2009 Scientific Report

Van Andel Research Institute | Scientific Report
Research Interests
Functioning as an antibody production core facility at VARI, our lab develops state-of-the-art services and technology platforms
for monoclonal antibody (mAb) production and characterization. Antibodies are primary tools of biomedical science. In basic
research, the characterization and analysis of almost any molecule involves the production of specific monoclonal or polyclonal
antibodies that react with it. Antibodies are also widely used in clinical diagnostic applications. Further, antibodies are making
rapid inroads into clinical treatment of a variety of diseases, driven by technological evolution from chimeric and humanized to
fully human antibodies.
Our technologies and services include antigen preparation and animal immunization; peptide design and coupling to protein
carriers; immunization with living or fixed cells; conventional antigen/adjuvant preparation; and immunizing a wide range of
antibody-producing models (including mice, rats, rabbits, and transgenic or knock-out mice). Our work also includes the
generation of hybridomas from spleen cells of immunized mice and rats; hybridoma expansion and subcloning; cryopreservation
of hybridomas; mAb isotyping; ELISA screening of hybridoma supernatants; mAb characterization by immunoprecipitation,
immunohistochemistry, immunofluorescence staining, western blot, FACS, and in vitro bioassays; conjugation of mAbs to
enzymes, biotin/streptavidin, or fluorescent reporters; and development of detection kits such as sandwich ELISA. We contract
our services to biotechnology companies, producing and purifying mAbs for their research and for diagnostic kit development.
We have also taken part over the past year in the following research projects.
• A single neutralizing mAb against the HGF/SF alpha domain. Hepatocyte growth factor/scatter factor (HGF/
SF) is a multifunctional heterodimeric polypeptide produced by mesenchymal cells; it is an effector of cells
expressing the Met tyrosine kinase receptor. We previously generated a cocktail of three or four neutralizing
mAbs against HGF/SF that significantly inhibited the HGF-Met signaling pathway in Met-expressing
cells. In a glioblastoma multiforme xenograft model, our cocktail showed potent inhibition of tumor growth.
Amgen and others have reported a single anti-HGF/SF b-subunit mAb that is able to inhibit biological activities
of HGF/SF; it is in early clinical trials. We hypothesized that two mAbs that react with different subunits
(a and b) of HGF/SF in combination would have stronger anti-tumor activity than any single antibody. Using
a unique immunization protocol, we have generated a mAb against the HGF/SF a subunit (designated HGF8)
that has neutralizing activity. Our current results show that HGF8 is able to block HGF/SF-induced scattering
of MDCK cells, and in collaboration with the Vande Woude lab, we have shown that HGF8 also significantly
inhibits the Met-HGF/SF signaling pathway in vitro using uPA and cell proliferation assays. The in vivo antitumor
activity of HGF8 is now under investigation in a brain tumor xenograft model using HGF/SF transgenic
mice established by the Vande Woude lab.
• Development of highly specific anti-Met mouse mAbs with potential application for clinical immunohistochemical
diagnosis. In collaboration with Beatrice Knudsen’s lab at the Fred Hutchinson Cancer Research Center,
we have developed a monoclonal antibody, designated MET4, with the goal of accurately and reproducibly
measuring MET in formalin-fixed paraffin-embedded (FFPE) tissues. MET4 was selected as the best probe
from a pool of MET-avid monoclonal antibodies, based on its specific staining pattern in FFPE preparations of
normal human prostate tissues. The reliability of MET4 immunohistochemistry was assessed by comparing
MET4-IHC in FFPE cell pellets with immunoblotting analysis, which demonstrated a high avidity of MET4 for
formalin-treated MET. These properties encourage further development of MET4 as a multipurpose molecular
diagnostic reagent to help guide the selection of individual patients being considered for treatment with MET
antagonistic drugs.
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