Abstract 2794: Expression of LAP, latency-associated peptide of TGFb, on immune cell subsets (2019)

Abstract LAP, latency-associated peptide of TGFβ1, is a known marker of regulatory T cells in the tumor microenvironment. TGFβ is a major tolerogenic cytokine that is co-opted by tumors to evade the immune system and is implicated in resistance to checkpoint inhibitors. LAP cages TGFβ in an inactive, latent state until it is activated and released by integrins, MMPs or other activators. Anti-LAP antibodies inhibit the release of TGFβ, inhibit tumor growth in mouse models (Gabriely et al., 2017), and have promise as novel cancer therapeutics. Anti-LAP antibodies are thought to mediate anti-tumor activity both by inhibiting the release of active TGFβ and by reducing the number of LAP+ immunosuppressive cells in the tumor microenvironment (TME). To provide insight into the cellular sources of TGFβ in the TME and further delineate the mechanism of action of anti-LAP antibodies in tumor growth inhibition we determined the expression profile of LAP on the surface of both Treg and other immune cell subsets derived from tumor, spleen, and human PBMCs. LAP expression on the surface of various mouse and human immune cell subsets was determined using flow cytometry. Tumor infiltrating leukocytes were profiled from CT26 and 4T1 syngeneic mouse tumor models. Expression of LAP was detected on a subset of regulatory T cells, M2 macrophages, dendritic cells, G-MDSC and M-MDSC populations in both CT26 and 4T1 tumors suggesting that each of these cell types are a potential source for TGFβ activation in the tumor microenvironment. Striking differences were detected in LAP expression on immune cell subsets in the TME versus spleens of CT26 tumor bearing mice, consistent with TGFβ expression being increased upon immune cell activation. Additionally, CD4+ cells were isolated from human PBMCs from normal healthy donors and activated using CD3/CD28 +IL2 stimulation. Consistent with previous reports, activated Foxp3+ T cells were positive for cell surface LAP. In addition, LAP expression was also assessed on various human macrophage subsets (M1, M2a, M2b and M2c) that were skewed from CD14+ cells isolated from PBMCs. LAP expression was detected in varying amounts on the surface of the different macrophage subsets with M2a macrophages demonstrating the highest expression. Taken together these results demonstrate that LAP is expressed on a variety of immune cells with known immunosuppressive function. The location of the LAP-TGFβ1 complex is of critical biological and clinical importance because, once the mature TGFβ1 cytokine, which has a short half-life in solution, is released, it acts locally, either in an autocrine or near paracrine fashion. These results warrant further research to determine the effectiveness of anti-LAP in inhibiting the release of TGFβ and its effects on immunosuppression in the tumor microenvironment. Citation Format: Stavros Kopsiaftis, Patricia E. Rao, Randall Burton, Jessie M. English, Barbara S. Fox, Kenneth J. Simon. Expression of LAP, latency-associated peptide of TGFb, on immune cell subsets [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 2794.