Silencing of GSTP1 gene by CpG island DNA hypermethylation in HBV-associated hepatocellular carcinomas.
OBJECTIVE
Glutathione S-transferases, enzymes that defend cells against damage mediated by oxidant and electrophilic carcinogens, may be critical determinants of cancer pathogenesis. In this report, we assess the role of epigenetic silencing of the GSTP1 gene, a gene encoding the pi-class glutathione S-transferase, in the pathogenesis of hepatitis B virus (HBV)-associated hepatocellular carcinomas (HCC). The cell lines Hep3B, HepG2, and a cohort of 43 HBV-associated HCC tissue specimens and corresponding nontumor tissues were subjected to analysis for GSTP1 epigenetic alteration and expression. GSTP1 "CpG" island DNA hypermethylation in the liver cell lines, and the tissue specimens were determined by methylation-specific PCR and correlated with expression of the gene using reverse-transcription PCR, immunoblotting, and immunohistochemistry.


RESULTS
GSTP1 CpG island DNA hypermethylation was detected in 28 of 43 (65.1%) HCC tissues and 4 of 40 (10%) corresponding nontumor tissues. GSTP1 protein was absent in those cases showing hypermethylation of the gene. Similarly, DNA from Hep3B and HepG2 cell lines displayed complete GSTP1 hypermethylation in the CpG island, and they failed to express GSTP1 mRNA and the corresponding protein product. Treatment of the cell lines with the DNA methyltransferase inhibitor 5-aza-deoxycytidine reversed the hypermethylation, and restored GSTP1 mRNA and polypeptide expression.


CONCLUSIONS
These data indicate that epigenetic silencing of GSTP1 gene expression by CpG island DNA hypermethylation is common in human HBV-associated HCC. In addition, somatic GSTP1 inactivation via CpG island hypermethylation may contribute to the pathogenesis of this malignancy.
Zhong
Sheng
S
Department of Clinical Oncology, Sir Y. K. Pao Centre for Cancer, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, N. T., Hong Kong SAR, China.
Tang
Mandy W
MW
Yeo
Winnie
W
Liu
Cuiling
C
Lo
Y M Dennis
YM
Johnson
Philip J
PJ
D000328:Adult; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000150:complications / Q000235:genetics* / Q000473:pathology; D018899:CpG Islands / Q000235:genetics*; D019175:DNA Methylation; D004273:DNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D000077209:Decitabine; D005260:Female; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D020868:Gene Silencing; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase / Q000235:genetics* / Q000378:metabolism; D006509:Hepatitis B / Q000150:complications; D006801:Humans; D007527:Isoenzymes / Q000235:genetics* / Q000378:metabolism; D008113:Liver Neoplasms / Q000150:complications / Q000235:genetics* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D007527:Isoenzymes; D012333:RNA, Messenger; D000077209:Decitabine; C496556:GSTP1 protein, human; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase; D001374:Azacitidine