Benzimidazoles: Novel Mycobacterial Gyrase Inhibitors from Scaffold Morphing

Type II topoisomerases are well conserved across the bacterial species, and inhibition of DNA gyrase by fluoroquinolones has provided an attractive option for treatment of tuberculosis (TB). However, the emergence of fluoroquinolone-resistant strains of <i>Mycobacterium tuberculosis</i> (<i>Mtb</i>) poses a threat for its sustainability. A scaffold hopping approach using the binding mode of novel bacterial topoisomerase inhibitors (NBTIs) led to the identification of a novel class of benzimidazoles as DNA gyrase inhibitors with potent anti-TB activity. Docking of benzimidazoles to a NBTI bound crystal structure suggested that this class of compound makes key contacts in the enzyme active site similar to the reported NBTIs. This observation was further confirmed through the measurement of DNA gyrase inhibition, and activity against <i>Mtb</i> strains harboring mutations that confer resistance to aminopiperidines based NBTIs and <i>Mtb</i> strains resistant to moxifloxacin. Structure–activity relationship modification at the C-7 position of the left-hand side ring provided further avenue to improve hERG selectivity for this chemical series that has been the major challenges for NBTIs

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

Left-Hand Side Exploration of Novel Bacterial Topoisomerase Inhibitors to Improve Selectivity against hERG Binding

Structure–activity relationship (SAR) exploration on the left-hand side (LHS) of a novel class of bacterial topoisomerase inhibitors led to a significant improvement in the selectivity against hERG cardiac channel binding with concomitant potent antimycobacterial activity. Bulky polar substituents at the C-7 position of the naphthyridone ring did not disturb its positioning between two base pairs of DNA. Further optimization of the polar substituents on the LHS of the naphthyridone ring led to potent antimycobacterial activity (Mtb MIC = 0.06 μM) against <i>Mycobacterium tuberculosis</i> (Mtb). Additionally, this knowledge provided a robust SAR understanding to mitigate the hERG risk. This compound class inhibits Mtb DNA gyrase and retains its antimycobacterial activity against moxifloxacin-resistant strains of Mtb. Finally, we demonstrate <i>in vivo</i> proof of concept in an acute mouse model of TB following oral administration of compound <b>19</b>

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

Thiazolopyridine Ureas as Novel Antitubercular Agents Acting through Inhibition of DNA Gyrase B

A pharmacophore-based search led to the identification of thiazolopyridine ureas as a novel scaffold with antitubercular activity acting through inhibition of DNA Gyrase B (GyrB) ATPase. Evaluation of the binding mode of thiazolopyridines in a Mycobacterium tuberculosis (Mtb) GyrB homology model prompted exploration of the side chains at the thiazolopyridine ring C-5 position to access the ribose/solvent pocket. Potent compounds with GyrB IC₅₀ ≤ 1 nM and Mtb MIC ≤ 0.1 μM were obtained with certain combinations of side chains at the C-5 position and heterocycles at the C-6 position of the thiazolopyridine core. Substitutions at C-5 also enabled optimization of the physicochemical properties. Representative compounds were cocrystallized with Streptococcus pneumoniae (Spn) ParE; these confirmed the binding modes predicted by the homology model. The target link to GyrB was confirmed by genetic mapping of the mutations conferring resistance to thiazolopyridine ureas. The compounds are bactericidal in vitro and efficacious in vivo in an acute murine model of tuberculosis

Novel N‑Linked Aminopiperidine-Based Gyrase Inhibitors with Improved hERG and in Vivo Efficacy against Mycobacterium tuberculosis

DNA gyrase is a clinically validated target for developing drugs against Mycobacterium tuberculosis (Mtb). Despite the promise of fluoroquinolones (FQs) as anti-tuberculosis drugs, the prevalence of pre-existing resistance to FQs is likely to restrict their clinical value. We describe a novel class of N-linked aminopiperidinyl alkyl quinolones and naphthyridones that kills Mtb by inhibiting the DNA gyrase activity. The mechanism of inhibition of DNA gyrase was distinct from the fluoroquinolones, as shown by their ability to inhibit the growth of fluoroquinolone-resistant Mtb. Biochemical studies demonstrated this class to exert its action via single-strand cleavage rather than double-strand cleavage, as seen with fluoroquinolones. The compounds are highly bactericidal against extracellular as well as intracellular Mtb. Lead optimization resulted in the identification of potent compounds with improved oral bioavailability and reduced cardiac ion channel liability. Compounds from this series are efficacious in various murine models of tuberculosis

Azaindoles: Noncovalent DprE1 Inhibitors from Scaffold Morphing Efforts, Kill Mycobacterium tuberculosis and Are Efficacious <i>in Vivo</i>

We report 1,4-azaindoles as a new inhibitor class that kills Mycobacterium tuberculosis <i>in vitro</i> and demonstrates efficacy in mouse tuberculosis models. The series emerged from scaffold morphing efforts and was demonstrated to noncovalently inhibit decaprenylphosphoryl-β-d-ribose2′-epimerase (DprE1). With “drug-like” properties and no expectation of pre-existing resistance in the clinic, this chemical class has the potential to be developed as a therapy for drug-sensitive and drug-resistant tuberculosis

4‑Aminoquinolone Piperidine Amides: Noncovalent Inhibitors of DprE1 with Long Residence Time and Potent Antimycobacterial Activity

4-Aminoquinolone piperidine amides (AQs) were identified as a novel scaffold starting from a whole cell screen, with potent cidality on Mycobacterium tuberculosis (Mtb). Evaluation of the minimum inhibitory concentrations, followed by whole genome sequencing of mutants raised against AQs, identified decaprenylphosphoryl-β-d-ribose 2′-epimerase (DprE1) as the primary target responsible for the antitubercular activity. Mass spectrometry and enzyme kinetic studies indicated that AQs are noncovalent, reversible inhibitors of DprE1 with slow on rates and long residence times of ∼100 min on the enzyme. In general, AQs have excellent leadlike properties and good in vitro secondary pharmacology profile. Although the scaffold started off as a single active compound with moderate potency from the whole cell screen, structure–activity relationship optimization of the scaffold led to compounds with potent DprE1 inhibition (IC₅₀ < 10 nM) along with potent cellular activity (MIC = 60 nM) against Mtb