Disulfiram is a potent modulator of multidrug transporter Cdr1p of Candida albicans.

To find novel drugs for effective antifungal therapy in candidiasis, we examined disulfiram, a drug used for the treatment of alcoholism, for its role as a potential modulator of Candida multidrug transporter Cdr1p. We show that disulfiram inhibits the oligomycin-sensitive ATPase activity of Cdr1p and 2.5mM dithiothreitol reverses this inhibition. Disulfiram inhibited the binding of photoaffinity analogs of both ATP ([alpha-(32)P]8-azidoATP; IC(50)=0.76 microM) and drug-substrates ([(3)H]azidopine and [(125)I]iodoarylazidoprazosin; IC(50) approximately 12 microM) to Cdr1p in a concentration-dependent manner, suggesting that it can interact with both ATP and substrate-binding site(s) of Cdr1p. Furthermore, a non-toxic concentration of disulfiram (1 microM) increased the sensitivity of Cdr1p expressing Saccharomyces cerevisiae cells to antifungal agents (fluconazole, miconazole, nystatin, and cycloheximide). Collectively these results demonstrate that disulfiram reverses Cdr1p-mediated drug resistance by interaction with both ATP and substrate-binding sites of the transporter and may be useful for antifungal therapy.

[1]  I. Paulsen,et al.  Major Facilitator Superfamily , 1998, Microbiology and Molecular Biology Reviews.

[2]  Stephen Naylor,et al.  Role of disulfiram in the in vitro inhibition of rat liver mitochondrial aldehyde dehydrogenase. , 2000, Biochemical pharmacology.

[3]  D. Keppler,et al.  Export pumps for anionic conjugates encoded by MRP genes. , 1999, Advances in enzyme regulation.

[4]  D. Clarke,et al.  Blockage of drug resistance in vitro by disulfiram, a drug used to treat alcoholism. , 2000, Journal of the National Cancer Institute.

[5]  M. Raymond,et al.  Functional Similarities and Differences between Candida albicans Cdr1p and Cdr2p Transporters , 2003, Antimicrobial Agents and Chemotherapy.

[6]  Z. Sauna,et al.  The molecular basis of the action of disulfiram as a modulator of the multidrug resistance-linked ATP binding cassette transporters MDR1 (ABCB1) and MRP1 (ABCC1). , 2004, Molecular pharmacology.

[7]  R. Béliveau,et al.  Molecular Interactions of Cyclosporin A with P-glycoprotein , 1997, The Journal of Biological Chemistry.

[8]  D. Sanglard,et al.  Susceptibilities of Candida albicans multidrug transporter mutants to various antifungal agents and other metabolic inhibitors , 1996, Antimicrobial agents and chemotherapy.

[9]  Z. Sauna,et al.  Evidence for a requirement for ATP hydrolysis at two distinct steps during a single turnover of the catalytic cycle of human P-glycoprotein. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[10]  F. Loor,et al.  Reversion of the P-glycoprotein-mediated multidrug resistance of cancer cells by FK-506 derivatives. , 1993, Anti-cancer drugs.

[11]  Smriti,et al.  Functional Characterization of Candida albicans ABC Transporter Cdr1p , 2003, Eukaryotic Cell.

[12]  C. Higgins,et al.  ABC transporters: from microorganisms to man. , 1992, Annual review of cell biology.

[13]  A. Goffeau,et al.  Functional Expression of Candida albicans Drug Efflux Pump Cdr1p in a Saccharomyces cerevisiae Strain Deficient in Membrane Transporters , 2001, Antimicrobial Agents and Chemotherapy.

[14]  L. Greenberger,et al.  Domain mapping of the photoaffinity drug-binding sites in P-glycoprotein encoded by mouse mdr1b. , 1991, The Journal of biological chemistry.

[15]  K. Kuchler,et al.  Mechanisms of resistance to azole antifungal agents in Candida albicans isolates from AIDS patients involve specific multidrug transporters , 1995, Antimicrobial agents and chemotherapy.

[16]  R. Cannon,et al.  Multiple efflux mechanisms are involved in Candida albicans fluconazole resistance , 1996, Antimicrobial agents and chemotherapy.

[17]  A. Goffeau,et al.  Molecular cloning and characterization of a novel gene of Candida albicans, CDR1, conferring multiple resistance to drugs and antifungals , 1995, Current Genetics.

[18]  W. Schaffner,et al.  A rapid, sensitive, and specific method for the determination of protein in dilute solution. , 1973, Analytical biochemistry.

[19]  J. Chick.  Safety Issues Concerning the Use of Disulfiram in Treating Alcohol Dependence , 1999, Drug safety.

[20]  K. Sugimoto,et al.  P-glycoprotein expression on normal and abnormally expanded natural killer cells and inhibition of P-glycoprotein function by cyclosporin A and its analogue, PSC833. , 1999, Blood.

[21]  I. Pastan,et al.  Characterization of the azidopine and vinblastine binding site of P-glycoprotein. , 1992, The Journal of biological chemistry.