Abstract 712: Characterization of exemestane metabolic pathways
Authors: Sun Dongxiao , Liu Ying , McCloskey Diane E. , Lazarus Philip 
Published: June 2012

Abstract
The current standard of care for early-stage ER+ breast cancer in post-menopausal women includes endocrine therapy with aromatase inhibitors (AIs) alone or sequentially following treatment with tamoxifen (TAM). Clinical trials indicate that AIs, including exemestane (EXE), are superior to TAM as first-line therapy for postmenopausal women with metastatic breast cancer and that AIs are associated with longer disease-free survival than therapy with TAM alone in adjuvant therapy. Previous studies demonstrated that UGT2B17 was the major enzyme involved in the glucuronidation of the major EXE metabolite, 17-dihydroexemestane (17-DHE), and that human liver specimens from subjects homozygous for the UGT2B17 deletion exhibited a ∼20-fold decrease in 17-DHE-glucuronide formation. The purpose of this study was to fully characterize the pathways involved in EXE metabolism. In assays that included a NADPH-regenerating system, the cytosolic and microsomal fractions of normal human liver were screened using ultra performance liquid chromatography-mass spectrometry (UPLC-MS/MS). In human liver cytosols (HLC), 17-DHE was the predominant metabolite. Using purified cytosolic enzyme, AKR1C3 was shown to exhibit significant 17-DHE formation activity. In human liver microsomes (HLM), in addition to the major metabolite 17-DHE, at least 4 other EXE metabolites (EMs) were observed: 6α/β,17β-dihydroxy-6α/β-hydroxymethylandrosta-1,4-diene-3-one (EMI), 6α/β-hydroxy-6α/β-hydroxymethyl-androsta-1,4-diene-3,17-dione (EMII), 17β-hydroxy-6-hydroxymtheylandrosta-1,4,6-triene-3-one (EMIII) and 6-hydroxymethyl-androsta-1,4,6-triene-3-17-dione (EMIV). The relative content of each metabolite varied significantly between HLM from different individuals. Among the 4 metabolites, EMIV was the major primary metabolite formed directly from EXE while EMIII was a primary metabolite of 17-DHE. Using EMIII and EMIV as substrates, EMI and EMII were shown to be secondary metabolites formed from EMIII and EMIV, respectively. When UDPGA was added to EXE or 17-DHE metabolism assays, 17-DHE-glucuronide was the only glucuronide formed from HLM; as expected, no glucuronides were observed when HLC were examined. Using microsomes from CYP450-over-expressing human lymphoblast cells, CYPs 1A1, 2A6, 2B6, 2C9, 2C19, 2D6, 2E1, 3A4 and 3A5 all exhibited activity against EXE to form EMIV and 17-DHE, and exhibited activity against 17-DHE to form EMIII. In addition to the above enzymes, CYP4A11 also exhibited metabolizing activity to form 17-DHE from EXE and EMIII from 17-DHE. CYPs 1A2 and 2A6 were found to metabolize EMIV to EMII, whereas CYP1A1 and 2C19 were found to metabolize EMIII to EMI. These data suggest that several enzymes contribute to EXE metabolism. Kinetic analysis will help better assess the importance of each to EXE metabolism and their potential role in EXE pharmacogenetics.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 712. doi:10.1158/1538-7445.AM2011-712