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

> **NIH NIH R01** · COLORADO STATE UNIVERSITY · 2021 · $636,059

## Abstract

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 ...

## Key facts

- **NIH application ID:** 10077828
- **Project number:** 5R01AI150223-02
- **Recipient organization:** COLORADO STATE UNIVERSITY
- **Principal Investigator:** CHAOPING CHEN
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $636,059
- **Award type:** 5
- **Project period:** 2020-01-01 → 2023-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10077828

## Citation

> US National Institutes of Health, RePORTER application 10077828, HTS Targeting HIV-1 Protease Autoprocessing for First in Class Drug Discovery (5R01AI150223-02). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10077828. Licensed CC0.

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