Design of C. parvum and T. gondii DHFR-TS Inhibitors

描述（由申请人提供）： 由微小隐孢子虫和刚地弓形虫引起的寄生虫感染对于免疫功能低下的患者、儿童和老年人来说是威胁生命的疾病。目前没有有效的治疗方法可用于隐孢子虫病和弓形虫病的治疗需要共同管理的sulfadoxine，化合物，许多患者有严重的不良反应。开发对病原体高度有效和高度选择性的新疗法具有直接重要性。已验证的药物靶标二氢叶酸还原酶-胸苷酸合酶（DHFR-TS）的晶体结构，DHFR-TS是原生动物中的一种双功能酶，来自C. parvum和T.为针对这些靶点的基于结构的药物设计提供了必要的证据。在这个建议的第一个目标中，我们将设计甲氧苄啶类似物与C. parvum DHFRTS结构。将通过比较致病性DHFR-TS和人DHFR的晶体结构来确定物种选择性元素。新的抑制剂将被建模成酶的结构，同时考虑配体诱导的构象变化，并预测结合模式。甲氧苄啶类似物的晶体结构将在迭代循环中指导未来的设计。在第二个目标中，我们将提高一个有前途的T的效力和选择性。弓形虫DHFR-TS抑制剂，利用T.弓形虫DHFR-TS结合亲脂性抑制剂。对T.弓形虫DHFR-TS抑制剂将利用物种选择性元件。在第三个目标中，我们将阐明DHFR中乙胺嘧啶抗性的结构基础，并设计一种新的乙胺嘧啶抗性抑制剂。弓形虫DHFR-TS. DHFR对乙胺嘧啶和其他抗微生物剂的耐药性是一个威胁性的问题，迫切需要能够抑制耐药酶的新疗法。我们将解决乙胺嘧啶抗性DHFR-TS的晶体结构结合到一种新的和令人兴奋的抑制剂，在初步研究中显示对抗性酶的抑制常数为350 nM。利用结构信息，我们将阐明耐药性的结构基础，并修改新的抑制剂，以获得更大的效力和选择性。

DESCRIPTION (provided by applicant): Opportunistic infections caused by Cryptosporidium parvum and Toxoplasma gondii represent life threatening diseases for immuno-compromised patients, children and the elderly. There are currently no effective treatments available for cryptosporidiosis and treatments for toxoplasmosis require the coadministration of sulfadioxine, a compound to which many patients have severe adverse reactions. The development of novel therapeutics that are highly potent and highly selective for the pathogen is of immediate importance. Crystal structures of the validated drug target, dihydrofolate reductase-thymidylate synthase (DHFR-TS), a bifunctional enzyme in protozoa, from C. parvum and T. gondii, provide essential evidence for structure-based drug design against these targets. In the first Aim of this proposal, we will design trimethoprim analogs to interact favorably with species-selective elements of the C. parvum DHFRTS structure. Species-selective elements will be determined by comparing crystal structures of pathogenic DHFR-TS and human DHFR. New inhibitors will be modeled into the structure of the enzyme, while accounting for ligand-induced conformational changes, and binding modes predicted. Crystal structures of trimethoprim analogs will guide future design in an iterative cycle. In the second Aim, we will improve the potency and selectivity of a promising T. gondii DHFR-TS inhibitor, using information from crystal structures of T. gondii DHFR-TS bound to lipophilic inhibitors. Designs for the improvement of T. gondii DHFR-TS inhibitors will take advantage of species-selective elements. In the third Aim, we will elucidate the structural basis of pyrimethamine resistance in DHFR and design potency and selectivity into a novel inhibitor of pyrimethamine-resistant T. gondii DHFR-TS. DHFR resistance to pyrimethamine and other antimicrobials is a threatening problem and novel therapeutics capable of inhibiting the resistant enzymes are desperately needed. We will solve a crystal structure of pyrimethamine-resistant DHFR-TS bound to a novel and exciting inhibitor that shows an inhibition constant of 350 nM against the resistant enzyme in preliminary studies. Using the structural information we will elucidate the structural basis of resistance and modify the novel inhibitor for greater potency and selectivity.