Fig. 1: Pre-clinical validation of AUTO3. | Nature Medicine

Fig. 1: Pre-clinical validation of AUTO3.

From: CAR T cells with dual targeting of CD19 and CD22 in pediatric and young adult patients with relapsed or refractory B cell acute lymphoblastic leukemia: a phase 1 trial

Fig. 1: Pre-clinical validation of AUTO3.The alternative text for this image may have been generated using AI.

a, AUTO3 is a dual CD19 and CD22 CAR T cell encoded on a single bicistronic γ-retroviral vector under the same promoter. The two CARs are separated by a self-cleaving 2A peptide that allows for equal expression, as shown in the flow cytometry plot. NT, non-transduced. b, Cytotoxicity of T cells co-cultured for 72 h with NT (left), CD19+ (middle) or CD22+ (right) SupT1 targets at a 1:1 effector : target (E:T) ratio. Data were normalized to NT T cells (n = 4 biologically independent samples). c, Comparison of cytotoxicity between AUTO3 and FMC63 CAR against Raji tumor cell lines. (left) Cytotoxicity of AUTO3 and FMC63 CAR T cells co-cultured with Raji for 24 h at a 1:1 E:T ratio (n = 4 biologically independent samples). (right) Cytotoxicity against mixed targets of WT Raji and CD19KO Raji cells at a 1:1 E:T for 72 h (n = 3 biologically independent samples). b,c, Horizontal lines represent the median of data points from healthy individual donors. ****P < 0.0001, ***P = 0.0001, one-way analysis of variance (ANOVA) with Dunnett’s post hoc test for multiple comparisons. d, In vivo efficacy of AUTO3 in a Nalm6 NSG mouse model after treatment with 5 × 106 CAR T cells. BLI, bioluminescent imaging. (left) Total flux radiance of tumor engraftment kinetics 15 d after treatment with CAR T cells (n = 5 FMC63 CAR and AUTO3, n = 3 NT, biologically independent animals). ***P = 0.0006, *P = 0.0437, two-way ANOVA with Bonferroni correction between FMC63 and AUTO3 over all six time points. (right) Percentage of CAR T cells in the bone marrow 6 d after CAR treatment (n = 5 biologically independent animals). Horizontal lines represent the median. P = 0.260 (NS, not significant), unpaired t-test. e, In vivo efficacy of AUTO3 in a CD19KO Nalm6 NSG model after treatment with 5 × 106 CAR T cells. (left) Tumor engraftment kinetics over 15 d (n = 5 biologically independent animals). ****P < 0.0001, *P = 0.0304, two-way ANOVA with Bonferroni correction between FMC63 and AUTO3 over all six time points. (right) CAR T cells in the bone marrow at d 15 (n = 5 biologically independent animals). Horizontal lines represent the median. *P = 0.0216, unpaired t-test.

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