Panel A shows a schematic of non-competitive CAIleR T cell activation where two CAIleR T cells engage Bet v1 antigen simultaneously through distinct binding sites. Panel B shows a bar graph of mCherry positive percent expression for WT and CAIleR 5, 8, 10, and 11 that reaches approximately 40 to 60 percent while WT remains near 0. Panel C shows a bar graph of CD25 positive CD71 positive percent expression across CAIleR combinations under no stimulation, Bet v1 alone, Bet v1 plus K562 CD64, and Bet v1 plus K562 CD64 plus mAb 10 and Bet v1 conditions. With Bet v1 alone, CAlleR 5 combined with CAlleR 10 as well as CAlleR 8 combined with CAlleR 10 reaches approximately 30 to 50 percent activation while combinations remain negative. With Bet v1 plus K562 CD64 plus mAb 10, all conditions where CAIleR 5 or CAlleR 8 where present approximately at least 50 percent activation.
CAlleR T cell activation by soluble Bet v1 through noncompetitive scFvs. (A) Schematic representation of two different CAlleR T cells interacting through noncompetitive binding of the scFv to Bet v1. (B) Transduction efficiency of anti-Bet v1 CAlleR T cells assessed by mCherry reporter expression by flow cytometry. (C) Cumulative data showing percentages of CD25+CD71+ CAlleR T cells (gated on mCherry+) after 48-h coculture with other CAlleR T cells (1:1 ratio) in presence or absence of K562 CD64 (1:5 ratio), Bet v1 (0.5 μg/ml) and mAb 10 (1 μg/ml). Data represent three biological independent experiments and are shown as the mean ± SEM in B and C. Exact P values were determined by two-way ANOVA with Tukey’s test in C. ****P < 0.0001.