Characterization of the HER-2/neu Oncogene by Immunohistochemical and Fluorescence in Situ Hybridization Analysis in Oral and Oropharyngeal Squamous Cell Carcinoma Atif J. Khan, Bonnie L. King, Benjamin D. Smith, Grace L. Smith, Michael P. DiGiovanna, Darryl Carter, and Bruce G. Haffty 1 Departments of Therapeutic Radiology [A. J. K., B. L. K., B. D. S., G. L. S., B. G. H.], Medical Oncology [M. P. D.], and Pathology [D. C.], Yale University School of Medicine, New Haven, Connecticut 06520-8040 ABSTRACT Purpose: The role of HER-2/neu in squamous cell car- cinoma (SCC) of the head and neck is not well defined. The purpose of the current study is to measure the frequency of HER-2/neu expression, to demonstrate HER-2/neu gene am- plification in the cases found to be positive for protein overexpression, and to investigate the prognostic signifi- cance of overexpression and/or amplification in SCC of the head and neck. Experimental Design: A cohort of 77 patients with SCC of the oral cavity or oropharynx, with stage III or IV disease and uniformly treated with surgical resection and postoperative radiation, served as the primary patient population for the study. Of these, 56 patients had ade- quate follow-up and paraffin-embedded specimens avail- able for analysis. Median follow-up was 6.1 years. Each of the paraffin-embedded specimens were immunohisto- chemically stained for HER-2/neu expression and graded for intensity of staining by a pathologist. All cases that demonstrated positive staining by immunohistochemistry were analyzed by fluorescence in situ hybridization (FISH) to assess HER-2/neu amplification status. Results: Five-year survival for the 56 evaluable patients was 40%, with 25% experiencing local relapse, 18% re- gional relapse, and 25% distant relapse. The percentage of tumors staining positive for HER-2/neu by immunohisto- chemistry was 17%. There was no statistically significant correlation between HER-2/neu and T stage, N stage, tumor grade, survival, or disease-free survival. HER-2/neu expres- sion did correlate with vascular endothelial growth factor expression. FISH analysis revealed four cases that were amplified for HER-2/neu. Of note, of the 4 amplified cases, 2 suffered regional relapse, 1 suffered distant metastasis, and all 4 expired by 5 years of follow-up. Conclusions: This is the first demonstration of HER-2/ neu gene amplification by FISH in SCC of the head and neck. FISH validates a previously contested controversial role for HER-2/neu gene overexpression in SCC of the head and neck. The prognostic significance and clinical implica- tions of HER-2/neu expression and amplification in head and neck cancer will require additional studies. INTRODUCTION The HER-2/neu oncogene (alternatively known as c-erbB-2 or neu) encodes a M r 185,000 glycoprotein, p185, with extracellular, transmembrane, and intracellular domains (1, 2). Both the HER-2/neu gene sequence, located on the short arm of chromosome 17, as well as the p185 protein share extensive sequence homology with the epidermal growth factor receptor, and the protein has been designated a member of the epidermal growth factor receptor tyrosine kinase superfamily. It is thought that overexpression of p185 leads to increased basal tyrosine kinase activity, thus transforming cells by chronically stimulat- ing signal transduction pathways (2). Overexpression of HER- 2/neu has been associated with advanced disease, metastasis, and poor clinical outcome in breast, ovarian, non-small cell lung, endometrial, salivary, and differentiated gastric carcino- mas (1, 3–5). Less is known about tumors of nonglandular origin. SCC 2 of the head and neck is a common epithelial neo- plasm, and a large proportion of these are derived from the mucosa of the oral cavity and oropharynx (6). The 5-year survival rate for patients with oral and oropharyngeal SCCs has remained steadfastly at 53% (6, 7), failing to improve despite significant research exploring the pathogenesis and management of these tumors. At present, therapeutic decisions are based on clinical-pathological parameters, including age, TNM stage, and histological grade. Although useful, these factors often fail to differentiate between more and less aggressive lesions. As such, Received 7/13/01; revised 10/25/01; accepted 11/17/01. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 To whom requests for reprints should be addressed, at Department of Therapeutic Radiology, Yale University School of Medicine, P. O. Box 208040, New Haven, CT 06520-8040. Phone: (203) 785-2959; Fax: (203) 785-4622. 2 The abbreviations used are: SCC, squamous cell carcinoma; VEGF, vascular endothelial growth factor; IHC, immunohistochemistry; FISH, fluorescence in situ hybridization; TNM, Tumor-Node-Metastasis. 540 Vol. 8, 540 –548, February 2002 Clinical Cancer Research Research. on November 29, 2021. © 2002 American Association for Cancer clincancerres.aacrjournals.org Downloaded from