Combining SiRPα decoy–coengineered T cells and antibodies augments macrophage-mediated phagocytosis of tumor cells
Evangelos Stefanidis, … , George Coukos, Melita Irving
Evangelos Stefanidis, … , George Coukos, Melita Irving
Published June 3, 2024
Citation Information: J Clin Invest. 2024;134(11):e161660. https://doi.org/10.1172/JCI161660.
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Research Article Immunology

Combining SiRPα decoy–coengineered T cells and antibodies augments macrophage-mediated phagocytosis of tumor cells

Abstract

The adoptive transfer of T cell receptor–engineered (TCR-engineered) T cells (ACT) targeting the HLA-A2–restricted cancer-testis epitope NY-ESO-1157–165 (A2/NY) has yielded favorable clinical responses against several cancers. Two approaches to improve ACT are TCR affinity optimization and T cell coengineering to express immunomodulatory molecules that can exploit endogenous immunity. By computational design we previously developed a panel of binding-enhanced A2/NY-TCRs including A97L, which augmented the in vitro function of gene-modified T cells as compared with WT. Here, we demonstrated higher persistence and improved tumor control by A97L–T cells. In order to harness macrophages in tumors, we further coengineered A97L–T cells to secrete a high-affinity signal regulatory protein α (SiRPα) decoy (CV1) that blocks CD47. While CV1-Fc–coengineered A97L–T cells mediated significantly better control of tumor outgrowth and survival in Winn assays, in subcutaneous xenograft models the T cells, coated by CV1-Fc, were depleted. Importantly, there was no phagocytosis of CV1 monomer–coengineered T cells by human macrophages. Moreover, avelumab and cetuximab enhanced macrophage-mediated phagocytosis of tumor cells in vitro in the presence of CV1 and improved tumor control upon coadministration with A97L–T cells. Taken together, our study indicates important clinical promise for harnessing macrophages by combining CV1-coengineered TCR–T cells with targeted antibodies to direct phagocytosis against tumor cells.

Authors

Evangelos Stefanidis, Aikaterini Semilietof, Julien Pujol, Bili Seijo, Kirsten Scholten, Vincent Zoete, Olivier Michielin, Raphael Sandaltzopoulos, George Coukos, Melita Irving

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Figure 7

CV1 monomer-coated human T cells are targeted for phagocytosis by NSG but not human macrophages, and engineered murine T cells are only depleted in C57BL/6 mice if the CD47 decoy is fused to an active Fc region.

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CV1 monomer-coated human T cells are targeted for phagocytosis by NSG bu...
(A) Schematic of lentiviral vectors encoding SiRPα monomer under 6xNFAT and of a retroviral vector encoding the A97L-TCR. (B) Inducible SiRPα monomer expression by A97L–T cells as detected by EGFP expression upon coculture with target tumor cells (n = 3). (C) A375 tumor control curves following ACT with A97L–T cells coengineered to express inSiRPα- or CV1 monomers under 6xNFAT (n ≥6; data are representative of 2 independent studies). (D) Evaluation of phagocytosis of A97L–T cells coengineered to express inSiRPα or CV1 monomers by BMDMs (n = 6). (E) Evaluation of phagocytosis of A97L–T cells coated with inSiRPα- or CV1 monomers by MDMs (n = 3). (F) Evaluation of phagocytosis of T cells coated with secreted CV1-nullFc by MDMs in vitro (n = 7). (G) Schematic of retroviral constructs encoding A4-Fc and A4 decoys. (H) Expression of A4-Fc and A4 monomer decoys in transduced mouse OT-I T cells, detected by EGFP (data are representative of 3 or more donors). (I) Flow cytometric detection of A4-Fc and A4 monomer binding on OT-I T cells by staining with anti-Fc Ab and anti–mouse CD47 Abs (data are representative of 3 donors). (J) Schematic of ACT against subcutaneous B16-OVA tumors and ex vivo analysis. (K) Frequency and number of mouse CD45.1+ cells in harvested tissues 6 days after ACT (n = 6; data are representative of 2 independent studies). Statistical analysis was done by 1-way ANOVA (B and K), 2-way ANOVA (C), or unpaired 2-tailed t test (D–F) with correction for multiple comparisons by post hoc Tukey’s test (B and K) or post hoc Šidák’s test (C). *P < 0.05; **P < 0.01; ***P < 0.001; ****P< 0.0001.