Inhibitors of Plasmodium liver infection

摘要 疟疾是由原生动物寄生虫疟原虫引起的。它始于人的疟原虫子孢子感染 肝脏。这一步骤对于寄生虫数量的扩大和随后的症状性红细胞周期至关重要。 间日疟原虫形成的休眠肝期是疟疾复发的主要原因。因此，抑制前红细胞 感染将防止疟疾病理和间日疟原虫复发。目前针对红细胞前阶段的药物有 严重的副作用或代价高昂。因此，迫切需要针对红细胞前阶段的新药。 我们建议启动一项药物化学努力，以优化恶性疟原虫cGMP依赖蛋白的抑制剂。 蛋白激酶(PKG)。PKG在子孢子侵入肝细胞和随后的肝期发育过程中是必不可少的。它的 三取代鹿茸(TSP)的化学抑制在组织培养试验和小鼠中防止肝脏感染。我们 假设对TSP物理性质的优化将产生对P. Berghei肝功能分期。我们的合作工作结合了红细胞前期生物学和 激酶、酶抑制剂的药物化学和计算机药物设计。 我们的具体目标是： 具体目标1：设计和合成新型TSP类似物。以恶性疟原虫为指导的药物化学技术 PKG(PfPKG)X射线晶体结构将用于合成预计具有更高效力和 渗透性特性。 具体目标2：测定TSP类似物的体外效力和肝期活性。对PfPKG酶的效力， 全细胞抗恶性疟原虫红细胞期和伯氏疟原虫感染HepG2细胞的活性，ADME， 在选择寄生虫选择性化合物进行测试时，将考虑毒性和专一性研究。 具体目标3：测定TSP类似物的体内药效。化合物(单一浓度，多次给药)将 通过口服和静脉注射感染了表达荧光素酶的伯氏假单胞菌子孢子的小鼠进行测试。 肝期感染将使用发光测量进行量化。显著抑制肝脏的化合物 寄生虫血症可能具有合理的药代动力学和药效学特性，适用于未来 优化。

ABSTRACT Malaria is caused by the protozoan parasite, Plasmodium. It begins with the infection by Plasmodium sporozoites of the liver. This step is essential for the expansion of parasite numbers and the subsequent symptomatic erythrocytic cycle. Dormant liver stages formed by P. vivax are the major cause of malaria relapses. Therefore, inhibition of pre-erythrocytic infection will prevent malaria pathology and relapses from P. vivax. Current drugs against pre-erythrocytic stages have significant side-effects or are expensive. Therefore, there is an urgent need for new drugs against pre-erythrocytic stages. We propose to initiate a medicinal chemistry effort to optimize an inhibitor of P. falciparum's cGMP-dependent protein kinase (PKG). PKG is essential for sporozoite invasion of hepatocytes and subsequent development in liver stages. Its chemical inhibition by a trisubstituted pyrolle (TSP) prevents liver infection in tissue culture assays and in mice. We hypothesize that optimization of TSP's physical properties will yield compounds with low dose efficacy against P. berghei liver stages. Our collaborative work combines expertise and experience in pre-erythrocytic stage biology and kinases, medicinal chemistry of kinase inhibitors and computational drug design. Our specific aims are: Specific Aim 1: Design and synthesize novel TSP analogs. Medicinal chemistry techniques guided by P. falciparum PKG (PfPKG) X-ray crystal structure will be used to synthesize analogs predicted to have improved potency and permeability properties. Specific Aim 2: Determine in vitro potency and liver stage activity of TSP analogs. Potency against PfPKG enzyme, whole-cell activity against P. falciparum erythrocytic stages and P. berghei sporozoite infection of HepG2 cells, ADME, toxicity and specificity studies will be considered in selecting parasite-selective compounds for testing in Aim 3 Specific Aim 3: Determine in vivo efficacy of TSP analogs. Compounds (single concentration, multiple dosing) will be tested through oral and intravenous administration to mice infected with luciferase-expressing P. berghei sporozoites. Liver stage infection will be quantified using luminescence measurements. Compounds that significantly inhibit liver parasitemia are likely to possess reasonable pharmacokinetic and pharmacodynamics properties, appropriate for future optimization.