Cloning of tissue-specific expressing genes in huma ovarian cancer cells and basic study of the adenovirus-mediaetd p16 gene therapy.

卵巢透明细胞腺癌是由于化疗耐药和/或早期淋巴结转移而预后不良的组织亚型。它有时会受到子宫内膜异位症和日本常见亚型的影响。另一方面，卵巢浆液性腺癌是一种常见的上皮性癌组织亚型，尽管其腹膜播散频繁，但对化疗仍高度敏感。我们尝试用差异显示技术筛选透明细胞腺癌中特异表达的基因，并将SP17基因作为候选基因。由于SP17基因的功能尚不清楚，因此将SP17基因全长插入腺病毒载体中，并感染浆液性腺癌细胞。作为进一步的验证性实验，将SP17基因的RNAi导入透明细胞腺癌中，并分析SP17基因的功能。在卵巢癌组织中进行MTS测定和细胞死亡检测 ...更多信息 cer细胞，有或没有治疗的紫杉醇，最有用的抗癌药物的卵巢癌。结果表明，SP17在卵巢透明细胞腺癌化疗耐药中起重要作用; P16 INK 4A基因是细胞周期蛋白依赖性激酶抑制剂，是肿瘤抑制基因之一，是细胞进入G1-S期的关键分子。约30%的上皮性卵巢癌中存在P16基因功能缺失，其原因包括杂合性缺失、启动子区甲基化和编码区错义突变。将其插入腺病毒载体，作为基因辅助治疗的基础实验。为了控制p16基因的功能，我们构建了缺失DNA结合域的ER基因与全长p16基因的复合体。该融合蛋白仅在雌激素处理条件下表达，被认为适合于体内应用。理论上，只有在雌激素存在的条件下，p16才能表达。由于雌激素受体内化携带p16蛋白，仅在雌激素阳性条件下观察到p16基因导致的G1期细胞阻滞。在雌激素存在的条件下，MTT法观察到野生型p16基因缺失的卵巢癌细胞因感染ER-p16融合蛋白而生长停滞。通过Western Blot分析证实了雌激素刺激的细胞内转换系统。报道了基因治疗应用的可能性。少

Clear cell adenocarcinoma of the ovary is the poor prognostic tissue subtypes due to the chemo-resistance and/or the early lymph node metastases. It is sometimes affected from the endometriosis and frequent subtypes in Japanese. On the other hand, serous adenocarcinoma of the ovary is common epithelial cancer tissue-subtypes, and it is highly sensitive for the chemotherapy though the peritoneal dissemination is frequent. We tried to identify the specifically expressing genes in clear cell adenocarcinoma by using Differential Display method and SP17 gene was identified as one candidate gene. The function of SP17 gene was unknown, so the full length of SP17 gene was inserted into the Adenovirus vector, and it was infected to the serous adenocarcinoma cells. As further confirmative experiment, RNAi interference of SP17 gene was introduced in the clear cell adenocarcinoma, and the function of SP17 gene was analyzed. MTS assay and Cell-death detection assay were performed in the ovarian can … More cer cells, with or without treatment of Paclitaxel, most useful anticancer drug for the ovarian cancers. As results, it was strongly suggested that SP17 plays an important role of chemotherapy-resistance of clear cell adenocarcinoma of the ovary.P16INK4A gene is a cyclin-dependent kinase inhibitor, one of the tumor suppressor genes, and a key molecule of cell cycle G1-S entry. P16 gene function was loss in about 30% of the epithelial ovarian cancers, due to the loss of heterozygosity, methylation of the promoter region and some missense mutations of coding regions. It was inserted into Adenovirus vector as a fundamental experiment of the gene supplementary therapy. In order to control the function of p16 gene, we made the construct of the ER gene which was lack of DNA binding domain combined with full length of p16 gene. The ER-p16 fusion protein which function would be expressed in only estrogen-treatment condition, was considered to suitable for in vivo application. Theoretically, p16 function was expressed only under the estrogen existence condition. The G1 cell cycle arrest due to p16 gene was observed only in estrogen positive condition because of the estrogen receptor internalization carrying p16 protein. The growth arrest of ovarian cancer cells lack of wild type p16 gene was observed by MTT assay only in estrogen existence condition due to infected ER-p16 fusion proteins. Intracellular switching system by the estrogen stimulus was confirmed by Western Blot analyses. The possibility to the gene therapy application was reported. Less