Synthesis, molecular modelling and CYP24A1 inhibitory activity of novel of (E)-N-(2-(1H-imidazol-1-yl)-2-(phenylethyl)-3/4-styrylbenzamides

CYP24A1 (25-hydroxyvitamin D-24-hydroxylase) is a useful enzyme target for a range of medical conditions including cancer, cardiovascular and autoimmune disease, which show elevated CYP24A1 levels and corresponding reduction of calcitriol (the biologically active form of vitamin D). A series of (E)-N-(2-(1H-imidazol-1-yl)-2-(phenylethyl)-3/4-styrylbenzamides have been synthesised using an efficient synthetic route and shown to be potent inhibitors of CYP24A1 (IC50 0.11–0.35 μM) compared with the standard ketoconazole. Molecular modelling using our CYP24A1 homology model showed the inhibitors to fill the hydrophobic binding site, forming key transition metal interaction between the imidazole nitrogen and the haem Fe3+ and multiple interactions with the active site amino acid residues

N-Cyclopropyl-(20R)-2-Methylene-19,26,27-trinor-25-aza-Vitamin D analogs and their uses

This invention discloses N-cyclopropyl-(20R)-2-methylene-19,26,27-trinor-25-aza-vitamin D analogs, and specifically N-cyclopropyl-(20R)-2-methylene-19,26,27-trinor-25-aza-1.alpha.-hydroxyvi- tamin D.sub.3 and pharmaceutical uses therefor. This compound exhibits relatively high binding activity and pronounced activity in arresting the proliferation of undifferentiated cells and inducing their differentiation to the monocyte thus evidencing use as an anti-cancer agent especially for the treatment or prevention of leukemia, colon cancer, breast cancer, skin cancer or prostate cancer

2alpha-Methyl and 2beta-Methyl Analogs of 19,26-Dinor-1alpha,25-Dihydroxyvitamin D3 and Their Uses

This invention discloses 2.alpha.-methyl and 2.beta.-methyl analogs of 19,26-dinor-1.alpha.,25-dihydroxyvitamin D.sub.3 and pharmaceutical uses therefor. These compounds exhibit in vitro biological activities evidencing use as an anti-cancer agent and for the treatment of skin diseases such as psoriasis as well as skin conditions such as wrinkles, slack skin, dry skin and insufficient sebum secretion. These compounds have little, if any, in vivo calcemic activity and therefore may be used to treat autoimmune disorders in humans as well as secondary hyperparathyroidism and renal osteodystrophy

Analysis of the binding sites of vitamin D 1α-hydroxylase (CYP27B1) and vitamin D 24-hydroxylase (CYP24A1) for the design of selective CYP24A1 inhibitors: homology modelling, molecular dynamics simulations and identification of key binding requirements

A homology model of human CYP27B1 was built using MOE and was further optimised by molecular dynamics simulations of the hCYP27B1 homology model and a hCYP27B1-SDZ-88357 complex. Docking results from the hCYP27B1-SDZ-88357 complex showed amino acids Arg107, Asn387 and Asp320 have an important role in binding interaction, with Asp320 part of the important acid-alcohol pair situated in the I-helix with the conserved sequence (A/G) GX (E/D) (T/S), which assumes an essential role in the binding of an oxygen molecule for catalysis. Additional docking experiments with selective hCYP27B1 or hCYP24A1 inhibitors using both the hCYP27B1 model and a triple mutant hCYP24A1 model provided further support for the importance of H-bonding interactions with the three identified active site amino acids. To confirm the role of Arg107, Asn387 and Asp320 in the active site of hCYP27B1 compounds were designed that would form H-bonding interactions, as determined from docking experiments with the hCYP27B1 model. Subsequent synthesis and CYP24A1 and CYP27B1 enzyme assays of the designed compounds 1a and 1b showed a ∼5-fold selectivity for CYP27B1 confirming the importance of Asp320 in particular and also Asn387 and Arg107 as important amino acids for CYP27B1 inhibitory activity

Novel styryl-indoles as small molecule inhibitors of 25-hydroxyvitamin D-24-hydroxylase (CYP24A1): Synthesis and biological evaluation

The synthesis of a series of imidazole styrylindoles and sulfonyl styrylindoles derivatives is described. Evaluation of binding affinity and inhibitory activity against CYP24A1 identified the imidazole styrylindoles as potent inhibitors with activity greater or comparable with the standard ketoconazole. Flexible alignment and docking studies of the inhibitors in the CYP24A1 enzyme active site confirmed that complete occupation of the vitamin D access tunnel is essential to inhibitory activity, allowing exposure to multiple hydrophobic binding interactions and optimal conformation for the interaction of the imidazole nitrogen lone pair and the active site haem

Small-molecule inhibitors of 25-hydroxyvitamin D-24-hydroxylase (CYP24A1): synthesis and biological evaluation

The synthesis of imidazole styrylbenzamide, tert-butyl styrylimidazole, and tert-butyl styrylsulfonate derivatives is described. Evaluation of binding affinity and inhibitory activity against CYP24A1 identified the imidazole styrylbenzamides as potent inhibitors of CYP24A1, having selectivity with respect to CYP27B1 comparable with or greater than that of the standard ketoconazole. Further evaluation of the 3,5-dimethoxy and 3,4,5-trimethoxy derivatives in chronic lymphocytic leukemia cells revealed that co-treatment of 1α,25-dihydroxyvitamin D3 plus inhibitor coordinately upregulated GADD45α and CDKN1A. Docking experiments on the inhibitors in the CYP24A1 enzyme active site suggest the compounds reach the active site through the vitamin D access tunnel and are exposed to multiple hydrophobic residues. The imidazole styrylbenzamides are optimally positioned to allow interaction of the imidazole with the heme, and, in the case of the methoxy derivatives, a hydrogen bond between the 3-methoxy group and Gln82 stabilizes the molecule in a favorable active conformation

Identification of the vitamin D receptor in various cells of the mouse kidney

The kidney is the major, if not sole, site for the production of 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3), the biologically active form of vitamin D that can stimulate calcium reabsorption in the kidney and may provide renoprotective benefits. The biological effects of 1,25(OH)2D3 are mediated through a nuclear hormone receptor, known as the vitamin D receptor (VDR). It is well accepted that the VDR is present in the distal renal convoluted tubule cells; however, whether VDR is present in other kidney cell types is uncertain. Using a highly specific and sensitive anti-VDR antibody, we determined its distribution in the mouse kidney by immunohistochemistry. Our results show that the VDR is not only present in the distal but is also found in the proximal tubules, but at 24-fold lower levels. The VDR was also found in the macula densa of the juxtaglomerular apparatus, glomerular parietal epithelial cells, and podocytes. In contrast, the VDR is either very low or absent in interstitial fibroblasts, glomerular mesangial cells, and juxtaglomerular cells. Thus, identification of VDR in the proximal tubule, macula densa, and podocytes suggests that 1,25(OH)2D3 plays a direct role in these cells under normal conditions