The Effect Of A Novel Histone Deacetylase Inhibitor Valproic Acid In Regulating Proliferation And Apoptosis In Multiple Myeloma KM3 Cells In Vitro

Background:Despite of the recent advances in the treatment of multiple myeloma(MM),it is still an incurable disease in the majority of patients.In the bone marrow microenvironment of MM,VEGF is essential for tumor growth and survival, and myeloma cells are known to produce VEGF and express VEGF receptors. Binding of VEGF₁₆₅to MM cells triggers VEGFR-1 tyrosine phosphorylation. Subsequently the downstream signaling pathways are activated,as follows:(1) PI3-kinase/protein kinase Cα(PKCα)-dependent cascade mediating MM cell migration on fibronectin,(2)MEK-extracellular signal-regulated protein kinase(ERK) pathway mediating MM cell proliferation.Now histone deacetylase inhibitors(HDAIs) have been regarded as a new kind of anticancer drug and several of them are currently being investigated in clinical trials.Valproic acid(VPA)is a shortchain fatty acid with a long history of clinical use as an anticonvulsant.However,VPA also inhibits histone deacetylases(HDACs)and induces apoptosis in selected solid tumors as well as in hematologic neoplasms.Objective:To define the effects of VPA on multiple myeloma cell line KM3 cells in vitro and the underlying mechanism of VPA in the treatment of MM were investigated.Methods:KM3 cells were cultured in RPMI1640 medium and treated with VPA in different concentration.The cell proliferation was measured by 3-[4, 5-dimehyl-2-thiazolyl]-2,5-diphenyl-2H-tetrazolium bromide(MTT)assay;the Annexinâ…¤and PI staining was used to detect the apoptosis rates;the mRNA level of VEGFR was determined by RT-PCR;and immunocytochemistry was used to detect the protein level of acetylated-histone H4(Ac-histone H4)and VEGFR.Results:The proliferation rate of KM3 cells was decreased in VPA- treated group in a time-dose dependent manner(P＜0.05).And the Annexinâ…¤and PI staining showed VPA can induce apoptosis of KM3 cells,treatment with VPA(0.5,1,2 and 4mmol/L) for 48h,the apoptosis rates of KM3 cells were(11.77±4.64)%,(22.13±1.20)%,(23.95±.2.57)%and(42.72±4.61)%respectively.The RT-PCR results demonstrated the KM3 cells only express VEGFR-1 and it was decreased in VPA(0.5,1,2 and 4 mmol/L for 48h)treated groups compared with control(P＜0.05).Results of immunocytochemistry show the protein of Ac-histone H4 was increased significantly in VPA(4 mmol/L for 48h)treated group whereas the VEGFR-1/Flt-1 was decreased (P＜0.05).Conclusion:VPA,as an histone deacetylase inhibitor,can increase the expression of Ac-histone H4 and inhibit the expression of VEGFR-1 in KM3 cells and it plays an important role in regulating proliferation and apoptosis of multiple myeloma cell line KM3 cells in vitro.