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Glioblastoma is the most common malignant brain cancer with dismal survival and prognosis. Temozolomide (TMZ) is a first-line chemotherapeutic agent for glioblastoma, but the emergence of drug resistance limits its anti-tumor activity. We previously discovered that the interferon inducible guanylate binding protein 3 (GBP3) is highly elevated and promotes tumorigenicity of glioblastoma. Here, we show that TMZ treatment significantly upregulates the expression of GBP3 and stimulator of interferon genes (STING), both of which increase TMZ-induced DNA damage repair and reduce cell apoptosis of glioblastoma cells. Mechanistically, relying on its N-terminal GTPase domain, GBP3 physically interacts with STING to stabilize STING protein levels, which in turn induces expression of p62 (Sequestosome 1), nuclear factor erythroid 2 like 2 (NFE2L2, NRF2), and O6-methlyguanine-DNA-methyltransferase (MGMT), leading to the resistance to TMZ treatment. Reducing GBP3 levels by RNA interference in glioblastoma cells markedly increases the sensitivity to TMZ treatment in vitro and in murine glioblastoma models. Clinically, GBP3 expression is high and positively correlated with STING, NRF2, p62, and MGMT expression in human glioblastoma tumors, and is associated with poor outcomes. These findings provide novel insight into TMZ resistance and suggest that GBP3 may represent a novel potential target for the treatment of glioblastoma.

Diffuse midline glioma (DMG) is a leading cause of brain tumor death in children. In addition to hallmark H3.3K27M mutations, significant subsets also harbor alterations of other genes, such asTP53andPDGFRA. Despite the prevalence of H3.3K27M, the results of clinical trials in DMG have been mixed, possibly due to the lack of models recapitulating its genetic heterogeneity. To address this gap, we developed human iPSC-derived tumor models harboring TP53R248Qwith or without heterozygous H3.3K27M and/or PDGFRAD842Voverexpression. The combination of H3.3K27M and PDGFRAD842Vresulted in more proliferative tumors when gene-edited neural progenitor (NP) cells were implanted into mouse brains compared to NP with either mutation alone. Transcriptomic comparison of tumors and their NP cells of origin identified conserved JAK/STAT pathway activation across genotypes as characteristic of malignant transformation. Conversely, integrated genome-wide epigenomic and transcriptomic analyses, as well as rational pharmacologic inhibition, revealed targetable vulnerabilities unique to the TP53R248Q; H3.3K27M; PDGFRAD842Vtumors and related to their aggressive growth phenotype. These includeAREG-mediated cell cycle control, altered metabolism, and vulnerability to combination ONC201/trametinib treatment. Taken together, these data suggest that cooperation between H3.3K27M and PDGFRA influences tumor biology, underscoring the need for better molecular stratification in DMG clinical trials.

Among children and adolescents in the United States (0 to 19 years old), brain and other central nervous system tumors are the second most common types of cancers, surpassed in incidence only by leukemias. Despite significant progress in the diagnosis and treatment modalities, brain cancer remains the leading cause of death in the pediatric population. There is an obvious unfulfilled need to streamline the therapeutic strategies and improve survival for these patients. For that purpose, preclinical models play a pivotal role. Numerous models are currently used in pediatric brain tumor research, including genetically engineered mouse models, patient-derived xenografts and cell lines, and newer models that utilize novel technologies such as genome engineering and organoids. Furthermore, extensive studies by the Children's Brain Tumor Network (CBTN) researchers and others have revealed multiomic landscapes of variable pediatric brain tumors. Combined with such integrative data, these novel technologies have enabled numerous applicable models. Genome engineering, including CRISPR/Cas9, expanded the flexibility of modeling. Models generated through genome engineering enabled studying particular genetic alterations in clean isogenic backgrounds, facilitating the dissection of functional mechanisms of those mutations in tumor biology. Organoids have been applied to study tumor-to-tumor-microenvironment interactions and to address developmental aspects of tumorigenesis, which is essential in some pediatric brain tumors. Other modalities, such as humanized mouse models, could potentially be applied to pediatric brain tumors. In addition to current valuable models, such novel models are anticipated to expedite functional tumor biology study and establish effective therapeutics for pediatric brain tumors.

Glioblastoma multiforme (GBM) is among the most aggressive, treatment-resistant cancers, and despite standard of care surgery, radiation and chemotherapy, is invariably fatal. GBM is marked by local and systemic immunosuppression, contributing to resistance to existing immunotherapies that have had success in other tumor types. Memory T cells specific for previous infections reside in tissues throughout the host and are capable of rapid and potent immune activation. Here, we show that virus-specific memory CD8 + T cells expressing tissue-resident markers populate the mouse and human glioblastoma microenvironment. Reactivating virus-specific memory T cells through intratumoral delivery of adjuvant-free virus-derived peptide triggered local immune activation. This delivery translated to antineoplastic effects, which improved survival in a murine glioblastoma model. Our results indicate that virus-specific memory T cells are a significant part of the glioblastoma immune microenvironment and may be leveraged to promote anti-tumoral immunity.

