AIDS Related Oral Candidiasis: Drugs, Sterols, and Fungal Cells

描述（由申请人提供）：致病性酵母菌白色念珠菌（Ca）是免疫功能低下人群（包括艾滋病患者）真菌感染的主要原因。通常用抗真菌药物治疗，最常见的是在艾滋病患者中广泛使用的唑类氟康唑。在这些患者中，有很大的可能性会产生唑耐药性。利用从艾滋病患者中分离出的口服耐药菌株，已经确定了主要的耐药机制，包括ERG11（一种编码麦角甾醇生物合成酶和偶氮靶的基因）的改变和外排泵的表达增加。麦角甾醇是真菌质膜中的主要甾醇；它的生物合成是唑类和许多其他抗真菌药物的靶标。真菌细胞与唑类化合物的相互作用是一个复杂的过程。代谢和环境的特定方面在体外影响药物/细胞相互作用，并具有重要的临床潜力。这项研究的总体目标是了解真菌细胞对唑类药物的反应。本研究探讨了UPC2转录因子对甾醇的输入、后续调控以及环境因素对细胞对药物反应的影响。本建议的具体目标是：1。表征氟康唑进入真菌细胞的特性。唑的输入对细胞/药物相互作用很重要；它可以通过被动或主动运输介导；以前也没有研究过。2. 分析UPC2的转录调控。一旦进入细胞，唑类药物抑制Erg 11p，改变固醇水平，并激活UPC2转录因子，UPC2转录因子调节固醇的生物合成和摄取。将分析UPC2基因的调控，以了解固醇代谢（包括摄取和生物合成）的变化如何与UPC2的表达相关。3. 表征特定环境因素对药物敏感性及细胞表面的影响。环境因素，包括pH值，氧水平，氮源和外源性固醇，将分析其对药物敏感性和细胞表面的影响，使用微生物学，生化和基因表达方法。4. 鉴定临床分离株的固醇代谢改变。已知的耐药机制并不适用于许多耐药菌株。目的是分析这些分离株的进口、UPC2调控和对环境因素的反应，以确定可能的新耐药机制。了解唑类药物与真菌细胞之间的相互作用是一个具有临床意义的问题，具有改善真菌感染和耐药性的诊断、治疗和预防的潜力。

DESCRIPTION (provided by applicant): The pathogenic yeast Candida albicans (Ca) is a major cause of fungal infections in immune-compromised populations including AIDS patients. It is usually treated with antifungal drugs, most commonly the azole fluconazole which is used extensively in AIDS patients. In these patients, there is a significant probability that azole resistance will develop. Using oral resistant isolates from AIDS patients, major mechanisms of resistance have been identified including alterations in ERG11 (a gene encoding an enzyme in ergosterol biosynthesis and target of the azoles) and increased expression of efflux pumps. Ergosterol is the major sterol in the fungal plasma membrane; and its biosynthesis is the target for azoles and many other antifungals. The interaction of fungal cells with azoles is a complex process. Specific aspects of metabolism and the environment influence drug/cell interactions in vitro and have the potential to be important clinically. The Overall Goal of this research is to understand how a fungal cell responds to azoles. This proposal investigates import, subsequent regulation of sterols by the UPC2 transcription factor, and the influence of environmental factors on the cellular response to drugs. The Specific Aims of this proposal are: 1. To characterize fluconazole import into the fungal cell. Azole import is important to the cell/drug interaction; it may be mediated by passive or active transport; and it has not been studied previously. 2. To analyze the transcriptional regulation of UPC2. Once within the cell, azoles inhibit Erg 11 p, altering sterol levels, and activating the UPC2 transcription factor, which regulates sterol biosynthesis and uptake. Regulation of the UPC2 gene will be analyzed to understand how changes in sterol metabolism, including uptake and biosynthesis are correlated with expression of UPC2. 3. To characterize the effect of specific environmental factors on drug susceptibility and the cell surface. Environmental factors, including pH, oxygen levels, nitrogen sources, and exogenous sterols, will be analyzed for their effect on the drug susceptibility and the cell surface, using microbiological, biochemical and gene expression approaches. 4. To characterize clinical isolates for alterations in sterol metabolism. The known mechanisms of resistance do not apply to many resistant strains. This aim will assay import, UPC2 regulation, and response to environmental factors to identify possible new resistance mechanisms in these isolates. Understanding the interactions between azoles and fungal cells is a clinically significant issue, with the potential for improving diagnosis, treatment and prevention of fungal infections and resistance.