Extended Data Fig. 5: ICN1 is unable to induce T-ALL generation in C80G mutant mice deficient in pre-TCR signalling.
From: Pre-TCR-targeted immunotherapy for T cell acute lymphoblastic leukemia
(A) Schematic diagram of experimental design for mouse T-ALL generation. Sublethally irradiated NSG (2H-Kd+) mice were subjected to i.v. injection with bone marrow lin− c-kit+ HPCs isolated from wild type (wt) or CD3ε C80G mutant C57BL/6 (H2-Kb+) mice and transduced with a lentiviral vector encoding either ICN1 and GFP or GFP alone as control. (B) Representative flow cytometry analysis of transduction efficiencies of wt (upper) and mutant (bottom) HPCs in (A). Numbers indicate percentages of transduced (GFP+) cells. (C) Representative CD8 and CD4 expression of electronically-gated mock-transduced (gate I; ICN1−) and ICN1-transduced (gate II; ICN1+) cells, derived from donor wt (upper) or C80G mutant (bottom) bone marrow HPCs engrafting the bone marrow of NSG mice at 5 weeks post-transplant. Results are representative of one out of at least 9 mice per group from two independent experiments. (D) Mean ± s.e.m. percentages of myeloid-lineage (CD11b+ or Gr1.1+), B-lineage (B220+), T-lineage (double positive; CD4+ CD8− and/or CD8+ CD4−) and Lin− (CD11b−, Gr1.1−, B220−, CD4−, CD8−) cells derived from ICN1+ (n = 5) and ICN1− (n = 4) wt or C80G (n = 5, each) mutant mouse bone marrow HPCs engrafting the bone marrow of NSG mice in (C). (E) Image of representative spleens obtained from mice shown in (C, D). (F) Mean percentages ± s.e.m. of ICN1+ cells derived from wt or C80G mutant HPCs engrafting the indicated organs of NSG mice (9 mice per group) at sacrifice, analysed by two-tailed unpaired t test. **P = 0.0022 (PB), ***P = 0.0005 (BM), ***P = 0.0003 (spleen), ****P < 0.0001 (liver), and *P = 0.0106 (thymus). (G) Kaplan-Meier survival curves of transplanted NSG mice in (F) were compared using Log-rank Mantle-Cox test. ****P < 0.0001.
