PILOT--ONCOGENES IN ORAL SQUAMOUS CELL CARCINOMA

试点——口腔鳞状细胞癌中的癌基因

基本信息

批准号：
6336495
负责人：
JEFFREY ALLEN ENGLER
金额：
$ 2.33万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
1999
资助国家：
美国
起止时间：
1999-08-01 至 2000-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6336495
关键词：
3T3 cells carcinoma genetic library genetic mapping head /neck neoplasm human genetic material tag natural gene amplification neoplasm /cancer genetics neoplastic transformation oncogenes squamous cell carcinoma tissue /cell culture

项目摘要

Identification of growth regulatory genes that are altered by mutation, gene amplification, or rearrangement is a major focus of cancer research. For the majority of human tumors, no such oncogene alteration has been found. That unidentified genes are activated is strongly suggested by cytogenetic and chromosome hybridization studies. For squamous cell carcinoma of the head and neck (SCCHN), up to eight new loci seem especially likely to harbor new oncogenes. Although many known oncogenes fail to transform NIH3T3 cells, the development of new assays has been hindered by the inability to efficiently transfect other cell types. Adenovirus-polylysine conjugate was found efficient for stable transfection of rat cells in vitro, facilitating several alternate screening strategies. In the initial screen we isolated breast cancer cDNAs that transform adenovirus E1a-immortalized rat kidney cells. One of these, named TAB (transforming gene amplified in breast cancer), was found amplified about 10-fold in breast cancer MCF-7 cells, and 2-5 fold in several primary breast cancers. Comparison of the full-length, about 400 amino acid open reading frame with sequences in the database failed to detect related genes or functional motifs. TAB was localized to chromosome 3p14.1-2 using somatic cell hybrid lines. The gene was further localized to a 100- 200kb interval in 3p14.1 using a complete YAC contig of 3p14. TAB maps within a region of 8Mb that represents the minimal region of loss of heterozygosity in renal cell carcinoma. The same region is lost from SCCHN and numerous other tumors. We propose to apply this new assay to SCCHN. A plasmid-based, cDNA expression library will be constructed using mRNA from SCCHN cell lines. The library will be introduced into adenovirus E1a-immortalized cells and transforming human cDNAs will be rescued from resulting transformed cell lines. These will be partially sequenced, and novel genes will be mapped to specific human chromosomes. Genes that map to loci previously implicated in SCCHN will be further characterized. These studies aim to isolate at least one new gene that undergoes somatic alteration in SCCHN. We propose to test primary SCCHN tumors for gene amplification of TAB. Since TAB represents a candidate gene relevant to allelic loss at 3p14, we propose to test for sequence alterations at the TAB locus in SCCHN tumors with loss of chromosome 3p.

鉴定因突变而改变的生长调节基因，基因扩增或重排是癌症研究的一个主要焦点。对于大多数人类肿瘤，尚未发现此类癌基因改变成立。强烈表明未识别的基因被激活细胞遗传学和染色体杂交研究。对于鳞状细胞头颈癌 (SCCHN)，出现多达 8 个新位点特别可能含有新的癌基因。尽管许多已知的癌基因无法转化 NIH3T3 细胞，由于无法有效转染其他细胞类型。腺病毒-聚赖氨酸缀合物发现对体外稳定转染大鼠细胞有效，促进几种替代筛选策略。在最初的筛选中，我们分离出了可转化的乳腺癌 cDNA 腺病毒E1a永生化大鼠肾细胞。其中之一，名为 TAB （在乳腺癌中扩增的转化基因），被发现扩增了在乳腺癌 MCF-7 细胞中增加 10 倍，在几种原发细胞中增加 2-5 倍乳腺癌。全长对比，约400个氨基酸开放数据库中序列的阅读框未能检测到相关基因或功能基序。 TAB 定位于染色体 3p14.1-2 使用体细胞杂交系。该基因进一步定位于100- 3p14.1 中的 200kb 间隔使用 3p14 的完整 YAC 重叠群。 TAB 地图在 8Mb 的区域内，代表丢失的最小区域肾细胞癌的杂合性。相同区域丢失 SCCHN 和许多其他肿瘤。我们建议将这种新测定法应用于 SCCHN。基于质粒的 cDNA 将使用来自 SCCHN 细胞系的 mRNA 构建表达文库。该文库将被导入腺病毒E1a永生化细胞中并转化的人类 cDNA 将从转化的细胞中被拯救出来线。这些将被部分测序，新的基因将被定位特定的人类染色体。先前映射到基因座的基因 SCCHN 中涉及的问题将得到进一步定性。这些研究旨在分离至少一个在 SCCHN 中经历体细胞改变的新基因。我们建议测试原发性 SCCHN 肿瘤的 TAB 基因扩增。由于 TAB 代表与 3p14 等位基因丢失相关的候选基因，我们提议测试 SCCHN 肿瘤中 TAB 基因座的序列改变 3p 染色体缺失。