Synthesis, biological evaluation and computational studies of pyrazole derivatives as Mycobacterium tuberculosis CYP121A1 inhibitors

A series of imidazole and triazole diarylpyrazole derivatives were prepared using an efficient 5-step synthetic scheme and evaluated for binding affinity with Mycobacterium tuberculosis (Mtb) CYP121A1 and antimycobacterial activity against Mtb H37Rv. Antimycobacterial susceptibility was measured using the spot-culture growth inhibition assay (SPOTi): the imidazoles displayed minimum inhibitory concentration (MIC₉₀) in the range of 3.95-12.03 μg mL⁻¹ (10.07-33.19 μM) with 11f the most active, while the triazoles displayed MIC₉₀ in the range of 4.35-25.63 μg mL⁻¹ (11.88-70.53 μM) with 12b the most active. Assessment of binding affinity using UV-vis spectroscopy showed that for the imidazole series, the propyloxy (11f) and isopropyloxy (11h) derivatives of the 4-chloroaryl pyrazoles displayed Mtb CYP121A1 type II binding affinity with K_d 11.73 and 17.72 μM respectively compared with the natural substrate cYY (K_d 12.28 μM), while in the triazole series, only the methoxy substitution with the 4-chloroaryl pyrazole (12b) showed good type II Mtb CYP121A1 binding affinity (K_d 5.13 μM). Protein-detected 1D ¹⁹F-NMR spectroscopy as an orthogonal strategy was used to evaluate ligand binding independent of perturbations at the haem. For imidazole and triazole compounds, perturbations were more intense than cYY indicating tighter binding and confirming that ligand coordination occurs in the substrate-binding pocket despite very modest changes in UV-vis absorbance, consistent with computational studies and the demonstrated potential anti-tuberculosis properties of these compounds.