Chidamide Inhibits Autophagy And Induces DNA Damage Response-related Apoptosis In Myeloma Cells

Bankground and Aim:It is increasingly apparent that tumor development depents not only on stable genetic alternations,but also on epigenetic changs,which can influence gene expression patterns.Epigenetic regulators such as histone acetyltransferase(HATs) and histone acetylase(HDACs) play an important role in gene expression.Modification of histone by acetylation affects transcription by changing chromatin condensation that modulates the accessibility of transcription factors to their target DNA and these alternation influence gene transcription.Overall,histone acetylation is being investigated as a therapeutic target.Two HDACi,vorinostat and romidepsin were FDA approved for use against refractory cutaneos T-cell lymphoma.Multiple myeloma is a malignant disease characterized by proliferation of clonal plasma cells in the bone marrow.Despite several approaches(such as proteasome inhibitors 、 immunomodulators and high-dose chemotherapy with Autologous Stem Cell(ASCT))have been applied in an effort to confront MM during the past few decades,chemoresistance that may arise during chemotherapy is one of the main causes of failure treatment.Chidamide is a new benzamide class of histone deacetylase inhibitor with marked anti-tumor activity,whichwas generally well tolerated in patients with advanced solid tumors or lymphomas in clinic phase Ⅰ tests.During numerous studies,Chidamide has exhibited significant clinical activity agaist lung, colorectal, esophageal, breast, gastric, prostate cancers,while the functions to inhibit or kill MM cells have not been found. In this study,we try to explore the effects of chidamide on MM cells and the related mechanisms.Methods:①We performed MTT assay to evaluate the proliferation of these two MM cell lines in the absence or presence of Chidamide. ② Cell apoptosis and cell cycle distribution were analyzed by flow cytometry. ③ The expression of SIRT1 and autophagy-related m RNA were detected by RT-PCR.④Western-blot was used to explore the protein expressions of p21、γH2AX、p ATM 、Cleaved Caspase-3、SIRT1 and H4K16 ac.⑤KU55933 inhibited the activity of ATM kinase.Results:①Chidamide reduced the viability of MM cells in a dose- and time-dependent manner. ②Chidamide arrested cells at G0/G1 phase.③ RT-PCR showed:The m RNA of SIRT1(class Ⅲ HDAC)、 ATG7 and LC3 decreased. ④Western-blot showed:Chidamide suppressed the expression of SIRT1(class Ⅲ HDAC) and increased histone H4 acetylation levels;The amount of autophagic proteins ATG7 and LC3 decreased;Chidamide effectively upregulated the protein expression of p21、γH2AX、p ATM and Cleaved Caspase-3. ⑤Pretreatment of MM cells with KU55933 suppressed γH2AX、p ATM and Cleaved Caspase-3 protein levels partly controled with chidamide singly,which indicated that the apoptosis of MM cells induced by Chidamide involved DNA damage responseConclusion:①The apoptosis of MM cells induced by Chidamide involved DNA damage response; ② Chidamide suppressed the expression of SIRT1(class Ⅲ HDAC) and increased histone H4 acetylation levels;③Chidamide effectively inhibited the process of autophagy in MM cells.