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. Author manuscript; available in PMC: 2023 Jan 23.
Published in final edited form as: Sci Transl Med. 2021 Dec 8;13(623):eabh1962. doi: 10.1126/scitranslmed.abh1962

Figure 5. CD38-CCR can signal through in-trans binding of the antigen expressed on accessory cells without off-tumor toxicity.

Figure 5.

(A) A schematic representation of the coculture cytotoxicity/proliferation assay is shown. Luciferase-positive BCMA+CD38− U266 cells were co-cultured with calcein-loaded 3T3-38+ cells and single- or double-targeting T cells were added in the co-culture. Lysis of U266 cells was determined by BLI and lysis of 3T3-CD38 cells by calcein release assay. (B) U266 cells and calcein-loaded 3T3-CD38 cells were cocultured at a 3:1 E:T ratio with mock, BCMA-ζ, BCMA-ζ+CD38Δ, BCMA-ζ+CD38-28, BCMA-28ζ, BCMA-28ζ+CD38-BB or BCMA-ζ+CD38-28BB CAR T cells and cell lysis was measured after 4 hours by BLI and a Calcein-AM release assay, respectively (n=4 per group). Statistical analysis was performed by a paired t-test. *p<0.05, **p<0.01. (C) Cell supernatants from (B) were harvested to measure cytokine secretion with a flow cytometry–based assay (n=4 per group). Statistical analysis was performed with paired t-tests. *p<0.05, **p<0.01, ***p<0.001. (D) Irradiated BCMA+CD38− U266 cells were co-cultured with irradiated 3T3-38+ cells. Mock, BCMA-ζ, BCMA-ζ+CD38Δ, BCMA-ζ+CD38-28, BCMA-28ζ, BCMA-28ζ+CD38-BB or BCMA-ζ+CD38-28BB CAR T cells were added in the co-culture and the expansion of CAR T cells was measured 7 days later. Statistical analysis was performed with one-way ANOVAs and subsequent multiple comparison, corrected by Turkey test. ****p<0.0001.