Performance and Operational Feasibility of Epstein-Barr Virus-Based Screening for Detection of Nasopharyngeal Carcinoma: Direct Comparison of Two Alternative Approaches.

PURPOSE: Two Epstein-Barr virus (EBV)-based testing approaches have shown promise for early detection of nasopharyngeal carcinoma (NPC). Neither has been independently validated nor their performance compared. We compared their diagnostic performance in an independent population. METHODS: We tested blood samples from 819 incident Taiwanese NPC cases (213 early-stage, American Joint Committee on Cancer version 7 stages I and II) diagnosed from 2010 to 2014 and from 1,768 controls from the same region, frequency matched to cases on age and sex. We compared an EBV antibody score using immunoglobulin A antibodies measured by enzyme-linked immunosorbent assay (EBV antibody score) and plasma EBV DNA load measured by real-time PCR followed by next-generation sequencing (NGS) among EBV DNA-positive individuals (EBV DNA algorithm). RESULTS: EBV antibodies and DNA load were measured for 2,522 (802 cases; 1,720 controls) and 2,542 (797 cases; 1,745 controls) individuals, respectively. Of the 898 individuals positive for plasma EBV DNA and therefore eligible for NGS, we selected 442 (49%) for NGS testing. The EBV antibody score had a sensitivity of 88.4% (95% CI, 86.1 to 90.6) and a specificity of 94.9% (95% CI, 93.8 to 96.0) for NPC. The EBV DNA algorithm yielded significantly higher sensitivity (93.2%; 95% CI, 91.3 to 94.9; P = 1.33 × 10-4) and specificity (98.1%; 95% CI, 97.3 to 98.8; P = 3.53 × 10-7). For early-stage NPC, the sensitivities were 87.1% (95% CI, 82.7 to 92.4) for the EBV antibody score and 87.0% (95% CI, 81.9 to 91.5) for the EBV DNA algorithm (P = .514). For regions with a NPC incidence of 20-100/100,000 person-years (eg, residents in southern China and Hong Kong), these two approaches yielded similar numbers needed to screen (EBV antibody score: 5,656-1,131; EBV DNA algorithm: 5,365-1,073); positive predictive values ranged from 0.4% to 1.7% and 1.0% to 4.7%, respectively. CONCLUSION: We demonstrated high sensitivity and specificity of EBV antibody and plasma EBV DNA for NPC detection, with slightly inferior performance of the EBV antibody score. Cost-effectiveness studies are needed to guide screening implementation.

Cigarette smoking increases the risk of nasopharyngeal carcinoma through the elevated level of IgA antibody against Epstein-Barr virus capsid antigen: A mediation analysis.

BACKGROUND: The study aims are to evaluate the associations between nasopharyngeal carcinoma (NPC) risk and cigarette smoking and to explore the effects of cigarette smoking on Epstein-Barr virus (EBV) infection for NPC risk. METHODS: 1235 male NPC cases and 1262 hospital-based male controls matched to cases were recruited across six collaborative hospitals between 2010 and 2014. Using a standardized questionnaire, information on cigarette smoking and other potential risk factors for NPC was obtained. Blood was collected and used for anti-EBV VCA IgA and anti-EBV EA-EBNA1 IgA testing using standard methods. Unconditional logistic regression analysis was used to estimate odds ratio (OR) with 95% confidence interval (CI) for each risk factor after adjusting for confounders. RESULTS: 63.6% of cases and 44.0% of controls reported ever smoking cigarettes. After full adjustment, current smokers had a significant 1.60-fold (95% CI = 1.30-1.97) and former smokers a borderline significant 1.27-fold (95% CI = 1.00-1.60) increased NPC risk compared to never smokers. NPC risk increased with increasing duration, intensity, and pack-years of cigarette smoking but not with age at smoking initiation. Among controls, anti-EBV VCA IgA seropositivity rate was higher in current smokers than never smokers (14.0% vs 8.4%; OR = 1.82; 95% CI = 1.19-2.79). Mediation analyses showed that more than 90% of the cigarette smoking effect on NPC risk is mediated through anti-EBV VCA IgA. CONCLUSION: This study confirms the association between long-term cigarette smoking and NPC and demonstrates that current smoking is associated with seropositivity of anti-EBV VCA IgA antibodies.

