Discovery of novel antivirulence drug targets in C. difficile by systematically demystifying its genetic network controlling toxin biosynthesis

Project Summary Clostridioides difficile infection (CDI), an antibiotic-associated diarrhea, is a leading cause of hospital-acquired infection and death. The traditional antibiotics metronidazole (MTZ) and Vancomycin (VAN) have been the mainstay treatment for CDI for over 30 years. However, these drugs are now associated with increased clinical failure. Postdoctoral research by the candidate revealed mechanisms of resistance to these agents and showed that resistance promotes poor clinical outcomes. Past research also highlighted the need for new CDI therapeutics that are microbiome-sparing and allow the gut microbiota to rebound during CDI therapy. This proposal tackles the need for microbiome-sparing CDI drugs, by seeking to explore genetic mechanisms that regulate the production of virulence toxins by C. difficile. The discovered gene determinants will represent potential drug targets and will help to fill the knowledge void concerning druggable metabolic and regulatory processes that control toxin production in C. difficile. The overarching hypothesis of aim 1 is that since toxin biosynthesis is a secondary metabolic step it will be subject to cellular genetic perturbations. We will identify gene determinants and cellular pathways that negatively affect toxin biosynthesis by high-throughput screening of transposon mutants. The key proteins will undergo druggability assessment and validated for effects on virulence using in vitro and in vivo methods. Aim 2 will adopt chemical genomic strategies on anti-virulence molecules to identify targets that are druggable. This study will advance fundamental scientific knowledge of C. difficile pathogenesis toward the discovery of new therapeutic agents. The K99 phase will be mentored by an exceptional panel of experts in C. difficile pathogenesis, drug discovery, and systems biology, to enable the development of skills in bioinformatics and high throughput screening. This support combined with the candidate’s expertise in molecular microbiology and protein biochemistry will facilitate the smooth transition to an independent scientist during the R00 phase. The institutional environment and support for career development are both strong. These studies will lay the foundation for future research aimed at developing innovative therapies that block target toxin biosynthesis in C. difficile and understanding of its pathogenic physiology.