IMPDH inhibitors for the treatment of Cryptosporidium infections

项目总结/摘要 隐孢子虫（Cryptosporidium），如C. parvum是原生动物寄生虫，对人类健康构成重大威胁。 幼儿和免疫功能低下的成人。这些生物体是潜在的生物恐怖分子， 也使大量人口丧失能力。目前隐孢子虫感染的治疗选择是有限的。 常用的抗寄生虫药物是无效的，唯一FDA批准的药物效果不佳， 没有疫苗。隐孢子虫依赖肌苷5 '-单磷酸脱氢酶 （IMPDH），如通过基因组分析所揭示的，用于产生鸟嘌呤核苷酸并因此存活。的 IMPDH基因可能来源于ε-变形杆菌，因此C. Parvum IMPDH（CpIMPDH）高度 与人类直系同源物不同因此，选择性CpIMPDH抑制剂可提供有效的抗肿瘤治疗。 对患者毒性最小的隐孢子虫病治疗策略。到目前为止，我们已经确定了 几种结构上不同的CpIMPDH抑制剂， 效力、对人IMPDH的选择性、在细胞培养模型中的抗寄生虫活性，并且对于一种化合物 系列，在急性隐孢子虫病小鼠模型中的体内功效。本研究的总体目标是进一步 改进CpIMPDH抑制剂以在隐孢子虫病的急性和慢性小鼠模型中实现功效， 支持选择性CpIMPDH抑制是C. parvum 感染，为研究界提供关于最佳化合物特性的指导， 体内功效和临床前候选物的选择以用于进一步开发。这些目标将通过以下方式实现： 追求三个具体目标：1）设计和在CpIMPDH测定中体外评价CpIMPDH抑制剂 抑制效力和选择性，C. parvum细胞培养感染模型和ADME性质（包括 溶解性、肠微粒体稳定性、细胞摄取、外排、毒性和UGT介导的葡萄糖醛酸化）; 2） 评估CpIMPDH抑制剂的体内药代动力学和急性毒性特性，包括体内 正常和C. parvum感染的小鼠; 3）评估优化的 CpIMPDH抑制剂在隐孢子虫病的急性和慢性动物模型中的疗效评估，通过成像 荧光寄生虫、粪便卵囊排泄和胃肠道大体和组织病理学。此外该 将检查CpIMPDH抑制剂对肠道细菌种群的影响，以更全面地评价 药效学该项目还将制定一项肝肠循环战略，以最大限度地 这类药物可在胃肠道内释放，同时降低全身暴露和毒性风险。 1

PROJECT SUMMARY/ABSTRACT Cryptosporidium species, such as C. parvum, are protozoan parasites that present a significant health threat to young children and immunocompromised adults. These organisms are potential bio-terrorism agents that could incapacitate large populations, as well. Current treatment options for Cryptosporidium infections are limited. Commonly used anti-parasitic drugs are ineffective, the only FDA-approved drug is poorly efficacious, and vaccines are unavailable. Cryptosporidium species rely on inosine 5’-monophosphate dehydrogenase (IMPDH), as revealed by genomic analysis, for producing guanine nucleotides and, hence, survival. The parasite likely obtained its IMPDH gene from ε-proteobacterium, so C. parvum IMPDH (CpIMPDH) is highly diverged from the human orthologs. Consequently, selective CpIMPDH inhibitors may provide an effective strategy for the treatment of cryptosporidiosis with minimum toxicity to patients. To date, we have identified several structurally distinct classes of CpIMPDH inhibitors 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 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 C. parvum infections, provide guidance for the research community with respect to optimal compound properties for in vivo efficacy and selection of pre-clinical candidates for further development. These goals will be achieved by pursuing three specific aims: 1) design 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 solubility, intestinal microsomal stability, cellular uptake, efflux, toxicity, and UGT-mediated glucuronidation); 2) assess the in vivo pharmacokinetic and acute toxicity properties of CpIMPDH inhibitors, including in vivo models of absorption and tissue distribution in both normal and C. parvum infected mice; 3) evaluate optimized CpIMPDH inhibitors in acute and chronic animal models of cryptosporidiosis assessing efficacy via imaging of fluorescent parasites, fecal oocyst excretion and gastrointestinal gross and histopathology. In addition, the impact of CpIMPDH inhibitors on commensal bacteria populations will be examined to more fully evaluate pharmacodynamics. This project will also develop a strategy for enterohepatic recycling in order to maximize gastrointestinal concentrations, while reducing systemic exposure and toxicity risk. 1