title: Infrequent DNA methylation of miR-9-1 and miR-9-3 in multiple myeloma.
abstract: OBJECTIVE
  The miR-9 family microRNAs (miRNAs) are tumour suppressor miRNAs implicated in carcinogenesis. We postulated that miR-9-1, miR-9-2 and miR-9-3 may be inactivated by aberrant promoter methylation in multiple myeloma (MM).
  
  
  METHODS
  Methylation of miR-9-1, miR-9-2 and miR-9-3 was studied by methylation-specific PCR (MSP) in six normal controls, including three each of healthy peripheral blood (PB) or bone marrow buffy coat, 10 MM cell lines, 62 primary MM marrow samples at diagnosis and 22 at relapse/progression.
  
  
  RESULTS
  MSP, verified by quantitative pyrosequencing, showed that the promoters of miR-9-3 and miR-9-1 were unmethylated in normal controls but methylated in 4 (40%) and 5 (50%) of 10 MM cell lines, respectively. However, the promoter of miR-9-2 was methylated in three normal PB buffy coat controls and in CD138-sorted healthy marrow plasma cells, indicating possibly tissue specific rather than tumour-specific methylation of miR-9-2, which was thus not studied further. In WL-2 cells, which were completely methylated for miR-9-3, 5-aza-2'-deoxycytidine treatment caused miR-9-3 promoter demethylation and pri-miR-9-3 re-expression. In primary samples, methylation of miR-9-3 was detected in 1 of 62 patients at diagnosis and 1 of 22 patients at relapse/progression. However, miR-9-1 methylation was absent in both primary samples at diagnosis and at relapse/progression.
  
  
  CONCLUSIONS
  Hypermethylation of miR-9-3 and miR-9-1 is tumour-specific in MM, leading to reversible miRNA silencing. Frequent methylation of miR-9-3 and miR-9-1 in cell lines, but not in primary samples, may be acquired during in vitro culture, and indicates an unimportant role of miR-9 methylation in myelomagenesis.
authors: Department of Haematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou Medical College, Xuzhou, Jiangsu Province, China.
authors: Qi
authors: Q
authors: Zhang
authors: Department of Medicine, Queen Mary Hospital, University of Hong Kong, Hong Kong, Hong Kong.
authors: Lu Qian
authors: LQ
authors: Wang
authors: Department of Medicine, Queen Mary Hospital, University of Hong Kong, Hong Kong, Hong Kong.
authors: Kwan Yeung
authors: KY
authors: Wong
authors: Department of Haematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou Medical College, Xuzhou, Jiangsu Province, China.
authors: Zhen Yu
authors: ZY
authors: Li
authors: Department of Medicine, Queen Mary Hospital, University of Hong Kong, Hong Kong, Hong Kong.
authors: Chor Sang
authors: CS
authors: Chim
mesh_terms: D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D014408:Biomarkers, Tumor / Q000235:genetics* / Q000378:metabolism; D016022:Case-Control Studies; D045744:Cell Line, Tumor; D019175:DNA Methylation; D015254:DNA Modification Methylases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000077209:Decitabine; D018450:Disease Progression; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic; D020022:Genetic Predisposition to Disease; D006801:Humans; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D009101:Multiple Myeloma / Q000235:genetics* / Q000378:metabolism / Q000628:therapy; D010641:Phenotype; D011401:Promoter Regions, Genetic; D012008:Recurrence; D016896:Treatment Outcome
chemical_list: D014408:Biomarkers, Tumor; D004791:Enzyme Inhibitors; C541874:MIRN92 microRNA, human; D035683:MicroRNAs; D000077209:Decitabine; D015254:DNA Modification Methylases; D001374:Azacitidine
keywords: CANCER RESEARCH; CELL BIOLOGY; MYELOMA