Identification of Small Molecules Exhibiting Oxacillin Synergy through a Novel Assay for Inhibition of <i>vraTSR</i> Expression in Methicillin-Resistant Staphylococcus aureus

Abstract

Methicillin-resistant <i>Staphylococcus aureus</i> (MRSA) strains that are resistant to all forms of penicillin have become an increasingly common and urgent problem threatening human health. They are responsible for a wide variety of infectious diseases ranging from minor skin abscesses to life-threatening severe infections. The <i>vra</i> operon that is conserved among <i>S. aureus</i> strains encodes a three-component signal transduction system (<i>vraTSR</i>) that is responsible for sensing and responding to cell wall stress. We developed a novel and multifaceted assay to identify compounds that potentiate the activity of oxacillin, essentially restoring efficacy of oxacillin against MRSA, and performed high-throughput screening (HTS) to identify oxacillin potentiators. HTS of 13,840 small-molecule compounds from an antimicrobial-focused Life Chemicals library, using the MRSA cell-based assay, identified three different inhibitor scaffolds. Checkerboard assays for synergy with oxacillin, reverse transcriptase PCR (RT-PCR) assays against <i>vraR</i> expression, and direct confirmation of interaction with VraS by surface plasmon resonance (SPR) further verified them to be viable hit compounds. A subsequent structure-activity relationship (SAR) study of the best scaffold with diverse analogs was utilized to improve potency and provides a strong foundation for further development.