BRAF抑制剂PLX4032和放疗联合治疗高级神经胶质瘤

The effect of combining BRAF inhibitor PLX4032 and radiation in advanced glioma

目的探讨BRAF抑制剂PLX4032联合放疗对高级别神经胶质瘤的疗效。方法选择两株细胞分别为AM-38(BRAF V600E)和GBM8(BRAF wildtype),用Cell Titre Glo方法检测细胞的IC50;用克隆形成实验检测PLX4032对不同放射剂量的增敏作用;用流式细胞仪检测PLX4032和放疗对细胞凋亡和周期的作用。结果 PLX4032显著抑制BRAF V600E细胞的增殖(IC_(50)=2μM)而对野生型细胞没有作用;克隆形成实验发现在BRAF V600E细胞上随着剂量增加,联合治疗组比单独放疗组的抑制作用更显著(P<0.05),而野生型细胞株上联合组和单独放疗组没有显著性差异(P>0.05);流式细胞分析发现在BRAF V600E突变细胞上,联合治疗组细胞凋亡数显著增加,与对照组和单独治疗组相比有显著差异(P<0.05),而在野生型细胞株上没有显著性差异;流式分析发现在BRAF V600E突变细胞上,与对照组相比,单独放疗组和联合治疗组在G0/G1期细胞数都减少,G2/M期细胞数增加,而且联合治疗组与单独放疗组相比有显著差异。结论 BRAF抑制剂PLX4032与放疗联合治疗BRAF V600E突变的高级别神经胶质瘤的疗效较好。

Objective To investigate the role of BRAF inhibitor PLX4032 in combination with radiotherapy on advanced glioma.Methods BRAF V600E-mutated cancer cell line（AM-38）and BRAFV600 E wild-type cancer cell line（GBM8）were obtained.We performed cell viability assay to detect the IC_（50） of the inhibitor PLX4032 by the Cell Titre Glo assay and clonogenic assays using combinations of radiation and PLX4032.Cell apoptosis and cell cycle were evaluated by flow cytometry.Results PLX4032 significantly inhibited the proliferation of BRAF V600 E cells（IC50=2μM）and had no effect on wild-type cells.By clonogenic assay,combination group had more significant inhibitory effect than radiotherapy alone group for BRAF V600 E cells with increasing dose（P〈0.05）,while there was no significant difference（P〉0.05）on the wild-type cell lines between radiotherapy alone group and the combination group.For apoptosis,flow cytometry analysis showed that cell apoptosis proportion of combination group were significantly different compared to the control group and alone group in the BRAF V600 E cells（P〈0.05）,whereas there＇s no significant differences in the wildtype cell lines.For the cell cylcle,flow analysis found that alone and combined groups have reduced the number of G0/G1 phase of the cell and caused the cell G2/M phase arrest compared to the control group in the BRAF V600 E mutated cell.There were significant difference between the combination group and alone groups.Conclusion These studies provide the pre-clinical rationale for clinical trials of concurrent radiotherapy and BRAF V600 E inhibitors.