The mibefradil derivative NNC55-0396, a specific T-type calcium channel antagonist, exhibits less CYP3A4 inhibition than mibefradil

A novel mibefradil derivative, NNC55-0396, designed to be hydrolysis-resistant, was shown to be a selective T-type Ca2+ channel inhibitor without L-type Ca2+ channel efficacy. However, its effects on cytochromes P450 (P450s) have not previously been examined. We investigated the inhibitory effects of NNC55-0396 toward seven major recombinant human P450s-CYP3A4, CYP2D6, CYP1A2, CYP2C9, CYP2C8, CYPC19, and CYP2E1 - and compared its effects with those of mibefradil and its hydrolyzed metabolite, Ro40-5966. Our results show that CYP3A4 and CYP2D6 are the two P450s most affected by mibefradil, Ro40-5966, and NNC55-0396. Mibefradil (IC50 = 33 +/- 3 nM, K-i = 23 +/- 0.5 nM) and Ro40-5966 (IC50 = 30 +/- 7.8 nM, K-i = 21 +/- 2.8 nM) have a 9- to 10-fold greater inhibitory activity toward recombinant CYP3A4 benzyloxy-4-trifluoromethylcoumarin-O- debenzylation activity than NNC55-0396 (IC50 = 300 +/- 30 nM, K-i = 210 +/- 6 nM). More dramatically, mibefradil (IC50 = 566 +/- 71 nM, K-i = 202 +/- 39 nM) shows 19-fold higher inhibition of CYP3A-associated testosterone 6 beta-hydroxylase activity in human liver microsomes compared with NNC55-0396 (IC50 = 11 +/- 1.1 mu M, K-i = 3.9 +/- 0.4 mu M). Loss of testosterone 6 beta-hydroxylase activity by recombinant CYP3A4 was shown to be time-and concentration-dependent with both compounds. However, NNC55-0396 (K-I = 3.87 mu M, K-inact = 0.061/min) is a much less potent mechanism-based inhibitor than mibefradil (K-I = 83 nM, K-inact = 0.048/min). In contrast, NNC55-0396 (IC50 = 29 +/- 1.2 nM, K-i = 2.8 +/- 0.3 nM) and Ro40-5966 (IC50 = 46 +/- 11 nM, K-i = 4.5 +/- 0.02 nM) have a 3- to 4-fold greater inhibitory activity toward recombinant CYP2D6 than mibefradil (IC50 = 129 +/- 21 nM, K-i = 12.7 +/- 0.9 nM). Our results suggest that NNC55-0396 could be a more favorable T-type Ca2+ antagonist than its parent compound, mibefradil, which was withdrawn from the market because of strong inhibition of CYP3A4.