Studies Of Effect And Mechanisms Of Benzamide Compound W-H4 On Acute Myeloid Leukemia In Vitro

SNX-2112 is a tetrahydro-indazol-substituted benzamide which targets Hsp90. In order to enhance its anti-tumor activity,we have synthesized a series of tetrahydro- indazol-substituted benzamide compounds. And we found W-H4 has greater anti-tumor activity in many human cancer cell lines. In this study, we investigated the in vitro inhibitory activity of W-H4 on the growth ofacute myeloid leukemia cells. This study provides a theory basis for the clinical application of W-H4 in the treatment of acute myeloid leukemia.The in vitro inhibitory activities of W-H4 on the growth of two acute myeloid leukemia cell lines HL60 and KG-1a were evaluated by CCK8 assay; We investigated the effect of W-H4 on cell apoptosisand cyclethrough flow cytometry; Then We examined mitochondrial membrane potential changes in HL60 and KG-1a cells induced by W-H4 with JC-1 fluorescent probe; And we examined the effects of W-H4 on the expression of apoptosis-related proteins, G1/S checkpoint regulatory proteins, Hsp90 client proteins and autophagy-ralated proteins through Western blot assay.Our results indicatethat W-H4 is a potent compound against acute myeloid leukemiain vitro, showing greateractivity in inhibiting the proliferation of two acute myeloid leukemia cell lines in a dose-dependent manner and the IC50 values of W-H4 for HL60, KG-1a at 48 h were 0.48, 0.38 μM respectively. W-H4 induced cell apoptosis in both cell lines and we found the cleavage of caspase-3, caspase-8, caspase-9 and PARP in a dose-dependent way after W-H4 treatment for 48 h. And W-H4 can also induced mitochondrial dysfunction and changes of the level of Bcl-2. The cell cycle was blocked in the G1 phase after exposing to W-H4 accompanied with the changes of expression levels of cell cycle proteins. Additionally, W-H4 inhibited Akt/mTO R pathway and induced autophagy.Taken together, these results reveal W-H4 induce apoptosis through multiple signaling pathways in HL60, KG-1a cells and showing strong anti-acute myeloid leukemia activity in vitro.