Structure‐Activity Relationship and Mode‐Of‐Action Studies Highlight 1‐(4‐Biphenylylmethyl)‐1H‐imidazole‐Derived Small Molecules as Potent CYP121 Inhibitors

Abstract

CYP121 of Mycobacterium tuberculosis (Mtb) is an essential target for the development of novel potent drugs against tuberculosis (TB). Besides known antifungal azoles, further compounds of the azole class were recently identified as CYP121 inhibitors with antimycobacterial activity. Herein, we report the screening of a similarity-oriented library based on the former hit compound, the evaluation of affinity toward CYP121, and activity against M. bovis BCG. The results enabled a comprehensive SAR study, which was extended through the synthesis of promising compounds and led to the identification of favorable features for affinity and/or activity and hit compounds with 2.7-fold improved potency. Mode of action studies show that the hit compounds inhibit substrate conversion and highlighted CYP121 as the main antimycobacterial target of our compounds. Exemplified complex crystal structures of CYP121 with three inhibitors reveal a common binding site. Engaging in both hydrophobic interactions as well as hydrogen bonding to the sixth iron ligand, our compounds block a solvent channel leading to the active site heme. Additionally, we report the first CYP inhibitors that are able to reduce the intracellular replication of M. bovis BCG in macrophages, emphasizing their potential as future drug candidates against TB.

Glycosylated Flavonoid Compounds as Potent CYP121 Inhibitors of Mycobacterium tuberculosis

“…Compound 2 , Catechin 5,7, -Di- O -Gallate, showed a −10.4 kcal/mol binding energy and formed stacking interaction with the ring of HIS343, as shown in ( Figure 1 and Figure S1 ). This residue is an experimentally annotated binding site residue that interacts with the heme group [ 21 ]. Moreover, like compound 1 , this compound was also involved in the formation of an H-bond with ASN74 to achieve greater binding stability.…”

“…Likewise, the (3S,6S)-3,6-bis (4-hydroxybenzyl) piperazine-2,5-dione compound is another strong inhibitor of the CYP121 protein (Kd of 10.5 µM) listed in the drug bank database [ 19 , 20 ]. Moreover, the mechanism of action of these inhibitors was deciphered as an interference with the substrate conversion for the CYP121 protein [ 21 , 22 ]. Furthermore, in silico approaches for rational drug design for CYP121 were demonstrated in several earlier studies.…”

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

“…The aldehydes (8) were reduced with NaBH 4 and the alcohols (9), obtained after extraction with EtOAc in yields of 68-98%, were converted to the chlorides (10) on treatment with SOCl 2 in CH 2 Cl 2 at 40 o C for 4 h. The crude chlorides (10) were obtained in very high yields (93-100%) except for the isopropyl/fluoride derivative (48%), and were used immediately in the next reaction, which involved the generation of the imidazole or triazole anion by reaction of imidazole or triazole with K 2 CO 3 in CH 3 CN at 45 o C for 1 h, followed by the addition of the chlorides and heating at 70 o C overnight (Scheme 1). The final imidazole (11) and triazole (12) products were obtained after purification by gradient column chromatography with generally good yields (Table 1). All compounds were evaluated for MIC 90 against Mtb H37Rv using the SPOTi assay as previously described.…”

“…4,5 CYP121A1 inhibitors with a range of structural scaffolds have been described by us [6][7][8][9] and others. [10][11][12] In all cases a basic nitrogen containing group with the potential to bind to the iron of the haem group is included, although indirect water-mediated binding with the haem is occasionally observed and/or binding of the nitrogen containing group with Gln395/Arg386, which would block access of cYY to the active site. [6][7][8][9][10][11][12] One of the more promising scaffolds is the diarylpyrazole series 6,9 with optimal binding observed for imidazole as the nitrogen containing binding group and substitution of one of the aryl rings with lipophilic alkyl groups (Figure 2).…”

“…[10][11][12] In all cases a basic nitrogen containing group with the potential to bind to the iron of the haem group is included, although indirect water-mediated binding with the haem is occasionally observed and/or binding of the nitrogen containing group with Gln395/Arg386, which would block access of cYY to the active site. [6][7][8][9][10][11][12] One of the more promising scaffolds is the diarylpyrazole series 6,9 with optimal binding observed for imidazole as the nitrogen containing binding group and substitution of one of the aryl rings with lipophilic alkyl groups (Figure 2). Presented here is a further design of these lead pyrazole derivatives to consider drug likeness, incorporating O-alkyl substitutions in one aryl ring to improve hydrophilic properties compared with the lead compounds, and the addition of a halide (F or Cl) in the second aryl ring for metabolic stability (Figure 2).…”