3.4 - A. Growth Factors and their Receptors

Several growth factor families that signal through receptor tyrosine kinases (RTKs) have been strongly implicated in breast tumorigenesis, particularly the EGF and IGF families, and there is some evidence to implicate two other types of growth factor/receptor pathways, namely the TGF-beta and wnt signaling pathways. Interestingly, the wnt and FGF pathways, which are frequently activated in "spontaneous" (=virally-induced) mouse models of mammary tumorigenesis, do not seem to play as dominant a role in human breast cancer.

A.1. Epidermal growth factor (EGF) system.
The EGF ligand/receptor family has been very convincingly implicated in breast cancer. The EGFs are potent mitogens, and downstream effectors of the pathway include c-myc and cyclin D. There are four closely related receptor tyrosine kinases (EGFR/HER/erbB1, erbB2/HER2/neu, erbB3 and erbB4) that bind ligands of the EGF family, including EGF, amphiregulin, heregulin, and transforming growth factor a. ErbB2/HER2 is the preferred heterodimerization partner for the other ErbB family members, and ErbB-containing heterodimers have a more potent and prolonged signaling ability which may explain why ErbB2 is the most frequently upregulated member of the ErbB family in breast cancer (Hynes, 2000). ErbB2 is amplified and/or overexpressed in 15-30% of breast cancers (Slamon et al., 1987). Elevated ErbB2 correlates with poor prognosis, lack of steroid receptors and aggressive tumor histology (Revillion et al., 1998). Increased ErbB2 may occur early in tumorigenesis and it is upregulated in up to 60% of DCIS, particularly in the comedo subtype (Revillion et al., 1998). Importantly, ErbB2 overexpression correlates with lack of response to endocrine therapy and chemotherapeutic agents. A monoclonal antibody, Herceptin, which targets the ErbB2 protein has shown some efficacy against advanced breast cancer. The EGF receptor (HER/ErbB1) is also overexpressed in 20-40% of breast tumors, and overexpression correlates with loss of ER and poorer prognosis, and may be more pronounced in metastases (Toi et al., 1991). ErbB3 is also upregulated in breast cancer, while overexpression of ErbB4 is uncommon and may actually be correlated with positive prognostic indicators (Stern, 2000). Several ligands for the EGF-Rs have been found to be upregulated in breast cancer (reviewed in Salomon et al., 1995). EGF protein is expressed in 15-30% of primary invasive breast cancers, and expression correlates with poor prognosis. Similarly, TGF-a is overexpressed in 30-50% of primary and metastatic breast cancers, and increased expression can already be seen in atypical ductal hyperplasias. Generally tumors that express TGF-a coexpress EGF receptors, suggesting an autocrine loop may be established. EGF and TGF-a expression appear to be negatively correlated with ER expression.

A. 2. Insulin-like growth factor (IGF) system.
IGF-I and IGF-II are pleiotropic growth factors whose actions are regulated by complexing with high affinity binding proteins (IGF-BPs) which determine access of the ligands to the high affinity cell surface receptors, IGF1R (an RTK) and IGF2R (mannose-6-phosphate receptor). The IGFs have mitogenic and antiapoptotic activities. They may be particularly important in tumorigenesis in preventing the apoptotic response to aberrant oncogene activation. Furthermore, the IGF and estrogen/ER pathways interact in a reciprocally positive way to enhance each other's activities. High IGF-II expression in breast tumors (mostly in the stromal compartment) is correlated with poor prognosis. Breast cancer patients have higher serum IGF-1 levels than controls and higher circulating IGF-1 is also associated with increased risk of developing breast cancer. IGF-R1 is significantly overexpressed and activated in breast cancers, though it is not clear that this correlates with poor prognosis. However, expression of the downstream signaling molecule IRS-1 is increased in breast cancer, correlates with ER status and does associate with poor prognosis in a subset of patients. For review, see (Zhang and Yee, 2000).

