Targeting synthetic lethality between non-homologous end joining and radiation in very-high-risk medulloblastoma.

Specific and biologically informed treatments for medulloblastoma, especially for the highly lethal TP53-mutant SHH subgroup, remain elusive, where radiotherapy is the primary treatment modality. Leveraging genome-wide CRISPR-Cas9 dropout screening in combination with lethal doses of radiotherapy, we identify loss of p53 as the main driver of radiation resistance in SHH medulloblastoma. A negative-selection CRISPR-Cas9 screen across multiple models of Trp53-deficient SHH medulloblastoma reveals a strong synthetic lethal interaction between components of the non-homologous end-joining pathway and radiation, particularly DNA-dependent protein kinase (DNA-PK) and its binding partners. Both genetic and pharmacological perturbation of DNA-PK enhance radiosensitivity in TP53-deficient SHH medulloblastoma, leading to cell death. In vivo treatment of both somatic and germline TP53-mutant SHH medulloblastoma models with peposertib, a small-molecule inhibitor of DNA-PK, significantly improves survival when combined with radiotherapy, strongly supporting further clinical investigation.