Mitotane increases the radiotherapy inhibitory effect and induces G2-arrest in combined treatment on both H295R and SW13 adrenocortical cell line
Mitotane increases the radiotherapy inhibitory effect and induces G2-arrest in combined treatment on both H295R and SW13 adrenocortical cell line
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DOI：
10.1677/erc.1.1315
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发表时间：
2008-06-01
影响因子：
3.9
通讯作者：
Stigliano, A.
中科院分区：
文献类型：
--
作者：
Cerquetti, L.;Bucci, B.;Stigliano, A.
米托坦，1，1-二氯-2-（邻氯苯基）-2-（对氯苯基）乙烷（o，p '-DDD）是一种具有肾上腺毒性作用的药物，能够阻断皮质醇的合成。这种药物和放射治疗也用于肾上腺癌治疗，即使它们在这种瘤形成中的生物学作用仍然未知。本文研究了o，p '-DDD和电离辐射（IR）对H295 R和SW 13肾上腺皮质癌细胞生长抑制和细胞周期紊乱的影响。两种细胞系均以6戈伊剂量照射，并分别用o，p ′-DDD 10（-5）M处理，并用IR/o，p ′-DDD组合处理。这种联合治疗诱导了两种肾上腺皮质癌细胞中细胞生长的不可逆抑制。细胞周期分析表明，IR单独及IR/o，p '-DDD联合作用可诱导细胞聚集于G（2）期。IR后120 h，细胞能恢复IR诱导的G（2）阻滞，而IR/o，p '-DDD处理的细胞仍阻滞于G（2）期。为了研究G（2）不可逆阻滞的分子机制，我们考虑了H295 R细胞系，该细胞系显示与显著的G（2）阻滞相关的最高细胞增殖抑制。在这些细胞中，细胞周期蛋白B1和Cdk 2蛋白检测蛋白质和Cdk 2激酶活性测定试剂盒测量。IR/o，p '-DDD处理的H295 R细胞中cyclin B1的含量增加，表现为Cdc 2-cyclin B1复合物的免疫共沉淀。激酶活性也显示在用组合疗法处理的细胞中增加。此外，在这些细胞中，p53的序列分析揭示了外显子8和9的大量缺失。IR/o，p '-DDD处理后，在恢复野生型p53的H295 R细胞中也发生了同样的G（2）期不可逆阻滞，表明该机制不是由p53通路介导的。
Mitotane, 1,1-dichloro-2-(o-chlorophenyl)-2-(p-chlorophenyl) ethane (o,p'-DDD) is an agent with adrenotoxic effect, which is able to block cortisol synthesis. This drug and radiotherapy are used also in adrenal cancer treatment even if their biological action in this neoplasia remains unknown. We investigated the effects of o,p'-DDD and ionizing radiations (I R) on cell growth inhibition and cell cycle perturbation in H295R and SW13 adrenocortical cancer cells. Both cell lines were irradiated at a 6 Gy dose and were treated with o,p'-DDD 10(-5) M separately and with IR/o,p'-DDD in combination. This combination treatment induced an irreversible inhibition of cell growth in both adrenocortical cancer cells. Cell cycle analysis showed that IR alone and IR/o,p'-DDD in combination induced the cell accumulation in the G(2) phase. At 120 h after IR, the cells were able to recover the IR-induced G(2) block while cells treated with IR/o,p'-DDD were still arrested in G(2) phase. In order to study the molecular mechanism involved in the G(2) irreversible arrest, we have considered the H295R cell line showing the highest inhibition of cell proliferation associated with a noteworthy G(2) arrest. In these cells, cyclin B1 and Cdk2 proteins were examined by western blot and Cdk2 kinase activity measured by assay kit. The H295R cells treated with IR/o,p'-DDD shared an increase in cyclin B1 amount as the coimmunoprecipitation of Cdc2-cyclin B1 complex. The kinase activity also shows an increase in the treated cells with combination therapy. Moreover, in these cells, sequence analysis of p53 revealed a large deletion of exons 8 and 9. The same irreversible block on G(2) phase, induced by IR/o,p'-DDD treatment, happened in H295R cells with restored wild-type p53 suggesting that this mechanism is not mediated by p53 pathway.