Abstract 4054: Macrophages engineered with cytokine switch receptors: Development of a modular platform for rebalancing inflammation in microenvironments

Abstract Motivation: Cytokines in tissue microenvironments regulate the balance between pro- and anti-inflammatory signals. Dysregulated cytokines cause deleterious immunosuppression or inflammation, which underpins the pathophysiology of solid tumors, chronic kidney disease, and more. Rebalancing inflammation/immunosuppression by rectifying cytokine signals offers a generalizable approach for treating numerous diseases. While doing so through cytokine blockade carries risks due to systemic administration, cellular immunotherapies offer a localized approach that could detect pathogenic cytokines then proportionately rebalance inflammation as needed. Specifically, macrophages are homeostatic regulators responsible for initiating and resolving inflammation. Here, we leveraged macrophages’ ability to regulate inflammation by equipping them with synthetic cytokine switch receptors (SR) that convert immunosuppressive M2 signals into pro-inflammatory M1 responses for solid tumor microenvironment conversion, or vice versa for inflammatory disease. We termed this platform “Engineered Microenvironment Converters” (EM-C) and evaluated its modular ability to target disease-associated cytokines. Methods: EM-Cs targeting IL10, TGFβ, IFNγ and IL17A were generated by expressing SR in primary human macrophages. M2-to-M1 SR were designed to convert IL10 or TGF-β into pro-inflammatory stimuli, and M1-to-M2 SR were designed to IFNγ or IL17A into immunosuppressive signals. The in vitro response of EM-Cs to their target cytokine was monitored using phenotypic characterization of surface molecules, measurement of cytokine production, mRNA sequencing, and biochemical analysis of downstream signaling. Co-culture assays with bystander cells were used to assess the ability of EM-Cs to alter their microenvironment. Results: Pro-inflammatory EM-Cs converted IL10 and TGFβ, two prevalent immunosuppressive cytokines in the TME, into pro-inflammatory signals. Unlike wildtype macrophages, these EM-Cs responded to IL10 or TGFβ with upregulated M1 markers and cytokines in a dose-dependent manner. Furthermore, EM-Cs repolarized bystander M2 macrophages towards a pro-inflammatory phenotype following co-culture. Similarly, anti-inflammatory EM-Cs responded IFNγ and IL17A, two cytokines canonically overexpressed in inflammatory disease, by upregulating M2 markers and inducing an anti-inflammatory environment. Conclusion: We present for the first time a novel immunotherapy platform that harnesses macrophages as “living converters” to locally regulate inflammation for oncology and inflammatory applications. By demonstrating EM-Cs in the M2-to-M1 and M1-to-M2 direction, this platform offers modularity in controlling the inflammatory status of tissue microenvironments without systemic cytokine antagonism. Citation Format: Chris Sloas, Yuhao Huangfu, Rehman Qureshi, Michael Ball, Thomas Condamine, Michael Klichinsky, Yumi Ohtani. Macrophages engineered with cytokine switch receptors: Development of a modular platform for rebalancing inflammation in microenvironments. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4054.