Patients with early-stage oropharyngeal cancer can be identified with label-free serum proteomics.

BACKGROUND: The increasing incidence of oropharyngeal squamous cell carcinoma (OPSCC) is mainly related to human papillomavirus (HPV) infection. As OPSCCs are often diagnosed at an advanced stage, mortality and morbidity remain high. There are no diagnostic biomarkers for early detection of OPSCC. METHODS: Serum from 25 patients with stage I-II OPSCC, and 12 healthy controls, was studied with quantitative label-free proteomics using ultra-definition MSE. Statistical analyses were performed to identify the proteins most reliably distinguishing early-stage OPSCCs from controls. P16 was used as a surrogate marker for HPV. P16-positive and P16-negative tumours were analysed separately. RESULTS: With two or more unique proteins per identification, 176 proteins were quantified. A clear separation between patients with early-stage tumours and controls was seen in principal component analysis. Latent structures discriminant analysis identified 96 proteins, most reliably differentiating OPSCC patients from controls, with 13 upregulated and 83 downregulated proteins in study cases. The set of proteins was studied further with network, pathway and protein-protein interaction analyses, and found to participate in lipid metabolism, for example. CONCLUSIONS: We found a set of serum proteins distinguishing early-stage OPSCC from healthy individuals, and suggest a protein set for further evaluation as a diagnostic biomarker panel for OPSCC.

Site-specific familial risk and survival of familial and sporadic head and neck cancer.

The vast majority of head and neck cancers (HNCs) are sporadic squamous cell carcinomas, smoking and heavy drinking being the main risk factors. However, little is known about the possible role of family history and the importance of inherited factors versus shared environment. We used Swedish population-based registries to study the family history of HNC. In order to estimate the risk for family members to get the same cancer, and the risk for cancer-specific death in patients with a family history of HNC compared with patients without a family history, multivariate Cox proportional hazards analyses were performed. A 1.43-fold increased risk for developing HNC in the first-degree relatives (FDRs) of HNC patients [hazard ratio (HR), 1.43; 95% CI, 1.28-1.61] was found, when compared with relatives of healthy controls. In spouses of patients with HNC, the risk for developing any HNC was moderately increased (HR, 1.25; 95% CI, 1.01-1.53), compared with spouses of healthy controls. In addition, a 1.34-fold increased risk for death of HNC was found in HNC patients with a family history of HNC (HR, 1.34; 95% CI, (1.03-1.73) compared with HNC patients without a family history. We found an increased risk for HNC in relatives and spouses of HNC patients, when compared with family members of healthy controls. This suggests that in addition to inherited factors, shared environmental factors have a significant role in the development of the cancer. Family history of HNC was associated with worse survival in a newly diagnosed HNC patient.

Oral squamous cell carcinoma patients can be differentiated from healthy individuals with label-free serum proteomics.

BACKGROUND: No blood biomarkers to detect early oral cavity squamous cell carcinoma (OSCC) without clinical signs exist - diagnosis is solely based on histology of a visible tumour. Most OSCC patients are diagnosed at advanced stage, which leads to significant morbidity and poor survival. Our aim was to find the serum screening or detection biomarkers in OSCC. METHODS: Serum samples from patients with OSCC treated at the Department of Otorhinolaryngology - Head and Neck Surgery, Helsinki University Hospital (Finland) were collected. Age- and gender-matched healthy individuals served as controls. Quantitative label-free proteomics in high definition MSE mode(HDMSE) was performed on 13 patients and 12 healthy samples. Various statistical analyses were performed on quantitative proteomics data to obtain the most influential proteins, which classify the patients vs healthy samples. RESULTS: In quantitative proteomic analysis (HDMSE), 388 proteins were quantified in our pilot study. A complete separation between cases and controls was seen in supervised and unsupervised classification techniques such as orthogonal projections on latent structure-discriminant analysis (OPLS-DA) and self-organising maps. Using OPLS-DA S-plot, we identified a set of eight proteins that completely separated OSCC patients from healthy individuals. CONCLUSIONS: Although the tumour stages varied from I to IVa, these potential biomarkers were able to identify all OSCCs demonstrating their sensitivity to detect tumours of all stages. We are the first to suggest a set of serum biomarkers in our pilot study to be evaluated further as a diagnostic panel to detect preclinical OSCC in risk patients.

