0020-71 IX/90 $3.00 + 0.00 Copyright 0 1990 Pergamon Press plc

Inr. J. Biochem.Vol. 22, No. 9, pp. 939-945, 1990 Printed in Great Britain. All rights reserved

MINIREVIEW EPIDERMAL

GROWTH S.

Department

FACTOR

BARKER

IN BREAST

CANCER

and G. P. VINSON

of Biochemistry, Medical College of St Bartholomew’s Hospital, Charterhouse London EClM 6BQ, England

Square,

(Received 24 January 1990)

INTRODUCTION It

has long been recognized that oestrogens play an important role in the development and progression of human breast cancer (Seibert and Lippman, 1982). In the normal breast it acts as a mitogen and also induces the progesterone receptor (PgR) which is responsible for the development and differentiation of the breast (Horwitz et al., 1985). More recently a great deal of evidence has arisen to suggest that the mitogenic effects of oestrogens are mediated in some part via peptide growth factors and their receptors (Lippman and Dickson, 1989). This review sets out to bring together the clinical and experimental evidence which has led to the realization that endocrine, (steroid hormone), control of growth and development is directly associated with autocrine- and paracrine-acting growth factors in the human breast. Oestrogen is an essential factor in the development of the normal breast (Laron et al., 1989). It acts via specific intracellular receptors (Gorski et al., 1986). Although there is still much to learn regarding the mode of action of the oestrogen receptor (ER) it is generally accepted that the “activated” hormonereceptor complex binds to specific oestrogen-response elements (EREs) of target cell DNA (Beato, 1989) thereby initiating the transcription of specific mRNAs such as that of the PgR (Toft and O’Malley, 1972; Savouret et al., 1989). ER can be detected in 60-80% of human breast cancers (McGuire et al., 1975) and the significance of the presence of ER and PgR in breast tumours has been emphasized by many authors because of its implications for the progression of the disease and its management (McGuire, 1980; Seibert and Lippmann 1982; Hubay et al., 1984). The presence of ER in breast tumours is used as a marker for predicting the likely response to endocrine (i.e. anti-oestrogen) therapy. Of patients with ER positive tumours 50% will show a beneficial response and 75% of those with ER + ve/PgR + ve tumours respond to therapy (Horwitz, 1987). The presence of ER has also been shown to reflect the grade of the tumour; the less differentiated more agressive tumours tend to be ER negative (Seibert and Lippman, 1982). EPIDERMAL

GROWTH

FACTOR

AND ITS RECEPTOR

Human epidermal growth factor (EGF) is a single chain polypeptide, comprising 53 amino acid residues

with a relative mol. wt of 6045 (Carpenter and Cohen, 1979). The notion that it may be important in the breast came from the detection of this peptide in human breast milk (Starkey and Orth, 1977). Subsequently EGF was shown to increase DNA replication and division in cells derived from human breast epithelium (Osborne et al., 1980; Imai et al., 1982). It also acts as a growth inhibitor of certain human carcinoma cells (Filmus et al., 1985). The EGF receptor (EGF-R), is a transmembrane glycoprotein (mol. wt 170,000) which possesses an intrinsic tyrosine kinase activity responsible for the transduction of signal to the cytoplasmic compartment (Carpenter, 1987; Schlessinger, 1988; Hsuan et al., 1989). The precise mechanism of signal transduction is not fully understood but binding of ligand is thought to lead to an association with phospholipase-C and the production of inositol 1,4,5 trisphosphate (IP,) (Schlessinger, 1988), while protein kinase-C negatively modulates EGF-R function (Cachet et al., 1984). At present the cellular substrates for the EGF-R tyrosine kinase are not known, although it is capable of phosphorylating a number of intracellular proteins in uitro, including PgR (GhoshDastidar et al., 1984). The EGF-R gene has recently been cloned and has considerable structural homology with other members of what is now regarded as a superfamily of membrane bound protein kinases which include a series of related cellular oncogene products (Hunter and Cooper, 1985; Yarden and Ullrich, 1988). Several other growth factors are present in breast tumours and are secreted by human breast cancer cells. These include transforming growth factor-alpha (TGF-alpha) (Derynck et al., 1987; Salamon et al., 1989) and beta (TGF-bet, 1 (Arteaga et al., 1988b; Roberts and Sporn, 1988) .md insulin-like growth factor I (IGF-I) (Lippman et al., 1986). TGF-alpha is of particular relevance as it acts through the EGF-R with an affinity similar to that of EGF itself (Todaro et al., 1980; Massague, 1983).

EGF AND EGF-R IN BREAST TISSUE

Since the original isolation of EGF from the sub-maxillary glands of the mouse (Cohen, 1962) and the purification of human EGF (Urogastrone) from urine (Gregory, 1975; Carpenter and Cohen, 939

