Chow Kai Uwe, Nowak Daniel, Boehrer Simone, Ruthardt Martin, Knau Andrea, Hoelzer Dieter, Mitrou Paris S, Weidmann Eckhart
Department of Internal Medicine III, Hematology and Oncology, Johann Wolfgang Goethe-University Hospital, 60590 Frankfurt am Main, Germany.
Biochem Pharmacol. 2003 Sep 1;66(5):711-24. doi: 10.1016/s0006-2952(03)00410-6.
Cytotoxic drugs mediate apoptotic tumor cell death by influencing key regulator proteins of programmed cell death. In clinical practice cytotoxic drug combinations are desired to potentiate tumor cell kill and to minimize side effects. Nevertheless, the molecular mechanisms underlying synergistic and antagonistic effects on tumor cells are still poorly understood. In order to elucidate these molecular mechanisms we established models of synergistic and antagonistic drug combinations within the same lymphoma cell lines. By combination index method we demonstrated that bendamustine in combination with either doxorubicin or mitoxantrone caused antagonistic effects on disruption of mitochondrial membrane potential as well as on the rate of apoptosis. In contrast the combination of bendamustine with cladribine acted synergistically on these parameters. By using the IC(50) (dosages causing 50% rate of apoptosis) the synergistic effect of the combination of bendamustine and cladribine was associated with an enhanced mitochondrial release of cytochrome c and Smac/DIABLO, by down-regulation of x-linked inhibitor of apoptosis (XIAP), cIAP1, Par-4 and Daxx as well as by a significantly increased activation of caspases-3, -6, -7, -8 and -9. At the same rate of apoptosis (IC(50)), the antagonistic combinations did not increase the release of cytochrome c or Smac/DIABLO, nor down-regulate the expression of XIAP, cIAP1, Par-4 and Daxx, nor increase the activation of caspases. The role of down-regulation of IAPs and of enforced caspase activation for synergism in this model was supported by the observation, that broad spectrum inhibition of caspases re-established expression of XIAP. Our study is the first to outline the molecular alterations caused by synergistic and antagonistic drug combinations within the same lymphoma cell model. The above described mechanisms were already assessable at a point where the effects of synergistic or antagonistic combinations could not yet be discriminated quantitatively by the level of apoptosis rate of the lymphoma cells.
细胞毒性药物通过影响程序性细胞死亡的关键调节蛋白来介导凋亡性肿瘤细胞死亡。在临床实践中，人们期望使用细胞毒性药物组合来增强对肿瘤细胞的杀伤作用并将副作用降至最低。然而，对肿瘤细胞产生协同和拮抗作用的分子机制仍知之甚少。为了阐明这些分子机制，我们在同一淋巴瘤细胞系中建立了协同和拮抗药物组合模型。通过联合指数法，我们证明苯达莫司汀与阿霉素或米托蒽醌联合使用对线粒体膜电位的破坏以及凋亡率产生拮抗作用。相比之下，苯达莫司汀与克拉屈滨联合使用对这些参数具有协同作用。通过使用IC(50)（导致50%凋亡率的剂量），苯达莫司汀和克拉屈滨联合使用的协同作用与细胞色素c和Smac/DIABLO的线粒体释放增加有关，这是通过下调凋亡抑制蛋白（XIAP）、细胞凋亡抑制蛋白1（cIAP1）、前列腺凋亡反应蛋白4（Par-4）和死亡相关蛋白（Daxx）以及显著增加半胱天冬酶-3、-6、-7、-8和-9的激活来实现的。在相同的凋亡率（IC(50)）下，拮抗组合并未增加细胞色素c或Smac/DIABLO的释放，也未下调XIAP、cIAP1、Par-4和Daxx的表达，也未增加半胱天冬酶的激活。在该模型中，对凋亡抑制蛋白的下调和半胱天冬酶的强制激活在协同作用中的作用得到了以下观察结果的支持，即对半胱天冬酶的广谱抑制重新建立了XIAP的表达。我们的研究首次概述了在同一淋巴瘤细胞模型中协同和拮抗药物组合引起的分子改变。上述机制在淋巴瘤细胞凋亡率水平尚无法定量区分协同或拮抗组合效果的阶段就已经可以评估了。
