MOLECULAR BASIS OF ESTROGEN ACTION IN OVARIAN CANCER

目的是确定与雌激素（E）相对应的分子事件 卵巢癌的反应性生长和侵袭性。 而雌激素 受体（ER）在大多数卵巢癌中表达， E和抗雌激素的作用尚未研究。 拟议 研究可能有助于制定治疗策略， 诊断标志物。 为了确定E和抗雌激素的作用，我们将使用模型卵巢 癌细胞系在生长和分化中表现出不同的ER功能， 转录调控和肿瘤组织中， 受体（PR）表达似乎与ER表达解偶联。的 具体目的是：（1）利用模型细胞系， E和抗雌激素对侵袭力的影响，通过使用Matrigel侵袭试验 以及对蛋白酶、组织蛋白酶D和120kDa标记物的分泌的影响， 北方印迹、免疫沉淀和Western印迹。 （二） 鉴定卵巢癌细胞和肿瘤中的ER变体和突变 显示ER功能改变的组织。 我们将使用聚合酶链 反应（PCR）和寡核苷酸引物来扩增序列， 编码ER的已知功能结构域。 直接测序，化学 错配切割和单链构象多态性 PCR产物的分析将用于检测ER变体， 突变。 (3)确定ER磷酸化模式是否改变 在卵巢癌细胞中，它们对雌激素的反应不同， 免疫沉淀和Western印迹32P标记的ER。

The goal is to define molecular events that correspond to estrogen (E) responsive growth and invasiveness in ovarian cancer. While estrogen receptor (ER) is expressed in most ovarian carcinomas, the molecular effects of E and antiestrogens have not been investigated. The proposed studies may facilitate the development of treatment strategies and diagnostic markers. To define the effects of E and antiestrogens, we will use model ovarian carcinoma cell lines that exhibit varied ER function in growth and in transcriptional regulation and in tumor tissue where progesterone receptor (PR) expression appears to be uncoupled from ER expression. The specific aims are: (1) Use the model cell lines to determine the effects of E and antiestrogens on invasiveness by using a Matrigel invasion assay and on secretion of the protease, Cathepsin D and the 120kDa marker by Northern blotting, immunoprecipitations and Western blotting. (2) Identify ER variants and mutations in ovarian carcinoma cells and tumor tissue that exhibit altered ER function. We will use polymerase chain reaction, (PCR) and oligonucleotide primers to amplify sequences that encode known functional domains of ER. Direct sequencing, chemical mismatch cleavage and single stranded conformational polymorphism analysis of PCR products will be used to detect ER variants and mutations. (3) Determine whether ER phosphorylation patterns are altered in ovarian carcinoma cells that differ in their response to estrogen by immunoprecipitations and Western blotting 32P-labelled ER.