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Importance: Positive margins are associated with poor prognosis among patients with oral tongue squamous cell carcinoma (SCC). However, wide variation exists in the margin sampling technique., Objective: To determine the effect of the margin sampling technique on local recurrence (LR) in patients with stage I or II oral tongue SCC., Design, Setting, and Participants: A retrospective study was conducted from January 1, 1986, to December 31, 2012, in 5 tertiary care centers following tumor resection and elective neck dissection in 280 patients with pathologic (p)T1-2 pN0 oral tongue SCC. Analysis was conducted from June 1, 2013, to January 20, 2015., Interventions: In group 1 (n = 119), tumor bed margins were not sampled. In group 2 (n = 61), margins were examined from the glossectomy specimen, found to be positive or suboptimal, and revised with additional tumor bed margins. In group 3 (n = 100), margins were primarily sampled from the tumor bed without preceding examination of the glossectomy specimen. The margin status (both as a binary [positive vs negative] and continuous [distance to the margin in millimeters] variable) and other clinicopathologic parameters were compared across the 3 groups and correlated with LR., Main Outcomes and Measures: Local recurrence., Results: Age, sex, pT stage, lymphovascular or perineural invasion, and adjuvant radiation treatment were similar across the 3 groups. The probability of LR-free survival at 3 years was 0.9 and 0.8 in groups 1 and 3, respectively (P = .03). The frequency of positive glossectomy margins was lowest in group 1 (9 of 117 [7.7%]) compared with groups 2 and 3 (28 of 61 [45.9%] and 23 of 95 [24.2%], respectively) (P < .001). Even after excluding cases with positive margins, the median distance to the closest margin was significantly narrower in group 3 (2 mm) compared with group 1 (3 mm) (P = .008). The status (positive vs negative) of margins obtained from the glossectomy specimen correlated with LR (P = .007), while the status of tumor bed margins did not. The status of the tumor bed margin was 24% sensitive (95% CI, 16%-34%) and 92% specific (95% CI, 85%-97%) for detecting a positive glossectomy margin., Conclusions and Relevance: The margin sampling technique affects local control in patients with oral tongue SCC. Reliance on margin sampling from the tumor bed is associated with worse local control, most likely owing to narrower margin clearance and greater incidence of positive margins. A resection specimen-based margin assessment is recommended.

Head and neck squamous cell carcinoma (HNSCC) has a variety of causes. Recently, the human papilloma virus (HPV) has been implicated in the rising incidence of oropharyngeal cancer and has led to variety of studies exploring the differences between HPV-positive and HPV-negative HNSCC. The calcium-activated chloride channel TMEM16A is overexpressed in a variety of cancers, including HNSCC, but whether or not it plays different roles in HPV-positive and HPV-negative HNSCC is unknown. Here, we demonstrate that TMEM16A is preferentially overexpressed in HPV-negative HNSCC and that this overexpression of TMEM16A is associated with decreased patient survival. We also show that TMEM16A expression is decreased in HPV-positive HNSCC at the DNA, RNA, and protein levels in patient samples as well as cell lines. We demonstrate that the lower levels of TMEM16A expression in HPV-positive tumors can be attributed to both a combination of copy number alteration and promoter methylation at the DNA level. Additionally, our cellular data show that HPV-negative cell lines are more dependent on TMEM16A for survival than HPV-positive cell lines. Therefore, we suspect that the down-regulation of TMEM16A in HPV-positive HNSCC makes TMEM16A a poor therapeutic target in HPV-positive HNSCC, but a potentially useful target in HPV-negative HNSCC.

