ID #1183 A Single-Cell Atlas of Spontaneous Leptomeningeal Dissemination in Medulloblastoma Across Molecular Subgroups

Abstract Background Medulloblastoma (MB) dissemination to the leptomeninges and spinal cord remains a major driver of treatment failure. A set of clinically relevant models replicating spontaneous CSF spread should facilitate in-depth understanding of the cellular and molecular determinants of spinal dissemination as related to MB subgroups. Methods Matching pairs of cerebellar primary tumors (CBTUMOR) and spontaneous spinal metastases (SPMET) were collected from 69 patient-derived orthotopic xenograft (PDOX) models. Single-cell RNA sequencing (scRNAseq) was applied to 21 MB models to examine cell components between the SPMET and CBTUMOR cells and to identify candidate driver genes pertaining to G3, G4 and SHH subgroups. Growth of tumor organoids from CBTUMOR and SPMET cells was also performed. Results Harvesting of candidate SPMET cells were successful in 46 of 69 PDOX models. ScRNAseq in 21 MB models revealed marked inter- and intra-subgroup heterogeneity, with the highest metastasis frequencies (SPMET/CBTUMOR%) reaching 35.9±32.6% (5 models 10%) in the 7 SHH, followed by 20.3±28.9% (3 models 10%) in 9 G3, 12.9±17.2% in 4 G4 (2 models 10%), and 0.52% in 1 WNT. In addition to identifying cellular composition differences between SPMET and CBTUMOR, three subtype enriched clusters were discovered in G3, two in G4 and one in SHH models accompanied by private and shared metastasis associated gene signatures in the SPMET cells. Among over-expressed genes, we found 23 G3-specific genes (e.g., GTSE1, H2-Q7, PMVK), 36 G4-specific genes (ID1, JUNB, SOX5), no SHH-specific genes, 14 genes shared between G3 and G4 (H2-D1, H2-K1, H2-T23), and 2 genes by G3 and SHH. From the 19 models, 15 CBTUMOR and 3 SPMET-derived tumor organoids were established. Conclusion A new set of spontaneous CSF metastasis MB models were created. ScRNAseq discovered novel subgroup-specific and highly individualized mechanisms, providing a framework for dissecting metastasis biology and developing strategies to target leptomeningeal dissemination.