Discovery of novel anti-HBV compounds targeting host factors

描述（由申请方提供）：本R21探索性提案使用综合方法发现适合进一步优化的候选先导药物，作为慢性B型肝炎病毒（HBV）感染的治疗药物。我们的长期目标是开发具有不同于目前可用药物的新作用机制的HBV生物合成的小分子抑制剂。HBV研究界存在广泛的共识，即成功开发新型HBV药物的关键在于靶向病毒生命周期中的步骤，而不是HBV聚合酶/逆转录酶介导的复制。目前可用的药物，包括1型干扰素和核苷类似物，不提供解决慢性携带者状态的显著可能性。现有的HBV治疗方案的缺点包括终身治疗的必要性和次优的患者接受度和依从性。显然，需要具有新作用机制的新药来找到导致完全根除HBV感染的治愈方法。抗病毒治疗的开发不针对病毒，而是针对病毒生物合成所必需的细胞成分，这可能代表了产生HBV治疗方法的一种新方法。核苷类似物与靶向宿主因子的新型药物的组合可能潜在地导致防止新的cccDNA合成。治疗相关化合物的靶标鉴定技术的最新进展以及适用于HTS分析的用于评价HBV转录的简单测定的可用性允许多种方法来发现HBV的新治疗剂。我们假设，有可能通过使用以下方法的组合特异性靶向对小瓶前基因组RNA合成至关重要的细胞因子来鉴定和设计HBV生物合成的抑制剂：针对HBV核壳启动子活性的高通量筛选（HTS）、计算机辅助药物设计（CADD）、化学生物学、靶点识别方法、靶向合理药物设计、药物化学，和评价显示在细胞培养物中抑制HBV转录和复制的最有希望的化合物的功效。为了实现这一目标，我们的具体目标如下：目标1。使用具有可用的初级HTS、反筛选HTS和次级病毒复制测定的化学文库的高通量筛选，鉴定HBV核衣壳启动子活性的先导或药物样低分子量抑制剂。目标二。设计和合成目标1中最有前途的HBV抗病毒化合物的类似物，以产生SAR并鉴定适合于基于细胞和基于光亲和性的靶标鉴定实验的化合物。如果时间允许，将使用补充有一组能够支持HBV转录和复制的核受体的非肝癌细胞进行初步靶点鉴定研究，并进行光亲和标记，然后进行LC-MS/MS分析。

DESCRIPTION (provided by applicant): This R21 exploratory proposal uses an integrated approach to discover lead candidates suitable for further lead optimization as therapeutics for chronic hepatitis B virus (HBV) infections. Our long-term objective is to develop small molecule inhibitors of HBV biosynthesis with a novel mechanism of action distinct from the currently available drugs. A broad consensus exists in the HBV research community that the key to the successful development of new novel classes of HBV medications lies in targeting steps in the viral life cycle other than replication mediated by the HBV polymerase/reverse transcriptase. Currently available medications, including type 1 interferon and nucleosides analogs, do not offer a significant probability for the resolution of the chronic carrier state. The shortcomings o the existing HBV therapeutic regiments include the necessity of life-long treatment and suboptimal patient acceptance and adherence. Clearly, new drugs with novel mechanisms of action are needed to find a cure leading to complete eradication of HBV infection. Antiviral therapy development targeting not viral, but cellular components essential for viral biosynthesis may represent a novel approach for the generation of HBV therapeutics. A combination of nucleoside analogs with the novel drugs targeting host factors may potentially lead to the prevention of new cccDNA synthesis. Recent advances in target identification techniques for therapeutically relevant chemical compounds and the availability of simple assays amenable to HTS analysis for the evaluation of HBV transcription allow for a multitude of methods to discover new therapeutic agents for HBV. We hypothesize that it will be possible to identify and design inhibitors of HBV biosynthesis by specifically targeting cellular factors that are essential for vial pregenomic RNA synthesis using a combination of approaches: high- throughput screening (HTS) against HBV nucleocapsid promoter activity, computer-aided drug design (CADD), chemical biology, target identification methods, target-oriented rational drug design, medicinal chemistry, and evaluation for efficacy of the most promising compounds shown to inhibit HBV transcription and replication in cell culture. To achieve this goal, our specific aims are as follo: Aim 1. Identify lead- or drug-like low molecular weight inhibitors of HBV nucleocapsid promoter activity using high- throughput screening of chemical libraries with available primary HTS, counter-screening HTS, and secondary viral replication assays. Aim 2. Design and synthesize analogs of the most promising HBV antiviral compounds from aim 1 to generate a SAR and identify compounds suitable for cell-based and photoaffinity-based target identification experiments. If time permits, pilot target identification studies using non-hepatoma cells complemented with a panel of nuclear receptors capable of supporting HBV transcription and replication, and photoaffinity labeling followed by LC-MS/MS analysis will be performed.