, which was established by in vitro selection against mitoxantrone . In order to elucidate a putative role of GPC3 in the drug-resistant phenotype, the mitoxantrone-resistant cell line EPG85-257RNOV was transfected with an expression vector construct carrying an anti-GPC3 hammerhead ribozyme . It could be demonstrated that in anti-GPC3 ribozyme-transfected cell clones, the GPC3-specific mRNA and corresponding protein expression levels were decreased to levels that are similar to those observed in nonresistant, parental cells . The anti-GPC3 ribozyme-containing clones reduced the mitoxantrone resistance level up to 21% of the original resistance and the crossresistance against etoposide to 33% of the original value . This reversal of drug resistance was accompanied by an increased cellular mitoxantrone accumulation in the anti-GPC3 ribozyme-expressing cells . In conclusion, it was verified that GPC3 is involved in the cellular protection against mitoxantrone in the atypical multidrug-resistant gastric carcinoma cell line EPG85-257RNOV.

Br J Cancer, 2003 Dec, 89 Suppl 3, S23 - 8
Resistance to chemotherapy in advanced ovarian cancer: mechanisms and current strategies; Vasey PA; Although treatment for advanced epithelial ovarian cancer has improved over recent years with the introduction of taxane-platinum chemotherapy, the majority of patients will relapse, and in most the disease remains incurable . A thorough understanding of drug resistance mechanisms is needed, as this remains the largest obstacle in treating patients with recurrent disease . Multidrug resistance proteins, mismatch repair processes and alterations in the p53 pathway are examples of properties within tumour cells that may lead to drug resistance . Novel agents designed to circumvent these mechanisms (e.g . PSC 833, ONYX-015 and ADP53) are currently being investigated for ovarian cancer patients . Further improvements may result from the optimisation of existing first-line regimens with more creative schedules, perhaps involving sequential or intraperitoneal administration of existing drugs, and the incorporation of newer noncross-resistant drugs.

Nat Biotechnol, 2004 Jan, 22(1), 62 - 9 Epub 2003 Dec 07.
Integration of chemical-genetic and genetic interaction data links bioactive compounds to cellular target pathways; Parsons AB et al.; Bioactive compounds can be valuable research tools and drug leads, but it is often difficult to identify their mechanism of action or cellular target . Here we investigate the potential for integration of chemical-genetic and genetic interaction data to reveal information about the pathways and targets of inhibitory compounds . Taking advantage of the existing complete set of yeast haploid deletion mutants, we generated drug-hypersensitivity (chemical-genetic) profiles for 12 compounds . In addition to a set of compound-specific interactions, the chemical-genetic profiles identified a large group of genes required for multidrug resistance . In particular, yeast mutants lacking a functional vacuolar H(+)-ATPase show multidrug sensitivity, a phenomenon that may be conserved in mammalian cells . By filtering chemical-genetic profiles for the multidrug-resistant genes and then clustering the compound-specific profiles with a compendium of large-scale genetic interaction profiles, we were able to identify target pathways or proteins . This method thus provides a powerful means for inferring mechanism of action.

J Biol Chem, 2004 Feb 20, 279(8), 6507 - 16 Epub 2003 Dec 01.
Increased calcium influx and ribosomal content correlate with resistance to endoplasmic reticulum stress-induced cell death in mutant leukemia cell lines; Zhang Y et al.; Cell clones were derived by treatment of HL-60 cells with stepwise increasing concentrations of econazole (Ec), an imidazole antifungal that blocks Ca2+ influx and induces endoplasmic reticulum (ER) stress-related cell death in multiple mammalian cell types . Clones exhibit 20- to more than 300-fold greater resistance to Ec . Unexpectedly, they also display stable cross-resistance to tunicamycin, thapsigargin, dithiothreitol, and cycloheximide but not doxorubicin, etoposide, or Fas ligand . Phenotypic analysis indicates that the cells display increased store-operated calcium influx and resistance to ER Ca2+ store depletion by Ec . E2R2, the most resistant clone, was observed to maintain protein synthesis levels after treatment with Ec or thapsigargin . Expression of GRP78, an ER-based chaperone, was induced by these ER stress treatments but to equal degrees in HL-60 and E2R2 cells . By using microarray analysis, at least 15 ribosomal protein genes were found to be overexpressed in E2R2 compared with HL-60 cells . We also found that ribosomal protein content was increased by 30% in E2R2 as well as other clones . The resistance phenotype was partially reversed by the ribosome-inactivating protein saporin . Therefore, increased store-operated calcium influx, resistance to ER Ca2+ store depletion, and overexpression of ribosomal proteins define a novel phenotype of ER stress-associated multidrug resistance.

Xenobiotica, 2003 Nov, 33(11), 1125 - 37
Stereoselective hepatic disposition of a diastereomeric pair of alphavbeta3 antagonists in rat; Prueksaritanont T et al.; 1 . The study investigated mechanisms underlying the stereoselective hepatic disposition observed in rats of a zwitterionic diastereomeric pair ((3S)-3-{(3R or 3S)-2-oxo-3-{3-(5,6,7,8-tetrahydro-1,8-naphthyridin-2-yl)propyl}pyrrolidin-1-yl}-3-quinolin-3-ylpropanoic acid) with different lipophilicities . 2 . In a recirculating isolated rat liver system, the more hydrophilic diastereomer II possessed biliary clearance, CLb, and bile-to-liver concentration ratio higher (about 10-30-fold) than the lipophilic zwitterion I, whereas both I and II exhibited comparably high concentration ratios between liver and perfusate . Although MK-571, a known multidrug resistance protein (MRP) inhibitor, significantly inhibited the CLb of both compounds, it did not inhibit their canalicular transport, as evident by unchanged concentration ratios between bile and liver of either I or II . 3 . Following an intravenous infusion of I or II to Sprague-Dawley rats, the biliary clearance calculated either based on plasma (CL(b,p)) or liver concentration (CL(b,l)), of II was much higher than that of I (about 5-50-fold) . In rats lacking multidrug resistance protein 2 (Mrp2) (Eisai hyperbilirubinemic rat, EHBR), the biliary excretion rate and CL(b,p) of II were also higher than the corresponding values for I . However, both CL(b,p) or CL(b,l) of either I or II were not reduced in EHBR, as compared with control SD rats . 4 . In the in vitro rat canalicular membrane vesicle study, I and II exhibited no differences in their inhibitory effect on the Mrp2 mediated ATP-dependent {3H}DNP-SG initial uptake (no inhibition at 10 microM and only about 40% inhibition at 100 microM) . 5 . Collectively, these results suggested that (1) the difference in the hepatic disposition between the two isomers was due primarily to the difference in their transport mechanism across the canalicular membrane and (2) Mrp2 did not play a major role in the observed differences in the biliary excretion of the diastereomers I and II in rats.

Breast, 2003 Feb, 12(1), 58 - 62
Predicting multidrug resistance-related protein and P-glycoprotein expression with technetium-99m tetrofosmin mammoscintigraphy; Liu TJ et al.; The purpose of this study was a retrospective survey of 40 patients with infiltrating ductal breast carcinoma to evaluate the relationships between the degree of accumulation of technetium-99m tetrofosmin (Tc-TF), multidrug resistance-related protein (MRP) expression and P-glycoprotein (Pgp) expression in breast cancer tissue . Immunohistochemical analysis (IHA) was performed on pathological specimens of the 40 breast cancers to determine Pgp and MRP expression . The results of IHA, were used as the basis for dividing the 40 breast cancers into four groups: A, 10 tumors with positive MRP and Pgp expressions; B, 10 tumors with positive MRP but negative Pgp expression; C, 10 tumors with negative MRP but positive Pgp expression; and D, 10 tumors with negative MRP and Pgp expression . All 40 patients had undergone Tc-TF mammoscintigraphy to calculate breast cancer uptake of Tc-TF to background uptake (T/B) ratios before IHA and surgery/biopsy . Of the four groups, group A had the lowest T/B ratios (1.15+/-0.10) and group D, the highest (2.19+/-0.15) (P<0.05) . The T/B ratios in groups B (1.36+/-0.27) and C (1.37+/-0.26) were intermediate between those of groups A and D . In addition, the T/B ratios were statistically significantly lower in group A than in group B or C, and statistically significantly higher in groups D than in groups B or C (P<0.05) . However, no significant difference in T/B ratio was found between groups B and C (P>0.05) . Our results indicate that Tc-TF mammoscintigraphy is helpful for in vivo determination of Pgp and MRP expression in breast cancers.

Am J Respir Crit Care Med, 2004 Mar 1, 169(5), 604 - 9 Epub 2003 Dec 04.
Cough-generated aerosols of Mycobacterium tuberculosis: a new method to study infectiousness; Fennelly KP et al.; The concentration and size distribution of infectious aerosols produced by patients with pulmonary tuberculosis (TB) has never been directly measured . We aimed to assess the feasibility of a method that we developed to collect and quantify culturable cough-generated aerosols of Mycobacterium tuberculosis . Subjects were recruited from a referral hospital and most had multidrug-resistant TB . They coughed into a chamber containing microbial air samplers while cough frequency was measured during two 5-minute sessions . Cough-generated aerosol cultures were positive in 4 of 16 subjects (25%) with smear-positive pulmonary TB . There was a rapid decrease in the cough-generated aerosol cultures within the first 3 weeks of effective treatment . Culture-positive cough aerosols were associated with lack of treatment during the previous week (p = 0.007), and there was a trend in the association with cough frequency (p = 0.08) . The size distributions of these aerosols were variable, but most particle sizes were in the respirable range . Quantification of viable cough-generated aerosols is feasible and offers a new approach to study infectiousness and transmission of M . tuberculosis and other airborne pathogens.

Brain Pathol, 2003 Oct, 13(4), 482 - 94
P-glycoprotein and multidrug resistance-associated protein mediate specific patterns of multidrug resistance in malignant glioma cell lines, but not in primary glioma cells; Bahr O et al.; Understanding and overcoming multidrug resistance (MDR) may be a promising strategy to develop more effective pharmacotherapies for malignant gliomas . In the present study, human malignant glioma cell lines (n=12) exhibited heterogeneous mRNA and protein expression and functional activity of the mdr gene-encoded P-glycoprotein (PGP) and MDR-associated protein (MRP) . Correlation between mRNA expression, protein levels and functional activity was strong . Inhibition of PGP activity by verapamil or PSC 833 enhanced the cytotoxic effects of vincristine, doxorubicin, teniposide and taxol . Inhibition of MRP activity by indomethacin or probenecid enhanced the cytotoxic effects of vincristine, doxorubicin and teniposide . The human cerebral endothelial cell line, SV-HCEC, exhibited the strongest PGP activity of all cell lines . Five primary human glioblastomas and one anaplastic astrocytoma displayed heterogenous protein levels of PGP and MRP-1 in tumor cells and of PGP in biopsy specimens in vivo, but no functional activity of these proteins upon ex vivo culturing . These data suggest that the glioma cell line-associated MDR-type drug resistance is a result of long-term culturing and that cerebral endothelial, but not glioma cells, may contribute to MDR-type drug resistance of gliomas in vivo.

