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Background Intratumoral heterogeneity is crucially involved in metastasis, resistance to therapy, and cancer relapse. Amplifications of the proto-oncogene MYC display notable heterogeneity at the single-cell level and are associated with a particularly dismal prognosis in high-risk medulloblastomas (MBs). The aim of this study was to establish the relevance of interclonal cross-talk between MYC-driven and non-MYC-driven MB cells. Methods We used fluorescence in situ hybridization, single-cell transcriptomics, and immunohistochemistry, in vitro isogenic cell models, non-targeted proteomics, mass spectrometry-based metabolite quantification, HUVECs tube formation assay, and orthotopic in vivo experiments to investigate interclonal cross-talk in MB. Results We found that the release of lactate dehydrogenase A (LDHA) from MYC-driven cells facilitates metastatic seeding and outgrowth, while secretion of dickkopf WNT signaling pathway inhibitor 3 from non-MYC-driven cells promotes tumor angiogenesis. This tumor-supporting interaction between both subclones was abrogated by targeting the secretome through pharmacological and genetic inhibition of LDHA, which significantly suppressed tumor cell migration. Conclusion Our study reveals the functional relevance of clonal diversity and highlights the therapeutic potential of targeting the secretome to interrupt interclonal communication and progression in high-risk MB.

Most lncRNAs display species-specific expression patterns suggesting that animal models of cancer may only incompletely recapitulate the regulatory crosstalk between lncRNAs and oncogenic pathways in humans. Among these pathways, Sonic Hedgehog (SHH) signaling is aberrantly activated in several human cancer entities. We unravel that aberrant expression of the primate-specific lncRNA HedgeHog Interacting Protein-AntiSense 1 (HHIP-AS1) is a hallmark of SHH-driven tumors including medulloblastoma and atypical teratoid/rhabdoid tumors. HHIP-AS1 is actively transcribed from a bidirectional promoter shared with SHH regulator HHIP. Knockdown of HHIP-AS1 induces mitotic spindle deregulation impairing tumorigenicity in vitro and in vivo. Mechanistically, HHIP-AS1 binds directly to the mRNA of cytoplasmic dynein 1 intermediate chain 2 (DYNC1I2) and attenuates its degradation by hsa-miR-425-5p. We uncover that neither HHIP-AS1 nor the corresponding regulatory element in DYNC1I2 are evolutionary conserved in mice. Taken together, we discover an lncRNA-mediated mechanism that enables the pro-mitotic effects of SHH pathway activation in human tumors.

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Introduction Resistance to chemotherapy is a common cause of treatment failure in cancer patients and a major problem facing current cancer research. Targeted modulation of oncogenic signaling pathways may be used to systematically characterize drug resistance mechanisms across tumor entities and may help to identify new therapeutic strategies. Since the transcription factor MYC is aberrantly activated in many cancers including pediatric malignant brain tumors, like medulloblastoma (MB), our study focused on MYC-related drug resistance. Methods and Results We performed high-throughput drug screening using our in-house semi-automated platform and identified the HDAC inhibitor Entinostat as a drug that shows promising effects in MYC-driven MB. Investigating genome-wide dCas9-based transcriptional activation screening, potential drug response modulators, mainly TGFB1/Erk/MKNK1 signaling including neural EGFL like 2 (NELL2), were discovered. For further validation, we stably overexpressed NELL2 in MYC-driven MB cells and treated overexpressing cells and the corresponding control cells with Entinostat. Using PI staining, cell cycle status was tracked. Entinostat treatment led to modest induction of cell death in MYC-driven MB control cells but only slightly increased cell death rate in MYC-driven MB cells with NELL2-overexpression. Conclusion We report that the combination of genetic and pharmacological approaches is a powerful approach to study drug resistance. Our data suggest that activation of the TGFB1/Erk/MKNK1 signaling pathway desensitizes MYC-driven MB cells to Entinostat. Synergistic targeting of TGFB1/Erk/MKNK1 signaling and MYC could therefore provide a novel therapeutic option in this aggressive MB subtype.