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: 3774353
• 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/3774353
• 关键词: MCF7 cell; antireceptor antibody; antisense nucleic acid; autocrine; blocking antibody; breast neoplasms; cell growth regulation; colorectal neoplasms; epidermal growth factor; estrogens; gene expression; genetic promoter element; growth factor receptors; hormone regulation /control mechanism; human tissue; molecular oncology; neoplastic process; neutralizing antibody; oncogenes; protooncogene; reporter genes; tissue /cell culture; transfection; transforming growth factors

Transforming growth factor a (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). TGFalpha 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 TGFalpha 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 70% of primary human breast tumors at a level that exceeded the level found in adjacent normal mammary epithelium.

转化生长因子 a (TGFalpha)、双调蛋白 (AR) 和 cripto-1 (CR-1) 是结构上和某些情况下功能上的蛋白质 与表皮生长因子 (EGF) 相关，TGFα 和 AR 可以结合 EGF 受体 (c-erb B)。 TGFα 已被间接 与许多不同人类的自分泌生长有关 癌细胞，例如乳腺癌和结肠肿瘤。然而，该规定 该生长因子的表达及其生物学干扰 活动尚未得到彻底检查。此外，相对水平 AR 和 CR-1 在这些恶性肿瘤中的表达和生物学功能 未知。目前的研究表明MCF-10A人类 乳腺上皮细胞对外源性 EGF 具有促有丝分裂反应， TGFα 或 AR 以及这些具有点突变的细胞的转化 c-Ha-ras 原癌基因但不与 c-erb B-2 原癌基因一起导致 内源性TGFα表达增加。此外， 在这些细胞中过度表达人类 TGFα cDNA 会导致它们的 体外转化。添加抗 EGF 受体阻断抗体 或者抗TGFα中和抗体可以部分或完全 抑制 Ha-ras 或 TGFalpha 转化的乳腺细胞的生长 表明外部自分泌环路在这些细胞中起作用。在 相比之下，Ha-ras 和 c-erb B-2 中 AR 表达均增加 转化的 MCF-10A 细胞和这些转化体的生长可以 被 AR 反义硫代磷酸寡核苷酸抑制 AR 在转化过程中充当自分泌中介 Ha-ras 和 erb B-2 均使用该途径。雌激素可以 增加雌激素中 TGFα mRNA 和蛋白的表达 反应性人乳腺癌细胞系，例如 MCF-7 或 ZR-75-1 细胞。 使用质粒在 MCF-7 或 ZR-75-1 细胞中进行瞬时转染测定 含有与氯霉素连接的 TGFα 启动子 乙酰转移酶（CAT）或荧光素酶基因已证明 雌激素生理浓度可诱导5至50倍 这些报告基因的活性增加，表明 TGFα启动子含有顺式雌激素反应元件 （ERE）。 MCF-7 或 ZR-75-1 细胞用重组双嗜性病毒感染 TGFα反义mRNA表达载体。这种反义的表达 mRNA 导致雌激素诱导的 TGFα 蛋白产生减少 并达到同等程度的雌激素诱导的抑制作用 这些细胞的增殖。 AR 的特异性 mRNA 和免疫反应性 和 CR-1 已在大约 50% 至 80% 的原发性和 转移性人类结直肠肿瘤，而只有 5% 的正常相邻结直肠肿瘤 结肠或肝脏组织表达这些基因。同样，免疫反应性 AR 和 大约 70% 的人类原发性乳腺肿瘤中检测到了 CR-1 超过邻近正常乳房的水平 上皮。