Folia Morphol (Warsz), 2003 Nov, 62(4), 493 - 5
The expression of metallothionein (MT) and proliferation intensity in ovarian cancers treated with cisplatin and paclitaxel; Surowiak P et al.; Metallothioneins (MT) represent low molecular weight proteins that are supposed to fulfil several functions . They participate in the cell cycle, protect cells from oxidative stress, control levels of heavy metals and participate in multidrug resistance processes, particularly in cases of alkylating drugs . The present study aimed at evaluation of proliferation intensity (Ki67, PCNA) in ovarian cancers treated using cisplatin and paclitaxel, as related to expression of MT . The experiments were performed on samples originating from 10 patients operated on due to ovarian cancer . The material originated from the first operations or second-look operations . All the patients were treated with cisplatin and paclitaxel . Immunocytochemical reactions using antibodies to MT, Ki67 and PCNA were performed in paraffin sections originating from the cases studied . Statistical analysis was performed using Statistica software . The studies demonstrated no relation between expression of MT on the one hand and intensity of proliferation before or after chemotherapy on the other hand (gamma correlation, p > 0.05) . The results indicate that expression of MT is not related to resistance to treatment using cisplatin and paclitaxel.

Cancer Chemother Pharmacol, 2004 Mar, 53(3), 220 - 4 Epub 2003 Dec 04.
Insulin-induced enhancement of antitumoral response to methotrexate in breast cancer patients; Lasalvia-Prisco E et al.; PURPOSE: It has been reported that insulin increases the cytotoxic effect in vitro of methotrexate by as much as 10,000-fold . The purpose of this study was to explore the clinical value of insulin as a potentiator of methotrexate . PATIENTS AND METHODS: Included in this prospective, randomized clinical trial were 30 women with metastatic breast cancer resistant to fluorouracil + Adriamycin + cyclophosphamide and also resistant to hormone therapy with measurable lesions . Three groups each of ten patients received two 21-day courses of the following treatments: insulin + methotrexate, methotrexate, and insulin, respectively . In each patient, the size of the target tumor was measured before and after treatment according to the Response Evaluation Criteria In Solid Tumors . The changes in the size of the target tumor in the three groups were compared statistically . RESULTS: Under the trial conditions, the methotrexate-treated group and the insulin-treated group responded most frequently with progressive disease . The group treated with insulin + methotrexate responded most frequently with stable disease . The median increase in tumor size was significantly lower with insulin + methotrexate than with each drug used separately . DISCUSSION: Our results confirmed in vivo the results of previous in vitro studies showing clinical evidence that insulin potentiates methotrexate under conditions where insulin alone does not promote an increase in tumor growth . Therefore, the chemotherapy antitumoral activity must have been enhanced by the biochemical events elicited in tumor cells by insulin . CONCLUSIONS: In multidrug-resistant metastatic breast cancer, methotrexate + insulin produced a significant antitumoral response that was not seen with either methotrexate or insulin used separately.

Oncol Rep, 2004 Jan, 11(1), 179 - 83
Expression of multidrug resistance-1 and multidrug resistance-associated protein genes in pediatric rhabdomyosarcoma; Gallego S et al.; Overexpression of multidrug resistance-1 (MDR-1), and multidrug resistance-associated protein (MRP) genes has been linked with resistance to chemotherapy in vitro and in vivo . Their role in chemotherapy resistance in pediatric rhabdomyosarcoma is unclear . The study was undertaken to analyze the expression of MDR-1 and MRP genes in the embryonal and the alveolar subtypes of rhabdomyosarcoma and to elucidate its clinical relevance . Twenty-three rhabdomyosarcoma samples were analyzed for the expression of MDR-1 and MRP genes using a semi-quantitative competitive RT-PCR assay . MRP gene expression was associated with a reduction in survival (p=0.02) . The overall survival of patients with tumors positive or negative for MRP expression were 50% (95% confidence interval, 30-70%) and 93% (95% confidence interval, 76-100%) respectively . In contrast, the expression of MDR-1 gene was not predictive of survival . These findings suggest that MRP expression could be a prognostic factor in patients with rhabdomyosarcoma.

Oncol Rep, 2004 Jan, 11(1), 133 - 6
Expression of the IAPs in multidrug resistant tumor cells; Notarbartolo M et al.; We have investigated the expression of the IAPs (inhibitory of apoptosis proteins) in the human HL-60 leukemia and in its multidrug resistant, P-glycoprotein (P-gp) over-expressing variant, HL-60R . HL-60R exhibits resistance to apoptosis induced from P-gp substrate drugs and also from other triggers (cisplatin, TNF-alpha, Fas ligation, TRAIL, IFN-gamma and serum starvation) not related to the multidrug transporter . Except for c-IAP-1 mRNA, HL-60R significantly over-expressed both the mRNAs and the proteins of all the IAPs studied, i.e . c-IAP-1, c-IAP-2, XIAP, NAIP and survivin . Determination of the DNA-binding capacity of NF-kappaB (p50 or p65 subunits) indicated that, while HL-60 cells show constitutive activation of p50 only, HL-60R cells contain the activated forms of both p50 and p65 . Since p65 is necessary to form the NF-kappaB heterodimers able to increase transcription, its presence in HL-60R cells might well correlate to their increased levels of IAPs and, possibly of P-gp, which, reportedly, are NF-kappaB target genes . These results underline the possible role that the coordinated over-expression of the different IAPs may play in tumor cell resistance to drug induced apoptosis . Inhibition of NF-kappaB might be a useful strategy to block their up-regulation.

Clin Cancer Res, 2003 Nov 15, 9(15), 5768 - 75
Manganese superoxide dismutase expression correlates with chemosensitivity in human gastric cancer cell lines; Hur GC et al.; PURPOSE: The purpose is to investigate the potential correlation between antioxidant enzyme (AOE) levels and resistance to anticancer drugs in human gastric carcinoma cell lines . EXPERIMENTAL DESIGN: Protein contents of AOEs such as manganese superoxide dismutase (MnSOD), copper/zinc superoxide dismutase, catalase, glutathione S-transferase-pi, p-glycoprotein, and multidrug resistance-associated protein were observed by Western blot analysis, and MnSOD activity was measured in six Korean gastric cancer cell lines . The direct correlation between AOEs and the chemosensitivity to doxorubicin (DOX), mitomycin C, 5-fluorouracil, and vinblastine was analyzed by cytotoxicity test . MnSOD was overexpressed by transient transfection of human MnSOD cDNA . RESULTS: Expressions of AOEs in gastric cancer cell lines were variable . MnSOD expression was related with the resistance to DOX and mitomycin C but not with that to 5-fluorouracil and vinblastine . In comparison, expressions of other AOEs, p-glycoprotein and multidrug resistance-associated protein, were not correlated with tumor sensitivity to any of the drugs used . Cell lines with a high MnSOD protein content showed higher MnSOD activity than those with a low MnSOD protein content . In addition, MnSOD overexpression increased the resistance of gastric carcinoma cells to DOX . CONCLUSIONS: MnSOD is an important factor of drug response to reactive oxygen species-generating anticancer drugs in the gastric cancer cells . Thus, measurement of MnSOD levels in clinical samples may provide an indication of subsequent treatment response of gastric cancer patients.

Acta Pharmacol Sin, 2003 Dec, 24(12), 1235 - 40
Down-regulation of survivin expression reversed multidrug resistance in adriamycin-resistant HL-60/ADR cell line; Wang L et al.; AIM: To investigate the ability of an antisense RNA eukaryotic expression plasmid pcDNA3.1/survivin in down-regulating the expression level of survivin mRNA and survivin protein and reversed multidrug resistance (MDR) in adriamycin-resistant HL-60/ADR cell line . METHODS: The expression of survivin mRNA was measured by RT-PCR and the expression of survivin protein was measured by Western blot . Caspase-3 activity was determined by Phar Mingen colorimetric assay kit . Apoptosis was assessed by flow cytometry . The chemosensitivity of HL-60/ADR cells to adriamycin (ADR) was measured by MTT assay . RESULTS: pcDNA3.1/survivin down-regulated the expression level of survivin mRNA and survivin protein obviously, and induced apoptosis of HL-60/ADR cells in a time-dependent manner during 12-48 h . After transient transfection with pcDNA3.1/survivin for 48 h, survivin mRNA decreased by 67 %, survivin protein decreased by 57 %, caspase-3 activity increased 4.37 times, and the apoptosis rate increased by 4.41 % compared with control . Compared with ADR alone, pcDNA3.1/survivin significantly reversed MDR in HL-60/ADR cells, the chemosensitivity of HL-60/ADR cells to ADR was increased to 5.36 folds . CONCLUSION: pcDNA3.1/survivin down-regulated the expression level of survivin mRNA and survivin protein obviously, the threshold of apoptosis was decreased and MDR was reversed.

J Am Chem Soc, 2003 Dec 10, 125(49), 15049 - 58
Mechanism of the glutathione transferase-catalyzed conversion of antitumor 2-crotonyloxymethyl-2-cycloalkenones to GSH adducts; Hamilton DS et al.; Human glutathione (GSH) transferase (hGSTP1-1) processes with similar kinetic efficiencies the antitumor agents 2-crotonyloxymethyl-2-cyclohexenone (COMC-6), 2-crotonyloxymethyl-2-cycloheptenone (COMC-7), and 2-crotonyloxymethyl-2-cyclopentenone (COMC-5) to 2-glutathionylmethyl-2-cyclohexenone, 2-glutathionylmethyl-3-glutathionyl-2-cycloheptenone, and 2-glutathionylmethyl-2-cyclopentenone, respectively . This process likely involves initial enzyme-catalyzed Michael addition of GSH to the COMC derivative to give a glutathionylated enol(ate), which undergoes nonstereospecific ketonization, either while bound to the active site or free in solution, to a glutathionylated exocyclic enone . Free in solution, GSH reacts at the exomethylene carbon of the exocyclic enone, displacing the first GSH to give the final product . This mechanism is supported by the observation of multiphasic kinetics in the presence of high concentrations of hGSTP1-1 and the ability to trap kinetically competent exocyclic enones in aqueous acid using COMC-6 and COMC-7 as substrates . That the exocyclic enone is formed by nonstereospecific ketonization of an enol(ate) species is indicated by the observation that COMC-6 (chirally labeled with deuterium at the exomethylene carbon) gives stereorandomly labeled exocyclic enone . The isozymes hGSTP1-1, hGSTA1-1, hGSTA4-4, and hGSTM2-2 catalyze the conversion of COMC-6 to final product with similar efficiencies (K(m) = 0.08-0.34 mM, k(cat) = 1.5-6.1 s(-)(1)); no activity was detected with the rat rGSTT2-2 isozyme . Molecular docking studies indicate that in hGSTP1-1, the hydroxyl group of Tyr108 might serve as a general acid catalyst during substrate turnover . The possible significance of these observations with respect to the metabolism of COMC derivatives in multidrug resistant tumors is discussed.

