Medical treatment research by the gene cluster (EPR-1, dCK, and TP) which guides antineoplastic drug susceptibility reinforcement

将潜在的Survivin反义基因EPR-1连接到含有金属硫蛋白启动子的载体pMTN2中。然后将pMTN2-EPR-1载体导入人结肠腺癌细胞HT29(Survivin表达阳性)。锌可诱导EPR-1高表达的HT29-AS33和HT29-AS49亚系细胞发生凋亡，并增加对顺铂和5-FU的体外敏感性。然后，结合今后的临床研究，构建了表达EPR-1的重组腺病毒载体(Ad.CMV-EPR-1)，并对其在裸鼠HT29裸鼠移植瘤上的体内抗肿瘤作用进行了评价。当移植瘤直径大于或等于8 mm时，给荷瘤小鼠一次性注射Ad.CMV-EPR1和Ad.CMV-LacZ。前者肿瘤形成率为对照组的1/4。提示其作用机制可能与诱导HT29细胞凋亡有关。此外，顺铂或5-FU联合Ad.CMV-EPR-1的疗效得到提高，肿瘤体积…降低分别增加25%和16%(EUJ癌症出版社)。此外，我们先前的研究表明，携带dThdPase(TP)表达载体的细胞对5‘-DFUR和5-FU更敏感(BR J癌症75,1997)。因此，在本研究的同时，将1.5kb的TPcDNA转移到逆转录病毒载体pNV7的EcoRI位点，获得了转染腺癌细胞MC38-TP和MC38-Neo的细胞系。Western印迹和ELISA法分析表明，在MC38-TP中可检测到TP的表达，而在对照组中未检测到TP的表达，且两个亚克隆的生长速度相同。由于前者在体外对5-FU、5‘-DFUR和卡培他滨的敏感性显著增加，体内实验也通过皮下接种MC38-TP小鼠模型进行，证实了5’-DFUR和卡培他滨对MC38-TP的化疗效果增强(数据提交)。这种与化疗敏感性相关的基因研究可能有助于为目前的癌症化疗提供有效的改进。较少

EPR-1 cDNA, a potential survivin antisense, was ligated into a plasmid vector (pMTN2) with metallothionein promoter. Then HT29 human colon adenocarcinoma cells (with positive survivin expression) were transfected with the pMTN2-EPR-1 plasmid vector. Apoptosis was induced in HT29-AS33 and HT29-AS49 subolones with high EPR-1 expression in the presence of Zinc and furthermore sensitivity to Cis-DDP and 5-FU in vitro was increased. Then, on the basis of future clinical development, EPR-1 expressing adenovirus vector (ad.CMV-EPR-1) was made and the in vivo antitumor treatment was assessed on nude mouse HT29 xenografts. When the xenografts were 8mm or more, Ad.CMV-EPR1 and Ad.CMV-LacZ were injected to the tumor bearing mice once. In former group tumor formation was reduced to 1/4 comparing to the controls. This, suggested that the effect was due to HT29 cell line apoptosis induction. In addition, efficacy of CDDP or 5-FU combination with Ad.CMV-EPR-1 was enhanced, with reduction in tumor vol … More ume of 25% and 16% respectively (Eur J Cancer, in Press). Moreover, we have previously shown that transfected cells with plasmid vector expressing dThdPase(TP) cDNA were more sensitive to 5'-DFUR and 5-FU(Br J Cancer 75,1997). Therefore, at the time of this study, TPcDNA (1.5 kb) was transferred to EcoRI site of a retroviral vector, -pNV7 and transfected murine adenocarcinoma cell lines of MC38-TP and MC38-Neo were obtained. In Western blot and ELISA analysis the expression of TP was detected in MC38-TP, but not in the controls and the growth rate of both subclones was identical. As increased sensitivity to 5-FU,5'-DFUR and Capecitabine was significantly observed in vitro for the former group, in vivo experiment was also studied using subcutaneously inoculated MC38-TP mouse models which confirmed the enhancement of 5'-DFUR and Capecitabine chemotherapeutic effect in MC38-TP (data in submission). Such chemosensitivity related gene studies may be useful to provide effective improvement in current cancer chemotherapy. Less