Abstract PURPOSE Actionable oncogenic fusion FGFR3-TACC3 (F3T3) is found in 3% of gliomas. Our goal was to characterize the clinical, radiological and molecular features of patients with F3T3-positive glioma. PATIENTS AND METHODS Overall, we screened 1112 gliomas by RT-PCR (861 WHO grade IV, 140 grade III and 111 grade II) and identified 50 F3T3-positive cases. We performed a radiological and radiomic case control study. RESULTS F3T3 fusion was exclusively found in IDH wild-type gliomas. F3T3 was mutually exclusive with the EGFR amplification (0/38 versus 143/336 of F3T3-negative cases, P=0.01), whereas it was associated with CDK4 amplification (7/33 versus 22/321 of F3T3-negative cases, P=0.04) and MDM2 amplification (6/33 versus 15/354 of F3T3-negative cases, P=0.03). F3T3-positive gliomas showed a longer overall survival than F3T3-negative gliomas (median OS 40.1 and 20.0 months, P=0.02), particularly in the grade IV subgroup (40.1 versus 19.0 months, P=0.006). Multivariate analysis showed that F3T3 fusion is an independent predictor of favorable outcome for glioblastoma patients. F3T-positive gliomas were associated with involvement in non-eloquent areas (r=0.31 P=0.007), cortico-subcortical regions and insula involvement (adjusted p<0.001), poorly defined enhancing margins (r=0.48 P<0.001), poorly defined non-enhancing margins (r=0.61 P<0.001), and higher proportion of edema (r=0.43 P<0.001). Radiomics analysis correctly classified F3T3-positive glioma with an AUC of 0.82. We compared different Cox proportional hazards models using Harrel’s C-Index. Although radiomics alone obtained a high C-Index (0.75, SD 0.04), the model combining clinical, genetic and radiomic data showed the highest C-index (0.81, SD 0.04). CONCLUSION Gliomas harboring F3T3 gene fusions show specific molecular features, distinct radiological and radiomic features and a less aggressive clinical evolution.
Abstract The standard of care for adult patients with gliomas, glioneuronal, and neuronal tumors consists of combinations of surgery, radiotherapy, and chemotherapy. For many systemic cancers, targeted treatments are a major part of the standard treatment; however, the predictive significance of most of the targets for treatment in systemic cancer is less well-established in central nervous system tumors. In 2023 the European Association for NeuroOncology (EANO) Guideline Committee presented evidence-based recommendations for rational testing of molecular targets for targeted treatments. From all targets reviewed, only testing for BRAF V600E mutations was of proven clinical benefit; despite regulatory approvals for tumor agnostic treatment of NTRK gene fusions and high tumor mutational burden (TMB) for patients with adult brain tumors, the evidence of clinical benefit for adult patients was still limited. This guideline has a modular structure, allowing regular updating of individual sections and adding new ones. The present version (Update 1) presents a review of the rationale of testing for PTEN, H3F3A, MTAP, RET and IDH, and presents an update of the text on TMB high and mismatch repair deficiency. It also presents an overview of the therapeutic yield of routine next-generation sequencing for mutations and fusion detection. The Supplemental File II accompanying this version contains an in-depth review of all targets, whereas, in the main manuscript, the final recommendations of the revised and new targets are presented. Updates will be made on a regular basis.
Abstract The heterogeneity within tumors has long been associated with therapy failure and disease progression. Recent advances in single-cell RNA-sequencing technologies have enabled us to dissect the cellular diversity in glioblastoma (GBM). However, how these cellular programs change longitudinally under therapy remains poorly understood. To address this question, we collected and profiled a large-scale longitudinal cohort of 59 matched IDH-wildtype GBM sample pairs provided by 7 centers worldwide by single-nucleus RNA-sequencing (snRNA-seq) and whole-exome/whole-genome sequencing. The majority of patients (51 patients) in this cohort received standard-of-care (temozolomide and radiation) following initial tumor resection. Leveraging this large-scale snRNS-seq dataset of 457,442 cells, we detected novel cellular states and associations between malignant and tumor microenvironment (TME) cells, and then performed longitudinal analyses. The recurrent samples showed significantly lower malignant cell fraction (p=0.002) and reciprocal increase in proportions of glio-neuronal TME cell types (oligodendrocytes, neurons and astrocytes). The TME composition, malignant cell state proportions and baseline expression programs were retained at recurrence more than expected by chance, but overall were not well conserved between primary and recurrent samples. A subset of pairs (12/59 pairs) enriched with glio-neuronal TME at recurrence showed a significant transcriptomic shift and was associated with better clinical course (p=0.02). The tumors that acquired radiation-related small deletion phenotype underwent a transition towards MES/Hypoxia phenotype (0/9 at primary, 6/9 at recurrence, p=0.02). We defined pairs as likely responders or non-responders to treatment based on the MGMT methylation status of the primary tumor sample. This uncovered diverging evolutionary trajectories in cellular programs between the two groups. Strikingly, the changes in malignant state frequency and baseline malignant expression profile were strongly associated with specific changes in the TME composition. Our findings based on high-resolution longitudinal snRNA-seq analyses highlight the diverse evolutionary trajectories in GBM that are shaped by TME changes and treatments.
