Relative Enzymatic Activity, Protein Stability, and Tissue Distribution of Human Steroid-Metabolizing UGT2B Subfamily Members**This work was supported by the Medical Research Council of Canada, the Fonds de la Recherche en Santé du Québec, and Endorecherche.

Abstract

Androgens and estrogens play major roles in cell differentiation, cell growth, and peptide secretion in steroid target tissues. In addition to the binding of these hormones to their receptors, formation and metabolism are important in the action of steroids. Metabolism of the potent steroid hormones includes glucuronidation, a major pathway of steroid elimination in liver and several steroid target tissues. Glucuronidation is catalyzed by UDP-glucuronosyltransferases (UGTs), which transfer the polar moiety from UDP-glucuronic acid to a wide variety of endogenous compounds, including steroid hormones. The UGT superfamily of enzymes is subdivided into two families, UGT1 and UGT2, on the basis of sequence homology. To date, six UGT2B proteins have been isolated, namely UGT2B4, UGT2B7, UGT2B10, UGT2B11, UGT2B15, and UGT2B17, all of which have been demonstrated to be active on steroid molecules, except for UGT2B10 and UGT2B11, for which no substrate was found. The relative activity of these enzymes on steroidal compounds remains unknown due to variable levels of UGT2B expression in different in vitro cell line models and various conditions of the enzymatic assays. Comparison of the glucuronidation rates of these enzymes requires a unique system for UGT2B protein expression, protein normalization, and enzymatic assays. In this study we have stably expressed UGT2B4, UGT2B7, UGT2B15, and UGT2B17 in the HK293 cell line, which is devoid of steroid UGT activity; characterized their kinetic properties relative to UGT protein expression; determined their transcript and protein stabilities; and established extensively their tissular distributions. UGT2B7 was demonstrated to glucuronidate estrogens, catechol estrogens, and androstane-3␣,17␤-diol more efficiently than any other human UGTB isoform. UGT2B15 and UGT2B17 showed similar glucuronidation activity for androstane-3␣,17␤-diol (30% lower than that of UGT2B7), whereas UGT2B17 demonstrated the highest activity for androsterone, testosterone, and dihydrotestosterone. UGT2B4 demonstrates reactivity toward 5␣-reduced androgens and catechol estrogens, but at a significantly lower level than UGT2B7, 2B15, and 2B17. Cycloheximide treatment of stably transfected HK293 cells demonstrated that the UGT2B17 protein is more labile than the other enzymes; the protein levels decrease after 1 h of treatment, whereas other UGT2B proteins were stable for at least 12 h. Treatment of stable cells with actinomycin D reveals that UGT2B transcripts are stable for 12 h, except for the UGT2B4 transcript, which was decreased by 50% after the 12-h incubation period. Tissue distribution of the UGT2B enzymes demonstrated that UGT2B isoforms are expressed in the liver as well as in several extrahepatic steroid target tissues, namely, kidney, breast, lung, and prostate. This study clearly demonstrates the relative activities and the major substrates of human steroid-metabolizing UGT2B enzymes, which are expressed in a wide variety of steroid target tissues. (Endocrinology 142: 778 -787, 2001)

Xenobiotic-Metabolizing Enzymes in Human Lung

“…No measurable expression of any UGT1A gene has been observed in human lung based on RT-PCR analyses [103]. UGT2B4, 2B7, 2B10, 2B11, 2B15, and 2B17 were identified in human lung [104]. It has been reported that beclomethasone dipropionate (BDP) stimulated the expression of mRNAs for UGT 2B4 and 2B11, and transporters such as multidrug resistance Pglycoprotein, multidrug resistance-associated protein 1 and 2 in cultured bronchial epithelial cells.…”

Xenobiotic Nuclear Receptor-Mediated Regulation of UDP-Glucuronosyltransferases

“…The UGT isoform expression is tissue-specific, which is reflected by the fact that each UGT family member is expressed in selective tissues, and each tissue harbors the expression of multiple UGT isoforms. UGT1A1, 1A3, 1A4, 1A6, 1A9, and all the UGT2B isoforms have been shown to be expressed in the liver [10,11], whereas the human intestine has been shown to express UGT1A1, 1A6, 1A8, 1A10, and UGT2B7 [10,12]. The mechanisms underlying the tissue-specific distribution of UGT isoforms, have not been fully understood.…”

Pilot study demonstrating potential association between breast cancer image‐based risk phenotypes and genomic biomarkers

“…Interestingly, these two enzymes are the ones with the highest activity on steroid hormones within this family. 47 The A allele of rs4148298, which was associated with higher UGT2B7 expression in liver, 35 was correlated with smaller power law β value, and thus lower mammographic density (Fig. 3).…”