Rational design of CNS-permeable cathepsin L inhibitors for treatment of chronic toxoplasmosis

项目摘要 慢性弓形虫（Tg）感染完全缺乏治疗选择， 10亿人面临再次感染弓形虫病的风险。先天性感染者或身体虚弱者 免疫系统特别容易受到表现为致命性脑炎的再活化弓形体病的影响， 心肌炎或视力丧失。作为感染性后葡萄膜炎的主要原因和 在美国的食源性死亡，重新激活弓形虫病可以大大减少高风险 个体通过消除Tg组织囊肿。我们使用新的遗传工具来确定一个重要的作用， 组织蛋白酶L（CPL）活性在慢性Tg感染，创造了一个令人兴奋的机会，利用 对抗复活的弓形虫病的新目标。为了开始解决这一关键的未满足需求，我们 基于其CNS抑制潜力鉴定了一种初始先导二肽腈CPL抑制剂， 进行了针对Tg和人组织蛋白酶L的主要SAR研究，证明我们可以实现 在20个类似物中对Tg酶的选择性提高超过100倍。在R21阶段， 使用基于结构的设计来进一步优化效力和选择性，沿着提高稳定性， 在体外模型中的渗透性，递送一种或多种有效的、选择性的、稳定的和细胞可渗透的引导物。在 在R33阶段，我们将在测量之前评估并进一步改进小鼠中PK和CNS代谢率 在已建立的慢性Tg感染的鼠治疗模型中的最大耐受剂量和功效。 这两个阶段都将采用为研究开发的第一种囊肿活力测定法。一旦成功 完成后，该项目将产生一种或多种Tg CPL抑制剂，可有效减少或消除组织 囊肿，从而为慢性Tg感染提供了一种潜在的新解决方案。 1

PROJECT SUMMARY The complete absence of treatment options for chronic Toxoplasma gondii (Tg) infection renders ~2 billion people at risk for reactivated toxoplasmosis. Congenitally infected individuals or those with weakened immune systems are particularly vulnerable to reactivated toxoplasmosis manifested as fatal encephalitis, myocarditis or loss of vision. As the leading cause of infectious posterior uveitis and second leading cause of foodborne deaths in the USA, reactivated toxoplasmosis could be substantially reduced in high-risk individuals by eliminating Tg tissue cysts. We used new genetic tools to identify an essential role for a cathepsin protease L (CPL) activity during chronic Tg infection, creating an exciting opportunity to exploit a new target for combatting reactivated toxoplasmosis. To begin addressing this key unmet need, we identified an initial lead dipeptide nitrile CPL inhibitor based on its potential for CNS penetrance and conducted primary SAR studies against Tg and human cathepsin L, demonstrating that we could achieve over 100-fold improvement in selectivity for the Tg enzyme in under 20 analogs. In the R21 phase we will use structure-based design to further optimize potency and selectivity along with improving stability and permeability in in vitro models, delivering one or more potent, selective, stable, and cell-permeable leads. In the R33 phase we will evaluate and further refine PK and CNS penetrance in mice before measuring maximum tolerated dose and efficacy in an established murine treatment model for chronic Tg infection. Both phases will feature first-of-their-kind assays for cyst viability developed for the studies. Upon successful completion, this project will yield one or more Tg CPL inhibitors effective for reducing or eliminating tissue cysts, thereby advancing a potential new solution for chronic Tg infection. 1