The Mechanism Of The Effect Of HDACi Combined With FLT3i On FLT3-ITD(+) Acute Myelocytic Leukemia Cell Lines MV4-11 And MOLM-13

Objective: This study was conducted to investigate whether histone deacetylase inhibitor(HDACi)combined with FLT3(FMS-like tyrosine kinase3)inhibitor can synergistically inhibit the proliferation of AML(Acute myeloid leukemia)cells with FLT3-ITD(Internal tandem duplication)and explore its related mechanisms,aiming to provide a preclinical basis for obtaining a more proven drug treatment modality for AML patients with FLT3-ITD mutation.Methods: Chidamide and quizartinib were administered to MV4-11 and MOLM-13 cell lines at different concentrations alone or in combination for 48 h.The following assays were performed: 1.CCK8 to detect cell proliferation ability;Compusyn 1.0 software was used to analyze the synergistic effect of the combination of the two drugs;2.The apoptosis rate and proliferation cycle of each group were detected by flow cytomety.3.The expression levels of FLT3,p-FLT3,P53,BCL-2,Bax and p-AKT proteins were detected by Western-blotting.All assays needed to be repeated at least three times,and statistical analysis of experimental results were performed using Graphpad Prism 8.4.2 software.The comparison between the two groups was performed by T-test,and the results were expressed as mean ± standard deviation(x ±s,%).p<0.05 was considered statistically significant,and p<0.01 was considered statistically significant.Results: 1.Chidamide and quizartinib had proliferation inhibitory effects on MV4-11 and MOLM-13 cell lines in a dose-dependent manner by CCK-8assay;chidamide combined with quizartinib had synergistic proliferation inhibitory effects on MV4-11 and MOLM-13 cell lines.2.Flow cytometry showed that chidamide and quizartinib alone could promote the apoptosis and block the cell cycle of MV4-11 and MOLM-13 cell lines.In a certain concentration range,chidamide combined with quizartinib make more effects.3.Western blot showed that compared with the blank control group,p-FLT3 was down-regulated in the quizartinib-containing group.P53 and Bax pro-apoptotic proteins were up-regulated,anti-apoptotic protein BCL-2 and p-AKT,a key downstream factor of PI3K/AKT signaling pathway were down-regulated in the three groups.Effects of protein P53,Bax,BCL-2 and p-AKT were most pronounced in the two-drug combination group.Conclusion: The combination of chidamide and quizartinib can promote the apoptosis and block the cell cycle of the AML cell,resulting in the suppression of the FLT3-ITD(+)AML cell,mainly through the downregulation of BCL-2 and p-AKT expression and the upregulation of P53 and Bax proteins.