Sonic hedgehog medulloblastoma encompasses a clinically and molecularly diverse group of cancers of the developing central nervous system. Here, we use unbiased sequencing of the transcriptome across a large cohort of 250 tumors to reveal differences among molecular subtypes of the disease, and demonstrate the previously unappreciated importance of non-coding RNA transcripts. We identify alterations within the cAMP dependent pathway (GNAS, PRKAR1A) which converge on GLI2 activity and show that 18% of tumors have a genetic event that directly targets the abundance and/or stability of MYCN. Furthermore, we discover an extensive network of fusions in focally amplified regions encompassing GLI2, and several loss-of-function fusions in tumor suppressor genes PTCH1, SUFU and NCOR1. Molecular convergence on a subset of genes by nucleotide variants, copy number aberrations, and gene fusions highlight the key roles of specific pathways in the pathogenesis of Sonic hedgehog medulloblastoma and open up opportunities for therapeutic intervention.

Glycine 34-to-tryptophan (G34W) substitutions in H3.3 arise in approximately 90% of giant cell tumor of bone (GCT). Here, we show H3.3 G34W is necessary for tumor formation. By profiling the epigenome, transcriptome, and secreted proteome of patient samples and tumor-derived cells CRISPR–Cas9-edited for H3.3 G34W, we show that H3.3K36me3 loss on mutant H3.3 alters the deposition of the repressive H3K27me3 mark from intergenic to genic regions, beyond areas of H3.3 deposition. This promotes redistribution of other chromatin marks and aberrant transcription, altering cell fate in mesenchymal progenitors and hindering differentiation. Single-cell transcriptomics reveals that H3.3 G34W stromal cells recapitulate a neoplastic trajectory from a SPP1⁺ osteoblast-like progenitor population toward an ACTA2⁺ myofibroblast-like population, which secretes extracellular matrix ligands predicted to recruit and activate osteoclasts. Our findings suggest that H3.3 G34W leads to GCT by sustaining a transformed state in osteoblast-like progenitors, which promotes neoplastic growth, pathologic recruitment of giant osteoclasts, and bone destruction. Significance: This study shows that H3.3 G34W drives GCT tumorigenesis through aberrant epigenetic remodeling, altering differentiation trajectories in mesenchymal progenitors. H3.3 G34W promotes in neoplastic stromal cells an osteoblast-like progenitor state that enables undue interactions with the tumor microenvironment, driving GCT pathogenesis. These epigenetic changes may be amenable to therapeutic targeting in GCT. See related commentary by Licht, p. 1794. This article is highlighted in the In This Issue feature, p. 1775

Gammaherpesviruses (GHVs) infect many animal species and are associated with lymphoproliferative disorders in some. Previously, we identified several novel GHVs in North American felids; however, a GHV had never been identified in Canada lynx (Lynx canadensis). We, therefore, hypothesized the existence of an unidentified GHV in lynx. Using degenerate nested and subsequently virus-specific PCR, we amplified and sequenced 3.4 kb of DNA from a novel GHV in lynx, which we named Lynx canadensis gammaherpesvirus 1 (LcaGHV1). Phylogenetic analysis determined that LcaGHV1 is a distinct GHV species belonging to the genus Percavirus. We then estimated the prevalence of LcaGHV1 in lynx by developing a PCR-based assay and detected LcaGHV1 DNA in 36% (95% CI: 22–53%) of lynx spleen DNA samples from Maine, USA and 17% (95% CI: 8–31%) from Newfoundland, Canada. The LcaGHV1 DNA sequences from Maine and Newfoundland lynx were nearly identical to each other (two nucleotide substitutions in 3.4 kb), suggesting that the unique lynx subspecies present on the island of Newfoundland (Lynx canadensis subsolanus) is infected with virus that very closely resembles virus found in mainland lynx. The potential ecologic and pathologic consequences of this novel virus for Canada lynx populations warrant further study.