CD47 signaling CAR-T cells have reduced proliferative capacity in vitro
(A) Schematic representation of CD47 signaling (CD47.CD28z) or non-signaling (ΔCD47) lentiviral CAR cassette where both constructs contain an anti-CD47 scFv, CD28 hinge, and CD28 TM domain. Additionally, the CD47.CD28z CAR contains CD28 and CD3ζ signaling domains. (B) Retention of endogenous CD47 expression on CD47.CD28z and ΔCD47 CAR-T cells was assessed by flow cytometry 7 days following transduction. (C and D) Expansion (C) and viability (D) of CAR-T cells prepared from three independent healthy donors was tracked for at least 7 days following transduction. T cells (no CAR) and TAG-72.4-1BB CAR-T cells were maintained in parallel. (E) CD47 expression on target cells, including OVCAR-3, MESOV, HLF, and HUF cells, was confirmed by flow cytometry (gray) compared to unstained control (red) where values represent average CD47 expression (percent viable) ± SD (n = 5).