Structural Characterization of Mycobacterium abscessus Phosphopantetheine Adenylyl Transferase Ligand Interactions: Implications for Fragment-Based Drug Design.

Anti-microbial resistance is a rising global healthcare concern that needs urgent attention as growing number of infections become difficult to treat with the currently available antibiotics. This is particularly true for mycobacterial infections like tuberculosis and leprosy and those with emerging opportunistic pathogens such as Mycobacterium abscessus, where multi-drug resistance leads to increased healthcare cost and mortality. M. abscessus is a highly drug-resistant non-tuberculous mycobacterium which causes life-threatening infections in people with chronic lung conditions such as cystic fibrosis. In this study, we explore M. abscessus phosphopantetheine adenylyl transferase (PPAT), an enzyme involved in the biosynthesis of Coenzyme A, as a target for the development of new antibiotics. We provide structural insights into substrate and feedback inhibitor binding modes of M. abscessus PPAT, thereby setting the basis for further chemical exploration of the enzyme. We then utilize a multi-dimensional fragment screening approach involving biophysical and structural analysis, followed by evaluation of compounds from a previous fragment-based drug discovery campaign against M. tuberculosis PPAT ortholog. This allowed the identification of an early-stage lead molecule exhibiting low micro molar affinity against M. abscessus PPAT (Kd 3.2 ± 0.8 µM) and potential new ways to design inhibitors against this enzyme. The resulting crystal structures reveal striking conformational changes and closure of solvent channel of M. abscessus PPAT hexamer providing novel strategies of inhibition. The study thus validates the ligandability of M. abscessus PPAT as an antibiotic target and identifies crucial starting points for structure-guided drug discovery against this bacterium.

Fragment-based discovery of a new class of inhibitors targeting mycobacterial tRNA modification.

Translational frameshift errors are often deleterious to the synthesis of functional proteins and could therefore be promoted therapeutically to kill bacteria. TrmD (tRNA-(N(1)G37) methyltransferase) is an essential tRNA modification enzyme in bacteria that prevents +1 errors in the reading frame during protein translation and represents an attractive potential target for the development of new antibiotics. Here, we describe the application of a structure-guided fragment-based drug discovery approach to the design of a new class of inhibitors against TrmD in Mycobacterium abscessus. Fragment library screening, followed by structure-guided chemical elaboration of hits, led to the rapid development of drug-like molecules with potent in vitro TrmD inhibitory activity. Several of these compounds exhibit activity against planktonic M. abscessus and M. tuberculosis as well as against intracellular M. abscessus and M. leprae, indicating their potential as the basis for a novel class of broad-spectrum mycobacterial drugs.

Development of Inhibitors against Mycobacterium abscessus tRNA (m1G37) Methyltransferase (TrmD) Using Fragment-Based Approaches.

Mycobacterium abscessus (Mab) is a rapidly growing species of multidrug-resistant nontuberculous mycobacteria that has emerged as a growing threat to individuals with cystic fibrosis and other pre-existing chronic lung diseases. Mab pulmonary infections are difficult, or sometimes impossible, to treat and result in accelerated lung function decline and premature death. There is therefore an urgent need to develop novel antibiotics with improved efficacy. tRNA (m1G37) methyltransferase (TrmD) is a promising target for novel antibiotics. It is essential in Mab and other mycobacteria, improving reading frame maintenance on the ribosome to prevent frameshift errors. In this work, a fragment-based approach was employed with the merging of two fragments bound to the active site, followed by structure-guided elaboration to design potent nanomolar inhibitors against Mab TrmD. Several of these compounds exhibit promising activity against mycobacterial species, including Mycobacterium tuberculosis and Mycobacterium leprae in addition to Mab, supporting the use of TrmD as a target for the development of antimycobacterial compounds.