Application of a Deuterium Replacement Strategy to Modulate the Pharmacokinetics of 7-(3,5-dimethyl-1H-1,2,4-triazol-1-yl)-3-(4-methoxy-2-methylphenyl)-2,6-dimethylpyrazolo[5,1-b]oxazole, a Novel CRF1 Antagonist

Abstract

Deuterium isotope effects were evaluated as a strategy to optimize the pharmacokinetics of 7-(3,5-dimethyl-1H-1,2,4-triazol-1-yl)-3-(4-methoxy-2-methylphenyl)-2,6-dimethylpyrazolo [5,1-b]oxazole (NVS-CRF38), a novel corticotropin-releasing factor receptor 1 (CRF 1 ) antagonist. In an attempt to suppress O-demethylation of NVS-CRF38 without losing activity against the CRF 1 receptor, the protons at the site of metabolism were replaced with deuterium. For in vitro and in vivo studies, intrinsic primary isotope effects (K H /K D ) were determined by the ratio of intrinsic clearance (CL int ) obtained for NVS-CRF38 and deuterated NVS-CRF38. In vitro kinetic isotope effects (K H /K D ) were more pronounced when CL int values were calculated based on the rate of formation of the O-desmethyl metabolite (K H /K D ∼7) compared with the substrate depletion method (K H /K D ∼2). In vivo isotope effects were measured in rats after intravenous (1 mg/kg) and oral (10 mg/kg) administration. For both administration routes, isotope effects calculated from in vivo CL int corresponding to all biotransformation pathways were lower (K H /K D ∼2) compared with CL int values calculated from the O-demethylation reaction alone (K H /K D ∼7). Comparative metabolite identification studies were undertaken using rat and human microsomes to explore the potential for metabolic switching. As expected, a marked reduction of the O-demethylated metabolite was observed for NVS-CRF38; however, levels of NVS-CRF38's other metabolites increased, compensating to some extent for the isotope effect.

Twelfth-Position Deuteration of Nevirapine Reduces 12-Hydroxy-Nevirapine Formation and Nevirapine-Induced Hepatocyte Death

“…No statistically significant changes were observed in 2- or 3-OHNVP formation using 50 μM 12-D 3 NVP vs NVP (Figure B,C, respectively) or with any of the other NVP concentrations tested (5, 10, 25, 100, 200, and 400 μM, data not shown). We found these results as well as the lack of change observed for both 2- and 3-OHNVP production in hepatocyte medium intriguing, as metabolic switching, in which other drug metabolite levels increase in response to decreased metabolism at one position, has been known to occur with deuterium substitution. ,,− Our observation may be due to differences in P450 contributions to metabolism at these different positions of the NVP scaffold. Since we did not see a significant decrease in 12-OHNVP with inhibitors that reduced production of 2-OHNVP and 3-OHNVP, this suggests that the enzymes responsible for 2- and 3-OHNVP production (CYP3A4 and CYP2B6, respectively) do not play a measurable role in the conversion of NVP to 12-OHNVP.…”

Effect of <em>N</em>-methyl deuteration on metabolism and pharmacokinetics of enzalutamide

“…Therefore, inaccuracies caused by other compensative pathways could be alleviated using the parent drug depletion method as in vitro evaluation model. Stringer et al 12 observed similar results when evaluating deuterated NVS-CRF-38 in vitro and in vivo.…”

Deuterium isotope effects in drug pharmacokinetics II: Substrate-dependence of the reaction mechanism influences outcome for cytochrome P450 cleared drugs

“…Cytochrome P450 enzymes (CYP) are responsible for over 90% of all metabolic clearance of drugs and xenobiotics, and three quarters of these reactions are attributable to five CYP isoforms (1A2, 2C9, 2C19, 2D6, and 3A4) with CYP3A4 contributing approximately 27% to the metabolism of all marketed drugs [ 4 ]. Thus, any deuteration strategy must consider the complex reaction mechanisms of these enzymes that can confound a deuteration strategy leading to a failure to achieve significant systemic pharmacokinetic gain even when metabolism by these enzymes may be rate-limiting in systemic clearance [ 5 – 8 ]. Examples of such mechanistic complexity include: a) Differences in reaction mechanisms of C-H bond cleavages such as the N- and O- dealkylation reactions, where single electron transfer and hydrogen atom abstraction mechanisms can have substantial differences in the magnitude of their intrinsic deuterium isotope effect [ 9 , 10 ]; b) Deuterium-induced metabolic switching to proximal or distal non-deuterated sites [ 11 , 12 ] which is possibly due to multiple binding orientations of a molecule within an active site, or freedom for a bound molecule to “tumble” within an active site because of the large active site cavity of some CYP enzymes, such that oxidation at a non-deuterated site compensates for decreased metabolism at the deuterated site resulting in loss of an isotope effect on the intrinsic clearance and a redistribution of the relative abundance of metabolites; and c) A rate limiting release of product resulting in masking of the intrinsic deuterium isotope effect ( H k/ D k ) on the intrinsic clearance ( H V m /K m / D V m /K m ) [ 13 , 14 ].…”

“…Using virtual molecular docking with CYPs 3A4 and 2C19, metabolic profiles and intrinsic clearance isotope effects with human liver microsomes and recombinant CYPs 3A4 and 2C19 with deuterated versions of 1a and 2a , we demonstrate the mechanistic complexities of CYP-catalyzed reactions where the rate limiting step may be determined by the substrate under consideration. The two chemotypes examined also provide an understanding of how to address a deuteration strategy for new chemical entities, and helps explain the numerous reports where deuteration has been largely ineffective in substantially altering the in-vivo pharmacokinetics of some CYP-cleared compounds [ 5 – 8 ].…”