GENETIC IDENTIFICATION OF ANTIFUNGAL DRUG TARGETS

这项提议有三个具体的短期目标：1.克隆C. 新生杆菌基因可能在发病机制中起重要作用； 构建这些基因的“敲除”突变；测试 在实验感染中导致新生隐球菌变异体。的目标是 这些实验是为了识别基因--无毒/无毒力的突变体 实验感染中的致病性，从而识别良好 潜在的毒品目标。 主要的关注点是氨基酸和维生素的特定子集。 生物合成基因。关注这些途径的原因是 许多营养缺乏菌是无毒的。这些途径中的基因选择是 由(I)它们在人类中的缺失和(Ii)高度有害的 特定突变体的体外表型。第二个重点是基因 只有在高温下才能生长的产品。这个 关注这些基因产品的原因是，生长的能力 高温对真菌致病毒力是必不可少的。 这项建议与健康有关，因为(I)它涉及新生芽孢杆菌， 最严重的致病真菌之一，以及(Ii)它集中在 关于确定抗真菌药物靶标的问题。目前的 现有的抗真菌药物很少(与抗菌药相比)。 数量，没有临床医生希望的那么有效，而且往往有 严重的副作用。因此，非常需要开发新的 抗真菌药。这项提议的目标是从基因上识别新的 抗真菌药物靶标。一旦确定了具体的新目标 人们将有可能将注意力集中在这些基因产品和用途上 新技术，如组合化学，以开发小 分子抑制剂。在杜克大学，这样的合作是可能的(参见 附信)。 该提案有三个广泛的长期/未来目标：1.检视 更多基因/突变体了解全球新生芽孢杆菌 发病机制；2.将这种方法应用于其他病原真菌；以及 3.)开发基因产品的小分子抑制剂 对致病是必不可少的。

This proposal has three specific, short term aims: 1.) cloning C. neoformans genes likely to be important for pathogenesis; 2.) constructing "knock-out" mutations in these genes; nd 3.) testing the resulting C. neoformans mutants in experimental infections. The goal of these experiments is to identify genes-mutants that are avirulent/non- pathogenic in experimental infections and thereby identify good potential drug targets. The primary focus is on a specific subset of amino acid and vitamin biosynthetic genes. The reason for focusing on these pathways is that many auxotrophs are avirulent. The choice of genes in these pathways is dictated by (i) their absence in humans and (ii) the highly deleterious in vitro phenotypes of specific mutants. The secondary focus is on gene products that are required for growth only at high temperature. The reason for focusing on such gene products is that the ability to growth at high temperatures is essential for fungal virulence-pathogenesis. This proposal is health related because (i) it deals with C. neoformans, one of the most serious of the pathogenic fungi, and (ii) it is focused on the identification of antifungal drug targets. The currently available antifungal drugs are few (compared to antibacterials) in number, are less effective than clinicians would like, and tend to have severe side effects. Therefore, there is a great need to develop new antifungals. The goal of this proposal is to genetically identify new antifungal drug targets. Once specific new targets have been identified it will be possible to focus attention on these gene products and use novel technologies, such as combinatorial chemistry, to develop small molecule inhibitors. Such collaborations are possible here at Duke (see attached letters). This proposal has three broad long term/future goals: 1.) to examine more genes/mutants to get a global picture of C. neoformans pathogenesis; 2.) to apply this approach to other pathogenic fungi; and 3.) to develop small molecule inhibitors of gene products that are essential for pathogenesis.