Evaluation of Rare and Common Variants from Suspected Familial or Sporadic Nasopharyngeal Carcinoma (NPC) Susceptibility Genes in Sporadic NPC.

BACKGROUND: Genetic susceptibility is associated with nasopharyngeal carcinoma (NPC). We previously identified rare variants potentially involved in familial NPC and common variants significantly associated with sporadic NPC. METHODS: We conducted targeted gene sequencing of 20 genes [16 identified from the study of multiplex families, three identified from a pooled analysis of NPC genome-wide association study (GWAS), and one identified from both studies] among 819 NPC cases and 938 controls from two case-control studies in Taiwan (independent from previous studies). A targeted, multiplex PCR primer panel was designed using the custom Ion AmpliSeq Designer v4.2 targeting the regions of the selected genes. Gene-based and single-variant tests were conducted. RESULTS: We found that NPC was associated with combined common and rare variants in CDKN2A/2B (P = 1.3 × 10-4), BRD2 (P = 1.6 × 10-3), TNFRSF19 (P = 4.0 × 10-3), and CLPTM1L/TERT (P = 5.4 × 10-3). Such associations were likely driven by common variants within these genes, based on gene-based analyses evaluating common variants and rare variants separately (e.g., for common variants of CDKN2A/2B, P = 4.6 × 10-4; for rare variants, P = 0.04). We also observed a suggestive association with rare variants in HNRNPU (P = 3.8 × 10-3) for NPC risk. In addition, we validated four previously reported NPC risk-associated SNPs. CONCLUSIONS: Our findings confirm previously reported associated variants and suggest that some common variants in genes previously linked to familial NPC are associated with the development of sporadic NPC. IMPACT: NPC-associated genes, including CLPTM1L/TERT, BRD2, and HNRNPU, suggest a role for telomere length maintenance in NPC etiology.

Enrichment of Human CCR6+ Regulatory T Cells with Superior Suppressive Activity in Oral Cancer.

Human oral squamous cell carcinoma (OSCC) constitutes an inflammatory microenvironment enriched with chemokines such as CCL20, which promote cancer cell invasion and tumor progression. We found that in OSCC there is a correlation between the expression of CCL20 and FOXP3 mRNA. Therefore, we hypothesized that OSCC may favor the recruitment and retention of regulatory T (Treg) cells that express the CCL20 receptor, CCR6. Interestingly, most (∼60%) peripheral blood Treg cells express CCR6, and CCR6+ Treg cells exhibit an activated effector/memory phenotype. In contrast, a significant portion (>30%) of CCR6- Treg cells were found to be CD45RA+ naive Treg cells. Compared to CCR6- naive or memory Treg cells, CCR6+ Treg cells exhibit stronger suppressive activity and display higher FOXP3 expression along with lower methylation at the Treg-specific demethylated region of the FOXP3 gene. This predominance of CCR6+ Treg cells was also found in the draining lymph nodes and tumor-infiltrating lymphocytes of OSCC patients with early or late clinical staging. Moreover, CCR6+ Treg cells isolated from tumor-infiltrating lymphocytes or draining lymph nodes maintained similar phenotypic and suppressive characteristics ex vivo as did their counterparts isolated from peripheral blood. These results suggest that CCR6 marks activated effector or memory Treg phenotypes with superior suppressive activity in humans.

Correction: Association between Ischemic Stroke and Iron-Deficiency Anemia: A Population-Based Study.

[This corrects the article DOI: 10.1371/journal.pone.0082952.].

Determining the critical effective temperature and heat dispersal pattern in monopolar radiofrequency ablation using temperature-time integration.

The radiofrequency ablation (RFA) lesion size is posited to be disproportionate to the total delivered energy, and temperature-time integration (TTI) may have a more critical effect on lesion size. The present study aimed to evaluate this hypothesis by determining the temperature threshold and temperature distribution over tissues during the RFA lesioning process. Using an ex vivo chicken tissue model and an in vivo rabbit model with RFA applied for 2 min under various target temperature settings, the resultant lesions were evaluated histologically using Masson's trichrome stain. The temperature distribution over the tissue during the RFA lesioning process was also determined using a VT02 Visual IR Thermometer. It was revealed that the thermal injury threshold for RFA in the chicken tissues was ~65°C, but that it ranged from 55-65°C in mammals. Using infra-red thermal imaging, the temperature gradient (from the center to the periphery) during the RFA lesioning process demonstrated a uniform heat diffusion pattern. This data supports the proposed hypothesis that TTI is a critical parameter in determining RFA lesion size and can be applied clinically using the following equation: [Target temperature - 55 (°C)] × time (sec) is proportional to RFA lesion size.

