Department of Pathology, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan.
Int J Radiat Oncol Biol Phys. 2011 Jun 1;80(2):540-8. doi: 10.1016/j.ijrobp.2010.12.065. Epub 2011 Mar 11.
Radioresistance is a major cause of treatment failure of radiotherapy (RT) in human cancer. We have recently revealed that acquired radioresistance of tumor cells induced by fractionated radiation is attributable to cyclin D1 overexpression as a consequence of the downregulation of GSK3β-dependent cyclin D1 proteolysis mediated by a constitutively activated serine-threonine kinase, AKT. This prompted us to hypothesize that targeting the AKT/GSK3β/cyclin D1 pathway may improve fractionated RT by suppressing acquired radioresistance of tumor cells.
Two human tumor cell lines with acquired radioresistance were exposed to X-rays after incubation with either an AKT inhibitor, AKT/PKB signaling inhibitor-2 (API-2), or a Cdk4 inhibitor (Cdk4-I). Cells were then subjected to immunoblotting, clonogenic survival assay, cell growth analysis, and cell death analysis with TUNEL and annexin V staining. In vivo radiosensitivity was assessed by growth of human tumors xenografted into nude mice.
Treatment with API-2 resulted in downregulation of cyclin D1 expression in cells with acquired radioresistance. Cellular radioresistance disappeared completely both in vitro and in vivo with accompanying apoptosis when treated with API-2. Furthermore, inhibition of cyclin D1/Cdk4 by Cdk4-I was sufficient for abolishing radioresistance. Treatment with either API-2 or Cdk4-I was also effective in suppressing resistance to cis-platinum (II)-diamine-dichloride in the cells with acquired radioresistance. Interestingly, the radiosensitizing effect of API-2 was canceled by overexpression of cyclin D1 whereas Cdk4-I was still able to sensitize cells with cyclin D1 overexpression.
Cyclin D1/Cdk4 is a critical target of the AKT survival signaling pathway responsible for tumor radioresistance. Targeting the AKT/GSK3β/cyclin D1/Cdk4 pathway would provide a novel approach to improve fractionated RT and would have an impact on tumor eradication in combination with chemotherapy.
放射抵抗是人类癌症放射治疗（RT）失败的主要原因。我们最近发现，分次照射诱导的肿瘤细胞获得性放射抵抗归因于 cyclin D1 的过表达，这是由于 AKT 介导的 GSK3β 依赖性 cyclin D1 蛋白水解的下调，AKT 是一种持续激活的丝氨酸-苏氨酸激酶。这促使我们假设，通过抑制肿瘤细胞获得性放射抵抗，靶向 AKT/GSK3β/cyclin D1 途径可能改善分次 RT。
用 AKT 抑制剂 AKT/PKB 信号转导抑制剂-2（API-2）或 Cdk4 抑制剂（Cdk4-I）孵育后，将两种具有获得性放射抵抗的人肿瘤细胞系暴露于 X 射线。然后进行免疫印迹、集落存活试验、细胞生长分析以及用 TUNEL 和膜联蛋白 V 染色进行细胞死亡分析。通过将人肿瘤异种移植到裸鼠中评估体内放射敏感性。
API-2 处理导致获得性放射抵抗细胞中 cyclin D1 表达下调。API-2 处理时，体外和体内的细胞放射抵抗完全消失，并伴有细胞凋亡。此外，通过 Cdk4-I 抑制 cyclin D1/Cdk4 足以消除放射抵抗。API-2 或 Cdk4-I 的处理对获得性放射抵抗细胞中顺铂（II）-二胺二氯的耐药性也有效。有趣的是，API-2 的放射增敏作用被 cyclin D1 的过表达所抵消，而 Cdk4-I 仍然能够使 cyclin D1 过表达的细胞敏感化。
cyclin D1/Cdk4 是负责肿瘤放射抵抗的 AKT 生存信号通路的关键靶标。靶向 AKT/GSK3β/cyclin D1/Cdk4 途径将为改善分次 RT 提供一种新方法，并结合化疗对肿瘤消除产生影响。
