BACKGROUND & AIMS:
:Cutaneous malignant melanoma is a life-threatening cancer with poor prognosis due to a high metastasis potential. The main obstacle in treatment of metastatic melanoma is the resistance to chemotherapy. Recent studies indicated that apoptosis is a common mechanism of action for various cytotoxic agents. As p53 plays an important part in apoptosis, we investigated the role of p53 in chemosensitivity of melanoma cells. Previously, we found that melanoma cell lines containing wild-type p53 have significantly higher response rates to chemotherapy than cell lines with a mutant p53 gene. To confirm the role of p53 in melanoma chemosensitivity further, we transfected an expression vector, pED1, which carries a mutant p53 gene, into a wild-type p53 melanoma cell line, MMAN. We examined the effect of mutant p53 on camptothecin-induced apoptosis and the expression of genes which are known to be involved in apoptosis or drug resistance, such as bcl-2, bax, bak, p21waf1, and P-glycoprotein. Our results indicate that overexpression of the mutant p53 increased the growth rate of MMAN cells, reduced the sensitivity to camptothecin, and lowered drug-induced apoptosis by 2-3-fold. Flow cytometry indicated that the camptothecin-induced apoptosis is not associated with G1 arrest. Furthermore, camptothecin treatment reduced bcl-2 and P-glycoprotein expression in wild-type p53 MMAN cells, but not cells overexpressing mutant p53. These results demonstrate that p53 mutational status is a determinant of melanoma chemosensitivity. p53 may downregulate bcl-2 and P-glycoprotein to induce apoptosis in melanoma cells after chemotherapy.
背景与目标:
:恶性黑色素瘤是一种威胁生命的癌症,因其高转移潜力而预后较差。治疗转移性黑色素瘤的主要障碍是对化学疗法的抵抗力。最近的研究表明,凋亡是各种细胞毒剂的共同作用机制。由于p53在凋亡中起重要作用,因此我们研究了p53在黑色素瘤细胞化学敏感性中的作用。以前,我们发现含有野生型p53的黑素瘤细胞系对化疗的反应率明显高于具有突变p53基因的细胞系。为了进一步证实p53在黑色素瘤化学敏感性中的作用,我们将携带突变p53基因的表达载体pED1转染到野生型p53黑色素瘤细胞系MMAN中。我们检查了突变体p53对喜树碱诱导的细胞凋亡的作用以及已知与细胞凋亡或耐药有关的基因的表达,例如bcl-2,bax,bak,p21waf1和P-糖蛋白。我们的结果表明,突变体p53的过表达提高了MMAN细胞的生长速率,降低了对喜树碱的敏感性,并使药物诱导的细胞凋亡降低了2至3倍。流式细胞仪表明喜树碱诱导的细胞凋亡与G1阻滞无关。此外,喜树碱处理可降低野生型p53 MMAN细胞中bcl-2和P-糖蛋白的表达,但不会过度表达突变型p53的细胞。这些结果表明,p53突变状态是黑色素瘤化学敏感性的决定因素。在化疗后,p53可能下调bcl-2和P-糖蛋白以诱导黑色素瘤细胞凋亡。