Study On The Role Of ZCCHC10 Gene In Acute Myeloid Leukemia

Background:ZCCHC10(zinc finger CCHC-containing 10)gene is located on chromosome 5q31.1.It has been reported that ZCCHC10 is deleted or methylated in lung cancer and acute myeloid leukemia(AML),suggesting that this gene is related to lung cancer and AML.Our laboratory has demonstrated that ZCCHC10 protein maintains the stability of p53 protein by inhibiting MDM2(murine double minute 2)-mediated p53 ubiquitination,and down-regulation of this gene can promote lung cancer growth and metastasis.But the role of ZCCHC10 in other cancers has not been reported.This paper aimed to study the role of ZCCHC10 in AML.Methods:The Oncomine database was used to analyze the expression differences of ZCCHC10 in bone marrow samples of AML patients and normal humans.Quantitative realtime PCR technology was used to detect the level of ZCCHC10 m RNA in bone marrow samples and cell lines of AML patients.Western blotting was used to detect the expression of proteins such as ZCCHC10.Lentivirus-mediated expression system was used to construct a stable cell line that overexpressed or interfered with ZCCCHC10.The cell viability was analyzed with CCK-8 kit.The cell clone formation ability was analyzed with methylcellulose medium,and the cell apoptosis rate was analyzed with cell flow cytometry.The constructed stable cell line was injected into SCID/NOD mice by tail vein injection,and then overall survival and leukemia burden were investigated.Results:1.Both Oncomine database analysis and clinical sample analysis showed that ZCCHC10 was low in AML.Except for OCI-AML5,the expression of ZCCHC10 protein in other AML cell lines was lower than that of human normal embryonic kidney cells HEK293,and ML2 cells hardly express ZCCHC10.Treatment with the methyltransferase inhibitor5-azacytidine significantly increased the expression level of ZCCHC10 in AML cell,indicating that the methylation of the ZCCHC10 gene promoter in AML cell is one of the reasons for the low expression of ZCCHC10.2.This study successfully constructed ML2 and OCI-AML3 stable cell lines that overexpress ZCCHC10 and OCI-AML3 stable cell lines that interfere with ZCCHC10.After overexpression of ZCCHC10,cell viability and clonal formation ability were significantly reduced,and cell apoptosis rate was significantly increased;on the contrary,after knockdown of ZCCHC10,cell viability and clonal formation ability increased,and apoptosis rate decreased.3.In AML cells,overexpression of ZCCHC10 increased the levels of p53,p21,and Bax proteins but decreased the level of bcl-2 protein.On the contrary,knockdown of ZCCHC10 decreased the protein levels of p53,p21,and Bax,but increased Bcl-2 protein level.4.The results of the AML xenograft mouse model showed that overexpression of ZCCHC10 can inhibit splenomegaly and the proliferation of leukemia cells in the bone marrow and spleen,and prolonged the survival time of the mice;and knockdown of ZCCHC10 led to splenomegaly and proliferation of AML in bone marrow and spleen,and shortened the survival time of the mice.Conclusion:1.ZCCHC10 gene is down-expressed in AML,and the promoter methylation of this gene may contribute to its down-regulation.2.ZCCHC10 can inhibit the viability and clonal formation ability of AML cells,and promote the apoptosis of AML cells.3.ZCCHC10 can up-regulate the expression of p53,p21 and Bax,and down-regulate the expression of Bcl-2.4.ZCCHC10 can inhibit the proliferation and infiltration of AML cells in the bone marrow and spleen of mice,and prolong the survival time of AML mice.In short,ZCCHC10 plays a tumor suppressive effect by activating the p53 signaling pathway in acute myeloid leukemia.