Methylation Profiling for Pediatric Pineal Parenchymal Tumors

Pineal parenchymal body tumors are rare central nervous system tumors with a variety of presentations ranging from well-differentiated low-grade tumors to undifferentiated highly aggressive tumors. Recent molecular classification has described the heterogeneity among these tumors, particularly among pineoblastomas (PB) and papillary tumors of intermediate differentiation (PPTID). Herein, we describe the methylation profile for 20 pediatric pineal parenchymal tumors using Illumina Infinium Methylation EPIC BeadChip array and correlate it with the patients’ clinical data. Our findings support that these tumors are diverse, incorporating novel entities with distinct clinical features and the presence of medulloblastomas outside the posterior fossa. Our Study included 20 formalin-fixed-paraffin embedded (FFPE) tumor samples. DNA was isolated with QIAamp DNA FFPE tissue kit (Qiagen) and quantified using the DENOVIX Fluorometer (ds DNA High Sensitivity). DNA quality was determined using the Illumina FFPE QC kit (Illumina Inc.). Bisulfite conversion of the extracted DNA was performed using an EZ DNA methylation kit (D5002, Zymo Research). The bisulfite-converted DNA was restored using the Infinium HD FFPE DNA Restore Kit (WG-321-1002, Illumina Inc.). Restored bisulfite-converted DNA was hybridized to the Illumina Infinium Human Methylation EPIC 850K bead chips and scanned using the Illumina iScan microarray scanner according to the manufacturer's recommendations (Illumina Inc.).