JS07.5.A THERAPEUTIC TIM-3 INHIBITION SUPPORTS NEUROLOGICAL FUNCTION POST-TUMOR CLEARANCE IN DMG PRECLINICAL MODELS

Abstract BACKGROUND Diffuse midline gliomas (DMGs) are among the most aggressive pediatric brain tumors, with very limited treatment options and poor outcomes. Radiotherapy, although still the standard approach, is not curative and often leads to long-term cognitive decline due to therapy-induced neuroinflammation. TIM-3 (T-cell immunoglobulin and mucin-domain containing-3) has recently been identified as a promising target for immunotherapy. In previous work, we demonstrated that blocking TIM-3 significantly improved survival in immunocompetent mouse models of DMG, with 50% of the treated animals achieving long-term remission and immune memory. However, the potential long-term effects of this treatment on brain function have not been explored. This study evaluates whether TIM-3 blockade impacts motor or cognitive abilities in long-term survivors, and investigates its effects on neuroinflammation and brain-resident immune and neuronal populations. MATERIAL AND METHODS We assessed motor skills and cognitive function in long-term surviving mice treated with anti-TIM-3 (anti-TIM-3-LTS) one- and two-months after tumor clearance. Motor coordination was evaluated using the rotarod, pole, and bar tests, while memory and learning were tested using the Novel Object Recognition Test (NORT), Fear Conditioning Test (FCT), and Morris Water Maze (MWM). Neuroinflammatory status was examined through immunostaining to detect reactive astrocytes, activated microglia, and persistent T-cell infiltration. Additionally, we analyzed gene expression levels of pro-inflammatory cytokines and chemokines (TNF-α, IL-1β, IFN-γ) and neuronal markers related to neurogenesis and synaptic function (Dcx, Syn1). RESULTS Anti-TIM-3-LTS mice performed similarly to non-tumor controls in all motor and cognitive tests, and no differences were observed between sexes. Histological analysis revealed no signs of ongoing neuroinflammation: astrocyte and microglia activation had resolved, and no chronic T-cell presence was found in the cortex, hippocampus or cerebellum. Expression levels of inflammatory cytokines and chemokines were comparable to those in control animals, and key neuronal markers remained stable, indicating no disruption of neurogenesis or synaptic integrity. CONCLUSION Our findings show that TIM-3 inhibition not only enhances survival in DMG without compromising neurological function, but also avoids long-term inflammatory damage typically associated with standard treatments. These results support TIM-3 as a strong candidate for immunotherapy in DMG, offering potential to improve outcomes while preserving patients’ quality of life.