S. BARKER and

940

1979) both EGF and TGF-alpha have been found in many human tissues by radioimmunoassay and immunolocalization. These include normal, benign and malignant breast tissue (Perroteau et al., 1986; Travers er al., 1988), and also breast fluid and milk (Connolly and Rose, 1988). However, the concentrations of these factors have not been found to correlate significantly with any clinical parameters of breast cancer. The EGF-R has also been identified in a wide variety of tissues (O’Keefe ef al., 1974) including all histological types of breast tissue (Barker et al., 1989; Moller et al., 1989). Assay procedures vary between laboratories but essentially they all estimate the specific binding of radioiodinated EGF within a particulate or membrane-enriched fraction after ultracentrifugation. Scatchard analysis of the data can yield either a linear regression line, indicating a single class of binding site, or a curvilinear plot which reflects the existence of two populations of receptor of high and low affinity for EGF. Cross-linking studies have led to the proposal that these correspond to a monomeric receptor and an activated homodimer, respectively (Yarden and Schlessinger, 1987; Hsuan et al., 1989) however, it is not yet clear how the incidence of these two receptor forms relates to other factors in breast cancer. In most studies workers have resorted to the use of a single point assay to overcome the problem of limited sample size (Nicholson et al., 1988b). By this method an estimate of the number of binding sites is obtained. A variety of concentrations of labelled ligand have been used, which has in turn led to differences in ranges of values obtained, and detection limits set. Nevertheless, a consistent pattern seems to have emerged with respect to EGF-R expression in human breast tissue.

EGF-R EXPRESSION

In any series of primary breast tumours analysed, EGF-R values range from undetectable to values between 50- and lOO-fold greater than the lowest detectable value. Those in the latter category represent only cu 5-20% of all EGFR positive tumours and probably only 225% result from gene amplification (Ro ef al., 1988; Lacroix et al., 1989). However, compared with tumours from tissues other than breast, the concentrations are generally relatively low (Neal et al., 1985; Ozawa et al., 1988; Bauknecht et al., 1989). Studies of non-malignant breast tissue (Pekonen et al., 1988; Barker et al., 1989) have indicated that such levels are not abnormal for breast tissue and of greater interest is the relationship between EGF-R expression and other prognostic markers in breast cancer. The most thoroughly characterized relationship is the inverse relationship between the presence of EGF-R and ER in primary breast carcinomas. It is rare for both receptor types to be detectable together in the same tumour, but where they are both present, their concentrations are inversely correlated (Perez et al., 1984; Sainsbury et al., 1985). In some studies (Battaglia et al., 1988a; Cappelletti et al., 1988; Delarue et al., 1988) but not all (Fitzpatrick et al.,

G.

P.

VINSON

1984; Wrba et al., 1988), PgR values have also shown a significant inverse relationship with EGF-R. This data is in agreement with the observation that the presence of EGF-R is positively correlated with the progression towards a less-differentiated tumour grade (Fitzpatrick et al., 1984; Delarue et al., 1988; Sainsbury et al., 1987). Thus, grade I (Bloom and Richardson, 1957) tumours tend to be ER positive/ EGF-R negative and grade III tumours ER negative/ EGF-R positive. The breast cancer cell-lines also reflect the ER:EGF-R relationship (Davidson et al., 1987). Oestrogen dependent cell-lines which over-express ER, such as MCF-7 cells, express only low levels of EGF-R while steroid hormone-independent ER-negative cell-lines (e.g. MDA-MB-231 cells) over-express EGF-R. This, together with the relationship with tumour grade, has led to speculation that in breast cancer the two receptor systems interact in some way, and that tumours may progress towards a steroid hormone-independent state under which control or mis-control of growth is effected by EGF-R and its polypeptide ligands. In contrast, it has been reported that EGF-R (and EGF-R mRNA) and ER levels are unrelated in normal breast, benign tumours and non-malignant tissues from the cancerous breast (Travers et al., 1988; Barker et al., 1989), and most samples are EGF-R positive. In addition, immunohistochemical analysis indicates that EGF-R is present in epithelial, myoepithelial and stromal cells (Moller et al., 1989). It is likely, therefore, that the inverse relationship found in primary carcinomas is abnormal and may be confined to the tumour itself. EGF-R in tumours is associated with a number of cellular proliferation activation factors such as the products of c-fos (Murayama et al., 1988; Wilding et al., 1988) and c-myc (Fernandez-Pol et al., 1987), and EGF has been shown to be capable of inducing these in vitro (Church and Buick, 1988). EGF is also positively correlated with the cell proliferation marker ~53, which has cell transforming properties (Cattoretti ef al., 1988). However, related cellular oncogenes such c-erb-B [coding for a truncated form of the EGF-R, Downward et al. (1984)], and the c-erb-B-2/HER2/neu oncogene [encoding a related but distinct tyrosine kinase activity, Coussens et al., (1985)] seem to co-exist with EGF-R in breast carcinomas but are expressed independently (Slamon et al., 1987; Guerin et al., 1989; Gullick, 1989).

PROGNOSTIC

SIGNIFICANCE

OF EGF-R

A number of studies have assessed the prognostic significance of EGF-R in relation to other markers with respect to disease free survival and overall survival. EGF-R has been found to be associated with early recurrence, irrespective of tumour sub-type (Sainsbury et al., 1988) and to be an indicator of poor prognosis (Macias et al., 1987; Sainsbury et al., 1987) associated with poorly differentiated, more invasive tumours. Some studies have shown EGF-R levels to be correlated with the frequency of cell mitoses (Spitzer et al., 1987) and the incidence of

Epidermal growth factor in breast cancer

lymph-node metastases (Battaglia et al., 1988b; Spitzer et al., 1988). The presence of EGF-R is also associated with a lack of response to anti-oestrogen therapy (Nicholson et al., 1988a, 1989). IN VITRO REGULATION