Cancer Control, 2003 Nov-Dec, 10(6), 469 - 77
Management of acute myelogenous leukemia in the elderly; Rathnasabapathy R et al.; BACKGROUND: Acute myelogenous leukemia (AML) is a hematopoietic neoplasm that primarily affects older adults . Despite scientific advances into the epidemiologic, genetic, and biological features of AML, this disease remains fatal to the majority of patients, particularly older individuals . METHODS: We review the biologic and clinical characteristics of AML in the elderly and the treatment options that have emerged for them during the past several years . RESULTS: Several biologic features of AML differ between older and younger individuals . Older patients often have disease that expresses multidrug resistance phenotype and cytogenetic abnormalities, which may be responsible in large part for the poor outcomes observed in older-aged subgroups . Traditional cytotoxic chemotherapy is associated with a low complete response rate and a high treatment-related mortality in older patients, which explains in part the poorer outcomes in cohorts over 60 years of age . Research into the pathophysiology of AML has revealed an abundance of intracellular signaling events that govern proliferation and survival of the malignant cell . Such discoveries have promoted recognition of new molecular and antigenic targets (eg, Flt-3 kinase, Ras, CD33 antigen) to which therapeutic development may be aimed . CONCLUSIONS: New therapies directed against these unique targets may add to the current arsenal of antileukemic regimens and improve response rates and survival in older patients.

Pharmacol Ther, 2003 Dec, 100(3), 257 - 90
Mechanisms involved in the induced differentiation of leukemia cells; Tsiftsoglou AS et al.; Despite the remarkable progress achieved in the treatment of leukemias over the last several years, many problems (multidrug resistance {MDR}, cellular heterogeneity, heterogeneous molecular abnormalities, karyotypic instability, and lack of selective action of antineoplastic agents) still remain . The recent progress in tumor molecular biology has revealed that leukemias are likely to arise from disruption of differentiation of early hematopoietic progenitors that fail to give birth to cell lineage restricted phenotypes . Evidence supporting such mechanisms has been derived from studying bone marrow leukemiogenesis and analyzing differentiation of leukemic cell lines in culture that serve as models of erythroleukemic (murine erythroleukemia {MEL} and human leukemia {K562} cells) and myeloid (human promyelocytic leukemia {HL-60} cells) cell maturation . This paper reviews the current concepts of differentiation, the chemical/pharmacological inducing agents developed thus far, and the mechanisms involved in initiation of leukemic cell differentiation . Emphasis was given on commitment and the cell lineage transcriptional factors as key regulators of terminal differentiation as well as on membrane-mediated events and signaling pathways involved in hematopoietic cell differentiation . The developmental program of MEL cells was presented in considerable depth . It is quite remarkable that the erythrocytic maturation of these cells is orchestrated into specific subprograms and gene expression patterns, suggesting that leukemic cell differentiation represents a highly coordinated set of events that lead to irreversible growth arrest and expression of cell lineage restricted phenotypes . In MEL and other leukemic cells, differentiation appears to be accompanied by differentiation-dependent apoptosis (DDA), an event that can be exploited chemotherapeutically . The mechanisms by which the chemical inducers promote differentiation of leukemic cells have been discussed.

J Pharm Sci, 2004 Jan, 93(1), 99 - 107
Do multidrug resistance-associated protein-1 and -2 play any role in the elimination of estradiol-17 beta-glucuronide and 2,4-dinitrophenyl-S-glutathione across the blood-cerebrospinal fluid barrier?
Lee YJ, Kusuhara H, Sugiyama Y.
The purpose of this study was to examine the role of multidrug resistance-associated protein-1 and -2 (Mrp1 and Mrp2) in the efflux transport of organic anions across the blood-cerebrospinal fluid (CSF) barrier . The CSF concentration of estradiol-17beta-glucuronide (E(2)17betaG) and 2,4-dinitrophenyl-S-glutathione (DNP-SG) in the CSF after intracerebroventricular and intravenous injection were compared between wild-type and Mrp1 gene knockout mice . There was no significant difference in the apparent CSF elimination rate constants of E(2)17betaG (0.158 and 0.145 min(-1)) and DNP-SG (0.116 and 0.0779 min(-1)) between wild-type and Mrp1 knockout mice, respectively . After intravenous administration of E(2)17betaG, its brain-to-serum and CSF-to-serum concentration ratios in Mrp1 knockout mice were not significantly different from those in the wild-type . Results from in vivo and in vitro studies using Eisai hyperbilirubinemic rats, in which Mrp2 is hereditarily deficient, were similar to those using normal rats . Quantitative polymerase chain reaction (PCR) showed that the expression level of Mrp4 and Mrp5 was several times higher than that of Mrp1, whereas the expression levels of Mrp2, Mrp3, and Mrp6 were negligible or low . Therefore, Mrp4 and Mrp5 may contribute to the efflux transport of E(2)17betaG and DNP-SG from the CSF .

Int J Biochem Cell Biol, 2004 Feb, 36(2), 247 - 57
Multidrug resistance protein 4 (MRP4/ABCC4) mediates efflux of bimane-glutathione; Bai J et al.; Multidrug resistance proteins (MRPs) are ATP-dependent export pumps that mediate the export of organic anions . ABCC1 (MRP1), ABCC2 (MRP2) and ABCC3 (MRP3) are all able to facilitate the efflux of anionic conjugates including glutathione (GSH), glucuronide and sulfate conjugates of xenobiotics and endogenous molecules . Earlier studies showed that ABCC4 functions as an ATP-driven export pump for cyclic AMP and cyclic GMP, as well as estradiol-17-beta-D-glucuronide . However, it was unclear if other conjugated metabolites can be transported by ABCC4 . Hence in this study, a fluorescent substrate, bimane-glutathione (bimane-GS) was used to further examine the transport activity of ABCC4 . Using cells stably overexpressing ABCC4, this study shows that ABCC4 can facilitate the efflux of the glutathione conjugate, bimane-glutathione . Bimane-glutathione efflux increased with time and >85% of the conjugate was exported after 15min . This transport was abolished in the presence of 2.5microM carbonylcyanide m-chlorophenylhydrasone (CCCP), an uncoupler of oxidative phosphorylation . Inhibition was also observed with known inhibitors of MRP transporters including benzbromarone, verapamil and indomethacin . In addition, 100microM methotrexate, an ABCC4 substrate or 100microM 6-thioguanine (6-TG), a compound whose monophosphate metabolite is an ABCC4 substrate, reduced efflux by >40% . A concentration-dependent inhibition of bimane-glutathione efflux was observed with 1-chloro-2,4-dinitrobenzene (CDNB) which is metabolized intracellularly to the glutathione conjugate, 2,4-dinitrophenyl-glutathione (DNP-GS) . The determination that ABCC4 can mediate the transport of glucuronide and glutathione conjugates indicates that ABCC4 may play a role in the cellular extrusion of Phase II detoxification metabolites.

Chin Med J (Engl), 2003 Nov, 116(11), 1644 - 8
Arsenic trioxide inhibits p-glycoprotein expression in multidrug-resistant human leukemia cells that overexpress the MDR1 gene; Wei H et al.; OBJECTIVE: To investigate the effects of arsenic trioxide (As(2)O(3)) on the apoptosis and p-glycoprotein (P-gp) expression of multidrug-resistant human leukemia cells . METHODS: Human multidrug-resistant leukemia cell line K562/ADM overexpressing the MDR1 gene, was used as the target cells . The cell proliferating activity was assessed using the MTT colorimetric assay . Cytomorphology was investigated under light, confocal and electron microscopes . DNA fragmentation was examined using agarose gel electrophoresis, while p-gp expression, cell cycle status and sub-G1 cells were determined using flow cytometry . RESULTS: Zero point five to 20 micromol/L As(2)O(3) inhibited the proliferation of K562/ADM cells, and K562/ADM cells were more sensitive to As(2)O(3) than the parental K562 cells . As(2)O(3)-induced apoptosis of K562/ADM cells was determined by the observance of typical morphological changes and the appearance of DNA ladder and sub-G1 cell populations . As(2)O(3) significantly inhibited the P-gp expression of K562/ADM cells, and synergistically enhanced the sensitivity of the drug-resistant cells to adriamycin . CONCLUSIONS: As(2)O(3) induces growth-inhibition and apoptosis, down-regulates P-gp expression and exerts a synergistic effect in combination with adriamycin in multidrug-resistant leukemia cells.

J Exp Ther Oncol, 2003 May-Jun, 3(3), 127 - 35
Modulation of resistance to idarubicin by the cyclosporin PSC 833 (valspodar) in multidrug-resistant cells; Lacayo NJ et al.; Idarubicin (IDA) is an anthracycline anticancer drug utilized in the treatment of acute leukemias . There are conflicting data published with regard to the cross-resistance of IDA in multidrug-resistant (MDR) cells expressing P-glycoprotein (P-gp) . We evaluated the cytotoxicity and cellular accumulation of IDA in a panel of anthracycline-selected MDR cell lines . Leukemia K562/R7 cells and sarcoma MES-SA/Dx5 cells expressing high levels of the MDR1 (ABCB1) gene were resistant to IDA (42-fold and 150-fold, respectively) . In both of these cell lines, resistance to IDA was equivalent to that for doxorubicin, the drug used to select for the MDR variants . The P-gp inhibitor PSC 833 (valspodar) at 2 microM completely restored sensitivity to IDA . IDA accumulation was decreased 12-fold in MES-SA/Dx5 cells vs parental cell line, and drug uptake was restored to control levels by PSC 833 . Reduced intracellular IDA was correlated with P-gp content by flow cytometry . Experiments in NIH3T3 murine cells transfected with the human MDR1 gene substantiated the findings of cross-resistance to IDA and reversal of resistance by PSC 833 . Our data indicate that IDA is a high-affinity substrate for P-gp.

