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Objective: To investigate the influence of adenosine (ADO)-induced epigenetic changes in colorectal cancer cells. Methods: SW480 cells were subjected to no treatment, ADO with a final concentration of 3 mmol/L, or 5-iodotubercidin (ITU) with a final concentration of 4 μmol/L for 72 h. The human MutL homolog 1 (hMLH1) gene methylation levels of the CpG islands were detected by bisulfite sequencing polymerase chain reaction; hMLH1 mRNA expression levels were detected by reverse transcription-polymerase chain reaction; hMLH1 protein, DNA (cytosine-5)-methyltransferase 1 (DNMT1), and DNA (cytosine-5)-methyltransferase 3A(DNMT3A) expression levels were detected by western blotting; and apoptosis rates were detected by flow cytometry. Results: The hMLH1 promoter methylation levels were significantly lower in the treatment groups than in the control group (P＜0.01). The mRNA expression levels increased dramatically (P＜0.01). The hMLH1 protein expression levels were significantly higher in the treatment groupsthan in the control group, especially in the ITU group (P＜0.01). The DNMT1 and DNMT3a expression levels were significantly lower than in the control group, especially the ITU group (P＜0.01). The apoptosis rates were significantly higher than the control group, especially the ITU group (P＜0.01).