Candida albicans is a predominant cause of opportunistic fungal infections in HIV-infected individuals. These infections can be present as topical infections which are readily treated with existing antifungal drugs or as disseminated infections which are extremely difficult to treat and result in mortality rates exceeding 50%. The treatment of disseminated disease often requires the use of the highly toxic drug amphotericin B. Other less toxic drugs ar available, such a s fluconazole, but their efficacy is unproven. There is an urgent need to develop highly specific antifungal directed at new molecular target. In this application, we propose to develop the proton pumping plasma membrane H+-ATPase of Candida Albicans as a target for a new class of antifungal agents. The H+-ATPase is an attractive target because it is an essential enzyme that represents 20-40% of the total plasma membrane protein and it has extracellularly expose regions that when modified attenuate its catalytic activity. In addition, the H+-ATPase is believed to play an important role in the pathogenicity of Candida through its effects on intracellular pH regulation, nutrient uptake, and medium acidification. Most importantly, an opportunity exists to jump start the drug discovery program and quickly develop lead compounds because of the discovery that omeprazole (Losec), a clinically-important anti-ulcer drug, which inhibits the gastric H+,K+-ATPase, blocks, Candida cell growth and is a potent inhibitor of the fungal H+-ATPase. ASTRA Hassle AB, the developer of omeprazole, has agreed to provide us with substituent analogy of omeprazole and other anti-ulcer drugs, like SCH28080, for specific anti-H+-ATPase reagent that can progress to clinical trials. The drug discovery program has been designed to permit efficient screening of compounds for anti-H+-ATPase activity. All initial screens will be established with a Saccharomyces strain expressing a hybrid Candida-Saccharomyces PMAI gene to allow the biochemistry and genetics of the Saccharomyces system to be exploited. Whenever possible, correlative experiments will be performed with Candida albicans. The drug discovery program will evaluate the mechanistic action of lead compounds thought biochemical and genetic analyses. Finally, the role of the H+-ATPase in dimorphism, which has been linked to pathogenicity, will be evaluated by the application of highly specific lead compounds as they arise, as well as by genetic manipulation of PMAI in Candida albicans and in a pseudohypha Saccharomyces model.
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