DNA methylation patterns in Uveal Melanoma FFPE samples correlate with survival

Abstract

Purpose : The present study examines correlations of DNA methylation patterns of human uveal (UM) melanoma with survival. UM is the most common primary intraocular malignancy and has a high propensity for metastatic spread. Monosomy 3 has long been known to be associated with the development of aggressive UM. The role of epigenetics in cancer research is emerging, though the epigenome of UM is less characterised. Epigenetic alterations involving DNA methylation can lead to cancer by various mechanisms, including induction of genomic instability, gene silencing and susceptibility to mutation.

Methods : Formalin fixed paraffin embedded (FFPE) samples from 24 UM patients undergoing enucleation of the eye in the period 1976-1989, were included and approved by the regional ethical committee (2013/803/REK). The inclusion was based on data from histopathology and from the Norwegian Cancer Registry and the Norwegian Cause of Death Registry. Survival cut-off: early relapse ≤5 years, late relapse ≥10 years and “Ctrl”/no relapse ≥17 years. DNA was isolated from the FFPE samples and further bisulphite converted and restored according to Illumina`s recommendations. Bisulphite converted DNA was then run on The Infinium HumanMethylation450 BeadChip assay (HM-450K). Bioinformatics analyses were conducted in R statistical software, including analysis of copy number variation (CNV). RNA was isolated from the FFPE samples and Clariom D arrays were used to verify changes in gene expression correlated to changes in methylation.

Results : Cluster analysis was performed using unsupervised hierarchical clustering. Our data shows subclustering correlated with survival and mortality, additionally our data provides insight of epigenetically deregulated (hypo/hyper methylated) genes and CNV. After limma analysis of subcluster, 4 samples of “Ctrl”/no relapse compared to 4 samples of early relapse, unraveled 335 significant probes (p-value≤0.05) with the potential of increasing our knowledge about the underlying mechanisms of UM.

Conclusions : The methylation profile of a given cancer can elucidate therapeutic targets, reveal biomarkers for early detection or identify high risk tumours. This is to our knowledge the first report of methylation
changes in FFPE samples derived from UMs correlated to survival.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.