Light-emitting diode-assisted narrow band imaging video endoscopy system in head and neck cancer.

BACKGROUND/AIMS: To validate the effectiveness of a newly developed light-emitting diode (LED)-narrow band imaging (NBI) system for detecting early malignant tumors in the oral cavity. METHODS: Six men (mean age, 51.5 years) with early oral mucosa lesions were screened using both the conventional white light and LED-NBI systems. RESULTS: Small elevated or ulcerative lesions were found under the white light view, and typical scattered brown spots were identified after shifting to the LED-NBI view for all six patients. Histopathological examination confirmed squamous cell carcinoma. The clinical stage was early malignant lesions (T1), and the patients underwent wide excision for primary cancer. This is the pilot study documenting the utility of a new LED-NBI system as an adjunctive technique to detect early oral cancer using the diagnostic criterion of the presence of typical scattered brown spots in six high-risk patients. CONCLUSIONS: Although large-scale screening programs should be established to further verify the accuracy of this technology, its lower power consumption, lower heat emission, and higher luminous efficiency appear promising for future clinical applications.

Preliminary study of acoustic analysis for evaluating speech-aid oral prostheses: Characteristic dips in octave spectrum for comparison of nasality.

BACKGROUND/PURPOSE: Acoustic analysis is often used in speech evaluation but seldom for the evaluation of oral prostheses designed for reconstruction of surgical defect. This study aimed to introduce the application of acoustic analysis for patients with velopharyngeal insufficiency (VPI) due to oral surgery and rehabilitated with oral speech-aid prostheses. METHODS: The pre- and postprosthetic rehabilitation acoustic features of sustained vowel sounds from two patients with VPI were analyzed and compared with the acoustic analysis software Praat. RESULTS: There were significant differences in the octave spectrum of sustained vowel speech sound between the pre- and postprosthetic rehabilitation. Acoustic measurements of sustained vowels for patients before and after prosthetic treatment showed no significant differences for all parameters of fundamental frequency, jitter, shimmer, noise-to-harmonics ratio, formant frequency, F1 bandwidth, and band energy difference. The decrease in objective nasality perceptions correlated very well with the decrease in dips of the spectra for the male patient with a higher speech bulb height. CONCLUSION: Acoustic analysis may be a potential technique for evaluating the functions of oral speech-aid prostheses, which eliminates dysfunctions due to the surgical defect and contributes to a high percentage of intelligible speech. Octave spectrum analysis may also be a valuable tool for detecting changes in nasality characteristics of the voice during prosthetic treatment of VPI.

Using temperature-time integration as a critical parameter in using monopolar radiofrequency ablations.

It is generally believed that radiofrequency energy delivered to the tissue determines the RFA lesion size. The purpose of this study was to re-evaluate this relationship and propose a potentially parameter of temperature-time integration as a better indicator of RFA lesion size. Using an Ex Vivo lesioning model, fixed 300 J RFA lesions were created under target temperature settings of 65, 75, and 85 °C. The lesion sizes were recorded and compared. Under the target temperature of 65 and 75 °C, the RFA procedures were sustained for a period of time after reaching the target temperature. The correlation between the lesion size and the sustained time (Ts) after reaching the target temperature was calculated. Under the same amount of energy output (300 J), the lesion size created under the three different target temperatures (65, 75, and 85 °C) differs significantly. When the target temperature was set to 75 °C, the correlation coefficient between the Ts and the lesion area and the maximal effective radius (Mer) were 0.913 and 0.971, respectively. When the target temperature was set to 65 °C, the correlation coefficient between the Ts and the lesion area and the Mer were 0.962 and 0.923, respectively. The RFA lesion size is not proportional to the total delivered energy. The Temperature-time integration appears to be a much better indicator that critically influences the lesion size.