Glioblastoma multiforme (GBM) is among the most aggressive, treatment resistant cancers, and despite standard of care surgery, radiation and chemotherapy, is invariably fatal. GBM is marked by local and systemic immunosuppression, contributing to resistance to existing immunotherapies that have had success in other tumor types. Memory T cells specific for previous infections reside in tissues throughout the host and these tissue resident memory T cells (TRM) are capable of rapid and potent immune activation. Here, we show that virus-specific memory CD8+ T cells expressing tissue resident markers populate mouse and human glioblastoma microenvironment. Reactivating virus-specific TRM through intra-tumoral delivery of adjuvant-free virus-derived peptide triggered local immune activation. This delivery translated to anti-neoplastic effects, which improved survival in a murine glioblastoma model. Our results indicate that virus-specific TRM are a significant part of the glioblastoma immune microenvironment and can be leveraged to promote anti-tumoral immunity.

Glioblastoma multiforme (GBM) is among the most aggressive, treatment resistant cancers, and despite standard of care surgery, radiation and chemotherapy, is invariably fatal. GBM is marked by local and systemic immunosuppression, contributing to resistance to existing immunotherapies that have had success in other tumor types. Memory T cells specific for previous infections reside in tissues throughout the host, including the brain, and are capable of rapid and potent immune activation. Here, we show that virus-specific memory CD8+ T cells expressing tissue resident markers populate the mouse and human glioblastoma microenvironment. Reactivating virus-specific memory T cells through intra-tumoral delivery of adjuvant-free virus-derived peptide triggered local immune activation. This delivery translated to anti-neoplastic effects, which improved survival in a murine glioblastoma model. Our results indicate that virus-specific memory T cells are a significant part of the glioblastoma immune microenvironment and may be leveraged to promote anti-tumoral immunity. Supported by UMN SPORE Program Project Planning grant (DM, CCC), NCI 1R01CA238439 (DM), Humor to Fight the Tumor Foundation (JN), NCI 5P30CA023108-42 (PR)

Background Cerebral bypass remains important for the treatment of complex cerebral aneurysms including dissecting, giant, and fusiform aneurysms not amenable to endovascular treatment or simple clip ligation. For such aneurysms involving the anterior communicating artery complex or its branches, distal anterior cerebral artery (ACA) A3-A3 side-to-side bypass represents a valuable treatment option. Distal ACA in situ anastomosis is recognized to be technically demanding mainly due to the relative depth and narrowness of the interhemispheric surgical corridor and type of anastomosis. Objective To demonstrate technical nuances of A3-A3 side-to-side in situ bypass surgery through case illustrations and operative videos. Methods Elements of the procedure relating to positioning, approach, and anastomosis which have evolved in the operative technique of the senior author were collated based on review of clinical case material, imaging and video recordings of ACA aneurysms treated with side-to-side in situ A3-A3 bypass procedure. Technical elements were contrasted with relevant literature. Results Nuances relative to patient positioning, selection of craniotomy variants, adjunctive intraoperative tools and microsurgical nuances of the side-to-side bypass procedure are reviewed. Three illustrative operative video cases, along with illustrations, are provided to complement the description of the nuances. Conclusion In the light of the inherent technical difficulty, as well as the rather limited case volumes, the technical tips provided may contribute to bringing additional refinement and simplicity to the A3-A3 bypass procedure.

BACKGROUND: Heterozygous mutations in Histone H3.3 (K27M, G34R/V) occur in 70–80% of pediatric high-grade gliomas and 78% of diffuse intrinsic pontine gliomas (DIPG). These mutations are also found in low-grade gliomas that progress to higher grade tumors. Epigenetic dysregulation resulting from altered methylation at or near the mutated sites is believed to drive malignant transformation H3 differs from H3.1 by only 5 amino acids, which includes a unique phosphorylation site, Serine 31. S31 is phosphorylated by Chk1 kinase at pericentromeric heterochromatin in mitosis and on the arms of missegregated chromosomes. Masking S31 phosphorylation by antibody injection or using a non-phosphorylatable S31A substitution prevents cell cycle arrest in response to chromosome missegregation. OBJECTIVE: To determine if H3.3 K27M or G34R/V mutations alter the regulation of S31 phosphorylation to promote chromosome missegregation, aneuploidy, and tumorigenesis. METHODS: Live cell imaging and an established mouse model of glioma. RESULTS: DIPG cells with the K27M mutations show decreased mitotic Ser31 phosphorylation and increased chromosome missegregation. In both model cell lines and normal astrocytes, K27M and G34R mutations abolish S31 phosphorylation. Expression of S31A, K27M, or G34R mutants results in increased chromosome missegregation and aneuploidy, whereas cells expressing phosphomimetic double mutants (K27M/S31E or G34R/S31E) divide normally. Using the RCAS/TVA mouse model, H3.3WT, K27M, and S31A constructs were introduced into the brains of newborn mice along with mouse BRAF(V600E). 0/13 tumors developed in the BRAF(V600E) + H3.3WT cohort; however, 80% (16/20) of H3.3K27M + BRAF(V600E) and 70% (7/10) H3.3S31A + BRAF(V600E) cohorts developed high-grade gliomas within 100 days. CONCLUSION: A single heterozygous H3.3 mutation is sufficient to generate aneuploid cells, allows them to reenter the cell cycle and to promote tumorigenesis. Restoring cell cycle checkpoint activity by modulating S31 phosphorylation may be a rational treatment for DIPG or to prevent the progression of low-grade gliomas with H3.3 mutations.