LIGANDS FOR THE ERAB-2: PURIFICATION, CLONING &BIO PROP

Project: Research project

Project Details

Description

The erbB-2 oncogene encodes a 185 kDa transmembrane protein which
has been suggested to be a growth factor receptor due to its homology
with the EGF receptor (EGFR). We have identified two ligands of the
erbB-2 oncoprotein, gp30 and p75. gp30 binds to EGFR and pl85erbB2, and
the p75 binds exclusively to pl85erbB-2. We have identified and purified the gp30 growth factor that is
secreted by MDA-231 human breast cancer cells. This growth factor binds
to the EGFR and induces NRK cell colony formation (1). We have reported
that gp30 is also a ligand for the pl85erbB-2 protein and induces growth
inhibition of cells with erbB-2 amplification and over expression. At
very low concentrations however, gp30 stimulated the proliferation of
SKBr-3 overexpressing erbB-2 breast cancer cells. We have demonstrated
that gp30 interferes directly with the binding of a specific erbB-2
antibody (4D5) which has been shown to induce growth inhibition of
overexpressing erbB-2 cells. We can utilize this specific competition
binding assay to detect other possible ligands for erbB-2, including the
p75 described in this proposal. Preliminary studies, with the
competitive erbB-2 receptor binding assay, suggest that SKBr-3 cells
secrete a putative erbB-2 ligand. Further-more, purification of this
erbB-2 ligand from SKBr-3 cells, using erbB-2 extracellular domain
coupled to an affinity column yielded a p75 ligand which competes with
4D5 of binding to erbB-2. We now intend to continue the biological, biochemical and molecular
characterization of gp30 and p75 by several approaches which include
protein sequencing to generate polyclonal and monoclonal antibodies, as
well as cDNA cloning and gene regulation. We also will compare their
sequence homology and biological activity. The prognostic, diagnostic and therapeutic implications of this
proposed work for breast, ovarian and possibly gastric cancers are of
substantial significance and will be explored systematically in this
project.
StatusFinished
Effective start/end date7/17/914/30/96

Funding

  • National Institutes of Health
  • National Institutes of Health: $225,067.00
  • National Institutes of Health
  • National Institutes of Health: $162,380.00

ASJC

  • Medicine(all)

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