This mode-of-action (MOA) core will support all 3 projects in our CETR. Our core's purpose is to divine the MOA of novel lead compounds identified by the projects and filtered through the in vitro and in vivo cores. Antimicrobial development requires knowledge of a drug target and MOA to enable downstream refinements and improvements of leads. We propose chemical-genetic strategies together with cutting edge tools and approaches to discern MOA of drug leads. We will exploit the principle of fitness testing where loss of one or both copies of a gene renders a mutant fungus or bacterium hyper- or hypo-sensitive to a specific drug. The hits from this analysis include genes encoding the specific drug target, or effectors ofthe drug's metabolism. We will capitalize on available mutant libraries that span the genomes of pathogenic fungi and bacteria, including Candida albicans and Staphlococcus aureus. Our strategy will allow us to screen leads across a comprehensive set of drug targets rather than just a single target. Thus, for antifungal candidates, drug induced growth alteration of strains in a library of heterozygous deletion mutants will reveal a potential drug target and MOA - a so-called haploin sufficiency profile in which strain-specific barcodes uniquely mark mutants, and those with altered-fitness are detected by lllumina sequencing. For antibacterial leads, high throughput screens will be used to assay mutant libraries of gram-positive and -negative species for drug sensitivity. Drug targets and MOAs will be confirmed with downstream genetic and biochemical studies. We have several leads in hand to begin our studies. From ongoing work by our group, we expect to study 2-4 leads (up to 10) annually for MOA. Our core is significant because it will generate vital new knowledge about promising antimicrobial drug leads. The insight on drug MOA will be needed to fuel development of new drugs against resistant infectious disease. Our research findings will exert a sustained and powerful influence on the field because it will improve the care of patients infected with resistant fungal and bacterial pathogens. These achievements will help the NIH CETR program realize its goals of improving the public's health.
Novel antimicrobial drug leads are urgently needed to combat emerging resistance of fungal and bacterial agents of infectious disease. Our core is designed to delineate the mode of action of novel antimicrobial drug leads so that the compounds can be refined, improved and readied in the long term for clinical use.