Inhibition of type II NADH dehydrogenase in Mycobacterium tuberculosis reveals collateral vulnerability in the electron transport chain

To sustain the anti-tuberculosis drug development pipeline, novel antibiotics must be developed both as standalone agents and as components of future combination regimens. To this end, tricyclic-spirolactams (TriSLa)-based inhibitors of mycobacterial type II NADH dehydrogenase (Ndh-2) represent a promising novel class of antibiotics, though activity is carbon source dependent. Initial studies established an enhanced understanding of the impact of TriSLa on mycobacteria using metabolomics and transcriptomics. Then, CRISPRi chemical-genetics and extensive in vitro TriSLa-combination studies showed that Ndh-2 inhibition resulted in rendering other components of the electron transport chain vulnerable, even in conditions when TriSLa was inactive alone. Finally, identified in vitro synergistic TriSLa-antibiotic combinations were validated to also give modestly improved in vivo efficacy. Together, this work expands our understanding of how Ndh-2 inhibition impacts bacterial physiology and reveals how it sensitizes the bacterium to electron transport chain inhibitors, providing a foundation for future TriSLa-based combination regimens.