title: Methylation of IRAK3 is a novel prognostic marker in hepatocellular carcinoma.
abstract: OBJECTIVE
  To examine the methylation levels of interleukin-1 receptor-associated kinase 3 (IRAK3) and GLOXD1 and their potential clinical applications in hepatocellular carcinoma (HCC).
  
  
  METHODS
  mRNA expression and promoter methylation of IRAK3 and GLOXD1 in HCC cells were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and methylation-specific PCR (MSP), respectively. Using pyrosequencing results, we further established a quantitative MSP (Q-MSP) system for the evaluation of IRAK3 and GLOXD1 methylation in 29 normal controls and 160 paired HCC tissues and their adjacent nontumor tissues. We also calculated Kaplan-Meier survival curves to determine the applications of gene methylation in the prognosis of HCC.
  
  
  RESULTS
  IRAK3 and GLOXD1 expression was partially restored in several HCC cell lines after treatment with 5-aza-2'-deoxycytidine (DNA methyltransferase inhibitor; 5DAC). A partial decrease in the methylated band was also observed in the HCC cell lines after 5DAC treatment. Using GLOXD1 as an example, we found a significant correlation between the data obtained from the methylation array and from pyrosequencing. The methylation frequency of IRAK3 and GLOXD1 in HCC tissues was 46.9% and 63.8%, respectively. Methylation of IRAK3 was statistically associated with tumor stage. Moreover, HCC patients with IRAK3 methylation had a trend toward poor 3-year disease-free survival (P < 0.05).
  
  
  CONCLUSIONS
  IRAK3 and GLOXD1 were frequently methylated in HCC tissues compared to normal controls and nontumor tissues. IRAK3 methylation was associated with tumor stage and poor prognosis of patients. These data suggest that IRAK3 methylation is a novel prognostic marker in HCC.
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Chih-Chi
authors: CC
authors: Kuo
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Yu-Lueng
authors: YL
authors: Shih
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Her-Young
authors: HY
authors: Su
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Ming-De
authors: MD
authors: Yan
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Chung-Bao
authors: CB
authors: Hsieh
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Chin-Yu
authors: CY
authors: Liu
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Wei-Ting
authors: WT
authors: Huang
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Mu-Hsien
authors: MH
authors: Yu
authors: Chih-Chi Kuo, Ya-Wen Lin, Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 114, Taiwan.
authors: Ya-Wen
authors: YW
authors: Lin
mesh_terms: D000328:Adult; D000368:Aged; D001483:Base Sequence; D014408:Biomarkers, Tumor / Q000235:genetics*; D006528:Carcinoma, Hepatocellular / Q000201:enzymology / Q000235:genetics* / Q000401:mortality / Q000473:pathology; D019175:DNA Methylation; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D056945:Hep G2 Cells; D006801:Humans; D053592:Interleukin-1 Receptor-Associated Kinases / Q000235:genetics*; D053208:Kaplan-Meier Estimate; D008113:Liver Neoplasms / Q000201:enzymology / Q000235:genetics* / Q000401:mortality / Q000473:pathology; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D009367:Neoplasm Staging; D011237:Predictive Value of Tests; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012189:Retrospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
chemical_list: D014408:Biomarkers, Tumor; D012333:RNA, Messenger; C507771:IRAK3 protein, human; D053592:Interleukin-1 Receptor-Associated Kinases
keywords: DNA methylation biomarker; GLOXD1; Hepatocellular carcinoma; IRAK3; Pyrosequencing; Quantitative methylation-specific polymerase chain reaction