Growth of tumours in vivo is thought to occur under the influence of autocrine and paracrine controls @porn and Roberts, 1985; Lippman et al., 1986; Stoschek and King, 1986). Most info~ation regarding possible regulatory mechanisms has been obtained from in vitro studies using breast cancer cell-lines. Generally these have been obtained from pleural effusions or ascites of patients with metastatic breast cancer. The best characterized of these in terms of receptor expression are MCF-7 (Horwitz et al., 1975) and T-47 D cells (Keydar et al., 1979). Work by Lippmann and colleagues with MCF-7 has established that these cells synthesize TGF--alpha and, as previously mentioned, this interacts with EGF-R. Furthermore, synthesis of this TGF-‘alpha’ is oestrogen-inducible (Lippman and Dickson, 1989). EGF is also produced by breast cancer cell lines (Murphy et al., 1988a) and as the binding of ligand to EGF-R causes downregulation of this receptor (Carpenter and Cohen, 1976; Stoscheck and Carpenter, 1984), long-term exposure to oestrogen-induced growth factors may explain the low or undetectable EGF-R concentrations found in primary carcinomas expressing high ER. While it is ctear that growth factors play an important role in the control of breast cancer cell proliferation the exact regulatory mechanisms involved remain unresolved. Although anti-oestrogens are able to inhibit oestrogen-stimulated proliferation in the oestrogen-sensitive cell line MCF-7, they have been shown to have no effect upon ~GF-stimulated growth (Cormier and Jordan, 1988). Moreover, oestrogen-stimulated MCF-7 cell proliferation is only partially or not at all attenuated by anti-EGF-Ror anti-TGF-‘alpha’-antibodies (Bates et al., 1988) while TGF-‘alpha’ directed proliferation is inhibited (Arteaga et al., 1988a) by these treatments. It is possible that other oestrogen-inducible growth factors such as IGF-I and related proteins (Huff et al., 1988) may be involved. Thus, breast cancer cell proliferation may be influenced by two mitogenic pathways, One involves possibly several different polypeptide growth factors acting alone or in concert. The second follows the mitogenic effects of oestrogen, which may also rely on the induction of some of these factors, and possibly other more direct intracellular effects on cell cycle progression factors such as c-myc and c-fos products. However, growth of MCF-7 cells in serum-free medium (i.e. in the complete absence of exogenous oestrogen and growth factors) can result in ceils which respond only minimally to added oestradiol compared with the stimulation of cell proliferation resulting from added IGF-I, EGF or TGF-‘alpha’ (Karey and Sirbasku, 1988). This suggests that oestradiol-stimulated mitogenesis requires additional serum-borne factors, such as estromedins (Ikeda et al., 1982), for maximum effect, while the poly-

941

peptide growth factors are able to act independently. However, despite this apparent independence, there is evidence that these pathways are interrelated by the inter-re~lation of their respective receptor moieties. Information regarding possible interactions between ER and EGF-R through the actions of their respective ligands has so far been contradictory. However, studies using MCF-7 cells indicate that E, can cause an increase in EGF-R levels with a concomitant decrease in ER, and that the anti-oestrogen hydroxytamoxifen causes a decrease in EGF-R (Berthois et al., 1989). In addition treatment of these cells with EGF was shown to increase ER and PgR, with a corresponding decrease in EGF-R levels, From these data the authors infer a parallel inverse relationship to that found in breast tumour biopsies. However, since binding of ligand to EGF-R causes receptor down-regulation, and TGF-alpha production is stimulated by E, and inhibited by antioestrogen treatment (Lippman and Dickson, 1989), it is not clear how far such a parallel extends. In contrast, other work has shown that treatment of MCF-7 cells with EGF has been shown to have no effect on ER mRNA levels (Read et al., 1989) so that the increase shown at the protein level may not result from an effect on transcription. Progestins also increase EGF-R (Murphy et al., 1986) EGF-R mRNA (Murphy et al., 1988b) and EGF mRNA (Murphy et al., 1988a) levels in T-47 D and other breast cancer cell lines. Conversely, EGF has been shown to diminish PgR levels in part by causing a decrease in ER (Cormier et al., 1989). Under these in vitro conditions EGF plays a dominant role (Sarup et al., 1988), capable of reversing any growth inhibitory effects of progestins. Although this suggests a possible role for progesterone and its receptor in the control of cell augmenting the mitogenic signal proliferation, by increasing levels of EGF and its receptor, progesterone is still regarded as a growth inhibitor under different conditions (Horwitz and Freidenberg, 1985). The present inability to establish a coherent and robust hypothesis regarding mechanisms of in vivo regulation is due to the complex nature of the problem and to problems inherent in extrapoIating data obtained from cell-line experiments to modeis for in zlivo regulation. The effectiveness of each potential candidate for in vivo ‘regulator’ is largely dependent upon the cell type and prior growth conditions (Read et al., 1989). Such differences in potential model systems are further accentuated by the “genetic instability” of cell lines during long-term culture (Katzenellenbogen ef al., 1987; Reddel et al., 1988). So while the true mechanism of steroid hormonegrowth factor interactions remains elusive this further highlights the difficulties encountered in the treatment of breast cancer. As tumours are known to be heterogeneous, being derived from possibly several different malignant cell types, a ho~one-responsive cancer may comprise both oestrogen-dependent and -independent cells (DeSombre et al., 1986). As a consequence, prolonged anti-oestrogen therapy may eventually fail due to the clonal selection of oestrogen-independent cell types (Jordan et al., 1989).

