Gastroenterology

Gastroenterology

Volume 124, Issue 5, May 2003, Pages 1337-1347
Gastroenterology

Basic-alimentary tract
Epidermal growth factor receptor-related protein: a potential therapeutic agent for colorectal cancer

https://doi.org/10.1016/S0016-5085(03)00264-6Get rights and content

Abstract

Background & Aims:

Epidermal growth factor receptor is frequently implicated in epithelial cancers and is, therefore, being considered as a potential target for therapy. Recently, we reported the isolation and characterization of epidermal growth factor receptor–related protein, a negative regulator of epidermal growth factor receptor. To discern whether epidermal growth factor receptor-related protein could be an effective therapeutic agent for colorectal cancer, we generated epidermal growth factor receptor–related protein fusion protein and studied its effect on the growth of colon cancer cells in vivo and in vitro. We also studied whether epidermal growth factor receptor-related protein expression is altered in colorectal cancer.

Methods:

A 55-kilodalton epidermal growth factor receptor–related protein fusion protein with V5 and His tags was generated in a drosophila expression system and subsequently purified by a His antibody affinity column. Rabbit polyclonal antibodies against epidermal growth factor receptor-related protein were used to examine the expression of epidermal growth factor receptor–related protein.

Results:

Epidermal growth factor receptor-related protein expression was found to be high in benign human colonic epithelium but low in adenocarcinoma. Exposure of the colon cancer cell lines HCT-116 and Caco-2 to purified recombinant epidermal growth factor receptor–related protein caused a marked inhibition of proliferation, as well as attenuation of basal and ligand-induced stimulation of epidermal growth factor receptor phosphorylation. Epidermal growth factor receptor-related protein-induced inhibition of proliferation of colon cancer cells was prevented by epidermal growth factor receptor–related protein antibodies. Reduced epidermal growth factor receptor phosphorylation was partly due to sequestration of epidermal growth factor receptor ligands by epidermal growth factor receptor-related protein, resulting in the formation of inactive heterodimers with epidermal growth factor receptor. Intratumoral or subcutaneous (away from the tumor site) injections of purified epidermal growth factor receptor–related protein caused regression of palpable colon cancer xenograft tumors in some severely compromised immunodeficient mice and arrested tumor growth in others.

Conclusions:

We propose that epidermal growth factor receptor-related protein inhibits cellular growth by attenuating epidermal growth factor receptor signaling processes and is an effective therapeutic agent for colorectal cancer.

Section snippets

Cell lines

Colon cancer cell lines HCT-116 and Caco-2 and monkey kidney COS-7 cells, obtained from the American Type Culture Collection (Rockville, MD), were maintained in Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum (FBS), penicillin (10,000 U/mL), streptomycin (10,000 U/mL), and amphotericin (25 μL/mL) at 37°C in an atmosphere of 95% air and 5% CO2.

Cell proliferation

This was assessed by 3-(4,5-dimethylthialzol-2yl)-2,5-diphenyltetrazolium bromide assay according to the procedure

Results

The fact that the ORF of ERRP contains a signal peptide composed of 24 amino acids, together with our observation that ERRP is present in the conditioned media derived from drosophila S2 cells (Figure 1), suggests that ERRP is a secretory protein. To further test this possibility that ERRP is a secretary protein, we generated a construct expressing polyhistidine and Myc-tagged ERRP protein. This construct or the vector plasmid (control) was subsequently transfected into COS-7 cells, and the

Discussion

Accumulating evidence suggests that downstream blocking of the EGFR signaling pathway is an effective therapeutic approach for the treatment and prevention of many epithelial malignancies, particularly those whose growth is regulated by the EGF family of peptides. This may be the most relevant area for use of ERRP. There are several strategies under investigation to block binding of growth factor to receptor. Previously, Wagner et al.25showed that transfection into pancreatic cancer cells

References (30)

  • M.B. Sporn et al.

    Autocrine growth factor and cancer

    Nature

    (1985)
  • Z. Culig et al.

    Regulation of prostatic growth and function by peptide growth factors

    Prostate

    (1996)
  • K. Khasharyarsha et al.

    EGF-receptor in neoplasia

    Cancer Metastasis Rev

    (1993)
  • W.J. Gullick

    Prevalence of aberrant expression of the epidermal growth factor receptor in human cancers

    Br Med Bull

    (1991)
  • E. Malecka-Panas et al.

    Differential activation of total and EGF receptor tyrosine kinase in rectal mucosa in patients with adenomatous polyps, ulcerative colitis and colon cancer

    Hepatogastroenterology

    (1997)
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    Supported by grants to Dr. Majumdar from the Department of Veterans Affairs and the National Institute on Aging (AG 14343).

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