Figure 3. SiRPα-Fc decoys are efficiently produced by gene-modified T cells.

(A) Schematic of retroviral constructs encoding inSiRPα-Fc, wtSiRPα-Fc, or CV1-Fc proteins with an EGFP reporter gene, and of a lentiviral construct encoding the A97L-TCR. (B) Strategy for T cell activation, dual virus transduction, and expansion. (C) Expression of SiRPα-Fc and A97L-TCR in cotransduced rested human CD8⁺ T cells as detected by EGFP and anti–human Vβ13.1 Ab, respectively (data are representative of 12 independent donors). (D) CD47-based ELISA detection of SiRPα-Fc secreted by engineered CD8⁺ T cells (n = 3). (E) Quantification of CD8⁺ T cell–secreted CV1-Fc by CD47-based ELISA after 24 hours of culture (n = 3). (F) Quantification of CV1-Fc accumulated in culture supernatants of engineered CD8⁺ T cells over time (representative results for 3 donors). (G) Binding of CD8⁺ T cell–secreted CV1-Fc on different CD47⁺ tumor cell lines (data are representative of 3 donors). (H) Frequency of effector and memory phenotypes of transduced and rested CD8⁺ T cells (n = 3) (T_E, effector; T_EM, effector memory; T_CM, central memory; T_N/SCM, naive/stem cell–like memory). (I) Expansion of engineered CD8⁺ T cells (n = 3). Statistical analysis was done by 1-way ANOVA (D and H), unpaired 2-tailed t test (E), or 2-way ANOVA (I) with correction for multiple comparisons by post hoc Tukey’s test on pooled donors (D, H, and I). ****P< 0.0001.