Accuracy of preoperative MRI to assess lateral neck metastases in papillary thyroid carcinoma.

Primary treatment of papillary thyroid carcinoma (PTC) with lateral lymph node metastasis is surgery, but the extent of lateral neck dissection remains undefined. Preoperative imaging is used to guide the extent of surgery, although its sensitivity and specificity for defining the number and level of affected lymph nodes on the lateral neck is relatively modest. Our aim was to assess the role of preoperative magnetic resonance imaging (MRI) in predicting the requisite levels of neck dissection in patients with regionally metastatic PTC, with a focus on Levels II and V. All patients with PTC and lateral neck metastasis who had undergone neck dissection at the Department of Otorhinolaryngology-Head and Neck Surgery, Helsinki University Hospital, Helsinki, Finland from 2013 to 2016 and had a preoperative MRI available were retrospectively reviewed. A head and neck radiologist re-evaluated all MRIs, and the imaging findings were compared with histopathology after neck dissection. In the cohort of 39 patients, preoperative MRI showed concordance with histopathology for Levels II and V as follows: sensitivity of 94 and 67%, specificity of 20 and 91%, positive predictive value of 56 and 75%, and negative predictive value of 75 and 87%, respectively. In PTC, MRI demonstrated fairly high specificity and negative predictive value for Level V metastasis, and future studies are needed to verify our results to omit prophylactic dissection of this level. Routine dissection of Level II in patients with regionally metastatic PTC needs to be considered, as MRI showed low specificity.

Expression of toll-like receptors in HPV-positive and HPV-negative oropharyngeal squamous cell carcinoma--an in vivo and in vitro study.

The incidence of oropharyngeal squamous cell carcinoma (OPSCC) has increased over the past decades in many western countries. This trend is mainly attributed to the human papillomavirus (HPV). Cancer-related actions of immunological defense systems are being intensively researched. Human toll-like receptors (TLRs) are a family of pattern recognition receptors that participate in the immunological defense against pathogens, but their actions are also linked to cancer. The expression of TLRs in cervical epithelium alters both during the clearance of HPV infection and the HPV-induced neoplasia, but the expression of TLRs has not been studied in OPSCC. Thirty-five paraffin-embedded, formalin-fixed, squamous cell carcinoma tissue specimens were analyzed for TLRs 2, 3, 4, 5, 7, and 9 and HPV and p16 statuses. The TLR 9 expression was lower in HPV-positive tumors compared with HPV-negative tumors. TLR 7 was expressed in all cancer specimens, but elevated expression was evident in HPV and/or p16-positive tumors. The majority of p16-positive tumors did not express TLR 5, whereas its expression was stronger in p16-negative tumors. The results of in vitro analysis of five human OPSCC cell lines and one human oral tongue squamous cell carcinoma cell line agree with the in vivo trends: low levels of TLR 5 and high levels of TLR 7 in p16-positive OPSCC. Overall, TLR 7 and 9 expression patterns are demonstrated here to relate to the HPV status in vivo and TLR 5 and 7 expression patterns to the p16 status in vivo and in vitro.

The changing surgical management of juvenile nasopharyngeal angiofibroma.

