T134: Allelic imbalance and epigenetic changes as a marker of tumor spreading into the adjacent tissue

Abstract

Non-small cell lung cancer (NSCLC) is characterized by multiple genetic alterations such as loss of heterozygosity (LOH), microsatellite instability (MSI), promoter hypermethylation and changes of miRNA expression. According to a field cancerization (FC) phenomenon the adjacent histologically normal tissue plays a role in tumor progression by triggering the transformation process. The aim of the study was the analysis of genetic alterations in tumor and adjacent tissue to determine the FC size and to reveal associations with clinico-morphological features of patients. The study group included 135 patients with NSCLC. From each patient 4 FFPE samples were analyzed: tumor, adjacent normal lung tissue at 2, 5, 10 cm. LOH/MSI analysis was evaluated by PCR using 7 microsatellite loci. Promoter hypermethylation in genes RASSF1A FHIT, DAPK1, CDH1, CD44, TIMP3, MGMT was investigated by methyl-sensitive PCR. The expression levels of miRNAs let-7a, miR-155, miR-205 were measured by real-time PCR. Our results demonstrated that LOH/MSI occurs only in tumor while promoter hypermethylation occurs also in adjacent tissue at 2, 5 cm, but not at 10 cm. The downregulation of let-7a, miR-155 in adjacent tissue is lower than in tumor. The levels of investigated miRNAs in adjacent tissue vary depending on tumor differentiation – in patients with differentiated tumors it is higher than in the group with poorly differentiated tumors. We postulate that FC size in NSCLC is at least 5 cm from tumor and includes only epigenetic but not structural (LOH/MSI) alterations. The evaluation of epigenetic changes in adjacent tissue (e.g., surgical margins) can potentially be used for postsurgical prognosis.