Department of Experimental Therapeutics, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.
Gynecol Oncol. 2011 Aug;122(2):402-9. doi: 10.1016/j.ygyno.2011.04.034. Epub 2011 May 18.
Cisplatin is a DNA-damaging antitumor agent that is highly effective in treating ovarian cancer. It activates the p53/p21 pathway for its cytotoxic mode of action, but it does not induce p21-dependent cell cycle arrest in G1. Therefore, we investigated this paradox, and used the model analog DAP as a positive control for p21-dependent G1-arrest.
Studies were conducted in p53-proficient ovarian A2780 tumor cells to examine Cdk activity, cell cycle distribution and DNA damage signaling after cisplatin or DAP in combination with the mitotic inhibitor nocodazole.
Cisplatin consistently induced transient S-phase arrest by inhibiting Cdk2/cyclin A complex in S-phase at 12 h and then a durable G2/M-arrest by inhibiting Cdc2/cyclin B complex at 12-18 h. These inhibitions were associated with Chk1 and Chk2 activation and resultant increase in inhibitory tyrosine phosphorylation of Cdk2 and Cdc2. Cisplatin also potently inhibited G1-phase Cdk4/cyclin D1 and Cdk2/cyclin E activities at ~18 h. In agreement, exposure of cisplatin-treated A2780, HCT-116(p53-/-) and HCT-116(p21-/-) tumor cells to nocodazole revealed limited G1-arrest that was dependent on p53 and p21. In contrast, the durable G1-arrest by DAP, which failed to activate Chk1 and Chk2, was unaffected by nocodazole.
Cisplatin induced G1-arrest, but at an attenuated level. This was primarily due to orchestration of Cdk inhibition in S-phase first, then in G2, and finally in G1 that effectively blocked cells in G2 and prevented cells from progressing and arresting in G1. These studies demonstrate that cisplatin unequivocally activates G1-checkpoint response, but the fidelity of G1-arrest is compromised by Chk1/2 activation and checkpoint response in S- and G2/M-phase.
顺铂是一种 DNA 损伤型抗肿瘤药物，在治疗卵巢癌方面非常有效。它通过激活 p53/p21 通路发挥细胞毒性作用，但不会诱导 p21 依赖性 G1 期细胞周期阻滞。因此，我们研究了这一悖论，并使用模型类似物 DAP 作为 p21 依赖性 G1 期阻滞的阳性对照。
在 p53 功能正常的卵巢 A2780 肿瘤细胞中进行研究，以研究顺铂或 DAP 与有丝分裂抑制剂诺考达唑联合使用后 Cdk 活性、细胞周期分布和 DNA 损伤信号。
顺铂通过在 12 小时时抑制 S 期的 Cdk2/周期蛋白 A 复合物，始终诱导短暂的 S 期阻滞，然后通过在 12-18 小时时抑制 Cdc2/周期蛋白 B 复合物，导致持久的 G2/M 期阻滞。这些抑制作用与 Chk1 和 Chk2 的激活以及随后 Cdk2 和 Cdc2 的抑制性酪氨酸磷酸化增加有关。顺铂还在约 18 小时时强烈抑制 G1 期 Cdk4/周期蛋白 D1 和 Cdk2/周期蛋白 E 的活性。顺铂处理的 A2780、HCT-116(p53-/-)和 HCT-116(p21-/-)肿瘤细胞暴露于诺考达唑，结果显示 G1 期阻滞有限，这依赖于 p53 和 p21。相比之下，DAP 诱导的持久 G1 期阻滞未能激活 Chk1 和 Chk2，不受诺考达唑影响。
顺铂诱导 G1 期阻滞，但程度较弱。这主要是由于首先在 S 期，然后在 G2 期，最后在 G1 期协调 Cdk 抑制，有效地阻止细胞进入 G2 期，并防止细胞进入 G1 期并在 G1 期停滞。这些研究表明，顺铂明确激活 G1 检查点反应，但 Chk1/2 激活和 S 期和 G2/M 期的检查点反应会损害 G1 期阻滞的保真度。
