Demethylation of Cancer/Testis Antigens and CpG ODN Stimulation Enhance Dendritic Cell and Cytotoxic T Lymphocyte Function in a Mouse Mammary Model

Figure 1

Methylation status of the P1A gene promoter and dose- and time-dependent P1A induction with 5-aza in 4T1 cells. 4T1, CT26, and A20 cell lines were treated with or without 3 μM 5-aza for 48 h, and (a) methylated levels were shown, −, not treated with 5-aza and +, 5-aza treated cells (M, methylated DNA; U, unmethylated DNA). (b) P1A mRNA levels detected by RT-PCR. (c) 5-aza dose response. 4T1 cells were treated for 48 h with 5-aza at concentrations of 0, 0.03, 0.1, 0.3, 1.0, 3, or 10 μM. Blank, negative control and P, positive control. (d) Time course 5-aza treatment at a fixed concentration of 3.0 μM. 4T1 cells were incubated with 5-aza for 0, 2, 6, 12, 24, 48, or 72 h. The P1A expression was detected with P1A-specific primers. GAPDH was amplified as a control to demonstrate cDNA integrity. The positive lane contained EL9611 tumor cells that express high level of P1A.