The pursuit of mechanism of action: Uncovering drug complexity in TB drug discovery

Whole cell-based phenotypic screens have become the primary mode of hit generation in tuberculosis (TB) drug discovery during the last two decades. Different drug screening models have been developed to mirror the complexity of TB disease in the laboratory. As these culture conditions are becoming more and more sophisticated, unraveling the drug target and the identification of the mechanism of action (MOA) of compounds of interest have additionally become more challenging. A good understanding of MOA is essential for the successful delivery of drug candidates for TB treatment due to the high level of complexity in the interactions between Mycobacterium tuberculosis (Mtb) and the TB drug used to treat the disease. There is no single "standard"protocol to follow and no single approach that is sufficient to fully investigate how a drug restrains Mtb. However, with the recent advancements in -omics technologies, there are multiple strategies that have been developed generally in the field of drug discovery that have been adapted to comprehensively characterize the MOAs of TB drugs in the laboratory. These approaches have led to the successful development of preclinical TB drug candidates, and to a better understanding of the pathogenesis of Mtb infection. In this review, we describe a plethora of efforts based upon genetic, metabolomic, biochemical, and computational approaches to investigate TB drug MOAs. We assess these different platforms for their strengths and limitations in TB drug MOA elucidation in the context of Mtb pathogenesis. With an emphasis on the essentiality of MOA identification, we outline the unmet needs in delivering TB drug candidates and provide direction for further TB drug discovery.