The management of juvenile nasopharyngeal angiofibroma (JNA) has changed during the last decades but it still continues to be a challenge for the multidisciplinary head and neck surgical team. The aim of this study was to review the used treatment approach and outcome of JNA in a single institution series of 27 patients diagnosed and treated during the years 1970-2009. All patients were male, with the median age of 17 years (range 11-33 years). Surgery was used as the primary treatment in every case. Surgical approaches varied, transpalatal approach (N = 14) being the most common approach used in this series. During the last decade various other techniques were applied, including endoscopic (N = 3) resection. Two patients were additionally treated with antiangiogenic agents and one patient with stereotactic radiotherapy. The primary recurrence rate was 37% and it seemed to correlate with vascular density of tumour and the surgical approach used. We suggest that the management of JNA should be planned by an experienced head and neck surgeon, as part of a multidisciplinary team, preferably in a tertiary referral setting, and the recent development of the available therapies should be taken into account to minimise the risk of recurrence.

Targeted Treatment With Radio Frequency Ablation for Lingual Tonsil.

OBJECTIVES: Benign enlargement of the lingual tonsils due to various causes may cause symptoms that warrant treatment. Conventional lingual tonsillectomy remains a challenging procedure, and there is no established standard procedure. We aimed to review the patients receiving different methods of lingual tonsil surgery for various indications at our institute. METHODS: Retrospective clinical data on all patients with an ablative operation of the tongue base during the 8-year period between 2007 and 2014 at the Helsinki University Hospital, Helsinki, Finland, were reviewed. The larger cohort comprised 35 patients, of whom 26 were men (74%). Ten patients had undergone solely lingual tonsil radio frequency ablation (LTRFA). The minimum follow-up time for all patients was 2 years. RESULTS: Of the 10 patients, 5 patients with LTRFA had been operated on because of symptomatic lingual tonsil hypertrophy and 5 because of periodic fever associated with possible lingual tonsil involvement. In 2 of the 5 patients with periodic fever, the fever cycles ended after the operation. Of the 5 patients, 3 patients with symptomatic lingual tonsil hypertrophy have been non-symptomatic after 1 to 3 treatment sessions. The last 2 patients continue to have persistent symptoms. There were no major complications. CONCLUSIONS: Development of new approaches for the management of various lingual tonsil conditions is warranted. Lingual tonsil volume reduction by LTRFA seems to be a treatment alternative with low morbidity but with limited curative effect only.

Tongue Cancer Patients Can be Distinguished from Healthy Controls by Specific N-Glycopeptides Found in Serum.

PURPOSE: There are no blood biomarkers to detect early-stage oral cavity squamous cell carcinoma (OSCC) prior to clinical signs. Most OSCC incidence is associated with significant morbidity and poor survival. The authors aimed to use mass-spectrometry (MS) technology to find specific N-glycopeptides potentially serving as serum biomarkers for preclinical OSCC screening. EXPERIMENTAL DESIGN: Serum samples from 14 patients treated for OSCC (stage I or stage IV) with 12 age- and sex-matched controls are collected. Quantitative label-free N-glycoproteomics is performed, with MS/MS analysis of the statistically significantly different N-glycopeptides. RESULTS: Combined with a database search using web-based software (GlycopeptideID), MS/MS provided detailed N-glycopeptide information, including glycosylation site, glycan composition, and proposed structures. Thirty-eight tryptic N-glycopeptides are identified, having 19 unique N-glycosylation sites representing 14 glycoproteins. OSCC patients, including stage I tumors, can be differentiated from healthy controls based on the expression levels of these glycoforms. N-glycopeptides of IgG1, IgG4, haptoglobin, and transferrin have statistically significant different abundances between cases and controls. CONCLUSIONS AND CLINICAL RELEVANCE: The authors are the first to suggest specific N-glycopeptides to serve as potential serum biomarkers to detect preclinical OSCC in patients. These N-glycopeptides are the lead candidates for validation as future diagnostic modalities of OSCC as early as stage I.

Association of BMI-1 and p16 as prognostic factors for head and neck carcinomas.

