Am J Infect Control, 1996 Aug, 24(4), 226 - 34 Status of tuberculosis infection control programs at United States hospitals, 1989 to 1992 . APIC . Association for Professionals in Infection Control and Epidemiology; Sinkowitz RL et al.; BACKGROUND: Recent nosocomial outbreaks have raised concern about the risk of Mycobacterium tuberculosis transmission in United States hospitals . METHODS: To determine current tuberculosis (TB) infection control practices, we surveyed a sample of approximately 3000 acute care facilities about the number of patients with drug-susceptible or multidrug-resistant TB (MDR-TB), health care worker (HCW) tuberculin skin test (TST) results, and compliance with the 1990 Centers for Disease Control and Prevention (CDC) TB guidelines . Analyses were restricted to one response per hospital . RESULTS: Personnel at 1494 (49.8%) hospitals returned a completed survey . Respondent hospitals had a mean of 881 HCWs (range 8 to 10,000) and 196 (range 6 to 2450) beds; 62% percent were community nonteaching hospitals . Of respondent hospitals providing data for 1989 through 1992, the proportion that cared for patients with TB or MDR-TB increased from 46.4% to 56.6% and 0.8% to 4.5%, respectively . The pooled mean HCW TST positivity rate at hire rose from 0.95% to 1.14%, and the pooled mean HCW TST conversion rate increased from 0.40% to 0.51% . In 1992, when we compared hospitals with zero, one to five, or six or greater patients with TB, the risk of a positive HCW TST result at hire or at routine testing significantly increased with increasing number of patients with TB . From 1989 through 1992, the number of hospitals reporting the use of surgical masks for HCW respiratory protection decreased from 96.8% to 66.8% . In 1992, 66% of the hospitals reported compliance with four or more of the AFB isolation room criteria specified in the 1990 CDC TB guidelines . CONCLUSIONS: Contrary to prior surveys, this study shows that many U.S . community hospitals admit patients with TB less frequently than do teaching hospitals, and infrequently admit patients with MDR-TB . Because the risk of HCW TST conversion varies with hospital characteristics, these data show the importance of performing a risk assessment, as recommended in the CDC TB guidelines, for each ward and hospital so that TB control measures can be individualized. Bull Cancer, 1996 Aug, 83(8), 609 - 18 {MDR (Multiple Drug Resistant) type of resistance to chemotherapy in clinical practice}; Oudard S et al.; Multifactorial resistance is the main mechanism of chemotherapy failure in cancers . Multidrug resistance (MDR) is related to the expression of a 170 kDa membrane glycoprotein, the so-called P-glycoprotein (P-gp) . This protein is able to extrude drugs of various structures and mechanisms out of the cytoplasm . P-gp is a pronostic value in hemopathy as well as in child sarcoma, osteosarcoma and neuroblastoma . Modulator agents of different generations are capable of inhibiting P-gp . MDR modulation is obtained in hemopathies and is associated with an eradication of the P-gp (+) cell clones . In solid tumors, clinical trials using verapamil or cyclosporin are not so convincing . It is likely that other mechanisms of resistance are responsible for tumor progression, such as the MRP system, glutathion and topoisomerases . A better knowledge of multifactorial resistance and drug synthesis counteracting these resistance mechanisms will allow to elaborate new therapeutic basis for cancer therapy. Br J Pharmacol, 1996 Aug, 118(8), 1879 - 85 Evidence for P-glycoprotein-modulated penetration of morphine-6-glucuronide into brain capillary endothelium; Huwyler J et al.; 1 . Morphine-6-glucuronide is one of the major metabolites of morphine . The potent analgesic action of this compound together with its potential lower apparent toxicity in man, when compared with morphine, indicated its clinical importance . 2 . Primary cultures of porcine brain capillary endothelial cells were used to study brain penetration of morphine-6-glucuronide . Biochemical characterization of the cell cultures revealed a marked enrichment in enzymatic activity of alkaline phosphatase (56 fold) and angiotensin converting enzyme (230 fold) as compared to whole brain tissue . By immunostaining the presence of vimentin, factor VIII, the tight junction associated protein ZO-1, and P-glycoprotein was shown . Functional characterization revealed that the carrier system responsible for transport of neutral amino acids was intact . 3 . Uptake and transport of morphine-6-glucuronide was marginal and in the range of the extracellular marker sucrose . However, uptake of morphine-6-glucuronide was enhanced significantly (P < 0.0001) in presence of the inhibitors of P-glycoprotein, verapamil or vincristine . The finding that morphine-6-glucuronide may serve as a substrate for P-glycoprotein was confirmed in multidrug-resistant P388 tumour cells . 4 . We conclude that penetration of the blood-brain barrier by morphine-6-glucuronide may depend on the expression of the product of the multidrug-resistance (MDR) gene in brain capillary endothelial cells. J Neurooncol, 1996 Aug, 29(2), 149 - 55 Effects of hypoxia on drug resistance phenotype and genotype in human glioma cell lines; Liang BC; Recurrent gliomas are most often treated by chemotherapy . However, these tumors typically acquire resistance to most drugs administered, and patients will usually die of recurrent tumor . Factors which may play a role include overexpression of putative multidrug resistance genes, such as the multidrug resistance gene 1 (MDR1), multidrug resistance associated protein gene (MRP), 06-alkylguanine, DNA alkyltransferase gene (06MT) and excision repair cross complementing gene 1 (ERCC1) . Tumor hypoxia has also been shown to be associated with drug resistance in other soft tissue tumors . Since gliomas have regions of diminished oxygenation, and have clinical resistance to chemotherapy, the relationship between phenotypic resistance to chemotherapy after hypoxic exposure and expression of drug resistance genes was investigated in glioma cell lines (U373 MG, PFAT-MT) . After a 24 hour exposure to hypoxia, drugs 1, 3-bis, 2-chloroethyl-1-nitrosurea (BCNU) and cis-diammine, dichloroplatinum II (CDDP) were administered, and cell survival was determined . Hypoxic exposure was associated with increased survival of the cell lines after administration of BCNU and CDDP, with resistance to BCNU 15 to 30-fold when compared to cells which did not undergo hypoxic exposure . Both tumor cell lines also showed some degree of resistance to CDDP, although not to the extent of BCNU (2 to 3-fold increased resistance) . The expression of the drug resistance genes was found to be unchanged when comparing cells which had undergone hypoxic exposure and those which had not . Thus, hypoxic exposure is associated with substantial drug resistance in brain tumor cell lines . The lack of correlation between the induced phenotype and known drug resistance genes suggests other mechanisms may be acting in these tumors in hypoxic conditions. Eur J Haematol, 1996 Aug, 57(2), 142 - 8 Maintenance therapy with interferon-alpha (IFN-alpha) versus IFN-alpha plus chemotherapy in multiple myeloma (MM) . The Greek Myeloma Study Group; Zervas K et al.; Results of studies using IFN-alpha treatment for maintaining remission and prolonging survival in multiple myeloma (MM) are in conflict and trials seeking optimum use for this biological response modifier are continuing . Between 1989 and 1993 a prospective randomized multicentre trial was undertaken to evaluate the role of the combination of IFN-alpha with chemotherapy (CT) in maintenance treatment of MM . For remission induction, in patients 65 yr or younger, we used VAD (group A) and for the remaining Melphalan and Prednisone (MP) (group B) . For maintenance, patients were randomized to receive IFN-alpha 3 x 10(6) i.u . s.c . t.i.w . (group I) or alternating monthly cycles of IFN-alpha and CT . The CT cycles were also alternated (VAD, MP, CP) in an effort to prevent the development of multidrug resistance . Median survival of the two maintenance groups from randomization (36 months for group I and 31 months for group II, p = 0.3) as well as response duration (13 months in group I and 15 months in group II, p = 0.95) were similar . Toxicities were more pronounced both with VAD induction and in the combination maintenance arm . The addition of chemotherapy to the IFN maintenance regimen in MM did not have an advantage over IFN alone. Nippon Hinyokika Gakkai Zasshi, 1996 Aug, 87(8), 1041 - 7 {Enhancement of cytotoxic effect of anticancer agents of renal cell carcinoma}; Okano M et al.; BACKGROUND: Human renal adenocarcinomas do not adequately respond to cancer chemotherapy . Their multidrug resistance is mainly conferred by the P-glycoprotein (P-gp) . In this study, we analyzed effects of P-gp modulators on enhancement of anticancer activities against human renal cell carcinomas . METHODS: ACHN/ADM human renal adenocarcinoma cells with a high level expression of P-gp and 28 surgical specimens of renal cell adenocarcinomas were recruited . Adriamycin (ADM) and vinblastin (VLB) were used as anticancer drugs, and verapamil (Ver) and cyclosporin A (CsA) were as P-gp modulators . The chemosensitivity was determined by the ATP-assay . RESULTS: Ver and CsA exhibited 1.5-fold and 6-fold increase, respectively, in the anticancer activities of ADM against ACHN/ADM cells . The anticancer activities of VLB were also enhanced by the modulators; 7-fold for Ver and 11-fold for CsA . In the chemosensitivity test of clinical specimens, the cancer for which the viability of the cells assessed by the ATP-assay was 50% or less than 50% after exposure to the anticancer drug with or without a P-gp modulator was defined as sensitive to the drug . Of the 14 clinical specimens exposed to anticancer drugs without Ver, only 3 tumors and 1 tumor were sensitive to ADM and VLB, respectively, whereas with Ver, 6 tumors and 4 tumors were sensitive to ADM and VLB, respectively . Of the other 14 clinical specimens exposed to anticancer drugs without CsA, only 3 tumors and no tumor were sensitive to ADM and VLB, respectively, whereas, with CsA, 9 tumors and 6 tumors were sensitive to ADM and VLB, respectively . CONCLUSION: This study indicate that Ver and CsA have effects on enhancement of the anticancer activities of ADM and VLB against human renal adenocarcinomas . The addition of Ver or CsA to chemotherapy will be a potential circumvention of P-gp-mediated multidrug resistance of renal cell adenocarcinomas. Rinsho Ketsueki, 1996 Aug, 37(8), 640 - 6 {Multidrug resistance in acute leukemia}; Takeshita A et al.; We examined the multidrug resistance (MDR) P-glycoprotein (P-gp) on normal bone marrow (BM) cells and acute myeloid leukaemia (AML) cells, using newly devised flow cytometric multi-parameter analysis with CD33, CD34 and MRK16 monoclonal antibodies . With biotinylated MRK16 and a Streptavidin-RED670 (SA-RED670) conjugate, we succeeded in the detection of a small amount of P-gp on these cells . In normal bone marrow cells, the percentage of P-gp positive CD34+CD33-, CD34+CD33+ and CD34-CD33+ cells were 12.2 (2.2% (mean +/- standard deviation), 6.3 +/- 3.1% and 1.4 +/- 0.9%, respectively . By the more precise list-mode analysis, myeloid lineage cells showed continuously regressing P-gp expression as they maturated from CD34+CD33- to CD34-CD33+ cells . In AML cells at diagnosis, CD34+ CD33- cells expressed P-gp strongly, CD34+CD33+ cells moderately, and CD34-CD33+ cells weakly, showing the same tendency as observed in normal BM cells . Blast cells from acute promyelocytic leukemia (APL), which mainly expressed CD34-CD33+ but no detectable CD34+CD33- at diagnosis, expressed less amount of P-gp than the other subtypes of AML . P-gp expression on these three phenotypes increased in relapsed cases, especially on the CD34+CD33- subpopulation. Exp Parasitol, 1996 Aug, 83(3), 295 - 303 Plasmodium falciparum: amplification and overexpression of pfmdr1 is not necessary for increased mefloquine resistance; Lim AS et al.; Amplification and overexpression of the pfmdr1 gene has been associated with mefloquine resistance in Plasmodium falciparum . We have selected for parasites more resistant to mefloquine from P . falciparum FAC8, a clone that has three copies of pfmdr1 . The parasite lines derived from this selection were up to threefold more resistant to mefloquine . The mefloquine-selected parasites were also more resistant to quinine and halofantrine, suggesting a multidrug resistance phenotype . In contrast to previous findings, the selection of P . falciparum FAC8 on mefloquine did not alter the copy number or the level of expression of pfmdr1 . Sequence analysis of pfmdr1 from the selected lines showed no change in the amino acids . These results show that alterations in pfmdr1 are not involved in mediating the increased mefloquine resistance observed in this clone. Nippon Rinsho, 1996 Aug, 54(8), 2276 - 90 {Hepatic metabolism and transport of bilirubin and other organic anions}; Adachi Y et al.; Most of bilirubin, bile acids and other organic anions are preferentially taken up by the liver and excreted into bile . Recently many transporters on the sinusoidal and canalicular membranes of the hepatocytes have been reported for each ligand . complementary DNA was cloned for human Na+/taurocholate cotransporting polypeptide (NTCP) which mediates sodium dependent secondary active hepatic uptake of bile acids . For the hepatic uptake of non-bile acid-organic anions such as bilirubin, at least 4 transporters are postulated, i.e., bilirubin/BSP binding protein (BBBP), organic anion binding protein (OABP), bilitranslocase, and organic anion transporting polypeptide (OATP) . In the hepatocytes, bilirubin is glucuronidated in the endoplasmic reticulum . The gene for UDP-glucuronosyltransferase (UGT) 1 family has been elucidated and differential splicing from several exons 1 (A to J) results in forming isozymes of UGT 1 including bilirubin UGT . At the canalicular membranes, two