Neutralizing monoclonal antibodies to hepatocyte growth factor/scatter factor (HGF/SF) display antitumor activity in animal models

Abstract

The hepatocyte growth factor (HGF/SF) receptor, Met, regulates mitogenesis, motility, and morphogenesis in a cell type-dependent fashion. Activation of Met via autocrine, paracrine, or mutational mechanisms can lead to tumorigenesis and metastasis and numerous studies have linked inappropriate expression of this ligand-receptor pair to most types of human solid tumors. To prepare mAbs to human HGF/SF, mice were immunized with native and denatured preparations of the ligand. Recloned mAbs were tested in vitro for blocking activity against scattering and branching morphogenesis. Our results show that no single mAb was capable of neutralizing the in vitro activity of HGF/SF, and that the ligand possesses a minimum of three epitopes that must be blocked to prevent Met tyrosine kinase activation. In vivo, the neutralizing mAb combination inhibited s.c. growth in athymic nu/nu mice of tumors dependent on an autocrine Met-HGF/SF loop. Importantly, growth of human glioblastoma multiforme xenografts expressing Met and HGF/SF were markedly reduced in the presence of HGF/SF-neutralizing mAbs. These results suggest interrupting autocrine and/or paracrine Met-HGF/SF signaling in tumors dependent on this pathway is a possible intervention strategy.

Figure 1

Neutralization of HGF/SF-mediated MDCK cells scattering. (A) Control without any treatment. (B) Human HGF/SF. (C) Human HGF/SF plus mAbs A-1, -5, and -7. (D) Human HGF/SF plus mAbs 7-2, -3, and -4.

Figure 2

Immunoprecipitation of HGF/SF from S-114 cell culture supernatant by mAbs. Lane 1, positive control, purified human HGF/SF; lanes 2–6, mAbs A-1, -4, -5, -7, and -10; lanes 7–9, mAbs 7-2, -3, and -4; and lane 10, positive serum from mouse immunized with native HGF/SF. The molecular weight standards are shown on the Left.

Figure 3

(a) Inhibition of C127 tumor growth by neutralizing mAb to HGF/SF. C-127 tumor cells were injected s.c. into athymic nude mice in PBS on day 0. Anti-HGF/SF mAbs A-1, -5, and -7 or mAb 7-2, -3, and -4 were administered either s.c. intratumor or i.p. every day for 20 days. One group of animals did not receive Ab. The values are an average of the size of five tumors in mm³. (b) Inhibition of U-118 glioblastoma tumor growth by neutralizing mAb to HGF/SF. U-118 human glioblastoma tumor cells were injected s.c. into athymic nude mice. On day 1, anti-HGF/SF mAbs A-1, -5, and -7 or mAbs 7-2 and -3 were administered either s.c. intratumor or i.p. twice a week for 10 weeks. One group of animals did not receive Ab. The values are an average size of six to seven tumors in mm³. (c) Tumor regression experiment using U-118 GBM cells. GBM cells were s.c. injected to athymic nude mice. After 30 days, animals were divided to five groups with average tumor size about 100 mm³. mAbs A-1, -5, and -7 or 7-2 and -3 were administered either s.c. intratumor or i.p. every 2 days until 10 weeks. One group of mice did not receive Ab. The values are an average size of eight to nine tumors in mm³.