Anti-Mycobacterial Activity of Bacterial Topoisomerase Inhibitors with Dioxygenated Linkers

Developing new classes of drugs that are active against infections caused by <i>Mycobacterium tuberculosis</i> is a priority for treating and managing this deadly disease. Here, we describe screening a small library of 20 DNA gyrase inhibitors and identifying new lead compounds. Three structurally diverse analogues were identified with minimal inhibitory concentrations of 0.125 μg/mL against both drug-susceptible and drug-resistant strains of <i>M. tuberculosis</i>. These lead compounds also demonstrated antitubercular activity in ex vivo studies using infected THP-1 macrophages with minimal cytotoxicity in THP-1, HeLa, and HepG2 cells (IC₅₀ ≥ 128 μg/mL). The molecular target of the lead compounds was validated through biochemical studies of select analogues with purified <i>M. tuberculosis</i> gyrase and the generation of resistant mutants. The lead compounds were assessed in combination with bedaquiline and pretomanid to determine the clinical potential, and the select lead (<b>158</b>) demonstrated in vivo efficacy in an acute model of TB infection in mice, reducing the lung bacterial burden by approximately 3 log₁₀ versus untreated control mice. The advancement of DNA gyrase inhibitors expands the field of innovative therapies for tuberculosis and may offer an alternative to fluoroquinolones in future therapeutic regimens.