Development of a AXL/PD-1 BiCE as novel immunotherapy agent

Only five to twenty percent of cancer patients respond to anti-PD1 therapy. Recent evidence suggests the involvement of tyrosine kinase receptor AXL in resistance to anti-PD1 therapy, and AXL inhibition sensitize tumors to anti-PD1. In this work we show that AXL expression is associated with exhaustion of CD8T+ cells in the multiple solid cancers, and thus hypothesized that development of Bi-specific-Cell-Engeger (BiCE) antibody that targets AXL and PD1 (BiCE AXL/PD1) will increase the interaction between the AXL-expressing tumor cell and CD8+ T cells and will enhance the anti-tumor lytic activity of the T cells against AXL-expressing tumor cells. To this end, we generated the BiCE AXL/PD-1 containing two single-chain variable (scFv) domains against AXL and PD-1 fused to Fc fragment and tested its ability to bind the target in vitro using ELISA and SPR, and to cells by flow cytometry. Moreover, using a co-culture system of human cell lines and native human peripheral blood mononuclear cells (PBMCs) we showed the ability of the BiCE to enhance the killing of tumor cells by PBMCs which also secrete IFNg. In vivo, the BiCE AXL/PD1 did not induce any major toxicity and demonstrated anti-tumor activity in humanized mice in two xenograft tumor models. We further developed a murine version of the AXL/PD1 BiCE and showed its anti-tumor activity in three mouse cancer models, and spatial multiplexing analysis confirmed the massive infiltration of CD8 T+ cells and their granzyme production in tumor treated with BiCE AXL/PD1 compared to the combination of anti-PD1 and anti-AXL. Altogether, our work shows that BiCE AXL/PD-1 is a novel potential therapeutic agent that can enhance the elimination of metastatic AXL-expressing tumor cells.