S. BARKER and G. P. VINSON

942 SYNOPSIS

REFERENCES

EGF is obviously a mediator of cellular proliferation in the human breast. It is likely that it is but one of many polypeptides factors which act locally in an autocrine and paracrine manner. Each factor probably has its own relative potency, as either a “competence” or “progression factor”, acting at a distinct part of the cell cycle to stimulate cells which express the relevant receptor protein to proceed to S-phase and cell division (Goustin et al., 1986; Yarden and Ullrich, 1988). Expression of these factors and their receptors may be under the control oestrogen. Under normal circumstances these interactions may be expected to be tightly regulated. Inhibitory factors such as TGF-beta (Sporn et al., 1986; Knabbe et al., 1987) and progesterone could provide the negative arm of this system. As EGF (Stoschek and Carpenter, 1984), oestrogen (Maxwell et al., 1987), and progesterone (Alexander et al., 1989) down-regulate their own receptors while initiating or preventing the transcription of other mRNAs it is possible that a balanced cycle of growth and differentiation exists in the normal breast. Only in abnormal cell-lines and tumour cells in oivo

Alexander I. E., Clarke C. L., Shine J. and Sutherland R. L. (1989) Progestin inhibition of progesterone receptor gene expression in human breast cancer cells. Molec. Endocr. 1377-1386. Arteaga C. L., Coronado E. and Osborne C. K. (1988a) Blockage of the epidermal growth factor receptor inhibits transforming growth factor ‘a:‘-induced but not estrogeninduced growth of hormone-dependent human breast cancer. Molec. Endocr. 2, 1064-1069. Arteaga C. L., Tandon A. K., Von Hoff D. D. and Osborne C. K. (1988b) Transforming growth factor ‘8’: potential autocrine growth inhibitor of oestrogen receptor-negative human breast cancer cells. Cancer Res. 48, 3898-3904.

would cell proliferation continue unabated as a result of a defect at some point in the control mechanism. Therein lies the potential for heterogeneity which is

seen in breast cancers, both between different patients and within the same tumour. In predominantly ER-positive/PgR positive cancers the defect may lie in mis-regulated ER expression, hence such tumours will probably have low or undetectable levels of EGF-R because of EGF/TGF-‘alpha’ induced downregulation. Seventy-five per cent of patients with these tumours will respond favourably to antioestrogen therapy. Predominantly EGF-R positive tumours may however have arisen from EGF-R gene amplification or from a defect in the ER system which would eliminate the inhibitory influence of the PgR, thus leading to unrestrained expression of EGF-R. Another possibility is that EGF-R may interact, perhaps even by heterodimerization, with oncogene products such as c-erbB-I and c-erbB-2 (King et al., 1988) which have extensive homology in the intracellular tyrosine-kinase domain. This could create an autonomous mitogenic signal which could evade the control normally provided by EGF binding. At present hormonal treatment of breast cancer is directed at the effects of oestrogen. However, with such endless potential for heterogeneity and the apparent adaptability of cancer cells to changing conditions. successful treatment of breast cancer will require a combined approach. An important part of this relies upon the characterization of ER-status, however anti-oestrogen therapy will need to be complemented by agents which can prevent growth factor stimulated mitogenesis. In conclusion, although EGF, per se, appears to play only a minor role in the progression of human breast cancer, information on its function and the regulation of its receptor will lead to more successful diagnosis and treatment. Acknowledgemen/-We Campaign for project

are grateful to the Cancer grant support.

Research

Barker S., Panahy C., Puddefoot J. R., Goode A. W. and Vinson G. P. (1989) Epidermal growth factor receptors and oestrogen receptors in the non-malignant part of the cancerous breast. Br. J. Cancer 60, 673477. Bates S. E., Davidson N. E., Valverius E. M., Freter C. E., Dickson R. B., Tam J. P., Kudlow J. E., Lippman M. E. and Salomon D. S. (1988) Expression of transforming growth factor ‘a’ and its messenger ribonucleic acid in human breast cancer: its regulation by estrogen and its possible functional significance. Molec. Endocr. 2, 543-555. Battaglia F., Polizzi G., Scambia G., Rossi S., Panici P. B., Iacobelli S., Crucitti F. and Mancuso S. (1988a) Receptors for epidermal growth factor and steroid hormones in human breast cancer. Oncology 45, 424427. Battaglia F., Scambia G., Rossi S., Panici P. B., Bellantone R., Polizzi G., Querzoli P., Negrini R., Iacobelli S., Crucitti F. and Mancuso. S. (1988b) Epidermal growth factor receptor in human breast cancer: correlation with steroid hormone receptors and axillary lymph node involvement. Eur. J. Cancer Clin. Oncol. 24, 1685-1690. Bauknecht T., Kohler M., Janz I. and Pfleiderer A. (1989) The occurrence of epidermal growth factor receptors and the characterisation of EGF-like factors in human ovarian, endometrial, cervical, and breast cancer. EGF receptors and factors in gynecological carcinomas. J. Cancer Res. Clin. Oncol. 115, 193-199. Beato M. (1989) Gene regulation by steroid hormones. Cell 56, 335-344 (120 refs.). Berthois Y., Dong X. F. and Martin P. M. (1989) Regulation of epidermal growth factor-receptor by oestrogen and anti-oestrogen in the human breast cancer cell-line MCF-7 Biochem. biophys. Res. Commun. 159, 126-131. Bloom N. D. and Richardson W. W. (1957) Histological grading and prognosis in breast cancer. Er. J. Cancer 11, 359-377. Cappelletti V., Brivio M., Miodini P., Granata G., Coradini D. and DiFronzo G. (1988) Simultaneous estimation of epidermal growth factor receptors and steroid receptors in a series of 136 resectable primary breast tumors. Tumor Biol. 9, 200-2 11. Carpenter G. (1987) Receptors for epidermal growth factor and other polypeptide mitogens. A. Rev. Eiochem 56, 881-914 (275 refs). Carpenter G. and Cohen S. (1976) ‘~51-labelled human epidermal growth factor: binding, internalisation, and degradation in human fibroblasts. J. cell. Biol. 71, 159-171. Carpenter G. and Cohen S. (1979) Epidermal growth factor. A. Rev. Biochem. 48, 198-216. Cattoretti G., Andreola S., Clemente C., D’Amato L. and Rilka F. (1988) Vimentin and p53 expression on epidermal growth factor receptor-positive, oestrogen receptor-negative breast carcinomas. Br. J. Cancer 57, 353-357.

