# Design and application of small molecules for the interception of quorum sensing in Staphylococcus aureus

> **NIH NIH F32** · UNIVERSITY OF WISCONSIN-MADISON · 2024 · $73,408

## Abstract

PROJECT SUMMARY
Quorum sensing (QS) is the process by which bacteria of the same species coordinate behavior at a high
population density. In many pathogenic bacteria, QS systems are used to regulate virulence. This proposal
focuses on the accessory gene regulator (agr) QS system found in the Gram-positive pathogen
Staphylococcus aureus, which uses agr to regulate an arsenal of toxins and biofilm formation. Small molecules
that inhibit agr QS have the potential to be used as therapeutics to combat S. aureus infection. Previous small
molecules that have been reported to inhibit agr QS suffer from low potencies, poorly characterized
mechanisms of action, and/or associated toxicities that render them impractical for use as chemical probes.
Herein I propose to develop a new class of small molecule agr inhibitors, study their biochemical mechanisms
of agr inhibition, and investigate their effects on S. aureus growth/invasion in human cells. In Aim 1, I will
develop a class of potent agr inhibitors through structure activity relationship (SAR) analyses on small
molecule agr modulators recently identified in our lab. I will design new synthetic routes to access the core
structure of two best-in-class small molecule inhibitors that represent the most potent small molecule inhibitors
of S. aureus agr QS to date. These redesigned syntheses will enable facile functional group variation for the
delineation of SARs and allow for installation of reactive groups for use in photoaffinity probes. My primary
goals will be to improve compound potency, membrane permeability, solubility, and stability. In Aim 2, I will
characterize the mechanisms of agr inhibition by these new small molecule scaffolds. Almost all known agr
inhibitors target autoinducer (AIP) binding to the transmembrane histidine kinase AgrC or intracellular response
regulator AgrA:DNA binding. Elucidating the mechanisms of agr inhibition by these new small molecules may
provide access to chemical probes that target other components of the agr QS system. I will use a series of in
vitro and cell-based assays to determine the target protein/pathway of these compounds, and photoreactive
probes to characterize ligand binding sites on these targets. In Aim 3, I will explore the effects of these small
molecule agr inhibitors on S. aureus growth/invasion in mammalian cells. S. aureus can use agr to evade the
immune system and cause persistent infection by infiltrating a wide range of human cell types. Macrophage
invasion assays will be used to evaluate the cell permeability of the lead small molecule agr inhibitors
developed in Aims 1 and 2, their ability to be phagocytosed, and whether inhibition of agr QS by these
compounds decreases macrophage invasion by S. aureus. Lastly, the inhibition/dysfunction of agr QS may
enable the development of S. aureus small colony variants (SCVs) that are less likely to evoke an immune
response. The ability of these small molecules to induce transition to SCVs by agr inhib...

## Key facts

- **NIH application ID:** 10995514
- **Project number:** 1F32GM155981-01
- **Recipient organization:** UNIVERSITY OF WISCONSIN-MADISON
- **Principal Investigator:** Alyssa Paige Clay
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $73,408
- **Award type:** 1
- **Project period:** 2024-12-01 → 2027-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10995514

## Citation

> US National Institutes of Health, RePORTER application 10995514, Design and application of small molecules for the interception of quorum sensing in Staphylococcus aureus (1F32GM155981-01). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/10995514. Licensed CC0.

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