DAPK promotor hypermethylation correlates with reduced Fas/FasL-induced apoptosis in colorectal cancer

Introduction: The death-associated protein kinase (DAPK) gene is a promoting regulator of programmed cell death and is frequently inactivated by promotor methylation in many tumor entities.

Aim: To determine the role of DAPK inactivation and its association with apoptosis in colorectal carcinogenesis.

Methods: We investigated fresh-frozen specimens obtained from 103 patients with primary colorectal carcinomas of advanced stage and microdissected samples of 22 formalin-fixed and paraffin-embedded Dukes'-A carcinomas showing normal colon mucosa, adenoma and carcinoma tissue on the same slice. Aberrant promotor methylation was determined using methylation-specific PCR. Expression of apoptotis-relevant proteins such as Bcl2, Bax, Fas, and FasL was investigated using immunohistochemistry.

Results: DAPK methylation was found in 58% of fresh-frozen colon carcinoma samples whereas it was higher in the microdissected carcinoma regions (80%). Adenomas showed aberrant methylation in 40% of cases. Interestingly, there was a significant increase in methylation frequency of low-grade adenomas compared with high-grade dysplasia. DAPK methylation rate of the normal colon mucosa was only 22%. In general, tissue showing DAPK hypermethylation failed to express Fas/FasL proteins, but showed an overexpression of the anti-apoptotic Bcl2 protein (p=0.015). Tumor lesions without DAPK methylation almost exclusively showed Fas/FasL coexpression but no detection of Bcl2. In addition, there was no significant association of DAPK methylation with tumor localization, stage (according to Dukes classification), age and gender.

In conclusion, our data suggest that DAPK hypermethylation plays a significant role in colorectal carcinogenesis and correlates with a reduction of Fas/FasL-induced apoptosis.