Epidermal

growth

fa ctor in breast

Church J. G. and Buick R. N. (1988) G-protein mediated epidermal growth factor signal transduction in a human breast cancer cell line. Evidence for two intracellular pathways distinguishable by pertussis toxin. J. biol. Chem. 263, 42424246. Cachet C., Gill G. N., Meisenhelder J., Cooper J. A. and Hunter T. (1984) C-kinase phosphorylates the epidermal growth factor receptor and reduces its epidermal growth factor-stimulated tyrosine protein kinase activity. J. biol. Chem. 259, 2553--2556. Cohen S. (1962) Isolation of a mouse submaxillary gland protein accelerating incisor eruption and eyelid opening in the newborn animal. J. biol. Chem. 237, 1555-1562. Connolly J. M. and Rose D. P. (1988) Epidermal growth factor-like proteins in breast fluid and human milk. Lifp Sci. 42, 1751ll756. Cormier E. M. and Jordan V. C. (1988) Contrasting ability of antiestrogens to inhibit MCF-7 growth stimulated by estradiol or epidermal growth factor. Eur. J. Cancer C/in. Oncol. 25, 5743. Cormier E. M.. Wolf M. F. and Jordan V. C. (1989) Decrease in estradiol-stimulated progesterone receptor production in MCF7 cells by epidermal growth factor and possible clinical implication for paracrine-regulated breast cancer growth. Cancer Res. 49, 576-580. Coussens L., Yang-Feng T. L., Liao Y. C., Chen E., Gray A., McGrath J., Seeburg P. H., Libermann T. A., Schlessinger J., Francke U., Levinson A. and Ullrich A. (1985) Tyrosine kinase receptor with extensive homology to EGF receptor shares chromosomal location with neu oncogene. Science 230, I 132-I 139. Davidson N. E., Gelmann E. P. and Lippman M. E. (1987) Epidermal growth factor receptor gene expression in estrogen receptor-positive and negative human breast cancer cell lines. Molec. Endow. 1, 216-223. Delarue J. C.. Friedman S. Mouriesse H., May-Levin F., Sancho Garnier H. and Contesso G. (1988) Epidermal growth factor receptor in human breast cancers: correlation with estrogen and progesterone receptors. Breast Cancer Res. Treur. 11, 173-l 78. Derynck R., Goeddel D. V., Ullrich A., Gutterman J. U., Williams R. D.. Bringman T. S. and Berger W. H. (1987) Synthesis of messenger RNAs for transforming growth factors ‘0~’ and ‘8’ and the epidermal growth factor receptor by human tumors. Cancer Res. 47, 707-7 12. DeSombre E. R., Thorpe S. M., Rose C., Blough R. R., Andersen K. W., Rasmussen B. B. and King W. J. (1986) Prognostic usefulness of estrogen receptor immunocytochemical assays for human breast cancer. Cancer Res. (suppl.) 46, 4256~4264s. Downward J.. Yarden Y., Mayes E. L. V., Scrace G., Totty N.. Stockwell P., Ullrich A., Schlessinger J. and Waterfield M. D. (1984) Close similarity of epidermal growth factor receptors and V-erb-B oncogene proten sequences. Nature 307, 521-527. Fernandez-Pal J. A., Talkad V. D., Klos D. J. and Hamilton P. D. (1987) Suppression of the EGF-dependent induction of c-myc proto-oncogene expression by transforming growth factor beta in a human breast carcinoma cell line. Biochem. biophys. Res. Commun. 144, 1197-1205. Filmus J., Pollak M. N., Cailleau R. and Buick R. N. (1985) MDA-468, as human breast cancer cell line with a high number of epidermal growth factor (EGF) receptors, has an amplified EGF receptor gene and is growth inhibited by EGF. Biochem. biophys. Res. Commun. 128, 898-905. Fitzpatrick S. L., Brightwell J., Wittliff J. L., Barrows G. H. and Schultz G. S. (1984) Epidermal growth factor binding by breast tumor biopsies and relationship to estrogen receptor and progestin receptor levels. Cancer Res. 44, 3448-3453.

