In Vitro Characterization of Ertugliflozin Metabolism by UDP-Glucuronosyltransferase and Cytochrome P450 Enzymes

Ertugliflozin is primarily cleared through UDP-glucurosyltransferase (UGT)-mediated metabolism (86%) with minor oxidative clearance (12%). In vitro phenotyping involved enzyme kinetic characterization of UGTs or cytochrome P450 enzymes catalyzing formation of the major 3-O-beta-glucuronide (M5c) and minor 2-O-beta-glucuronide (M5a), monohydroxylated ertugliflozin (M1 and M3), and des-ethyl ertugliflozin (M2) metabolites in human liver microsomes (HLMs). Fractional clearance (f(CL)) from HLM intrinsic clearance (CLint) indicated a major role for glucuronidation (f(CL) 0.96; CLint 37 mu l/min per milligram) versus oxidative metabolism (f(CL) 0.04; CLint 1.64 mu l/min per milligram). Substrate concentration at half-maximal velocity (K-m), maximal rate of metabolism (V-max), and CLint for M5c and M5a formation were 10.8 mu M, 375 pmol/min per milligram, and 34.7 pl/min per milligram and 41.7 mu M, 94.9 pmol/min per milligram, and 2.28 pl/min per milligram, respectively. Inhibition of HLM CLint with 10 mu M digoxin or tranilast (UGT1A9) and 3 mu M 16 beta-phenyllongifolol (UGT2B7/UGT2B4) resulted in fraction metabolism (f(m)) estimates of 0.81 and 0.19 for UGT1A9 and UGT2B7/UGT2B4, respectively. Relative activity factor scaling of recombinant enzyme kinetics provided comparable f(m) for UGT1A9 (0.86) and UGT2B7 (0.14). K-m and V-max for M1, M2, and M3 formation ranged 73.0-93.0 OA and 24.3-116 pmol/min per milligram, respectively, and was inhibited by ketoconazole (M1, M2, and M3) and montelukast (M2). In summary, ertugliflozin metabolism in HLMs was primarily mediated by UGT1A9 (78%) with minor contributions from UGT2B7/UGT2B4 (18%), CYP3A4 (3.4%), CYP3A5 (0.4%), and CYP2C8 (0.16%). Considering higher ertugliflozin oxidative metabolism (f(CL) 0.12) obtained from human mass balance, human systemic clearance is expected to be mediated by UGT1A9 (70%), UGT2B7/UGT2B4 (16%), CYP3A4 (10%), CYP3A5 (1.2%), CYP2C8 (0.5%), and renal elimination (2%). SIGNIFICANCE STATEMENT This manuscript describes the use of orthogonal approaches (i.e., enzyme kinetics, chemical inhibitors, and recombinant enzymes) to characterize the fraction of ertugliflozin metabolism through various UDP-glucuronosyltransferase (UGT) and cytochrome P450 (CYP) enzyme-mediated pathways. Phenotyping approaches routinely used to characterize CYP hepatic fractional metabolism (f(m)) to estimate specific enzymes contributing to overall systemic clearance were similarly applied for UGT-mediated metabolism. Defining the in vitro metabolic disposition and f(m) for ertugliflozin allows risk assessment when considering potential victim-based drug-drug interactions perpetrated by coadministered drugs.