High throughput screening and drug discovery for antagonists of the Ebola VP40 protein assembly

埃博拉 VP40 蛋白组装拮抗剂的高通量筛选和药物发现

基本信息

批准号：
10760573
负责人：
Jason Douglas Salter
金额：
$ 29.9万
依托单位：
OYAGEN, INC.
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-10 至 2025-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10760573
关键词：
Address Antiviral Agents Area Back Biogenesis Biological Assay Cause of Death Cell Cycle Cell membrane Cell-Mediated Cytolysis Cells Characteristics Chemicals Communities Complex Country Critical Pathways Cytoplasm Democratic Republic of the Congo Disease Outbreaks Diversity Library Dose Dryness Ebola Ebola virus Evaluation Filovirus Future Genetic Transcription Homo Immune Targeting Immune response Immune system Immunize In Vitro Individual Infection Intervention Lead Libraries Life Link Location Membrane Pathology Patients Persons Pharmaceutical Chemistry Pharmaceutical Preparations Plasma Cells Population Powder dose form Probability Production Promega Property Protein Binding Domain Proteins Publishing Recording of previous events Reporter Research Risk Services Structure Symptoms Testing Texas Therapeutic Therapeutic Agents Transcript Transfection Travel Triage Vaccination Viral Viral Genome Viral Physiology Viral Proteins Virion Virus Virus Assembly Virus Replication Virus-like particle World Health Organization Zaire Ebola virus antagonist antiviral drug development assay development biosafety level 4 facility cell type cytotoxic cytotoxicity drug candidate drug discovery effective therapy extracellular vesicles genetic variant high-throughput drug screening imaging system indexing inhibitor innovation medical countermeasure membrane assembly migration mutant new outbreak novel novel strategies novel therapeutic intervention novel therapeutics pandemic potential particle preclinical development previous outbreak protein crosslink protein protein interaction sangivamycin screening small molecule small molecule inhibitor social success targeted treatment

项目摘要

PROJECT SUMMARY This R43 proposal answers the call of the RFA PA-22-176 for assay development and chemical probe screening by addressing the unmet need for development of antiviral treatments for Ebola patients through an innovative high-content small molecule screen for antagonists of the Ebola VP40 matrix protein. There is a need for a fast-acting therapy that is independent of the immune system and targets essential viral proteins. Such a novel therapeutic is anticipated to enhance the survival probability for infected people in hot zones of an Ebola outbreak. We chose to target EBOV VP40 because it is absolutely required for EBOV particle assembly at the cell membrane, is capable of budding virus- like particles (VLPs) when expressed in isolation and VP40 protein-protein interaction domains have been structurally determined. Further guiding this application is our published pilot screen which established proof-of-concept by demonstrating the accessibility of Ebola VP40 protein-protein interactions to sangivamycin, a dual acting small molecule antagonist of both EBOV VP40 assembly of virions and the viral replication machinery. Given this success and due to anticipated complications with efficacy and MOA studies inherent to compounds with dual targets, our Specific Aims propose to screen a library of ~123,000 small molecule compounds to identify antagonists of VP40 accumulation at the cell membrane for VLP formation and release from cells through a fully automated, quantitative, and high-content assay. The assay has been vetted to quantify the effect of small molecules on the cellular distribution of a fluorescent VP40 expressed in 293T cells. Hits validated as dose-dependent by qHTS and displaying low cytotoxicity will be further prioritized based on their absolute requirement for VP40 in antiviral mechanism of action through counter screening with the VP40- independent minigenome assay. Lead compounds also will be prioritized by their favorable ADMET profiles. Our proposed critical path anticipates identifying 2-4 dose-dependent, VP40- selective antagonists with potent antiviral activity that display low cytotoxicity for future medicinal chemistry and preclinical development.

项目摘要该R43提案回答了RFA PA-22-176呼吁测定开发和化学探针筛查通过解决抗病毒的未满足需求通过创新的高含量小分子筛选为埃博拉患者的治疗用于埃博拉病毒VP40基质蛋白的拮抗剂。需要快速疗法这与免疫系统无关，并靶向必需的病毒蛋白。这样的预计新颖的治疗方法可以提高感染者的生存概率埃博拉疫情的热区。我们选择针对EBOV VP40，因为它绝对是 EBOV颗粒组件在细胞膜处所需的需要，能够发芽的病毒 - 像颗粒（VLP）一样，分别表示和VP40蛋白质蛋白质相互作用域已在结构上确定。进一步指导此申请是我们的已发表的飞行员屏幕，通过证明埃博拉病毒VP40蛋白质 - 蛋白质相互作用与Sangivamycin的可及性，这是一种双重作用 EBOV VP40病毒体和病毒的小分子拮抗剂复制机制。鉴于这一成功，并且由于预期的并发症我们的具体目的是具有双重目标的化合物固有的功效和MOA研究建议筛选约123,000个小分子化合物的库以识别拮抗剂 VP40在细胞膜上积聚的VLP形成并从细胞中释放通过全自动，定量和高含量测定法。该测定是审查以量化小分子对荧光的细胞分布的影响 VP40在293T细胞中表达。 QHTS依赖于剂量并显示的命中率低细胞毒性将根据其对VP40的绝对要求进一步优先考虑通过使用VP40独立的反筛查来抗病毒机理微型素组测定法。铅化合物也将由其优惠的助理优先考虑概况。我们提出的临界路径预计确定2-4剂量依赖性VP40- 具有有效抗病毒活性的选择性拮抗剂，表现出低细胞毒性的未来药物化学和临床前开发。