Fig 6. HS20 inhibited HGF-induced tumor spheroid formation.

(A) Representative photographs of Hep3B and Huh-7 spheroid. Hep3B and Huh-7 cells (5000 cell/well in 6-well low attachment plate) were treated with 50ng/ml HGF alone or co-cultured with 50 μg/mL HS20 for 20 days. Human IgG was used as negative control. Scale bar indicates 50 μm. (B) The spheroid volume in each group described in (A). Each dot represents a spheroid. P**<0.01 and P***<0.001. (C) Western blot to detect the expression of total c-Met and phosphorylated c-Met in spheroid. Hep3B cells and Huh-7 cells (5000 cell/well in a 6-well low attachment plate) were co-cultured with 50ng/ml HGF and 50 μg/mL HS20 for 20 days in a low attachment plate. Human IgG was used as a negative control. (D) BALB/c nu/nu mice were subcutaneously inoculated with 10x10⁶ Hep3B cells. When tumors reached an average volume of 100 mm³, mice were grouped and intravenously administered 25mg/kg HS20 twice a week. Values are mean ± SE from different mice. P*<0.05. n = 4 for each group. PBS was used as vehicle. Arrows indicate antibody injection. (E) BALB/c nu/nu mice were subcutaneously inoculated with 5x10⁶ HepG2 cells. When tumors reached an average volume of 100 mm³, mice were grouped and intravenously administered 20mg/kg HS20 twice a week. Values are mean ± SE from different mice. P*<0.05. n = 5 for each group. PBS was used as vehicle. Arrows indicate antibody injection.