Antagonism between Epidermal Growth Factor and Phorbol Ester Tumor Promoters in Human Breast Cancer Cells

Osborne, C. Kent; Hamilton, Barbara; Nover, Marianne; Ziegler, Jeanette

doi:10.1172/jci110144

Cited by 55 publications

(21 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Tumor promoters inhibit EGF-induced mitogenesis in human fibroblasts (34), rat hepatoma cells (29), and human hepatoma cells (37) and block EGF-induced cellular adhesion in PC12 phaeochromocytoma cells (8) and EGF-induced production of diacylglycerol in A431 cells (43). We confirm that TPA can inhibit EGF-induced mitogenesis in human fibroblasts and present a possible mechanism.…”

supporting

confidence: 73%

Effects of epidermal growth factor and 12-O-tetradecanoylphorbol-13-acetate on metabolism of the epidermal growth factor receptor in normal human fibroblasts.

Decker¹

1984

Mol. Cell. Biol.

View full text Add to dashboard Cite

The biosynthesis, phosphorylation, and degradation of the epidermal growth factor (EGF) receptor were examined in normal human fibroblasts. The receptor was initially synthesized as an Mr = 160,000 immature form which matured to an Mr = 170,000 form in a monensin-sensitive manner. Tunicamycin treatment led to the accumulation of an Mr = 130,000 protein. The receptor was phosphorylated on serine and threonine residues in normally growing and quiescent cells, and treatment with EGF or the tumor promoter 12-0-tetradecanoylphorbol-13-acetate (TPA) resulted in a two-to threefold increase in receptor-bound phosphate. EGF increased the amount of phosphoserine and phosphothreonine and caused the appearance of a minor amount of phosphotyrosine. TPA increased the levels of phosphoserine and phosphothreonine exclusively. Prior treatment with TPA inhibited the EGF-dependent appearance of phosphotyrosine in the receptor. Analysis of tryptic phosphopeptides revealed that six of the seven major peptides were common to the receptor from cells treated with EGF or TPA. EGF strongly stimulated [3H]thymidine incorporation in confluent cells, increased final saturation density three to fourfold, and increased whole-cell levels of phosphotyrosine about threefold. Treatment of cells with TPA before addition of EGF inhibited all three of these EGF-dependent responses. EGF also decreased the receptor half-life from 15 h to 1 h, but this was not inhibited by TPA. TPA alone had no detectable effect on the receptor half-life.Epidermal growth factor (EGF) binds to a specific plasma membrane receptor protein which functions in mediating EGF-induced mitogenesis. Responses to EGF binding to its receptor include activation of a receptor-associated, tyrosine-specific protein kinase (9, 10), increased phosphorylation of the receptor protein (15, 26) and of other unidentified proteins (11, 21), increased nutrient transport (1, 24), transient influx of Ca2+ and Na2+ ions (38, 45), changes in membrane phospholipid metabolism (40, 43), and ultimately cell division. The primary event is probably an activation of the receptor protein kinase, which could induce the other phenomena through phosphorylation of appropriate protein substrates. EGF-dependent stimulation of the EGF receptor-associated protein kinase has been demonstrated in vitro in membrane preparations from many different cell types (4, 13, 18) and in vivo for the A431 epidermoid carcinoma cell line (26) and 3T3 cells (11). This unusual tyrosine-specific protein kinase activity has also been found associated with the receptors for platelet-derived growth factor (PDGF) (17), insulin (30), and somatomedin C (28) and with the transforming gene products of a number of oncogenic viruses (2). In vivo phosphorylation of the EGF receptor from A431 cells is of a complex nature; phosphate has been found on serine, threonine, and tyrosine residues (12, 15}. The insulin receptor has also been shown to contain these three phosphoamino acids (31). The functional significance of receptor phosphorylation is...

show abstract

supporting

confidence: 73%

Effects of epidermal growth factor and 12-O-tetradecanoylphorbol-13-acetate on metabolism of the epidermal growth factor receptor in normal human fibroblasts.

Decker¹

1984

Mol. Cell. Biol.