J Med Assoc Thai, 2003 Oct, 86(10), 953 - 63
Drug-resistant tuberculosis among prisoners of three prisons in Bangkok and the vicinity; Tansuphasiri U et al.; The purpose of this study was to determine the prevalence of drug-resistant tuberculosis and some factors associated with drug resistance among prisoners of three prisons in Bangkok and the vicinity . Susceptibility testing to four first-line antituberculous drugs was performed on 165 M . tuberculosis strains isolated from prisoners of three prisons including Klongprem Central (KC) prison, Bangkwang Central (BC) prison and the Correctional Institution (CI) for Male Drug Addicts . Of 165 smear positive tuberculosis (TB) cases with drugs susceptibility results, resistance to one or more drugs was 49.7 per cent . Resistance to one, two, three, and four drugs was 20.0, 13.3, 4.2 and 12.1 per cent, respectively . Multidrug resistant tuberculosis (MDR-TB) was 18.8 per cent . Patients classified as primary and acquired drug resistant were 6.7 and 50.0 per cent . The primary drug resistance to one or more drugs among prisoners at KC, BC and CI were 42.5, 36.4 and 53.9 per cent, respectively and MDR-TB were 8.2, 3.0, and 7.7 per cent, respectively . Of several factors analyzed in the present study, only a history of previous TB treatment was significantly associated with drug resistance (p < 0.05) . In conclusion, the results indicate the high prevalence of drug-resistant tuberculosis and the seriousness of the TB problem in prisons . The public health sector and prison authorities should work in close collaboration and co-ordination to continue improving TB case detection . Directly Observed Treatment Short course (DOTS) is highly recommended . Moreover, discharged prisoners with tuberculosis should be appropriately referred to hospitals or TB control centers.

Cytometry, 1996 Jan 1, 23(1), 78 - 81
Probenicid inhibition of fluorescence extrusion after MCB-staining of rat-1 fibroblasts; Poot M et al.; The intracellular fluorescence level of cells stained continuously with monochlorobimane was monitored by flow cytometry in order to assess the initial rate of glutatione to monochlorobimane conjugation as a measure of glutathione S-transferase activity . In addition to a rapid initial increase and a plateau level, a decline in fluorescence intensity was found upon prolonged flow cytometric monitoring . Exposure to probenicid, an inhibitor of an ATP-dependent organic anion pump, prevented this decrease . Incubation with vanadate and verapamil was without effect . Thus, extrusion of fluorescentglutathione-conjugate perturbs the proportionality between initial glutathione level and monochlorobimane-dependent fluorescence intensity . Monitoring by flow cytometry the decrease in monochlorobimane-dependent fluorescence may be useful to detect multidrug resistant cells.

Trop Med Int Health, 2003 Dec, 8(12), 1068 - 73
Molecular assessment of drug resistance in Plasmodium falciparum from Bahr El Gazal province, Sudan; Anderson TJ et al.; AIMS: To assess resistance to chloroquine (CQ) and sulphadoxine/pyrimethamine (SP) in a Sudanese parasite population . METHODS: Recurrent security problems in Akuem, Sudan, prevented us from conducting a classical in vivo treatment efficacy study . Instead we genotyped key mutations in the chloroquine resistance transporter (pfcrt), the multidrug resistance gene (pfmdr1), dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) . We genotyped the K76T mutation in pfcrt and the N86Y mutation in (pfmdr) by restriction digestion of fluorescent end-labelled polymerase chain reaction (PCR) products, while we genotyped codons 16, 51, 59, 108 and 164 in dhfr and codons 436, 437, 540, 581 and 613 in dhps by primer extension in 100 blood samples . RESULTS: Sixty-three percent of parasites carried the 76T mutation at pfcrt critical for CQ resistance, while 31% carried the 86Y mutation at pfmdr that is associated with, although not essential, for CQ resistance . We found five dhfr alleles: 60% of infections contained wild-type dhfr alleles, 3% had one mutation, 34% had two mutations, while 3% had three mutations . We found three dhps alleles: 47% were wild type, 44% had one mutation, while 9% had two mutations . CONCLUSIONS: We expect high levels of treatment failure (RI-RIII) with CQ (20-40%) and predict efficient treatment with SP . However, dhfr alleles with three mutations (51I, 59R, 108N) are present as are dhps alleles with two mutations (437G, 540E) . Successful treatment with SP is therefore likely to be short-lived.

Biochem Soc Trans, 2003 Dec, 31(Pt 6), 1372 - 7
Protecting the genome: defence against nucleotide glycation and emerging role of glyoxalase I overexpression in multidrug resistance in cancer chemotherapy; Thornalley PJ; Glycation of nucleotides in DNA forms AGEs (advanced glycation end-products) . Nucleotide AGEs are: the imidazopurinone derivative dG-G {3-(2'-deoxyribosyl)-6,7-dihydro-6,7-dihydroxyimidazo{2,3-b}purin-9(8)one}, CMdG ( N (2)-carboxymethyldeoxyguanosine) and gdC (5-glycolyldeoxycytidine) derived from glyoxal, dG-MG {6,7-dihydro-6,7-dihydroxy-6-methylimidazo-{2,3-b}purine-9(8)one}, dG-MG(2) { N (2),7-bis-(1-hydroxy-2-oxopropyl)deoxyguanosine} and CEdG { N (2)-(1-carboxyethyl)deoxyguanosine} derived from methylglyoxal, and dG-3DG { N (2)-(1-oxo-2,4,5,6-tetrahydroxyhexyl)deoxyguanosine} derived from 3-deoxyglucosone and others . Glyoxal and methylglyoxal induce multi-base deletions, and base-pair substitutions - mostly occurring at G:C sites with G:C-->C:G and G:C-->T:A transversions . Suppression of nucleotide glycation by glyoxalase I and aldehyde reductases and dehydrogenases, and base excision repair, protects and recovers DNA from damaging glycation . The effects of DNA glycation may be most marked in diabetes and uraemia . Mutations arising from DNA glycation may explain the link of non-dietary carbohydrate intake to incidence of colorectal cancer . Overexpression of glyoxalase I was found in drug-resistant tumour cells and may be an example of an undesirable effect of the enzymatic protection against DNA glycation . Experimental overexpression of glyoxalase I conferred resistance to drug-induced apoptosis . Glyoxalase I-mediated drug resistance was found in human leukaemia and lung carcinoma cells . Methylglyoxal-mediated glycation of DNA may contribute to the cytotoxicity of some antitumour agents as a consequence of depletion of NAD(+) by poly(ADP-ribose) polymerase, marked increases in triosephosphate concentration and increased formation of methylglyoxal . S - p -Bromobenzylglutathione cyclopentyl diester is a cell-permeable glyoxalase I inhibitor . It countered drug resistance and was a potent antitumour agent against lung and prostate carcinoma . Glyoxalase I overexpression was also found in invasive ovarian cancer and breast cancer.

Biochemistry, 2003 Dec 9, 42(48), 14099 - 113
Identification of the structural and functional boundaries of the multidrug resistance protein 1 cytoplasmic loop 3; Westlake CJ et al.; Multidrug resistance protein (MRP) 1 is a member of the ABCC branch of the ATP binding cassette (ABC) transporter superfamily that can confer resistance to natural product chemotherapeutic drugs and transport a variety of conjugated organic anions, as well as some unconjugated compounds in a glutathione- (GSH-) dependent manner . In addition to the two tandemly repeated polytopic membrane-spanning domains (MSDs) typical of ABC transporters, MRP1 and its homologues MRP2, -3, -6, and -7 contain a third NH(2)-terminal MSD . The cytoplasmic loop (CL3) connecting this MSD, but apparently not the MSD itself, is required for MRP1 leukotriene C(4) (LTC(4)) transport activity, substrate binding and appropriate trafficking of the protein to the basolateral membrane . We have used a baculovirus dual-expression system to produce various functionally complementing fragments of MRP1 in insect Sf21 cells to precisely define the region in CL3 that is required for activity and substrate binding . Using a parallel approach in polarized MDCK-I cells, we have also defined the region of CL3 that is required for basolateral trafficking . The CL3 NH(2)- and COOH-proximal functional boundaries have been identified as Cys(208) and Asn(260), respectively . Cys(208) also corresponds to the NH(2)-proximal boundary of the region required for basolateral trafficking in MDCK-I cells . However, additional residues downstream of the CL3 COOH-proximal functional boundary extending to Lys(270) were found to be important for basolateral localization . Finally, we show that regions in CL3 necessary for LTC(4) binding and transport are also required for binding of the photoactivatable GSH derivative azidophenacyl-GSH.

Int J Cancer, 2004 Jan 10, 108(2), 212 - 8
Analysis of gene expression profiles in human HL-60 cell exposed to cantharidin using cDNA microarray; Zhang JP et al.; Cantharidin is a natural toxin that has antitumor properties and causes leukocytosis as well as increasing sensitivity of tumor cells resistant to other chemotherapeutic agents . There is limited information, however, on the molecular pharmacological mechanisms of cantharidin on human cancer cells . We have used cDNA microarrays to identify gene expression changes in HL-60 promyeloid leukemia cells exposed to cantharidin . Cantharidin-treated cells not only decreased expression of genes coding for proteins involved in DNA replication (e.g., DNA polymerase delta), DNA repair (e.g., FANCG, ERCC), energy metabolism (e.g., isocitrate dehydrogenase alpha, ADP/ATP translocase), but also decreased expression of genes coding for proteins that have oncogenic activity (e.g., c-myc, GTPase) or show tumor-specific expression (e.g., phosphatidylinositol 3-kinase) . In contrast, these treated cells overexpressed several genes that encode intracellular and secreted growth-inhibitory proteins (e.g., BTG2, MCP-3) as well as proapoptotic genes (e.g., ATL-derived PMA-responsive peptide) . Our findings suggest that alterations in specific genes functionally related to cell proliferation or apoptosis may be responsible for cantharidin-mediated cytotoxicity . We also found that exposure of HL-60 cells to cantharidin resulted in the decreased expression of multidrug resistance-associated protein genes (e.g., ABCA3, MOAT-B), suggesting that cantharidin may be used as an oncotherapy sensitizer, and the increased expression of genes in modulating cytokine production and inflammatory response (e.g., NFIL-3, N-formylpeptide receptor), which may partly explain the stimulating effects on leukocytosis . Our data provide new insight into the molecular mechanisms of cantharidin .

