Optimizing IMPDH inhibitors for the treatment of cryptosporidiosis

描述（由申请人提供）：隐孢子虫种类，包括小隐孢子虫和人隐孢子虫，是一种原生动物寄生虫，尤其对免疫功能低下的个体和幼儿构成健康威胁。这些生物也是潜在的生物恐怖主义代理人，可以使大量人口丧失能力。目前的治疗方案有限。常用的抗寄生虫药物是无效的，唯一fda批准的药物效果很差，而且没有疫苗。基因组分析表明隐孢子虫依靠肌苷5′-单磷酸脱氢酶（IMPDH）产生鸟嘌呤核苷酸，从而生存。此外，疟原虫的IMPDH基因似乎是通过横向转移从一个ε-变形菌中获得的，因此，疟原虫的IMPDH （CpIMPDH）与人类的同源基因高度分化。因此，选择性CpIMPDH抑制剂可能为隐孢子虫病的治疗提供了一种有效的策略，并且对患者的毒性最小。迄今为止，已经发现了几种结构不同的CpIMPDH抑制剂，它们在细胞培养模型中表现出优异的酶效、对人IMPDH的选择性、抗寄生虫活性，其中一种化合物系列在急性隐孢子虫病小鼠模型中表现出体内疗效。本研究的总体目标是进一步完善CpIMPDH抑制剂，以达到对隐孢子虫病急性和慢性小鼠模型的疗效，以支持选择性抑制CpIMPDH是这种原生动物感染的可行治疗策略的假设，为研究界提供关于体内疗效的最佳化合物特性的指导，并进一步提供CpIMPDH作为分子靶点的临床前验证。这些目标将通过追求三个具体目标来实现：1)设计、合成和体外评估CpIMPDH抑制剂的CpIMPDH抑制效力和选择性、小孢子虫细胞培养感染模型和ADME特性（包括小鼠和人类肠道和肝脏微粒体稳定性、细胞摄取、外排和毒性以及ugt介导的糖醛酸化）；2)评估CpIMPDH抑制剂的体内药代动力学和急性毒性特性，包括在未感染和细小疟原虫感染小鼠的口服吸收和组织分布的体内模型；3)评估优化后的CpIMPDH抑制剂在隐孢子虫病急性和慢性动物模型中的作用，评估粪便卵囊排泄和胃肠道大体和组织病理学。此外，CpIMPDH抑制剂对共生菌群的影响也将被检查，以更充分地评估药效学效应，因为许多细菌具有与CpIMPDH相似的impdh。该项目还将制定限制药物在胃肠道分布的策略。由于这一策略也适用于其他胃肠道疾病，

DESCRIPTION (provided by applicant): Cryptosporidium species, including C. parvum and C. hominis, are protozoan parasites that present a health threat particularly to immunocompromised individuals and young children. These organisms are also potential bio-terrorism agents that could incapacitate large populations. Current treatment options are limited. Commonly used antiparasitic drugs are ineffective, the only FDA-approved drug is poorly efficacious, and vaccines are unavailable. Genomic analysis has revealed that Cryptosporidium species rely on inosine 5'- monophosphate dehydrogenase (IMPDH) for producing guanine nucleotides and, hence, survival. It also appears that the parasite obtained its IMPDH gene from an ε-proteobacterium by lateral transfer, so C. parvum IMPDH (CpIMPDH) is highly diverged from the human orthologs. Therefore, selective CpIMPDH inhibitors may provide an effective strategy for the treatment of cryptosporidiosis with minimum toxicity to the patient. To date, several structurally distinct classes of CpIMPDH inhibitors have been identified that have demonstrated excellent enzymatic potency, selectivity over human IMPDH, anti-parasitic activity in a cell culture model and, for one compound series, in vivo efficacy in an acute cryptosporidiosis mouse model. The overall goals of this proposed study are to further refine CpIMPDH inhibitors to achieve efficacy in both acute and chronic mouse models of cryptosporidiosis to support the hypothesis that selective CpIMPDH inhibition is a viable treatment strategy for this protozoan infection, provide guidance for the research community with respect to optimal compound properties for in vivo efficacy and provide further pre-clinical validation of CpIMPDH as a molecular target. These goals will be achieved by pursuing three specific aims: 1) design, synthesis and in vitro evaluation of CpIMPDH inhibitors in assays of CpIMPDH inhibitory potency and selectivity, a C. parvum cell culture infection model and ADME properties (including mouse and human intestinal and liver microsome stability, cellular uptake, efflux and toxicity, and UGT-mediated glucuronidation); 2) assess the in vivo pharmacokinetic and acute toxicity properties of CpIMPDH inhibitors, including in vivo models of oral absorption and tissue distribution in both non-infected and C. parvum infected mice; 3) evaluate optimized CpIMPDH inhibitors in acute and chronic animal models of cryptosporidiosis assessing fecal oocyst excretion and gastrointestinal gross and histopathology. In addition, the impact of CpIMPDH inhibitors on commensal bacteria populations will also be examined to more fully evaluate pharmacodynamics effects, since many bacteria have IMPDHs that are similar to CpIMPDH. This project will also develop strategies for limiting drug distribution to the gastrointestinal tract. Since this strategy could be applicable to other gastrointestinal diseases, such as irritable bowel disease and colon cancer, this work has broad implications beyond the treatment of cryptosporidiosis.