Medulloblastoma is the most common brain tumor in early childhood. Molecular profiling of medulloblastomas has resulted in a classification system including at least 4 major subtypes of disease, defining prognosis in patients treated largely on traditional chemotherapy and irradiation-based protocols. The Head Start Consortium has developed treatment protocols designed to avoid irradiation and its debilitating long-term side effects. The first three Head Start trials showed promising results; however, prospective genetic data are still lacking to evaluate whether molecular alterations can predict patient response to irradiation-avoiding treatment strategies. The Head Start 4 Protocol is a multi-national trial that incorporates non-mandatory, prospective tumor and blood collection for analysis in an attempt to address this question. Currently, 100% of patients enrolled to date have submitted samples for WES. Paired blood and tumor samples (FFPE) from the first 10 medulloblastoma patients had WES (SureSelect Human All Exon V6+COSMIC) performed utilizing an Illumina Hiseq 4000. Data were processed using varscan v2.4.1 somatic command on tumor/normal pairs and annotated using Variant Effect Predictor from ensembl. Results were filtered at a threshold of 20 read depth or 40% variant allele frequency. Low-impact mutations were removed. Mutation profiles were compared with pediatric medulloblastoma gene mutation data from the PedcBioPortal and PeCan databases. A total of 903 mutation events were identified across 10 patients with each patient demonstrating mutations in one or more genes previously reported to be mutated in medulloblastoma. Two samples harbored SUFU mutations similar to those reported previously. CONCLUSIONS: These data show the feasibility of prospective high-dimensional mutational analysis using non-mandatory tissue submissions for pediatric medulloblastoma patients enrolled on the Head Start 4 Protocol. Copy-number and methylation analysis is ongoing. FUNDING: Thrasher Research Fund (JF, AC). R01CA108633, R01CA169368, RC2CA148190, U10CA180850-01 (NCI), Brain Tumor Funders Collaborative Grant, and OSUCCC (AC).