Fig. 6: T cells expressing the CD19/CD22 Bi-ChTCR have excellent sensitivity for both antigens and have potent antitumor activity.

a,b, Left: western blot of LAT pTyr²²⁰, actin and LAT in T cells expressing the indicated receptors after stimulation (Stim.) with Nalm-6 CD19⁺CD22^KO (a) or Nalm-6 CD19^KOCD22⁺ (b) cells. Right, heat map of the mean band intensity of LAT pTyr²²⁰ normalized to LAT (n = 3 independent experiments). c, Flow plots depicting CD19 and CD22 expression levels on Nalm-6 cells after gene KO. d, Left: flow histograms of CTV dilution measured after coculture with Nalm-6 CD19^KOCD22^KO (DKO, control) or Nalm-6 CD19^lowCD22^KO cells. Right: frequency of divided cells (CTV^low cells) after coculture with Nalm-6 CD19^lowCD22^KO cells (mean ± s.d.) (n = 3 independent donors). P values were calculated using a two-way ANOVA. e,f, IL-2 (e) and IFNγ (f) concentrations in supernatants after coculture with Nalm-6 CD19^lowCD22^KO cells (mean ± s.d.) (n = 3 independent donors). P values were calculated using a two-way ANOVA. g, Left: flow histograms of CTV dilution after coculture with Nalm-6 DKO cells or Nalm-6 CD19^KOCD22^low cells. Right: frequency of divided cells (CTV^low cells) after coculture with Nalm-6 CD19^KOCD22^low cells (mean ± s.d.) (n = 3 independent donors). P values were calculated using a two-way ANOVA. h,i, IL-2 (h) and IFNγ (i) concentrations in supernatants after coculture with Nalm-6 CD19^KOCD22^low cells (mean ± s.d.) (n = 3 independent donors). P values were calculated using a two-way ANOVA. j, Schematic of NSG mice engrafted with a mixture of CD19⁺CD22⁺ and CD19^lowCD22^low Nalm-6 GFP-ffLuc⁺ cells and treated with 2 × 10⁶ T cells expressing CD19 and CD22 BBζ CARs (1:1 ratio) or the CD19/CD22 Bi-ChTCR. Mouse icon created with BioRender.com. k, Representative bioluminescence images of Nalm-6 GFP-ffLuc⁺ tumor-bearing mice. l, Tumor burden (mean ± s.d. radiance (photons per s per cm² per sr)) of tumor-bearing mice (n = 5 mice in the control group and 7 mice per treatment group). m, Kaplan–Meier survival of Nalm-6 GFP-ffLuc⁺ tumor-bearing NSG mice. P value was calculated using a log-rank test. n,o, Frequencies of CD4⁺ (n) and CD8⁺ (o) T cells expressing the CARs or Bi-ChTCR in the blood at the indicated time points (n = 7 mice per treatment group, % of live⁺lymphocyte⁺).

Source data