Raju Uma, Nakata Eiko, Mason Kathy A, Ang K Kian, Milas Luka
Department of Experimental Radiation Oncology, The University of Texas, M. D. Anderson Cancer Center, Houston, Texas 77030, USA.
Cancer Res. 2003 Jun 15;63(12):3263-7.
Flavopiridol, a cyclin-dependent kinase (cdk) inhibitor, can cause cell cycle arrest, induce apoptosis in cancer cells, and inhibit tumor cell growth in vivo. The present study investigated the in vitro radiosensitizing effect of flavopiridol and the underlying molecular mechanisms in a murine ovarian cancer cell line, OCA-I. Flavopiridol inhibited cell growth in a dose-dependent manner and enhanced cell radiosensitivity assessed by the clonogenic cell survival assay. A flavopiridol dose of 300 nM, given for 1 day, enhanced radiosensitivity by a factor of 2.1. Clonogenic cell survival after split-dose radiation showed that flavopiridol inhibited repair from radiation damage. In addition, flavopiridol treatment (300 nM, 1 day) resulted in decreased levels of Ku70 and Ku86 proteins that play a role in DNA repair processes, suggesting that DNA repair processes may have been disrupted by this agent. Flow cytometry analysis showed that flavopiridol (300 nM, 1 day) accumulated the cells in G(1) and G(2) phases, with a significant reduction in the S phase component. This cell cycle redistribution is likely another mechanism underlying flavopiridol-induced cell radiosensitivity. Flavopiridol down-regulated cyclin D1 and cyclin E protein levels and also inhibited phosphorylation of retinoblastoma protein, which is inconsistent with the observed cell cycle arrest. Among the cdks tested, cdk-9, the catalytic subunit of positive transcription elongation factor b, was significantly down-regulated by flavopiridol, suggesting that flavopiridol may modulate cellular transcription processes. Furthermore, flavopiridol on its own induced apoptosis in the OCA-I cells, whereas in combination with radiation, exerted no additional increase in apoptosis. Taken together, our data show that flavopiridol strongly augmented the response of ovarian carcinoma cells to radiation and that the underlying mechanisms included inhibition of sublethal DNA damage repair and cell cycle redistribution. At the molecular level, transcriptional regulation by flavopiridol may have been involved.
黄酮哌啶醇是一种细胞周期蛋白依赖性激酶（cdk）抑制剂，可导致细胞周期停滞，诱导癌细胞凋亡，并在体内抑制肿瘤细胞生长。本研究调查了黄酮哌啶醇在小鼠卵巢癌细胞系OCA-I中的体外放射增敏作用及其潜在的分子机制。黄酮哌啶醇以剂量依赖性方式抑制细胞生长，并通过克隆形成细胞存活试验评估增强了细胞放射敏感性。给予300 nM黄酮哌啶醇1天，放射敏感性提高了2.1倍。分次照射后的克隆形成细胞存活情况表明，黄酮哌啶醇抑制了辐射损伤的修复。此外，黄酮哌啶醇处理（300 nM，1天）导致在DNA修复过程中起作用的Ku70和Ku86蛋白水平降低，表明该药物可能破坏了DNA修复过程。流式细胞术分析表明，黄酮哌啶醇（300 nM，1天）使细胞在G(1)期和G(2)期积累，S期成分显著减少。这种细胞周期重新分布可能是黄酮哌啶醇诱导细胞放射敏感性的另一种机制。黄酮哌啶醇下调细胞周期蛋白D1和细胞周期蛋白E蛋白水平，还抑制视网膜母细胞瘤蛋白的磷酸化，这与观察到的细胞周期停滞不一致。在所测试的cdk中，正转录延伸因子b的催化亚基cdk-9被黄酮哌啶醇显著下调，表明黄酮哌啶醇可能调节细胞转录过程。此外，黄酮哌啶醇自身诱导OCA-I细胞凋亡，而与辐射联合使用时，并未使凋亡进一步增加。综上所述，我们的数据表明黄酮哌啶醇强烈增强了卵巢癌细胞对辐射的反应，其潜在机制包括抑制亚致死性DNA损伤修复和细胞周期重新分布。在分子水平上，可能涉及黄酮哌啶醇的转录调控。