main ATP-dependent organic anion transporters have been reported, i.e., canalicular bile salt transporter (cBST) for bile acids and canalicular multispecific organic anion transporter (cMOAT) for non-bile acid organic anions . Recently multidrug resistance protein (MRP) is reported closely related to or identical to cMOAT . These canalicular ATP-dependent transporters are called ABC (ATP-binding cassette) transporters. Tuber Lung Dis, 1996 Aug, 77(4), 315 - 21 Bacille Calmette Guérin immunization of health care workers exposed to multidrug-resistant tuberculosis: a decision analysis; Stevens JP et al.; SETTING: North American health care workers with exposure to multidrug-resistant tuberculosis . OBJECTIVE: To evaluate the relative utilities of bacille Calmette Guerin (BCG) immunization and post-infection chemoprophylaxis for the protection of health care workers exposed to multidrug-resistant Mycobacterium tuberculosis . DESIGN: Decision analysis using SMLTREE software and published data for probabilities . RESULTS: BCG vaccination was preferred by a small margin over post-infection chemoprophylaxis . Sensitivity analysis revealed that possible changes in probability values used tended to tilt the result towards use of BCG vaccination . The threshold for protective efficacy of BCG vaccination was 26% . CONCLUSIONS: BCG vaccination should be considered for health care workers in environments where there is a substantial risk of exposure to multidrug-resistant tuberculosis. Eur J Biochem, 1996 Aug 1, 239(3), 796 - 804 Expression, subcellular distribution and response to phorbol esters of protein kinase C (PKC) isozymes in drug-sensitive and multidrug-resistant KB cells evidence for altered regulation of PKC-alpha; Cloud-Heflin BA et al.; Protein kinase C (PKC) comprises a family of related phospholipid-dependent serine/threonine protein kinases . PKC has been implicated in the induction and maintenance of the multidrug-resistance (MDR) phenotype but the role of different isozymes is not well understood . We compared the expression and subcellular distribution, and membrane association and down-regulation induced by phorbol esters, of individual PKC isozymes in drug-sensitive KB-3 and multidrug-resistant KB-V1 human carcinoma cell lines . Immunoblotting with isozyme-specific antibodies indicated the presence of PKC alpha (cytosol only) . PKC beta (membrane only) . PKC epsilon (mainly membrane associated) and PKC zeta (both fractions) . PKC delta and PKC gamma were not detected . The expression levels of PKC beta . PKC epsilon and PKC zeta were unchanged in KB-V1 cells; PKC alpha was modestly increased ( approximately 65%) in the resistant cells as further determined by enzyme assay . The cytosolic nature and increased expression of PKC alpha were confirmed by immunofluorescent localization studies . Revertant cells, obtained by culturing KB-V1 cells in a drug-free medium, regained drug sensitivity with a loss of P-glycoprotein and a concomitant decrease in expression of PKC alpha, KB-V1 cells were found to differ markedly from KB-3 cells with respect to the translocation and down-regulation specifically of PKC alpha upon exposure to 12-O-tetradecanoyl-1-phorbol-13-acetate (TPA) . Treatment with 30 nM TPA for 24 h completely depleted KB-3 cells of PKC alpha whereas 1 microM TPA was required to deplete KB-V1 cells of PKC alpha . Similar results were obtained when phorbol-12, 13-dibutyrate was used instead of TPA . Defective TPA-mediated down-regulation of PKC alpha was also observed in another PKC alpha-overexpressing MDR cell line . KB-A1 . Importantly, cellular uptake of radiolabeled phorbol ester was similar for both drug-sensitive and MDR cells . Sensitive and resistant cells exhibited similar expression levels of RACK1, a PKC-binding protein important in activation-induced translocation . These findings further highlight the importance of PKC alpha in the MDR phenotype, and suggest that this isozyme may be expressed in a modified form or be subject to an altered regulation in MDR cells. Exp Hematol, 1996 Aug, 24(10), 1258 - 64 Monoclonal antibodies against P-glycoprotein, an MDR1 gene product, inhibit interleukin-2 release from PHA-activated lymphocytes; Raghu G et al.; P-glycoprotein (Pgp), a product of the human MDR1 gene, is a member of the ABC superfamily of transporters responsible for the trafficking of biologically active substances across the membrane . In tumors, Pgp is associated with multidrug resistance (MDR), the phenomenon characterized by the ability of cells to efflux structurally diverse lipophilic compounds . It has been demonstrated that Pgp is also expressed on various types of normal human tissues and cells, including hematopoietic stem cells, T, B, and natural killer (NK) cells . The normal physiologic function of Pgp in immune cells is unclear . In this study, we used highly specific and nontoxic monoclonal antibodies (mAbs) against external epitopes of Pgp (mAb UIC2, its monovalent Fab fragments, and mAb MRK16) to inhibit Pgp-mediated efflux and investigate a possible role of Pgp in activated T lymphocytes . We found that the treatment of phytohemagglutinin (PHA)-stimulated peripheral blood leukocytes (PBL) with these mAbs resulted in a significant reduction of interleukin-2 (IL-2) levels in the culture . Early activation events, as measured by intracellular calcium flux, expression of the CD69 early activation marker, and expression of IL-2 mRNA, were not affected by anti-Pgp mAbs . These results suggest that the Pgp efflux pump may be involved in the transport of IL-2 in T lymphocytes. Cancer Res, 1996 Aug 1, 56(15), 3490 - 4 Interaction of P-glycoprotein with protein kinase C in human multidrug resistant carcinoma cells; Yang JM et al.; Indirect evidence has suggested that P-glycoprotein (P-gp), the multidrug transporter, is phosphorylated by protein kinase C (PKC) and that phosphorylation modulates its transport function . To address the first premise more directly, ie., that P-gp is phosphorylated by PKC, we investigated the interaction between P-gp and PKC in sensitive and multidrug resistant MCF-7 and KB human carcinoma cell lines . We found that P-gp and PKC were coimmunoprecipitated from the multidrug-resistant cell lines MCF-7/AdrR and KB-V-1, using antibodies to either protein . The association between the two proteins was enhanced by phorbol 12-myristate 13-acetate, an analogue of diacylglycerol that induces translocation of PKC to the plasma membrane . The anti-P-gp immunoprecipitates contained PKC activity as measured by direct phosphorylation reactions . The interaction of PKC with P-gp displayed isozyme specificity: PKC-alpha, -beta, gamma, -epsilon, and -phi, but not -delta, -mu, -zeta, -lambda, were found to coimmunoprecipitate with P-gp . These studies indicate that P-gp closely interacts with PKC and serves as a substrate, and that specific isozymes of this kinase may be involved in the phosphorylation of the multidrug transporter. Cancer Res, 1996 Aug 1, 56(15), 3577 - 82 Transfection of glutathione S-transferase (GST)-pi antisense complementary DNA increases the sensitivity of a colon cancer cell line to adriamycin, cisplatin, melphalan, and etoposide; Ban N et al.; The goal of this study was to demonstrate that glutathione S-transferase (GST)-pi is directly involved in the intrinsic and acquired resistance of cancer cells to anticancer drugs . To this end, GST-pi antisense cDNA was transfected into the cultured human colon cancer cell line M7609, which expresses an innately high level of GST-pi and shows intrinsic drug resistance, and into an M7609 strain with acquired resistance to Adriamycin (ADR;i.e., M7609/ADR cells) . The changes in the sensitivity of these transfectants to various anticancer drugs were investigated . The intracellular concentrations of GST-pi in M7609/anti-1 cells and M7609/anti-2 cells, two clones that were established by transfection of GST-pi antisense cDNA into M7609 cells, were decreased to approximately half of those detected in the parent cells (M7609) and in the control cells transfected with vector alone (M7609/pLJ) . The sensitivities of the antisense transfectants in relation to ADR, cisplatin, melphalan, and etoposide were increased -3.3-fold, 2.3-fold, 2.2-fold, and 2.1-fold, respectively, compared with those of M7609 and M7609/pLJ . On the other hand, the sensitivities of the antisense transfectants to Taxol, vincristine, 5-fluorouracil, and mitomycin C were not significantly changed . Similarly, the transfection of antisense cDNA into M7609/ADR cells resulted in the reduction of intracellular GST-pi concentration (by about half) and an increased sensitivity to ADR (4.4-fold), but no increase in 5-fluorouracil sensitivity . Thus, GST-pi is considered to be a multidrug resistance factor that is responsible for both the intrinsic and acquired resistance of cancer cells to anticancer drugs such as ADR, cisplatin, melphalan, and etoposide. Cancer Res, 1996 Aug 1, 56(15), 3461 - 7 Proton nuclear magnetic resonance spectroscopy reveals cellular lipids involved in resistance to adriamycin and taxol by the K562 leukemia cell line; Le Moyec L et al.; Proton nuclear magnetic resonance spectroscopy was performed on whole cells to study lipids and metabolites in Adriamycin- and Taxol-resistant K562 cells expressing multidrug resistance (MDR) and their sensitive counterparts . With one-dimensional spectra, both resistant cell lines showed lower fatty acid methylene:methyl ratios and higher choline:methyl ratios than sensitive cells . Using two-dimensional COSY spectra, a decrease in the glutamine content was evidenced in resistant cells . When these cells were maintained in culture medium without the drug, the fatty acid signals were partially recovered . Adriamycin-resistant K562 cells were also treated for 4 days with a high dose of verapamil, a MDR-reversing agent . The nuclear magnetic resonance spectra of verapamil-treated cells also showed partial recovery of fatty acid signals . These results could be paralleled with the reversion of the resistant phenotype, as evidenced by measuring the inhibiting concentration of Adriamycin and vinblastine in K562adr cells cultured without the drug or after short-term exposure to verapamil . Conversely, P-glycoprotein and mRNA expression and DNA amplification of the mdr gene were not modified when compared to resistant cells, suggesting that the MDR phenotype could be partially reversed independently of the mdr gene amplification and expression . These results demonstrate the role of lipids in the resistance phenomenon. Eur J Nucl Med, 1996 Aug, 23(8), 980 - 6 Primary breast cancer imaging with technetium-99m sestamibi and its relation with P-glycoprotein overexpression; Moretti JL et al.; The aim of this preliminary study was to evaluate retrospectively sestamibi scintigraphy in relation to the presence of the 170-kDa P-glycoprotein (Pgp), which represents an expression of multidrug resistance in patients with primary breast cancer . Fifteen women (age range 37-76 years) were referred for technetium-99m sestamibi scintigraphy because of suspicious breast lesions detected by mammography and ultrasonography, and subsequently assessed by fine-needle aspiration . Scintigraphy was performed 30 min following the injection of 500 MBq 99mTc-sestamibi . Three planar anterior and oblique images were obtained with the patient in the supine position . Excised tumours were assessed for cytosolic CA 15.3, oestrogen (OR) and progesterone (PR) receptors and c-erb B2 neu oncogene . Pathology revealed that only 13 of the 15 patients had malignant tumours . The two benign tumours were sestamibi-negative and Pgp-positive . Sestamibi scintigraphy was positive in 10 of the 13 malignant lesions (including nine of ten infiltrating ductal carcinomas) . Two of the three lesions with false-negative scintigraphy were Pgp-negative; in one of these cases histology revealed an invasive lobular carcinoma and in the other, mucinous adenocarcinoma . The third false-negative lesion was a Pgp-positive infiltrating ductal carcinoma which was c-erb B2 neu-negative but CA 15.3-, OR- and PR-positive . This preliminary study confirms that the resistance to chemotherapy which may occur in patients with primary breast cancer can be a cause of negative sestamibi scintigraphy. Curr Genet, 1996 Aug, 30(3), 212 - 7 The unusual inheritance of multidrug-resistance factors in Saccharomyces; Shallom JM et al.; The yeast PDR5 locus encodes a 160-kDa member of the ABC family of transport proteins . Strains bearing a deletion of this locus are drug hypersensitive . Resistant revertants arise when cells are plated on cycloheximide medium . About one-third of these are cross resistant to other agents, including oligomycin, fluconazole and sulfometuron methyl . Most of the revertants exhibit linkage to the PDR5 locus and map in three locations . Curiously, the multi-drug resistance is not due to a single mutation . Most of the revertants behave as though they contained several tightly linked resistance factors. Leukemia, 1996 Aug, 10(8), 1274 - 82 Multidrug resistance-1 (MDR1) expression and functional dye/drug efflux is highly correlated with the t(8;21) chromosomal translocation in pediatric acute myeloid leukemia; Pearson L et al.; Resistance to chemotherapy is a major problem in acute myeloid leukemia (AML) . An important resistance mechanism in adult AML is active drug efflux mediated by the multidrug resistance protein-1 (MDR1) . To determine if MDR1 is important in childhood AML, we examined MDR1 expression and functional dye/drug efflux in 20 pediatric/adolescent AML patients; results were correlated with cytogenetics and clinical outcome . Using flow cytometry, MDR1 protein expression on the leukemic blasts was measured with the antibody MRK16, while efflux was measured by extrusion of the fluorescent dye DiO(C2)3 in the presence/absence of cyclosporin A (CsA) . Six of 20 cases expressed MDR1 . While all six MDR1+ cases were efflux+, three of 14 MDR1- cases also demonstrated efflux . Both MDR1 and efflux were strongly correlated with the t(8;21) . All six MDR1 +/efflux+ cases and 2/3 MDR1 -/efflux+ cases had a t(8;21), while no MDR1-/efflux- cases had a t(8;21) (P < 0.0005) . This correlation between MDR1, efflux, and the t(8;21) in pediatric AML was not found in 11 adult t(8;21) cases similarly studied . Although the clinical relevance of MDR1 in pediatric AML awaits larger studies, our results suggest a biologic subset of pediatric AML patients may benefit from regimens which include MDR1-reversing agents or non-MDR1 substrates. J Urol, 1996 Aug, 156(2 Pt 1), 506 - 11 Expression patterns of multidrug-resistance (MDR1), multidrug resistance-associated protein (MRP),glutathione-S-transferase-pi (GST-pi) and DNA topoisomerase II (Topo II) genes in renal cell carcinomas and normal kidney; Kim WJ et al.; PURPOSE: Expression levels of the multidrug-resistance (mdr1), multidrug resistance-associated protein (MRP), glutathione-S-transferase-pi (GST-pi) and DNA topoisomerase II (Topo II) genes in normal kidney and renal cell carcinomas were analyzed to study the complexity of the roles of these genes . MATERIALS AND METHODS: The reverse transcription-polymerase chain reaction (RT-PCR) assay was used with beta 2 microglobulin (beta 2 m) as the internal control . RESULTS: In normal kidneys, the expression levels of the 4 genes in individual normal kidney samples correlated significantly with one another . Comparisons of the expression levels between normal kidneys and renal cell carcinomas showed that only the mean MRP gene expression level was higher in renal cell carcinomas than in normal kidneys (p = 0.018) . The expression patterns of the 4 genes in renal cell carcinomas differed markedly for nonpapillary and papillary tumors . The mean MRP/beta 2 m ratio for the papillary type was significantly lower than that for the nonpapillary alveolar type carcinoma (p = 0.004) . The 4 genes showed moderate positive correlations with one another in alveolar type renal carcinoma similar to the correlations observed in normal kidneys . In contrast, in papillary type, MRP expression was inversely correlated with mdr1 and Topo II expression . CONCLUSION: Differences in cytogenetic changes, origins and natural histories between papillary and nonpapillary carcinoma may be associated with these distinct expression patterns of the resistance-related genes . Further study is required to clarify whether the differences in the expression patterns between these 2 structural types of carcinoma affect their chemosensitivities and clinical outcomes. Biochemistry, 1996 Jul 30, 35(30), 9728 - 36 Topological folding and proteolysis profile of P-glycoprotein in membranes of multidrug-resistant cells: implications for the drug-transport mechanism; Zhang M et al.; P-glycoprotein (Pgp)1 is a polytopic membrane protein and functions as an energy-dependent drug efflux pump . It is responsible for multidrug resistance (MDR) in cancer cell lines . Recently, the topological structure of Pgp has been investigated . However, the results are in dispute . A major question concerning the Pgp topology is the membrane orientation of the loop linking TM4 and TM5 (loop 4) and the loop linking TM8 and TM9 (loop 8) . In this study, we generated polyclonal antibodies specific to these two loops . In combination with a panel of other well-characterized site-specific polyclonal- and monoclonal antibodies of Pgp, we tested the membrane orientation of these two loops of Pgp in multidrug-resistant cells using immunocytochemistry and proteolysis/membrane protection assay . Our results showed that (1) both loops 4 and 8 are located extracellularly whereas other domains, such as the ATP-binding sites, are in the cytoplasm and (2) proteolysis of Pgp is not a random event and the trypsin-sensitive sites are cleaved in orders . Since the Pgp was not genetically manipulated in this study, in contrast to previous studies, we believe that naturally expressed Pgp molecules have an unconventional topology . We speculate that this alternate topology of Pgp may represent a different functional state of the protein . Further detailed analysis of Pgp topology will help to understand the fundamental mechanism of drug transport by Pgp. Biochem Pharmacol, 1996 Jul 26, 52(2), 213 - 7 Study of P-glycoprotein functionality in living resistant K562 cells after photolabeling with a verapamil analogue; Mankhetkorn S et al.; To our knowledge, this is the first study to investigate the modification of P-glycoprotein functionality in living resistant cells after photolabeling . For this purpose, four new photoactive verapamil analogues were synthesized . These compounds have the same efficacy as verapamil to increase pirarubicin (pira) incorporation into living multidrug resistant (MDR) K562 cells and to sensitize them to the cytotoxic effect of this anthracycline derivative, indicating that they act as typical MDR modifiers in MDR cells . These compounds were used to photolabel P-glycoprotein (P-gp) in living resistant cells . Irradiation did not result in photodamage to cells, and P-gp functionality was verified by the ability of living cells to incorporate pira . The irradiation of resistant cells, 10(6)/mL, in the presence of a verapamil analogue at concentrations equal to or higher than 3 microM yielded 70% inhibition of P-gp functionality . Our data provide the first evidence that the binding of a verapamil analogue to P-gp is not sufficient to completely inhibit the efflux of this anthracycline . The cells were, subsequently, cultured for several days . Resistance was progressively recovered with time, with the treated cells being just as resistant as before photolabeling after 6 days. J Biol Chem, 1996 Jul 19, 271(29), 17147 - 51 Thyroid hormone export regulates cellular hormone content and response; Ribeiro RC et al.; Actions of thyroid hormones (THs) are determined by intracellular free hormone concentration . Here we report that enhanced TH extrusion via a saturable, cold-sensitive mechanism lowers intracellular TH and causes TH resistance in hepatoma cells . Since these cells overexpress multidrug resistance P-glycoproteins and TH extrusion and resistance are blunted by verapamil, P-glycoproteins may mediate this resistance . Verapamil-inhibitable TH efflux was also found in primary hepatocytes, cardiocytes, and fibroblasts . These findings demonstrate that TH extrusion can modulate TH availability and action in mammalian cells. Int J Cancer, 1996 Jul 17, 67(2), 256 - 63 Characterization and epitope mapping of several new anti-P-glycoprotein monoclonal antibodies; Shapiro AB et al.; Monoclonal antibodies (MAbs) were raised against partially purified Class I P-glycoprotein from multidrug-resistant Chinese hamster ovary CHRB30 cells . Fifteen stable monoclonal hybridoma cell lines were established, and the secreted antibodies were classified into 8 groups on the basis of banding pattern on immunoblots of P-glycoprotein digested with cyanogen bromide or partially digested with proteases . One representative of each group was tested further for several activities . Six of the 8 recognized human P-glycoprotein in the multidrug-resistant SKVLBI cell line . None of the antibodies recognized P-glycoprotein in unfixed cells, suggesting that all recognize cytoplasmic epitopes or extracellular epitopes not accessible in native P-glycoprotein . All 8 antibodies were able to immunoprecipitate P-glycoprotein from non-denaturing detergent solution . The linear epitopes of the antibodies were mapped to 11-27 amino acids . Two of the antibodies had epitopes in the linker region, 3 in the N-terminal nucleotide binding domain, 2 in the C-terminal nucleotide binding domain and 1 in the predicted cytoplasmic loop between predicted transmembrane helices 8 and 9 . These antibodies, with known epitopes, could have uses for P-glycoprotein detection, structure/function studies, purification and quantitation. Int J Cancer, 1996 Jul 17, 67(2), 238 - 47 Tumor necrosis factor alpha is a powerful apoptotic inducer in lymphoid leukemic cells expressing the P-170 glycoprotein; Malorni W et al.; Multidrug resistance (MDR) is a phenomenon by which tumor cells exposed to a single anti-proliferative agent acquire resistance to other structurally and functionally unrelated drugs . The classical form of MDR is caused by a plasma-membrane protein currently named P-glycoprotein or P-170 encoded by the human mdr-1 gene in its functional isoform . In vitro cell lines expressing P-170 usually also present phenotypic and functional alterations . In the present study we report that the cytotoxicity mediated by tumor necrosis factor alpha (TNF alpha) in MDR variants of the human T-lymphoblastoid CEM cell line is associated with apoptosis (programmed cell death) . Susceptibility of MDR cells to apoptosis was increased upon cycloheximide + TNF alpha sequential treatment, whereby the impairment of protein synthesis due to the former agent was followed by the effect of cytokine exposure . Massive apoptosis of P-170-positive cells, but not of controls, was also obtained by depletion of nutrients (i.e., serum starvation) . In contrast, TNF-alpha exerted a similar apoptotic effect in epithelial (MCF-7) or myeloma (S8226) drug-sensitive/ -resistant cell pairs . However, the MDR variant of myeloma S8226 was more sensitive to the cytostatic effect of TNF alpha than the parental drug-sensitive cell line . These results suggest that the presence of the MDR phenotype may be associated with increased histotype-dependent cell susceptibility to specific, protein-synthesis-independent, apoptotic pathways. Cancer Res, 1996 Jul 15, 56(14), 3307 - 14 Location of a protease-hypersensitive region in the multidrug resistance protein (MRP) by mapping of the epitope of MRP-specific monoclonal antibody QCRL-1; Hipfner DR et al.; Multidrug resistance protein (MRP) is a Mr 190,000 integral membrane phosphoglycoprotein which has been shown by transfection studies to confer multidrug resistance . We have previously raised and characterized a panel of MRP-specific monoclonal antibodies (MAbs) which detect distinct epitopes in the MRP molecule (D . R . Hipfner et A, Cancer Res., 54 . 5788-5792, 1994), and, in the present study, we have identified the epitope of one of these, MAb QCRL-1 . Immunoblot analysis of MRP fragments generated by digestion with formic acid or trypsin suggested that the MAb QCRL-1 epitope was located in the region connecting the two halves of MRP . Subsequent analyses of a series of truncated bacterial glutathione S-transferase fusion proteins containing segments of human MRP further localized the MAb QCRL-1 epitope to a region encompassing amino acids 903-956 . Similar experiments with an analogous segment of murine MRP demonstrated that MAb QCRL-1 was highly specific for the human protein . The reactivity of MAb QCRL-1 with a series of overlapping hexapeptides and heptapeptides within this region identified the human MRP-specific heptapeptide SSYSGDI (corresponding to amino acids 918-924) as the epitope, and this peptide was shown to specifically inhibit MAb QCRL-1 binding to MRP . The results of these studies confirm that this epitope has a cytoplasmic location consistent with the topology of MRP predicted from hydrophobicity analyses . These experiments also revealed the presence of a number of protease-sensitive sites on either side of the MAb QCRL-1 epitope in the cytoplasmic domain connecting the two halves of MRP . Future epitope-mapping studies with other MRP-specific MAbs win provide additional insights into the topology of MRP, and may help to identify functionally important regions of this protein . Moreover, definition of the epitope recognized by MAb QCRL-1 as well as the other MAbs will facilitate the use of these reagents for immunohistological studies of MRP expression in drug-resistant tumors. Nucleic Acids Res, 1996 Jul 15, 24(14), 2829 - 34 De novo generation of simple sequence during gene amplification; Kirschner LS; Mammalian cells that have undergone gene amplification and/or gene rearrangement have been used as resources to gain insight into the questions of chromosome structure and dynamics . The multidrug resistant murine cell line J7.V2-1 has been shown previously to contain two distinct forms of the highly amplified mdr2 gene, a member of the mouse gene family responsible for the multidrug resistant (MDR) phenotype {Kirschner, L . S . (1995) DNA Cell Biol . 