Biochem Pharmacol, 2003 Dec 15, 66(12), 2381 - 95
Ecteinascidin-743 drug resistance in sarcoma cells: transcriptional and cellular alterations; Shao L et al.; A human chondrosarcoma cell line, CS-1, was treated successively with increasing concentrations of the marine chemotherapeutic Ecteinascidin-743 (ET-743), yielding a variant cell line displaying a significant degree of resistance to the cytotoxic action of this drug . Various experiments were performed to discern molecular aberrations between the parent and resistant cell line, and also identify potential molecular markers indicative of drug resistance . Although no significant differences in the levels of membrane transporters such as P-glycoprotein or multidrug resistance protein 1 (MRP1) were detected, the cell migratory ability of the ET-743-resistant cell variant was reduced, as was its attachment capability to gelatin-coated cell culture dishes . Staining of the actin-containing cytoskeleton with fluorescent-labeled phalloidin revealed marked differences in the cytoskeleton architecture between the parent and ET-743-resistant CS-1 cell lines . Comparison of serum-free conditioned medium from both cell lines showed conspicuous differences in the levels of several proteins, including a quartet of high molecular weight proteins (> or =140 kDa) . The protein sequences of two of these high molecular weight proteins, present at significantly higher concentrations in conditioned medium obtained from the parent cell line, corresponded to subunits of types I and IV collagen . Analysis of type I collagen alpha1 chain mRNA revealed a significantly lower level in the ET-743-resistant CS-1 cell line . Thus, prolonged exposure to ET-743 may cause changes in cell function through cytoskeleton rearrangement and/or modulation of collagen levels.

Acta Trop, 2003 Dec, 89(1), 47 - 53
Randomised efficacy and safety study of two 3-day artesunate rectal capsule/mefloquine regimens versus artesunate alone for uncomplicated malaria in Ecuadorian children; Gomez EA et al.; The combination of artesunate and mefloquine is one of the most effective treatments against multidrug-resistant falciparum malaria . Experience in children is however limited . The objective of this study was to compare the efficacy and safety of two artesunate/mefloquine combinations with artesunate monotherapy in Ecuadorian children . A total of 150 children with an age between 2 and 12 years, confirmed to have uncomplicated falciparum malaria, were randomly selected and divided in three treatment groups of 50 patients each . Group 1 received 50 mg rectal capsules alone (40 mg/kg total dose) administered over 6 days . Group 2 received 50 mg rectal capsules (30 mg/kg total dose) for 3 days combined with mefloquine (20 mg/kg total dose) on day 1 . Group 3 was treated with 50 mg rectal capsules (30 mg/kg total dose) for 3 days, combined with mefloquine on days 1 and 3 (15-17 mg/kg total dose) . Patients were continuously followed up and controlled by clinical and laboratory examinations for 7 days as well as on days 14, 21 and 28 . An additional parasite examination was performed at 2 months following therapy . Clearance of parasitaemia was comparable between treatment groups . These were 9.2, 9.2 and 8.3 h for Groups 1, 2 and 3, respectively . Cure rates at day 28 were 76, 96 and 94% and after 2 months 60, 88 and 80%, respectively . There were no adverse events (AEs) reported during the study . Vital signs and laboratory examinations revealed no changes of clinical relevance . It can be concluded that the combination of artesunate rectal capsules with mefloquine is effective and safe . Starting concomitant administration already on day 1 is well tolerated . This combination significantly reduces the incidence of recrudescence compared to artesunate monotherapy . Comparing the two tested artesunate/mefloquine regimens, a total mefloquine dose of 20 mg/kg seems to be more effective compared to a total dose of 15-17 mg/kg . Further studies seem to be warranted.

Biochim Biophys Acta, 2003 Nov 20, 1639(3), 213 - 24
P-glycoprotein-mediated multidrug resistance phenotype of L1210/VCR cells is associated with decreases of oligo- and/or polysaccharide contents; Fiala R et al.; Multidrug resistance of murine leukaemic cell line L1210/VCR (obtained by adaptation of parental drug-sensitive L1210 cells to vincristine) is associated with overexpression of mdr1 gene product P-glycoprotein (Pgp)-the ATP-dependent drug efflux pump . 31P-NMR spectra of L1210 and L1210/VCR cells (the latter in the presence of vincristine) revealed, besides the decrease of ATP level, a considerable lower level of UDP-saccharides in L1210/VCR cells . Histochemical staining of negatively charged cell surface binding sites (mostly sialic acid) by ruthenium red (RR) revealed a compact layer of RR bound to the external coat of sensitive cells . In resistant cells cultivated in the absence or presence of vincristine, the RR layer is either reduced or absent . Consistently, resistant cells were found to be less sensitive to Concanavalin A (ConA) . Moreover, differences in the amount and spectrum of glycoproteins interacting with ConA-Sepharose were demonstrated between sensitive and resistant cells . Finally, the content of glycogen in resistant cells is lower than in sensitive cells . All the above facts indicate that multidrug resistance of L1210/VCR cells mediated predominantly by drug efflux activity of Pgp is accompanied by a considerable depression of oligo- and/or polysaccharides biosynthesis.

J Control Release, 2003 Dec 5, 93(2), 151 - 60
Drug delivery to resistant tumors: the potential of poly(alkyl cyanoacrylate) nanoparticles; Vauthier C et al.; Simultaneous cellular resistance to multiple lipophilic drugs represents a major problem in cancer chemotherapy . This drug resistance may appear clinically either as a lack of tumor size reduction or as the occurrence of clinical relapse after an initial positive response to antitumor treatment . The resistance mechanism can have different origins either directly linked to specific mechanisms developed by the tumor tissue or connected to the more general problem of distribution of a drug towards its targeted tissue . The purpose of this paper is to summarize the results of the use of poly(alkyl cyanoacrylate) nanoparticles to overcome multidrug resistance (MDR) phenomena at both the cellular and the non-cellular level.

Epilepsia, 2003 Nov, 44(11), 1388 - 96
Major vault protein, a marker of drug resistance, is upregulated in refractory epilepsy; Sisodiya SM et al.; PURPOSE: The molecular basis of drug resistance in epilepsy is being explored . Two proteins associated with drug resistance in cancer, P-glycoprotein and multidrug resistance-associated protein 1, are upregulated in human epileptogenic pathologies . Other proteins associated with resistance in cancer include major vault protein (MVP) and breast cancer resistance protein (BCRP) . We hypothesized that these proteins would also be upregulated in human epileptogenic pathologies . METHODS: Hippocampal sclerosis (HS), focal cortical dysplasia (FCD), and dysembryoplastic neuroepithelial tumor (DNT) were studied by using immunohistochemistry for MVP and BCRP . Nonepileptogenic control and histologically normal brain adjacent to epileptogenic tissue were used for comparison . RESULTS: MVP and BCRP were expressed ubiquitously in brain capillary endothelium . Ectopic upregulation of MVP was seen in hilar neurons in HS, dysplastic neurons in FCD, and lesional neurons in DNT . Only in HS cases were rare extralesional neurons immunoreactive . Glial upregulation was not seen . There was no qualitative upregulation of BCRP . CONCLUSIONS: These results show that more than one resistance protein may be upregulated in a given epileptogenic pathology and may contribute to drug resistance . Determination of the types, amounts, and distribution of such proteins will be necessary for rational treatment for drug resistance in epilepsy.

Epilepsia, 2003 Dec, 44(12), 1479 - 86
Brain access and anticonvulsant efficacy of carbamazepine, lamotrigine, and felbamate in ABCC2/MRP2-deficient TR- rats; Potschka H et al.; PURPOSE: Different adenosine triphosphate (ATP)-driven multidrug transporters have been described to be expressed in the luminal membrane of blood-brain barrier (BBB) endothelial cells . At this site, multidrug transporters have been suggested to restrict penetration of drugs into the brain . Increasing evidence suggests that overexpression of different multidrug transporters occurs in the region of the epileptic focus of pharmacoresistant epilepsy patients . Based on the assumption that antiepileptic drugs (AEDs) are substrates of these transporters, this overexpression may limit access of AEDs to epileptic neurons and may contribute to drug-refractoriness . In a recent study, overexpression of multidrug resistance protein 2 (ABCC2; MRP2) was reported in BBB endothelial cells of epileptic focal tissue from pharmacoresistant patients . With brain microdialysis, we recently demonstrated that the AED phenytoin is subject to transport by ABCC2 at the BBB, whereas phenobarbital does not seem to be a substrate of ABCC2 . METHODS: We investigated whether ABCC2 is functionally involved in transport of the AEDs carbamazepine (CBZ), lamotrigine (LTG), and felbamate (FBM) across the BBB . The distribution of these AEDs into the brain of ABCC2-deficient TR- rats was determined . RESULTS: AED concentrations in plasma and brain extracellular space of these mutant rats did not differ significantly from those of rats of the corresponding background strain . In the amygdala-kindling model of epilepsy, the anticonvulsant efficacy of LTG and FBM was comparable in both groups of rats . In contrast, CBZ exhibited a higher anticonvulsant activity in kindled ABCC2-deficient rats as compared with nonmutant rats . CONCLUSIONS: In this present study, the microdialysis results gave no evidence that ABCC2 function modulates entry of CBZ, LTG, and FBM into the CNS of naive rats . However, ABCC2 deficiency was associated with an increased anticonvulsant response of CBZ in the kindling model . Future investigations are planned to identify the underlying mechanism for this difference, clarifying whether a pharmacokinetic difference is detectable only when brain access of CBZ is compared in kindled ABCC2-deficient rats and kindled nonmutant rats, which may have an increased expression of ABCC2 in response to seizures . The data substantiate that ABCC2-deficient TR- rats are a useful tool for defining the role of ABCC2 for transport of AEDs, and give evidence that the use of kindled TR- rats may provide important supplementary information.

J Comput Aided Mol Des, 2003 May-Jun, 17(5-6), 291 - 7
MCASE study of the multidrug resistance reversal activity of propafenone analogs; Klopman G et al.; A database containing 130 propafenone type chemicals which have been tested for their multidrug resistance (MDR) reversal activity was compiled . Using the Multiple Computer-Automated Structure Evaluation (MCASE) program to analyze this database, an underlying relationship between MDR reversal activity and octanol/water partition coefficient was found . An MDR reversal model was created based on this database by the baseline activity identification algorithm (BAIA) of the MCASE program . The main phamacophores relevant to MDR reversal activity were identified.

Apoptosis, 1999 Aug, 4(4), 233 - 7
Caveolin-1, a metastasis-related gene that promotes cell survival in prostate cancer; Thompson TC et al.; Metastasis represents the ultimate target in cancer therapy as this complex biological process is the direct cause of mortality for a variety of human malignancies . The current high level of mortality from prostate cancer results in large part from the inexorable growth of overt or occult metastasis present at the time of diagnosis . Currently, there are no curative therapies for metastatic prostate cancer . To better understand the metastatic phenotype in prostate cancer, we developed a strategy to identify mRNAs that are expressed differentially in cell lines derived from primary versus metastatic mouse prostate cancer using differential display-PCR . In using this system a number of metastasis-related sequences were identified including a cDNA that encodes caveolin-1 . Caveolin-1 was found to be overexpressed not only in metastatic mouse prostate cancer, but also in human metastatic disease . Recent studies have indicated that suppression of caveolin-1 expression induces androgen sensitivity in high caveolin-1, androgen-insensitive mouse prostate cancer cells derived from metastases . Conversely, overexpression of caveolin-1 leads to androgen insensitivity in low caveolin, androgen-sensitive mouse prostate cancer cells . Caveolin-1, therefore, is both a metastasis-related gene as well as a candidate androgen resistance gene for prostate cancer in man . Interestingly, recent studies also point to a potential role for caveolin-1 in the resistance of various malignancies to multiple antineoplastic agents . The linkage of caveolin-1 expression with the androgen-resistant phenotype in prostate cancer and the multidrug resistance phenotype in various solid tumors establishes a novel paradigm for understanding these clinically important and now potentially related processes in malignant progression.

