Combined Bcl-2/mammalian target of rapamycin inhibition leads to enhanced radiosensitization via induction of apoptosis and autophagy in non-small cell lung tumor xenograft model.
Combined Bcl-2/mammalian target of rapamycin inhibition leads to enhanced radiosensitization via induction of apoptosis and autophagy in non-small cell lung tumor xenograft model.
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DOI：
10.1158/1078-0432.ccr-09-0589
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发表时间：
2009-10-01
期刊：
影响因子：
--
通讯作者：
Lu B
中科院分区：
文献类型：
--
作者：
Kim KW;Moretti L;Mitchell LR;Jung DK;Lu B
放射治疗在非小细胞肺癌的治疗中起着核心作用。然而，这种模式的有效性通常是有限的，因为抗性来自细胞死亡的缺陷。我们研究了通过Bcl-2抑制剂ABT-737同时上调细胞凋亡和通过mTOR抑制剂雷帕霉素同时上调自噬是否可以用于增强体外H460细胞的放射敏感性和异种移植模型中的生长延迟。体外研究证实，ABT-737和雷帕霉素分别诱导细胞凋亡和自噬。ABT-737诱导裂解的半胱天冬酶-3（细胞凋亡的标志物）和雷帕霉素与GFP-LC 3的点状定位增加（自噬的特征）相关。在克隆形成试验中，ABT-737/雷帕霉素组合显著增强了H460细胞对辐射的敏感性（DER=2.47，p=0.002）。此外，ABT-737/雷帕霉素/放射的组合在小鼠异种移植模型中显示出显著的肿瘤生长延迟。体内免疫组织化学染色显示，与单独放射对照组相比，联合治疗产生超过100%的半胱天冬酶-3活性（凋亡）增加和6倍的p62蛋白水平降低（指示自噬通量）。此外，与单独辐射相比，细胞增殖（Ki 67染色）降低了77%（p=0.001），血管密度（vWF染色）降低了67.5%（p=0.09）。在人脐内皮细胞中的另外的体外研究表明，联合治疗也显著减少小管形成。这些结果表明，细胞凋亡和自噬的同时诱导增强了体外和肺癌异种移植模型中的放射治疗。进一步的研究是必要的，以评估这种策略在肺癌患者中的临床潜力。
Radiotherapy has a central role in the treatment of non-small-cell lung cancer. Effectiveness of this modality, however, is often limited as resistance results from defects in cell death. We investigated whether simultaneous upregulation of apoptosis, via Bcl-2 inhibitor ABT-737, and autophagy, via mTOR inhibitor rapamycin, can be used to enhance radiosensitivity of H460 cells in vitro and growth delay in a xenograft model. In vitro studies confirmed that ABT-737 and rapamycin induce apoptosis and autophagy, respectively. ABT-737 induced cleaved caspase-3, a marker of apoptosis, and rapamycin correlated with an increase in punctate localization of GFP-LC3, characteristic of autophagy. The combination ABT-737/rapamycin markedly enhanced sensitivity of H460 cells to radiation (DER=2.47, p=0.002) in clonogenic assay. In addition, the combination ABT-737/rapamycin/radiation showed a dramatic tumor growth delay in a mouse xenograft model. In vivo immunohistochemistry staining showed that combination therapy yielded over a 100% increase in caspase-3 activity (apoptosis) and a 6-fold decrease in p62 protein level (indicative of autophagic flux) as compared to radiation alone control group. Moreover, cell proliferation (Ki67 staining) was reduced by 77% (p=0.001) and vascular density (vWF staining) by 67.5% (p=0.09) compared to radiation alone. Additional in vitro studies in human umbilical endothelial cells indicated that combined therapy also significantly decrease tubule formation. These results suggest that concurrent induction of apoptosis and autophagy enhances radiation therapy both in vitro and in lung cancer xenograft models. Further investigations are warranted to assess the clinical potential of such strategy in lung cancer patients.