CARCINOGENICITY OF B RING UNSATURATED ESTROGENS

有关雌激素替代疗法(ERT)的主要担忧是长期使用会显著增加患乳腺癌的风险。ERT最常用的制剂是倍美力，这是一种主要由B环不饱和雌激素组成的制剂，包括马琳(Eq)和马豆素(Eqn)的结合形式。我们的初步研究表明，在几个表达细胞色素P4501B1(CYP1B1)的人乳腺来源细胞系中，Eqn的代谢受ah受体调节，到4-羟基代谢物。从这些4-羟基代谢物中衍生出的半喹酮和对苯二酚具有加合作用并导致自由基的产生，可能与癌症的发生有关。我们假设雌激素参与了致癌的启动和促进阶段，并且类固醇雌激素B环的芳香性增加了致癌效力。我们广泛的长期目标是确定类固醇雌激素，包括B环不饱和雌激素，Eq和Eqn，是否通过儿茶酚雌激素的代谢激活而致癌。我们的具体目标是：1)在一系列永生化的肿瘤和非肿瘤来源的人类乳腺细胞系中表征Eq和Eqn的代谢。Eq和Eqn的生物活化途径包括偶联物的水解、还原为17β-二氢形式和羟基化为邻苯二酚雌激素。2)以Eq、Eqn及其17α和17β二氢形式为底物，测定人细胞色素P450的细胞色素P450的合成活性。3)建立乳腺上皮细胞表达人CYP1B1的转基因小鼠系；4)检测Eq和Eqn对人CYP1B1转基因小鼠DNA损伤和乳腺肿瘤发生率的影响。本文描述的研究将提供有关人乳腺上皮细胞中人类酶代谢B-环不饱和雌激素的新结果，并可能提供支持Eq、Eqn和内源性代谢激活在启动人类乳腺癌变中的作用的机制数据。

The major concern regarding estrogen replacement therapy (ERT) is the significant increase in the risk of breast cancer that accompanies long-term use. The most commonly used formulation for ERT is Premarin, a preparation consisting largely of B-ring unsaturated estrogens including conjugated forms of equilin (Eq) and equilenin (Eqn). Our preliminary studies show Ah-receptor-regulated metabolism of Eqn to 4-hydroxylated metabolites in several human breast-derived cell lines expressing cytochrome P4501B1 (CYP1Bl). Semiquinones and quinones derived from these 4-hydroxy metabolites, which are adductive and lead to free radical production, may be involved in carcinogenesis. We hypothesize that estrogens are involved in both the initiation and promotion phases of carcinogenesis, and that aromaticity of the B-ring of steroidal estrogens increases carcinogenic potency. Our broad, long-term goal is to determine whether steroidal estrogens, including the B-ring unsaturated estrogens, Eq and Eqn, are carcinogenic through metabolic activation via catechol estrogens. 0ur Specific Aims are to: 1) Characterize Eq and Eqn metabolism in a series of immortalized tumor- and non-tumor-derived human breast-cell lines. Pathways of Eq and Eqn bioactivation involving hydrolysis of conjugates, reduction to 17beta-dihydro forms and hydroxyation to catechol estrogens will be investigated. 2) Determine the catechol synthetic activities of human cytochromes P450 of the CYP1, CYP2, and CYP3 families with Eq, Eqn, and their 17alpha- and 17beta-dihydro forms as substrates. 3) Establish transgenic mouse lines expressing human CYP1B1 in the mammary epithelium, 4) Determine the effects of treatment with Eq and Eqn on DNA damage and the incidence of mammary-gland tumors in human CYP1B1-transgenic mice. The studies described here will provide novel results regarding the metabolism of the B-ring unsaturated estrogens by human enzymes in breast epithelial cells, and may provide mechanistic data supporting a role of metabolic activation of Eq, Eqn, and endogenous in the initiation of carcinogenesis in the human breast.