Aim: To investigate the effects of DNA methylation on the expression of tumor suppressor genes and proto-oncogene in human colon cancer cell lines.
Methods: Three colon cancer cell lines (HT-29, SW1116 and Colo-320) treated with different concentrations of DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine (5-aza-dC) were used to induce DNA demethylation. The expressions of p16(INK4A), p21(WAF1), APC and c-myc genes were observed by using RT-PCR. The methylation status of p16(INK4A) promoter in HT-29 cells was also determined by methylation-specific PCR (MSP).
Results: Weak expressions of p16(INK4A) and APC in the three colon cancer cells were detected, and p21(WAF1) expression was not found in SW1116 and Colo-320 cells before treatment. After treatment of 1 micromol/L but not 10 micromol/L of 5-aza-dC, the methylation level of p16(INK4A) gene promoter decreased significantly, and the hypomethylation led to the up-regulation of p16(INK4A) gene transcription in HT-29 cells. In the cell lines of SW1116 and Colo-320, p16(INK4A) and APC mRNA expressions were obviously enhanced after treatment of either 10 micromol/L or 5 micromol/L 5-aza-dC for 24 h. However, no evidence was found that methylation regulated the expression of p21(WAF1) and c-myc genes in human colon cancer cell lines.
Conclusion: Expression of p16(INK4A) and APC genes is regulated by DNA methylation in three human colon cancer cell lines.