14, 47-59} . Characterization of both forms of the gene revealed that one form corresponded to the wild-type structure of the gene, whereas the other represented a rearrangement . Investigation of this altered gene demonstrated a deletion of 1.6 kb of the wild-type sequence, and replacement of this region with a poly(AT) tract that appears to have been generated de novo . Analysis of the native sequence in this region demonstrated the absence of repetitive elements, but was notable for the presence of two long stretches of polypurine: polypyrimidine strand asymmetry . Analysis of mdr2 transcripts in this cell line revealed that nearly all of the mRNA is transcribed from the rearranged form of the gene . This message is unable to code for a functional mdr2 gene product, owing to a deletion of the fourth exon during this event . Mechanisms of the rearrangement, as well as the significance of this curious effect on transcription, are discussed. Biochem J, 1996 Jul 15, 317 ( Pt 2), 515 - 22 Effects of steroids and verapamil on P-glycoprotein ATPase activity: progesterone, desoxycorticosterone, corticosterone and verapamil are mutually non-exclusive modulators; Orlowski S et al.; P-glycoprotein (P-gp) is a membranous ATPase responsible for the multidrug resistance (MDR) phenotype . Using membrane vesicles prepared from the highly resistant cell line DC-3F/ADX we studied the influence of P-gp ATPase activity of four progesterone derivatives which specifically bind to P-gp and reverse MDR . Progesterone and desoxycorticosterone stimulate P-gp ATPase activity with, respectively, apparent concentrations giving half-maximal activation of 20-25 microM and 40-50 microM, and activation factors of 2.3 (at 100 microM progesterone) and 1.8 (at 170 microM desoxycorticosterone) . Hydrocortisone above 100 microM stimulates P-gp ATPase activity while corticosterone has no apparent stimulating effect . Our data are consistent with the location of the binding sites for the progesterone derivatives on the P-gp membranous domain . The effects of these steroids on verapamil-stimulated P-gp ATPase activity support a non-competitive mechanism, i.e . the binding sites for verapamil and steroids are mutually non-exclusive for P-gp ATPase modulation . A similar non-competitive inhibition of progesterone-stimulated P-gp ATPase activity by desoxycorticosterone or by corticosterone leads to the conclusion that these steroids, although sharing related structures, have distinct modulating sites on P-gp . As expected from their mutually non-exclusive interactions on P-gp, progesterone and verapamil when mixed induce a synergistic modulation of P-gp ATPase activity . Since drug transport by P-gp is believed to be coupled to its ATPase activity, a corresponding synergistic effect of these two modulators for the inhibition of P-gp-mediated drug resistance can be expected. Blood, 1996 Jul 15, 88(2), 633 - 44 The novel anthracycline annamycin is not affected by P-glycoprotein-related multidrug resistance: comparison with idarubicin and doxorubicin in HL-60 leukemia cell lines; Consoli U et al.; A major factor in limiting the efficacy of anthracyclines is overexpression of the MDR1-encoded p-glycoprotein (p-gp) . A new analogue less affected by p-gp is annamycin (ANN), an anthracycline antibiotic with high affinity for lipid membranes and significantly more activity than doxorubicin (DOX) . We investigated whether ANN was affected by p-gp-mediated multidrug resistance (MDR) by comparing the cellular accumulation and retention of ANN, idarubicin (IDR), and DOX in the p-gp-negative human leukemia cell lines (HL-60S) and its DOX-selected p-gp-positive subline (HL-60/DOX) with and without verapamil (VER) . As expected, HL-60/DOX cells showed lower DOX uptake than HL-60S cells; coincubation with VER (10 mmol/L) increased uptake 2.6-fold restoring it to 100% of uptake in HL-60S cells . IDR uptake increased 1.5-fold in the presence of VER, but ANN was not affected . Coincubation with VER increased DOX retention in HL-60/DOX cells 2.8-fold and IDR retention 1.4-fold; unchanged ANN retention indicated that ANN may overcome p-gp . In the cytotoxicity assay to correlate intracellular anthracycline content with antitumor activity, we found ANN to be less potent than DOX and IDR In sensitive cells, ID 50 being the drug concentration that inhibits cell growth by 50% but its resistance index (RI; ID50 resistant cells divided by ID50 sensitive cells) was lower than that of IDR and DOX (2.6 v 40 and 117.5) . Coincubation in the presence of VER resulted in 4.5-fold and 2-fold RI decreases of DOX and IDR, respectively, whereas ANN did not change, further confirming ANN's ability to circumvent p-gp-mediated MDR . Confocal microscopy studies of IDR, ANN, and DOX showed higher intracellular drug compartmentalization for DOX in HL-60/DOX cells incubated in the presence of VER . This study provided evidence that, unlike DOX and IDR, ANN is not affected by p-gp-mediated MDR. Biochem Pharmacol, 1996 Jul 12, 52(1), 127 - 31 Expression of the low-density lipoprotein receptor, HMG-CoA reductase, and multidrug resistance (Mdr1) genes in colorectal carcinomas; Vitols S et al.; Some malignant cells have elevated uptake of plasma low-density lipoprotein (LDL) . We determined the expressions in colorectal cancers of the LDL receptor gene, of the gene for the rate-limiting enzyme in cholesterol synthesis, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, and of the multidrug resistance gene (mdr1) by quantitative RNA-RNA solution hybridisation . LDL receptor RNA levels in tumor tissue exceeded those in normal mucosa in 20 of 23 patients (2-11-fold higher in 17 of 23 patients), with a mean +/- SD of 7.8 +/- 5.8 copies/cell in tumor tissue vs 3.5 +/- 2.5 in normal mucosa (P = 0.002) . The HMG-CoA reductase gene was similarly expressed in tumor and normal tissue, with means and SD of 2.0 +/- 1.3 copies/cell versus 2.2 +/- 1.9 (pi = 21) . Mdr1 RNA was undetectable ( < 0.15 copies/cell) in 5 of 20 tumors, with a mean +/- SD of 1.0 +/- 1.1 copies/cell vs 1.6 +/- 1.7 in normal mucosa . Expression of all three genes was, in most cases, higher in normal liver than in liver metastasis of colorectal carcinomas or normal colon mucosa . The results may form the basis for using LDL as a drug carrier for treatment of colorectal carcinomas, and may indicate that drug resistance in these tumors is not due to overexpression of the mdr1 gene. Hum Gene Ther, 1996 Jul 10, 7(11), 1309 - 22 Expression of the human multidrug resistance and glucocerebrosidase cDNAs from adeno-associated vectors: efficient promoter activity of AAV sequences and in vivo delivery via liposomes; Baudard M et al.; Recombinant adeno-associated viruses (rAAV) are attractive tools for gene therapy . We designed plasmids in which the human multidrug resistance gene (hMDR1) cDNA was placed downstream from portions of the 5' end of AAV including either a 234-bp cassette or the entire AAV p5 promoter . The drug-resistant phenotype conferred by the P-glycoprotein (Pgp) efflux pump encoded by the hMDR1 cDNA was used to select NIH-3T3 cells transfected with these plasmids . The 234-bp region alone showed promoter activity similar in strength to that of the entire p5 promoter or the retroviral Harvey murine sarcoma virus long terminal repeat (LTR); this result demonstrates that the 234-bp cassette might be used as a small and efficient promoter in rAAV designed to express large genes approaching the packaging limit of AAV particles . After transfection of AAV-MDR1 vectors, the integration of MDR1 sequences into the host cell genome was demonstrated by fluorescent in situ hybridization (FISH) . In addition, Southern analysis of low-molecular-weight DNA extracted from drug-resistant cells grown under continuous selection pressure indicated the persistence of nonintegrated AAV-MDR1 plasmids . Coordinate expression of Pgp and human glucocerebrosidase (hGC) was observed in drug-selected NIH-3T3 cells transfected with a bicistronic vector in which MDR1 cDNA was linked to hGC cDNA via the encephalomyocarditis internal ribosome entry site sequence . Moreover, following a single intravenous injection of the bicistronic vector complexed to cationic liposomes into recipient mice, delivery of MDR1 and GC cDNAs was achieved in all the organs we tested . Our results demonstrate that the efficiency of liposomes as vehicles for in vitro and in vivo gene delivery, the advantages of AAV-vectors, and the use of MDR1 as a selectable marker might be successfully combined in gene therapy protocols. Proc Natl Acad Sci U S A, 1996 Jul 9, 93(14), 6929 - 34 ATP-dependent uptake of natural product cytotoxic drugs by membrane vesicles establishes MRP as a broad specificity transporter; Paul S et al.; MRP is a recently isolated ATP-binding cassette family transporter . We previously reported transfection studies that established that MRP confers multidrug resistance {Kruh, G . D., Chan, A., Myers, K., Gaughan, K., Miki, T . & Aaronson, S . A . (1994) Cancer Res . 54, 1649-1652} and that expression of MRP is associated with enhanced cellular efflux of lipophilic cytotoxic agents {Breuninger, L . M., Paul, S., Gaughan, K., Miki, T., Chan, A., Aaronson, S . A . & Kruh, G . D . (1995) Cancer Res . 55, 5342-5347} . To examine the biochemical mechanism by which MRP confers multidrug resistance, drug uptake experiments were performed using inside-out membrane vesicles prepared from NIH 3T3 cells transfected with an MRP expression vector . ATP-dependent transport was observed for several lipophilic cytotoxic agents including daunorubicin, etoposide, and vincristine, as well as for the glutathione conjugate leukotriene C4 (LTC4) . However, only marginally increased uptake was observed for vinblastine and Taxol . Drug uptake was osmotically sensitive and saturable with regard to substrate concentration, with Km values of 6.3 microM, 4.4 microM, 4.2 microM, 35 nM, and 38 microM, for daunorubicin, etoposide, vincristine, LTC4, and ATP, respectively . The broad substrate specificity of MRP was confirmed by the observation that daunorubicin transport was competitively inhibited by reduced and oxidized glutathione, the glutathione conjugates S-(p-azidophenacyl)-glutathione (APA-SG) and S-(2,4-dinitrophenyl)glutathione (DNP-SG), arsenate, and the LTD4 antagonist MK571 . This study establishes that MRP pumps unaltered lipophilic cytotoxic drugs, and suggests that this activity is an important mechanism by which the transporter confers multidrug resistance . The present study also indicates that the substrate specificity of MRP is overlapping but distinct from that of P-glycoprotein, and includes both the neutral or mildly cationic natural product cytotoxic drugs and the anionic products of glutathione conjugation . The widespread expression of MRP in tissues, combined with its ability to transport both lipophilic xenobiotics and the products of phase II detoxification, indicates that the transporter represents a widespread and remarkably versatile cellular defense mechanism. Int J Cancer, 1996 Jul 3, 67(1), 129 - 37 Cellular pharmacology of idarubicinol in multidrug-resistant LoVo cell lines; Toffoli G et al.; Idarubicin (IDA) is a daunorubicin (DAU) analog that is being used to treat a variety of human malignancies . The major circulating metabolite of IDA is idarubicinol (IDOL) . After administration of IDA to patients, the systemic exposure to IDOL is greater than IDA . We investigated the cytotoxic effect of IDOL in the LoVo human colon-carcinoma cell line and its derivative multidrug-resistant (MDR) sub-lines . In LoVo-sensitive cells, the extracellular IDOL concentration inhibiting cell growth by 50% (IC50) was about 2-fold higher than IDA IC50 but lower than DAU IC50 . After continuous exposure of the LoVo parental cells to 20 nM IDOL, 5 drug-resistant clones were obtained . All these clones exhibited an MDR phenotype, indicating that IDOL is involved in multidrug resistance . The resistance index (RI) to IDOL was investigated in LoVo MDR sub-lines obtained by IDOL (LoVo-IDOL-1), IDA (LoVo-IDA-1) and DOX (LoVo-DOX-1) selection . In spite of the drug used for their selection, all the MDR sub-lines exhibited an RI to IDOL lower than DAU and only 2-fold higher than IDA . In LoVo-IDOL-1 cells the RI was 5, 11 and 32 for IDA, IDOL and DAU respectively . Differences in the RI were explained by the greater intracellular tolerance exhibited by MDR cells to DAU than to IDOL and IDA . In the LoVo-IDOL-1 sub-line, the intracellular drug concentration inhibiting cell growth by 50% (IC50int) was higher than in the sensitive cells by 11.4-, 4.7- and 2.8-fold for DAU, IDOL, and IDA respectively . Differences in the intracellular tolerance were explained by the different intracellular distribution of DAU compared with IDOL and IDA . While DAU had a higher nuclear location in LoVo-sensitive cells than in resistant cells, IDOL and IDA maintained the same distribution both in sensitive and in resistant cells . In conclusion, contrary to what has been observed for other derivative metabolites of anthracyclines, the metabolism of IDA to IDOL must not be considered an inactivation pathway . IDOL is a potent inhibitor of cell growth and retains good activity in MDR cells. Biokhimiia, 1996 Jul, 61(7), 1320 - 32 {Mitochondrial transport of nucleic acids . Participation of the benzodiazepine receptor}; Zorov DB; The models of mitochondrial transmembrane nucleic acid transfer are discussed . According to this hypothesis, mitochondria can exchange their nucleic acids by two possible mechanisms including either intermitochondrial fusion and fission or transmembrane transport . In the latter case, important roles for mitochondrial benzodiazepine receptor and factors that regulate activities of its components (mitochondrial porin and adenine nucleotide translocator) are suggested . Nucleic acids can be transported through a Ca(2+)-dependent pore that can be one of the functional states of mitochondrial exchange of genetic material . Problems like cellular aging, apoptosis, proliferation, mitochondrial diseases, multidrug resistance, intracellular traffic, and mitochondrial heredity are discussed considering an important role of mitochondrial benzodiazepine receptor in these processes. Pediatr Pathol Lab Med, 1996 Jul-Aug, 16(4), 551 - 61 Ependymomas in children express the multidrug resistance gene: immunohistochemical and molecular biologic study; Chou PM et al.; In view of the poor response of ependymomas to chemotherapy, it may be hypothesized that these tumors have intrinsic drug resistance to some chemotherapeutic agents . The expression of drug resistance may be specific to a single agent or may involve multiple drugs . Among several mechanisms of drug resistance, P-glycoprotein (Pgp) has been the subject of considerable attention in clinical practice . In order to assess the possible participation of Pgp in the chemotherapeutic resistance of ependymomas, 42 biopsy specimens from 35 patients with ependymoma seen at our institutions were studied by immunohistochemistry with two monoclonal antibodies: C219 (Signet) and UIC-2 (Dr . Roninson's gift) . In addition, four cases were studied by polymerase chain reaction after reverse transcription to detect transcripts of Pgp . Our results showed that in 35 samples there was a positive reaction for Pgp with both antibodies; two biopsy samples were positive only with C219 and three others with UIC-2; the remaining two samples were negative with both antibodies . Of the four cases studied by RT-PCR, three showed MDR1 transcripts . These results support our hypothesis of Pgp-mediated intrinsic multidrug resistance in these tumors. Ann Ital Med Int, 1996 Jul-Sep, 11(3), 211 - 5 {Therapy of tuberculosis today: to prescribe is not enough . A case report}; Rogasi PG et al.; Tuberculosis is a re-emerging disease not