A. 3. Fibroblast growth factor (FGF) system.
The FGF ligand/receptor family affects cell growth and motility and have pro-angiogenic properties. The family has been convincingly implicated in mammary tumorigenesis in mouse models, where the mouse mammary tumor virus frequently integrates near to and activates FGF family members. Aberrant expression of FGFs and their receptors are seen in human breast cancer since FGFR1 is amplified in ~20% and FGFR4 in ~40% of human breast cancers, and elevated FGF-8 is found in a subset of malignant tumors. However, to date these changes have not been convincingly linked to increased proliferation or angiogenesis in the tumors nor do they correlate with prognosis (Dickson et al., 2000).

A. 4. Hepatocyte growth factor (HGF) system.
Hepatocyte growth factor/scatter factor and its receptor c-met are important in the regulation of cell proliferation, survival and motility. Serum HGF levels are increased in ~27% of primary breast cancer patients, and the elevated HGF was significantly associated with tumor size, nodal status and histological evidence of venous invasion, and well as predicting poor prognosis (Toi et al., 1998). C-Met was found to be overexpressed in 25% of node-negative invasive ductal carcinomas, and correlated with increased risk of metastatic disease, suggesting that establishment of an HGF/Met autocrine loop may promote invasion and/or metastasis (Camp et al., 1999). However, this is still a rather controversial area.

A. 5. Vascular endothelial growth factor (VEGF) system.
VEGFs are highly potent endothelial cell mitogens and regulators of vascular permeability that signal through a family of RTKS, including c-flt and c-flk. Expression of VEGF in breast tumors is significantly associated with decreased relapse-free survival and overall survival, suggesting a role for tumor-derived VEGF in promoting tumor angiogenesis (Gasparini, 2000).

A. 6. Transforming growth factor-b (TGF-b) system.
The TGF-bs are a family of pleiotropic growth regulators that play complex roles in carcinogenesis due to their ability both to inhibit epithelial cell proliferation, and to promote invasion and metastasis. The ligands activate membrane-bound serine-threonine kinases (TbRI and TbRII) and signal through activation and nuclear translocation of cytoplasmic Smad proteins. Mutational inactivation of TGF-b system components is very rare in breast cancer. However, decreased expression of the type II TGF-b receptor (TbRII) in epithelial hyperplasias without atypia is associated with a significantly increased risk of subsequently developing invasive breast cancer (Gobbi et al., 1999). Similarly reduced TbRII staining seen in 25-40% of DCIS and IBC is associated with more aggressive disease (Gobbi et al.). Conversely, increased TGF-b1 expression in tumors and metastases correlates with poor prognosis (Gold, 1999). The data are consistent with a model in which TGF-bfunctions as a tumor suppressor early in the tumorigenic process, and as an oncogene in the later stages.

A. 7. The Wnt system.
Wnts are a family of 19 highly related secreted signaling proteins that play critical roles in development. They activate a signaling pathway that involves the proto-oncogene b-catenin and the tumor suppressor protein, APC. Wnt1 was identified as the first mammary oncogene in the mouse, as a site of insertion of MMTV. Overexpression of WNT2, WNT4, WNT5A, WNT10B and WNT13, though not WNT1, has been described in some human breast tumors, and interestingly for WNT2, the pattern of expression switches from stromal only in normal tissue, to stroma plus epithelium in tumors (reviewed in Brown, 2001). As yet however, the overexpression of WNTs has not been convincingly associated with any biological or clinical parameters of tumorigenesis. Few if any mutations have been found in the downstream mediators b-catenin, APC or axin in breast cancers. However, increased cytoplasmic and nuclear staining for b-catenin (which reflects activation of the Wnt signaling path) has been seen in up to 60% of human breast cancer specimens, and this staining pattern correlates with poor prognosis (Lin et al., 2000). The staining pattern was also highly associated with cyclin D overexpression, suggesting that activation of the Wnt pathway may lead to dysregulation of cyclin D.