Potent synergic effect between ibuprofen and azoles on Candida resulting from blockade of efflux pumps as determined by FUN-1 staining and flow cytometry.

OBJECTIVES Resistance to antifungals often relates to efflux pumps exporting drugs; several modulators may block them, reverting resistance. Verapamil, beta-oestradiol and progesterone, known efflux pump inhibitors of human neoplastic cells, and ibuprofen were tested as potential modulators of resistance of Candida spp. METHODS Forty-two clinical isolates of Candida (38 fluconazole-resistant), two ATCC type strains and two C. albicans strains with known mechanisms of fluconazole resistance were incubated with subinhibitory concentrations of the modulators. After exposure, MICs of fluconazole, itraconazole and voriconazole were re-determined. Simultaneously, yeasts exposed to modulators were stained with FUN-1 and analysed by flow cytometry. 3H-labelled itraconazole was also used to study efflux in the presence and absence of modulators. RESULTS Fluconazole MICs decreased in most strains after exposure to modulators, including control strains with documented efflux overexpression. No significant MIC variation was noticed for: all C. krusei strains tested, for the resistant strain by target change, for susceptible strains, and for a very few other clinical isolates. Reverted resistant phenotypes showed cross-resistance to itraconazole and to voriconazole, which was also reverted by the modulators. For these strains, an increase in FUN-1 staining and increased accumulation of 3H-labelled itraconazole were noticed after incubation with modulators. CONCLUSIONS Resistance related to overexpression of efflux pumps was common among clinical isolates and could be reverted by the assayed modulators, particularly ibuprofen. The mechanism of resistance in all tested C. krusei and in a few other strains seems, however, to be of a different nature. Ibuprofen is a promising compound in association with azoles, deserving future clinical trials. FUN-1 proved to be a good marker of efflux in Candida.

[1]  T. C. White,et al.  Clinical, Cellular, and Molecular Factors That Contribute to Antifungal Drug Resistance , 1998, Clinical Microbiology Reviews.

[2]  L. Drago,et al.  Antimycotic Activity and Phagocytosis Effects of Econazole in Combination with Ibuprofen Isobuthanolammonium Against Vaginal Strains , 2000, Journal of chemotherapy.

[3]  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.

[4]  Robert,et al.  Progesterone interacts with P-glycoprotein in multidrug-resistant cells and in the endometrium of gravid uterus. , 1989, The Journal of biological chemistry.

[5]  J. Lopez-Ribot,et al.  Prevalence of Molecular Mechanisms of Resistance to Azole Antifungal Agents in Candida albicans Strains Displaying High-Level Fluconazole Resistance Isolated from Human Immunodeficiency Virus-Infected Patients , 2001, Antimicrobial Agents and Chemotherapy.

[6]  W. T. Beck,et al.  Chloroquine enhancement of anticancer drug cytotoxicity in multiple drug resistant human leukemic cells. , 1986, Biochemical pharmacology.

[7]  J. M. Ford,et al.  Pharmacology of drugs that alter multidrug resistance in cancer. , 1990, Pharmacological reviews.

[8]  S. Sen Gupta,et al.  Mechanisms and clinical impact of antifungal drug resistance. , 1994, Journal of medical and veterinary mycology : bi-monthly publication of the International Society for Human and Animal Mycology.

[9]  S. Costa-de-Oliveira,et al.  Susceptibility to fluconazole of Candida clinical isolates determined by FUN-1 staining with flow cytometry and epifluorescence microscopy. , 2001, Journal of medical microbiology.

[10]  R. Haugland,et al.  Development of the FUN-1 family of fluorescent probes for vacuole labeling and viability testing of yeasts , 1997, Applied and environmental microbiology.

[11]  A. Rodrigues,et al.  Antifungal activity of local anesthetics against Candida species. , 2000, Infectious diseases in obstetrics and gynecology.

[12]  D. Sanglard,et al.  Cloning of Candida albicans genes conferring resistance to azole antifungal agents: characterization of CDR2, a new multidrug ABC transporter gene. , 1997, Microbiology.

[13]  N. Gow,et al.  Correlation between rhodamine 123 accumulation and azole sensitivity in Candida species: possible role for drug efflux in drug resistance , 1996, Antimicrobial agents and chemotherapy.

[14]  T. C. White,et al.  Resistance Mechanisms in Clinical Isolates of Candida albicans , 2002, Antimicrobial Agents and Chemotherapy.

[15]  I. Polacheck,et al.  Activity of phenothiazines against medically important yeasts , 1987, Antimicrobial Agents and Chemotherapy.

[16]  A. Rodrigues,et al.  Germ tube formation changes surface hydrophobicity of Candida cells. , 1999, Infectious diseases in obstetrics and gynecology.

[17]  G. Wishart,et al.  Adequate tumour quinidine levels for multidrug resistance modulation can be achieved in vivo. , 1992, European journal of cancer.

[18]  P. Twentyman.  Cyclosporins as drug resistance modifiers. , 1992, Biochemical pharmacology.

[19]  E. Scott,et al.  Demonstration of synergy with fluconazole and either ibuprofen, sodium salicylate, or propylparaben against Candida albicans in vitro , 1995, Antimicrobial agents and chemotherapy.

[20]  M. Milgroom,et al.  Fungicide Resistance—Lessons for Herbicide Resistance Management? , 1995, Weed Technology.

[21]  A. Rodrigues,et al.  Antifungal activity of ibuprofen alone and in combination with fluconazole against Candida species. , 2000, Journal of medical microbiology.

[22]  K. Marr,et al.  Mechanism of Increased Fluconazole Resistance in Candida glabrata during Prophylaxis , 2004, Antimicrobial Agents and Chemotherapy.

[23]  J. Zalcberg,et al.  Rapid functional assay for multidrug resistance in human tumor cell lines using the fluorescent indicator fluo-3. , 1991, Journal of the National Cancer Institute.