only in developing but also in industrialized countries . The multidrug resistance of Mycobacterium tuberculosis is an increasing problem, mainly due to the poor compliance of the patients on drug regimens . The authors discuss the case of a 28-year-old male affected by pulmonary tuberculosis who did not improve after a 3-month period of oral tuberculosis treatment . The switch to intravenous therapy was rapidly followed by clinical and microbiological improvement . The patient subsequently admitted that he had not complied with oral treatment because after healing he would be sent back to prison . This case demonstrates that, even in the hospital setting, noncompliance to antitubercular therapy is not unfrequent, as demonstrated by other case reports . The authors discuss a great amount of literature that strongly supports the introduction of directly observed therapy for the treatment of all tuberculosis patients. Pharmacol Res, 1996 Jul-Aug, 34(1-2), 47 - 57 Prediction of blood cyclosporine concentrations in haematological patients with multidrug resistance by one-, two- and three-compartment models using Bayesian and non-linear least squares methods; Wu G et al.; The blood cyclosporine (CsA) concentration-time profile in each of 24 adult haematological patients with multidrug resistance taking the first course of CsA treatment was fitted by one-, two- and three-compartment models to obtain relevant pharmacokinetic parameters . The pharmacokinetic parameters obtained were implemented into the PKS program (Abbottbase Pharmacokinetic System) as the population pharmacokinetic parameters used to predict blood CsA concentrations in adult haematological patients with multidrug resistance . The predictions of blood CsA concentrations by one-, two- and three-compartment models using the Bayesian method (BM) and the non-linear least squares method (NLLSM) were evaluated employing 11 patients who took the second course of CsA treatment . While the Akaike's information criterion (AIC) favoured the two-compartment model to describe CsA concentration-time profiles in patients taking the first and second courses of CsA treatment, the predictive performance analyses showed that both two- and three-compartment models were better than the one-compartment model for prediction, but the three-compartment model was slightly superior to the two-compartment model . The results also show that the predictions using BM were slightly better than those using NLLSM . Several factors affecting BM predictions and the possible difference among AIC, BM and predictive performance analyses were also addressed. Trans R Soc Trop Med Hyg, 1996 Jul-Aug, 90(4), 418 - 21 Proguanil polymorphism does not affect the antimalarial activity of proguanil combined with atovaquone in vitro; Edstein MD et al.; Clinical studies have shown proguanil (PROG) combined with atovaquone (ATQ) to be an effective and safe antimalarial combination for the treatment of multidrug-resistant falciparum malaria . PROG is a prodrug, which undergoes hepatic metabolism to its pharmacologically active metabolite cycloguanil (CYC) . Individuals exhibit genetic polymorphism with respect to PROG, and can be phenotyped as either extensive metabolizers (EMs) or poor metabolizers (PMs) by measuring their PROG/CYC concentration ratio in plasma following PROG/ATQ administration . PMs produce lower plasma concentrations of CYC than EMs and thus may be more susceptible to prophylaxis or treatment failure . Both PROG and CYC potentiate the activity of ATQ in vitro . The antimalarial activity ex vivo of Thai patients' plasma samples obtained from EMs and PMs given concurrent PROG and ATQ was studied using the K1 isolate of Plasmodium falciparum . This isolate is resistant to PROG and CYC, but sensitive to ATQ . Maximum inhibitory dilution profiles of the patients' plasma samples containing PROG and ATQ from EMs and PMs were similar . These findings indicate that differences in plasma drug concentrations between EMs and PMs did not alter the antimalarial activity in vitro against the K1 isolate . The phenotypic status of individuals is not an important issue in the treatment of patients with PROG/ATQ. Trans R Soc Trop Med Hyg, 1996 Jul-Aug, 90(4), 415 - 7 Artemether or artesunate followed by mefloquine as a possible treatment for multidrug resistant falciparum malaria; Bunnag D et al.; Plasmodium falciparum in south-east Asia is highly resistant to chloroquine and sulfadoxine-pyrimethamine . Mefloquine used to be the chemosuppressant drug of choice in areas with chloroquine resistance . However, sensitivity to this drug has recently decreased in Thailand, Cambodia and Myanmar, and there is no suitable single alternative drug . We therefore investigated possible alternative combination therapies for multidrug resistant falciparum malaria . 120 male Thai patients at Makarm Malaria Clinic, Chantaburi, in eastern Thailand were allocated at random to receive either oral artemether (group A) or artesunate (group B) at a single dose of 300 mg on day 1, both followed by mefloquine, 750 and 500 mg at 24 and 30 h, respectively . Follow-up was on days 1, 2, 7, 14, 21, 28, 35 and 42 . Patients in both groups had a rapid initial response to treatment; in most cases parasitaemia was cleared within 24 h, and fever was cleared within 24 h in 62% and 76.7% of the patients in groups A and B, respectively . 58 patients in group A and 57 in group B completed follow-up and cure rates were 98% and 97%, respectively . Reinfection could not be excluded for the 3 patients with recrudescences; all were cured with a repeated course of treatment . No serious adverse effect was observed in either group, only mild and transient nausea, vomiting and loss of appetite, with no significant difference between the 2 groups . These results suggest that a single oral dose of 300 mg of either artemether or artesunate followed by 1250 mg of mefloquine in 2 divided doses is effective against multiple drug resistant falciparum malaria . Either regimen can be considered as a suitable 'stand-by' in endemic areas of multiple drug resistant falciparum malaria. Pathol Res Pract, 1996 Jul, 192(7), 768 - 80 Molecular mechanisms of multidrug resistance in cancer chemotherapy; Nooter K et al.; The occurrence of multidrug resistance (MDR) is one of the main obstacles in the successful chemotherapeutic treatment of cancer . MDR cell lines are resistant to the so-called naturally occurring anti-cancer drugs, such as anthracyclines, Vinca alkaloids and epipodophyllotoxins, but are not cross-resistant to alkylating agents, antimetabolites and cisplatin . So far, three separate forms of MDR have been characterized in more detail: classical MDR, non-Pgp MDR and atypical MDR . Although all three MDR phenotypes have much in common with respect to cross-resistance patterns, the underlying mechanisms certainly differ . Atypical MDR is associated with quantitative and qualitative alterations in topoisomerase II alpha, a nuclear enzyme that actively participates in the lethal action of cytotoxic drugs . Atypical MDR cells do not overexpress P-glycoprotein, and are unaltered in their ability to accumulate drugs . In this review we will focus on classical and non-Pgp MDR . The molecular mechanism of classical and non-Pgp MDR is transcriptional activation of membrane-bound transport proteins . These transport proteins belong to the ATP-binding cassette (ABC) superfamily of transport systems . The classical MDR phenotype is characterized by a reduced ability to accumulate drugs, due to activity of an energy-dependent uni-directional, membrane-bound, drug-efflux pump with broad substrate specificity . The classical MDR drug pump is composed of a transmembrane glycoprotein (P-glyco-protein-Pgp) with a molecular weight of 170 kD, and is, in man, encoded by the so-called multidrug resistance (MDR1) gene . Typically, non-Pgp MDR has no P-gly-coprotein expression, yet has about the same cross-resistance pattern as classical MDR . This non-Pgp MDR phenotype is caused by overexpression of the multidrug resistance-associated protein (MRP) gene, which encodes a 190 kD membrane-bound glycoprotein (MRP) . MRP probably works by direct extrusion of cytotoxic drugs from the cell and/or by mediating sequestration of the drugs into intracellular compartments, both leading to a reduction in effective intracellular drug concentrations . For the classical MDR phenotype, evidence is accumulating that it plays a role indeed, in clinical drug resistance, especially in some hematological malignancies (acute myeloid leukemia, multiple myeloma and non-Hodgkin's lymphoma) and solid tumors (soft tissue sarcomas and neuroblastoma) . The association of MRP with clinical drug resistance has not been elaborated, yet, and studies on MRP expression in human cancer have just begun . We found that overexpression of MRP, as determined by RNase protection assay as well as by immunohistochemistry, occurs in several human cancers, among which are cancer of the lung, esophagus, breast and ovary, and leukemias . Further studies are indicated to establish whether elevated MRP expression at diagnosis is an unfavorable prognostic factor for clinical outcome of chemotherapy. Haematologica, 1996 Jul-Aug, 81(4), 295 - 301 Overcoming PGP-related multidrug resistance . The cyclosporine derivative SDZ PSC 833 can abolish the resistance to methoxy-morpholynil-doxorubicin; Michieli M et al.; BACKGROUND: The results obtained so far in studies designed to neutralize P glycoprotein (PGP)-related multidrug resistance (MDR) by using MDR reversal agents, have not yet fulfilled the promise of the experiments which were performed in vitro . In order to improve PGP-related MDR neutralization, we tested in vitro the activity of the cyclosporine derivative SDZ PSC 833 (PSC) together with doxorubicin (DOX) and with two new DOX derivatives named 4' iodo 4' deoxy-doxorubicin (IODODOX) and methoxy-morpholynil-doxorubicin (MMDOX, FCE 23762) using four different human cell lines and their multi-drug resistant variants . METHODS: Anthracycline toxicity was evaluated by using the MTT method after a 7-day culture with continuous exposure to the antitumor drugs with or without the addition of PSC . RESULTS: PSC significantly downmodulated the toxicity of all three anthracyclines in all the four cell systems . However, despite the great increase caused by PSC in the toxicity of DOX and a more modest effect on the toxicity of the two DOX derivatives, this MDR reversal agent could only completely block the PGP mediated MMDOX resistance whereas DOX refractoriness was only decreased . CONCLUSIONS: The combination of MMDOX or IODODOX with PSC 1.6 microM is more efficient than the combination of DOX plus PSC for the full reversion of PGP-mediated drug resistance . Careful clinical studies are required to evaluate if these associations can also effectively and safely neutralize MDR in vivo. Anticancer Drugs, 1996 Jul, 7(5), 568 - 78 Ranking of P-glycoprotein substrates and inhibitors by a calcein-AM fluorometry screening assay; Tiberghien F et al.; In order to compare the capacities of a variety of compounds to interfere with P-glycoprotein (Pgp) function, a novel assay was set up to work on a large screening scale . The model assay measures the capacity of parental sensitive (Par) and multidrug-resistant (MDR) cells to efflux a small fixed amount of acetoxymethyl calcein (calcein-AM) after their pretreatment with concentration ranges of known Pgp modulators . This microplate cytometry-based assay was performed with two different pairs of cell lines, the human lymphocytic leukemia CEM cells and the murine monocytic leukemia P388 cells . For a given Pgp-expressing MDR cell line, a Pgp modulator EC50 was defined as the concentration required to restore half of the calcein retention shown by similarly treated Par cells . With both MDR-P388 and MDR-CEM cells, EC50 comparisons ranked five reference Pgp modulators as follows: SDZ 280-446 > SDZ PSC 833 > cyclosporin A > verapamil > vinblastine . Further use of the MDR-CEM cells could rank 15 Pgp modulators for their capacity to interfere with calcein-AM efflux as follows: SDZ 280-446 1.9 x > SDZ PSC 833 8.3 x > cyclosporin A 3.8 x > amiodarone 1.1 x > quinacrine 1.6 x > verapamil 1.4 x > quinidine 1.1 x > vinblastine 11 x > vincristine 2 x > chloroquine > beta-lumicolchicine > or = gamma-lumicolchicine > or = colchicine > etoposide > or = doxorubicin . This calcein-AM assay should open the way for ranking large numbers of novel structures for their potential Pgp modulator properties, particularly for an efficient screening of Pgp function antagonists, but it does not allow defining whether their inhibition may be competitive or not. Biochem Mol Biol Int, 1996 Jul, 39(4), 687 - 96 Chlorpromazine transport in membrane vesicles from multidrug resistant CCRF-CEM cells; Syed SK et al.; The mechanism by which chlorpromazine (2-chloro-10-(3-dimethylaminopropyl)-phenothiazine) reverses P-glycoprotein (P-gp2) mediated multidrug resistance was investigated using membrane vesicles prepared from human CCRF-CEM leukaemia cells . Chlorpromazine was transported in an ATP-dependent manner into membrane vesicles prepared from vinblastine resistant (VBL1000) cells but not from drug-sensitive cells . The chlorpromazine uptake was sensitive to osmotic pressure, indicating true transport into the vesicle lumen . The ATP-dependent chlorpromazine uptake was inhibited about 30% by the addition of ammonium chloride, indicating that a pH or electrical gradient could not account for the majority of ATP-dependent chlorpromazine uptake . The results of this study show that chlorpromazine is actively transported my P-glycoprotein and that chemosensitization by phenothiazines may occur by competition of these agents for active transport of anticancer agents by P-glycoprotein. Pharm Res, 1996 Jul, 13(7), 1073 - 7 Effect of cyclosporin analogues and FK506 on transcellular transport of daunorubicin and vinblastine via P-glycoprotein; Tanaka K et al.; PURPOSE: P-glycoprotein-mediated transcellular transport of anticancer agents and the inhibitory