EMBO J, 2003 Dec 1, 22(23), 6175 - 81
Three-dimensional structure of the bacterial multidrug transporter EmrE shows it is an asymmetric homodimer; Ubarretxena-Belandia I et al.; The small multidrug resistance family of transporters is widespread in bacteria and is responsible for bacterial resistance to toxic aromatic cations by proton-linked efflux . We have determined the three-dimensional (3D) structure of the Escherichia coli multidrug transporter EmrE by electron cryomicroscopy of 2D crystals, including data to 7.0 A resolution . The structure of EmrE consists of a bundle of eight transmembrane alpha-helices with one substrate molecule bound near the centre . The substrate binding chamber is formed from six helices and is accessible both from the aqueous phase and laterally from the lipid bilayer . The most remarkable feature of the structure of EmrE is that it is an asymmetric homodimer . The possible arrangement of the two polypeptides in the EmrE dimer is discussed based on the 3D density map.

Blood, 2004 Apr 1, 103(7), 2727 - 37 Epub 2003 Nov 20.
Small lymphocytic lymphoma, marginal zone B-cell lymphoma, and mantle cell lymphoma exhibit distinct gene-expression profiles allowing molecular diagnosis; Thieblemont C et al.; Non-germinal center small B-cell lymphomas represent a heterogeneous group of non-Hodgkin lymphomas, the most frequent histologic subtypes being small lymphocytic lymphoma (SLL), splenic marginal zone B-cell lymphoma (MZL), and mantle cell lymphoma (MCL) . In order to identify genomic signatures specific for each disease, we analyzed 128 primary tumors using high-density microarrays . Several clusters of genes significantly discriminated the 3 histologic subtypes . Genes associated with cell adhesion, angiogenesis, and inhibition of apoptosis were up-regulated in SLL . Genes associated with intracellular signaling via the AKT1 pathway were up-regulated in splenic MZL . Genes associated with cell cycle control and multidrug resistance were up-regulated in MCL . Using 44 genes selected within the gene clusters discriminant for the 3 lymphoma subtypes, we generated a class prediction score that allowed us to classify the 3 entities in 96% of the cases, including borderline cases . Whereas specific transcriptional profiles easily distinguished all MZL samples, SLL samples, and most of the MCL samples into separate groups, few MCL cases exhibited MZL-type transcriptional profiles . This study demonstrates that SLL, splenic MZL, and MCL possess specific transcriptional profiles that may be relevant to the pathogenesis and the diagnosis of these histologic subtypes.

FEBS Lett, 2003 Nov 27, 555(1), 102 - 5
Multidrug resistance ABC transporters; Chang G; Clinical multidrug resistance is caused by a group of integral membrane proteins that transport hydrophobic drugs and lipids across the cell membrane . One class of these permeases, known as multidrug resistance ATP binding cassette (ABC) transporters, translocate these molecules by coupling drug/lipid efflux with energy derived from the hydrolysis of ATP . In this review, we examine both the structures and conformational changes of multidrug resistance ABC transporters . Together with the available biochemical and structural evidence, we propose a general mechanism for hydrophobic substrate transport coupled to ATP hydrolysis.

Cancer Biother Radiopharm, 2003 Oct, 18(5), 791 - 801
The multidrug resistance of in vitro tumor cell lines derived from human breast carcinoma MCF-7 does not influence pentavalent technetium-99m-dimercaptosuccinic Acid uptake; Denoyer D et al.; The main causes of multidrug resistance (MDR) are overexpression of P-glycoprotein (P-gp) and multidrug resistance-associated protein isoform 1 (MRP1) often associated with high levels of glutathione (GSH) . We investigated whether MDR phenotype can influence Tc-99m-(V)-DMSA {pentavalent technetium-99m-dimercaptosuccinic acid} entry by comparing its uptake with that of Tc-99m-sestamibi (MIBI) on an in vitro model of sensitive (MCF-7) and variant resistant cell lines . Drug resistance was assessed by immunoblotting, GSH measurement, and 3-{4,5-dimethylthiazol-2-yl}-2,5,diphenyl tetrazolium bromide (MTT) assay . To correlate MDR phenotype with tracer accumulation, uptakes were performed with and without P-gp and MRP1 inhibitors and after GSH modulation . Similar accumulation of Tc-99m-(V)-DMSA was observed in all cell lines and the use of MDR reversals did not enhance its uptake . Our results demonstrate clearly that Tc-99m-(V)-DMSA uptake is not related to either P-gp and MRP1 expression, or GSH levels . In contrast, Tc-99m-MIBI accumulation is inversely proportional to the cell MDR phenotype . The combination of Tc-99m-(V)-DMSA and Tc-99m-MIBI may be a useful tool for noninvasive detection of malignant sites and their chemoresistance status.

J Org Chem, 2003 Nov 28, 68(24), 9506 - 9
Total synthesis of dendroamide a: oxazole and thiazole construction using an oxodiphosphonium salt; You SL et al.; The total synthesis of dendroamide A (1), a multidrug-resistance reversing bistratamide-type peptide-derived macrocycle, has been accomplished in 19% yield . Fmoc-protected amino acids were condensed into appropriately protected dipeptides which were treated with bis(triphenyl)oxodiphosphonium trifluoromethanesulfonate to afford oxazoles and thiazolines (oxidized to thiazoles) with high chemo- and stereoselectivity . The convergent condensation of three heterocyclic amino acids followed by macrocyclization afforded the natural product.

Yao Xue Xue Bao, 2003 Aug, 38(8), 565 - 70
Experimental chemotherapy against xenografts derived from multidrug resistant KBv200 cells and parental drug-sensitive KB cells in nude mice by annonaceous acetogenin 89-2; Fu LW et al.; AIM: Annonaceous acetogenin 89-2 was obtained from atemoya plant . To investigate the effect of 89-2 on experimental chemotherapy against xenografts derived from multidrug resistant KBv200 cells and parental drug-sensitive KB cells . METHODS: Cytotoxicity was determined by tetrazolium (MTT) assay . The models of KB and KBv200 xenografts in nude mice were established to investigate the effect of 89-2 on experimental chemotherapy against cancer in vivo . Mechanistic experiments were conducted to examine the function of P-gp by Fura 2-AM assay . RESULTS: The compound 89-2 showed potent cytotoxicity in KBv200 and KB cells, and the mean IC50 of 89-2 to KBv200 and KB cells was 48.7 and 64.6 nmol.L-1, respectively . The IC50 of 89-2 to multidrug resistant (MDR) cells was similar to that to the parental drug-sensitive cells (P < 0.05) . In the models of KBv200 and KB cell xenografts in nude mice, 89-2 (0.90 mg.kg-1, q2d x 6) exhibited 52.3% and 56.5% in inhibiting the growth of xenografts, respectively . The toxicity was endurable . The intracellular accumulation of Fura-2 in KBv200 cells increased to 1.66, 2.03, and 2.74-fold, respectively, by addition of 12.8, 64 and 320 nmol.L-1 of 89-2 . CONCLUSION: Both MDR KBv200 cells and parental drug-sensitive KB cells were sensitive to the treatment of 89-2 in vitro and in vivo . The mechanism of overcoming MDR was associated with the decrease of P-gp function MDR cells.

J Biol Chem, 2004 Feb 13, 279(7), 5734 - 8 Epub 2003 Nov 19.
ATP-binding cassette transporters are enriched in non-caveolar detergent-insoluble glycosphingolipid-enriched membrane domains (DIGs) in human multidrug-resistant cancer cells; Hinrichs JW et al.; In this study we show that P-glycoprotein in multidrug-resistant 2780AD human ovarian carcinoma cells and multidrug resistance-associated protein 1 in multidrug-resistant HT29col human colon carcinoma cells are predominantly located in Lubrol-based detergent-insoluble glycosphingolipid-enriched membrane domains . This localization is independent of caveolae, since 2780AD cells do not express caveolin-1 . Although HT29col cells do express caveolin-1, the ATP-binding cassette transporter and caveolin-1 were dissociated on the basis of differential solubility in Triton X-100 and absence of microscopical colocalization . While both the multidrug resistance-associated protein 1 and caveolin-1 are located in Lubrol-based membrane domains, they occupy different regions of these domains.

Biochem Biophys Res Commun, 2003 Nov 28, 311(4), 891 - 6
Upregulation in the expression of multidrug resistance protein Mrp1 mRNA and protein by increased bilirubin production in rat; Cekic D et al.; Earlier studies suggest that Mrp1 may mediate ATP-dependent cellular extrusion of unconjugated bilirubin (UCB) . We studied the serial responses of expression of Mrp1 mRNA and protein in rats with increased bilirubin production due to hemolysis induced by phenylhydrazine (PHZ) treatment . Mrp1 mRNA was analyzed by quantitative PCR and protein by Western blot . Hepatic expression of Mrp1 mRNA and protein peaked at day 3 of PHZ treatment . Splenic expression of Mrp1 mRNA peaked within 24h and returned to baseline at day 5 whereas Mrp1 protein expression peaked at day 3 . Pretreatment with heme-oxygenase inhibitor, tin mesoporphyrin, blunted the increase in serum UCB and erased the overexpression of Mrp1 both in liver and spleen . Thus, the upregulation of Mrp1 in hemolysis is mediated by UCB and/or other products of heme oxygenase, further supporting a role of Mrp1 in UCB transport and protection from its cellular toxicity.

