Synergistic Anti-tumor Activity and Mechanism of Ibrutinib Combined with BH3 Analogue ABT737

OBJECTIVE To investigate the synergistic anti-tumor effect and mechanism of ibrutinib combined with BH3 analogue ABT737. METHODS Human non-small cell lung cancer cell lines A549 and H1299, and glioma cells U251 and U87 were used as subjects. SRB method was used to detect cell proliferation after 24 h of single or combined action of different concentrations of ibutinib(20, 15, 10, 7.5, 5 μmol·L^-1) and BH3 analogues ABT737(20, 15, 10, 7.5, 5 μmol·L^-1), cell survival rate and combined index was calculated. The clone formation assay was used to detect the characteristic of stemness after the combination treatment of 10 μmol·L^-1 ibrutinib and 10 μmol·L^-1 ABT737 for 120 h. Cell spheroidization test was used to investigate the effect of the combination treatment of two drugs on cell spheroidization ability for 7 d. PI staining together with flow cytometry was used to detected effect of the combination treatment of 10 μmol·L^-1ibrutinib and 10 μmol·L^-1ABT737 on apoptosis of U87 and U251 cells. RT-PCR was used to detect the effect of the combination treatment of the two drugs on the mRNA levels of stem cell markers Sox2, Nanog, and Oct4 in U87 and U251 cells for 24 h. Western blotting was used to detect the expression of brain glioma stem cell marker CD44 protein. RESULTS Different concentrations of ibrutinib and BH3 analogue ABT737 had synergistic effects in non-small cell lung cancer and glioma, and the combined group had a higher proliferation inhibition rate than the single group(P<0.05). The combination of two drugs could inhibit the ability of cell clone formation and synergistically induce cell apoptosis. At the same time, the combination of the two drugs could inhibit the mRNA expression level of tumor stem cell marker Sox2 and reduce the expression level of CD44 protein. CONCLUSION Ibrutinib combined with ABT737 can synergistically inhibit the proliferation of solid tumor cells and promote apoptosis, the mechanism is possibly by inhibiting the expression of tumor stem cell proteins.