乳腺癌中雌激素受体DNA结合功能区的检测及意义

Study of estrogen receptor DNA binding domain and its significance in patients with breast cancer

目的：发展检测雌激素受体（ＥＲ）ＤＮＡ结合功能区（ＤＢＤ）的方法，探讨其在内分泌治疗乳腺癌方面的临床意义．方法：根据ＥＲ特异地与雌激素应答元件（ＥＲＥ）相结合的原理，合成含有ＥＲＥ核心序列的ＤＮＡ片段，经３２Ｐ标记后作为探针．选取经激素结合法测定ＥＲ、ＰＲ（孕激素受体）同时阳性的１９例乳腺癌组织标本，制备乳腺癌组织和细胞株ＭＣＦ－７的蛋白抽提液．应用凝胶滞留法检测ＥＲＥ探针与抽提液中ＥＲ的结合能力作为反映ＥＲＤＢＤ功能的指标．结果：在制备的ＭＣＦ－７细胞蛋白抽提液中，放射自显影结果显示ＥＲ－ＥＲＥ复合物带（ＥＲ－ＥＲＥ＋），而在对照组结肠癌细胞中未出现此带（ＥＲ－ＥＲＥ－），应用竞争法发现此复合物带密度减弱．在１９例筛选的乳腺癌标本中，１３例ＥＲ－ＥＲＥ（＋），随访发现，１２例患者内分泌治疗效果好，３ａ内未见有复发转移灶；６例ＥＲ－ＥＲＥ（－），其中４例２ａ内出现肺部转移．结论：应用ＥＲＥ探针能直接检测ＥＲＤＢＤ功能，其检测结果可为临床乳腺癌患者内分泌治疗及其预后判断提供可靠依据．

AIM: To develop a method of detecting the effect of estrogen receptor (ER) DNA binding domain (DBD) and to investigate its clinic significance in endocrine therapy for breast cancer. METHODS: In the light of the mechanism that ER can specifically bind to its estrogen response element (ERE), we synthesized the DNA segment containing core sequence of ERE, which was then labeled with 32P]ATP to be used as a probe. Protein extracts were prepared from ERcontaining human breast cancer cell line MCF7 and 19 patients' frozen tissues of tumors which were confirmed both ER (+) and PR (+) by hormonebinding assay. Using gel retardation assay, ERERE complexes were detected, which were used to represent the function of ER DBD. RESULTS: The results of autoradiograph showed that there was a band intensity of ERERE complexes (ERERE+) in the human breast cancer cell line MCF7 while in the colon cancer cells no band intensity of ERERE complexes (ERERE-) was observed. It was also found that when unlabeled ERE into the binding reaction, the band intensity decreased. Our research revealed that in 12 of the 13 cancer cases with ERERE (+) there was no recurrence or metastasis within 3 years and 4 of the 6 cases with ERERE (-) had lung metastasis within 2 years. CONCLUSION: Application of ERE probe can directly evaluate the function of ER DBD the results of which can be used as reliable indexes in clinic endocrine therapy and prognosis estimation.