SUMMARY
Tuberculosis (TB) is the leading cause of death from a single infection worldwide, with an annual death toll of
over 1.6 million lives. TB is curable and drug regimens can be above 90% effective if completed but are long and
toxic leading to noncompliance and emergence of resistance. New treatment options relying on novel and
unexplored targets are scarce. The BPaL regimen recently approved by the FDA features, for the first time in
many decades, drugs against new Mtb targets (ATP synthase) and with novel mode of action (nitroimidazoles).
Nevertheless, more inhibitors against previously unexplored targets are urgently needed to sustain the TB drug
pipeline, as resistance is already detected to components of BPaL. Both shortages, in effective antibiotics and
unexplored targets, are even more glaring in the field of non-tuberculous mycobacterial (NTM) diseases which
are on a rise and are recognized now as an emerging global threat in both immunocompromised and
immunocompetent individuals. Mycobacterial lipoamide dehydrogenase (Lpd) represents one of the unexplored
but Mtb-validated targets and serves in at least 4 enzyme complexes in Mtb: in pyruvate dehydrogenase (PDH),
-ketoacid dehydrogenase, branched chain keto-acid dehydrogenase, and peroxynitrite reductase/peroxidase.
Lpd is a genetically validated target: Mtb with lpd deleted does not survive in a mouse model of TB infection and
a conditionally regulated lpd Mtb strain is cleared in mice, when Lpd expression is suppressed in either the acute
or chronic infection. This application leverages the knowledge of the chemical biology of Mtb Lpd to explore our
newly identified tight binding inhibitors (TBI) of Mtb Lpd by structure-guided analysis, enzyme kinetics and
binding assays to define Lpd binding site features responsible for tight binding interactions and extensions of the
on-target residence time (t1/2). We aim to define what drives antibacterial activity of Lpd TBI, how t1/2 contributes
to whole cell activity and selectivity, and how we can design/predict better analogs with improved t1/2 to enable
further efficacy improvements. We will develop a framework for progression of TBIs from an isolated target to its
cellular PDH complex target and its whole cell activity against TB in culture and during host infection. Extension
of those studies to NTM pathogens and testing against Mycobacterium avium and M. abscessus Lpd orthologs
will lead to scaffold progression and SAR development of a novel whole cell active inhibitor of NTM. Collectively
our studies will explore the SAR of Lpd TBI in vitro, define vulnerability of mycobacterial Lpd to chemical inhibition
in vivo, advance our understanding of ways to improve inhibitors' efficacy through its target's t1/2, contribute to
rational prediction of in vivo efficacy and characterize structural features of mycobacterial PDH that define its
distinct macromolecular organization and sustain its functional diversity.