Fig. 3: Macrophage suppression of ImmTAC-redirected T cells in vitro is contact-dependent and does not rely on known individual checkpoint interactions.

a Schematic diagram of ImmTAC-redirected E:T + M2 cell-cell contact-dependent or no contact co-cultures using transwell inserts b MFI of CD69 expression by ImmTAC-redirected CD3⁺ CD8⁺ T cells (mean ± SEM, n = 4 biological replicates, Tukey’s multiple comparison’s test, exact two-sided p values are given). c, d Representative flow cytometry plots (c) and quantification of early and late apoptotic targets (d) in M2-T cell-cell contact and contact-free co-cultures (Annexin V⁺, 7AAD^+/-), (mean ± SEM, n = 4 biological replicates, Tukey’s multiple comparison’s test, exact two-sided p values are given). T cell checkpoint receptors blocked in ImmTAC-redirected cell-cell contact M2 co-cultures using the indicated neutralizing antibodies. CD69 expression fold-change relative to ImmTAC-redirected E:T co-culture with IgG isotype control (e), IFN-γ release fold-change relative to IgG (E:T) controls (f), quantification of early and late apoptotic targets (g) [n = 3 for anti-TIM-3, anti-TIGIT, and anti-PD1 + anti-LAG-3 + anti-CTLA-4, n = 4 for anti-LILRB1, n = 6 for anti-PD-1 and anti-PD-L1, n = 7 for anti-CTLA-4 and anti-LAG-3, n = 10 for anti-IgG for (e) and (f) and n = 11 for anti-IgG for (g) all replicates are biological replicates and data shown is mean ± SEM]. Dashed lines represent IgG Ctrl mean. Abbreviations: E = Effectors, T = Targets, M2 = M2 macrophages, MFI = Mean Fluorescence Intensity.