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HTS Targeting HIV-1 Protease Autoprocessing for First in Class Drug Discovery

HTS 靶向 HIV-1 蛋白酶自动处理以实现一流药物发现

基本信息

批准号：
10553592
负责人：
CHAOPING CHEN
金额：
$ 51.07万
依托单位：
COLORADO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-01-01 至 2024-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10553592
关键词：
Affinity Binding Biochemical Biological Biological Assay Biological Sciences Catalogs Catalytic Domain Cells Chemicals Collaborations Collection Combined Modality Therapy Drug Design Drug resistance Enzyme-Linked Immunosorbent Assay Enzymes FDA approved Fractionation Genetic Goals HIV HIV-1 HIV-1 protease Lead Life Longevity Mediating Medical Michigan Microbe Molecular Conformation Mutation Natural Products Patients Peptide Hydrolases Performance Phenotype Predisposition Process Production Prognosis Protease Inhibitor Proteolysis RNA-Directed DNA Polymerase Reaction Reporting Resistance Sagittaria Structure Technology Therapeutic Therapeutic Agents Time Treatment Efficacy Universities Viral Virion Virus Inhibitors analog antiretroviral therapy assay development combat drug discovery follow-up high throughput screening improved inhibitor insight novel novel therapeutic intervention novel therapeutics pol Gene Products resistant strain response screening small molecule small molecule libraries therapeutic development therapeutic target therapy outcome

项目摘要

HTS Targeting HIV-1 Protease Autoprocessing for First in Class Drug Discovery Project Summary This proposal is in response to PAR-17-438: Assay development and screening for discovery of chemical probes or therapeutic agents (R01). The goal of this project is to carry out high-throughput screens and follow-up characterizations to identify molecules inhibiting HIV-1 protease (PR) autoprocessing with modes of action (MOAs) different from the currently available protease inhibitors (PIs). In the infected cell, HIV-1 PR is initially synthesized as part of the Gag-Pol polyprotein precursor with its proteolysis activity tightly suppressed. During late stage of virion production, the precursor self-catalyzes the cleavage reactions that lead to liberation of the free mature PR in a temporospatially regulated fashion. The FDA-approved HIV-1 PIs primarily target the catalytic site of the mature PR and they are significantly less effective at suppressing precursor-mediated autoprocessing, suggesting that these two forms of HIV-1 PR are enzymatically different. Also, the emergence of PI resistant strain in patients treated with PI-containing combination antiretroviral therapy (cART) is an ongoing problem that diminishes treatment efficacy, which warrants the need for new therapeutics. This project seeks to find novel autoprocessing inhibitors targeting the precursor at regions not recognized by the currently available PIs. Towards this goal, we have established a cell-based functional assay that has faithfully recapitulated the autoprocessing phenotypes observed with proviral constructs. This assay has also, for the first time, made it possible to screen for autoprocessing inhibitors by HTS using AlphaLISA (amplified luminescent proximity homogeneous assay ELISA) technology. Our pilot screens of ~26K small molecule compounds displayed satisfactory performance with Z’ factors >0.45, S/N ratios >10, and hit rates <0.1% although no confirmed hit was identified. Therefore, we plan to screen a collection of natural product extracts (40K extracts, 5-25 compounds per extract, totaling ~0.6 million chemicals) with a wild type and two PI resistant precursors in collaboration with Dr. David Sherman at University of Michigan Life Sciences Institute (Aim 1). In parallel, we will team up with Drs. Thomas Chung and Ian Pass at Sanford Burnham Prebys Medical Discovery Institute to screen their ~350K small molecule library (Aim 2). These HTS campaigns will hopefully identify a handful confirmed compounds that will be subjected to a battery of established secondary and tertiary assays (Aim 3) in order to find novel autoprocessing inhibitors that are different from the current HIV-1 PIs in their MOA. This next generation of therapeutic probes, when used in combination with the current PIs, will implement a new therapeutic approach: targeting a vital enzyme (HIV-1 protease) at two distinct functional states (precursor and mature PR) and at different regions (non-catalytic and catalytic sites) at the same time. Such a strategy is expected to drastically increase difficulty (genetic barrier) for HIV-1 to evolve viable strains simultaneously resistant to inhibitors from both classes to resist the resistance.

HTS靶向HIV-1蛋白酶自动加工，用于同类新药的发现