cancer

943

Ghosh-Dastidar P., Coty W. A., Griest R. E., Woo D. D. and Fox C. F. (1984) Progesterone receptor sub-units are high affinity substrates for phosphorylation by epidermal growth factor receptor. Proc. natn. Acad. Sci. V.S.A 81, 16541658. Gorski J., Welshons W. V., Sakai D., Hansen J., Walent J., Kassis J., Shull J., Stack G. and Campen C. (1986) Evolution of a model of estrogen action. Rec. Prog. Harm. Res. 2, 297-329. Gregory H. (1975) Isolation and structure of urogastrone and its relationship to epidermal growth factor. Narure 257, 325-327. Guerin M., Gabillot M., Mathieu M. C., Travagh J. P., Spielman M. M., Andrieu N. and Riou G. (1989) Structure and expression of c-erbB-2 and EGF receptor genes in inflammatory and non-inflammatory breast cancer: prognostic significance. Int. J. Cancer 43, 201-208. Gullick W. J. (1989) Expression of the c-erbB-2 proto-oncogene protein in human breast cancer. Rec. Results Cancer Res. 113, 51-55. Goustin A. S., Leof E. B., Shipley G. D. and Moses H. L. (1986) Growth factors and cancer. Cancer Res. 46, 1015-1029 (267 refs). Horwitz K. B. (1987) The structure and function of progesterone receptors in breast cancer. J. Steroid Biochem. 27, 447457 (141 refs). Horwitz K. B. and Freidenberg G. R. (1985) Growth inhibition and increase of insulin receptors in antiestrogen-resistant T47D,, human breast cancer cells by progestins: implications for endocrine therapies. Cancer Res. 45, 167-173. Horwitz K. B., Costlow M. E. and McGuire W. L. (1975) MCF-7: a human breast cancer cell line with estrogen, androgen, progesterone, and glucocorticoid receptors. Steroids 26, 785-795. Horwitz K. B., Wei L. L., Sedlacek S. M. and D’Arville C. N. (1985) Progestin action and progesterone receptor structure in human breast cancer: a reveiw. Rec. Prog. Horm. Res. 41, 2499316 (231 refs). Hsuan J. J., Panayotou G. and Waterfield M. D. (1989) Structural basis for epidermal growth factor receptor function.Progr. Growth Factor Res. 1, 23-32 (50 refs). Hubay C. A. Arafah B., Gordon N. H., Guyton S. P. and Crowe J. P. 1984) Hormone receptors: and update and application. Surg. C/in. North Am. 64, I15551172. Huff K. K., Knabbe C., Lindsay R., Lippman M. E. and Dickson R. B. (1988) Multihormone regulation of insulinlike growth factor-I-related protein in MCF-7 human breast cancer cells. Molec. Endocr. 2, 200-208. Hunter T. and Cooper J. A. (1985) Protein-tyrosine kinases. A. Rec. Biochem. 54, 897-930. Imai Y., Leung C. K. H., Friesen H. G. and Shiu R. P. C. (1982) Epidermal growth factor receptors and effect of epidermal growth factor on growth of human breast cancer cells in long-term tissue culture. Cancer Res. 42, 43944398. Ikeda T., Liu Q.-F., Danielpour D., Officer L. B., Iio M., Leland F. E. and Sirbasku D. A. (1982) Identification of estrogen-inducible growth factors (estromedins) for rat and human mammary tumor cells in culture. In Vifro 18, 961-907. Jordan V. C., Robinson S. P. and Welshons W. V. (1989) Resistance to anti-oestrogen therapy. In Drug Resisrance (Edited bv Kessel D.). CRC Press. Boca Raton. Karey K. P.-and Sirbasku D. A. (1988) Differential responsiveness of human breast cancer cell lines MCF-7 and T47D to growth factors and 17 ‘p’-estradiol. Cancer Res. 48, 40834092. Katzenellenbogen B. S., Kendra K. L., Norman M. J. and Berthois Y. (I 987) Proliferation, hormone responsiveness, and estrogen receptor content of MCF-7 human breast cancer cells grown in the short-term and long-term absence of estrogens. Cancer Res. 47, 43554360.