effect of cyclosporin analogs and FK506 were investigated . METHODS: The transcellular transport of daunorubicin and vinblastine by monolayers of LLC-GA5-COL150 cells which overexpressed P-glycoprotein was measured in the presence and absence of cyclosporins or FK506 . RESULTS: Cyclosporins and FK506 inhibited P-glycoprotein-mediated transport of daunorubicin and vinblastine in the order of cyclosporin D, dihydrocyclosporin D > cyclosporin A > FK506 > cyclosporin C, dihydrocyclosporin C . The intracellular accumulation of the anticancer agents was highly associated with the transporting function of P-glycoprotein . The inhibitory effect of cyclosporin D was concentration-dependent . The inhibitory effect of the modulators on P-glycoprotein was not correlated with the immunosuppressive activity, but was correlated with their lipophilicity . CONCLUSIONS: In the transcellular transport system, lipophilicity may be one of the determinants for the inhibitory effect of various multidrug resistance modulators on the P-glycoprotein-mediated transport. Pharm Res, 1996 Jul, 13(7), 963 - 77 Carrier-mediated intestinal transport of drugs; Tsuji A et al.; Recent advances in the field of carrier-mediated intestinal absorption of of amino acids, oligopeptides, monosaccharides, monocarboxylic acids, phosphate, bile acids and several water-soluble vitamins across brush-border and basolateral membranes are summarized . An understanding of the molecular and functional characteristics of the intestinal membrane transporters will be helpful in the utilization of these transporters for the enhanced oral delivery of poorly absorbed drugs . Some successful examples of the synthesis of prodrugs recognized by the targeted transporters are described . Functional expression of the multidrug resistance gene product, P-glycoprotein, as a primary active transporter in the intestinal brush-border membrane leads to net secretion of some drugs such as anticancer agents in the blood-to-luminal direction, serving as a secretory detoxifying mechanism and as a part of the absorption barrier in the intestine. Biol Pharm Bull, 1996 Jul, 19(7), 971 - 6 Transport mechanisms of anthracycline derivatives in human leukemia cell lines: uptake of pirarubicin, daunorubicin and doxorubicin by K562 and multidrug-resistant K562/ADM cells; Nagasawa K et al.; We studied the uptake mechanisms of anthracycline derivatives, pirarubicin (THP), daunorubicin (DNR) and doxorubicin (ADR), in K562 and multidrug-resistant K562/ADM cells, which overexpress a multidrug efflux pump P-glycoprotein (P-gp) . The uptake of THP, DNR and ADR by K562 or K562/ADM cells was time-, temperature- and concentration-dependent . The THP and ADR uptake by the parental cells was not affected by treatment with 4 mM 2,4-dinitrophenol (DNP) alone or DNP plus a P-gp specific inhibitor, cyclosporin A (CyA, 10 microM), while the DNR uptake in the DNP treatment group was significantly greater than that in the control group . There was no difference in the uptake of THP between DNP-pretreated K562 cells and DNP plus CyA-pretreated K562/ADM cells . The uptake of DNR or ADR was almost equal in both types of cell treated with DNP alone . Every kinetic constant for THP, DNR and ADR uptake by the sensitive cells was approximately equal to that in the resistant cells, respectively, under the above conditions . THP uptake was noncompetitively inhibited and stimulated on simultaneous treatment and preloading, respectively, of DNR or ADR in each type of cell . ADR showed noncompetitive inhibition of DNR uptake by either type of cell . Therefore, it was suggested that a common carrier-mediated transport system was involved in the uptake of THP, DNR and ADR, and that their binding sites in the carrier might be different from one another in both K562 and K562/ADM cells. J Photochem Photobiol B, 1996 Jul, 34(2-3), 177 - 82 Intracellular pH does not affect drug extrusion by P-glycoprotein; Goda K et al.; The intracellular pH (pH(i)) of cells exhibiting multidrug resistance (MDR) related to the expression of the P-glycoprotein (Pgp) is often more alkaline than that of the parental cells, as also observed for the KB-V1/KB-3-1 system in this paper . The possible role of an elevated pH(i) in Pgp-related MDR has been investigated by shifting back the pH(i) of the MDR+ cells to a more acidic value using the mobile proton ionophore carbonylcyanide m-chlorophenylhydrazone (CCCP) . The influence of CCCP-evoked delta pH(i) on relative daunorubicin (DNR) accumulation was similar in the case of several Pgp positive and negative cell lines, in view of flow cytometric and radioactive drug accumulation studies and measuring DNR levels in the medium in a flow-through system . Our data argue against a significant effect of pH(i) on Pgp pumping efficiency . However, an indirect connection between pH(i) regulation and the MDR phenotype is suggested by the fact that acidification of the external medium in the presence of verpamil could be observed exclusively in MDR+ cells. Antimicrob Agents Chemother, 1996 Jul, 40(7), 1695 - 8 Identification for mar mutants among quinolone-resistant clinical isolates of Escherichia coli; Maneewannakul K et al.; Quinolone-resistant clinical Escherichia coli isolates were examined for mutations in the marRAB operon of the multiple antibiotic resistance (mar) locus . Among 23 strains evaluated, 8 were chosen for further study: 3 that showed relatively high levels of uninduced, i.e., constitutive, expression of the operon and 5 with variable responses to induction by salicylate or tetracyclines . The marR genes, specifying the repressor of the operon, cloned from the three strains constitutively expressing the operon did not reduce the level of expression of beta-galactosidase from a marO::lacZ transcriptional fusion and were therefore mutant; however, marR genes cloned from the five other clinical strains repressed LacZ expression and were wild type . All three mutant marR genes contained more than one mutation: a deletion and a point mutation . Inactivation of the mar locus in the three known marR mutant strains with a kanamycin resistance cassette introduced by homologous recombination reduced resistance to quinolones and multiple antibiotics . These findings indicate that mar operon mutations exist in quinolone-resistant clinical E . coli isolates and contribute to quinolone and multidrug resistance. Clin Sci (Lond), 1996 Jul, 91(1), 93 - 8 Modulation of multidrug resistance gene (mdr-1) with antisense oligodeoxynucleotides; Liu C et al.; 1 . Multidrug resistance is the major obstacle to successful cancer chemotherapy . Circumventing multidrug resistance therefore represents a high priority for clinical anti-cancer treatment . Among many reversal strategies, antisense oligodeoxynucleotides may offer a molecular targeting tool for overcoming cellular multidrug resistance . 2 . Two 17-mer phosphorothioate antisense oligomers, complementary to the 5' end of the ATG initiator codon-containing region and loop-forming site (located at nucleotides 991-1007 from the first ATG codon) in mdr-1 cDNA sequence, were synthesized . The purpose was to study their effects on the function and expression of P-glycoprotein and mdr-1 gene . 3 . The results showed that 10 mumol/l antisense oligomers could significantly inhibit the growth of multidrug resistant K562/Adm cells cultured in adriamycin-containing medium . No such effect was observed for parental (sensitive) K562/S cells . Intracellular daunorubicin accumulation increased greatly in the K562/Adm cells after they were treated with oligomers for 48 h and P-glycoprotein synthesis was strikingly reduced . 4 . Further investigation with {alpha-32P}dCTP incorporation by the reverse transcriptase-polymerase chain reaction method revealed that antisense oligomers could result in a reduction in the level of mdr-1 mRNA, probably through hindering mdr-1 gene transcription . 5 . The high reversal efficiency and specificity of antisense oligomers in regulating mdr-1 gene expression suggest a potential clinical application in gene therapy for drug resistant malignancies. Microbiology, 1996 Jul, 142 ( Pt 7), 1855 - 62 Analysis of putative ABC transporter genes in Mycoplasma hyopneumoniae; Blanchard B et al.; A previously described DNA probe specific for Mycoplasma hyopneumoniae (I-141) was fully sequenced and found to consist of 1618 bp and to contain two tandemly repeated ORFs . The deduced amino acid sequence of the two ORFs showed significant homologies with ATP-binding cassette (ABC) transporter proteins, particularly those of the eukaryotic multidrug resistance (MDR) protein family (up to 21% identity and 47% similarity) . A somewhat lower homology was evident with the secretion protein HlyB of the RTX-haemolysin from Escherichia coli . The location of the two ORFs on the M . hyopneumoniae chromosome was downstream of the rrl gene encoding the 23S rRNA, but transcribed in the opposite direction . PCR amplification and subsequent chromosomal analysis by Southern blot hybridization of several M . hyopneumoniae strains showed that all field strains contained the two putative ABC transporter genes . However, some culture collection strains derived from strain J had lost these genes as the result of a 2221 bp deletion. Br J Haematol, 1996 Jul, 94(1), 23 - 33 Expression of the multidrug resistance-associated protein (MRP) mRNA and protein in normal peripheral blood and bone marrow haemopoietic cells; Legrand O et al.; We studied the expression of multidrug resistance-associated protein (MRP) in normal haemopoietic cells from peripheral blood and bone marrow . The MRP mRNA levels were estimated by RT/PCR and in situ hybridization (ISH) assay, and the protein levels by flow cytometry . 21 samples of peripheral blood and 21 samples of bone marrow (11 normal bone marrow donors, 10 patients in complete remission after chemotherapy for large cell lymphoma or acute myeloid leukaemia) were analysed . In peripheral blood the mean MRP mRNA level in CD3+ cells was statistically higher than in the other cells (3-fold by the methods used) . The levels of MRP in CD3+ varied from one individual to another (4.5-34.8 units by RT/PCR and 5-23 grains/cell by ISH); however, this was proportional to the variation in all the cell lineages of same individual (r = 0.84) . In bone marrow the mean MRP levels of the various cell lineages (including CD34+) were similar to the basal level in HL60 cells . Individual expression levels were again variable; however, there was no difference between untreated normal bone marrow and post chemotherapy normal bone marrow . MRP protein expression was determined by flow cytometry with the monoclonal antibody MRPm6 . The CD4+ lymphocytes exhibited a higher MRP protein expression than the other cell lineages, including CD8+ cells . There was a good correlation between the three methods used (RT/PCR and ISH, P = 0.0001, r = 0.87; RT/PCR and flow cytometry, P = 0.0001, r = 0.85; ISH and flow cytometry, P = 0.002, r = 0.67). Nippon Sanka Fujinka Gakkai Zasshi, 1996 Jul, 48(7), 522 - 8 {An immunohistological study on expression of glutathione S-transferase pi (form) in dysplastic and neoplastic human uterine cervix lesions}; Satoh T et al.; The glutathione S-transferase (GST) pi has been studied in association with the mechanisms of multidrug resistance and as a marker for malignant tumors . In this study, specimens from 92 cases of cervical neoplasms and 10 cases of normal squamous epithelium adhering to myoma were stained immunohistochemically with a rabbit polyclonal antibody to GST-pi . In 6 cases of normal squamous epithelium, the intermediate layer was positively stained with the GST-pi antibody . In all 20 cases of dysplasia, the cells with koilocytotic atypia were stained positively . In all 10 cases of carcinoma in situ and all 16 cases of stage Ia squamous cell carcinoma, various intensities of GST-pi staining were demonstrated . Forty-six specimens of stage Ib or more squamous cell carcinoma were positive for GST-pi binding except only one case . In general, squamous cell carcinoma of the uterine cervix is resistant to chemotherapeutic agents . GST-pi is most frequently stained in cervical squamous cell carcinoma as compared with ovarian or endometrial carcinoma . In conclusion, these results suggest that GST-pi may be a marker for cervical squamous cell carcinoma. Anticancer Res, 1996 Jul-Aug, 16(4A), 2079 - 82 Multidrug resistance-associated protein (MRP) gene expression in human lung cancer; Narasaki F et al.; Multidrug resistance-associated protein (MRP) is a 190 kD transmembrane protein and a potentially important drug-transporter protein in human cancers . While the MRP gene is expressed in normal cells and tissues, the expression in solid tumors is not sufficiently determined . MRP and mdr1 mRNA expressions were examined in normal lung parenchyma and in tumor tissues from six small cell lung cancer (SCLC) patients who had received preoperative chemotherapy and eleven nonsmall cell lung cancer (NSCLC) patients . The reverse transcriptase polymerase chain reaction was used . Normal lung tissues and all SCLCs expressed abundant levels of MRP mRNA, while the NSCLCs expressed a wide range of levels from low to high . Most tumor tissues coexpressed both MRP and mdr1, but the levels of mdrl expression was low except in two SCLCs and one NSCLC . MRP is more likely than mdr1 to be one of the clinical multidrug resistance mechanisms found in lung cancer. Anticancer Res, 1996 Jul-Aug, 16(4A), 1719 - 25 Influence of culture media and multidrug resistance on the wheat germ agglutinin (WGA) glycocytochemical expression of two human glioblastoma cell lines; Camby I et al.; Over the last two decades, many studies carried out with the aid of lectins have firmly established that cell glycans usually change in the course of the normal processes of growth and development, as well as in pathological situations . We describe here the in vivo binding expression of wheat germ agglutinin (WGA) to the U87 and U373 human glioblastoma cell lines exposed to various culture media i.e., media supplemented