Abstract 5545: Neutralization of interleukin-10 by genetically engineered macrophages increase cancer cell death in metastatic colorectal cancer

Abstract

Abstract Background Interleukin-10 (IL-10) is an immunosuppressive cytokine that inhibits innate and adaptive antitumor immune responses in the tumor microenvironment (TME). Antibody neutralization of IL-10 in organotypic tumor slice cultures (TSC) in colorectal cancer liver metastases (CRCLM) leads to cancer cell death through. To target IL-10 blockade to the TME, we genetically engineered macrophages (GEM) to produce an IL-10 neutralizing antibody which we hypothesized would reactivate tumor infiltrating immune cells and increase cancer cell death in TSC. Methods We created a lentiviral construct to transduce GEMs with a publicly available neutralizing IL-10 antibody sequence (BT-063). CD14+ monocytes were isolated from healthy donor peripheral blood mononuclear cells and differentiated into macrophages, which were transduced with lentivirus to create control (cGEMs) and anti-IL-10 GEMs (αIL10 GEMs). To test the efficacy of αIL10 GEMs, punch biopsies (6 mm) were obtained from resected CRCLM, sliced to 250 um thickness with a vibratome and placed in individual culture wells. Slices were treated in triplicates with either 20ug/mL of human monoclonal αIL10 (αIL10), 1e5 cGEMs or αIL10 GEMs + 20ug/mL αIL10. GEMs and tumor cells were visualized with a Leica SP8X confocal microscope and apoptosis was measured with SR-FLICA fluorescent poly-caspase assay kit. Supernatant interferon-gamma (IFN-γ) levels were measured with Bioplex immunoassay and IL-10 neutralizing antibody was quantified with an enzyme-linked immunosorbent assay. Results Allogenic GEMs were transduced with BT-063-T2A-CD19t at 31% efficiency. cGEMs and αIL10 GEMs persisted in tumor slice culture for at least 6 days. αIL10 GEM-treated slice cultures contained 6.6ng/mL and 6.5ng/mL of αIL10 antibody after 3 and 6 days, respectively. At day 6 in culture, cancer cell apoptosis was increased in the αIL10 treated groups with 7%, 15%, 25%, 31% and 39% of cells were apoptotic in the no treatment control, cGEM, αIL10, cGEM + αIL10 and αIL10 GEM groups, respectively (One-way ANOVA, p = 0.0004). Elevation in IFN-γ was observed on day 6 in slices treated cGEMs (7.2ng/mL+4.3) and αIL10 GEMs (9.2ng/mL+5) compared to isotype control. Conclusion GEMs transduced with lentivirus encoding IL-10 neutralizing antibody maintain its production and are associated with increased cancer cell apoptosis. Citation Format: Kevin P. Labadie, Shannon A. Kreuser, Katherine J. Brempelis, Xiuyun Jiang, Sara K. Daniel, Courtney A. Crane, Venu G. Pillarisetty. Neutralization of interleukin-10 by genetically engineered macrophages increase cancer cell death in metastatic colorectal cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5545.