ABCG2 promoter methylation. (A) Cells were harvested at plateau phase, and the DNA was extracted and assayed by bisulfite DNA sequencing analysis. The figure shows the methylation status of the putative CpG island of the ABCG2 promoter in 4 cell lines. Filled circles represent methylated groups and the open circles, demethylated CpG. (B) H929/5aza are cells treated with 100 nM 5-aza-2′-deoxycytidine for 72 hours and the ABCG2 promoter was assayed by bisulfite sequencing. 5-Aza-2′-deoxycytidine was able to augment ABCG2 transcription in low ABCG2-expressing H929 cells but had no effect in moderate-expressing 8226 cells. (C) CD138-selected MM patient samples were assayed for ABCG2 promoter methylation after bisulfite conversion using real-time quantitative PCR. The percentage of alleles that was methylated inversely correlated with ABCG2 mRNA expression. Error bars represent the standard deviation for 3 separate experiments.