PO-391 DNA methylation as a probable cause of SPRR3 loss in esophageal cancer

Introduction Esophageal cancer (EC) is the eighth most frequent cancer and the sixth leading cause of cancer-related deaths worldwide, with squamous cell carcinoma (ESCC) corresponding to 80% of the cases. SPRR3 is a differentiation-associated gene that belongs to SPRR (small proline rich) family that showed a gradual loss of expression in malignant transformation of the healthy oesophagus into ESCC. However, studies of the molecular mechanisms involved in SPRR3 silencing are limited. Thus, the aim of this study is to examine DNA methylation as a regulatory mechanism of SPRR3 expression in ESCC. Material and methods Three CpG sites of SPRR3 (A, B for promotor region and C for 5´UTR region) were analysed by pyrosequencing in esophageal cancer cell lines (TE-1, TE-13 and OE21) treated with the demethylating agent decitabine, and in tumour and matched normal surrounding mucosa from patients with ESCC. RT-qPCR was performed to evaluated SPRR3 , DNMT1, DNMT3A and DNMT3B expression in the same samples. Results and discussions Decitabine treatment was able to induce SPRR3 demethylation and reestablished esophagin expression in the three cell lines tested. The methylation of all CpG sites analysed was significantly higher in tumours in comparison with the adjacent tissues (p SPRR3 methylation to distinguish the adjacent mucosa from tumour samples using ROC curve analyses was significant for each of the CpG sites examined (A, sensitivity 93.10% and specificity=86.21%, p SPRR3 mRNA expression for all CpG sites evaluated (A and B p SPRR3 hypermethylation, we evaluated DNMTs expression (higher expression in tumour in comparision with the normal surrounding mucosa, p DNMT1 , p=0.0005 DNMT3A and p DNMT3B ) which are inversely correlated with SPRR3 expression ( DNMT1 p=0.0013, DNMT3A p=0.0108 and DNMT3B p SPRR3 methylation and DNMTs mRNA expression for all CpG sites evaluated exceptd for DNMT3A and Site C. Conclusion Thus, our data provide evidences that DNA methylation induced by DNMTs is a possible mechanism involved for SPRR3 silencing in ESCC.