944

S. BARKER and G. P. VINSON

Keydar I., Chen L., Karyby S., Weiss F. R., DeLarea J., Radu M., Chaitcik S. and Brenner H. J. (1979) Establishment and characterisation of a cell line of human breast carcinoma origin. Eur. J. Cancer 15, 659670. King C. R., Borrello I., Bellot F., Comoglio P. and Schlessinger J. (1988) EGF binding to its receptor triggers a rapid tyrosine phosphorylation of the erbB-2 protein in the mammary tumor cell line SK-BR-3. EMBO Ji 7, 164771651. Knabbe C., Lippman M. E., Wakefield L. M., Flanders K. C.. Kasid A., Derynck R. and Dickson R. B. (1987) Evidence that transforming growth factor-‘/?’ is a hormonally regulated negative growth factor in human breast cancer cells. CeN 48, 417428. Lacroix H., Iglehart J. D.. Skinner M. A. and Kraus M. H., (1989) Overexpression of erbB-2 or EGF receptor proteins present in early stage mammary carcinoma is detected simultaneously in matched primary tumors and regional metastases. Oncogene 4, 145- 15 I. Laron Z., Kauli R. and Pertzelan A. (1989) Clinical evidence on the role of oestrogens in the development of the breasts. Proc. Royal Sot. Edinburgh 95B, 13.-22. Lippman M. E. and Dickson R. B. (1989) Mechanisms of growth control in normal and malignant breast epithehum. Rec. Prog. Horm. Res. 45, 383440 (459 refs). Lippman M. E., Dickson R. B., Bates S., Knabbe C., Huff K., Swain S., McManaway M., Bronzert D., Kasid A. and Gelmann. E. P. (1986) Autocrine and paracrine growth regulation of human breast cancer. Breast Cancer Res. Treat. 7, 59-70 (61 refs). Macias A.. Azavedo E., Hagerstrom T., Klintenberg C., Perez R. and Skoog L. (1987) Prognostic significance of the receptor for epidermal growth factor in human mammary carcinomas. Anticancer Res. 7, 459464. Massague J. (1983) Epidermal growth factor-like transforming growth factor. II. Interaction with epidermal growth factor receptors in human placenta membranes and A431 cells. J. biol. Chem. 258, 13614-13260. Maxwell B. L., McDonnell D. P., Conneely 0. M., Schulz T. Z., Greene G. L. and O’Malley B. W. (1987) Structural organisation and regulation of the chicken estrogen receptor. Molec. Endow. 1, 25535. McGuire W. L. (1980) An update on oestrogen and progesterone receptors in prognosis for primary and advanced breast cancer. In Hormones and Cancer (Edited by Iacobelli S.), pp. 3377343. Raven Press, New York. McGuire W. L., Carbone P. P.. Sears M. E. and Ecsher G. C. (1975) Estrogen Receptors in breast cancer: an overview. In Oeswogen Receptors in Breast Cancer (Edited by McGuire W. L., Carbone P. P. and Vollmer E. P.), pp. I-7. Raven Press, New York. Moller P., Mechtersheimer G., Kaufmann M., Moldenhauer G., Momberg F., Mattfeldt T. and Otto H. F. (1989) Expression of epidermal growth factor receptor in benign and malignant primary tumours of the breast. Virchows Archs A 414, 157-164. Murayama Y.. Kurdta S. and Mishim Y. (1988) Regulation of human estrogen receptor gene, epidermal growth factor receptor gene. and oncogens by estrogen and antiestrogen in MCF-7 breast cancer cells. Cancer Detect. Prev. 13, 103-107. Murphy L. C., Murphy, Dubik D., Bell G. I. and Shiu R. P. C. (1988a) Epidermal growth factor gene expression in human breast cancer cells: regulation of expression by progestins. Cancer Res. 48, 45554560. _ Murohv L. C., Murohv L. J. and Shiu R. P. C. (1988b) Prbgestin regulation -of EGF-receptor mRNA accumu: lation in T-47D human breast cancer cells. Biochem. biophys. Res. Commun. 150, 192-196. Murphy L. J., Sutherland R. L., Stead B., Murphy L. C. and Lazarus L. (1986) Progestin regulation of epidermal growth factor receptor in human mammary carcinoma cells. Cancer Res. 46, 728-734.

Neal D. E., Bennett M. K., Hall R. R., Marsh C., Abel P. D., Sainsbury J. R. C. and Harris A. L. (1985) Epidermalgrowth-factor receptors in human bladder cancer: comparison of invasive and superficial tumours. Lancet i, 366-367. Nicholson S., Halcrow P., Sainsbury J. R. C., Angus B., Chambers P., Farndon J. R. and Harris A. L. (1988a) Epidermal growth factor receptor (EGFr) status associated with failure of primary endocrine therapy in elderly postmenopausal patients with breast cancer. Br. J. Cancer 58, 810-814. Nicholson S., Sainsbury J. R., Needham G. K., Chambers P., Farndon J. R. and Harris A. L. (1988b) Quantitative assays of epidermal growth factor receptor in human breast cancer; cut-off points of clinical relevance. Inf. J. Cancer ‘42, 36-4 1. Nicholson S., Halcrow P., Farndon J. R.. Sainsbury J. R. C., Chambers P. and Harris A. L. (1989) Expression of epidermal growth factor receptors associated with lack of response to endocrine therapy in recurrent breast cancer. Lancer i, 1822185 O’Keefe E., Hollenberg M. D. and Cuatrecasas P. (1974) Epidermal growth factor. Characteristics of specific binding in membranes from liver, placenta, and other target tissues. Archs Biochem. Biophys. 164, 518-526. Osborne C. K., Hamilton B.. Titus G. and Livingston R. B. (1980). Epidermal growth factor stimulation of human breast cancer cells in culture. Cancer Res. 40, 2361-2366. Ozawa S., Ueda M., Ando N., Abe 0. and Shimuzu N. (1988) Epidermal growth factor receptors in cancer tissues of esophagus, lung, pancreas, colorectum, breast and stomach. jop. J. C&tier Res. (Gann) 79, 1201-1207. Pekonen F.. Partanen S.. Makinen T. and Rutanen E. M. (1988) Receptors for epidermal growth factor and insulin-like growth factor I and their regulation to steroid receptors in human breast cancer. Cancer Res. 48, 1343-l 347. Perez R., Pascual M., Macias A. and Lage A. (1984) Epidermal growth factor receptors in human breast cancer. Breasr Cancer Res. Treat. 4, 189-193. Perroteau I., Salamon D., DeBortoli M., Kidwell W., Hazarika P., Pardue R., Dedman J. and Tam J. (1986) Immunological detection and quantification of alpha transforming growth factors in human breast carcinoma cells. Breast Cancer Res. Treat. 7, 201&210. Read L. D., Green G. L. and Katzenellebogen B. S. (1989) Regulation of estrogen receptor messenger ribonucleic acid and protein levels in human breast cancer cell lines by sex steroid hormones, their antagonists, and growth factors. Molec. Endocr 3, 295-304. Reddel R. R., Alexander I. E., Koga M., Shine J. and Sutherland R. L. (1988) Genetic instability and the development of steroid hormone insensitivity in cultured T 47D human breast cancer cells. Cancer Res. 48, 43404347. Ro J., North S. M., Gallick G. E., Hortobagyi G. N., Gutterman J. U. and Blick M. (1988) Amplified and overexpressed epidermal growth factor receptor gene in uncultured primary human breast carcinoma. Cancer Res. 48, 161-164. Roberts A. B. and Sporn M. B. (1988) Transforming growth factor ‘b’. Adt. Cancer Res. $1, 107-145 (207 refs). Sainsburv J. R. C.. Farndon J. R.. Harris A. L. and Sherbet G. V.- (1985) Epidermal growth factor receptors on human breast cancers. Br. J. Surg. 72, 186-188. Sainsbury J. R. C., Farndon J. R., Needham G. K., Malcolm A. J. and Harris A. L. (1987) Epidermial-growth factor receptor status as predictor of early recurrence of and death from breast cancer. Luncer i, 139881402. Sainsbury J. R. C., Nicholson S., Angus B., Farndon J. R., Malcolm A. J. and Harris A. L. (1988) Epidermal growth factor receptor status of histological sub-types ofbreast cancer. Br. J. Cancer. 58, 458460.

