BACKGROUND: In this study the total and phosphorylated amount of epidermal 
growth factor receptor 1 (EGFR) and 2 (HER2) were measured together with EGFR 
ligands in tissue samples of breast cancer patients in order to investigate 
interrelations and possible prognostic values.
METHODS: Samples of malignant and non-cancer autologous reference tissue were 
collected from 415 breast cancer patients. The tissue samples were cut and 
either paraffin-embedded or homogenized in a lysis buffer to extract the 
proteins. HER2 was measured using both immunohistochemistry (IHC)/fluorescence 
in situ hybridization (FISH) and ADVIA Centaur. Phosphorylated HER2 and EGFR 
(pHER2, pEGFR), total EGFR and the ligands: epidermal growth factor (EGF), 
transforming growth factor-α (TGFα), amphiregulin (AREG), heparin-binding 
EGF-like growth factor (HB-EGF), betacellulin (BTC) and epiregulin (EREG) were 
measured using the Luminex.
RESULTS: The HER2 positivity rate was determined to be 25.2% by the Centaur 
method vs. 15.8% by IHC and FISH. HER2, HB-EGF, TGFα and AREG were upregulated 
in cancer tissue as compared with autologous reference tissue while EGFR, pEGFR 
and EGF were downregulated (p<10-6). pEGFR in autologous reference tissue was 
negatively correlated to the number of positive lymph nodes and to the tumor 
size (p=0.0007 and p=0.001, respectively) and furthermore, decreased in the 
group of mastectomy operated patients as compared with the lumpectomy group 
(p<10-6). HB-EGF in cancer tissue was positively associated with high grade 
tumors (p<10-6) and pHER2, HB-EGF and BTC were associated with poor disease free 
survival (p=0.017, p=0.012 and p=0.0026, respectively).
CONCLUSIONS: Our study demonstrated a profound activation of the EGFR system. 
HB-EGF was increased by factor 10 in cancer tissue and related to the biological 
aggressiveness of the tumors, and pHER2, HB-EGF and BTC were associated with 
poor clinical outcome.