Intragenic DNA Methylation Concomitant with Repression of ATP4B and ATP4A Gene Expression in Gastric Cancer is a Potential Serum Biomarker
0301 basic medicine
0303 health sciences
Reverse Transcriptase Polymerase Chain Reaction
Down-Regulation
Adenocarcinoma
DNA Methylation
Real-Time Polymerase Chain Reaction
3. Good health
Gene Expression Regulation, Neoplastic
H(+)-K(+)-Exchanging ATPase
03 medical and health sciences
Gastric Mucosa
Stomach Neoplasms
Biomarkers, Tumor
Tumor Cells, Cultured
Humans
CpG Islands
RNA, Messenger
DNA Primers
DOI:
10.7314/apjcp.2012.13.11.5563
Publication Date:
2013-03-14T06:28:41Z
AUTHORS (3)
ABSTRACT
Based on our previous report on gastric cancer which documented ATP4A and ATP4B mRNA down- regulation in gastric tumors relative to normal gastric tissues, we hypothesized that epigenetic mechanisms could be responsible. ATP4A and ATP4B mRNA expression in gastric cancer cell lines AGS, SNU638 and NUGC-3 was examined using reverse transcriptase PCR (RT-PCR). AGS cells were treated with TSA or 5'-AzaDC and methylation specific PCR (MSP) and bisulfite sequencing PCR (BSP) analysis were performed. MSP analysis was on DNA from paraffin embedded tissues sections and plasma. Expression analysis revealed downregulation of ATP4A and ATP4B genes in gastric cancer cell lines relative to normal gastric tissue, while treatment with 5'-AzaDC re-activated expression of both. Search for CpG islands in their putative promoter regions did not indicate CpG islands (CGI) but only further downstream in the bodies of the genes. Methylation specific PCR (MSP) in the exon1 of the ATP4B gene and exon7 in ATP4A indicated methylation in all the gastric cancer cell lines tested. MSP analysis in tumor tissue samples revealed methylation in the majority of tumor samples, 15/19, for ATP4B and 8/8 for ATP4A. There was concordance between ATP4B and ATP4A down-regulation and methylation status in the tumour samples tested. ATP4B methylation was detectable in cell free DNA from gastric cancer patient's plasma samples. Thus ATP4A and ATP4B down-regulation involves DNA methylation and methylated ATP4B DNA in plasma is a potential biomarker for gastric cancer.
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