View full text Add to dashboard Cite

show abstract

“…The ZR-75-1 cell line was obtained from C. Kent Osborne (The University of Texas Health Science Center at San Antonio) (11,12) in January 1992 and designated ZR-75-1-Tx to distinguish it from the ZR-75-1 cell line that we obtained from the American Type Culture Collection (Rockville, MD) in September 1988 (3, 7) and which we now designate ZR-75-1-Ro.…”

Section: Methodsmentioning

confidence: 99%

“…These findings led to the investigation of a third subline of ZR-75-1 cells (11,12), henceforth called ZR-75-1-Tx cells. We now report that in these cells IL-6 disrupts both cell-cell and cell-substratum adhesion: cells round up and are unable to translocate directionally; however, they continue to proliferate at an undiminished rate.…”

mentioning

confidence: 99%

Cell-adhesion-disrupting action of interleukin 6 in human ductal breast carcinoma cells.

Tamm

Kikuchi

Cardinale

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Recombinant baculovirus-derived interleukin 6 (IL-6) disrupts the attachment of human ductal breast carcinoma subline ZR-75-1-Tx cells to neighbors and the substratum in culture without inhibiting the proliferation ofthe cells. The nonadherent cells lack pseudopodia and do not translocate directionally. These findings stand in contrast to the earlier observations in the Ro subline of ZR-75-1 cells in which 11-6 induces cell-cell separation without detachment of the cells from the substratum, with the cells displaying pseudopodia, increased motility, and decreased proliferation. The IL-6-induced ZR-75-1-Tx cell detachment and rounding are reversible by incubation of the treated cells in IL-6-free medium for several days. The distinctive changes induced by 11-6 in ZR-75-1-Ro cells are similarly reversible. Either acidic fibroblast growth factor or phorbol 12-myrisate 13-acetate can replace serum as a cofactor in IL-6-induced ZR-75-1-Tx cell detachment. Our findings indicate that genetic changes can occur in breast carcinoma cells that through cytokine action markedly affect cell structure, adhesiveness, and motility.Infiltrating ductal breast carcinoma is characterized by disordered cell adherence (1). Cell distribution in cytological preparations varies from minimally dyshesive multilayered sheets to small piled groups and isolated cells. Both the degree of dyshesion and extent of nuclear alteration depend on the differentiation of the carcinoma (1). Poorly differentiated tumors are characterized by the presence of many dyshesive groups and isolated cells. The other cardinal feature of ductal breast carcinoma is that once obvious neoplasia develops genetic defects multiply and accumulate (2).Earlier studies identified interleukin 6 (IL-6) as a cytokine that can act on epithelioid breast carcinoma cells in culture to induce conversion to a fibroblastoid phenotype accompanied by cell-cell separation and increased cell motility with cells in preformed colonies moving apart (3-5). These studies were carried out with T-47D (6) and ZR-75-1 (7) (henceforth called ZR-75-1-Ro subline) cells, in which IL-6 suppressed cell proliferation. An examination of IL-6 action on the clone B subline of ZR-75-1 cells (8, 9) revealed a modified response: IL-6 did not cause cell-cell separation in preformed colonies or inhibit the proliferation of clone B cells, but it did prolong the interval between mitosis and readherence of daughter cells, presumably because of somewhat decreased adhesivity of the IL-6-treated cells (10).These findings led to the investigation of a third subline of ZR-75-1 cells (11, 12), henceforth called ZR-75-1-Tx cells. We now report that in these cells IL-6 disrupts both cell-cell and cell-substratum adhesion: cells round up and are unable to translocate directionally; however, they continue to proliferate at an undiminished rate. MATERIALS AND METHODSCells. The ZR-75-1 cell line was obtained from C. Kent Osborne (The University of Texas Health Science Center at San Antonio) (11,12) in January 1992 and ...

show abstract

“…Physiological regulators of cell growth and differentiation, such as peptide growth factors (2, 4, 10, 11), hematopoietic regulatory proteins (5, 12-14), tumor necrosis factor types a and ,3 (8, 9), interferons (15) . In addition, PMA also alters the morphology of MCF-7 cells and PMA-treated cells exhibit the morphological characteristics of secretory cells (27,28 (vol/vol) heat-inactivated fetal bovine serum, L-glutamine, penicillin (60.6 Jug/ml), and streptomycin (100 ,ug/ml). …”

mentioning

confidence: 99%

“…Biologically active phorbol esters, such as phorbol 12-myristate 13-acetate (PMA), are potent tumor-promoters in vivo, and they elicit and modulate a wide variety of biological and biochemical responses in vivo as well as in vitro (24)(25)(26). It has been known for some time that PMA inhibits the growth of human breast adenocarcinoma cell line MCF-7 (27). In addition, PMA also alters the morphology of MCF-7 cells and PMA-treated cells exhibit the morphological characteristics of secretory cells (27,28).…”

mentioning

confidence: 99%

Amphiregulin: a bifunctional growth-modulating glycoprotein produced by the phorbol 12-myristate 13-acetate-treated human breast adenocarcinoma cell line MCF-7.

