Objective: To study the effects of FLT3-ITD length on 32D cell proliferation, apoptosis and sensitivity to FLT3 inhibitor, so as to provide references for stepwise therapy of FLT3-ITD mutated acute myeloid leukemia patients.
Methods: Three different FLT3-ITD mutants with same or adjacent insert sites were selected and constructed in an eukaryotic expression vector. FLT3-ITD mutants stably expressed 32D cell strains were selected with the help of lentivirus system and IL3 free cell culture medium. The proliferation and apoptosis of 32D cell strains after AC220 treatment were detected.
Results: FLT3-ITD mutants (ITD1, ITD2 and ITD3) stably expressed 32D cell strains were constructed successfully. In the absence of IL3 factor, the proliferation number of ITD1, ITD2 and ITD3 cell strains were mounted up to 2.3 folds, 3.7 folds, and 4.3 folds after 48 hours, respectively. Under the exposure of FLT3 inhibitor AC220, the IC50 values was 0.183, 0.446 and 0.836 nmol/L, and apoptosis rates was 88.6%, 34.2% and 16.1%, respectively.
Conclusion: FLT3-ITD mutant expressed cell strains with longer ITD show higher capacity of proliferation and higher tolerance to AC220 treatment.
题目: FLT3-ITD插入长度对32D细胞增殖、凋亡及靶向抑制剂敏感性的影响.
目的: 研究FLT3-ITD突变体ITD插入长度对细胞增殖能力、凋亡及靶向抑制剂敏感性的影响，为临床分级治疗FLT3-ITD突变型急性髓系白血病提供数据参考.
方法: 从先前解析出的ITD序列中挑选3个位置相同或相邻的不同大小的ITD序列，构建FLT3-ITD真核表达质粒；利用慢病毒包被系统建立FLT3-ITD稳定表达32D细胞株；检测FLT3-ITD细胞株细胞增殖情况，经靶向抑制剂AC220处理后的其增殖抑制及凋亡情况.
结果: 成功构建了ITD1、ITD2和ITD3 三株FLT3-ITD稳定表达32D细胞株；ITD1、ITD2和ITD3细胞株在无IL3因子培养基生长条件下48 h后的增殖倍数分别是2.3、3.7和4.3倍；在FLT3小分子抑制剂AC220加药条件下，ITD1、ITD2和ITD3细胞株增殖抑制IC50值分别是0.183、0.446和0.836 nmol/L，凋亡率分别是88.6%、34.2%和16.1.
结论: 插入ITD长度长的FLT3-ITD突变体表达细胞在增殖能力和耐受靶向抑制剂AC220的能力方面均增强.