Genomic findings underscore the heterogeneity of head and neck squamous cell carcinoma (HNSCC). Identification of mutations that predict therapeutic response would be a major advance. We determined the mutationally altered, targetable mitogenic pathways in a large HNSCC cohort. Analysis of whole-exome sequencing data from 151 tumors revealed the phosphoinositide 3-kinase (PI3K) pathway to be the most frequently mutated oncogenic pathway (30.5%). PI3K pathway-mutated HNSCC tumors harbored a significantly higher rate of mutations in known cancer genes. In a subset of human papillomavirus-positive tumors, PIK3CA or PIK3R1 was the only mutated cancer gene. Strikingly, all tumors with concurrent mutation of multiple PI3K pathway genes were advanced (stage IV), implicating concerted PI3K pathway aberrations in HNSCC progression. Patient-derived tumorgrafts with canonical and noncanonical PIK3CA mutations were sensitive to an mTOR/PI3K inhibitor (BEZ-235), in contrast to PIK3CA-wild-type tumorgrafts. These results suggest that PI3K pathway mutations may serve as predictive biomarkers for treatment selection.

We sought to identify biomarkers of antitumor activity in patients with locally advanced head and neck cancer treated with therapy containing cetuximab, an epidermal growth factor receptor (EGFR) inhibitor. Patients with stage III-IVB head and neck cancer received cisplatin, docetaxel, and cetuximab (TPE) followed by radiotherapy, cisplatin, and cetuximab (XPE) and maintenance cetuximab in a phase II clinical trial. Serum and tissue biomarkers were examined for treatment-related changes and for association with clinical outcomes. Concentrations of 31 cytokines, chemokines and growth factors were measured before and after 3 cycles (9 weeks) of induction TPE using multi-analyte immunobead-based profiling (Luminex Corp., Austin, TX), with selected analytes validated by a single analyte enzyme-linked immunosorbent assay. Tumor biomarkers included phosphorylated signal transducer and activator of transcription-3 (pSTAT3), EGFR and human papillomavirus (HPV). Thirty-one patients had baseline biomarkers and 25 had paired samples, pre- and post-TPE. Adjusting for false discovery, 14 analytes including MCP1c, IP-10, Leptin, interleukin (IL)-5, Eotaxin, IL-6, G-CSF, CXCL5 changed significantly post TPE induction. Serum vascular endothelial growth factor (VEGF) and IL-6 levels were associated with tumor response as assessed by positron emission tomography and progression-free survival, however, the association was not significant after adjustment for false discovery. Analytes were not associated with toxicities, smoking history, HPV status, EGFR amplification, or pSTAT3 tumor protein levels. Baseline serum biomarkers, in particular VEGF and IL-6, were identified as potentially useful prognostic markers of cetuximab-containing therapy. Validation is warranted in future studies specifically designed to detect biomarker associations., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Purpose: Epidermal growth factor receptor (EGFR) overexpression is correlated with decreased survival in head and neck cancer (HNC) where the addition of EGFR inhibition to standard chemoradiation approaches has improved treatment responses. However, the basis for the limited efficacy of EGFR inhibitors in HNC is incompletely understood. G-protein-coupled receptors (GPCR) have been shown to be overexpressed in HNC where GPCR activation induces HNC growth via both EGFR-dependent and -independent pathways. We hypothesized that targeting GPCR-induced EGFR-independent signaling would improve the efficacy of EGFR inhibition., Experimental Design: Using a high-throughput phosphoproteome array, we identified proteins that were phosphorylated in HNC cells where EGFR expression was downmodulated by RNA interference (RNAi) in the presence or absence of a GPCR ligand. We confirmed the findings from the array by Western blotting followed by in vitro and in vivo phenotypic assays., Results: p70S6K phosphorylation was elevated approximately sixfold in EGFR siRNA-transfected cells treated with a GPCR ligand. In addition to RNAi-mediated EGFR downmodulation, GPCR-mediated phosphorylation of p70S6K was modestly increased by EGFR inhibitor cetuximab approved by the Food and Drug Administration. Biopsies from cetuximab-treated patients also displayed increased phospho-p70S6K staining compared with pretreatment biopsies. HNC cells were growth inhibited by both genetic and pharmacologic p70S6K targeting strategies. Furthermore, p70S6K targeting in combination with cetuximab resulted in enhanced antitumor effects in both in vitro and in vivo HNC models., Conclusions: These results indicate that increased phosphorylation of p70S6K in cetuximab-treated patients may be due to increased GPCR signaling. Therefore, the addition of p70S6K targeting strategies may improve treatment responses to EGFR inhibition., (©2011 AACR.)

