新酪氨酸激酶抑制剂BGJ398对白血病细胞株KG-1细胞的抑制效应研究

Inhibitory Eefects of the novel tyrosine kinase inhibitor BGJ398 against human leukemic cell line KG- 1 cells

【摘要】目的探讨针对FGFRl的酪氨酸激酶抑制剂BGJ398对人急性髓系白血病细胞株KG-1细胞的影响及可能的作用机制。方法CCK-8法检测BGJ398对KG-1细胞增殖的影响；流式细胞术AnnexinV／PI双重染色法检测BGJ398对细胞凋亡的影响；RQ．PCR检测细胞凋亡相关基因的表达；Westernblot法检测凋亡相关蛋白、FGFRlOP2-FGFRl融合蛋白及信号通路分子磷酸化水平的表达变化。结果BGJ398能有效抑制KG-1细胞增殖，抑制率呈剂量依赖性升高，并能诱导细胞凋亡。BGJ398作用KG．1细胞48h后，与对照组比较凋亡相关基因Bcl。2表达下降（0．342±0．054对1．026±0．165，t=3．94，P=0．017），caspase．3表达上调（0．456±0．189对1．008±0．091，t=16．44，P〈0．001），差异均有统计学意义。与对照组相比，BGJ398作用组caspase-3活化蛋白表达增加，同时Bcl-2蛋白表达下凋；FGFRlOP2-FGFRl融合蛋白水平及AKT、S6K1磷酸化水平下捌，差异均有统计学意义∽值均〈0．01），但ERK磷酸化表达水平无明显改变。结论BGJ398能有效抑制KG-1细胞增殖并诱导细胞凋亡，其机制可能与抑制FGFR1表达、下调Bcl-2水平、促进Caspase-3活化及抑制AKT和S6K磷酸化有关。

Objective To explore the effects and possible mechanisms of the novel pan-FGFR inhibitor BGJ398 on KG-1 cells in vitro. Methods Effects of BGJ398 on cells proliferation were detected by CCK-8, the apoptosis was assessed by Annexin V-FITC. Reverse transcriptionquantitative polymerase chain reaction （q-PCR） analysis was used to detect the expression of apoptosis-related genes B cell lymphoma-2 （Bcl-2） and caspase-3. Western blotting analysis was performed to explore the proteins expression levels of Bcl-2, caspase-3 and the expression of p-AKT, p-S6K, p-ERK and FGFRI. Results BGJ398 effectively inhibited cell proliferation by dose-dependent manners. BGJ398（1.4 p.mol/L） induced apoptosis of KG-1 cells by 36.4%, compared with 4.5% in the control group（P〈 0.001）. Treatment with BGJ398 at 1.4 μmol/L led to significant increases in the expression levels of caspase-3, and decreases in the expression of Bcl-2 （P 〈 0.005）. In accordance with these results, Western blot analysis further confirmed the increased expression of Bcl-2 protein along with elevated caspase-3 activity. In addition, BGJ398 markedly down-regulated FGFR1OP2-FGFR1 fusion protein, p-AKT and p-S6K expression, but not p- ERK expression. Conclusion Novel pan-FGFR inhibitor BGJ398 substantially suppressed KG- 1 cell growth and induced apoptosis by inhibiting the expression of FGFR1, p-AKT, p-S6K and regulating apoptosis-related proteins.