Synthesis and CYP24A1 inhibitory activity of N-(2-(1H-imidazol-1-yl)-2-phenylethyl)arylamides
Graphical abstract
A series of N-(2-(1H-imidazol-1-yl)-2-phenylethyl)arylamides were prepared and evaluated for their inhibitory activity against human cytochrome P450C24A1 (CYP24A1) hydroxylase. The styryl derivative (11c) displayed enhanced activity (IC50 = 0.3 μM) compared with the standard ketoconazole, providing a useful lead.
Introduction
Calcitriol (1α,25-(OH)2D3), the biologically active metabolite of vitamin D binds to the vitamin D receptor (VDR), which mediates tissue-specific effects including enhancement of intestinal calcium transport, maintenance of skeletal health and epidermal integrity and immune cell regulation (e.g., macrophages).1 Replacement calcitriol therapy has been used in chronic kidney disease, hyperproliferative skin diseases and osteoporosis, with clinical trials for various forms of cancer and autoimmune conditions ongoing.2, 3 Although the potential of vitamin D as an antiproliferative drug has been realised in psoriasis and parathyroid cell hyperplasia, an anticancer treatment incorporating vitamin D remains a challenge owing to increased drug resistance.3 Evidence is growing that this resistance is caused by up-regulation of the cytochrome P450 enzyme, CYP24A1, in tumours resulting in accelerated metabolism of calcitriol and derivatives.4, 5 Inhibitors of CYP24A1 are expected to extend the half-life of calcitriol thereby enhancing endogenous levels of calcitriol, or vitamin D analogues, which may result in stabilized CYP24A1 and enhanced stability of vitamin D compounds. Small molecule inhibitors of CYP24A1 have been described by our group6, 7 and others,8, 9, 10, 11 with the non-specific P450 inhibitor ketoconazole8 (Fig. 1) used as a standard for comparison. The most promising CYP24A1 inhibitors described to date are VID-4009 (IC50 = 15.1 nM compared with ketoconazole IC50 = 126 nM in human epidermal keratinocytes) and the vitamin D derived sulfoximes10 (IC50 = 7.4 nM compared with ketoconazole IC50 = 312 nM in V79-CYP24A1 recombinant cell line expressing human CYP24A1), which are based on the structure of the natural substrate, calcitriol.
Here we report the synthesis and inhibitory activity of a series of N-(2-(1H-imidazol-1-yl)-2-phenylethyl)arylamides. The scaffold of the benzofuran series (7) were based on VID-400, while the phenylmethyl (11b) and styryl (11c) derivatives combined pharmacophores of VID-400 and calcitriol.
Section snippets
Chemistry
The synthesis of N2-[2-phenyl-2-(1H-imidazoyl)ethyl]-benzo[b]furan-2-carboxamide derivatives (7a–e), was achieved using a five-step reaction (Scheme 1) and involved a modification of the procedure described by Schuster and Egger12 and Moenius et al.13
The substituted ethyl benzo[b]furan-2-carboxylates (2) were prepared by the reaction of the appropriate salicylaldehyde (1) with ethyl bromoacetate in anhydrous DMF at 130 °C for 6 h according to the methodology previously described.14 After
CYP24A1 inhibitory activity
The imidazole derivatives 7a–e and 11a–c were evaluated for their calcitriol metabolism (CYP24A1) inhibitory activity using a cell based assay employing a recombinant cell line expressing human CYP24A1 enzyme (V79-CYP24),18 using radiolabelled [3H-1β]-calcitriol as the substrate and ketoconazole as the standard for comparison. In the benzofuran series (7), the 4-fluorophenyl (7d) and unsubstituted (7a) derivatives were the most potent (IC50 = 2.8 and 4.0 μM, respectively), with introduction of the
Docking
To investigate the possible binding mode of this series of compounds, we have performed a set of molecular docking simulations, using our model of the CYP24A1.19 In the benzofuran analogues series (Fig. 2) it is possible to observe how the imidazole N3 complexes the Haem–Fe, while the phenyl ring is placed in a small hydrophobic pocked created by Ile242, Val391, Thr330, and Ala326. The benzofuran ring is placed in a hydrophobic channel and it establishes a series of interactions with Ile149,
Materials and methods: chemistry
1H and 13C NMR spectra were recorded with a Brucker Avance DPX500 spectrometer operating at 500 and 125 MHz, with Me4Si as internal standard. Mass spectra were determined by the EPSRC mass spectrometry centre (Swansea, UK). Microanalyses were determined by Medac Ltd (Surrey, UK). Flash column chromatography was performed with silica gel 60 (230–400 mesh) (Merck) and TLC was carried out on precoated silica plates (kiesel gel 60 F254, BDH). Melting points were determined on an electrothermal
Acknowledgements
For A.S.A. and M.S.G. we acknowledge the Embassy of the Arab Republic of Egypt for the award of a Ph.D. scholarship. For S.W.Y. we would like to acknowledge the ORS Awards Scheme for a United Kingdom Scholarship.
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Cited by (18)
Synthesis, molecular modelling and CYP24A1 inhibitory activity of novel of (E)-N-(2-(1H-imidazol-1-yl)-2-(phenylethyl)-3/4-styrylbenzamides
2017, Bioorganic and Medicinal ChemistryCitation Excerpt :To investigate the possible binding mode of this series of compounds, we have performed a set of molecular docking simulations, using our model of CYP24A1.24,30 We have previously shown the importance of the styrylphenyl structure for inhibitory activity and that the imidazole was essential for metal-ligand interaction with the haem group of the enzyme.23–25 Compounds 11a–11f have electron withdrawing and electron donating substituents on the phenyl ring.
Novel styryl-indoles as small molecule inhibitors of 25-hydroxyvitamin D-24-hydroxylase (CYP24A1): Synthesis and biological evaluation
2014, European Journal of Medicinal ChemistryCitation Excerpt :Our previous research on non-vitamin D CYP24A1 inhibitors [20–22] indicated the importance of a hydrophobic group or chain to allow complete fill of the hydrophobic channel of the CYP24A1 enzyme active site. In the development of the azole inhibitors, a styryl group was found to be optimal for inhibitory activity [21] (Fig. 1) resulting in compounds with activity comparable with the standard inhibitor ketoconazole (Fig. 1). Combining the styryl and azole haem binding group with an indole group, to mimic the calcitriol C/D ring unit, resulted in the design of the herein described indole imidazole series.
25-Hydroxyvitamin D-24-hydroxylase (CYP24A1): Its important role in the degradation of vitamin D
2012, Archives of Biochemistry and Biophysics
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Present address: Medicinal Chemistry Department, Faculty of Pharmacy, Assiut University, Assiut, Egypt.