title: Early B-cell factor 3 (EBF3) is a novel tumor suppressor gene with promoter hypermethylation in pediatric acute myeloid leukemia.
abstract: BACKGROUND
  Pediatric acute myeloid leukemia (AML) comprises up to 20% of all childhood leukemia. Recent research shows that aberrant DNA methylation patterning may play a role in leukemogenesis. The epigenetic silencing of the EBF3 locus is very frequent in glioblastoma. However, the expression profiles and molecular function of EBF3 in pediatric AML is still unclear.
  
  
  METHODS
  Twelve human acute leukemia cell lines, 105 pediatric AML samples and 30 normal bone marrow/idiopathic thrombocytopenic purpura (NBM/ITP) control samples were analyzed. Transcriptional level of EBF3 was evaluated by semi-quantitative and real-time PCR. EBF3 methylation status was determined by methylation specific PCR (MSP) and bisulfite genomic sequencing (BGS). The molecular mechanism of EBF3 was investigated by apoptosis assays and PCR array analysis.
  
  
  RESULTS
  EBF3 promoter was hypermethylated in 10/12 leukemia cell lines. Aberrant EBF3 methylation was observed in 42.9% (45/105) of the pediatric AML samples using MSP analysis, and the BGS results confirmed promoter methylation. EBF3 expression was decreased in the AML samples compared with control. Methylated samples revealed similar survival outcomes by Kaplan-Meier survival analysis. EBF3 overexpression significantly inhibited cell proliferation and increased apoptosis. Real-time PCR array analysis revealed 93 dysregulated genes possibly implicated in the apoptosis of EBF3-induced AML cells.
  
  
  CONCLUSIONS
  In this study, we firstly identified epigenetic inactivation of EBF3 in both AML cell lines and pediatric AML samples for the first time. Our findings also showed for the first time that transcriptional overexpression of EBF3 could inhibit proliferation and induce apoptosis in AML cells. We identified 93 dysregulated apoptosis-related genes in EBF3-overexpressing, including DCC, AIFM2 and DAPK1. Most of these genes have never been related with EBF3 over expression. These results may provide new insights into the molecular mechanism of EBF3-induced apoptosis; however, further research will be required to determine the underlying details. Our findings suggest that EBF3 may act as a putative tumor suppressor gene in pediatric AML.
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. taoyanfang1982@163.com.
authors.forename: Yan-Fang
authors.initials: YF
authors.lastname: Tao
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. xulixiao2013@hotmail.com.
authors.forename: Li-Xiao
authors.initials: LX
authors.lastname: Xu
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. drlujun_sz@163.com.
authors.forename: Jun
authors.initials: J
authors.lastname: Lu
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. hsy139@126.com.
authors.forename: Shao-Yan
authors.initials: SY
authors.lastname: Hu
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. baseff77@gmail.com.
authors.forename: Fang
authors.initials: F
authors.lastname: Fang
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. cl2012@sohu.com.
authors.forename: Lan
authors.initials: L
authors.lastname: Cao
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. xiaopfdr@gmail.com.
authors.forename: Pei-Fang
authors.initials: PF
authors.lastname: Xiao
authors.affiliation: Department of Gastroenterology, the 5th Hospital of Chinese PLA, Yin chuan, China. du_xiaojuan123@163.com.
authors.forename: Xiao-Juan
authors.initials: XJ
authors.lastname: Du
authors.affiliation: Department of Cell and Molecular Biology, Cancer Institute (Hospital), Chinese Academy of Medical Sciences, Peking Union Medical College, Beijing, China. sunlichao_1980@hotmail.com.
authors.forename: Li-Chao
authors.initials: LC
authors.lastname: Sun
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. szlizhiheng@hotmail.com.
authors.forename: Zhi-Heng
authors.initials: ZH
authors.lastname: Li
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. 570843000@qq.com.
authors.forename: Na-Na
authors.initials: NN
authors.lastname: Wang
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. sky2skeleton@163.com.
authors.forename: Guang-Hao
authors.initials: GH
authors.lastname: Su
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. lyh072006@hotmail.com.
authors.forename: Yan-Hong
authors.initials: YH
authors.lastname: Li
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. ligangmailbox@hotmail.com.
authors.forename: Gang
authors.initials: G
authors.lastname: Li
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. 623863179@qq.com.
authors.forename: He
authors.initials: H
authors.lastname: Zhao
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. ray8302880@163.com.
authors.forename: Yi-Ping
authors.initials: YP
authors.lastname: Li
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. rdxyy@163.com.
authors.forename: Yun-Yun
authors.initials: YY
authors.lastname: Xu
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. huitingzhousdfey@163.com.
authors.forename: Hui-Ting
authors.initials: HT
authors.lastname: Zhou
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. 170330856@qq.com.
authors.forename: Yi
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. sunnysnow12@163.com.
authors.forename: Mei-Fang
authors.initials: MF
authors.lastname: Jin
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. flykf@163.com.
authors.forename: Lin
authors.initials: L
authors.lastname: Liu
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. xueming_zhu@aliyun.com.
authors.forename: Xue-Ming
authors.initials: XM
authors.lastname: Zhu
authors.affiliation: Translational Research Center, Second Hospital, The Second Clinical School, Nanjing Medical University, Nanjing, China. ni_jian2008@163.com.
authors.forename: Jian
authors.initials: J
authors.lastname: Ni
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. wj196312@vip.163.com.
authors.forename: Jian
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. xing_feng66@hotmail.com.
authors.forename: Feng
authors.initials: F
authors.lastname: Xing
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. wenlizhao69@163.com.
authors.forename: Wen-Li
authors.initials: WL
authors.lastname: Zhao
authors.affiliation: Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China. panjian2008@163.com.
authors.forename: Jian
authors.initials: J
authors.lastname: Pan
mesh_terms: D000293:Adolescent; D000367:Age Factors; D017209:Apoptosis / Q000235:genetics; D045744:Cell Line, Tumor; D002648:Child; D002675:Child, Preschool; D016000:Cluster Analysis; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D020869:Gene Expression Profiling; D015973:Gene Expression Regulation, Leukemic; D016147:Genes, Tumor Suppressor; D018922:HL-60 Cells; D006801:Humans; D053208:Kaplan-Meier Estimate; D015470:Leukemia, Myeloid, Acute / Q000175:diagnosis / Q000235:genetics* / Q000401:mortality; D008297:Male; D011379:Prognosis; D011401:Promoter Regions, Genetic; D015398:Signal Transduction; D014157:Transcription Factors / Q000235:genetics*
chemical_list: C000605678:EBF3 protein, human; D014157:Transcription Factors