Inhibition of Testosterone Glucuronidation by Imatinib in Human Liver Microsomes Characterized for UGT2B17 Expression

Background and Objectives Glucuronidation is a major route of elimination for orally administered testosterone (T). Expression of UGT2B17 is highly variable and its gene deletion is associated with changes in T levels. We hypothesized that inhibition of UGT2B17 by drugs such as tyrosine kinase inhibitors could result in elevated T levels. Since high T levels (e.g., during T therapy) are associated with side effects, e.g., venous thromboembolism (1), the specific aims of this project were a) to determine the inhibition potential of imatinib for T‐glucuronidation in human liver microsomes (HLMs) identified as UGT2B17 expressers and non‐expressers, and b) to predict in vivo inter‐individual variability in imatinib‐T interaction. Method Protein abundance of T‐glucuronidation enzymes, UGT2B17 (major) and UGT2B15 (minor) was measured in HLMs (n=4) using a validated LC‐MS/MS proteomics method. T‐glucuronidation activity were measured in HLMs (0.1 mg/ml) by incubating T (5 μM) with varying concentrations of imatinib (0–100 μM) in presence of the cofactor, UDPGA (2.5 mM). Reactions were quenched by acetonitrile, centrifuged, and T‐glucuronide in supernatant was analyzed by LC‐MS/MS. IC 50 values of imatinib were determined in UGT2B17 expressers and non‐expressers. The in vivo fold change for T area under the curve (AUC) was predicted using a) the proteomics‐based individual fraction metabolized (f m ) data, b) imatinib steady‐state plasma concentration [I], and c) inhibitory constant (K i ) for imatinib. Results Protein abundance of UGT2B17 was highly variable across individuals, which led to corresponding variability in the estimated UGT2B17 f m for T‐glucuronidation (0.9, 0.82, 0.72 and 0.0). Imatinib was a potent inhibitor of T‐glucuronidation in high UGT2B17 expressers (IC 50 = 0.37 μM), but it was a weak inhibitor in the non‐expressers (IC 50 = 25.4 μM), where T is metabolized by UGT2B15 (Fig. 1). Weak inhibition of UGT2B15 by imatinib was also confirmed by repeating the inhibition assay using a UGT2B15 probe substrate, oxazepam. By integrating the individual f m and [I]/Ki (i.e., 5.1) data, inhibition of T‐glucuronidation by imatinib was predicted to alter the in vivo T AUC by 4, 3.2, 2.5, and 0 fold, respectively. Discussion Because inhibition of T‐glucuronidation by imatinib is primarily mediated by UGT2B17 in the liver, imatinib (at ~5 uM) could be used as a selective inhibitor with T as a positive control for in vitro UGT2B17 phenotyping assays. Potent inhibition of UGT2B17 by imatinib is also important in vivo as high UGT2B17 expressers could experience up to 4‐fold increase in the T AUC. Our data predicts that imatinib steady‐state concentrations, can lead to selective drug interactions (expressers vs. non‐expressers) with UGT2B17 substrates such as testosterone, vorinostat and hydroxy‐exemestane. Support or Funding Information Department of Pharmaceutics, University of Washington and NIH R01 HD081299 This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .