Glucan Binding to Azole Drugs: A Novel Resistance Mechanism in Candida albicans

描述（由申请方提供）：致病性酵母菌白色念珠菌（Ca）是免疫功能低下人群口腔和全身真菌感染以及女性阴道感染的常见原因。念珠菌感染通常用唑类药物治疗，这些患者人群中经常出现唑类耐药。唑类耐药的主要机制包括外排泵表达增加和麦角固醇生物合成酶的改变。然而，这些耐药机制尚未在许多耐药临床分离株中确定。最近，在念珠菌属生物膜中发现了唑类耐药的第三种机制-在粘膜表面和植入的医疗器械上形成的细胞和细胞外物质的基质。这些生物膜中的高水平的唑类耐药性是由于细胞壁中与唑类药物结合的β-1，3葡聚糖的产生增加。我们的假设是，唑类的β-1，3-葡聚糖结合不仅是生物膜中耐药性的一个组成部分，而且也是抗生素临床耐药分离株中耐药性的一个组成部分。唑类与细胞壁的葡聚糖结合将阻止药物到达细胞质，增加耐药性。总体目标是评价唑类与β-1，3-葡聚糖结合作为念珠菌耐药性组分的作用。具体而言，该提案研究了β-1，3葡聚糖结合如何影响真菌细胞中的药物积累，并评估了临床分离株集合中的β-1，3葡聚糖结合，包括没有已知耐药机制的分离株。本申请的具体目的是：1.确定β-1，3葡聚糖结合对氟康唑在白念珠菌中蓄积的影响。真菌细胞的唑积累可能是几个竞争过程的结果，包括输入和流出（内部积累），也可能是β-1，3葡聚糖结合（外部积累）。将在改变细胞葡聚糖含量的条件下评价细胞中放射性标记的FLC蓄积。2.评估药物敏感性改变的临床分离株中β-1，3葡聚糖的结合。将在尚未确定已知耐药机制的耐药临床分离株中评估唑与β-1，3葡聚糖的结合以及细胞中β-1，3葡聚糖水平。唑类和真菌细胞之间的相互作用在可预见的未来将继续成为临床上重要的问题。(-1唑类药物的，3-葡聚糖结合是这些相互作用的一个新的和重要的方面。这一过程的完整表征，加上我们对药物/细胞相互作用其他方面的理解，有可能有助于改善真菌感染和耐药性的诊断、治疗和预防。致病性酵母菌白色念珠菌在口腔、阴道和血液中引起严重的人类疾病。本研究拟分析抗真菌药物氟康唑与念珠菌之间的相互作用。白色念珠菌特别是，它将测试真菌细胞壁作为海绵的假设，吸收药物，从而允许细胞在药物存在下持续存在。

DESCRIPTION (provided by applicant): The pathogenic yeast Candida albicans (Ca) is a frequent cause of oral and systemic fungal infections in immune-compromised people and of vaginal infections in women. Candida infections are usually treated with azole drugs, and azole resistance arises frequently in these patient populations. The major mechanisms of azole resistance include increased expression of efflux pumps and alterations in enzymes in ergosterol biosynthesis. However, these mechanisms of resistance have not been identified in many resistant clinical isolates. Recently, a third mechanism of azole resistance has been found in Candida biofilms - matrices of cells and extracellular material that forms on mucosal surfaces and implanted medical devices. The high levels of azole resistance in these biofilms is due to increased production of (-1,3 glucan in the cell wall that binds to azole drugs. Our Hypothesis is that (-1,3 glucan binding of azoles is a component of resistance not only in biofilms, but in planktonic clinically resistant isolates. Glucan binding of azoles to the cell wall would prevent the drug from reaching the cytoplasm, increasing resistance. The Overall Goal is to evaluate the contribution of azole binding to (-1,3 glucan as a component of drug resistance in Candida. Specifically, this proposal investigates how (-1,3 glucan binding affects drug accumulation in fungal cells, and it evaluates (-1,3 glucan binding in a collection of clinical isolates, including isolates with no known resistance mechanisms. The Specific Aims of this application are: 1. To determine the effect of (-1,3 glucan binding on fluconazole accumulation in Candida albicans. Azole accumulation by fungal cells is likely to be the result of several competing processes, including import and efflux (internal accumulation), and possibly (-1,3 glucan binding (external accumulation). Radiolabeled FLC accumulation in cells will be evaluated under conditions that alter the glucan content of the cells. 2. To assess (-1,3 glucan binding in clinical isolates with altered drug susceptibilities. Azole binding to (-1,3 glucans, and (-1,3 glucan levels in the cells will be assessed in resistant clinical isolates in which no known mechanism of resistance has been identified. The interactions between azoles and fungal cells will continue to be clinically significant issues for the foreseeable future. (-1,3 glucan binding of azole drugs is a new and important aspect of these interactions. A full characterization of this process, together with our understanding of the other facets of drug/cell interactions, has the potential to contribute to improvements in diagnosis, treatment and prevention of fungal infections and of resistance. PUBLIC HEALTH RELEVANCE The pathogenic yeast Candida albicans causes significant human disease in the mouth, vagina and blood stream. This proposal will analyze the interaction between the antifungal drug fluconazole and C. albicans. In particular, it will test the hypothesis that the fungal cell wall acts as a sponge, soaking up the drug, thus allowing the cells to persist in the presence of drug.