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 alpha (ER alpha(nuc); P < 0.001) levels were significantly increased in HNSCC tumors (n = 56) compared with adjacent mucosa (n = 30), whereas nuclear estrogen receptor beta (ER beta(nuc)) levels did not differ (P = 0.67). Patients with high ER alpha(nuc) and EGFR tumor levels had significantly reduced progression-free survival compared with patients with low tumor ER alpha(nuc) and EGFR levels (hazards ratio, 4.09; P = 0.01; Cox proportional hazards). In contrast, high ER beta(nuc) tumor levels were not associated with reduced progression-free survival alone or when combined with EGFR., Conclusions: ER alpha and ER beta 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.

Bradykinin has been shown to promote growth and migration of head and neck squamous cell carcinoma (HNSCC) cells via epidermal growth factor receptor (EGFR) transactivation. It has also been reported that bradykinin can cause the induction of cyclooxygenase-2 (COX-2), a protumorigenic enzyme, via the mitogen-activated protein kinase (MAPK) pathway in human airway cells. To determine whether COX-2 is up-regulated by bradykinin in HNSCC, the current study investigated bradykinin-induced EGFR transactivation, MAPK activation, and COX-2 expression in human HNSCC cells. Bradykinin induced a concentration- and time-dependent induction of COX-2 protein in HNSCC, which was preceded by phosphorylation of EGFR and MAPK. These effects were abolished by the B2 receptor (B2R) antagonist HOE140 but not by the B1 receptor (B1R) antagonist Lys-[Leu(8)]des-Arg(9)-bradykinin. COX-2 induction was accompanied by increased release of prostaglandin E(2). No effect of a B1R agonist (des-Arg(9)-bradykinin) on p-MAPK or COX-2 expression was observed. B2R protein was found to be expressed in all four head and neck cell lines tested. Immunohistochemical analysis and immunoblot analysis revealed that B2R, but not B1R, was significantly overexpressed in HNSCC tumors compared with levels in normal mucosa from the same patient. In HNSCC cells, the bradykinin-induced expression of COX-2 was inhibited by the EGFR kinase inhibitor gefitinib or mitogen-activated protein kinase kinase inhibitors (PD98059 or U0126). These results suggest that EGFR and MAPK are required for COX-2 induction by bradykinin. Up-regulation of the B2R in head and neck cancers suggests that this pathway is involved in HNSCC tumorigenesis.

Although cardiac sarcomas are rare in comparison to their soft tissue counterparts, they are the second most common type of primary cardiac neoplasm. Of the few hundred cases reported, most has been based on autopsy series. A series of 27 cardiac sarcomas removed at surgery for curative and diagnostic intent were reviewed for clinicopathologic features with correlation to available postoperative follow-up data in 17 patients. There were 6 angiosarcomas, 6 myxofibrosarcomas, 3 malignant peripheral nerve sheath tumors, 3 leiomyosarcomas, 2 synovial sarcomas, 1 epithelioid hemangioendothelioma, 1 chondrosarcoma, 1 osteosarcoma, and 4 poorly differentiated sarcomas. There was a wide age and size range with slight female predilection. There were 20 cases that arose in the atria/pulmonary vessels, 4 in the ventricles, 1 in mitral valve, and 2 in epi/pericardium. There was a slight left predilection. The histologic grade was low in 4, moderate in 3, and high in 20 cases. Six high-grade and 1 low-grade tumors were also treated with adjuvant chemotherapy and/or radiation. In 17 patients with follow-up data, 6 of 12 patients with high-grade tumor died (4 within 5 days of the initial surgery, 1 in 21 months, and 1 in 131 months), and 1 patient with moderate-grade tumor and all 4 patients with low-grade tumor were alive without evidence of disease at the end of follow-up. Tumor grade appeared to be prognostically important in cardiac sarcoma. Long survival was achieved in patients who survived the initial surgery well.

