Fig. 1

a Docking studies. The derived molecular model was created using ZDOCK. The models were created by PEPFOLD3 software, version 9.16, and figures were generated with a UCSF Chimera. Ligplot+ & Pocketquery generated representation of the interaction between VGB3 and VEGFR1D2. The polar and van der Waals interactions are indicated in green and orange, respectively. The Pocketquery results identify side chains of VGB3 residues that may be involved in important interactions (Polar and van der Waals interactions) at the complex interface, as listed in Table S3. b MST curve. The result of binding of VGB3 to VEGFR1D2 at a stoichiometry of 1:1. The assay was performed using a fixed concentration of fluorescently labeled VEGFR1D2 and the initial concentration of ligand was 0.25 mM. c The effect of VGB3 on VEGF-stimulated cell proliferation. The effect of VGB3 on cell proliferation was determined on endothelial cells as well as 4T1 tumor cells. The cells with (dark grey) and without VEGF (light grey) were treated with various concentration ranges (0–0.061 μM) of VGB3 and then an MTT assay was performed to measure cell viability at two time points (24 and 48 h). Data points are mean ± SEM, obtained by Prism 6. Unpaired two-tailed t-test (to compare the differences between No-VEGF and VEGF in each concentration) and one-way ANOVA with Tukey multiple comparison; n = 6, NS no significant. d ELISA-based displacement assay. Bars represent mean ± SEM, obtained by Prism 6; one-way ANOVA with Tukey multiple comparison. e VGB3 inhibits angiogenesis in vitro. VGB3 inhibits migration of wounded HUVEC monolayers compared with the control. VGB3 decoy assessment in HUVEC Cytodex 3D bead sprouting assay which followed by embedding into the collagen gel in the presence of VEGF (0.75 pM) as stimulator following exposure to increasing concentrations of VGB3. Analytical results of mean data are shown for each concentrations. Bars represent mean ± SEM, obtained by Prism 6. Unpaired two-tailed t-test and one-way ANOVA with Tukey multiple comparison, NS no significant. f VGB3 inhibits 4T1 metastatic breast cancer growth in vivo. Significant inhibition of tumor growth occurred in animals treated with VGB3 when compared with control. Data points are mean ± SEM, obtained by Prism 6 in two-way ANOVA statistical analysis; n = 6. The differences between VGB3 treatments and PBS controls are demonstrated; *P ≤ 0.05 (0.02 mg/kg of VGB3 to PBS control), ***P ≤ 0.001 (0.2 mg/kg of VGB3 to PBS control), no significant (0.1 mg/kg and 0.5 mg/kg of VGB3 to PBS control). The average body weight of each group was measured until the size of the tumor in the control animals reached the endpoint of the study (from day 13 till 31), then presented as mean ± SEM. The data indicated no significant (NS) differences between VGB3 treatments and PBS controls in two-way ANOVA statistical analysis. g Mechanistic basis for VGB3-mediated tumor inhibition in vivo. Data were analyzed by ImageJ, and visualized as columns bars using Prism 6. Results shown are representative or mean ± SEM (n = 6). The differences between VGB3 treatments (0.2 mg/kg) and PBS controls were assessed by unpaired two-tailed t-test