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

转化生长因子α(TGFα)、双调节蛋白(AR)和 CRIPTO-1(CR-1)是一种结构上和某些情况下的蛋白质 与TGFα中表皮生长因子(EGF)功能相关 AR可与EGF受体(c-erb B)结合。TGFBeta已经被 间接牵涉到一些人的自分泌增长 不同的人类癌细胞，如乳腺和结肠肿瘤。 然而，这种生长因子和蛋白的表达调控 对其生物活性的干扰还不彻底 检查过了。此外，表达和生物学上的相对水平 AR和CR-1在这些恶性肿瘤中的功能尚不清楚。现在 研究表明，MCF-10A人乳腺上皮细胞 对外源性EGF、TGFα或AR有丝分裂反应 C-Ha-ras点突变对这些细胞的转化作用 原癌基因但不与c-erb B-2原癌基因一起导致 内源性转化生长因子α表达增加。此外， 人转化生长因子α基因在这些细胞中的过表达导致其 体外转化。添加抗EGF受体阻断剂 抗体或抗TGFα中和抗体可以部分或 完全抑制Ha-ras或TGFα转化细胞的生长 乳腺细胞提示外自分泌环路是可操作的 在这些牢房里。相反，AR在两个Ha-ras中的表达都增加 和c-erb B-2转化的MCF-10A细胞及其生长 AR反义硫代磷酸对转化子的抑制作用 证明AR具有自分泌功能的寡核苷酸 两者都利用的转化途径中的中介 HA-RAS和ERB-2。雌激素可增加血管内皮生长因子的表达 雌激素敏感型人乳腺癌细胞株的mRNA和蛋白表达 如MCF-7或ZR-75-1细胞。MCF-7细胞的瞬时转染法 或使用含有转化生长因子α启动子的质粒的ZR-75-1细胞 连接到氯霉素乙酰转移酶(CAT)或 荧光素酶基因已经证明了生理浓度 雌激素可以诱导血管活性增加5到50倍 这些报告基因表明，TGFpha启动子包含一个 顺式作用雌激素反应元件(S)(ERE)。MCF-7或ZR-75-1 重组两性亲性转化生长因子α反义核酸感染细胞 MR NA表达载体。这种反义mRNA的表达导致了 减少雌激素诱导的转化生长因子α蛋白的产生并转化为 雌激素对小鼠肺组织增殖抑制的等效性 这些细胞。AR和CR-1的特异性mRNA和免疫反应性 在大约50%到80%的原发和转移癌中检测到 人类大肠肿瘤，而只有5%的正常邻近结肠或 肝组织表达这些基因。同样，免疫阳性的AR和CR- 在大约80%的原发人类乳腺肿瘤中检测到1 水平超过邻近正常乳腺的水平 上皮组织。

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.