Lee Lung-Ta, Schally Andrew V, Liebow Charles, Lee Ping-Ping, Lee Po-Huang, Lee Ming-Ting
Department of Nutrition and Health, ToKo University, Taiwan, P.R. China.
Anticancer Res. 2008 Sep-Oct;28(5A):2599-605.
Protein phosphorylation/dephosphorylation of tyrosine residues is an important regulatory mechanism in cell growth and differentiation. Previously it has been reported that RC-160, an octapeptide analog of somatostatin, and [D-Trp6]LHRH, an agonist of luteinizing hormone-releasing hormone (LHRH), stimulate receptor-mediated activity of tyrosine phosphatases (PTP) and reverse growth promotion of the tyrosine kinase (PTK) class of oncogenes in tumor cells. The effect of RC-160 and [D-Trp6]LHRH on protein phosphorylation was further examined in surgical specimens of human carcinomas. Protein extracts of human ovarian, liver, breast and prostate tumor samples were preincubated with epidermal growth factor (EGF) (10(-7) M) with or without [D-Trp6]LHRH or RC-160 (10(-6) M) at 25 degrees C for 2 h, followed by incubation for 10 min with [gamma-32p]ATP. SDS-PAGE, Western blotting, autoradiography and densitometry were then used to quantify the phosphorylation level of individual protein bands. It was found that EGF enhanced, and [D-Trp6]LHRH and RC-160 reduced phosphorylation of a prominent 300-kDa band. Two proteins (65 and 60 kDa), involved in growth control in tumor cell lines, were also identified in this study. The homology of substrate phosphorylation between induced PTK and PTP in the presence of hormones provided evidence that these substrates might be identical or related in tumors. These findings, along with the previous cell culture results, suggest that many solid tumors may respond to treatment with analogues of somatostatin and LHRH. Collectively, the results further support the hypothesis that these 60-, 65- and 300-kDa protein substrates may be involved in growth-message transduction.
酪氨酸残基的蛋白质磷酸化/去磷酸化是细胞生长和分化中的一种重要调节机制。此前有报道称，生长抑素的八肽类似物RC-160和促黄体生成素释放激素（LHRH）的激动剂[D-Trp6]LHRH，可刺激受体介导的酪氨酸磷酸酶（PTP）活性，并逆转肿瘤细胞中酪氨酸激酶（PTK）类癌基因的促生长作用。在人类癌组织手术标本中进一步研究了RC-160和[D-Trp6]LHRH对蛋白质磷酸化的影响。将人卵巢、肝脏、乳腺和前列腺肿瘤样本的蛋白质提取物在25℃下与表皮生长因子（EGF）（10⁻⁷ M）预孵育2小时，加入或不加入[D-Trp6]LHRH或RC-160（10⁻⁶ M），然后与[γ-³²P]ATP孵育10分钟。接着使用SDS-PAGE、蛋白质印迹法、放射自显影和光密度测定法来量化各个蛋白条带的磷酸化水平。发现EGF增强了，而[D-Trp6]LHRH和RC-160降低了一条明显的300 kDa条带的磷酸化。本研究还鉴定了两种参与肿瘤细胞系生长控制的蛋白质（65 kDa和60 kDa）。在激素存在下诱导的PTK和PTP之间底物磷酸化的同源性提供了证据，表明这些底物在肿瘤中可能相同或相关。这些发现以及先前的细胞培养结果表明，许多实体瘤可能对生长抑素和LHRH类似物的治疗有反应。总体而言，结果进一步支持了这些60 kDa、65 kDa和300 kDa蛋白质底物可能参与生长信号转导的假说。
