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

人卵巢癌细胞组织特异性表达基因的克隆及腺病毒介导的p16基因治疗的基础研究。

• 批准号: 15591730
• 负责人: KANUMA Tatsuya
• 金额: $ 1.6万
• 依托单位: Gunma University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15591730/
• 关键词: Ovarian cancer; Clear cell carcinoma; Chemo-resistance; SP17; Gene therapy; Adenovirus; Estrogen receptor; p16INK4A gene; 卵巣癌; 癌遺伝子; Differential Display法; エストロゲン受容体; 癌抑制遺伝子

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

卵巢的透明细胞腺癌是由于化学抗性和/或早期淋巴结转移而导致的预后组织亚型不良。有时会受到子宫内膜异位症的影响，日语中经常受到亚型的影响。另一方面，卵巢的浆液性腺癌是常见的上皮性癌组织 - 含量，尽管腹膜传播经常是对化学疗法的高度敏感的。我们试图通过使用差分显示方法来鉴定清晰细胞腺癌中特定表达的基因，而SP17基因被鉴定为一个候选基因。 SP17基因的功能尚不清楚，因此将SP17基因的全长插入到腺病毒载体中，并将​​其感染到浆液性腺癌细胞中。正如进一步确认的实验一样，在透明细胞腺癌中引入了SP17基因的RNAi干扰，并分析了SP17基因的功能。 MTS分析和细胞死亡检测测定法在卵巢罐中进行……更多的CER细胞，或者不使用紫杉醇治疗，这是卵巢癌最有用的抗癌药物。结果，强烈建议SP17在卵巢的透明细胞腺癌的抗化疗法抗性中起重要作用。P16INK4A基因是一种依赖细胞周期蛋白依赖性激酶抑制剂，肿瘤抑制基因之一，以及cyle g1-s canter of cyle cane the thecy canee can in can can can in can of 30％的关键分子，是of 30％的of the of cyith of。杂合性丧失，启动子区域的甲基化以及编码区域的一些错义突变。它被插入腺病毒载体中，作为基因补充疗法的基本实验。为了控制p16基因的功能，我们制造了ER基因的构造，该基因缺乏DNA结合结构域与p16基因的全长结合在一起。 ER-P16融合蛋白的功能仅在雌激素处理条件下表达，被认为适合体内应用。从理论上讲，p16功能仅在雌激素存在条件下表达。由于携带p16蛋白的雌激素受体内在化，仅在雌激素正条件下观察到p16基因引起的G1细胞周期停滞。由于感染了ER-P16融合蛋白，仅在雌激素存在条件下，MTT测定法观察到卵巢癌细胞的生长停滞缺乏野生型p16基因。通过蛋白质印迹分析证实了雌激素刺激的细胞内切换系统。报告了基因治疗应用的可能性。较少的

项目成果

Usefulness of liquid-based cytology specimens for the immunocytochemical study of p16 expression and human papillomavirus testing : a comparative using simulataneously smapled histology materials.

液基细胞学标本在 p16 表达和人乳头瘤病毒检测的免疫细胞化学研究中的用途：使用同时采样的组织学材料进行比较。

• 发表时间: 2004
• 期刊: Cancer Cytopathology 102
• 作者: Yoshida T;Fukuda T;Sano T;Kanuma T;Owada N;Nakajima T
• 通讯作者: Nakajima T

Sperm protein 17 infulences the tissue-specific malignancy of clear cell adenocarcinoma in human eptithelial ovarian cancer.

精子蛋白 17 影响人上皮卵巢癌中透明细胞腺癌的组织特异性恶性肿瘤。

• 期刊: Gynecologic Oncology
• 影响因子: 4.7
• 作者: Tomoko Saito;Tatsuya Kanuma;Tomohiro Tamura;Leri S. Faried;Hiroshi Aoki;Takashi Minegishi
• 通讯作者: Takashi Minegishi