Novel Aryl Substituted Pyrazoles as Small Molecule Inhibitors of Cytochrome P450 CYP121A1: Synthesis and Antimycobacterial Evaluation

Abstract

Three series of biarylpyrazole imidazole and triazoles are described, which vary in the linker between the biaryl pyrazole and imidazole/triazole group. The imidazole and triazole series with the short −CH2– linker displayed promising antimycobacterial activity, with the imidazole–CH2– series (7) showing low MIC values (6.25–25 μg/mL), which was also influenced by lipophilicity. Extending the linker to −C(O)NH(CH2)2– resulted in a loss of antimycobacterial activity. The binding affinity of the compounds with CYP121A1 was determined by UV–visible optical titrations with KD values of 2.63, 35.6, and 290 μM, respectively, for the tightest binding compounds 7e, 8b, and 13d from their respective series. Both binding affinity assays and docking studies of the CYP121A1 inhibitors suggest type II indirect binding through interstitial water molecules, with key binding residues Thr77, Val78, Val82, Val83, Met86, Ser237, Gln385, and Arg386, comparable with the binding interactions observed with fluconazole and the natural substrate dicyclotyrosine.

Design and Synthesis of Imidazole and Triazole Pyrazoles as Mycobacterium Tuberculosis CYP121A1 Inhibitors

“…Hansch analysis (Figure ) showed a clear correlation between the lipophilicity of the compounds and MIC, as previously observed for the pyrazole lead compounds . This increased lipophilicity may facilitate increased drug uptake across the Mtb lipid‐rich cell wall resulting in enhanced antimycobacterial activity.…”

“…Thionyl chloride was the chlorinating reagent used to convert the alcohols to the chlorides ( 9 ), different equivalents of thionyl chloride were used to optimize the reaction conditions (1, 4 and 10 equivalents), and the maximum yield was obtained when 10 equivalents of the reagent were used. Reaction of the chlorides ( 9 ) with either the potassium salt of imidazole or triazole, prepared in situ by treatment of imidazole or triazole with potassium carbonate in acetonitrile at 45 °C for 1 h, and overnight reflux at 70 °C gave the required final imidazole ( 10 a and 10 b ) and triazole ( 11 a and 11 b ) pyridyl‐pyrazole derivatives (Scheme ) in yields of 67, 54, 48 and 55 % respectively.…”

“…For CYP121A1, the typical low‐spin ligand free spectrum shows five characteristic peaks; a protein peak from the aromatic amino acid residues at 280 nm, a Soret peak at 416.5 nm, the β band at 538 nm, the α band at 565 nm, and small charge transfer band at ∼650 nm owing to a residual amount of high‐spin heme iron. A heme iron low‐spin (LS) to high‐spin (HS) type I shift occurs as cYY binds to CYP121A1, with the Soret band shifting from 416.5 to 393 nm (the cYY K D is 5.82±0.16 μM). Other features of note in the spectral titration are apparent isosbestic points at approximately 420 nm and 492 nm, small red shifts in the alpha and beta band maxima at ∼572 and 547 nm, and loss of the small high‐spin signal at ∼650 nm.…”

“…The requirement for two phenyl rings was also explored in Series 2 by preparation of a small series of imidazole/triazole derivatives, with one of the phenyl rings replaced with a methyl group (Figure ). Finally, our previous research showed a direct correlation between lipophilicity and MIC . Therefore in Series 3 one of the phenyl rings is further substituted with a range of alkyl substituted phenyl and biaryl groups to determine structure‐activity relationships (Figure ).…”

“…Our previous research identified derivatives of (1,3‐diphenyl)‐4‐((1,3‐imidazol‐1‐yl)methyl)‐1 H ‐pyrazole and 1‐((1,3‐diphenyl‐1 H ‐pyrazol‐4‐yl)methyl)‐1 H ‐triazole as inhibitors of CYP121A1 ( e. g . 1 , 2 and 3 , Figure ), representing lead compounds for further development. Diphenyl pyrazole derivatives were found to interact with the active site of CYP121A1 in a manner similar to fluconazole and the natural substrate cYY (Figure ), interacting indirectly with the heme via an interstitial water molecule (Figure ).…”