Mechanism of gemcitabine enhances ABT-199 induced apoptosis in Philadelphia chromosome-positive acute lymphoblastic leukemia:a preliminary study / 白血病·淋巴瘤

ABSTRACT

Objective To investigate the effects of gemcitabine and ABT-199 on proliferation inhibition and apoptosis induction of Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL) cell line SUP-B15, and to explore its synergistic mechanism. Methods SUP-B15 cells in logarithmic growth phase were treated with gemcitabine (0.025 and 0.050 μmol/L), ABT-199 (0, 0.5, 1.0, 2.0, 4.0, 8.0 μmol/L) or two drugs for 24 h. Cell proliferation was detected by CCK-8 method, apoptosis was detected by flow cytometry (FCM), mitochondrial membrane potential was detected by JC-1 method, and expression of mitochondrial apoptosis pathway-related protein was analyzed by Western blot. Results The 50 ％ inhibitory concentration (IC50) of SBT-B15 cells treated with ABT-199 for 24 h was (4.13±0.89) μmol/L. However, gemcitabine (0.025, 0.050 μmol/L) significantly enhanced the inhibitory effect of ABT-199 on proliferation of SUP-B15 cells, the IC50 values were (2.23 ±0.73) and (1.15 ±0.45) μmol/L, respectively. The results of FCM assay showed that compared with the monotherapy group [(7.33±1.54)％], 0.025 umol/L gemcitabine combined with ABT-199 (1.0 and 2.0 μmol/L) acted on SUP-B15 cells for 24 h, the proportions of apoptotic cells were (32.42±1.45) ％and (44.33±1.86) ％, the difference was statistically significant (F=70.78, P<0.001);compared with the monotherapy group [(9.60 ±2.76) ％], 0.05 μmol/L gemcitabine combined with ABT-199 (1.0 and 2.0 μmol/L) acted on SUP-B15 cells for 24 h, the proportion of apoptotic cells increased to (47.63 ± 3.81) ％ and (58.73 ±4.33) ％, respectively, and the difference was statistically significant (F= 79.21, P<0.001). The JC-1 experiment showed that treated with ABT-199 and gemcitabine for 12 h, the percentage of depolarizing cell was significantly higher than that in single agent group, and the difference was statistical significant (P<0.001). Western blot showed that the anti-apoptotic proteins bcl-2, bcl-xL and Mcl-1 decreased after treated by gemcitabine combined with ABT-199 for 12 h. Conclusion Gemcitabine could enhance the proliferation inhibition and induce apoptosis of Ph+ALL cells by ABT-199, and its mechanism may be related to down-regulation of anti-apoptosis-related proteins.