Detection of genetic and epigenetic alterations and DNA- and albumin-adducts in paired tissue and plasma of HCC patients

2087

The development and progression of hepatocellular carcinoma (HCC) is a multistep process, in which genetic, epigenetic and environmental factors are deeply involved. Several studies have demonstrated that DNA isolated from serum or plasma of cancer patients contains the same genetic and epigenetic aberrations as DNA isolated from an individual’s tumor tissue. In the present study, 39 paired plasma and frozen dissected tumor tissue of HCC patients were collected from Taiwan. p53 mutations in exon 5-8 were analyzed in tissue DNA by the Surveyor™ Mutation Detection Kit followed by sequencing. Mutant p53 protein was determined by immunohistochemical staining of the tissue paraffin sections. 10 of 39 tissue DNA samples (26%) were positive for p53 mutations; 12 of 39 samples (31%) were positive for mutant p53 protein by immunohistochemistry. Results by both methods were consistent in 80% of samples. We are currently analyzing plasma DNA samples from subjects with p53 mutations in tumor. Analysis of hypermethylation of p16 in the same tumor DNA samples found 24 of 39 samples (62%) positive by MSP (Methylation Specific PCR). DNA from the plasma of all 39 subjects was tested for p16 promoter methylation and 12/39 (30%) were positive. AFB1- and PAH- DNA and albumin adducts in tissues and serum were detected by immunohistochemistry and ELISA, respectively. AFB1-Alb adducts in subjects with low, medium and high levels of AFB1- DNA adducts in tumor tissues were 43.9, 58.8 and 69.7 fmol/mg respectively (P-value =0.06). No correlation was found between PAH-DNA adducts level in tissues and PAH-Alb in serum. This primary data shows that genetic, epigenetic and environmental exposure biomarkers in serum may help in estimating risk for development of HCC.