CHROMOSOME ALTERATIONS AND PROTO-ONCOGENE TRANSPOSITION IN CARCINOGENESIS

致癌过程中的染色体改变和原癌基因转座

• 批准号: 5201499
• 负责人: N C POPESCU
• 金额: --
• 依托单位: DIVISION OF CANCER EPIDEMIOLOGY AND GENETICS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/5201499
• 关键词: HeLa cells; Kaposi's sarcoma; adeno associated virus group; carcinogenesis; chromosome deletion; chromosome translocation; cytogenetics; digital imaging; fluorescent dye /probe; gene therapy; genetic mapping; genetic markers; genetic transduction; human genetic material tag; human tissue; in situ hybridization; molecular oncology; neoplasm /cancer genetics; protooncogene; tissue /cell culture; transfection; transfection /expression vector

Molecular cytogenetics investigations utilizing fluorescence in situ hybridization (FISH) and digital imaging analysis has resulted in significant progress in cancer molecular cytogenetics, mapping of viral sequences and single-copy cancer-related genes. Kaposi's sarcoma (KS) is a neoplasia of rising incidence due to it's association with infection by human immunodeficiency virus and AIDS. Until recently, the only cultured KS cells available were hyperplastic normal diploid cells. Two tumorigenic KS cell lines, one established from a patient with AIDS and another derived from a patient with renal transplant-associated KS who also had an acute infection with cytomegalovirus, have been isolated and were examined cytogenetically. Deletion and translocation of the short arm of chromosome 3 at region 3p14 was identified in both cell lines. Nonrandom alterations in these lines may be a key contributing lesion to the process of neoplastic development, and thus provide a cytogenetic marker leading to the isolation of specific molecular probes for detecting tumor cells in KS. In gene therapy, the delivery system is critical for a successful outcome. The ability of wild type adeno- associated virus (AAV) to integrate site-specifically into chromosome 19 and to infect a variety of cell types, including nondividing cells, makes this agent unique for gene therapy. To examine if AAV recombinants retained the site-specific integration associated with the wild type virus, the integration sites were determined by FISH. FISH analysis of HeLa clones transfected with recombinant AAV demonstrated that in contrast to the wild type virus, the recombinant virus did not specifically integrate in chromosome 19, but integrated nonrandomly to regions of proto-oncogenes. These results are important in evaluating a delivery system for gene therapy. Localization of genes in the human genome has been essential for advances in the molecular characterization of chromosome alterations and understanding of their role in cancer development. The direct visualization of the fluorescent signal on banded metaphase chromosomes is critical to the resolution of mapping. A procedure was developed in our laboratory based on contrast enhanced digital images of DAPI banded chromosomes. With this procedure immune gamma interferon, HER4/erbB-4, Syrian hamster Tp53 and human macrophage metalloelastase gene have been mapped. The localization of these genes will facilitate linkage analysis, identification of gene interactions as well as the analysis of specific cancer cell chromosomal rearrangements involving their loci.

利用原位荧光进行分子细胞遗传学研究 杂交（FISH）和数字成像分析已产生 癌症分子细胞遗传学、病毒图谱研究取得重大进展 序列和单拷贝癌症相关基因。 卡波西肉瘤 (KS) 是一种由于与感染相关而发病率不断上升的肿瘤 人类免疫缺陷病毒和艾滋病。 直到最近，唯一 培养得到的KS细胞均为增生性正常二倍体细胞。 二 致瘤 KS 细胞系，一种从艾滋病患者建立的细胞系 另一种来自肾移植相关 KS 患者，他 还患有巨细胞病毒急性感染，已被隔离并 进行了细胞遗传学检查。 短片的删除和易位 在两个细胞系中均鉴定出 3 号染色体臂的 3p14 区域。 这些线的非随机改变可能是导致病变的关键 肿瘤发展的过程，从而提供细胞遗传学 导致分离特定分子探针的标记 检测 KS 中的肿瘤细胞。在基因治疗中，传递系统是 对于取得成功的结果至关重要。 野生型腺病毒的能力 相关病毒 (AAV) 定点整合到 19 号染色体上 并感染多种细胞类型，包括非分裂细胞，使得 这种药物对于基因治疗来说是独一无二的。 检查 AAV 是否​​重组 保留与野生型相关的位点特异性整合 病毒，通过 FISH 确定整合位点。 FISH 分析 用重组 AAV 转染的 HeLa 克隆证明， 与野生型病毒相比，重组病毒没有 特异性整合到 19 号染色体上，但非随机整合到 原癌基因区域。 这些结果对于评估很重要 基因治疗的传递系统。 基因在人体中的定位 基因组对于分子表征的进步至关重要 染色体改变及其在癌症中的作用的理解 发展。 荧光信号的直接可视化 带状中期染色体对于定位的分辨率至关重要。 我们的实验室开发了一种基于对比度增强的程序 DAPI 带状染色体的数字图像。 通过此程序免疫 γ 干扰素、HER4/erbB-4、叙利亚仓鼠 Tp53 和人巨噬细胞 金属弹性蛋白酶基因已被定位。 这些基因的定位 将有助于连锁分析、基因相互作用的鉴定 以及特定癌细胞染色体重排的分析 涉及他们的基因座。