Induction of UDP-Glucuronosyltransferase 2B15 Gene Expression by the Major Active Metabolites of Tamoxifen, 4-Hydroxytamoxifen and Endoxifen, in Breast Cancer Cells

Abstract

We previously reported upregulation of UGT2B15 by 17b-estradiol in breast cancer MCF7 cells via binding of the estrogen receptor a (ERa) to an estrogen response unit (ERU) in the proximal UGT2B15 promoter. In the present study, we show that this ERa-mediated upregulation was significantly reduced by two ER antagonists (fulvestrant and raloxifene) but was not affected by a third ER antagonist, 4-hydroxytamoxifen (4-OHTAM), a major active tamoxifen (TAM) metabolite. Furthermore, we found that, similar to 17b-estradiol, 4-OHTAM and endoxifen (another major active TAM metabolite) elevated UGT2B15 mRNA levels, and that this stimulation was significantly abrogated by fulvestrant. Further experiments using 4-OHTAM revealed a critical role for ERa in this regulation. Specifically; knockdown of ERa expression by anti-ERa small interfering RNA reduced the 4-OHTAM-mediated induction of UGT2B15 expression; 4-OHTAM activated the wild-type but not the ERU-mutated UGT2B15 promoter; and chromatin immunoprecipitation assays showed increased ERa occupancy at the UGT2B15 ERU in MCF7 cells upon exposure to 4-OHTAM. Together, these data indicate that both 17b-estradiol and the antiestrogen 4-OHTAM upregulate UGT2B15 in MCF7 cells via the same ERa-signaling pathway. This is consistent with previous observations that both 17b-estradiol and TAM upregulate a common set of genes in MCF7 cells via the ER-signaling pathway. As 4-OHTAM is a UGT2B15 substrate, the upregulation of UGT2B15 by 4-OHTAM in target breast cancer cells is likely to enhance local metabolism and inactivation of 4-OHTAM within the tumor. This represents a potential mechanism that may reduce TAM therapeutic efficacy or even contribute to the development of acquired TAM resistance.

Emerging roles for UDP-glucuronosyltransferases in drug resistance and cancer progression

“…This is the case for UGT2B7 induced by several anti-cancer agents in liver cell models, for UGT2B15 induced by tamoxifen and UGT2B17 induced by exemestane in breast cancer-cell models, as well as for UGT1A4 induced by fulvestrant in breast cancer and liver cell models. [42][43][44][45][46] The notion that induction of UGT expression may lead to acquired drug resistance is further supported by the following studies. Breast cancer-cell models that were rendered resistant to methotrexate by prolonged exposure to the drug also displayed enhanced expression of several UGT1As-particularly UGT1A6-as well as enhanced glucuronidation activity.…”

Exemestane and Its Active Metabolite 17-Hydroexemestane Induce UDP-Glucuronosyltransferase (UGT) 2B17 Expression in Breast Cancer Cells

“…We recently showed that UGT2B15 can be induced by tamoxifen and its active metabolite 4-OH tamoxifen via the ERU (Chanawong et al, 2015). Because UGT2B15 glucuronidates and inactivates 4-OH-tamoxifen, its intratumoral induction by tamoxifen generates a potential regulatory feedback loop that might impact on therapeutic efficacy and promote the acquisition of drug resistance (Chanawong et al, Fig.…”

“…Human UGT2B17 supersomes (In Vitro Technologies, Noble Park North, VIC, Australia) were included as positive controls in each assay. Chromatography and quantification of glucuronide band intensities were conducted as previously reported (Chanawong et al, 2015;Wijayakumara et al, 2015). Fold induction of glucuronidation activity in drugtreated cells was calculated relative to vehicle-treated cells.…”

“…Immunosignals were detected with the SuperSignalWest Pico Chemiluminescent kit (Thermo-Fisher Scientific, Waltham, MA) and an ImageQuant LAS 4000 luminescent image analyzer (GE Healthcare, Chalfont St. Giles, United Kingdom). Quantitation of band intensity and background subtraction were performed using MultiGauge version 3.0 (Fujifilm Corporation, Tokyo, Japan) as described elsewhere (Chanawong et al, 2015).…”

Glutamic acid ameliorates estrogen deficiency–induced menopausal-like symptoms in ovariectomized mice

“…G6649; Sigma Chemical Co) and fulvestrant (no. I4409; Sigma Chemical Co) were dissolved in DMSO and prepared at a dose of 1 μmol/L, respectively, according to previous reports [22,23].…”