Department of Radiation Oncology, The University of Alabama, Birmingham, AL 35249, USA.
Radiother Oncol. 2011 Oct;101(1):183-9. doi: 10.1016/j.radonc.2011.05.083. Epub 2011 Jun 30.
Limited stage small cell lung cancer (SCLC) represents a minority of SCLC. Despite extensive clinical trials, standard treatment remains cisplatin-based chemotherapy and thoracic irradiation (TI). This study focused on the interaction of cisplatin/radiation with the anti-human DR5 monoclonal antibody TRA-8 in SCLC cells. TRA-8 binds specifically to DR5 and has been shown to activate apoptosis.
Four human SCLC cell lines were utilized for experimentation (SCLC-41, SCLC-58, SCLC-68, and SCLC-74). Immunoblot analysis was used to determine relative protein levels of DR5, DR4 and pro-caspase 8 for each cell line. Using a tetrazolium-based assay (XTT), the IC(50) values for cisplatin with or without TRA-8 were determined for the SCLC cell lines. Four SCLC lines were assayed with a combination of TRA-8 (10 μg/ml), 2 Gy radiation and various concentrations of cisplatin. Apoptosis was evaluated using Annexin V-FITC and cleaved caspase immunoblotting. Using a SCLC-58 subcutaneous xenograft model, treatment began 21 d after tumor cell injection. Treatment included weekly cisplatin (4 mg/kg) and radiation of 1 Gy (24 h after cisplatin) and TRA-8 (200 μg) was administered i.p. twice weekly for three weeks.
Immunoblot analysis showed similar levels of DR5 for all cell lines with variable levels of DR4. Various concentrations of TRA-8 antibody (≤ 10 μg/ml) induced no significant cytotoxicity in the SCLC cell lines. The in vitro combination treatment with TRA-8 (10 μg/ml), 1.25 μg/ml cisplatin and 2 Gy radiation showed increased cytotoxicity when compared to combinations without TRA-8. Furthermore, the triple combination demonstrated the greatest amount of apoptosis as measured by Annexin V staining. The in vivo studies showed the combination of 1G y, cisplatin and TRA-8 extended the tumor doubling time to 44 d as compared to any doublet treatment groups that ranged from 12 to 20 d. Analysis of survival data showed 100% of the combination group (RT+cisplatin+TRA-8) were alive 65 d after treatment began whereas all doublet treatment groups showed 50% or less survival.
These studies showed increased cytotoxicity when TRA-8 was added to radiation/cisplatin in SCLC. This effect was demonstrated in vitro and in vivo. TRA-8 represents a promising new agent in the treatment of SCLC.
局限期小细胞肺癌（SCLC）占 SCLC 的少数。尽管进行了广泛的临床试验，标准治疗仍然是基于顺铂的化疗和胸部照射（TI）。本研究集中于顺铂/辐射与抗人 DR5 单克隆抗体 TRA-8 在 SCLC 细胞中的相互作用。TRA-8 特异性结合 DR5，并已被证明能激活细胞凋亡。
使用四种人 SCLC 细胞系进行实验（SCLC-41、SCLC-58、SCLC-68 和 SCLC-74）。免疫印迹分析用于确定每个细胞系的 DR5、DR4 和 pro-caspase 8 的相对蛋白水平。使用四唑盐测定法（XTT），确定 SCLC 细胞系中顺铂与 TRA-8 联合应用的 IC50 值。用 TRA-8（10μg/ml）、2Gy 辐射和不同浓度的顺铂联合检测四种 SCLC 系。用 Annexin V-FITC 和切割的 caspase 免疫印迹法评估细胞凋亡。使用 SCLC-58 皮下异种移植模型，在肿瘤细胞注射后 21 天开始治疗。治疗包括每周顺铂（4mg/kg）和 1Gy 辐射（顺铂后 24 小时），每周两次腹腔注射 TRA-8（200μg），持续三周。
免疫印迹分析显示，所有细胞系的 DR5 水平相似，DR4 水平不同。≤10μg/ml 的 TRA-8 抗体浓度在 SCLC 细胞系中没有引起明显的细胞毒性。与不含 TRA-8 的联合治疗相比，体外联合治疗（10μg/mlTRA-8、1.25μg/ml 顺铂和 2Gy 辐射）显示出更高的细胞毒性。此外，三重联合显示出最大的 Annexin V 染色测量的细胞凋亡量。体内研究表明，与 12 至 20 天的任何双药治疗组相比，1Gy、顺铂和 TRA-8 的联合治疗将肿瘤倍增时间延长至 44 天。生存数据分析显示，联合治疗组（RT+cisplatin+TRA-8）在治疗开始后 65 天的存活率为 100%，而所有双药治疗组的存活率均为 50%或更低。
这些研究表明，在 SCLC 中，当 TRA-8 与放射治疗/顺铂联合应用时，细胞毒性增加。这种效应在体外和体内都得到了证实。TRA-8 是治疗 SCLC 的一种很有前途的新药物。