CONCLUSIONS: BMI-1 is an upstream repressor of tumor suppressor p16 and their inverse expression patterns have been linked with patient survival in OPSCC. In this material only p16 remained a relevant prognostic marker in OPSCC. OBJECTIVES: HNSCC tumors carry variable phenotypes and clinical outcomes depending on their anatomical location. In OPSCC, expression of tumor suppressor p16 is used as a surrogate marker of HPV infection and has prognostic value. There are no good prognostic biomarkers for HNSCC tumors of other anatomical locations. AIM: To study the expression patterns of p16 and BMI-1 in not only oropharyngeal but also oral, hypopharyngeal, and laryngeal squamous cell carcinomas and to clarify their putative connections with clinical parameters, survival, and each other. METHOD: Hospital records on 130 patients (59 OPSCC, 18 OSCC, 20 HPSCC, and 33 LSCC) diagnosed between 1997-2008 at the Helsinki University Hospital, Finland, were reviewed. BMI-1 and p16 expressions were studied by immunohistochemistry. RESULTS: Sixty-eight per cent of OPSCC expressed p16 and expression correlated with lower age, lower T- and higher N-category, and with improved OS and DFS. BMI-1 expression was most prevalent in OPSCC and LSCC, but had no clinical correlations. No correlation between p16 and BMI-1 expression was found.

Toll-like receptors 3, 7, and 9 in Juvenile nasopharyngeal angiofibroma.

UNLABELLED: Juvenile nasopharyngeal angiofibroma (JNA) is a rare, benign tumor affecting adolescent males. The etiology of JNA as well as the causes determining the variable growth patterns of individual tumors remains unknown. Toll-like receptors (TLRs) are part of the innate immune response to microbes; by recognition of distinct features, they link to induction of pro-inflammatory signaling pathways. We immunostained TLR 3, 7, and 9 in 27 JNA specimens of patients treated at the Helsinki University Central Hospital, Helsinki, Finland, during the years 1970-2009. RESULTS: TLR 3, 7, and 9 expressions were found in stromal and endothelial cells of JNA, and their expression levels varied from negative to very strong positive. TLR 3 expression was found to have a significant correlation with the clinical stage of JNA. CONCLUSIONS: The present results propose a putative role of TLRs in the growth process of JNA.

Reconstruction of facial nerve after radical parotidectomy.

CONCLUSION: Most patients benefitted from immediate facial nerve grafting after radical parotidectomy. Even weak movement is valuable and can be augmented with secondary static operations. Post-operative radiotherapy does not seem to affect the final outcome of facial function. OBJECTIVES: During radical parotidectomy, the sacrifice of the facial nerve results in severe disfigurement of the face. Data on the principles and outcome of facial nerve reconstruction and reanimation after radical parotidectomy are limited and no consensus exists on the best practice. METHOD: This study retrospectively reviewed all patients having undergone radical parotidectomy and immediate facial nerve reconstruction with a free, non-vascularized nerve graft at the Helsinki University Hospital, Helsinki, Finland during the years 1990-2010. There were 31 patients (18 male; mean age = 54.7 years; range = 30-82) and 23 of them had a sufficient follow-up time. RESULTS: Facial nerve function recovery was seen in 18 (78%) of the 23 patients with a minimum of 2-year follow-up and adequate reporting available. Only slight facial movement was observed in five (22%), moderate or good movement in nine (39%), and excellent movement in four (17%) patients. Twenty-two (74%) patients received post-operative radiotherapy and 16 (70%) of them had some recovery of facial nerve function. Nineteen (61%) patients needed secondary static reanimation of the face.

Tenascin-C, GLUT-1, and syndecan-2 expression in juvenile nasopharyngeal angiofibroma: correlations to vessel density and tumor stage.

BACKGROUND: Juvenile nasopharyngeal angiofibroma (JNA) is a highly vascular tumor with some characteristics resembling those of vascular malformations. The significance of different growth and angiogenesis promoting factors for the etiology of JNA remains unsolved. METHODS: We analyzed the immunoexpressions of glucose transporter 1 (GLUT-1), tenascin-C (TNC), and syndecan-2 in a series of 27 patients with JNA and compared these with each other and the clinical data to investigate their possible connections and role in the angiogenesis and growth of JNA. RESULTS: We found that frequent stromal TNC expression had a strong correlation with vessel density and tumor stage and endothelial GLUT-1 expression, when present, correlated with higher tumor stage. Stromal TNC and stromal GLUT-1 expressions were also found to correlate with each other. CONCLUSIONS: The immunoexpression of stromal TNC correlated with vascular density and higher tumors stage, which supports the idea of TNC having a role in the tumorigenesis of JNA putatively by promoting angiogenesis.

