Single nCounter assay for prediction of MYCN amplification and molecular classification of medulloblastomas: a multicentric study

Purpose Medulloblastoma is the most frequent pediatric malignant brain tumor, and is divided into four main subgroups: WNT, SHH, group 3, and group 4. MYCN amplification is an important medulloblastoma prognostic biomarker. We aimed to molecular classify and predict MYCN amplification in a single assay. Methods It was included 209 medulloblastomas from 205 patients (Brazil, Argentina, and Portugal), divided into training ( n = 50) and validation ( n = 159) sets. A nCounter assay was carried out using a custom panel for molecular classification, with additional genes, including MYCN . nSolver 4.0 software and the R environment were used for profiling and MYCN mRNA analysis. MYCN amplification by FISH was performed in 64 cases. Results The 205 medulloblastomas were classified in SHH (44.9%), WNT (15.6%), group 3 (18.1%) and group 4 (21.4%). In the training set, MYCN amplification was detected in three SHH medulloblastomas by FISH, which showed significantly higher MYCN mRNA counts than non-FISH amplified cases, and a cutoff for MYCN amplification was established ( X ¯ + 4σ = 11,124.3). Applying this threshold value in the validation set, we identified MYCN mRNA counts above the cutoff in three cases, which were FISH validated. Conclusion We successfully stratified medulloblastoma molecular subgroups and predicted MYCN amplification using a single nCounter assay without the requirement of additional biological tissue, costs, or bench time.