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Abstract 2725: Galectin-9 drives TIM-3 mediated immune suppression

Purpose: Following recent Immunotherapy successes by targeting the checkpoints PD1, PD-L1, and CTLA-4, additional checkpoint inhibitors and their counter-structures are being explored as therapeutic targets to reverse T cell exhaustion and increase T-cell anti-tumor activity. One such inhibitory receptor, TIM3, has been reported to be expressed on severely dysfunctional T cells in preclinical cancer models and in multiple human cancers. While TIM3 expression is significantly upregulated on TILs, including exhausted CD8+ T cells and Tregs, constitutive expression of TIM3 on myeloid cells has also been shown to negatively regulate anti-tumor activity of T cells. Inhibitory signaling in T-cells is activated by engagement of TIM3 by Galectin 9 (GAL9, LGALS9) and putatively by ligands CEACAM1, HMGB1, and Phosphatidylserine. GAL9 has also been shown to induce T cell apoptosis in the TME. Given the complex nature of TIM3 biology, we sought to characterize expression of TIM3 and its ligands across multiple tumor indications. Experimental Design: TIM3, LGALS9, CEACAM1, HMGB1, and signatures capturing T and myeloid populations were examined in TCGA RNASeq data. TIM3 and LGALS9 in various cell subsets was evaluated using public single-cell RNASeq. Multiplex immunofluorescence (mIF) staining was performed on tumor TMAs procured from commercial sources. Results: Analysis of TCGA data showed that TIM3 expression strongly correlates with macrophages and with T cells. TIM3 expression strongly correlates with LGALS9 but not with CEACAM1 or HMGB1 across essentially all TCGA indications. LGALS9 expression was observed to correlate more strongly with T cells, macrophages, and IFNγ expression signatures compared to HMGB1 and CEACAM1, suggesting it may be the most relevant ligand as adaptive mechanism of resistance to cytotoxic activity. Interestingly, the genes most correlated with TIM3 and its ligands were largely distinct between cell types in the single-cell data. This allows for cell type-specific expression signatures to predict levels of these genes from bulk tumor expression data. Such signatures combined with overall survival data from TCGA suggest that TIM3+ T cells are active in immune response and that TIM3+ macrophages have a negative predictive value. Experiments to confirm findings for TIM3 and its ligands from TCGA using mIF analysis are ongoing. Results will be reported in detail in the poster. Conclusions: These results suggest that interaction of TIM3 and GAL9, as opposed to its other ligands, may be an important mechanism of immune suppression in the TME. Thus, blocking TIM3/GAL9 interaction would potentially reactivate anti-tumor activity by modulating both T cell intrinsic and extrinsic functions involving multiple immune cells subsets. Future correlative and clinical investigations should consider the potential combined role of TIM3 and Gal9 for development of next generation IO therapeutics. Citation Format: Amit M. Deshpande, Rachel Fontana, Yue Zhang, Mary Ruisi, Dong Zhang, Tilo Senger, P. Alexander Rolfe. Galectin-9 drives TIM-3 mediated immune suppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2725.