TRANSFORMING GROWTH FACTORS IN NEOPLASTIC TRANSFORMATION

肿瘤转化中的转化生长因子

• 批准号: 2090828
• 负责人: HAROLD L MOSES
• 金额: $ 70.86万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 2000-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2090828
• 关键词: DNA binding protein; breast neoplasms; cell growth regulation; clone cells; complementary DNA; gene expression; genetic promoter element; genetic regulation; genetic transcription; genetically modified animals; in situ hybridization; keratin; laboratory mouse; laboratory rabbit; metallothionein; molecular oncology; neoplastic transformation; oncogenes; point mutation; polymerase chain reaction; southern blotting; transfection; transforming growth factors; tumor suppressor genes

In the past project period it was demonstrated that TGFbeta1 rapidly suppresses expression of the protooncogene c-myc at the level of transcriptional initiation in keratinocytes, and that expression of c-myc is necessary for proliferation of these cells. We have identified the TGFbeta control element (TCE) in the human c-myc promoter and further shown that the product of the retinoblastoma tumor susceptibility gene (pRB) is probably necessary for TGFbeta suppression of c-myc, at least in certain cell types. Preliminary data indicate that the tumor suppressor, p53, can also regulate c-myc transcription. Studies on carcinoma cell lines have shown that abnormalities in the rb gene correlate with the loss of the growth inhibitory response to TGFbeta1. Also in the past project period, studies with MMTV-TGFbeta1 transgenic mouse lines have shown a phenotype of inhibition of end-bud growth and branching morphogenesis. Additional studies have shown that focal administration or local overproduction of TGFbeta1 can cause increased connective tissue formation, including angiogenesis, and immunosuppression. Studies with MMTV-TGFalpha transgenic lines in another laboratory have demonstrated that misregulation of TGFalpha in breast epithelial cells leads to hyperplasia and an increased incidence of spontaneous and DMBA-induced tumorigenesis. Based on this and other data, it is hypothesized that in early steps of mammary carcinogenesis, over-expression of TGFalpha promotes, and over-expression of TGFbetas retards or suppresses, the carcinogenic process. With mutations in the carcinoma cells that result in loss of TGFbeta growth inhibition, endogenous TGFbetas then accelerate tumor progression through paracrine effects on stroma, including angiogenesis, and local immunosuppression. It is further hypothesized that the specific alterations that can cause loss of the growth inhibitory response to the TGFbetas are rb deletions, p53 mutations, and/or c-myc misregulation. These hypotheses will be tested through the following specific aims: (1) Continued studies on determining the role of pRB and p53 in the TGFbeta pathway for suppression of c-myc transcription and growth inhibition; characterizing a newly isolated cDNA clone encoding a protein that specifically binds the TCE; and characterizing the effect of p53 on c-myc transcription. (2) Transfection studies to explore the effect of c-myc, pRB and p53 on TGFbeta1 growth inhibition and the effect of TGFbeta1 over-expression in selected carcinoma cell lines. (3) Transgenic mouse studies on development and susceptibility to tumor formation resulting from misregulation of TGFbeta1S223,225, including crossbreeding of homozygous MMTV-TGFalpha and MMTV-TGFbeta1S223,225 lines to determine effect of transgene coexpression on mammary tumor formation; examining the effect of expression of TGFalpha and TGFbeta1S223,225 under the control of the whey acidic protein (WAP) promoter; crossbreeding MMTV-myc and MMTV- TGFbeta1S223,225 or WAP-myc and WAP-TGFbeta1S223,225 transgenic mice to determine effects on breast tumor formation; and experiments with cytokeratin and metallothionein promoters to determine the effects of misregulating TGFbeta1S223,255 on different tissues.

在过去的项目期间，已经证明TGF β 1可以迅速地 抑制原癌基因c-myc的表达， 角质形成细胞转录起始和c-myc表达 是这些细胞增殖所必需的。 我们已经确定了 人c-myc启动子中的TGF β控制元件（TCE），并且进一步 显示视网膜母细胞瘤肿瘤易感基因的产物 (pRB)可能是TGF β抑制c-myc所必需的，至少 在某些细胞类型中。 初步数据显示肿瘤 抑制基因p53也可以调节c-myc转录。 研究 癌细胞系显示Rb基因的异常 与对TGF β 1的生长抑制反应的丧失相关。 同样在过去的项目期间，MMTV-TGF β 1转基因的研究 小鼠品系显示出抑制末端芽生长的表型， 分枝形态发生 其他研究表明， TGF β 1的施用或局部过度产生可引起增加的 结缔组织形成，包括血管生成，以及 免疫抑制 MMTV-TGF α转基因株系的研究 另一个实验室已经证明， 乳腺上皮细胞导致增生， 自发和DMBA诱导的肿瘤发生。 基于这个和其他 数据，假设在乳腺癌发生的早期阶段， TGF α的过度表达促进TGF β的过度表达， 延缓或抑制致癌过程。 突变的 导致TGF β生长抑制丧失的癌细胞， 内源性TGF β然后通过旁分泌加速肿瘤进展 对间质的影响，包括血管生成和局部免疫抑制。 进一步假设，可以引起 对TGF β的生长抑制应答的丧失是Rb缺失， p53突变和/或c-myc失调。 这些假设将是 通过以下具体目标进行测试：（1）继续研究 确定pRB和p53在TGF β通路中的作用， 抑制c-myc转录和生长抑制;表征 一个新分离的cDNA克隆，编码一种特异性结合 TCE;和表征p53对c-myc转录的影响。 (2)转染研究以探索c-myc、pRB和p53对细胞凋亡的影响。 TGF β 1生长抑制和TGF β 1过表达在人乳腺癌中的作用 选择的癌细胞系。 (3)转基因小鼠研究 发展和对肿瘤形成的易感性， TGF β 1 S223，225的失调，包括纯合子的杂交 MMTV-TGF α和MMTV-TGF β 1 S223，225细胞系，以确定 转基因共表达对乳腺肿瘤形成的影响 TGF α和TGF β 1 S223，225的表达受 乳清酸性蛋白（WAP）启动子;杂交MMTV-myc和MMTV- TGF β 1 S 223，225或WAP-myc和WAP-TGF β 1 S 223，225转基因小鼠， 确定对乳腺肿瘤形成的影响;以及实验 细胞角蛋白和金属硫蛋白启动子，以确定 在不同的组织上错误调节TGF β 1 S223，255。