Mol Microbiol, 2003 Nov, 50(4), 1141 - 53
ParG, a protein required for active partition of bacterial plasmids, has a dimeric ribbon-helix-helix structure; Golovanov AP et al.; The ParG protein (8.6 kDa) is an essential component of the DNA partition complex of multidrug resistance plasmid TP228 . ParG is a dimer in solution, interacts with DNA sequences upstream of the parFG genes and also with the ParF partition protein both in the absence and presence of target DNA . Here, the solution nuclear magnetic resonance structure of ParG is reported . The ParG dimer is composed of a folded domain formed by two closely intertwined C-terminal parts (residues 33-76), and two highly mobile tails consisting of N-terminal regions (residues 1-32) . The folded part of ParG has the ribbon-helix-helix (RHH) architecture similar to that of the Arc/MetJ superfamily of DNA-binding transcriptional repressors, although the primary sequence similarity is very low . ParG interacts with DNA predominantly via its folded domain; this interaction is coupled with ParG oligomerization . The dimeric RHH structure of ParG suggests that it binds to DNA by inserting the double-stranded beta-sheet into the major groove of DNA, in a manner similar to transcriptional repressors from the Arc/MetJ superfamily, and that ParG can function as a transcriptional repressor itself . A new classification of proteins belonging to the Arc/MetJ superfamily and ParG homologues is proposed, based on the location of a conserved positively charged residue at either the beginning or at the end of the beta-strand which forms part of the DNA recognition motif.

J Neurochem, 2003 Nov, 87(4), 1010 - 23
P-glycoprotein in blood-brain barrier endothelial cells: interaction and oligomerization with caveolins; Jodoin J et al.; P-glycoprotein (P-gp), an adenosine triphosphate (ATP)-binding cassette transporter which acts as a drug efflux pump, is highly expressed at the blood-brain barrier (BBB) where it plays an important role in brain protection . Recently, P-gp has been reported to be located in the caveolae of multidrug-resistant cells . In this study, we investigated the localization and the activity of P-gp in the caveolae of endothelial cells of the BBB . We used an in vitro model of the BBB which is formed by co-culture of bovine brain capillary endothelial cells (BBCEC) with astrocytes . Caveolar microdomains isolated from BBCEC are enriched in P-gp, cholesterol, caveolin-1, and caveolin-2 . Moreover, P-gp interacts with caveolin-1 and caveolin-2; together, they form a high molecular mass complex . P-gp in isolated caveolae is able to bind its substrates, and the caveolae-disrupting agents filipin III and nystatin decrease P-gp transport activity . In addition, mutations in the caveolin-binding motif present in P-gp reduced the interaction of P-gp with caveolin-1 and increased the transport activity of P-gp . Thus, P-gp expressed at the BBB is mainly localized in caveolae and its activity may be modulated by interaction with caveolin-1.

J Neurochem, 2003 Nov, 87(4), 820 - 30
P-glycoprotein (ABCB1) but not multidrug resistance-associated protein 1 (ABCC1) is induced by doxorubicin in primary cultures of rat astrocytes; Mercier C et al.; At least two drug efflux pumps involved in multidrug resistance, P-glycoprotein (P-gp) and multidrug resistance-associated protein 1 (Mrp1), are expressed in rat astrocyte primary cultures . The aim of this study was to compare the expression of P-gp and Mrp1 in primary cultures exposed to 50 or 500 ng/mL doxorubicin (DOX) . Among the two P-gp genes expressed in rodents, mdr1a and mdr1b, a time- and dose-dependent increase in mdr1b mRNA levels was revealed by northern blot analysis . This up-regulation was inhibited by actinomycin D and occurred as early as 2 h after exposure to 50 or 500 ng/mL DOX, whereas mdr1a and mrp1 transcripts were not modified by the DOX exposure . In addition, DOX also strongly enhanced, in a time- and dose-dependent manner, P-gp but not Mrp1 expression . Moreover, DOX raised the cellular efflux of vincristine, a substrate for both P-gp and Mrp1 . This efflux was inhibited by the P-gp modulators PSC833 and GW918, but not by the Mrp1 modulator MK571 . On the other hand, a 24-h exposure to 500 ng/mL DOX, but not 50 ng/mL DOX, induced apoptosis in primary cultures of rat astrocytes . Fumonisin B1, a ceramide synthase inhibitor, reduced DOX-induced apoptosis, suggesting that de novo synthesis of the ceramide regulatory pathway might be involved in DOX-induced apoptosis . Moreover, western blot analysis showed that fumonisin B1 was not able to decrease the overexpression of P-gp induced by DOX . Our results provide evidence that DOX up-regulates a functional P-gp in primary cultures of rat astrocytes and might cause astrocyte apoptosis via the ceramide pathway.

Biochemistry, 2003 Nov 25, 42(46), 13659 - 66
Multidrug resistance to HIV-1 protease inhibition requires cooperative coupling between distal mutations; Ohtaka H et al.; The appearance of viral strains that are resistant to protease inhibitors is one of the most serious problems in the chemotherapy of HIV-1/AIDS . The most pervasive drug-resistant mutants are those that affect all inhibitors in clinical use . In this paper, we have characterized a multiple-drug-resistant mutant of the HIV-1 protease that affects indinavir, nelfinavir, saquinavir, ritonavir, amprenavir, and lopinavir . This mutant (MDR-HM) contains six amino acid mutations (L10I/M46I/I54V/V82A/I84V/L90M) located within and outside the active site of the enzyme . Microcalorimetric and enzyme kinetic measurements indicate that this mutant lowers the affinity of all inhibitors by 2-3 orders of magnitude . By comparison, the multiiple-drug-resistant mutant only increased the K(m) of the substrate by a factor of 2, indicating that the substrate is able to adapt to the changes caused by the mutations and maintain its binding affinity . To understand the origin of resistance, three submutants containing mutations in specific regions were also studied, i.e., the active site (V82A/I84V), flap region (M46I/I54V), and dimerization region (L10I/L90M) . None of these sets of mutations by themselves lowered the affinity of inhibitors by more than 1 order of magnitude, and additionally, the sum of the effects of each set of mutations did not add up to the overall effect, indicating the presence of cooperative effects . A mutant containing only the four active site mutations (V82A/I84V/M46I/I54V) only showed a small cooperative effect, suggesting that the mutations at the dimer interface (L10I/L90M) play a major role in eliciting a cooperative response . These studies demonstrate that cooperative interactions contribute an average of 1.2 +/- 0.7 kcal/mol to the overall resistance, most of the cooperative effect (0.8 +/- 0.7 kcal/mol) being mediated by the mutations at the dimerization interface . Not all inhibitors in clinical use are affected the same by long-range cooperative interactions between mutations . These interactions can amplify the effects of individual mutations by factors ranging between 2 and 40 depending on the inhibitor . Dissection of the energetics of drug resistance into enthalpic and entropic components provides a quantitative account of the inhibitor response and a set of thermodynamic guidelines for the design of inhibitors with a lower susceptibility to this type of mutations.

Eur Respir J, 2003 Nov, 22(5), 833 - 7
Multidrug-resistant tuberculosis: eight years of surveillance in France; Robert J et al.; The aim of this study was to evaluate the annual prevalence of multidrug-resistant tuberculosis (MDRTB) and to describe the characteristics of the patients with MDRTB in France . Annual questionnaire surveys from 1992-1999 were mailed to all French microbiological laboratories performing mycobacterial cultures . A total of 264 distinct patients were reported to the National Reference Centre for Resistance of Mycobacteria to Antituberculosis Drugs during the 8-yr surveillance period resulting in a mean annual prevalence of MDRTB of 0.6% . A mean of 16% of the MDRTB patients were reported over several subsequent years . The majority of patients were male (69.7%), foreign-born (55.7%), with a previous history of treatment (65.9%), and pulmonary involvement (92.8%) with smear-positive results (59.1%) . Human immunodeficiency virus (HIV) coinfection was present in 20.8% of the patients . Strains were resistant only to isoniazid and rifampin in 37.9% of the cases, and additional resistance to both streptomycin and ethambutol was present in 25.8% . HIV coinfection and female status were statistically associated with primary resistance, whereas smear-positive results were associated with secondary resistance . Foreign-birth and smear-positive results were associated with a chronic status . The prevalence of multidrug-resistant tuberculosis is low in France (<1%) . However, a substantial proportion of patients remain positive for several years, suggesting nonoptimal management . Therefore, as recommended by the World Health Organization, a few reference teams, working in collaboration with national associations of physicians and microbiologists, should be established to improve the outcome of multidrug-resistant tuberculosis.

Pharm Res, 2003 Oct, 20(10), 1581 - 90
Sensitization of cells overexpressing multidrug-resistant proteins by pluronic P85; Batrakova EV et al.; PURPOSE: This study evaluated the chemosensitizing effects of Pluronic P85 (P85) on cells expressing multidrug resistance-associated proteins, MRPI and MRP2 . METHODS: Cell models included MRP1- and MRP2-transfected MDCKII cells as well as doxorubicin-selected COR-L23/R cells overexpressing MRP1 . Effects of P85 on cellular accumulation and cytotoxicity of vinblastine and doxorubicin were determined . Mechanistic studies characterized the effects of P85 on ATP and reduced glutathione (GSH) intracellular levels as well as MRP ATPase and glutathione-S-transferase (GST) activities in these cells . RESULTS: Considerable increases of vinblastine and doxorubicin accumulation in the cells overexpressing MRP1 and MRP2 in the presence of P85 were observed, although no statistically significant changes in drug accumulation in the parental cells were found . P85 treatment caused an inhibition of MRP ATPase activity . Furthermore, P85 induced ATP depletion in these cells similar to that previously reported for Pgp-overexpressing cells . In addition, reduction of GSH intracellular levels and decrease of GST activity were observed following P85 treatment . Finally, significant enhancement of cytotoxicity of vinblastine and doxorubicin by P85 in MRP-overexpressing cells was demonstrated . CONCLUSIONS: This study suggests that P85 can sensitize cells overexpressing MRP1 and MRP2, which could be useful for chemotherapy of cancers that display these resistant mechanisms.

Sichuan Da Xue Xue Bao Yi Xue Ban, 2003 Oct, 34(4), 684 - 7
{Expression of multidrug resistance-associated protein 1 in osteosarcoma and its relationship with clinicopathologic characteristics}; Tu C et al.; OBJECTIVE: To detect the expression of multidrug resistance-associated protein 1 (MRP1) in human ostesarcoma and to explore its relationship with clinicopathologic characteristics . METHODS: Six normal bone specimens and 45 osteosarcomas from patients were analysed for MRP1 expression by immunohistochemistry . The expression levels of MRP1 with the clinicopathologic parameters of the patients were examined using Chisquare test . RESULTS: The expression of MRP1 was observed in 32 (71.11%) osteosarcoma specimens . More(P < 0.05) specimens expressed MRP1 in high-grade osteosarcomas(30/38, 78.95%) than in low-grade osteosarcomas (2/7, 28.57%) . The correlation coefficient between expression of MRP1 and grade of pathology was 0.844 . In a limited number of patients, the expression of MRP1 was not related to the the age and sex of the patients, Enneking surgical stage, osteosarcoma size, serum concentration of ALP and duration before diagnosis . No expression of MRP1 in 6 normal bone specimens was observed . CONCLUSION: MRP1 is expressed in osteosarcoma and its expression is positively correlated with the malignancy of osteosarcoma.

