Li Li, Piloto Obdulio, Kim Kyu-Tae, Ye Zhaohui, Nguyen Ho Bao, Yu Xiaobing, Levis Mark, Cheng Linzhao, Small Donald
Department of Oncology, Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD, USA.
Br J Haematol. 2007 Apr;137(1):64-75. doi: 10.1111/j.1365-2141.2007.06525.x.
Activating mutation of FLT3 by internal tandem duplications (ITDs) in the juxtamembrane region is the most common molecular aberration found in acute myeloid leukaemia (AML). In this study, a lentiviral vector containing two promoters achieved consistent and efficient co-expression of FLT3/ITD and GFP in transduced human CD34(+) haematopoietic stem/progenitor cells (HSPCs). When cultured in medium containing stem cell factor, thrombopoietin and FLT3 ligand (FL), FLT3/ITD-transduced cells demonstrated enhanced self-renewal and survival potential, unaffected by the withdrawal of FL. These cells retained a CD34(+)CD38(-/dim) immunophenotype, typical of HSPCs. Compared to cells transduced with a vector expressing GFP alone, FLT3/ITD-transduced HSPCs had a higher fraction of cells in active cell cycle. FLT3/ITD-transduced HSPCs were more sensitive to the induction of cytotoxicity by CEP-701, a selective FLT3 inhibitor, indicating a rapid 'addiction' to signalling through this oncogenic pathway. The FLT3/ITD-transduced HSPCs showed increased expression of Pim-1, c-Myc and Cyclin D3 (CCND3), each of which may contribute to the altered genetic programme instituted by FLT3/ITD signalling. Taken together, these results indicate that FLT3/ITD mutations may contribute to leukaemic transformation of normal HSPCs by prolonging survival, promoting proliferation and partially blocking differentiation. CEP-701 may act as a potent therapeutic agent for AML stem cells harbouring FLT3/ITD mutations.
近膜区内部串联重复（ITD）导致的FLT3激活突变是急性髓系白血病（AML）中最常见的分子畸变。在本研究中，一种含有两个启动子的慢病毒载体在转导的人CD34(+)造血干/祖细胞（HSPCs）中实现了FLT3/ITD和GFP的一致且高效的共表达。当在含有干细胞因子、血小板生成素和FLT3配体（FL）的培养基中培养时，转导了FLT3/ITD的细胞表现出增强的自我更新和存活潜力，不受FL撤除的影响。这些细胞保留了HSPCs典型的CD34(+)CD38(-/dim)免疫表型。与仅转导表达GFP载体的细胞相比，转导了FLT3/ITD的HSPCs处于活跃细胞周期的细胞比例更高。转导了FLT3/ITD的HSPCs对选择性FLT3抑制剂CEP-701诱导的细胞毒性更敏感，表明通过这种致癌途径的信号传导存在快速的“成瘾性”。转导了FLT3/ITD的HSPCs显示出Pim-1、c-Myc和细胞周期蛋白D3（CCND3）的表达增加，其中每一种都可能促成由FLT3/ITD信号传导引发的遗传程序改变。综上所述，这些结果表明FLT3/ITD突变可能通过延长存活、促进增殖和部分阻断分化来促成正常HSPCs的白血病转化。CEP-701可能作为一种有效的治疗剂用于治疗携带FLT3/ITD突变的AML干细胞。
