Fig. 5

Toxin is the determinant causing tumor inhibition by E8-PE38. a SDS-PAGE analysis of purified E8 (16 kDa), E8-PE38 (60 kDa) and E8-PE38 mut (60 kDa). b Cell viability assay for MKN45 and TMK1 cells treated with E8, E8-PE38 and E8-PE38 mut (n = 3, ****p < 0.0001 compared with E8 or E8-PE38 mut, two-way ANNOVA). E8 nanobody alone and E8-PE38 mut did not show cytotoxic effect on cell proliferation. c MKN45 tumor growth curves from mice treated with PBS, E8-PE38 (0.6 mg/kg), E8-PE38 mut (0.6 mg/kg) or equal molar E8 nanobody (n = 5-6, ****p < 0.001 as compared with all three control groups, two-way ANOVA). E8 alone or E8-PE38 mut did not have any tumor inhibitory effect. E8-PE38 significantly suppressed tumor growth. d Tumor weight from treated mice at the end of treatment in b (n = 5-6, ***p < 0.001, one-way ANOVA). e Body weight during the treatment from four groups in c. f and g Ki67 immunostaining in tumor tissues collected from c. Tumor tissues treated with E8-PE38 markedly inhibited the Ki67 expression and no obvious change was detection in three control groups (n = 5-6, *p < 0.05, ***p < 0.001, one-way ANOVA). Scale bars: 200 μm. h Tumor growth curves from TMK1 tumors treated with PBS, E8-PE38 (0.4 mg/kg), E8-PE38 mut (0.4 mg/kg) or equal molar E8 nanobody (n = 5, ****p < 0.0001 as compared with all three control groups, two-way ANOVA). i Survival curves of treated mice from h (n = 5, **p < 0.01, Log-rank (Mantel-Cox) test). j Tumor growth curves from MNK45 tumor bearing mice received the treatment with PBS, 5-FU (25 mg/kg), E8-PE38 (0.4 mg/kg), and combination therapy (5-FU + E8-PE38) (n = 6, ***p < 0.001, ****p < 0.0001, two-way ANOVA). k Survival curves of tumor-bearing mice treated in j (n=6, **p<0.01, ***p<0.001, Log-rank (Mantel-Cox) test)