Identification of small molecules exhibiting oxacillin synergy through a novel assay for inhibition of vraTSR expression in methicillin resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (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 vra operon that is conserved among S. aureus strains encodes a three-component signal transduction system (vraTSR) that is responsible for sensing and responding to cell-wall stress. We developed a novel and multi-faceted 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, RT-PCR assays against vraR expression and direct confirmation of interaction with VraS by surface plasmon resonance (SPR) further verified them to be viable hit compounds. 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