Antibacterial Discovery via Phenotypic DNA-Encoded Library Screening

[Image: see text] The global rise of multidrug resistant infections poses an imminent, existential threat. Numerous pipelines have failed to convert biochemically active molecules into bona fide antibacterials, owing to a lack of chemical material with antibacterial-like physical properties in high-throughput screening compound libraries. Here, we demonstrate scalable design and synthesis of an antibacterial-like solid-phase DNA-encoded library (DEL, 7488 members) and facile hit deconvolution from whole-cell Escherichia coli and Bacillus subtilis cytotoxicity screens. The screen output identified two low-micromolar inhibitors of B. subtilis growth and recapitulated known structure–activity relationships of the fluoroquinolone antibacterial class. This phenotypic DEL screening strategy is also potentially applicable to adherent cells and will broadly enable the discovery and optimization of cell-active molecules.