Amplified MYCN, GLI2 and mutant TP53 medulloblastoma are vulnerable to the translation inhibitor homoharringtonine

Background: Medulloblastoma (MB) is the most common malignant pediatric brain tumor derived from transformed neuroepithelial stem (NES) cells, and is comprised of four main subgroups: WNT, SHH, Group 3, and Group 4. SHH MB tumors with amplification of MYCN and GLI2 and mutation of TP53 (MGT) are resistant to inhibitors of the SHH activator, SMO, and exhibit extremely poor prognosis. Given the poor druggability of MYCN and GLI2, we sought to identify new therapeutic targets by performing a drug screen. However, there are no reliable MGT MB cell lines that grow in culture. To circumvent this issue, we utilized a human induced pluripotent stem cell (iPSC) model of MB to generate MGT MB tumors in vivo which can subsequently be cultured in vitro to perform therapeutic screens along with an isogenic NES cell control line. Method: iPSCs from a healthy adult were differentiated towards NES cells, transduced with doxycycline-inducible MYCN and GLI2 and constitutive expression of TP53R248Q and implanted orthotopically into immunocompromised (NSG) mice. Resulting MGT tumors and control (empty vector) NES cells were analyzed by RNAseq and proteomics. MGT tumors were cultured in vitro and, along with control NES cells, were subjected to a drug screen (360 unique compounds from the SBP Oncology Dose Library (ODL3), and CTD2 Informer Set). MGT MB PDX tumors were maintained in NSG mice and analyzed for cell viability in response to drug treatments in 384 well plates with CellTiter-Glo cell viability assay. Results: MGT NES cells were fully penetrant within 35 days post injection in NSG mice. MGT tumor lines had transcriptomes similar to MB SHH subgroup and were resistant to in vitro treatment with chemotherapy drugs cisplatin and cyclophosphamide. The drug screen showed MGT tumors were more sensitive than control NES cells to homoharringtonine (HHT, inhibitor of translation elongation), MLN4924 (inhibitor of NEDD8 Activating Enzyme), and PI-103 (inhibitor of PI3K & mTOR). With three separate MGT MB PDX tumors, MLN4924 and PI-103 failed to reduce viability. In contrast, HHT was effective at reducing viability in all three MGT PDX tumors in a dose dependent manner and IC50s are in the nanomolar range (29.61nM/31.10nM/7.319nM)). Proteomic analysis of MGT NES cell tumors and control NES cells untreated or treated with HHT reveal that two of the 10 proteins most downregulated by HHT in MGT tumors but not control cells, are MYCN and GLI2. In support of this, HHT (1.52nM) induced lower viability in tumors with MYCN and GLI2 (+dox, 50%)) than tumors without (-dox, 93%). Conclusion: Here, we show for the first time that the human stem cell model of MB can be used to identify therapeutic targets that are effective against MB PDX tumors. Specifically, targeting translation elongation is a potential therapeutic target to treat MGT MB tumors.