Extended Data Fig. 1: In vitro and in vivo activities of IL-2^wt or IL-2^nα.

(a–c) Binding kinetics of anti-human B7H3 antibody (hz20G5) fused with either IL-2^wt or IL-2^nα detected by bio-layer interferometry (BLI). IL-2^wt (A) and IL-2^nα (B) binding kinetics to human IL-2Rα, human IL-2Rβγ, mouse IL-2Rα and mouse IL-2Rβγ. Table summarizing binding affinity of each molecule (C). (d) Binding kinetics of antibody hz20G5 to human B7H3 and mouse B7H3, showing high affinity of hz20G5 to human B7H3 but no binding activity to mouse B7H3 antigen. (e) Dose response curves of pSTAT5% in CD4⁺CD25^-T cell and NK cells treated with serial diluted IL-2^wt or IL-2^nα (experiment performed in duplicates). (f) Individual MC38 tumor growth curves in each treatment group (related to Fig. 1c). (g–i) Three repeated experiments to confirm in vivo efficacy and safety in MC38 tumor-bearing mice. Average tumor volume changes, body weight changes, and individual tumor growth curves were shown for each experiment. High dose (10 mg/kg) of IL-2 agonists was used in one experiment (G), and the remaining experiments used 3mg/kg. n = 7 (G) or n = 6 (H-I) mice per group. (j–k) Evaluation of wet lung weight in wild-type C57BL/6 mice (J) or Rag1-/- C57BL/6 mice (K) after a single administration of hIgG, IL-2^wt or IL-2^nα at indicated dosage 5 days post treatment. n = 5 (J) or n = 8 (K) mice per group. Mean + s.e.m. is presented, and p-values are calculated using two-way ANOVA for tumor growth curves and one-way ANOVA for bar graphs.

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