with either 10% (FCS10) or 1% (FCS1) fetal calf serum with or without 10 n Mol/l 17 beta-oestradiol (E2) . After exposure to chemotherapeutic agents, the resistant variants (CR) developed by the two cell lines were also investigated . The quantitative cytochemical assessment of WGA binding was assessed by means of a cell image processor, which was also used to determine ploidy level (on Feulgen-stained nuclei) by means of DNA histogram typing (DHT) . Our results clearly demonstrate that when U373 cells are cultured with E2, this steroid can modify the expression of WGA binding, whereas U87 cells were unaffected . Similarly, lowering the FCS level enhanced the WGA binding of the U373 cell line . Multidrug-resistant cell variants were associated with both aneuploidy and a dramatic decrease in cytochemical WGA expression. Anticancer Res, 1996 Jul-Aug, 16(4A), 1675 - 81 CG5/Dx human breast cancer cell line: characterization of a new doxorubicin-resistant variant; Gibelli N et al.; By continuous exposure of CG5 human breast cancer cell line to increasing doxorubicin (Dx) concentrations, a multidrug-resistant (MDR) subline (CG5/Dx) was obtained . The resistant variant showed P-glycoprotein (P-gp) expression and a lower intracellular doxorubicin level than the parental cells . CG5/Dx cells were 19.4 fold more resistant to Dx than CG5 cells and showed a cross-resistance to some structurally related and unrelated compounds . Differences in kinetics, biological and ultrastructural features between the two cell lines were investigated . The CG5/Dx cells grew more slowly, produced higher CEA levels and showed a reduced progesterone receptor (PgR) content than the parental cells . Ultrastructural studies revealed differences involving, polyribosomes, rough endoplasmic reticulum, {mitochondria} and cytoskeleton. Hepatology, 1996 Jul, 24(1), 248 - 52 Experimental Mallory body formation is accompanied by modulation of the expression of multidrug-resistance genes and their products; Preisegger KH et al.; Mallory bodies (MBs) are characteristic morphological features of alcoholic hepatitis and are also found in other chronic liver disorders and hepatocellular neoplasms . MBs can be produced in mouse liver by chronic administration of the porphyrinogenic drugs griseofulvin (GF) and 3,5-diethoxy-carbonyl-1,4-dihydrocollidine (DDC) . The mechanisms causing the formation of MBs are poorly understood, and the significance of MB formation during the course of liver disease remains unclear . We investigated the relationship between the mechanisms underlying the formation of MBs and the regulation of multidrug resistance (mdr) genes and their products, the P-glycoproteins (Pgp) . Immunofluorescence microscopy using the monoclonal antibody C219 revealed an increase of Pgp expression in almost all hepatocytes after 3 to 8 days of feeding mice DDC- and GF-containing diets . However, after approximately 4 weeks of DDC and approximately 8 weeks of GF feeding, when the first small MBs appeared and loosening and diminution of keratin intermediate filament (KIF) cytoskeleton occurred in some hepatocytes, a decrease or loss of Pgp staining in affected hepatocytes was observed . After feeding mice DDC for 6 weeks and GF for 12 weeks, many hepatocytes contained MBs and displayed a disruption of the immunohistochemically demonstrable KIF meshwork . Double immunofluorescence microscopy with the keratin polyclonal antibody and the mab C219 at this time point revealed a complete loss of Pgp staining in affected cells, although remaining hepatocytes with unaltered KIF meshwork showed a strong reaction with the C219 antibody . Northern blot analyses revealed a significant increase of mdr2 mRNA and, to a lesser extent, of mdr1a mRNA in the livers of DDC- and GF-fed animals. Blood, 1996 Jul 1, 88(1), 309 - 18 Correlation of P-glycoprotein expression and function in childhood acute leukemia: a children's cancer group study; Ivy SP et al.; Clinical drug resistance may be attributed to the simultaneous selection and expression of genes modulating the uptake and metabolism of chemotherapeutic agents . P-glycoprotein (P-gp) functions as a membrane-associated drug efflux pump whose increased expression results in resistance to anthracyclines, epipodophyllotoxins, vinca alkaloids, and some alkylating agents . This type of resistance occurs as both de novo and acquired resistance to therapy for leukemia . We have studied P-gp expression and function in childhood acute leukemias by developing a series of doxorubicin- and vincristine-selected CEM, T-cell lymphoblastoid cell lines that recapitulate the low levels of expression and resistance seen clinically . These cell lines have been used to develop flow cytometric assays for the semiquantitative measurements of P-gp expression with the MRK16 monoclonal antibody and P-gp function using the enhanced retention of rhodamine 123 in the presence of verapamil, a resistance modulator . Kolmogorov-Smirnov statistics, represented by the D measurement, are used to determine the difference in level of P-gp expression by comparing MRK16 staining to an IgG2a isotype control . When D is > 0.09, there is an excellent correlation (R = 0.82) between P-gp expression and function . The evaluation of 107 bone marrow specimens from 84 children with lymphoblastic or myelogenous leukemia showed a statistically significant (P = .004) increase in P-gp function at relapse . P-gp expression at relapse, however, approached but did not reach a significant level (P = .097) . Using this methodology, we can identify patients with levels of P-gp expression and function that we can define clinically, as well as children with discordant multidrug resistance phenotypes . This study supports the role of P-gp-mediated drug resistance in childhood leukemia and confirms that P-gp expression and function are measurable in their leukemic blasts . These assays provide the means for the in vitro testing of resistance modulators and the monitoring of in vivo response to treatment with these agents. Jpn J Cancer Res, 1996 Jul, 87(7), 765 - 72 Overexpression of multidrug resistance protein gene in human cancer cell lines selected for drug resistance to epipodophyllotoxins; Koike K et al.; Overexpression of either the multidrug resistance 1 (MDR1) gene or multidrug resistance protein (MRP) gene is involved in acquisition of multidrug-resistant phenotypes in human cancer cells . In this study we examined whether selection for resistance to the epipodophyllotoxins, etoposide/teniposide (VP16/VM26), could induce overexpression of MDR1 or MRP . We have previously isolated two VP16/VM26-resistant KB cell lines . Two VP16/VM26-resistant KB cell lines, KB/VM-1 and KB/ VM-4, which were selected by stepwise exposure to VM26 had decreased accumulation of {3H}VP16 and increased levels of MRP, but no apparent expression of MDR1 gene was observed . Another VP16/VM26-resistant KB cell line, KB/VP-4, which was further isolated from a VP16-resistant KB cell line, KB/VP-2, had decreased accumulation of {3H}VP16 and showed overexpression of MRP gene, but not that of MDR1 gene . We also isolated a VP16-resistant cell line, IN157/VP-1, from a human glioma cell line IN157 . IN157/VP-1 cells showed decreased accumulation of {3H}VP16 and overexpression of MRP gene, but not of MDR1 . These findings suggest that selection for resistance to VP16/VM26, preferentially induces overexpression of MRP gene. Jpn J Cancer Res, 1996 Jul, 87(7), 757 - 64 Transduction of the macrophage colony-stimulating factor gene into human multidrug resistant cancer cells: enhanced therapeutic efficacy of monoclonal anti-P-glycoprotein antibody in nude mice; Sone S et al.; To develop a therapeutic modality for overcoming multidrug-resistant (MDR) cancer with anti-MDR1 antibody, we examined the effect of macrophage colony-stimulating factor (M-CSF) gene transfection into MDR AD10 cells on therapy of MDR cancer with anti-MDR1 antibody (MRK17) in nude mice . MDR human ovarian cancer (AD10) cells were transduced with the human M-CSF gene inserted into an expression vector to establish gene-modified cells capable of producing low (ML-AD10), intermediate (MM-AD10) nd high (MH-AD10) amounts of M-CSF . Systemic administration of MRK17 resulted in significant dose-dependent inhibition of subcutaneous growth of ML-AD10 tumors . In contrast, systemic administration of recombinant M-CSF in combination with MRK17 did not augment the therapeutic efficacy of MRK17 alone, but rather promoted the growth of the parent AD10 cells . To test the efficacy of in vivo M-CSF gene therapy combined with antibody, we mixed the parent AD10 cells with MH-AD10 cells producing a large amount of M-CSF, and inoculated the mixed cells subcutaneously . Treatment with MRK17 inhibited growth of the mixed cells more than that of the parent cells alone . Thus, combined therapy with anti-MDR1 mAb and M-CSF gene modification of MDR cancer cells may provide a new immunotherapeutic modality for overcoming MDR in humans. Br J Cancer, 1996 Jul, 74(2), 187 - 93 Daunorubicin and doxorubicin but not BCNU have deleterious effects on organotypic multicellular spheroids of gliomas; Kaaijk P et al.; In the present study organotypic multicellular spheroids (OMS) were used to study the effects of chemotherapeutic agents on malignant gliomas . Compared with the frequently used cell line models, OMS have several advantages with respect to the preservation of the cellular heterogeneity and the structure of the original tumour . OMS prepared from seven glioma specimens were treated with 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), daunorubicin or doxorubicin . After exposure to these drugs, the histology and cell proliferation of the OMS were analysed by immunohistochemistry and image analysis . Furthermore, the expression of P-glycoprotein (P-gp) and multidrug resistance-related protein (MRP), which both can contribute to resistance to daunorubicin and doxorubicin, were immunohistochemically investigated . We found that OMS from gliomas are sensitive for daunorubicin and doxorubicin but not for BCNU in terms of tissue destruction and decrease in cell proliferation . In addition, all gliomas were P-gp and MRP negative, which is in accordance with the sensitivity for daunorubicin and doxorubicin . Considering the potential use of several new alternative drug delivery methods, such as intratumoural implantation of drug-impregnated polymers or liposomal encapsulation of cytostatic drugs, daunorubicin and doxorubicin might be effective in the treatment of malignant gliomas. Chest, 1996 Jul, 110(1), 279 - 81 A death associated with therapy for nosocomially acquired multidrug-resistant tuberculosis; Weltman AC et al.; Treatment of multidrug-resistant tuberculosis is difficult and has been associated rarely with severe side effects . We report the nosocomial transmission of multidrug-resistant tuberculosis to a health-care worker who was seronegative for HIV infection . She died because of liver failure associated with treatment for active multidrug-resistant tuberculosis. Br J Cancer, 1996 Jul, 74(1), 43 - 8 An improved method of encapsulation of doxorubicin in liposomes: pharmacological, toxicological and therapeutic evaluation; Gokhale PC et al.; We describe here an improved method of encapsulating doxorubicin in liposomes using phosphatidylcholine, cholesterol and synthetic tetramyristoyl cardiolipin . With this new composition of lipids the entrapment of doxorubicin was found to be > 90% . Cytotoxicity studies using vincristine-resistant HL-60/VCR leukaemia cells showed that liposome-encapsulated doxorubicin reverses multidrug resistance 5-fold compared with conventional doxorubicin and at levels equivalent to that obtained using liposomes with natural cardiolipin . In normal mice, liposome-encapsulated doxorubicin was much less toxic than the conventional drug . A dose of 25 mg kg-1 i.v . of conventional doxorubicin produced 100% mortality in mice by day 14, whereas liposomal doxorubicin exhibited only 10% mortality by day 60 . Liposomal doxorubicin demonstrated enhanced anti-tumour activity against murine ascitic L1210 leukaemia compared with conventional doxorubicin . At a dose of 15 mg kg-1, liposomal doxorubicin increased the median life span with 12 of 18 long-term (60 days) survivors compared with only 3 of 18 with conventional drug . Mice injected i.v . with liposomal doxorubicin had plasma levels 44-fold higher than conventional doxorubicin, producing significantly higher (P < 0.02) area under the plasma concentration curve . An altered tissue distribution was also observed with liposomal doxorubicin; cardiac tissue demonstrating at least 2-fold lower levels with liposomal doxorubicin probably accounting for its lower toxicity . This altered pharmacokinetics of liposome-encapsulated doxorubicin, providing enhanced therapeutic advantage and the ability to modulate multidrug resistance, could be useful in a clinical setting. J Histochem Cytochem, 1996 Jul, 44(7), 679 - 85 Cellular localization of P-glycoprotein in brain versus gonadal capillaries; Stewart PA et al.; P-glycoprotein, the multidrug resistance protein that actively transports a wide variety of lipophilic substrates out of cancer cells, has recently been described in some normal tissues, including the endothelium of the brain and testes . Here we show that P-glycoprotein is also expressed in ovarian endothelium . In ovarian capillaries, the immunolabeled protein was detected with two monoclonal antibodies to P-glycoprotein . It was shown to be membrane-bound and to transport a known P-glycoprotein substrate . Expression of P-glycoprotein in endothelial cells suggests that this transport protein plays a role in enhancing or restricting vascular permeability to lipophilic molecules . If it does, then its role may be predicted from its site of expression on the luminal or abluminal face of the capillary wall . In the region of the endothelial nucleus, endothelial membranes are sufficiently far apart that they can be distinguished at the light microscopic level . Confocal examination of tissue sections double