Molina Panadero, I.; Moreno Rodriguez, A.; de la Cruz, M.; Sanchez, P.; Tomas Gallardo, L.; Samernate, T.; Sencanski, M.; Glisic, S.; Genilloud, O.; Nonejuie, P.; Perez Pulido, A. J.; Hmadcha, A.; Smani, Y.

High-throughput screening studies provide an additional approach to discovering repurposed drugs for antimicrobial treatments. In this work, we report the identification of ENOblock, an anticancer drug, as a novel antibiotic class. We computationally and experimentally validated that ENOblock synergizes with colistin, the last resort antibiotic, the colistin. Additionally, we identified enolase as the potential bacterial target for ENOblock. The in silico and in vitro antibacterial activity of ENOblock translated into potent in vivo efficacy in animal infection model. Collectively, the preclinical data support the selection of ENOblock as a promising candidate for antimicrobial development, with the potential to address the urgent threat of infections caused by Acinetobacter baumannii.