Epidermal

growth

factor

Salamon D. S., Kidwell W. R., Kim N., Ciardello F., Bates S. E., Valverius E., Lippman M. E., Dickson R. B. and Stampfer M. (1989) Modulation by estrogen and growth factors of transforming growth factor-alpha and epidermal growth factor receptor expression in normal and malignant human mammary epithelial cells. Recent Results Cancer Res. 113, 5742. Sarup J. C., Rao K. V. and Fox C. F. (1988) Decreased progesterone binding and attenuated progesterone action in cultured human breast carcinoma cells treated with epidermal growth factor. Cancer Res. 48, 5071-5078. Savouret J. F.. Misrahi M., Loosfeh H., Atger M., Bailly A. Perrot-Applanat M., Vu Hai M. T., Guiochon-Mantel A., Jolivet A., Lorenzo F., Logeat F., Pichon M. F., Bouchard P. and Milgrom E. (1989) Molecular and cellular biology of mammalian progestrone receptors. Rec. Prog. Horm. Res. 45, 65-120. Schlessinger J. (1988) The epidermal growth factor receptor as a multifunctional allosteric protein. Biochemisfry 27, 3119-3123 (58 refs). Seibert K. and Lippman M. (1982) Hormone receptors in breast cancer, I. In Clinics in Oncology (Edited by Baum M.), pp. 735-794. Saunders Eastbourne (247 refs.). Slamon D. J., Clark G. M., Wong S. G., Levin W. J., Ullrich A. and McGuire W. L. (1987) Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science 235, 177-182 (45 refs). Spitzer E., Grosse R., Kunde D. and Schmidt H. E. (1987) Growth of mammary epithelial cells in breast-cancer biopsies correlates with EGF binding. Int. J. Cancer 39, 279-282. Spitzer E., Koepke K., Kunde D. and Grosse R. (1988) EGF binding is quantitatively related to growth in node-positive breast cancer. Breast Cancer Res. Treat. 12, 4549. Sporn M. B. and Roberts A. B. (1985) Autocrine growth factors and cancer. Nature 313, 745-747 (79 refs). Sporn M. B., Roberts A. B., Wakefield L. M. and Assoian R. K. (I 986) Transforming growth factor-‘p’: biological

in breast

cancer

945

function and chemical structure. Science 233, 532-534 (63 refs). Starkey R. and Orth D. N. (1977) Radioimmunoassay of human epidermal growth factor. J. Clin. Endocr. Metab. 45, 1144-J 153. Stoscheck C. M. and Carpenter G. (1984) “Downregulation” of EGF receptors: direct demonstraion of receptor degradation in human fibroblasts. J. Cell Biol. 98, 1048-1053. Stoscheck C. M. and King L. E. Jr (1986) Role of epidermal growth factor in carcinogenesis. Cancer Res. 46, 1030-1037 (156 refs). Todaro G. L., Fryling C. and De Larco J. E. (1980) Transforming growth factors produced by certain human tumor cells: polypeptides that interact with epidermal growth factor receptors. Proc. natn. Acad. Sri. U.S.A. 77, 5258-5262. Toft D. 0. and O’Malley B. W. (1972) Target tissue receptors for progesterone: the influence of estrogen. Endcrinology 90, 104L1045. Travers M. l?; Barrett-Lee P. J., Berger U., Luqmani Y. A., Gazet J.-C.. Powles T. J. and Coombes R. C. (1988) Growth factor expression in normal, benign, and malignant breast tissue. BMJ 2%, 1621-1624. Wilding G., Lippman M. E. and Gelmann, E. P. (1988) Effects of Steroid hormones and peptide growth factrors on protooncogene c-fos expression in human breast cancer cells. Cancer Res. 48, 802-805. Wrba F.. Reiner A., Markis-Ritzinger E. and Holzner J. H., Reiner G. and Spona J. (1988) Prognostic significance of immunohistochemical parameters in breast carcinomas. Parh. Res. Pratt. 183, 277-283. Yarden Y. and Schlessinger J. (1987) Self-phosphorylation of epidermal growth factor receptor: evidence for a model of intermolecular allosteric activation. Biochemistry 26, 14341442. Yarden Y. and Ullrich A. (1988) Growth factor receptor tyrosine kinases. A. Ret. Biochem. 57, 443478 (284 refs).