Background: Siglec-15 is an immunosuppressive sialic acid-binding Ig-like lectin expressed by myeloid cells, tumor associated macrophages (TAMs) and some human tumors. Siglec-15 expressed by TAMs inhibits anti-tumor immune responses by engaging unknown immune checkpoint(s) on T cells. Interactions between Siglec-15 on TAMs and sialy Tn (sTn) antigen found on tumor cells contributes to the immunosuppressive tumor microenvironment. Notably, the mutually exclusive expression of Siglec-15 and the checkpoint ligand PD-L1 by cancer cells emphasizes Siglec-15 as an attractive target for cancer immunotherapy.
Methods: Anti-Siglec-15 antibodies were cloned from B cells derived from rabbits immunized with human Siglec-15. Cells were cultured at clonal density, and IgG antibodies in supernatants were evaluated for binding to human, murine and cynomolgus Siglec-15. Variable-regions from positive hits were sequenced, cloned, and expressed as recombinant rabbit human IgG1 Fc chimeras. Anti-Siglec-15 chimeric antibodies were evaluated in a panel of functional and phenotypic assays using primary human macrophages and T cells, and then prioritized for evaluation in murine tumor models.
Results: Thirty-one rabbit anti-Siglec-15 clones were expressed as rabbit-human IgG1 chimeras based on binding to recombinant human, cynomolgus and murine Siglec-15 proteins, binding to Siglec-15 expressing cell lines, and lack of binding to other Siglec family members. A subset of these clones inhibited the binding of Siglec-15 to sTn. The top 5 clones were identified in functional screens modeling Siglec-15 mediated immune suppression. The clones rescued the NFAT promotor activity of a T cell reporter cell line as well as the proliferative and IFN-γ response of anti-CD3 activated human T cells from the inhibitory activity of recombinant Siglec-15-Fc protein. Selected clones also relieved M2c-macrophage-mediated immune suppression in M2c/T cell coculture assays by restoring T-cell proliferation and IFN-γ secretion. The prioritized clones are currently under evaluation in in vivo tumor models for PK and efficacy.
Conclusions: We identified novel anti-Siglec-15 antibodies that restore T cell effector function following suppression by recombinant and cell-expressed Siglec-15. These data support further development of these anti-Siglec 15 antibodies as an anti-cancer immunotherapy.
Citation Format: Myriam Bouchlaka, Huyen Dinh, Sam Lam, Valerie Wall, Darbie Whitman, Ramya Chandrasekaran, Johanna Harshman, Texia Loh, Lauren Loh, Tom Graddis, Kamal D. Puri, Peter Probst. Discovery and preclinical characterization of anti-Siglec-15 antibodies that rescue T cells from macrophage-mediated immune suppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1638.