REP3123, a novel inhibitor of methionyl tRNA synthetase from C. difficile
Abstract number: P577
Critchley I., Jarvis T., Guiles J., Sun X., Evans R., Davies D., Green L., Ochsner U., Young C., Citron D., Bell S., Drolet D., Gill S., Janjic N.
Objectives:C. difficile-associated disease (CDAD) is caused by overgrowth of toxin-producing strains of C. difficile (CD) following disruption of normal gut flora. Currently, there are few therapeutic options and clinical problems are associated with disease recurrence and the persistence of spores in the environment. In this report, we describe biochemical, crystallographic, microbiological and in vivo efficacy properties of REP3123, a novel diaryldiamine inhibitor of CD methionyl tRNA synthetase (MetRS).
Methods: REP3123 was synthesised and purified as a single active enantiomer. CD MetRS was expressed in E. coli and purified to homogeneity. The enzyme was crystallised by sitting drop vapor diffusion and structures were solved by molecular replacement. Microbiological testing was done according to CLSI guidelines. Toxins A and B were detected by semi-quantitative immunoassays, Western blot and cytotoxicity assays. In vivo efficacy of REP3123 was evaluated in the hamster model of CDAD.
Results: The tight binding of REP3123 to MetRS (Ki=20 pM) is competitive with methionine but cooperative with ATP. Inhibitor binding is accompanied with a conformational change in the enzyme in which the inhibitor occupies the methionine binding pocket and a newly created hydrophobic pocket. REP3123 is highly active against CD, including the NAP1/027 outbreak strain (MIC range, 0.251.0 mg/L). In contrast, REP3123 is inactive against Gram-negative bacteria and many anaerobic bacteria that constitute normal gut flora such as Actinomyces, Bacteroides, Bifidobacterium and Lactobacillus species. As a protein synthesis inhibitor, REP3123 inhibits the production of CD toxins A and B under a variety of culture conditions. REP3123 is also highly effective in inhibiting spore formation. In vivo, REP3123 exhibits low oral bioavailability and superior efficacy to vancomycin in the hamster model of CDAD.
Conclusion: REP3123 is a novel mechanism-of-action antibacterial agent that exhibits potent and selective inhibition of CD with limited potential for disruption of normal gut flora. REP3123 effectively inhibits toxin production and sporulation, exhibits low oral bioavailability and protects against CDAD in vivo. With these unique features, REP3123 represents a promising new development candidate for the treatment of CDAD.
|Session name:||18th European Congress of Clinical Microbiology and Infectious Diseases|
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