Kyprianou N, Rock S
Department of Surgery, University of Maryland School of Medicine, Baltimore 21201, USA.
Anticancer Res. 1998 Mar-Apr;18(2A):897-905.
Previous studies have demonstrated that androgen-independent prostate cancer cancer cells undergo apoptosis in response to ionizing irradiation. The p53 protein controls cell cycle arrest and apoptosis by acting as a checkpoint control that halts the cell cycle in G1, while DNA damage is present. In this study the effect of overexpression of mutant p53 protein, on radiation-induced apoptotic cell death of human prostate cancer cells PC-3 was investigated.
PC-3 cells were transfected with the plasmid encoding the mutant p53 sequence, and the neomycin resistance gene. Selected transfectant clones, were characterized at the molecular level (gene integration, and level of mRNA and protein expression) and cloned transfectants expressing high levels of p53 protein were treated with increasing doses of ionizing irradiation. The cellular response to radiation was determined on the basis of: a) clonogenic survival (colony forming ability of irradiated cells); b) induction of apoptosis as determined by the terminal transferase assay; c) apoptotic DNA fragmentation; and d) induction of expression of genes associated with prostate-apoptosis.
Both mutant p53 transfectant and parental PC-3 cells underwent apoptosis in response to ionizing irradiation following similar kinetics of induction of DNA fragmentation. In addition, the magnitude of induction of expression of prostate apoptosis associated genes, SGP-2 and TGF-beta, was similar in the mutant p53 overexpressing and parental PC-3 cells and coincidental with DNA fragmentation.
These findings seriously challenge the involvement of p53 in radiation-induced apoptosis in human prostate cancer cells and suggest that p53 mutations provide no selective advantage in the development of radioresistance of prostate tumor cells within the context of p53 independent apoptotic pathway.
先前的研究表明，雄激素非依赖性前列腺癌细胞在受到电离辐射后会发生凋亡。p53蛋白通过作为一种检查点控制机制来控制细胞周期停滞和凋亡，该机制在存在DNA损伤时使细胞周期在G1期停止。在本研究中，研究了突变型p53蛋白过表达对人前列腺癌细胞PC-3辐射诱导的凋亡性细胞死亡的影响。
用编码突变型p53序列的质粒和新霉素抗性基因转染PC-3细胞。对筛选出的转染克隆在分子水平（基因整合、mRNA和蛋白表达水平）进行鉴定，并用递增剂量的电离辐射处理表达高水平p53蛋白的克隆转染子。根据以下方面确定细胞对辐射的反应：a）克隆形成存活率（受辐射细胞的集落形成能力）；b）通过末端转移酶测定法确定的凋亡诱导；c）凋亡性DNA片段化；d）与前列腺凋亡相关基因表达的诱导。
突变型p53转染子和亲本PC-3细胞在受到电离辐射后均发生凋亡，DNA片段化诱导动力学相似。此外，在过表达突变型p53的PC-3细胞和亲本PC-3细胞中，前列腺凋亡相关基因SGP-2和TGF-β的表达诱导幅度相似，且与DNA片段化同时发生。
这些发现严重质疑了p53在人前列腺癌细胞辐射诱导凋亡中的作用，并表明在p53非依赖性凋亡途径的背景下，p53突变在前列腺肿瘤细胞放射抗性的发展中没有提供选择性优势。
