Abstract
Mycobacterium abscessus is the most common rapidly growing non-tuberculous mycobacteria to cause pulmonary disease in patients with impaired lung function such as cystic fibrosis. M. abscessus displays high intrinsic resistance to common antibiotics and inducible resistance to macrolides like clarithromycin. As such, M. abscessus is clinically resistant to the entire regimen of front-line M. tuberculosis drugs, and treatment with antibiotics that do inhibit M. abscessus in the lab results in cure rates of 50% or less. Here, we identified epetraborole (EPT) from the MMV pandemic response box as an inhibitor against the essential protein leucyl-tRNA synthetase (LeuRS) in M. abscessus. EPT protected zebrafish from lethal M. abscessus infection and did not induce self-resistance nor against clarithromycin. Contrary to most antimycobacterials, the whole-cell activity of EPT was greater against M. abscessus than M. tuberculosis, but crystallographic and equilibrium binding data showed that EPT binds LeuRSMabs and LeuRSMtb with similar residues and dissociation constants. Since EPT-resistant M. abscessus mutants lost LeuRS editing activity, these mutants became susceptible to misaminoacylation with leucine mimics like the non-proteinogenic amino acid norvaline. Proteomic analysis revealed that when M. abscessus LeuRS mutants were fed norvaline, leucine residues in proteins were replaced by norvaline, inducing the unfolded protein response with temporal changes in expression of GroEL chaperonins and Clp proteases. This supports our in vitro data that supplementation of media with norvaline reduced the emergence of EPT mutants in both M. abscessus and M. tuberculosis. Furthermore, the combination of EPT and norvaline had improved in vivo efficacy compared to EPT in a murine model of M. abscessus infection. Our results emphasize the effectiveness of EPT against the clinically relevant cystic fibrosis pathogen M. abscessus, and these findings also suggest norvaline adjunct therapy with EPT could be beneficial for M. abscessus and other mycobacterial infections like tuberculosis.
Highlights
Mycobacterium abscessus is a nontuberculous mycobacterium that commonly causes chronic lung disease, especially among patients with cystic fibrosis [1]
We showed that EPT targets the editing domain of the leucyl-tRNA synthetase (LeuRS) and that escape mutants lost LeuRS editing activity, making these mutants susceptible to misaminoacylation with leucine mimics
Combination therapy of EPT and norvaline limited the rate of EPT resistance in both M. abscessus and M. tuberculosis, and this was the first study to demonstrate improved in vivo efficacy of EPT and norvaline compared to EPT in a murine model of M. abscessus pulmonary infection
Summary
Mycobacterium abscessus is a nontuberculous mycobacterium that commonly causes chronic lung disease, especially among patients with cystic fibrosis [1]. AaRSs are enzymes that aminoacylate tRNAs with their cognate amino acids. All aaRSs have an aminoacylation domain while some are bifunctional and contain an editing domain. Some aaRSs, have evolved an editing domain to ensure the correct amino acid is ligated to its tRNA [13,14]. This domain is critical for aaRSs that must distinguish their cognate amino acid from structurally similar amino acids like branched-chain amino acids. Leucyl-tRNA synthetases (LeuRSs) are examples of aaRSs that rely on their editing domains to limit misaminoacylation of tRNA with near-cognate amino acids and maintain the fidelity of the genetic code
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