EXTRACHROMOSOMAL DNA IN PATIENTS' OVARIAN CANCERS

卵巢癌患者的染色体外 DNA

• 批准号: 2099242
• 负责人: BRAD E. WINDLE
• 金额: $ 11.02万
• 依托单位: CTRC RESEARCH FOUNDATION
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-07-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2099242
• 关键词: dissection; drug resistance; extrachromosomal DNA; female; genetic mapping; human subject; human tissue; natural gene amplification; neoplasm /cancer genetics; neoplastic process; oncogenes; ovary neoplasms; polymerase chain reaction; women's health

Ovarian cancer is one of the most frequent causes of cancer deaths in women. It was responsible for the deaths of more than 12,000 women last year in the United States. About one in 70 women will develop the disease. One percent of al female deaths in this country are due to ovarian cancer. We are proposing a novel method to identify amplified genes which may be responsible for tumor progression and drug resistance in these patients by targeting extrachromosomal DNA. Extrachromosomal DNA is thought to play a pivotal role in tumorigenesis since amplified oncogenes and drug-resistant genes are often located in extrachromosomal DNA in tumor cells and tumor cell lines. The largest form of extrachromosomal DNA is the double minute chromosome (DM). We now have documented that DMs are frequently found in metaphase spreads from ovarian tumors obtained from fresh biopsy specimens. Therefore, prolonged culture will be avoided since DMs are often lost in vitro. We have developed a strategy to isolate and identify extrachromosomally located (i.e. on DMs amplified oncogenes or drug resistance genes. These amplified oncogenes or drug resistant genes present on DMs can be specifically isolated by chromosome microdissection and the fragments amplified in vitro by polymerase chain reaction (PCR). Initially, we will use PCR-based strategies on DMs to identify the precise region on metaphase chromosomes where the amplified genes of the DMs originate by using gene mapping techniques on normal cell chromosomal preparations. After determining the chromosomal map position of the DM we would be able to determine if we have identified probable new uncharacterized oncogenes and/or drug resistance genes. At that time specific clones could be isolated and their gene sequences ultimately determine. The following specific aims are proposed: 1) Utilize current state-of-the-art chromosome microdissection techniques to determine the incidence of known oncogene amplification on DMs and develop a genetic fingerprinting techniques to characterize DMs. 2) To use chromosome microdissection techniques and PCR amplification to characterize amplified oncogenes and drug-resistant genes in human tumors.

卵巢癌是癌症死亡最常见的原因之一 女性。 它是造成12,000多名女性死亡的原因 在美国。 大约有70个女性会发展 疾病。 这个国家的女性死亡中有百分之一。 卵巢癌。 我们正在提出一种新的方法来识别放大 可能导致肿瘤进展和耐药性的基因 在这些患者中，通过靶向外肌体DNA。 外染色体 自被扩增以来，DNA被认为在肿瘤发生中起关键作用 癌基因和耐药基因通常位于外染色体中 肿瘤细胞和肿瘤细胞系中的DNA。 最大的外染色体DNA形式是双分钟染色体 （DM）。 我们现在已经记录了DMS经常在中期发现 从新鲜活检标本获得的卵巢肿瘤中扩散。 因此，将避免长期文化，因为DMS经常丢失 体外。 我们已经制定了一种隔离和识别外体的策略 位于（即在DMS上放大癌基因或耐药基因。 DMS上存在的扩增的癌基因或耐药基因可以是 特异性通过染色体显微解剖和片段分离 通过聚合酶链反应（PCR）在体外扩增。 最初，我们 将在DMS上使用基于PCR的策略来确定 中期染色体，其中DMS的扩增基因由 在正常细胞染色体制剂上使用基因映射技术。 确定DM的染色体图位置后，我们将能够 确定我们是否已经确定了可能的新的未表征的癌基因 和/或耐药性基因。 当时特定的克隆可能是 分离及其基因序列最终决定。 提出了以下具体目标： 1）利用当前的最新染色体微分解技术 确定DMS和 开发一种遗传指纹技术来表征DMS。 2）使用染色体显微解剖技术和PCR扩增 表征了人类的扩增癌基因和耐药基因 肿瘤。