Int J Cancer, 2004 Jan 1, 108(1), 146 - 51
Reversal of breast cancer resistance protein (BCRP/ABCG2)-mediated drug resistance by novobiocin, a coumermycin antibiotic; Shiozawa K et al.; Breast cancer resistance protein (BCRP/ABCG2) of an ATP-binding cassette half-transporter confers resistance against mitoxantrone and camptothecin derivatives of topotecan and irinotecan . Novobiocin, a coumermycin antibiotic, is known to enhance anticancer drug sensitivity of cancer cells in vitro and in vivo, the mechanism of which remains undetermined . Here we focused on drug efflux pump and examined whether novobiocin reversed drug resistance in multidrug-resistant cells highly expressing BCRP . To explore the reversal mechanisms, intracellular drug accumulation was measured by flow cytometry, and a topotecan transport study using plasma membrane vesicles was performed . We used PC-6/SN2-5H2 small cell lung cancer and MCF-7/MX breast cancer cells selected with SN-38 of the active irinotecan metabolite and mitoxantrone, respectively, and the BCRP cDNA transfectant MCF-7/clone 8 cells . These cells expressed high levels of BCRP mRNA but not other known transporters . Compared to the parental PC-6 cells, PC-6/SN2-5H2 cells were 141-, 173- and 57.2-fold resistant to topotecan, SN-38 and mitoxantrone, respectively . Novobiocin at 60 microM decreased the degree of the above resistance by approximately 26-fold in PC-6/SN2-5H2 cells, and similarly reversed resistance in MCF-7/MX, MCF-7/clone 8 and un-selected NCI-H460 cells highly expressing BCRP . Furthermore, novobiocin increased the intracellular topotecan accumulation in these cells and inhibited the topotecan transport into the membrane vesicles of PC-6/SN2-5H2 cells . No effects of novobiocin in these assay were observed in the parental PC-6 and MCF-7 cells . The kinetic parameters in the transport study indicated that novobiocin was a inhibitor for BCRP, resulting in competitive inhibition of BCRP-mediated topotecan transport . These findings suggest that novobiocin effectively overcomes BCRP-mediated drug resistance at acceptable concentrations .

Int J Cancer, 2004 Jan 1, 108(1), 78 - 85
Altered conjugate formation and altered apoptosis of multidrug-resistant human leukemia cell line affects susceptibility to killing by activated natural killer (NK) cells; Treichel RS et al.; Most leukemias that exhibit P-glycoprotein (P-gp)-associated multidrug resistance (MDR) exhibit reduced susceptibility to immune cytotoxicity mediated by natural killer (NK) cells . To explore this phenomenon we investigated N6/ADR, a doxorubicin-selected, P-gp-positive variant of the human acute lymphoblastic leukemia (ALL) cell line NALM6 . Each stage of the NK cytolytic pathway, (binding, activation and killing) was evaluated to identify the alterations responsible for the reduced cytotoxicity of the variant relative to its drug-sensitive parental line . The major cause of the decreased susceptibility to NK cytolysis was found to be reduced conjugate formation by the MDR variant . Activation of NK effectors by parental and MDR cells with concomitant release of tumor necrosis factor-alpha (TNF-alpha) correlated with conjugate formation . N6/ADR was also more resistant than NALM6 to antibody-dependent cellular cytotoxicity and to cytotoxic factors released from NK cells as measured both by 51Cr-release and by DNA fragmentation . This is the first report of a P-gp-positive leukemic line that exhibits reduced conjugate formation as well as increased resistance to NK-mediated killing mechanisms . Our results suggest caution in the use of NK-based immunotherapy as an alternative treatment for multidrug-resistant leukemias .

Cell Mol Life Sci, 2003 Oct, 60(10), 2164 - 77
Multiple flavonoid-binding sites within multidrug resistance protein MRP1; Trompier D et al.; Recombinant nucleotide-binding domains (NBDs) from human multidrug resistance protein MRP1 were overexpressed in bacteria and purified to measure their direct interaction with high-affinity flavonoids, and to evaluate a potential correlation with inhibition of MRP1-mediated transport activity and reversion of cellular multidrug resistance . Among different classes of flavonoids, dehydrosilybin exhibited the highest affinity for both NBDs, the binding to N-terminal NBD1 being prevented by ATP . Dehydrosilybin increased vanadate-induced 8-N3-{alpha-32P}ADP trapping, indicating stimulation of ATPase activity . In contrast, dehydrosilybin strongly inhibited leukotriene C4 (LTC4) transport by membrane vesicles from MRP1-transfected cells, independently of reduced glutathione, and chemosensitized cell growth to vincristine . Hydrophobic C-isoprenylation of dehydrosilybin increased the binding affinity for NBD1, but outsite the ATP site, lowered the increase in vanadate-induced 8-N3-{alpha-32P}ADP trapping, weakened inhibition of LTC4 transport which became glutathione dependent, and induced some cross-resistance . The overall results indicate multiple binding sites for dehydrosilybin and its derivatives, on both cytosolic and transmembrane domains of MRP1.

Mol Cancer Ther, 2003 Nov, 2(11), 1207 - 14
Involvement of oligosaccharide changes in alpha5beta1 integrin in a cisplatin-resistant human squamous cell carcinoma cell line; Nakahara S et al.; Multiple mechanisms are involved in the resistance of cancer cells to cisplatin, including the expression of multidrug resistance-associated protein (MRP) and enhanced DNA repair . Here, we report findings to show that oligosaccharide changes in alpha5beta1 integrin are associated with cisplatin resistance in a head and neck squamous cell carcinoma cell line, HSC-2 . Cisplatin-resistant HSC-2 (HSC-2/CR) cells were established by stepwise treatment with various concentrations of cisplatin . The oligosaccharides containing beta1, 6-N-acetylglucosamine (beta1-6GlcNAc) branching, detected by leukoagglutinating phytohemagglutinin (L(4)-PHA) lectin blot, were found to be dramatically decreased in alpha5beta1 integrin immunoprecipitated from HSC-2/CR cells . To better understand the mechanisms underlying cisplatin resistance and oligosaccharide alteration, we analyzed the downstream signaling of alpha5beta1 integrin, one of the target glycoproteins of beta1-6GlcNAc transferase {UDP-GlcNAc:alpha-D-mannoside beta1, 6-N-acetylglucosaminyltransferase (GnT-V)} . Cell adhesion to fibronectin and phosphorylation of focal adhesion kinase (FAK), which are associated with alpha5beta1 integrin and involved in a cell survival signaling, were found to be increased in the cisplatin-resistant cells . Enhancement of the inhibition of cell adhesion and FAK phosphorylation also support the above data in GnT-V transfectants of HSC-2 cells . Interestingly, the differences in sensitivity to cisplatin and FAK phosphorylation between cisplatin-sensitive and -resistant cells were completely abolished by treatment with a neutral antibody of alpha5beta1 integrin . These results suggest that modification of oligosaccharides of alpha5beta1 integrin represents one of the possible mechanisms of drug resistance in head and neck cancer cells.

Mol Cancer Ther, 2003 Nov, 2(11), 1195 - 205
Taxane-based reversal agents modulate drug resistance mediated by P-glycoprotein, multidrug resistance protein, and breast cancer resistance protein; Brooks TA et al.; Overexpression of ATP-binding cassette transport proteins, including P-glycoprotein (Pgp), multidrug resistance (MDR) protein (MRP-1), and breast cancer resistance protein (BCRP), is a well-characterized mechanism of MDR in tumor cells . Although the cytotoxic taxanes paclitaxel and docetaxel are substrates for Pgp-mediated efflux, the semisynthetic taxane analogue ortataxel inhibits drug efflux mediated by Pgp as well as, as we recently demonstrated, MRP-1 and BCRP . Nevertheless, ortataxel is not optimal for development as a clinical MDR modulator because of its cytotoxicity {corrected} . We sought to identify noncytotoxic taxane-based broad-spectrum modulators from a library of noncytotoxic taxane-based reversal agents (tRAs) designed by eliminating the C-13 side chain of the taxane molecule, which inhibits microtubule depolymerization . Twenty tRAs, selected based on modulation of paclitaxel cytotoxicity in Pgp-overexpressing MDA435/LCC6(mdr1) cells, were studied for modulation of retention and cytotoxicity of substrates of MRP-1 and BCRP as well as Pgp in established cell lines overexpressing each of these transporters . Four tRAs modulated MRP-1 and 17 modulated BCRP in addition to Pgp . The four broad-spectrum tRAs strongly modulated daunorubicin and mitoxantrone efflux and enhanced their cytotoxicity in cell lines overexpressing the three MDRs, decreasing IC(50) values by as much as 97% {corrected} . These tRAs, especially tRA 98006, have promise for development as clinical broad-spectrum MDR modulators and warrant more preclinical analysis to determine pharmacokinetic interactions and efficacy.

J Acquir Immune Defic Syndr, 2003 Dec 1, 34(4), 398 - 402
Colinearity of reverse transcriptase inhibitor resistance mutations detected by population-based sequencing; Gonzales MJ et al.; High-level resistance to multiple drugs is often detected by directly sequencing uncloned polymerase chain reaction products (population-based sequencing) . It is not known, however, if this method of identifying mutations gives an accurate picture of individual viral genomes . To determine how often multidrug-resistant isolates consist of clones containing every mutation present in the population-based sequence, a mean of 2.8 molecular clones was sequenced from the plasma of 25 heavily treated persons whose population-based sequence contained multiple reverse transcriptase (RT) inhibitor resistance mutations (71 clones) . The 25 population-based sequences contained a mean of 5.7 nucleoside reverse transcriptase inhibitor (NRTI) resistance mutations and 1.2 nonnucleoside reverse transcriptase inhibitor (NNRTI) resistance mutations . The 71 clones contained a mean of 5.3 NRTI resistance mutations and 1.0 NNRTI resistance mutations . Sequences of clones closely resembled the population-based sequence: 36 (51%) clones had each of the RT inhibitor mutations present in the population-based sequence, 25 (35%) had all but 1 RT inhibitor mutation, 4 (6%) had all but 2 RT inhibitor mutations, 3 (4%) had all but 3 RT inhibitor mutations, and 3 (4%) had all but 4 RT inhibitor mutations . Phenotypic testing of 29 clones showed that most clones were resistant to nearly all NRTIs and that those with NNRTI resistance mutations were also resistant to multiple NNRTIs . These data show that in heavily treated persons, most RT inhibitor resistance mutations are present in the same viral genomes (colinear) and that multidrug resistance often occurs within individual clones as well as within virus populations.

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