Aminopyrazinamides: Novel and Specific GyrB Inhibitors that Kill Replicating and Nonreplicating <i>Mycobacterium tuberculosis</i>

Aminopyrazinamides originated from a high throughput screen targeting the <i>Mycobacterium smegmatis</i> (Msm) GyrB ATPase. This series displays chemical tractability, robust structure–activity relationship, and potent antitubercular activity. The crystal structure of Msm GyrB in complex with one of the aminopyrazinamides revealed promising attributes of specificity against other broad spectrum pathogens and selectivity against eukaryotic kinases due to novel interactions at hydrophobic pocket, unlike other known GyrB inhibitors. The aminopyrazinamides display excellent mycobacterial kill under <i>in vitro</i>, intracellular, and hypoxic conditions

Methyl-Thiazoles: A Novel Mode of Inhibition with the Potential to Develop Novel Inhibitors Targeting InhA in Mycobacterium tuberculosis

InhA is a well validated Mycobacterium tuberculosis (Mtb) target as evidenced by the clinical success of isoniazid. Translating enzyme inhibition to bacterial cidality by targeting the fatty acid substrate site of InhA remains a daunting challenge. The recent disclosure of a methyl-thiazole series demonstrates that bacterial cidality can be achieved with potent enzyme inhibition and appropriate physicochemical properties. In this study, we report the molecular mode of action of a lead methyl-thiazole, along with analogues with improved CYP inhibition profile. We have identified a novel mechanism of InhA inhibition characterized by a hitherto unreported “Y158-out” inhibitor-bound conformation of the protein that accommodates a neutrally charged “warhead”. An additional novel hydrophilic interaction with protein residue M98 allows the incorporation of favorable physicochemical properties for cellular activity. Notably, the methyl-thiazole prefers the NADH-bound form of the enzyme with a <i>K</i>_d of ∼13.7 nM, as against the NAD⁺-bound form of the enzyme

Structure Guided Lead Generation for <i>M. tuberculosis</i> Thymidylate Kinase (Mtb TMK): Discovery of 3‑Cyanopyridone and 1,6-Naphthyridin-2-one as Potent Inhibitors

<i>M. tuberculosis</i> thymidylate kinase (Mtb TMK) has been shown in vitro to be an essential enzyme in DNA synthesis. In order to identify novel leads for Mtb TMK, we performed a high throughput biochemical screen and an NMR based fragment screen through which we discovered two novel classes of inhibitors, 3-cyanopyridones and 1,6-naphthyridin-2-ones, respectively. We describe three cyanopyridone subseries that arose during our hit to lead campaign, along with cocrystal structures of representatives with Mtb TMK. Structure aided optimization of the cyanopyridones led to single digit nanomolar inhibitors of Mtb TMK. Fragment based lead generation, augmented by crystal structures and the SAR from the cyanopyridones, enabled us to drive the potency of our 1,6-naphthyridin-2-one fragment hit from 500 μM to 200 nM while simultaneously improving the ligand efficiency. Cyanopyridone derivatives containing sulfoxides and sulfones showed cellular activity against <i>M. tuberculosis</i>. To the best of our knowledge, these compounds are the first reports of non-thymidine-like inhibitors of Mtb TMK