Carbon ion radiotherapy eradicates medulloblastomas with chromothripsis in an orthotopic Li-Fraumeni patient-derived mouse model

Abstract

Background Medulloblastomas with chromothripsis developing in children with Li-Fraumeni Syndrome (germline TP53 mutations) are highly aggressive brain tumors with dismal prognosis. Conventional photon radiotherapy and DNA-damaging chemotherapy are not successful for these patients and raise the risk of secondary malignancies. We hypothesized that the pronounced homologous recombination deficiency in these tumors might offer vulnerabilities that can be therapeutically utilized in combination with high linear energy transfer carbon ion radiotherapy. Methods We tested high-precision particle therapy with carbon ions and protons as well as topotecan with or without PARP inhibitor in orthotopic primary and matched relapsed patient-derived xenograft models. Tumor and normal tissue underwent longitudinal morphological (MRI), cellular (markers of neurogenesis and DNA damage-repair) and molecular characterization (whole-genome sequencing). Results In the primary medulloblastoma model, carbon ions led to complete response in 79% of animals irrespective of PARP inhibitor within a follow-up period of 300 days post-irradiation, as detected by MRI and histology. No sign of neurologic symptoms, impairment of neurogenesis or in-field carcinogenesis was detected in repair-deficient host mice. PARP inhibitors further enhanced the effect of proton irradiation. In the post-radiotherapy relapsed tumor model, median survival was significantly increased after carbon ions (96 days) versus control (43 days, p<0.0001). No major change in the clonal composition was detected in the relapsed model. Conclusion The high efficacy and favorable toxicity profile of carbon ions warrants further investigation in primary medulloblastomas with chromothripsis. Post-radiotherapy relapsed medulloblastomas exhibit relative resistance compared to treatment-naïve tumors, calling for exploration of multimodal strategies.

Long-read sequencing of diagnosis and post-therapy medulloblastoma reveals complex rearrangement patterns and epigenetic signatures

“…We developed workflows and algorithms to analyze both genetic and epigenetic alterations in these samples (Methods). Making use of short-read data generated at 45x-48x coverage for these samples 16,18,19 (Table S2), we discovered single-nucleotide variants (SNVs) as well as short insertions and deletions (InDels), where ONT reads have limitations due to their relatively high error rate. As expected, germline variant calling confirmed a TP53 mutation (TP53:c.395A>T, p.Lys132Met, rs1057519996), consistent with Li-Fraumeni Syndrome, coupled with somatic inactivation of the wild-type TP53 allele through deletion in the tumor samples.…”

“…We further sought to integrate the transcriptomic data with the long ONT reads to look for supporting data for gene fusion events (see Table S7), previously described to be prevalent in SHH-Medulloblastoma 44 . We inferred gene fusion events from transcriptomic reads using Arriba on the primary tumor, and identified 127 putative gene fusion pairs of which 103 pairs are supported by genomic evidence, either directly through individual chimeric read alignments of ONT reads near the fusion breakpoints (53) or by tracing SVs called from long and short genomic reads (19) or both (31) (Methods). Breaking down predictions by Arriba confidence shows increased traceability for higher confidence fusion calls (Figure 3F).…”

A synergistic interaction between HDAC‐ and PARP inhibitors in childhood tumors with chromothripsis

“…LFS_MB_P, LFS_MB_1R and LFS_MB_2R were generated and sequenced by Milena Simovic 21 . RCMB18, BT084 and ICB984 PDX models 22 were kindly provided and previously characterized by M. Kool (DKFZ).…”

“…Copy number changes were reported on genes with more than 0.15 in log2 fold change. For whole genome sequencing data, 5,21 copy number analysis was performed using ACESeq (https://www.biorxiv.org/content/10.1101/ 210807v1). Paired tumor-normal alignment files processed using the DKFZ one touch pipeline (https://pubmed.ncbi.nlm.nih.gov/28803971/ ) were provided to ACESeq for allele specific copy number estimation.…”

“…To use representative medulloblastoma models, we established spheroid cultures freshly isolated from patient-derived xenografts. All lines included in the screen were molecularly characterized by methylation arrays (data newly generated for our study) or previously characterized by methylation arrays and/or whole-genome sequencing 19,21,32 (Figure S1).…”