Shoyab

McDonald

Bradley

et al. 1988

Proc. Natl. Acad. Sci. U.S.A.

315

233

View full text Add to dashboard Cite

A glycoprotein, termed amphiregulin (AR), inhibits growth of several human carcinoma cells in culture and stimulates proliferation of human fibroblasts and certain other tumor cells. It has been purified to apparent homogeneity from serum-free conditioned medium of MCF-7 human breast carcinoma cells that had been treated with phorbol 12-myristate 13-acetate. AR is, a single-chain extremely hydrophilic glycoprotein containing cysteines in disulfide linkage(s) that are essential for biological activity; it is stable between pH 2 and pH 12 and after heating for 30 min at 560C but unstable at 1OOC.The apparent molecular weights of AR and N-Glycanasetreated AR are 14,000 and 15,000, respectively, as assessed by gel chromatography, and 22,500 and %14,000, respectively, as determined by polyacrylamide gel electrophoresis. Treatment of AR with N-Glycanase, O-Glycanase, or neuraminidase does not affect its activity. The pI of AR is =7.8. The aminoterminal amino acid sequence of AR has been determined, and no significant sequence homology between AR and other proteins was found. The molecule thus appears to be a distinct growth regulatory protein.Cellular growth and differentiation appear to be initiated, promoted, maintained, and regulated by a multiplicity of stimulatory, inhibitory, and synergistic factors and hormones. The alteration and/or breakdown of the cellular homeostasis mechanism is a fundamental cause of growthrelated diseases including neoplasia (1-11). Physiological regulators of cell growth and differentiation, such as peptide growth factors (2, 4, 10, 11), hematopoietic regulatory proteins (5, 12-14), tumor necrosis factor types a and ,3 (8, 9), interferons (15) . In addition, PMA also alters the morphology of MCF-7 cells and PMA-treated cells exhibit the morphological characteristics of secretory cells (27,28 (vol/vol) heat-inactivated fetal bovine serum, L-glutamine, penicillin (60.6 Jug/ml), and streptomycin (100 ,ug/ml). Growth Modulatory Assay with ',I-Labeled DeoxyuridineIncorporation into DNA. The assays were performed in 96-well flat-bottomed plates (Falcon, catalog number 3072). Human epidermoid carcinoma of the vulva cells (A431) were used as test cells for growth inhibitory activity (GIA) and human forearm fibroblasts (SS) were used as indicator cells for growth stimulatory activity. A total of 3.5 x 104 cells in 50 tkl of DMEM, supplemented with 5% (vol/vol) heatinactivated fetal bovine serum, penicillin (60.6 pug/ml), streptomycin (100 tug/ml), and glutamine (test medium), was placed in all wells except peripheral wells. Three hours later, 50 jal of the test sample in test medium was added to each well; control wells received only 50 ,il of test medium. Three wells were used for each concentration of test sample. Plates were incubated at 370C for 2-3 days. After this, 100 t1d of solution of 125I-labeled deoxyuridine (Amersham) [4 Ci/mg; 0.5 Ci/ml (2 ,ul/ml in test medium); 1 Ci = 37 GBq] was added to each well and plates were incubated at 370C. After 4-6 hr, samples were processed as...

show abstract

Antagonism between Epidermal Growth Factor and Phorbol Ester Tumor Promoters in Human Breast Cancer Cells

Cited by 55 publications

References 34 publications

Effects of epidermal growth factor and 12-O-tetradecanoylphorbol-13-acetate on metabolism of the epidermal growth factor receptor in normal human fibroblasts.

Effects of epidermal growth factor and 12-O-tetradecanoylphorbol-13-acetate on metabolism of the epidermal growth factor receptor in normal human fibroblasts.

Cell-adhesion-disrupting action of interleukin 6 in human ductal breast carcinoma cells.

Amphiregulin: a bifunctional growth-modulating glycoprotein produced by the phorbol 12-myristate 13-acetate-treated human breast adenocarcinoma cell line MCF-7.

Contact Info

Product

Resources

About