Development of Lead Inhibitors of HIV Vif Binding to Antiretroviral A3G Through Medicinal Chemistry

项目摘要 HIV宿主限制性因子APOBEC 3G（A3 G）可以通过诱导灾难性的免疫应答来抑制HIV。 病毒基因组的突变HIV Vif蛋白通过与A3 G结合并指导 蛋白酶体降解途径。拟议的SBIR第一阶段的意义 研究表明，Vif仍然是一个新的抗病毒靶点，其临床潜力尚未完全发挥。 探讨了尽管有很多关于Vif的学术研究，但只有有限的努力去识别 Vif的小分子拮抗剂。OyaGen公司是目前唯一一家 识别以有意义的方式与Vif特异性相互作用的化学支架， 药物开发先前的抗Vif药物发现努力的固有限制是使用 在初级药物筛选中，“命中”可能是由于一种以上的作用机制。我们 设计了一种活细胞淬灭FRET（FqRET）报告基因测定，能够选择膜- 使用Vif突变体的Vif和A3 G之间蛋白质-蛋白质相互作用的渗透性拮抗剂 保留野生型与A3 G的结合，但不再与延伸蛋白C结合， A3 G降级。从110 K小分子药物样化合物库的筛选中， 确定了165个优先进行进一步分析的人。虽然一些化合物具有剂量依赖性， A3 G的Vif结合的拮抗剂中，一种化合物（下文称为C5）是无细胞毒性的 并且显示出剂量依赖性：i）抑制Vif依赖性A3 G降解，ii）抑制 假型HIV复制和iii）促进病毒体中A3 G掺入增加。这里 我们提出了一个创新药物发现的关键路径，以优化一种新的抗HIV先导化合物， 从C5开始，预期进行II期SBIR药代动力学分析、体内疗效试验， 国家研究开发扶持性研究。第一阶段SBIR目标将侧重于结构-活性关系（SAR） 用于抗HIV化合物的命中-先导优化的C5支架的药物化学， 抗病毒活性的机制是由于其作为Vif拮抗剂的功能， 依赖于VIF的降解。

Project Summary The HIV host restriction factor APOBEC3G (A3G) can inhibit HIV by inducing catastrophic mutations in the viral genome. The HIV Vif protein protects HIV by binding to A3G and directing it to the proteasomal degradation pathway. The significance of the proposed SBIR phase I research is that Vif remains a new antiviral target whose clinical potential has yet to be fully explored. Despite much academic research on Vif, only a limited effort has gone toward identifying small molecule antagonists of Vif. OyaGen, Inc. is the only commercial entity currently pursuing the identification of chemical scaffolds that specifically interact with Vif in meaningful ways for drug development. An inherent limitation of prior anti-Vif drug discovery efforts has been the use of primary drug screens in which ‘hits’ may be due to more than one mechanisms of action. We designed a live cell quenched FRET (FqRET) reporter assay that enables selection of membrane- permeable antagonists of protein-protein interactions between Vif and A3G using a Vif mutant that retains wild type binding to A3G but no longer binds to Elongin C and no longer facilitates A3G degradation. From a screen of a 110K small molecule library of drug-like compounds we identified 165 prioritized his for further analysis. Although a few compounds were dose-dependent antagonists of Vif binding of A3G, one compound, hereafter referred to as C5, was non-cytotoxic and displayed dose-dependent: i) inhibition of Vif-dependent A3G degradation, ii) inhibition of pseudotyped HIV replication and iii) promoted an increased incorporation of A3G in virions. Here we propose an innovative drug discovery critical path to optimize a novel anti-HIV lead compound from C5 in anticipation of phase II SBIR pharmacokinetic analysis, in vivo efficacy testing, and IND-enabling studies. The Phase I SBIR Aims will focus on structure-activity relationship (SAR) medicinal chemistry of the C5 scaffold for hit-to-lead optimization of anti-HIV compound whose antiviral activity is mechanistically due to its function as a Vif antagonist that protects A3G from Vif-dependent degradation.