Molecular Cytogenetics of Solid Tumors

实体瘤的分子细胞遗传学

• 批准号: 7291776
• 负责人: NICOLAE POPESCU
• 金额: --
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 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/7291776
• 关键词: Molecular; Cytogenetics; Solid; Tumors

The research program of the Molecular Cytogenetics Section of the Laboratory of Experimental Carcinogenesis is aimed at the identification and characterization of genomic modifications associated with initiation and progression of the neoplastic phenotype. Using a combined approach based on integrated use of molecular cytogenetics and molecular biology, our group identified and examined a number of recurrent chromosome alterations that led to the discovery of several new cancer-relevant genes, and to the detection of alterations in a number of known ones. Analysis of these alterations may provide markers useful for the prognosis and diagnosis of cancer and potential targets for therapy. In the past year significant progress has been made molecular cytogenetics and molecular biology of cancer cells. New recurrent chromosome alterations and evidence regarding the role of DLC-1 gene in embryonic development and human breast cancer metastasis. were identifid.. The mouse DLC-1 gene was isolated, and the exon-intron organization was characterized. An intragenic polymorphic microsatellite marker was identified that was useful for linkage mapping and LOH analysis.To provide an animal model system for investigating the biological functions of DLC-1 in vivo, we successfully used homologous recombination in embryonic stem cells to generate mice with a disrupted DLC-1 gene. Mice heterozygous for the disrupted allele were viable and phenotypically normal, although with reduced levels of DLC-1 mRNA. No homozygous mutant progeny was obtained from mating of heterozygous animals, indicating that DLC-1 deficiency resulted in embryonic lethality. Analysis of timed pregnancies showed that DLC-1-/- embryos did not survive beyond 10.5 days gestation , and histological examination revealed defects in the neural tube, brain, heart, and placenta. In situ hybridization localization of DLC-1 mRNA in the wild type mouse embryo showed a widespread expression in embryonic and extraembryonic tissues as at the time when abnormalities were found in mutant embryos. This distribution is consistent with a role for DLC-1 in normal development. Cultured fibroblasts from homozygous mutant embryos displayed alterations in the organization of actin filaments and focal adhesions. These results suggest that the DLC-1 protein plays an important role in the assembly of the cytoskeleton and cell adhesion complexes and loss of DLC-1 may interfere with development by adversely affecting cell adhesion and migration Identification of molecular signatures characteristic of the biological mechanisms involved in the metastasis spread of cancer is required for the development of therapeutic interventions able to abrogate the process. Previously , we demonstrated that restoration of DLC-1 expression in cell lines derived from metastasic breast adenocarcinomas lacking endogenous gene expression caused significant growth inhibition and prevented the development of tumors in athymic nude mice.In collaboaration with Dr. Steve Goodison from University of Florida and his colleagues the role of DLC-1 in metastasis. was examined. Monoclonal cell lines M4A4 and NM2C5 are spontaneously occurring sublines of the MDA-MB235 which exhibit many phenotypic differences in growth, invasion, dissemination and spontaneous metastatic efficiency from an orthotopic site. The common origin of these cell lines enables the comparative investigation of cellular and molecular events in the metastatic process in a stable and isogeneic model. A 171-gene expression signature that correlated with metastatic phenotype highlighted several GTPase signaling components. One of these components, DLC 1 gene, was found down-regulated in the metastatic relative to the non-metastatic cells.

实验癌变实验室分子细胞遗传学研究室的研究项目旨在鉴定和表征与肿瘤表型启动和进展相关的基因组修饰。使用基于分子细胞遗传学和分子生物学的综合使用的综合方法，我们小组识别并检查了一些反复发生的染色体改变，这些改变导致了几个新的与癌症相关的基因的发现，并检测到了一些已知基因的改变。对这些改变的分析可能为癌症的预后和诊断提供有用的标记物，并为治疗提供潜在的靶点。在过去的一年里，分子细胞遗传学和癌细胞的分子生物学取得了重大进展。关于DLC-1基因在胚胎发育和人类乳腺癌转移中的作用的新的重复染色体改变和证据。被确认为..。克隆了小鼠DLC-1基因，并对其外显子-内含子结构进行了鉴定。为了为研究DLC-1的体内生物学功能提供动物模型，我们成功地利用胚胎干细胞中的同源重组产生了DLC-1基因缺失的小鼠。杂合子被破坏的等位基因的小鼠是存活的和表型正常的，尽管DLC-1mRNA水平降低。杂合子动物交配未获得纯合子突变后代，表明DLC-1基因缺失导致胚胎死亡。对定时妊娠的分析表明，DLC-1-/-胚胎不能存活超过10.5天，组织学检查显示神经管、大脑、心脏和胎盘存在缺陷。DLC-1mRNA在野生型小鼠胚胎中的原位杂交定位表明，当突变胚胎出现异常时，DLC-1mRNA在胚胎和胚外组织中广泛表达。这种分布与DLC-1在正常发育中的作用是一致的。来自纯合子突变胚胎的培养成纤维细胞显示肌动蛋白细丝和局部粘连的组织发生了变化。这些结果表明，DLC-1蛋白在细胞骨架和细胞黏附复合体的组装中起着重要作用，DLC-1的缺失可能通过对细胞黏附和迁移产生不利影响而干扰发育。识别癌症转移和扩散所涉及的生物学机制特征的分子特征对于开发能够消除这一过程的治疗干预措施是必要的。此前，我们与佛罗里达大学的Steve Goodison博士及其同事合作，证明了在缺乏内源性基因表达的转移性乳腺癌细胞系中，DLC-1的表达恢复会导致显著的生长抑制，并阻止裸鼠肿瘤的发展。被检查过了。单抗细胞系M4A4和NM2C5是MDA-MB235的自发亚系，在生长、侵袭、扩散和原位自发转移效率方面表现出许多表型差异。这些细胞系的共同起源使得能够在稳定和同源的模型中对转移过程中的细胞和分子事件进行比较研究。与转移表型相关的171个基因表达特征突出了几个GTPase信号成分。其中DLC-1基因在转移细胞中的表达较非转移细胞下调。