Abstract P2-04-12: Targeting of phosphatidylserine by monoclonal antibodies enhances the activity of immune checkpoint lag3 targeting antibodies in murine breast tumors
Authors: Gray MJ , Gong J , Hutchins JT , Freimark BD 
Published: March 2017

Abstract
Previous preclinical modeling has demonstrated that antibodies targeting the programmed cell death ligand 1 (PD-1) have activity in murine breast cancers (BC), and this activity is significantly enhanced when combined with phosphatidylserine (PS) targeting antibodies. This suggested that the addition of PS targeting antibodies could be capable of augmenting anti-tumorigenic properties of other checkpoint inhibitors and alternative immune activating therapies in BC. The ability of PS targeting antibodies to enhance the activity of anti-PD-1 therapy occurs in part through increasing tumor infiltrating lymphocytes (TILs), boosting the Th1 immuno-profile, and the suppression of tumor promoting cytokines induced by anti-PD-1 therapy. PS normally resides in the inner leaflet of the plasma membrane in many types of cells. Conditions that incite cellular stress in the tumor microenvironment, such as ROS, hypoxia, and irradiation, promote PS externalization and exposure on tumor associated endothelial cells and tumor cells, where it is recognized by specific receptors, including members of TIM and TAM family. This PS recognition promotes an innate system driven immunosuppressive condition in part by promoting the recruitment of myeloid derived suppressor cells (MDSCs), M2-like macrophages, and suppressing dendritic cells maturation, while inducing anti-inflammatory cytokines. To identify additional immuno-therapeutic targets that may have activity when used in combination with PS targeting antibodies, we employed bioinformatics analysis on tumors from murine triple negative breast cancer (TNBC) treated with PS and PD-1 targeting antibodies, alone or in combination. Interestingly, expression of the lymphocyte activation gene 3 (LAG3) increased in each treatment arm, suggesting it may be a potential target for combinational therapy in breast cancers. To test this hypothesis, immune competent mice harboring the murine TNBC line E0771 were treated with a PS targeting antibody or a LAG3 targeting antibody as single agent or in combination. Our data demonstrate that while PS-blocking and anti-LAG3 therapies each have efficacy in E0771 as single agents, combinational treatment significantly improved growth inhibition and was capable of increasing TILs, including CD8 + and CD3 + T-cells, while reducing the population of MDSCs. Overall, our data suggest that LAG3 targeting may also represent a viable option for the treatment of breast cancer and that the addition of PS targeting antibodies to LAG3 therapy can effectively increase the anti-tumor and immune-activating effects mediated by additional T-cell checkpoint therapies.
Citation Format: Gray MJ, Gong J, Hutchins JT, Freimark BD. Targeting of phosphatidylserine by monoclonal antibodies enhances the activity of immune checkpoint lag3 targeting antibodies in murine breast tumors [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P2-04-12.