Epigenetic Regulation Of DACT3 Expression And Its Clinical Significance In Acute Myeloid Leukemia

Acute myeloid leukemia(AML)is a clonal malignant proliferative disease of myeloid primordial cells in hematopoietic system,including all non lymphocytic acute leukemia.AML is the most common acute leukemia in adults.At present,chemotherapy is still the main method,but70% of patients will relapse and develop into refractory leukemia.Recent genetic analysis using next-generation sequencing(NGS)has greatly helped judge gene mutation in AML patients,and identify targets for AML therapy.Targeting epigenetic enzymes for AML therapies include inhibitors of DNMT,HDAC,LSD1,EZH2,DOT1 L,PRMT5,BET,KAT,MEN1-MLL,et al.However,some clinical trials of treating AML with epigenetic drugs alone have shown low response rates.The Wnt/beta-catenin pathway is required for the development of leukemia stem cells(LSC)in AML.In AML,abnormal activation of Wnt signal and downstream effectors has been confirmed.Wnt antagonists in AML,including s FRP1,s FRP2,s FRP4,s FRP5,DKK1 and DKK3,have different degrees of methylation and transcriptional inhibition.These molecules have been proved to be associated with poor prognosis and regulated by epigenetic drugs,suggesting that they may be involved in the pathogenesis of AML and are potential therapeutic targets.The human DACT gene family consists of three members,namely DACT1,DACT2 and DACT3.More and more studies have shown that DACT gene is down-regulated and promoter hypermethylation in a variety of solid tumors,which is a potential Wnt antagonist and tumor suppressor gene.However,the expression and role of DACT gene in AML are still unknown.Firstly,the present study comprehensively analyzed the expression of DACT gene in AML using RNA-seq,q RT-PCR Western blot technology and combined with TCGA database,identified the functional member DACT3,and explored its clinical significance.Then,the possible mechanism of down-regulation of DACT3 expression in AML was investigated from the two aspects that DNA methylation and histone acetylation.Finally,the biological function of DACT3 in AML was determined using AML cell model in vitro and animal experiment in vivo.The main experimental results are as follows:1.Expression characteristics of DACT3 in acute myeloid leukemia:Compared with DACT1 and DACT2,DACT3 was the most differentially expressed member in AML,suggesting that DACT3 plays a major role in AML.The m RNA and protein expression levels of DACT3 decreased significantly in AML,and the m RNA and protein levels were positively correlated.More importantly,the low expression of DACT3 is associated with poor prognosis in patients with AML.It is worth noting that,the m RNA level of DACT3 was low at new diagnosis,increased after complete remission,and decreased again at relapse.2.The possible mechanism of epigenetic regulation of DACT3 in acute myeloid leukemia:The down-regulation of DACT3 expression in AML is not entirely dependent on promoter methylation,but also affected by histone deacetylation.The class I HDACs play a major regulatory role in the expression of tumor suppressor gene DACT3.Further studies showed that the combination of epigenetic drug chidamide/azacytidine could increase DACT3 m RNA and protein levels,and promote AML cell apoptosis by inhibiting effective Wnt signal transduction in AML cell lines.3.Biological function of DACT3 in acute myeloid leukemia:DACT3 could improve the sensitivity of AML cells to adriamycin by promoting the inhibition of cell proliferation induced by adriamycin in different AML cell lines.The down-regulation of DACT3 expression in AML was related to the activation of Wnt signaling pathway,indicating that DACT3 is a negative regulator of Wnt signaling pathway in AML.In addition,DACT3 could also inhibit the proliferation of AML cells and the growth of subcutaneous transplanted tumors in severe immune deficiency(SCID)mice.DACT3 inhibited the growth of AML cells mainly through down-regulating the Wnt/β-catenin signaling pathway.In conclusion,this study found the down-regulation of DACT3 expression in AML and its correlation with the poor prognosis of AML for the first time,and revealed the epigenetic regulatory molecular mechanism of down-regulation of DACT3 expression.The study also showed that DACT3 promoted the apoptosis of AML cells and inhibited cell growth.These results not only deepen our understanding of the expression regulation mechanism and biological function of DACT3 as a tumor suppressor gene in AML,but also provide new clues for improving the treatment of AML,that is,DACT3 may be a potential target for AML treatment.This paper contains 35 figures,3 tables,143 references.