The human epidermal growth factor receptor (HER) family of receptor tyrosine 
kinase has been extensively studied in breast cancer; however, systematic 
studies of EGFR gene amplification and protein overexpression in breast 
carcinoma are lacking. We studied EGFR gene amplification by chromogenic in situ 
hybridization (CISH) and protein expression by immunohistochemistry in 175 
breast carcinomas, using tissue microarrays. Tumors with >5 EGFR gene copies per 
nucleus were interpreted as positive for gene amplification. Protein 
overexpression was scored according to standardized criteria originally 
developed for HER-2. EGFR mRNA levels, as measured by Affymetrix U133 Gene Chip 
microarray hybridization, were available in 63 of these tumors. HER-2 gene 
amplification by fluorescence in situ hybridization (FISH) and protein 
overexpression by immunohistochemistry were also studied. EGFR gene 
amplification (copy number range: 7-18; median: 12) was detected in 11/175 (6%) 
tumors, and protein overexpression was found in 13/175 (7%) tumors. Of the 11 
tumors, 10 (91%) with gene amplification also showed EGFR protein overexpression 
(2+ or 3+ by immunohistochemistry). The EGFR mRNA level, based on Affymetrix 
U133 chip hybridization data, was increased relative to other breast cancer 
samples in three of the five tumors showing gene amplification. Exons 19 and 21 
of EGFR, the sites of hotspot mutations in lung adenocarcinomas, were screened 
in the 11 EGFR-amplified tumors but no mutations were found. Three of these 11 
tumors also showed HER-2 overexpression and gene amplification. Approximately 6% 
of breast carcinomas show EGFR amplification with EGFR protein overexpression 
and may be candidates for trials of EGFR-targeted antibodies or small inhibitory 
molecules.