Discovery of Fully Human Anti-MET Monoclonal Antibodies with Antitumor Activity against Colon Cancer Tumor Models In Vivo

Abstract

The receptor tyrosine kinase MET is a major component controlling the invasive growth program in embryonic development and in invasive malignancies. The discovery of therapeutic antibodies against MET has been difficult, and antibodies that compete with hepatocyte growth factor (HGF) act as agonists. By applying phage technology and cell-based panning strategies, we discovered two fully human antibodies against MET (R13 and R28), which synergistically inhibit HGF binding to MET and elicit antibody-dependent cellular cytotoxicity. Cell-based phosphorylation assays demonstrate that R13 and R28 abrogate HGF-induced activation of MET, AKT1, ERK1/2, and HGF-induced migration and proliferation. FACS experiments suggest that the inhibitory effect is mediated by "locking" MET receptor in a state with R13, which then increases avidity of R28 for the extracellular domain of MET, thus blocking HGF binding without activating the receptor. In vivo studies demonstrate that the combination of R13/28 significantly inhibited tumor growth in various colon tumor xenograft models. Inhibition of tumor growth was associated with induction of hypoxia. Global gene expression analysis shows that inhibition of HGF/MET pathway significantly upregulated the tumor suppressors KLF6, CEACAM1, and BMP2, the negative regulator of phosphatidylinositol-3-OH-kinase PIK3IP1, and significantly suppressed SCF and SERPINE2, both enhancers of proliferation and invasiveness. Moreover, in an experimental metastasis model, R13/28 increased survival by preventing the recurrence of otherwise lethal lung metastases. Taken together, these results underscore the utility of a dual-antibody approach for targeting MET and possibly other receptor tyrosine kinases. Our approach could be expanded to drug discovery efforts against other cell surface proteins.

Antitumor activity of an anti‐CD98 antibody

“…18 IGN-LNG-12 and -54 are proprietary patient-derived lung tumor xenograft lines that were established at Igenica Biotherapeutics. IGN-LNG-12 is a metastatic poorly differentiated (Grade 3) NSCLC.…”

Hepatocyte Growth Factor Enhances Alternative Splicing of the Krüppel-like Factor 6 (KLF6) Tumor Suppressor to Promote Growth through SRSF1

“…As noted, a recent study using global gene expression analysis to identify HGF-regulated targets found that HGF inhibition significantly upregulates the KLF6 tumor suppressor (43). This finding indicated that HGF might exert its growth-promoting activity, in part, by inhibiting the growth-suppressive KLF6.…”

New Horizons in Therapeutic Antibody Discovery: Opportunities and Challenges versus Small-Molecule Therapeutics

“…90 Examples where cell-based panning has been successfully used are the isolation by subtractive selection of scFvs specific to the activated conformation of integrin αIIbβ3 91 and OncoMed's use of a hybrid cell panning/recombinant protein strategy to isolate different anti-MET antibodies that synergistically inhibit HGF binding to MET. 92 The use of cells in selections potentially also allows selection of clones with specific phenotypic characteristics such as the ability to internalize, affect proliferation, and recognize novel cell type-specific (e.g., tumor) antigen forms. 90 …”