457 Semaphorin7a expression in breast cancer promotes susceptibility to immune checkpoint blockade

Abstract

OBJECTIVES/GOALS: The goal of this study is to decipher the mechanisms by which breast cancers evade the immune system to facilitate metastasis through the lymphatic vasculature. We believe this is caused, in part, by semaphorin 7A (SEMA7A) as its expression is associated with increased lymphatic vessels, tumor-associated macrophages, and metastasis in breast cancer. METHODS/STUDY POPULATION: We have observed that breast cancer cells, lymphatics, and macrophages exhibit SEMA7A-dependent expression of PD-L1 in vitro, which may make SEMA7A+ tumors more sensitive to anti-PD-1/PD-L1 immunotherapies. Therefore, we utilized multiple orthotopic, immunocompetent mouse models to determine if SEMA7A expression makes tumors more susceptible to immune checkpoint blockade. E0771 and 66cl4 mouse mammary carcinoma cells were injected orthotopically into the #4 mammary fat pads. Once tumors reached 50-100 mm^3 mice were injected intraperitoneally with 250ug of anti-PD-1, anti-PD-L1, or IgG control every third day. Tumors were measured with calipers every other day and harvested for flow cytometry to determine the effect of immune checkpoint blockade on the TME in SEMA7A+ tumors. RESULTS/ANTICIPATED RESULTS: We reveal that growth of SEMA7A overexpressing (OE) tumors, but not controls, was significantly slowed with both anti-PD-1 and anti-PD-L1 treatments. Flow cytometric analysis of cells from the TME revealed increased LECs, TAMs, and PoEMs in SEMA7A+ tumors, compared to controls – all populations had higher PD-L1 expression, which was decreased with both anti-PD-1 or anti-PD-L1. We also observed a decrease in PD-L1 expression on the tumor cells with treatment. Furthermore, LEC, TAM, and PoEM presence was decreased within the TME alongside increased presence of activated CD4 and CD8 T cells. Collectively, our results suggest that SEMA7A expression in breast cancers activates a major pathway associated with immune evasion and helps to establish a pro-tumor microenvironment. DISCUSSION/SIGNIFICANCE: SEMA7A expression establishes a pro-tumor microenvironment, which can be targeted with readily available FDA approved drugs such as immune checkpoint-based therapies. Since SEMA7A+ breast cancers have high rates of metastasis, more specific treatments for these patient populations should be explored.