Roa W, Chen H, Alexander A, Gulavita S, Thng J, Sun X J, Petruk K, Moore R
Department of Radiation Oncology, Cross Cancer Institute, Edmonton, Alberta, Canada.
Clin Invest Med. 2005 Apr;28(2):55-63.
Anti-apoptotic proteins, such as Bcl-2 and Bcl-xL, are frequently over-expressed in human malignancies, and this is correlated with resistance to chemotherapeutic drugs and gamma- radiation. Recently identified small organic molecules capable of inhibiting Bcl-2 and/or Bcl-xL function, may enhance radiation sensitivity of cancer cells in which they are over expressed. We examined whether specific blockade of the BH3-domain binding to Bcl-xL could sensitize cancer cells to gamma- radiation.
Human non-small-cell lung cancer H460 cells with wild-type p53 and H1792 cells with mutant p53 were exposed to various doses of radiation and/or BH3I-1 and for different points of time to BH3I-1 treatment. XTT and clonogenic survival assays were used to evaluate the growth-inhibitory effects of the antagonist BH3I-1, ionizing radiation or both. Western blot analysis was used to examine the cellular effect of the expression of Bcl-xL, Bax, and p53. Apoptosis and cell cycle distribution were analyzed by confocal microscopy with Hoechst 33258 staining and cytochrome c, and flow cytometry, respectively.
BH3I-1 appeared to induce a dose- and time-dependent apoptosis in H460 and H1792 cells, regardless of p53 status. After 2 days of BH3I-1 treatment, the cells that remained attached were exposed to ionizing radiation. Followed by clonogenic assay, BH3I-1 treatment enhanced the radiation sensitivity of H1792 surviving cells with mutant p53, but not in H460 cells with wild-type p53. A transient time-dependent cell cycle blockade at G2-M phase was identified for H1792 cells without subsequent modification of cell cycle distribution.
These findings suggest a potential role for the small molecule inhibitor as a novel radiation sensitizer in non-small cell lung cancer.
抗凋亡蛋白，如Bcl-2和Bcl-xL，在人类恶性肿瘤中经常过度表达，这与对化疗药物和γ射线的抗性相关。最近发现的能够抑制Bcl-2和/或Bcl-xL功能的小分子有机化合物，可能会增强其过度表达的癌细胞对辐射的敏感性。我们研究了BH3结构域与Bcl-xL结合的特异性阻断是否能使癌细胞对γ射线敏感。
将具有野生型p53的人非小细胞肺癌H460细胞和具有突变型p53的H1792细胞暴露于不同剂量的辐射和/或BH3I-1，并在不同时间点进行BH3I-1处理。采用XTT和克隆形成存活试验评估拮抗剂BH3I-1、电离辐射或两者的生长抑制作用。蛋白质免疫印迹分析用于检测Bcl-xL、Bax和p53表达的细胞效应。分别通过用Hoechst 33258染色和细胞色素c的共聚焦显微镜以及流式细胞术分析细胞凋亡和细胞周期分布。
无论p53状态如何，BH3I-1似乎在H460和H1792细胞中诱导剂量和时间依赖性凋亡。BH3I-1处理2天后，将仍附着的细胞暴露于电离辐射。随后进行克隆形成试验，BH3I-1处理增强了具有突变型p53的H1792存活细胞对辐射的敏感性，但对具有野生型p53 的H460细胞没有增强作用。在H1792细胞中发现了G2-M期短暂的时间依赖性细胞周期阻滞，随后细胞周期分布没有改变。
这些发现表明小分子抑制剂作为一种新型辐射增敏剂在非小细胞肺癌中具有潜在作用。