Abstract Neurofibromatosis type 1 (NF-1) is the most common cancer predisposition syndrome in which 15-20% of affected individuals develop glioma. Large scale DNA and RNA bulk profiling showed the molecular complexity of NF-1 glioma with the tumor cellular ecosystem constituted by multiple malignant phenotypes and heterogenous immune microenvironment. However, the composition and function of infiltrating cells was hidden in the bulk tumor, and the extended granularity of NF-1 glioma tumor microenvironment (TME) remained still unexplored. Here, we collected glioma samples from 46 NF-1 patients including 22 high-grade (HGG) and 24 low-grade (LGG) tumors, and we analyzed their gene expression by single nuclei RNA sequencing. A total of 239,044 single cells were classified into tumor and non-tumor components by integrating multiple computational approaches (including genomic copy number inference, gene signature enrichment, and clustering). We defined the pattern of intra-tumor heterogeneity of NF-1 glioma cells using non-negative matrix factorization and derived 7 malignant meta-programs (MPs) that we respectively defined as Neuronal-like, EMT, Astrocyte-like, Dividing Radial Glia-like, Ependymal-like, Immune, and Glycolytic/Hypoxic-like. These MPs recapitulated normal brain cell subtypes, thereby reflecting broad cell plasticity. The non-tumor cell compartment (121,364 cells, 51%) was dissected for the characterization of the cell types that populate the TME of NF-1 glioma. We identified different subpopulations exhibiting specific immune functions within myeloid and lymphoid components. Different glioma ecomodules were highlighted by comparing the relative composition of the TME across the tumors. Recruitment and activation of cytotoxic CD8+ T cells and natural killers by an active crosstalk with dendritic and pro-inflammatory myeloid cells defined an immune-supportive phenotype that could mediate a potential anti-tumor response in low-grade NF-1 glioma (LGG immune high). Conversely, regulatory T cell infiltration and effector T cell exhaustion induced immune suppression in a low-grade glioma immune dysfunctional ecomodule. The absence of lymphocytes characterized a large set of cold tumors, mostly including high-grade glioma. Together, the complex interplay of tumor cell states with different TME compartments elucidated the existence of separate ecomodules in NF-1 glioma, with the LGG immune high TME associated with Neuronal-like and the LGG immune dysfunctional with Ependymal-like tumor cells. The Ependymal-like state also exhibits maximal association with brain-specific normal cells, including oligodendrocytes, neurons and astrocytes, whereas the HGG are enriched with Dividing Radial Glia- and Glycolytic/Hypoxic-like tumor cell states. The elucidation of different ecomodules provides novel insights for the application of targeted therapies in NF-1 glioma patients. Citation Format: Luciano Garofano, Fulvio D'Angelo, Michael Oh, Michele Ceccarelli, Franck Bielle, Marc Sanson, Anna Lasorella, Antonio Iavarone. Identification of distinct tumor-TME ecomodules in glioma from neurofibromatosis type 1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6938.
Abstract Glioblastoma multiforme (GBM) is the most aggressive form of primary brain tumor, with no curative treatment options. Multiple studies have characterized at single cell resolution the GBM as being composed of transcriptional cell states interconnected with components in the tumor immune microenvironment (TME). Our group proposed and validated the first single cell guided functional classification of GBM in four tumor-intrinsic cell states which informed clinical outcome and delivered therapeutic options. However, single cell technologies are unable to unravel the spatial relationships among the cell states of GBM and between GBM cell states and TME. Spatially resolved transcriptomic technologies are emerging as powerful tools to reconstruct the spatial architecture of a tissue. We performed spatial transcriptomics of multicellular regions of interest (ROI) in 6 primary IDH wild-type GBM and 2 recurrent GBM with both CosMx Spatial Molecular Imager, which analyzes 1,000 RNA probes and 64 proteins at single cell resolution, and GeoMx Digital Spatial Profiler which profiles the whole transcriptome (~18,000 genes) at ROI resolution. The development of computational tools aimed to integrate spatial proximity and CosMx derived single-cell transcriptomics revealed spatial segregation of the tumor cell clones and cellular states and highlighted recurrent patterns of cell states, distinct TME cell types associated with coherent histopathological features across multiple samples. The development of a spatial informed intercellular communication algorithm and the reconstruction of ligand-receptor-target networks will allow the discovery of tumor cell states-TME cross-talks and the biological signaling regulated by these interactions that are driving the heterogeneity of GBM and therefore potentially therapeutically targetable. Analysis of matched regions of interest profiled by GeoMx and spatial proteomics with CosMx further cross-validated the spatial ecosystem of glioblastoma as reconstructed at single-cell resolution. Our studies established a scalable approach to resolve the transcriptional heterogeneity of GBM and reconstruct the architecture of GBM cell states and tumor microenvironment. Citation Format: Bruno Adabbo, Simona Migliozzi, Luciano Garofano, Fulvio D'Angelo, Pedro Davila, Sakir H. Gultekin, Daniel Bilbao Cortes, Benjamin Currall, Sion L. Williams, Marc Sanson, Franck Bielle, Anna Luisa Di Stefano, Michele Ceccarelli, Anna Lasorella, Antonio Iavarone. Reconstruction of the spatial ecosystem of glioblastoma reveals relationships between tumor cell states and microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1151.
