Cellular Growth Response to Epidermal Growth Factor in Colon Carcinoma Cells with an Amplified Epidermal Growth Factor Receptor Derived from a Familial Adenomatous Polyposis Patient

M. E. Gross, M. A. Zorbas, V. J. Danels, M. G. Brattain, L. C. Yeoman

Research output: Contribution to journalArticle

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Abstract

The receptor binding and cellular growth responses to exogenousepidermal growth factor (EGF) were studied using the DiKi cell lineestablished from a familial adenomatous polyposis patient. The numberof cell membrane EGF receptors on DiKi cells, as measured by competitive radioligand binding assays and Scatchard analysis of 125I-EGF binding isotherms, was calculated to be 4.8 x 106 receptors/cell. An acidprcwash step performed prior to ligand binding assays did not revealadditional receptor numbers. A single. Ion-affinity receptor populationwas identified by Scatchard analysis, with an apparent A, of 4.6 UM.Thisresult was confirmed by radioligand binding studies performed in thepresence and absence of the receptor-antagonist monoclonal antibody528 IgG that binds predominantly to the low-affinity form of the EGFreceptor. DiFi cells at 50-60% confluence, when exposed to 50 nMexogenous FGF, exhibited a rapid but partial (30%) reduction in theircell membrane-associated receptor, characteristic of sequestration. Exposure of DiFi cells to 50 nM EGF for longer periods of time (4 h) did not result in any further reduction in EGK-receptor number. The cellulargrowIh response of DiFi cells to exogenous EGF was studied in monolayercultures as well as in a soft agarose assay. Inhibition of soft agar colonyformation was observed at exogenous EGF concentrations greater than1.7 nM. and inhibition of monolayer growth occurred at EGF concentrations greater than I nm. In immune complex kinase assays, the DiFireceptor showed similar specific activity to that from the well-characterized A431 cell line. Additionally, phosphorylation of the receptor ontyrosine was qualitatively similar to that of A43I cells, further suggestingthat the DiFi receptors identified by EGF-binding studies were Biologically functional.

Original languageEnglish (US)
Pages (from-to)1452-1459
Number of pages8
JournalCancer Research
Volume51
Issue number5
StatePublished - Mar 1991

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Adenomatous Polyposis Coli
Epidermal Growth Factor Receptor
Epidermal Growth Factor
Intercellular Signaling Peptides and Proteins
Colon
Carcinoma
Growth
Radioligand Assay
Competitive Binding
Growth Factor Receptors
Antigen-Antibody Complex
Sepharose
Agar
Phosphotransferases
Immunoglobulin G
Phosphorylation
Cell Membrane
Ions
Ligands
Cell Line

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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Cellular Growth Response to Epidermal Growth Factor in Colon Carcinoma Cells with an Amplified Epidermal Growth Factor Receptor Derived from a Familial Adenomatous Polyposis Patient. / Gross, M. E.; Zorbas, M. A.; Danels, V. J.; Brattain, M. G.; Yeoman, L. C.

In: Cancer Research, Vol. 51, No. 5, 03.1991, p. 1452-1459.

Research output: Contribution to journalArticle

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abstract = "The receptor binding and cellular growth responses to exogenousepidermal growth factor (EGF) were studied using the DiKi cell lineestablished from a familial adenomatous polyposis patient. The numberof cell membrane EGF receptors on DiKi cells, as measured by competitive radioligand binding assays and Scatchard analysis of 125I-EGF binding isotherms, was calculated to be 4.8 x 106 receptors/cell. An acidprcwash step performed prior to ligand binding assays did not revealadditional receptor numbers. A single. Ion-affinity receptor populationwas identified by Scatchard analysis, with an apparent A, of 4.6 UM.Thisresult was confirmed by radioligand binding studies performed in thepresence and absence of the receptor-antagonist monoclonal antibody528 IgG that binds predominantly to the low-affinity form of the EGFreceptor. DiFi cells at 50-60{\%} confluence, when exposed to 50 nMexogenous FGF, exhibited a rapid but partial (30{\%}) reduction in theircell membrane-associated receptor, characteristic of sequestration. Exposure of DiFi cells to 50 nM EGF for longer periods of time (4 h) did not result in any further reduction in EGK-receptor number. The cellulargrowIh response of DiFi cells to exogenous EGF was studied in monolayercultures as well as in a soft agarose assay. Inhibition of soft agar colonyformation was observed at exogenous EGF concentrations greater than1.7 nM. and inhibition of monolayer growth occurred at EGF concentrations greater than I nm. In immune complex kinase assays, the DiFireceptor showed similar specific activity to that from the well-characterized A431 cell line. Additionally, phosphorylation of the receptor ontyrosine was qualitatively similar to that of A43I cells, further suggestingthat the DiFi receptors identified by EGF-binding studies were Biologically functional.",
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