B7-H3/CD16 BiKE induced antitumor effect mediated by NK cells in vitro and in vivo. a Construct diagrams of B7-H3/CD16 BiKE, anti-B7-H3 scFv, and anti-CD16 scFv. b Purity of B7-H3/CD16 BiKE determined by size-exclusion chromatography. c Cytotoxicity of B7-H3/CD16 BiKE (red line), anti-B7-H3 scFv (blue line) and anti-CD16 scFv (black line) against different tumor cell lines by PBMC as effectors. d Comparison of ADCC induced by B7-H3/CD16 BiKE and anti-B7-H3 IgG 8H9. PBMCs were used as effectors and A549 cells were used as targets. Representative graphs are shown. For the experiments c–d, the p values of the difference between BiKE and control groups were analyzed using ANOVA. e–h Therapeutic effects of B7-H3/CD16 BiKE in A549 and NCI-H23 xenografted mice. Antibodies were administered (100 μg per dose) intravenously twice a week for a total of four doses. Human PBMCs (10 million per iv injection) were delivered intravenously twice. Tumor growth in A549-injected mice e or HCT 116-injected mice g was measured using digital caliper, and tumor area was calculated (n = 5). Mean values per treatment group are shown. Survival curves of A549-injected mice f or HCT 116-injected mice (h) are shown. The p values of treatment were analyzed using ANOVA. The p values of survival curves were analyzed using the log-rank (Mantel–Cox) test.