Abstract 581: Determining the influence of galectin-3 binding properties on its suppression of tumor antigen-specific CD8+ T cells
Authors: Alexandra B. Pucsek, Todd D. Armstrong, Elizabeth M. Jaffee
Journal: Cancer Research (2019)

Abstract Improving the efficacy of immunotherapeutic cancer treatments requires increased understanding of the factors that contribute to a suppressive tumor microenvironment (TME). Recently, galectin-3 has emerged as an important player in the development and progression of cancer, favoring immune suppression and dysfunction at disease sites. While this protein is consistently identified as a tumor-promoting factor, its unique properties make the mechanisms of its functions difficult to study, and impede the development of therapies against it. Galectin-3 binds β-galactoside carbohydrate residues through a highly-conserved C-terminal domain (CRD) characteristic of mammalian galectins. It is made unique by its ability to self-associate into homo-oligomers through a disordered N-terminal domain (NTD). We have previously shown that galectin-3 knockout in a mouse model of tumor tolerance improves effector function of antitumor CD8+ T cells and increases duration of tumor-free survival, suggesting a suppressive role for galectin-3 in antitumor immunity. Our current work aims to elucidate how the distinct biochemical properties of galectin-3 contribute to this suppression. To this end, we are developing a new assay to model the suppression of CD8+ T cells by galectin-3 in vitro. Our assay utilizes TCR transgenic CD8+ T cells specific for rodent Erbb2, the Her-2/neu (Neu) oncogene. These cells are isolated from either normal transgenic mice or those deficient in galectin-3. The T cells are stimulated in culture, then co-cultured with NT2.5 Neu-expressing tumor cells. We use flow cytometry to analyze the levels of CD8+ effector molecules interferon gamma (IFNγ) and Granzyme B, as well as cell surface binding of galectin-3 after co-culture. To further elucidate the roles of carbohydrate binding and self-association in this process, we have developed recombinant mutant galectin-3 proteins deficient in carbohydrate binding through CRD point mutation, lacking the ability to self-associate through NTD truncation, or both. By introducing these recombinant proteins into the co-culture with normal or galectin-3 deficient transgenic T cells, we expect to be able to learn whether galectin-3-mediated suppression is more dependent on carbohydrate binding, self-association, or both properties in synergy. Current therapies targeting galectin-3 focus only on its carbohydrate binding properties through use of small molecules and glycomimetics, but these have been of limited efficacy. We hypothesize that self-association of galectin-3 through the NTD is an equally important factor in its unique ability among galectins to contribute to an immunosuppressive TME. Learning how these properties each contribute to the mechanism of galectin-3-mediated immunosuppression will allow us to refine strategies for breaking tumor tolerance, contributing to improved outcomes for cancer immunotherapies. Citation Format: Alexandra B. Pucsek, Todd D. Armstrong, Elizabeth M. Jaffee. Determining the influence of galectin-3 binding properties on its suppression of tumor antigen-specific CD8+ T cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 581.