labeled for P-glycoprotein and nuclei confirmed that, in brain, P-glycoprotein is expressed only on luminal membranes . This location is consistent with its putative role in protecting the neuropil from circulating lipophilic molecules . In both testicular and ovarian endothelium, however, P-glycoprotein is expressed on both luminal and abluminal membranes . This localization suggests that it acts to exclude P-glycoprotein substrates from the endothelial cells themselves. Cancer Res, 1996 Jul 1, 56(13), 3010 - 20 Methods to detect P-glycoprotein-associated multidrug resistance in patients' tumors: consensus recommendations; Beck WT et al.; Multidrug resistance (MDR), especially that associated with overexpression of MDR1 and its product, P-glycoprotein (Pgp), is thought to play a role in the outcome of therapy for some human tumors; however, a consensus conclusion has been difficult to reach, owing to the variable results published by different laboratories . Many factors appear to influence the detection of Pgp in clinical specimens, including its low and heterogeneous expression; conflicting definitions of detection end points; differences in methods of sample preparation, fixation, and analysis; use of immunological reagents with variable Pgp specificity and avidity and with different recognition epitopes; use of secondary reagents and chromogens; and differences in clinical end points . Also, mechanisms other than Pgp overexpression may contribute to clinical MDR . The combined effect of these factors is clearly important, especially among tumors with low expression of Pgp . Thus, a workshop was organized in Memphis, Tennessee, to promote the standardization of approaches to MDR1 and Pgp detection in clinical specimens . The 15 North American and European institutions that agreed to participate conducted three preworkshop trials with well-characterized MDR myeloma and carcinoma cell lines that expressed increasing amounts of Pgp . The intent was to establish standard materials and methods for a fourth trial, assays of Pgp and MDR1 in clinical specimens . The general conclusions emerging from these efforts led to a number of recommendations for future studies: (a) although detection of Pgp and MDR1 is at present likely to be more reliable in leukemias and lymphomas than in solid tumors, accurate measurement of low levels of Pgp expression under most conditions remains an elusive goal; (b) tissue-specific controls, antibody controls, and standardized MDR cell lines are essential for calibrating any detection method and for subsequent analyses of clinical samples; (c) use of two or more vendor-standardized anti-Pgp antibody reagents that recognize different epitopes improves the reliability of immunological detection of Pgp; (d) sample fixation and antigen preservation must be carefully controlled; (e) multiparameter analysis is useful in clinical assays of MDR1/Pgp expression; (f) immunostaining data are best reported as staining intensity and the percentage of positive cells; and (g) arbitrary minimal cutoff points for analysis compromise the reliability of conclusions . The recommendations made by workshop participants should enhance the quality of research on the role of Pgp in clinical MDR development and provide a paradigm for investigations of other drug resistance-associated proteins. Cancer Res, 1996 Jul 1, 56(13), 2904 - 7 H19 gene overexpression in atypical multidrug-resistant cells associated with expression of a 95-kilodalton membrane glycoprotein; Doyle LA et al.; The multidrug resistance phenotype of human breast carcinoma MCF-7/AdrVp cells is characterized by overexpression of a 95-kilodalton membrane glycoprotein (p95), accompanied by a marked reduction in intracellular anthracycline accumulation, without overexpression of P-glycoprotein or the multidrug resistance protein . We discovered that the mRNA of the H19 gene is overexpressed in MCF-7/AdrVp cells relative to parental MCF-7 cells or drug-sensitive MCF-7/AdrVp revertant cells . H19 is an imprinted gene with an important role in fetal differentiation, as well as a postulated function as a tumor suppressor gene . Another p95-overexpressing multidrug-resistant cell line, human lung carcinoma NCI-H1688, also displays high levels of H19 mRNA . In contrast, several multidrug-resistant cell lines that overexpress P-glycoprotein or the multidrug resistance protein do not have higher levels of H19 mRNA than their drug-sensitive counterparts . This is the first report of H19 gene overexpression accompanying any form of drug resistance . The association of H19 and p95 gene expression in drug resistance warrants further study. Leukemia, 1996 Jul, 10 Suppl 3, S39 - S45 MDR1, MRP, topoisomerase IIalpha/beta, and cyclin A gene expression in acute and chronic leukemias; Beck J et al.; Gene expression was analyzed by cDNA-PCR at the mRNA level in bone marrow samples (>80% blasts) from ALL (28 primary, 22 first relapses, 10 recurrent relapses), from AML (14 primary, 23 relapses), In peripheral blood lymphocytes from CLL (five untreated, 10 treated), in one CML in blast crisis in the course of the disease (four samples), and in bone marrow samples from healthy donors (12 specimens) . We found low mean MDR1 expression in primary ALL, first relapses of ALL, and primary AML . Significantly higher mean relative MDR1 expression levels were seen in recurrent relapses of ALL, and in the group of relapsed state AML . MDR1 expression measured intermediate in bone marrow samples from healthy donors . The CLL lymphocytes showed generally relatively high MDR1 expression levels . MRP gene expression measured very similar in primary ALL, first relapses of ALL, primary AML, and normal bone marrow . Significantly increased MRP mRNA levels were observed in the groups of recurrent ALL and relapsed state AML . CLL lymphocytes also showed high MRP expression levels . A combined increase of MDRI (about 20-fold) and MRP (about four-fold) was monitored in samples obtained from the CML in blast crisis after chemotherapy . While no significant differences of the mean topoisomerase IIbeta mRNA levels were found throughout, a significantly decreased topoisomerase IIalpha gene expression was measured in first and recurrent relapses of ALL . In CLL lymphocytes either the expression of the topoisomerase IIalpha gene was not detectable by cDNA-PCR, or it measured very low . Topoisomerase IIalpha gene expression was correlated to cyclin A gene expression in the samples of acute leukemias, Indicating the link of topoisomerase IIalpha expression to the proliferative activity of these leukemic blast cells . Our results point to a potentially multifactorial emergence of multidrug resistance in particular states and types of leukemias. Leukemia, 1996 Jul, 10 Suppl 3, S32 - S38 MDR1 reversal: criteria for clinical trials designed to overcome the multidrug resistance phenotype; Hegewisch-Becker S; Multidrug resistance (MDR) is a widely studied mechanism of cellular resistance to structurally unrelated cytotoxic agents . The MDR phenotype is related to the overexpression of the MDR1 gene product, P-glycoprotein (P-gp), a transmembrane drug efflux pump . The capacity to compete with the cytotoxic drug for the active outward transport process, thus inhibiting the activity of the P-gp pump, has been demonstrated for numerous non-cytotoxic compounds, termed MDR modulators . The possibility of modulating the activity of the P-gp pump has initiated numerous clinical trials, using a wide range of chemosensitizers . This article reviews these substances, discusses problems that may arise in connection with the concurrent administration of P-gp modulators and chemotherapeutic agents and provides guidelines for the design of future clinical trials . Furthermore, now data are presented concerning the potential of idarubicin to overcome the MDR phenotype. Leukemia, 1996 Jul, 10 Suppl 3, S10 - S17 Clinical relevance of drug resistance genes in malignant disease; Filipits M et al.; Drug resistance is a major reason for the failure of anticancer chemotherapy . Multidrug resistance has been recognized as an important type of resistance and can be due to various mechanisms . Here we review the published data on the presence and clinical significance of these mechanisms in solid tumors and hematological malignancies . We also refer to new treatment strategies resulting from the knowledge of the various mechanisms of drug resistance present in malignant diseases. Cancer, 1996 Jul 1, 78(1), 63 - 9 Cisplatin, epirubicin, and vindesine with or without lonidamine in the treatment of inoperable nonsmall cell lung carcinoma: a multicenter randomized clinical trial; Ianniello GP et al.; BACKGROUND: Lonidamine (LND) is an indazol-carboxylic acid derivative that selectively inhibits the energy metabolism of neoplastic cells, and increases the permeability of cell membranes . In vitro studies have demonstrated that LND can potentiate the oncolytic activity of cytotoxic drugs and is able to reverse the acquired multidrug resistance of neoplastic cells . Some clinical trials have suggested a synergism of LND with alkylating agents, cisplatin, and anthracyclines in various solid tumors . METHODS: From June 1990 to June 1993, 158 previously untreated patients with Stage IIIB and IV nonsmall cell lung cancer (NSCLC) were enrolled into a multicentric randomized trial to evaluate the addition of LND to a cisplatin-epirubicin-vindesine regimen . Eighty patients in the control arm (A) received cisplatin, 60 mg/m2 intravenously (i.v.); epirubicin, 60 mg/m2 i.v.; and vindesine, 3 mg/m2 i.v . (PEV), on Day 1 every 4 weeks, whereas 78 patients in the experimental arm (B) received the same regimen with the addition of LND from 75 mg orally three times on Day 1 to 150 mg orally three times on Day 7+ until tumor progression occurred . RESULTS: The experimental treatment achieved a significantly higher proportion of major responses in comparison with the control regimen (43% vs . 24%; P=0.02) . The addition of LND apparently potentiated the activity of this cytotoxic treatment, particularly in patients with metastatic disease (overall response rate, 39% vs . 17%) . The median time to progression (5 vs . 8 months; P=0.0007) and the median survival time (7.6 vs . 11 months; P=0.0013) were also statistically improved in Arm B . The acute toxicity of the 2 treatments was low: only 6% of patients in Arm A and 4% of patients in Arm B had to withdraw from treatment due to Grade 4 World Health Organization toxicity . The main additional side effects related to the administration of LND were epigastralgia, myalgia, asthenia, and orchialgia . However, these symptoms were mild and controlled by the concomitant administration of low doses of steroids . CONCLUSIONS: The mild acute toxicity of the PEV regimen and the acceptable and nonoverlapping additional side effects of LND render our experimental therapy worthy of consideration for the management of NSCLC patients with poor performance status or low tolerance to more aggressive therapeutic approaches. Proc Natl Acad Sci U S A, 1996 Jun 25, 93(13), 6743 - 8 The human multidrug resistance-associated protein functionally complements the yeast cadmium resistance factor 1; Tommasini R et al.; A Saccharomyces cerevisiae strain with a disrupted yeast cadmium resistance factor (YCF1) gene (DTY168) is hypersensitive to cadmium . YCF1 resembles the human multidrug resistance-associated protein MRP (63% amino acid similarity), which confers resistance to various cytotoxic drugs by lowering the intracellular drug concentration . Whereas the mechanism of action of YCF1 is not known, MRP was recently found to transport glutathione S-conjugates across membranes . Here we show that expression of the human MRP cDNA in yeast mutant DTY168 cells restores cadmium resistance to the wild-type level . Transport of S-(2,4-dinitrobenzene)-glutathione into isolated yeast microsomal vesicles is strongly reduced in the DTY168 mutant and this transport is restored to wild-type level in mutant cells expressing MRP cDNA . We find in cell fractionation experiments that YCF1 is mainly localized in the vacuolar membrane in yeast, whereas MRP is associated both with the vacuolar membrane and with other internal membranes in the transformed yeast cells . Our results indicate that yeast YCF1 is a glutathione S-conjugate pump, like MRP, and they raise the possibility that the cadmium resistance in yeast involves cotransport of cadmium with glutathione derivatives. J Biol Chem, 1996 Jun 21, 271(25), 14712 - 6 The multidrug resistance-associated protein (MRP) subfamily (Yrs1/Yor1) of Saccharomyces cerevisiae is important for the tolerance to a broad range of organic anions; Cui Z et al.; We have cloned and characterized a Saccharomyces cerevisiae gene YRS1 that complements the phenotype of the mutant sensitive to the anionic drug reveromycin A . The YRS1 gene, which is identical to the recently identified YOR1 gene, encodes a protein with extensive homology to the human multidrug resistance-associated protein (MRP) and the yeast cadmium factor (Ycf1) . A chromosomal deletion of YRS1 lead to viable Deltayrs1 cells, which exhibited hypersensitivity to reveromycin A . Elevation of the YRS1 gene dosage in wild type cells conferred increased resistance to reveromycin A . By analyzing the effect of YRS1 disruption and overexpression it was demonstrated that Yrs1 is involved in the detoxification of a wide range of the organic anions that contain carboxyl group(s) but none of the other type of toxic compounds examined . Fluorescence-activated cell sorter analysis indicated the increased accumulation of the anionic fluorescent compound rhodamine B in Deltayrs1 cells . The expression of YRS1 was induced strikingly by reveromycin A . These results suggest that Yrs1 is a multispecific organic anion transporter important for tolerance against toxic environmental organic anions . Yrs1 had an overlapping specificity with Ycf1 in the resistance to cadmium. J Biol Chem, 1996 Jun 21, 271(25), 14981 - 8 Coordinated induction of MRP/GS-X pump and gamma-glutamylcysteine syntheta