Beschreibung:
Abstract Purpose of Study: Diffuse midline glioma (DMG) is an aggressive brain tumor and the leading cause of pediatric death caused by cancer. Despite great strides in the understanding of this disease, survival is still dismal. One of the objectives of our lab is to modulate the tumor microenvironment (TME) towards a proinflammatory phenotype to render these tumors amenable to immunotherapy. TIM-3 is a member of the TIM family of immunoregulatory proteins expressed on multiple immune cell types, including T-cells, NK, myeloid populations, and microglia, regulating adaptive and innate immunity. Therefore, the aim of this project is to study the antitumor effect of the anti-TIM-3 monoclonal antibody and its effect on the DMG tumor microenvironment. Experimental Procedures: TIM-3 expression in DMG patients was analyzed using total mRNA sequencing data and single-cell RNAseq data. To perform all the experiments, DIPG murine cell lines were used (NP53 and XFM). For in vivo experiments, cells were injected into the pons of mice using a screw-guided system. The antibody was administered intracranially (25µg) with the same system and two times intraperitoneally (10mg/kg) 3, 7, and 11 days after the cell implantation respectively. Tumor immune populations, chemokines, and cytokines were analyzed by flow cytometry. Results: In silico assessment of TIM-3 expression in DMG mRNA and single-cell datasets showed a robust expression of this gene mainly in microglia and macrophages uncovering this molecule as a potential target in DMGs. In vivo studies showed that TIM-3 blockade with an antibody significantly increased the overall survival of two DMG immunocompetent orthotopic models, led to long-term survivors (50%), and showed immune memory. TIM-3 inhibition resulted in significant increase in the number and proliferative state of microglia, NK, and CD8+ cells and higher levels of IFNγ, GrzB and TNFα corresponding to NK and T-cell activate phenotypes. Interestingly, there was a decrease in the Treg population, which caused an increase in the pro-inflammatory CD8/Treg ratio. Chemokine studies demonstrated an augmentation of CCL5, CCL2 chemotactic chemokines, and CXCL10, IL-1β and IFN-γ pro-inflammatory cytokines in the tumor microenvironment of treated mice. Additionally, DCs, CD4+, and CD8+ cells were increased in treated draining lymph nodes and of functional significance, expressed higher amounts of pro-inflammatory cytokines than in control mice. Interestingly, the depletion NK, CD4 and CD8 immune populations did not completely abrogate the treatment efficacy. However, microglia and macrophages depletion with an anti-CSF1R resulted in a total loss of efficacy indicating a critical role of these populations in the effect of TIM-3 blockade. Conclusions: These data uncover TIM-3 as a potential target for the treatment of DMG and its role as an immune regulator of the DMG tumor microenvironment. Citation Format: Iker Ausejo-Mauleon, Sara Labiano, Virginia Laspidea, Daniel de la Nava, Oren Becher, Mariella G Filbin, Fernando Pastor, Marta M Alonso. Modulation of tumor microenvironment with TIM-3 blockade improves survival in diffuse midline glioma models. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6369.