TIM-3 blockade as a therapeutic approach for diffuse intrinsic pontine glioma

Abstract

Diffuse intrinsic pontine gliomas (DIPG) is an aggressive brain tumor and the leading cause of pediatric death caused by cancer. Despite great strides in understanding this disease, prognosis is dismal, with over 90% of patients dying within two years of diagnosis and a median overall survival time of 9-12 months. These grim statistics underscore that DIPG is an unmet clinical need. In this doctoral thesis, I have evaluated the local administration of an anti-TIM-3 antibody in anti-TIM-3 antibody in syngeneic orthotopic models of DIPG as a therapeutic approach for this disease. Our work uncovered, through in silico studies in patient datasets (whole RNAseq and scRNAseq) and samples (by multiplex IF) that TIM-3 was robustly expressed in tumor cells and tumor microenvironment, mainly in microglia and macrophages, suggesting this molecule as a potential therapeutic target in DIPGs. Mechanistic studies showed that TIM-3 provided intrinsic survival cues to the tumor cell while modulating the tumor microenvironment when expressed in the myeloid compartment. In vivo studies showed that TIM-3 blockade significantly increased the overall survival of DIPG immunocompetent orthotopic models, led to long-term survivors, and showed immune memory. TIM-3 inhibition resulted in an increase in the number and proliferative state of microglia, NK cells, and CD8+ T cells and higher levels of IFN, GrzB, and TNF&945; 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-1;, and IFN- pro-inflammatory cytokines in the tumor microenvironment of treated mice. Additionally, DCs, CD4+, and CD8+ T 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 of NK cells, CD4+, and CD8+ T cells immune populations did not completely abrogate the treatment efficacy. However, microglia and macrophages depletion with an anti-CSF1R resulted in a total loss due to a loss of microglia and CD8 T cells pro-inflammatory populations, chemokines, and cytokines indicating a critical role of these populations in the therapeutic effect of TIM-3 blockade. This study provides a new and previously unstudied view of DIPG treatment. TIM-3 blockade emerges as an exciting alternative to classical immune checkpoints, such as PD-1, that did not obtain the desired results in DIPG clinical trials. Moreover, the lack of other effective therapies for these devastating pediatric brain tumors makes these pre-clinical results especially promising and offers strong support for initiating a clinical trial with an anti-TIM-3 antibody for the treatment of DIPG.