ROLE OF EGF-RELATED PEPTIDES IN THE PATHOGENESIS OF BREAST AND COLON CANCER

EGF 相关肽在乳腺癌和结肠癌发病机制中的作用

• 批准号: 5200978
• 负责人: D S SALOMON
• 金额: --
• 依托单位: DIVISION OF CANCER BIOLOGY AND DIAGNOSIS
• 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/5200978
• 关键词: MCF7 cell; antireceptor antibody; antisense nucleic acid; autocrine; blocking antibody; breast neoplasms; cell growth regulation; chloramphenicol acetyltransferase; colon neoplasms; epidermal growth factor; estrogens; gene expression; genetic promoter element; growth factor receptors; hormone regulation /control mechanism; human genetic material tag; human tissue; messenger RNA; neoplastic process; neoplastic transformation; neutralizing antibody; protein structure function; protooncogene; receptor binding; transforming growth factors

Transforming growth factor alpha (TGFalpha), amphiregulin (AR) and cripto-1 (CR-1) are proteins that are structurally and in some cases functionally related to epidermal growth factor (EGF) in that TGFalpha and AR can bind to the EGF receptor (c-erb B). TGFbeta has been circumstantially implicated in the autocrine growth of a number of different human carcinoma cells such as breast and colon tumors. However, the regulation of expression of this growth factor and interference with its biological activity have not been thoroughly examined. Moreover, the relative levels of expression and biological function of AR and CR-1 in these malignancies are unknown. The present studies have demonstrated that MCF-10A human mammary epithelial cells are mitogenically responsive to exogenous EGF, TGFalpha or AR and that transformation of these cells with a point-mutated c-Ha-ras protooncogene but not with a c-erb B-2 protooncogene results in an increase in the expression of endogenous TGFalpha. Furthermore, overexpression of a human TGFalpha cDNA in these cells leads to their in vitro transformation. Addition of an anti-EGF receptor blocking antibody or an anti-TGFalpha neutralizing antibody can partially or completely inhibit the growth of the Ha-ras or TGFalpha transformed mammary cells suggesting that an external autocrine loop is operative in these cells. In contrast, AR expression is increased in both Ha-ras and c-erb B-2 transformed MCF-10A cells and the growth of these transformants can be inhibited by AR antisense phosphorothioate oligonucleotides demonstrating that AR is functioning as an autocrine intermediary in the transformation pathway that is utilized by both Ha-ras and erb B-2. Estrogens can increase the expression of TGF` mRNA and protein in estrogen-responsive human breast cancer cell lines such as MCF-7 or ZR-75-1 cells. Transient transfection assays in MCF-7 or ZR-75-1 cells using a plasmid containing the TGFalpha promoter ligated to either the chloramphenicol acetyltransferase (CAT) or luciferase genes have demonstrated that physiological concentrations of estrogens can induce a 5-to 50-fold increase in the activity of these reporter genes, suggesting that the TGFalpha promoter contains a cis-acting estrogen-responsive element(s) (ERE). MCF-7 or ZR-75-1 cells were infected with a recombinant amphotropic TGFalpha antisense mRNA expression vector. Expression of this antisense mRNA lead to a reduction in estrogen-induced TGFalpha protein production and to an equivalent degree of inhibition of estrogen-induced proliferation in these cells. Specific mRNA and immunoreactivity for AR and CR-1 have been detected in approximately 50% to 80% of primary and metastatic human colorectal tumors, whereas only 5% of normal adjacent colon or liver tissue express these genes. Likewise, immunoreactive AR and CR- 1 was detected in approximately 80% of primary human breast tumors at a level that exceeded the level found in adjacent normal mammary epithelium.

转化生长因子α（TGF α）、双调蛋白（AR）和 cripto-1（CR-1）是在结构上并且在某些情况下 在TGF α中与表皮生长因子（EGF）功能相关 AR可与EGF受体（c-er B B）结合。TGF β已经 在一定程度上与一些自分泌生长有关， 不同的人类癌细胞，如乳腺癌和结肠癌。 然而，这种生长因子表达的调节和 干扰其生物活性， 考察此外，表达和生物学的相对水平 AR和CR-1在这些恶性肿瘤中的功能尚不清楚。本 研究表明MCF-10A人乳腺上皮细胞 对外源性EGF、TGF α或AR有促有丝分裂反应， 用点突变的c-Ha-ras转化这些细胞， 原癌基因而不是c-er B B-2原癌基因导致 增加内源性TGF α的表达。 此外，委员会认为， 人TGF α cDNA在这些细胞中的过表达导致其 体外转化 加入抗EGF受体阻断剂 抗体或抗TGF α中和抗体可以部分或 完全抑制Ha-ras或TGF α转化的 乳腺细胞，表明外部自分泌回路是有效的 在这些细胞中。相反，在Ha-ras基因中AR表达增加， 和c-er B B-2转化的MCF-10 A细胞，并观察这些细胞的生长 AR反义硫代磷酸酯能抑制转化子的表达 寡核苷酸证明AR是作为一种自分泌 转化途径中的中间体， Ha-ras和erB B-2. 雌激素可增加TGF β 1的表达。 人乳腺癌雌激素敏感细胞系mRNA和蛋白的表达 如MCF-7或ZR-75-1细胞。MCF-7中的瞬时转染测定 或ZR-75-1细胞，使用含有TGF α启动子的质粒 连接到氯霉素乙酰转移酶（CAT）或 荧光素酶基因已经证明， 雌激素可以诱导5- 50倍的活性增加， 这些报告基因，表明TGF α启动子含有一个 顺式作用雌激素反应元件（ERE）。MCF-7或ZR-75-1 用重组双嗜性TGF α反义核酸感染细胞 mRNA表达载体。 这种反义mRNA的表达导致了一种 减少雌激素诱导的TGF α蛋白产生， 雌激素诱导的增殖抑制的等效程度， 这些细胞。AR和CR-1的特异性mRNA和免疫反应性具有 在约50%至80%的原发性和转移性 人类结直肠肿瘤，而只有5%的正常相邻结肠或 肝组织表达这些基因。 同样，免疫反应性AR和CR- 1在约80%的原发性人类乳腺肿瘤中检测到， 超过邻近正常乳腺组织水平的水平 上皮