Conventional or real-time RT-PCR was performed in normal tissue samples, cell lines and HCC samples, and sequencing analysis and methylation-specific PCR in cell lines and HCC samples.
hDAB2IPA was the predominant isoform, being expressed in the majority of tissues examined. The expression of hDAB2IPA was silenced or down-regulated but could be restored by 5-aza-2′-deoxycytidine treatment in liver cancer cell lines. The reactivation of hDAB2IPA was associated with promoter demethylation. The correlation between promoter methylation and hDAB2IPA expression was confirmed in eight pairs of matched HCC samples. Further, the methylation of the hDAB2IPA promoter in HCC was confirmed in an additional 53 pairs of patient samples. More than 80 % of HCC samples showed hDAB2IPA promoter methylation, compared to 11.5 % in the corresponding adjacent normal tissue (p < 0.0001, χ2).
Our data suggest that hDAB2IPA is the dominant isoform expressed in normal tissues. Its expression is suppressed in HCC, consistent with its role as a tumor suppressor gene, mainly by promoter methylation.