RETINOID INHIBITION OF BREAST CANCER GROWTH

类维生素A用作抗肿瘤剂或化学预防剂， 包括乳腺癌患者在内的临床试验。 我们发现 类维生素A只能抑制雌激素受体（ER）阳性细胞的生长， 细胞 我们现在发现乳腺癌细胞具有独特的 视黄酸核受体-α（RAR-α）的同种型， 必须通过雌激素来提高水平，以允许类维生素A抑制 增长 具体目的1：确定人类乳腺中是否存在ER阳性 癌活组织检查标本导致RAR和RXR mRNA水平增加。 ER阴性乳腺癌细胞系的生长难治性 抑制RA，并具有降低RAR-α mRNA水平，而ER- 阳性细胞系表达高水平的RAR-α mRNA特异性目的2： 证明RA通过RAR-alpha调节基因发挥着至关重要的作用 在RA抑制乳腺癌生长方面。 2A）确定是否 用A横切RA难治性ER阴性乳腺癌细胞 RAR-α表达载体和随后增加的RAR-α表达 α导致生长的RA抑制。2B）证明抑制 使用选择性RAR-α拮抗剂的RAR-α功能的结果 2C）选择性抑制野生型 利用RAR-α显性负突变体的RAR-α型功能 导致细胞对RA介导的生长抑制不敏感。 具体目标3：我们发现人乳腺癌细胞系表达一种独特的 受雌二醇转录调节的RAR-α亚型 在ER阳性细胞中。 我们将识别和描述RAR-α 在人乳腺癌细胞中表达同种型;分离物，序列 并在功能上表征了5 ′-上游调控区（ 启动子区）的RAR-α基因，并确定调节 雌激素介导的RAR-α基因调节的元件 表情 具体目标4：阐明 雌激素对人乳腺RAR-α基因转录调控 癌细胞 利用分离的启动子确定是否 雌二醇通过与启动子结合， 雌激素反应元件（ERE）或刺激或抑制的水平， 其他因素反过来又调节基因活性。 具体目标五： 研究乳腺癌中特定RAR-α亚型的mRNA水平 石蜡包埋的癌细胞及其周围间质 从患者获得的浸润性导管癌标本，和 将样本的ER状态与RAR-α mRNA的水平相关联。 具体目标6：确定RA是否也抑制体内生长 和/或ER转染的MDA-MB-231细胞的转移潜力。

Retinoids are employed as anti-neoplastic or chemopreventative agents in clinical trials including women with breast carcinoma. We found retinoids only inhibit the growth of estrogen receptor (ER)-positive cells. We have now found that breast carcinoma cells possess a unique isoform of the retinoic acid nuclear receptor-alpha (RAR-alpha) whose levels must be enhanced by estrogens to allow retinoids to inhibit growth. Specific Aim 1: Determine whether ER positivity in human breast carcinoma biopsy specimens results in increased RAR and RXR mRNA levels. ER negative breast carcinoma cell lines are refractory to growth inhibitory by RA, and possess decreased RAR-alpha mRNA levels, while ER- positive cell lines express high RAR-alpha mRNA level Specific Aim 2: Demonstrate that RA modulation of genes via RAR-alpha plays a vital role in RA inhibition of breast carcinoma growth. 2A) Determine if transection of RA-refractory ER-negative breast carcinoma cells with a RAR-alpha expression vector and subsequent increased expression of RAR- alpha results in RA-inhibition of growth. 2B) Demonstrate that inhibition of RAR-alpha function utilizing a selective RAR-alpha antagonist results in resistance to RA-inhibition of growth 2C) Selective inhibition of wild type RAR-alpha function utilizing a RAR-alpha dominant negative mutant results in cells refractory to RA-mediated inhibition of growth. Specific Aim 3: We found human breast cancer cell lines express a unique isoform of RAR-alpha which is transcriptionally regulated by estradiol in ER-positive cells. We will identify and characterize the RAR-alpha isoform(s) expressed in human breast carcinoma cells; isolate, sequence and functionally characterize the 5'-upstream regulatory region (the promoter region) of the RAR-alpha gene and identify the regulatory elements responsible for estrogen-mediated modulation of RAR-alpha gene expression. Specific Aim 4: Elucidate the mechanism(s) involved in estrogen regulation of RAR-alpha gene transcription in human breast carcinoma cells. Ascertain utilizing the isolated promoter whether estradiol enhances genes transcription by binding to the promoter via an estrogen response element (ERE) or stimulates or represses the levels of other factors which in turn modulate gene activity. Specific Aim 5: Study the mRNA levels of the specific RAR-alpha isoform(s) in breast carcinoma cells and the surrounding stroma in paraffin embedded infiltrating ductal carcinoma specimens obtained from patients, and correlate the ER-status of the specimen and the level of RAR-alpha mRNA. Specific Aim 6: Determine whether RA also inhibits the in vivo growth and/or metastatic potential of ER-transfected MDA-MB-231 cells.