Systems-level analysis of clinically different phenotypes of juvenile nasopharyngeal angiofibromas.

OBJECTIVES/HYPOTHESIS: To explore the molecular genetic background of juvenile nasopharyngeal angiofibromas and to identify biological processes and putative factors determining the different growth patterns of these tumors. STUDY DESIGN: By comparing copy number and gene expression level changes of two clinically different phenotypes of juvenile nasopharyngeal angiofibromas, we aimed to find processes essential in the growth and development of these tumors. Based on the results and prior knowledge of the proteins significance for growth, we studied the expression of tyrosine kinase SYK in 27 tumor samples. METHODS: Comparative genomic hybridization and gene expression analyses were performed for the two tumor samples, and protein expression of SYK was studied in 27 samples by immunohistochemical staining. RESULTS: Between low- and high-stage juvenile nasopharyngeal angiofibromas, 1,245 genes showed at least a two-fold change in expression. The corresponding proteins of these transcripts were enriched in different biological processes. Protein kinase SYK was expressed in all 27 samples, and its intensity significantly correlated with tumor stage. CONCLUSIONS: Because the molecular genetic background of juvenile nasopharyngeal angiofibroma is unknown, our aim was to investigate genomic alterations that could associate to low- and high-stage tumors. We were able to identify gene expression changes that relate to particular biological processes, but assessing clinically relevant molecular profiles still requires further characterization. Due to the low incidence of juvenile angiofibroma, in the future a combination of molecular profiling data from several studies would be useful in understanding the molecular background of the disease.

Stem cell-related proteins C-KIT, C-MYC and BMI-1 in juvenile nasopharyngeal angiofibroma--do they have a role?

Juvenile nasopharyngeal angiofibroma (JNA) is a highly vascular tumour, occurring almost exclusively in adolescent males. Histogenesis of JNA remains unclear, two optional theories proposing either fibrous or vascular tissue as the tissue of origin. Stem cell-related proteins have been discussed to possibly participate in the growth of these tumours. In our study, we reviewed retrospective clinicopathological data of 26 JNA patients. By immunohistochemistry, we investigated the cellular distribution and expression levels of stem cell-related proteins C-KIT, C-MYC and BMI-1 and their correlation with cell and vessel density of the tumour. Contrary to earlier reports, we detected C-KIT expression in addition to stromal cells also in endothelial cells. The C-KIT expression was more dominant in slit vessels than large vessels. A significant correlation was found between endothelial immunoexpression of C-KIT and cellular density of the tumour. C-MYC and BMI-1 expression was detected in stromal cells only. Due to our finding of C-KIT expression in both stromal and endothelial cells and the strong correlation between the endothelial C-KIT expression and cellular density, we suggest that, besides the stromal tissue, the vascular component might take part in the neoplastic growth of JNA.

Juvenile nasopharyngeal angiofibroma: no evidence for inheritance or association with familial adenomatous polyposis.

Juvenile nasopharyngeal angiofibromas (JNAs) are rare tumors with prominent vascularity and locally destructive growth. The pathogenesis of JNA is largely unknown. A causal association between JNA and familial adenomatous polyposis has been suggested. Twenty-one patients diagnosed with juvenile angiofibroma filled out a detailed patient questionnaire. No patients reported any relatives with nasopharyngeal angiofibroma or familial adenomatous polyposis. No significant regional clustering suggestive for founder effect could be identified. We believe that if there were a strong genetic predisposition or association with familial adenomatous polyposis, it should have been seen in this patient sample.