title: Global promoter methylation analysis reveals novel candidate tumor suppressor genes in natural killer cell lymphoma.
abstract: OBJECTIVE
  To identify tumor suppressor genes epigenetically silenced by promoter hypermethylation in extranodal natural killer cell lymphoma (NKCL).
  
  
  METHODS
  Promoter methylation was analyzed with global and locus-specific methylation assays in NKCL cases and NK cell lines. Gene expression profiles were used to identify genes for which aberrant promoter methylation was associated with transcriptional silencing. Selected DNA methylations were validated by RRBS, pyrosequencing, or q-MSP. Decitabine treatment was performed to evaluate reactivation of methylated genes. The tumor suppressor effect of silenced genes was evaluated functionally by reintroducing them into NK cell lines.
  
  
  RESULTS
  We observed significant promoter hypermethylation in most NKCL samples compared with normal NK cells. Correlation of global promoter methylation with gene expression profiles identified 95 genes with strong evidence for being silenced because of promoter methylation, including BCL2L11 (BIM), DAPK1, PTPN6 (SHP1), TET2, SOCS6, and ASNS. Known tumor suppressor genes were significantly overrepresented in this set of genes. Decitabine treatment of NK cell lines was associated with reexpression of all 10 selected methylated and silenced genes. Ectopic expression of frequently silenced BIM in two BIM-nonexpressing NK cell lines led to increased apoptosis and eventual elimination of BIM-transduced cells. It also sensitized these cell lines to chemotherapy-induced apoptosis. Similarly, reintroduction of SOCS6 significantly inhibited growth in SOCS6-nonexpressing NK cell lines. NK cell lines lacking ASNS expression showed increased sensitivity to treatment with l-asparaginase. Reintroduction of ASNS reduced drug sensitivity.
  
  
  CONCLUSIONS
  Promoter region hypermethylation is frequent in NKCL, and aberrantly methylated genes are pathologically and clinically significant.
authors.affiliation: Department of Pathology, City of Hope National Medical Center, Duarte, California.
authors.forename: Can
authors.initials: C
authors.lastname: Küçük
authors.affiliation: Department of Pathology, City of Hope National Medical Center, Duarte, California. Department of Clinical Laboratory, Peking University Third Hospital, Beijing, China.
authors.forename: Xiaozhou
authors.initials: X
authors.lastname: Hu
authors.affiliation: Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska.
authors.forename: Bei
authors.initials: B
authors.lastname: Jiang
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, Nebraska.
authors.forename: David
authors.initials: D
authors.lastname: Klinkebiel
authors.affiliation: Department of Laboratory Medicine, University of California San Francisco School of Medicine, San Francisco, California.
authors.forename: Huimin
authors.initials: H
authors.lastname: Geng
authors.affiliation: Department of Pathology, City of Hope National Medical Center, Duarte, California.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Gong
authors.affiliation: Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska.
authors.forename: Alyssa
authors.initials: A
authors.lastname: Bouska
authors.affiliation: Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska.
authors.forename: Javeed
authors.initials: J
authors.lastname: Iqbal
authors.affiliation: Département de Pathologie, Groupe Henri-Mondor Albert-Chenevier, Inserm U955, Université Paris Est, Créteil, France.
authors.forename: Philippe
authors.initials: P
authors.lastname: Gaulard
authors.affiliation: Department of Pathology, City of Hope National Medical Center, Duarte, California.
authors.forename: Timothy W
authors.initials: TW
authors.lastname: McKeithan
authors.affiliation: Department of Pathology, City of Hope National Medical Center, Duarte, California. jochan@coh.org.
authors.forename: Wing C
authors.initials: WC
authors.lastname: Chan
mesh_terms: D019175:DNA Methylation / Q000235:genetics*; D016147:Genes, Tumor Suppressor; D006801:Humans; D054391:Lymphoma, Extranodal NK-T-Cell / Q000235:genetics*; D011401:Promoter Regions, Genetic; D059467:Transcriptome