Squamous cell carcinoma of the head and neck (SCCHN) is a leading cause of cancer deaths worldwide. Epidermal growth factor receptor (EGFR), an upstream mediator of signal transducer and activator of transcription (STAT)-3 is overexpressed in a variety of cancers, including SCCHN. Therapies such as monoclonal antibodies and tyrosine kinase inhibitors targeting EGFR have demonstrated limited antitumor efficacy, which may be explained, in part, by persistent STAT3 activation despite EGFR inhibition. STAT3 activation induces expression of target genes in SCCHN, including Bcl-X(L), a mediator of antiapoptotic activity. Bcl-X(L) is commonly overexpressed in SCCHN where it correlates with chemoresistance, making it a potential therapeutic target. Targeting the EGFR-STAT3-Bcl-X(L) pathway at several levels, including the upstream receptor, the intracellular transcription factor, and the downstream target gene, has not been investigated previously. Using erlotinib, an EGFR-specific reversible tyrosine kinase inhibitor in combination with a STAT3 transcription factor decoy, we found enhanced antitumor effects in vitro and in vivo. The combination of the STAT3 decoy and gossypol, a small molecule targeting Bcl-X(L), also yielded enhanced inhibition of cell proliferation. The triple combination of erlotinib, STAT3 decoy, and gossypol further enhanced cell growth inhibition and apoptosis in vitro, and it down-regulated signaling molecules further downstream of the EGFR-STAT3 signaling pathway, such as cyclin D1. These results suggest that combined targeting of several components of an oncogenic signaling pathway may be an effective therapeutic strategy for SCCHN.

Background: A subset of Chronic Rhinosinusitis (CRS) patients are those with eosinophilic CRS (ECRS). These patients remain the most refractory to medical and surgical intervention, and are thought to reflect an inflammatory process arising from a variety of causes. Recently the role of fungus in ECRS, defined as the presence of fungi in the eosinophilic mucin of CRS, has been proposed as causal in many cases. Other proposed causes of ECRS include bacterial superantigen induction of inflammation and aspirin sensitivity., Design: Retrospective review., Methods: Histopathology from patients with previously diagnosed ECRS was prospectively re-reviewed for bacterial and fungal presence and correlated with patients' demographic data. The study population was comprised of 55 specimens with ECRS from 34 patients, over a 3-year period from a single investigator at the University of Pittsburgh Medical Center., Results: Histologic presence of bacteria was shown in 34 of 55 (62%) of all ECRS specimens. Specimens with fungal colonization were more often associated with bacterial presence than without bacteria (17 of 22 [77%] vs. 5 of 22 [23%], P < .02)., Conclusion: Bacteria were present in almost two-thirds of all specimens, and most cases of fungal presence were accompanied by bacterial presence, potentially providing support for bacteria as a source of superantigen. Other possible reasons for the high rate of bacterial and fungal colonization of ECRS include increased pathogen exposure, or failure to eliminate pathogens.

Immunophenotypic analysis is useful in distinguishing reactive from neoplastic lymphoproliferations, particularly when tissue is limited or histologic findings are equivocal. Surface membrane immunoglobulin (SmIg) light chain restriction in B cells is especially helpful in documenting clonality, and the loss of SmIg by B cells in extramedullary sites also has been used as a criterion to support the presence of lymphoma. However, we identified 3 cases of benign follicular hyperplasia (in 101 cases analyzed) with profound expansions (56%-88% of the B cells) of SmIg light chain-negative B cells without clonality by immunoglobulin heavy chain gene polymerase chain reaction. Thus, although uncommonly encountered, lack of SmIg light chain expression by B cells should not necessarily be interpreted as indicative of lymphoma. Interestingly, 2 of the 3 patients with these "aberrant" expansions were HIV+, and such patients are at heightened risk for the development of lymphoma. Therefore, there is the potential for misdiagnosing lymphoma if flow cytometric data are interpreted inappropriately in isolation.