顺铂增强替尼泊甙杀伤小细胞肺癌细胞系的作用

Enhancement of cisplatin on cytocidal effect of tineposide against small cell lung cancer cell line

目的通过顺铂(CDDP)与替尼泊甙(VM-26)联合作用,探讨顺铂增强替尼泊甙对小细胞肺癌细胞株杀伤作用的机制。方法用MTT法,测定H446小细胞肺癌细胞24 h抑制率;AO/EB双荧光染色观察细胞活率;琼脂糖凝胶电泳检测DNA断裂条带;半定量RT-PCR及蛋白印迹检测DNA拓扑异构酶Ⅱ(TopoⅡ)及特化蛋白1(SP1)的表达。结果CDDP与VM-26联合应用有明显的协同效应;二药合用与单独用药相比,细胞活率明显降低,DNA损伤程度增加。H446细胞经CDDP单独作用可以促进SP1、TopoⅡα和TopoⅡβmRNA及蛋白的表达;经VM-26单独作用后,TopoⅡα和TopoⅡβmRNA及蛋白表达降低,而SP1表达无明显改变;二药合用后可见TopoⅡαmRNA及蛋白的表达比单独应用CDDP下调,SP1 mRNA及蛋白的表达比单用VM-26升高,但与单用CDDP相比没有明显变化。结论CDDP通过上调SP1促进H446细胞中TopoⅡ表达,为TopoⅡ抑制剂类药物VM-26提供更多的作用靶点,能够明显提高对小细胞肺癌的抑制作用。

Objective To investigate the mechanism of cisplatin enhanceing the effect of tineposide killing small cell lung cancer cell line by combination of the two drug. Methods The microeulture tetrazolium （MTT） assay was used to determine the inhibition rates of CDDP combined with VM-26. Acridine orange/ethidium bromide （AO/EB） fluorescent staining was used to show the cell vitality rates, DNA disruption ladder were applied to reveal cell apoptosis. The mRNA and protein level of topoisomerase Ⅱ（Topo Ⅱ ） and transcript factor SP1 were measured by semi-quantitative RT-PCR and western blot. Results There is synergistic effect between the two drug. The cell vitality rates were decreased in combination group than that of CDDP or VM-26 used alone, the corn- bination treatment group resulted in more serious DNA strand breakage. The expression of Topo Ⅱα, β and SP1 mRNA and protein both increased in CDDP treatment group, while the Topo Ⅱα,β expression decreased and the SP1 expression has no obviously change in VM-26 treatment group. In combination group, Topo Ⅱα expression were decreased comparing with CDDP used has no obviously change as comparing with sion through up-regulating SP1 expression, small cell lung cancer cell. alone, SP1 expression increased comparing with VM-26 used alone, but CDDP used alone. Conclusion CDDP could enhance Topo Ⅱ expreswhich offer more target for Topo Ⅱ inhibitor VM-26 to act on killing