Epigenetic modifiers such as decitabine (DAC), a DNA methyltransferase inhibitor, have the potential benefit of combination therapy with conventional anti-cancer drugs. This study was aimed to clarify whether DAC at the low concentration without cytotoxic effects exerts synergistic effects with conventional anti-cancer drugs in human colorectal cancer cell line, HT29 cells low-sensitive to DAC. DAC at very low concentration below its IC20 synergistically enhanced cytotoxicity of oxaliplatin (L-OHP), and the enhancement was the most remarkable for L-OHP among several anti-cancer drugs tested. DAC was found to be metabolized and incorporated into DNA in HT29 cells. Combination of L-OHP with DAC did not increase the protein expression of gamma H2A.X, the earliest indicator of DNA damage, in HT29 cells. On the other hand, although DAC alone did not produce reactive oxygen species (ROS), DAC significantly enhanced ROS production by L-OHP in HT29 cells. Furthermore, enhanced cytotoxicity of L-OHP by DAC was cancelled with the presence of N-acetyl-L-cysteine, a ROS scavenger. The mRNA expression of ROS-generating enzymes was significantly increased by combination in HT29 cells. Taken together, indirect enhancement of ROS production by DAC at the low concentration with negligible cytotoxicity should be at least involved in synergistic effects with L-OHP in HT29 cells with intrinsic resistance to DAC. (C) 2018 Elsevier Inc. All rights reserved.