NORE1B, a candidate tumor suppressor, is epigenetically silenced in human hepatocellular carcinoma.
OBJECTIVE
In human hepatocellular carcinoma (HCC) the ras-proto-oncogene is rarely mutated. We therefore studied the possible inactivation of the putative tumor-suppressors and ras-associating proteins, NORE1A, NORE1B, and RASSF1A in HCCs by mutation or epigenetic gene silencing through promoter-CpG hypermethylation.


METHODS
SSCP-analyses, sequencing, and methylation-specific PCR were performed in 28 fibrotic/cirrhotic livers and 40 HCCs.


RESULTS
The sequence of NORE1A/B exhibited no deviations and that of the RASSF1A gene a non-silent polymorphism ( approximately 10% of cases) and a missense mutation (one HCC). Both alterations may affect the growth-inhibiting capability of RASSF1A. Epigenetic inactivation of NORE1B was found in 62% of the HCCs and in hepatocarcinoma-cell lines due to considerable promoter-methylation of the gene. Methylation was detected also for RASSF1A in HCCs and hepatocarcinoma cell-lines. As a result, 97% of the HCCs revealed epigenetic silencing of NORE1B, RASSF1A, or both. In contrast every third fibrotic/cirrhotic liver only exhibited silencing of one or both genes.


CONCLUSIONS
The candidate tumor suppressor genes NORE1B and RASSF1A are epigenetically down-regulated alone in at least 62%, or in combination in 97% of the HCCs studied. This indicates a frequent and critical event in hepatocarcinogenesis, which may allow HCCs to subverse growth-control in the presence of an unaltered Ras.
Macheiner
Doris
D
Department of Medicine I, Division: Institute of Cancer Research, Medical University of Vienna, Borschkegasse 8a, A-1090 Vienna, Austria.
Heller
Gerwin
G
Kappel
Sonja
S
Bichler
Christoph
C
Stättner
Stefan
S
Ziegler
Barbara
B
Kandioler
Daniela
D
Wrba
Fritz
F
Schulte-Hermann
Rolf
R
Zöchbauer-Müller
Sabine
S
Grasl-Kraupp
Bettina
B
D048868:Adaptor Proteins, Signal Transducing; D000328:Adult; D000368:Aged; D051017:Apoptosis Regulatory Proteins; D006528:Carcinoma, Hepatocellular / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D002455:Cell Division; D045744:Cell Line, Tumor; D019175:DNA Methylation; D004273:DNA, Neoplasm / Q000235:genetics; D005260:Female; D020868:Gene Silencing; D016147:Genes, Tumor Suppressor; D005914:Globins / Q000235:genetics; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D008297:Male; D008875:Middle Aged; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics*; D016133:Polymerase Chain Reaction; D000090063:Proto-Oncogene Mas
D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D004273:DNA, Neoplasm; C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; C454042:RASSF5 protein, human; D005914:Globins; D020559:Monomeric GTP-Binding Proteins