Calcium Dependent Protein Kinase 1 as a drug target for T. gondii and C. parvum

描述(由申请人提供)： 需要新的疗法来治疗由微小隐孢子虫和弓形虫引起的感染。弓形虫的钙依赖蛋白激酶1(CDPK1)被认为在弓形虫的侵袭过程中起关键作用，因此是弓形虫病的药物靶点。由此推测，微小隐孢子虫的同源CDPK可能在隐孢子虫感染中起关键作用。我们已经解决了TgCDPK1和CpCDPK1的晶体结构，并表明活性部位对不抑制哺乳动物激酶的“凹凸不平”的激酶抑制剂(BKI)敏感。这种不同的敏感性是由于在哺乳动物蛋白激酶中存在的TgCDPK1和CpCDPK1的ATP结合位置上没有庞大的门卫侧链。因此，这些BKI为抑制TgCDPK1和CpCDPK1提供了极大的选择性，而不是人的激酶。此外，BKI化合物在其他工作过程中给药时，对小鼠的毒性最小。我们的初步结果表明，基于一个已知支架的多个BKI化合物可以抑制TgCDPK1和CpCDPK1，并在中低纳米分子浓度下抑制弓形虫和微小弧菌的细胞侵袭。突变的TgCDPK1与Met网守一起在弓形虫细胞中的表达导致对BKI效应的抗性，表明BKI通过CDPK1抑制细胞进入。我们将开发口服生物可用、足够有效、无毒性的化合物，并治愈弓形虫和/或微小弓形虫的动物模型。到这个项目结束时，我们希望确定和鉴定2到4个先导化合物，作为治疗隐孢子虫病和弓形虫病的潜在药物。该项目将启动新药的开发，以治疗由两种寄生原虫--隐孢子虫和弓形虫--引起的疾病，这两种寄生虫通常通过食用不纯净的食物或水传播。我们已经确定了一类化合物，这些化合物专门抑制这些寄生虫用来入侵人类细胞的一种特定蛋白质。我们将设计和表征这类化合物，这些化合物对人类无毒，但在治疗感染方面有效。 相关性：该项目将启动新药开发，以治疗由两种寄生原虫--隐孢子虫和弓形虫--引起的疾病，这两种寄生虫通常通过食用不纯净的食物或水传播。我们已经确定了一类化合物，这些化合物专门抑制这些寄生虫用来入侵人类细胞的一种特定蛋白质。我们将设计和表征这类化合物，这些化合物对人类无毒，但在治疗感染方面有效。

DESCRIPTION (provided by applicant): New therapeutics are needed for infections caused by Cryptosporidium parvum and Toxoplasma gondii. Calcium dependent protein kinase 1 (CDPK1) of T. gondii is thought to be critical for the invasion process of T. gondii and thus is a drug target for toxoplasmosis. By inference, the homologous CDPK of C. parvum is likely to be critical for infection by cryptosporidium. We have solved the crystal structures of TgCDPK1 and CpCDPK1 and have shown the active sites are susceptible to "bumped" kinase inhibitors (BKI) that do not inhibit mammalian kinases. This differential sensitivity is due to the absence of a bulky gatekeeper side chain in the ATP binding site of both TgCDPK1 and CpCDPK1 that is present in mammalian protein kinases. Thus, these BKI offer tremendous selectivity for inhibition of TgCDPK1 & CpCDPK1 vs. human kinases. Furthermore, BKI compounds have shown minimal toxicity in mice when administered in the course of other work. Our preliminary results show that multiple BKI compounds based on a known scaffold can inhibit TgCDPK1 & CpCDPK1 and also inhibit T. gondii and C. parvum cell invasion at low-mid nanomolar concentrations. Expression of a mutant TgCDPK1 with a Met gatekeeper in T. gondii cells leads to resistance to the BKI effect, demonstrating the BKI inhibits cell entry via CDPK1. We will develop compounds that are orally bioavailable, sufficiently potent, lack toxicity, and cure animal models of T. gondii and/or C. parvum. By the end of this project we expect to identify and characterize 2 to 4 leads for evaluation as potential drugs for cryptosporidiosis and toxoplasmosis. This project will initiate development of new drugs to treat diseases caused by two parasitic protozoa, Cryptosporidium and Toxoplasma that are commonly transmitted by consumption of impure food or water. We have identified a class of chemical compounds that specifically inhibit a particular protein used by these parasites to invade human cells. We will design and characterize compounds of this class that are nontoxic to humans but effective in treating infection. Relevance: This project will initiate development of new drugs to treat diseases caused by two parasitic protozoa, Cryptosporidium and Toxoplasma, that are commonly transmitted by consumption of impure food or water. We have identified a class of chemical compounds that specifically inhibit a particular protein used by these parasites to invade human cells. We will design and characterize compounds of this class that are nontoxic to humans but effective in treating infection.