title: Methylation of miR-34a, miR-34b/c, miR-124-1 and miR-203 in Ph-negative myeloproliferative neoplasms.
abstract: BACKGROUND
  MicroRNA (miR) miR-34a, -34b/c, -124-1 and -203 are tumor suppressor miRs implicated in carcinogenesis.
  
  
  METHODS
  We studied DNA methylation of these miRs in Philadelphia-negative (Ph-ve) myeloproliferative neoplasms (MPNs). Methylation-specific PCR (MSP), verified by direct sequencing of the methylated MSP products, was performed in cell lines, normal controls and diagnostic marrow samples of patients with MPNs.
  
  
  RESULTS
  Methylation of these miRs was absent in the normal controls. miR-34b/c were homozygously methylated in HEL cells but heterozygously in MEG-01. In HEL cells, homozygous miR-34b/c methylation was associated with miR silencing, and 5-aza-2'-deoxycytidine treatment led to re-expression of both miR-34b and miR-34c, consistent with that both miRs are under the regulation of the same promoter CpG island. miR-34a was heterozygously methylated in MEG-01 and K-562. miR-203 was completely unmethylated in K-562 and SET-2 but no MSP amplification was found in both HEL and MEG-01, suggestive of miR deletion. In primary samples, four each had miR-34b/c and -203 methylation, in which two had concomitant methylation of miR-34b/c and -203. miR-34a was methylated in one patient and none had methylation of miR-124-1. Seven patients (15.6%) had methylation of at least one of the four miRs. miR methylation did not correlate with clinical parameters, disease complications or JAK2 V617F mutation.
  
  
  CONCLUSIONS
  This is the first report of miR hypermethylation in MPNs. miR-203 hypermethylation is not specific to Ph+ve leukemias but also present in Ph-ve MPNs. miR-34b/c methylation was associated with reversible miR silencing. There was no correlation of miR methylation with clinical demographic data or outcome.
authors.affiliation: Department of Medicine, Queen Mary Hospital, The University of Hong Kong, Hong Kong. jcschim@hku.hk
authors.forename: Chor Sang
authors.initials: CS
authors.lastname: Chim
authors.forename: Thomas S
authors.initials: TS
authors.lastname: Wan
authors.forename: Kwan Yeung
authors.initials: KY
authors.lastname: Wong
authors.forename: Tsz Kin
authors.initials: TK
authors.lastname: Fung
authors.forename: Hans G
authors.initials: HG
authors.lastname: Drexler
authors.forename: Kit Fai
authors.initials: KF
authors.lastname: Wong
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001374:Azacitidine / Q000494:pharmacology; D001483:Base Sequence; D019046:Bone Marrow Neoplasms / Q000235:genetics*; D016022:Case-Control Studies; D045744:Cell Line, Tumor; D019175:DNA Methylation / Q000187:drug effects / Q000235:genetics*; D003710:Demography; D005260:Female; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism*; D008875:Middle Aged; D008969:Molecular Sequence Data; D009196:Myeloproliferative Disorders / Q000235:genetics*; D010677:Philadelphia Chromosome
chemical_list: D035683:MicroRNAs; D001374:Azacitidine
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