Anti-vascular endothelial growth factor receptor-1 antagonist antibody as a therapeutic agent for cancer.

PURPOSE: Vascular endothelial growth factor receptor-1 (VEGFR-1) plays important roles in promotion of tumor growth by mediating cellular functions in tumor vascular endothelium and cancer cells. Blockade of VEGFR-1 activation has been shown to inhibit pathologic angiogenesis and tumor growth, implicating VEGFR-1 as a potential therapeutic target for the treatment of cancer. We have thus developed a VEGFR-1 antagonist human monoclonal antibody designated as IMC-18F1 and evaluated its antitumor activity in preclinical experimental models to show the therapeutic potential of the antibody for cancer treatment in clinic. EXPERIMENTAL DESIGN: Human IgG transgenic mice were used for generation of anti-VEGFR-1 antibodies. Anti-VEGFR-1-specific blocking antibodies were identified using solid-phase binding and blocking assays. Inhibitory antitumor cell activity of IMC-18F1 was assessed in cell-based kinase and growth assays. Pharmacokinetic/pharmacodynamic studies were done to determine the association of antibody blood level with antitumor efficacy of the antibody in vivo. Antitumor efficacy of the anti-VEGFR-1 antibodies as monotherapy and in combination with cytotoxic agents was evaluated in human breast cancer xenograft models. RESULTS: A fully human neutralizing antibody, IMC-18F1, was shown to be a high-affinity (KD=54 pmol) inhibitor of VEGFR-1 ligand binding (VEGF-A, VEGF-B, and placental growth factor). IMC-18F1 inhibited ligand-induced intracellular activation of VEGFR-1 and mitogen-activated protein kinase signaling and prevented ligand-stimulated in vitro growth of breast cancer cells. In vivo, IMC-18F1 suppressed the growth of human breast tumor xenografts in association with reduced mitogen-activated protein kinase and Akt activation, reduced tumor cell proliferation, and increased tumor cell apoptosis. Pharmacokinetic/pharmacodynamic studies established a plasma elimination half-life of 5 days for IMC-18F1 and a steady-state trough plasma therapeutic threshold of 88 microg/mL. Importantly, inhibition of mouse and human VEGFR-1 with MF1 and IMC-18F1, respectively, enhanced the antitumor efficacy of cytotoxic agents commonly used to treat breast cancer. CONCLUSIONS: Based on preclinical validation studies, IMC-18F1 anti-VEGFR-1 has potential to provide clinical benefit to cancer patients.

Targeting the platelet-derived growth factor receptor alpha with a neutralizing human monoclonal antibody inhibits the growth of tumor xenografts: implications as a potential therapeutic target.

Platelet-derived growth factor receptor alpha (PDGFRalpha) is a type III receptor tyrosine kinase that is expressed on a variety of tumor types. A neutralizing monoclonal antibody to human PDGFRalpha, which did not cross-react with the beta form of the receptor, was generated. The fully human antibody, termed 3G3, has a Kd of 40 pmol/L and blocks both PDGF-AA and PDGF-BB ligands from binding to PDGFRalpha. In addition to blocking ligand-induced cell mitogenesis and receptor autophosphorylation, 3G3 inhibited phosphorylation of the downstream signaling molecules Akt and mitogen-activated protein kinase. This inhibition was seen in both transfected and tumor cell lines expressing PDGFRalpha. The in vivo antitumor activity of 3G3 was tested in human glioblastoma (U118) and leiomyosarcoma (SKLMS-1) xenograft tumor models in athymic nude mice. Antibody 3G3 significantly inhibited the growth of U118 (P=0.0004) and SKLMS-1 (P <0.0001) tumors relative to control. These data suggest that 3G3 may be useful for the treatment of tumors that express PDGFRalpha.

Increased production of a secreted glycoprotein in engineered CHO cells through amplification of a transcription factor.

Expression levels of reporter protein driven by Mouse Mammary Tumor Virus Promoter system were improved by expressing its specific transcription factor (glucocorticoid receptor) from a different expression vector. The vector that expresses glucocorticoid receptor (GR) also contained dihydrofolate reductase (dhfr) gene as a selection marker. In the presentstudy we amplified the glucocorticoid receptor gene (gr)along with the dhfr gene by adapting the cell lines to increasing concentrations of methotrexate, an antifolate analog. Stepwise increases in the volumetric titers of a secreted reporter glycoprotein, Secreted Alkaline Phosphatase (SEAP), were observed in recombinant Chinese hamster ovary (CHO) cellsgrowing in increased concentrations of methotrexate. Western andRT-PCR analysis showed that this increase in volumetric titers is associated with higher levels of GR expressed in CHO cellsgrowing in increased concentration of methotrexate. A stablytransfected cell line growing in 10(-6) M methotrexate wasgrown in suspension culture and induced with 10(-7) Mdexamethasone. The SEAP volumetric titers reached a peak of approximately 23 mug ml(-1) on the 5th day after induction.Inducing these cells with increasing concentrations of dexamethasone resulted in increased specific productivity. These high volumetric productivities were further increased in fed-batch bioreactors.