Human Cancer Biology Cross-Talk between Estrogen Receptor and Epidermal Growth Factor Receptor in Head and Neck Squamous Cell Carcinoma

Purpose: This study aimed to characterize estrogen receptor expression and signaling in head and neck squamous cell carcinoma (HNSCC) cell lines and patient tissues, and to evaluate estrogen receptor and epidermal growth factor (EGF) receptor (EGFR) cross-activation in HNSCC. Experimental Design: Estrogen receptor expression and signaling in HNSCC cell lines were assessed by immunoblotting. In vitro proliferation and invasion were evaluated in HNSCC cell lines in response to estrogen receptor and EGFR ligands or inhibitors. Estrogen receptor and EGFR protein expression in patient tissues was assessed by immunohistochemical staining. Results: Phospho–mitogen-activated protein kinase (P-MAPK) levels were significantly increased following combined estrogen and EGF treatment. Treatment of HNSCC cells with estrogen and EGF significantly increased cell invasion compared with either treatment alone, whereas inhibiting these two pathways resulted in reduced invasion compared with inhibiting either pathway alone. EGFR (P = 0.008) and nuclear estrogen receptor α (ERαnuc; P < 0.001) levels were significantly increased in HNSCC tumors (n = 56) compared with adjacent mucosa (n = 30), whereas nuclear estrogen receptor β (ERβnuc) levels did not differ (P = 0.67). Patients with high ERαnuc and EGFR tumor levels had significantly reduced progression-free survival compared with patients with low tumor ERαnuc and EGFR levels (hazards ratio, 4.09; P = 0.01; Cox proportional hazards). In contrast, high ERβnuc tumor levels were not associated with reduced progression-free survival alone or when combined with EGFR. Conclusions: ERα and ERβ were expressed in HNSCC, and stimulation with estrogen receptor ligands resulted in both cytoplasmic signal transduction and transcriptional activation. Estrogen receptor and EGFR cross-talk was observed. Collectively, these studies indicate that estrogen receptor and EGFR together may contribute to HNSCC development and disease progression. (Clin Cancer Res 2009;15(21):6529–40) Epidermal growth factor receptor (EGFR) is overexpressed in 40% to 90% of head and neck squamous cell carcinoma (HNSCC), and EGFR overexpression is associated with reduced HNSCC patient survival (1, 2). The EGFR-targeted chimeric monoclonal antibody cetuximab (C225, ImClone) has been Food and Drug Administration–approved for the treatment of HNSCC. Although EGFR is overexpressed in many HNSCC, clinical response to cetuximab and other EGFR-targeted therapies has been modest in clinical trials (3–5). In addition, response to EGFR-targeted treatment has not positively correlated with tumor EGFR levels in several studies (5–7). These data suggest that signaling pathways working in parallel or in concert with EGFR may modulate tumor response to EGFR-targeted therapies. The mechanisms of acquired or de novo resistance to EGFR targeting in EGFR-expressing tumors are incompletely understood. Estrogen receptor signaling independent of EGFR and/ or in concert with EGFR has been reported for cancers of the lung and esophagus (8–11), and combined EGFR and estrogen receptor targeting in lung cancer has been previously reported by our group to be a more effective antitumor therapy than targeting either alone (9). EGFR overexpression is common in HNSCC (12–14), and the role of EGFR signaling in HNSCC growth and invasion has been well established (15, 16). In contrast, reports of estrogen receptor α (ERα) and estrogen receptor β (ERβ) expression in HNSCC are conflicting, and reports characterizing estrogen receptor function in HNSCC are scarce. In addition, reported Authors' Affiliations: Departments of Otolaryngology, Pharmacology and Chemical Biology, and Pathology, University of Pittsburgh, Pittsburgh, Pennsylvania Received 4/6/09; revised 6/30/09; accepted 7/14/09; published OnlineFirst 10/13/09. Grant support: Oral Cancer Center at the University of Pittsburgh and NIH P50CA097190 SPORE in Head and Neck Cancer. 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. Requests for reprints: Laura P. Stabile, University of Pittsburgh, E1340 Thomas Starzl Biomedical Science Tower, 200 Lothrop Street, Pittsburgh, PA 15261. Phone: 412-623-2015; Fax: 412-648-1945; E-mail: [email protected]. F 2009 American Association for Cancer Research. doi:10.1158/1078-0432.CCR-09-0862 6529 Clin Cancer Res 2009;15(21) November 1, 2009 www.aacrjournals.org Research. on April 13, 2017. © 2009 American Association for Cancer clincancerres.aacrjournals.org Downloaded from Published OnlineFirst October 13, 2009; DOI: 10.1158/1078-0432.CCR-09-0862