L-756423

Mycobacterium tuberculosis (Mtb) remains a major global health threat due to its survival under host-induced stress and the rise of drug-resistant strains. One critical factor contributing to this resilience is its ability to efficiently repair DNA damage. To survive double-stranded DNA breaks, Mtb utilizes two pathways: homologous recombination (HR) and non-homologous end joining (NHEJ). During dormancy, when HR is less active, Mtb predominantly relies on NHEJ, a pathway involving Ku and LigaseD. LigaseD catalyses DNA ligation through its ligase domain (LigDom), which comprises two subdomains: the NTase, harboring the ATP-binding site essential for adenylation, and the OB domain, which interacts with DNA. LigDom is highly conserved and exhibits low mutation rates in resistant strains, making it an attractive drug target. Using a structure-based drug repurposing approach, FDA-approved drugs were screened, identifying L-756423 as a potent inhibitor with a lowest binding free energy. Molecular dynamics simulation of the L-756423 bound LigDom complex reveals a stable domain structure with significant fluctuations in the interdomain region, suggesting movement in the OB domain that may support efficient binding. To validate these findings, Indinavir, a clinically approved HIV protease inhibitor with structural similarity to L-756423, was selected for experimental testing. Indinavir induces conformational flexibility in LigDom, and also competitively inhibits ATP binding thus blocking the adenylation. Functional microbiological assays confirmed that under oxidative stress, when NHEJ is critical, indinavir-treated cells displayed reduced survival, consistent with impaired DNA repair. These findings indicate that L-756423/Indinavir may hold therapeutic potential against latent tuberculosis and provide a rationale for further experimental investigation.