Targeting DHFR to Design Antimicrobial Agents

真菌感染已成为严重疾病的一个重要且不断增加的原因 死亡。光滑假丝酵母正在成为一种致命的真菌病原体， 对流行病原体假丝酵母菌已经造成的死亡有重要意义 白念珠菌。不幸的是，光肩星天牛对几种可用的 抗真菌药物，包括两性霉素B和唑类化合物。 治疗光滑毛囊线虫感染是非常必要的。此外，由于 系统性真菌感染通常会迅速走向死亡和正式诊断。 需要关键的时间，理想的治疗方法也应该是广谱抗真菌药 对主要的真菌病原体白念珠菌也有效的药剂。在基础上建设 以前在病原体中靶向二氢叶酸还原酶(DHFR)的成功，我们 合成了一类多功能的DHFR抑制剂。我们最初的系列中的几个 抑制剂对真菌DHFR酶有很强的抑制作用，选择性好 抗哺乳动物酶，在培养的生物中具有良好的抗真菌活性 几乎没有明显的哺乳动物细胞毒性。我们还明确了我们的一项 抑制剂与光肩星天牛DHFR结合，并测定其结构，分辨率为1.6？在……里面 这个应用程序的第一个目标是，我们建议使用结构指导的方法来设计 并合成有效和选择性地针对光肩星天牛DHFR的新类似物。在一个 第二个平行目标，我们将确定化合物在细胞和动物中的功效， 阐明抗性机制并确定抗性酶的结构。在一个 第三个目标，我们将评估这些抑制剂对白色念珠菌的抑制作用，并确定晶体 白念珠菌DHFR强效和选择性抑制剂的结构。在第四个目标中，我们 建议生成一种有效的、选择性的广谱抑制剂，该抑制剂具有 对两种假丝酵母菌均有良好的抗真菌活性，且无毒性 对抗哺乳动物细胞。

Fungal infections have become a significant and increasing cause of severe illness and death. Candida glabrata is emerging as a lethal fungal pathogen, contributing significantly to the mortality already caused by the prevalent pathogen, Candida albicans. Unfortunately, C. glabrata is inherently resistant to several of the available antifungal therapeutics, including amphotericin B and the azole compounds. Therapeutics to treat C. glabrata infections are critically necessary. Furthermore, since systemic fungal infections often proceed rapidly toward death and formal diagnosis requires critical time, an ideal therapeutic would also be a broad spectrum antifungal agent that is also effective against the primary fungal pathogen, C. albicans. Building on previous success to target dihydrofolate reductase (DHFR) in pathogenic organisms, we have synthesized a class of versatile DHFR inhibitors. Several of an initial series of our inhibitors exhibit strong potency against the fungal DHFR enzymes, good selectivity against the mammalian enzyme, good antifungal activity in cultures of the organisms and little appreciable mammalian cell toxicity. We have also crystallized one of our inhibitors bound to C. glabrata DHFR and determined the structure to 1.6 ¿ resolution. In the first aim of this application, we propose to use a structure-guided approach to design and synthesize new analogs that potently and selectively target C. glabrata DHFR. In a second parallel aim, we will determine efficacy of the compounds in cells and animals, elucidate resistance mechanisms and determine structures of resistant enzymes. In a third aim, we will evaluate these inhibitors against C. albicans and determine crystal structures of potent and selective inhibitors with C. albicans DHFR. In a fourth aim, we propose to generate a potent and selective broad spectrum inhibitor that exhibits excellent antifungal activity against both Candida species and maintains a lack of toxicity against mammalian cells.