CD44-P185HER2 INTERACTION IN OVARIAN CANCER PROGRESSION

CD44-P185HER2 在卵巢癌进展中的相互作用

• 批准号: 6150044
• 负责人: Lilly YW Bourguignon
• 金额: $ 22.58万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-01 至 2003-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6150044
• 关键词: CD44 molecule; cell adhesion; cell line; cell motility; cell proliferation; clinical research; cytoskeleton; female; flow cytometry; gene interaction; human subject; human tissue; immunofluorescence technique; in situ hybridization; laboratory mouse; laboratory rabbit; neoplastic cell; neoplastic process; oncogenes; ovary neoplasms; protein isoforms; site directed mutagenesis; transfection

Ovarian cancer is the leading cause of gynecologic malignancy deaths in the United States. In particular, ovarian carcinomas appears to be the most lethal tumor of the female genital tract and continues to be the major cause of mortality in female cancer patients. Metastases from the ovary are common by the time the ovarian enlargement leads to patient symptoms (at advanced stages III-IV). The biological factors affecting the ability of ovarian tumor cells to migrate and implant at distant locations within the peritoneal cavity remain unclear. In the proposed research, we plan to test the hypothesis that the CD44 isoform-p185HER2 interaction with the cytoskeleton plays an important role in promoting oncogenic signals required for tumor cell adhesion, motility, proliferation and growth during human ovarian cancer progression. This hypothesis clearly has important implications for the spread and progression of human ovarian cancers. Specifically, we plan to use a variety of biochemical, immunological and molecular biological techniques to elucidate the structural and functional association between the CD44s-p185HER2 complex and the cytoskeleton. We will also employ in vitro mutagenesis to construct CD44s deletion (or site- directed) mutants that lack certain portions (or replace certain amino acids) of the cytoplasmic domain required for the binding of the cytoskeleton. These mutants polypeptides will be expressed in ovarian tumor cells and their structural changes will be correlated with CD44s- p185HER2-mediated oncogenic properties (e.g. cell adhesion, motility, cell proliferation and growth). In the second part of the proposal, we plan to identify CD44 variant (CD44v) isoforms associated with human ovarian cancers using RT-PCR, DNA sequence analyses and in situ hybridization. We will clone the unique variant exons into standard CD44 cDNA and express these constructs in CD44v-negative (p185HER2- positive) ovarian epithelial cells. These CD44v transfectants will then be analyzed for their functional properties (e.g. CD44v-p185HER2 interaction, adhesion, tumor cell motility, proliferation and growth). We also plan to downregulate CD44v-p185HER2-mediated oncogenic signals by using dominant-negative mutants of Shc and/or Grb2. These mutant polypeptides will be expressed in ovarian tumor cells and their structural changes will be correlated with various oncogenic properties. Finally, specific antibodies against the "fusion" proteins containing CD44 variant exons (related to ovarian cancer cells) will be prepared. The co-expression of certain CD44v isoforms with p185HER2 in ovarian carcinomas will be analyzed during ovarian cancer progression. We believe that the new information obtained from this proposal may establish the CD44v-p185HER2 complex as an important tumor marker for early detection and evaluation of oncogenic potential, as well as allow the development of new drug targets to inhibit tumor cell motility and proliferation/growth.

卵巢癌是#年妇科恶性肿瘤死亡的主要原因。 美国。特别是，卵巢癌似乎是 女性生殖道最致命的肿瘤，一直是 女性癌症患者死亡的主要原因。卵巢癌的转移 卵巢肿大导致患者的卵巢是常见的 症状(晚期III-IV期)。影响人体健康的生物因素 卵巢肿瘤细胞异地迁移和种植能力的研究 腹膜腔内的位置仍不清楚。在建议的 研究中，我们计划检验CD44亚型-p185HER2的假设 与细胞骨架的相互作用在促进 肿瘤细胞黏附、移动所需致癌信号 人卵巢癌进展过程中的增殖和生长。这 假设显然对传播有重要的影响， 人类卵巢癌的进展。具体地说，我们计划使用 生物化学、免疫学和分子生物学的多样性 阐明结构和功能关联的技术 CD44S-p185HER2复合体与细胞骨架之间的关系。我们还将 利用体外诱变构建CD44s缺失(或位点- 缺乏某些部分(或取代某些氨基)的突变体 酸)结合所需的细胞质结构域 细胞骨架。这些突变体多肽将在卵巢中表达 肿瘤细胞及其结构变化将与CD44s- P185HER2介导的致癌特性(如细胞黏附、运动、 细胞增殖和生长)。在建议的第二部分，我们 计划鉴定与人类相关的CD44变异体(CD44v)亚型 应用RT-PCR、DNA序列分析和原位杂交技术检测卵巢癌 杂交。我们将把独特的变异外显子克隆到标准的CD44中 并在CD44v阴性(p185HER2-)中表达 阳性)卵巢上皮细胞。然后这些CD44v转染体将 分析其功能特性(例如CD44v-p185HER2 相互作用、黏附、肿瘤细胞运动、增殖和生长)。 我们还计划下调CD44v-p185HER2介导的致癌信号 利用Shc和/或Grb2的显性负性突变体。这些突变体 多肽将在卵巢肿瘤细胞中表达，其 结构变化将与各种致癌特性相关。 最后，针对含有“融合”蛋白的特定抗体 将准备CD44变异外显子(与卵巢癌细胞有关)。 某些CD44v亚型与p185HER2在卵巢中的共表达 癌症将在卵巢癌进展过程中进行分析。我们 相信从这项提案中获得的新信息可能 建立CD44v-p185HER2复合体作为重要的肿瘤标志物 早期检测和评估致癌潜力，以及允许 抑制肿瘤细胞运动的新药靶点的开发 增殖/生长。