Huang Z, Guengerich F P, Kaminsky L S
Department of Environmental Health and Toxicology, School of Public Health, University at Albany, SUNY, NY 12201-0509, USA.
Carcinogenesis. 1998 May;19(5):867-72. doi: 10.1093/carcin/19.5.867.
The cytochrome P450 (P450) enzymes that catalyse metabolism of the estrogen, estrone (E1), to the putative carcinogen 16alpha-hydroxy E1 (16alpha-OHE1) in humans were determined. The potential of the most abundant circulating form of estrogen, estrone 3-sulfate (E1S), to be the substrate was also investigated. Human liver microsomal sulfatases convert E1S to E1, an essential prerequisite for formation of 16alpha-OHE1 from added E1S in this system. E1 metabolism to 16alpha-OHE1 in a panel of 15 human liver microsomal preparations correlated with total P450 concentrations (r2 = 0.63) and with activities associated with P450 forms CYP3A4 and 3A5 (r2 = 0.72). E1 16alpha-hydroxylase activity in human liver microsomes was inhibited by 75% by monoclonal anti human CYP3A4/5 antibodies at 4 mg antibody/nmol total P450, and by troleandomycin, a specific CYP3A4/5 inhibitor. Rates of E1 metabolism to 16alpha-OHE1 were 1.6-fold higher when E1 was generated in situ from E1S than when E1 was added. Microsomal preparations of cDNA expressed CYP3A4 or 3A5, with NADPH-P450-reductase co-expressed, both metabolized E1 to 16alpha-OHE1, and added cytochrome b5 increased the rates 5.1- and 7.5-fold, respectively. In these systems rates of E1 metabolism to 16alpha-OHE1 were 2.8-fold higher when E1 was generated in situ from E1S than when E1 was added. Kinetic values for E1 metabolism to 16alpha-OHE1 by human liver microsomes and for the expressed CYP3A4 system were Km 154 and 172 microM, respectively, and Vmax 238 pmol/min/nmol total P450 and 1050 pmol/min/nmol CYP3A4, respectively. Thus, formation of the putative carcinogen 16alpha-OHE1 is catalysed by CYP3A4 and 3A5 and stimulated by cytochrome b5. E1S is not a substrate but formation of E1 from E1S in situ stimulates formation of 16alpha-OHE1, possibly because E1S is more water soluble and in situ generation of E1 provides for facilitated exposure of E1 to the P450 substrate binding sites. Blocking of the pathway of E1 to 16alpha-OHE1 could provide a therapeutic approach for diminishing the risk of estrogen dependent breast cancer.
确定了在人体内催化雌激素雌酮（E1）代谢为推定致癌物16α-羟基E1（16α-OHE1）的细胞色素P450（P450）酶。还研究了雌激素最丰富的循环形式硫酸雌酮3-硫酸酯（E1S）作为底物的可能性。人肝微粒体硫酸酯酶将E1S转化为E1，这是在该系统中由添加的E1S形成16α-OHE1的必要前提。在一组15种人肝微粒体制剂中，E1代谢为16α-OHE1与总P450浓度相关（r2 = 0.63），并与P450形式CYP3A4和3A5相关的活性相关（r2 = 0.72）。人肝微粒体中的E1 16α-羟化酶活性在4 mg抗体/nmol总P450时被单克隆抗人CYP3A4/5抗体抑制75%，并被特异性CYP3A4/5抑制剂三乙酰竹桃霉素抑制。当E1由E1S原位生成时，E1代谢为16α-OHE1的速率比添加E1时高1.6倍。共表达NADPH-P450还原酶的cDNA表达的CYP3A4或CYP3A5的微粒体制剂均将E1代谢为16α-OHE1，添加细胞色素b5分别使速率提高5.1倍和7.5倍。在这些系统中，当E1由E1S原位生成时，E1代谢为16α-OHE1的速率比添加E1时高2.8倍。人肝微粒体和表达的CYP3A4系统将E1代谢为16α-OHE1的动力学值分别为Km 154和172 μM，Vmax分别为238 pmol/min/nmol总P450和1050 pmol/min/nmol CYP3A4。因此，可以推定致癌物16α-OHE1的形成由CYP3A4和CYP3A5催化，并受到细胞色素b5的刺激。E1S不是底物，但E1S原位生成E1刺激了16α-OHE1的形成，可能是因为E1S更易溶于水，E1的原位生成便于E1暴露于P450底物结合位点。阻断E1转化为16α-OHE1的途径可能为降低雌激素依赖性乳腺癌风险提供一种治疗方法。