Extramedullary plasmacytoma of the nasopharynx: A case report and review of the literature.

Plasmacytoma is an extremely rare and discrete solitary mass of neoplastic monoclonal plasma cells. Extramedullary plasmacytoma (EMP) tends to occur during the fifth and seventh decades of life and is rarely diagnosed in younger patients. Only four cases of EMP have been previously reported in relatively young patients. Here we report a 15-year-old patient presenting with long-term nasal obstruction, who was found to have EMP of the nasopharynx. The patient was treated with surgery followed by radiotherapy with a satisfactory outcome. To the best of our knowledge, this study describes the the youngest individual with nasopharyngeal EMP to be reported in the literature.

Skewed distribution of IL-7 receptor-α-expressing effector memory CD8+ T cells with distinct functional characteristics in oral squamous cell carcinoma.

CD8(+) T cells play important roles in anti-tumor immunity but distribution profile or functional characteristics of effector memory subsets during tumor progression are unclear. We found that, in oral squamous carcinoma patients, circulating CD8(+) T cell pools skewed toward effector memory subsets with the distribution frequency of CCR7(-)CD45RA(-)CD8(+) T cells and CCR7(-) CD45RA(+)CD8(+) T cells negatively correlated with each other. A significantly higher frequency of CD127(lo) CCR7(-)CD45RA(-)CD8(+) T cells or CCR7(-)CD45RA(+)CD8(+) T cells among total CD8(+) T cells was found in peripheral blood or tumor infiltrating lymphocytes, but not in regional lymph nodes. The CD127(hi) CCR7(-)CD45RA(-)CD8(+) T cells or CCR7(-)CD45RA(+)CD8(+) T cells maintained significantly higher IFN-γ, IL-2 productivity and ex vivo proliferative capacity, while the CD127(lo) CCR7(-)CD45RA(-)CD8(+) T cells or CCR7(-)CD45RA(+)CD8(+) T cells exhibited higher granzyme B productivity and susceptibility to activation induced cell death. A higher ratio of CCR7(-)CD45RA(+)CD8(+) T cells to CCR7(-)CD45RA(-)CD8(+) T cells was associated with advanced cancer staging and poor differentiation of tumor cells. Therefore, the CD127(lo) CCR7(-)CD45RA(-)CD8(+) T cells and CCR7(-)CD45RA(+)CD8(+) T cells are functionally similar CD8(+) T cell subsets which exhibit late differentiated effector phenotypes and the shift of peripheral CD8(+) effector memory balance toward CCR7(-)CD45RA(+)CD8(+) T cells is associated with OSCC progression.

Association between ischemic stroke and iron-deficiency anemia: a population-based study.

BACKGROUND: Very little is known about the relationship between non-sickle cell anemia and stroke. The purpose of this study is to evaluate the association of iron-deficiency anemia (IDA) with stroke based on a nationwide coverage database in Taiwan. METHODS: The case-control study subjects were obtained from the Taiwanese Longitudinal Health Insurance Database 2000. We included 51,093 subjects with stroke as cases and randomly selected 153,279 controls (3 controls per case) in this study.Separate conditional logistic regression analyses were used to calculate the odds ratio (OR) for having been previously diagnosed with IDA between cases and controls.We further analyzed the association between stroke and IDA by stroke subtype. RESULTS: Results showed that 3,685 study subjects (1.81%) had been diagnosed with IDA prior to the index date; of those subjects, 1,268 (2.48%) were cases and 2,417 (1.58%) were controls (p<0.001). Conditional logistic regression shows that the OR of having previously received an IDA diagnosis among cases was 1.49 (95% CI: 1.39~1.60; p < 0.01) that of controls after adjusting for monthly income, geographic region, hypertension, diabetes, coronary heart disease, atrial fibrillation, heart failure, hyperlipidemia, tobacco use disorder, and alcohol abuse/alcohol dependency syndrome. Furthermore, the adjusted OR of prior IDA for cases with ischemic stroke was found to be 1.45 (95% CI: 1.34~1.58) compared to controls. However, we did not find any significant relationship between IDA and subarachnoid/intracerebral hemorrhage even adjusting for other confounding factors (OR=1.17, 95% CI=0.97~1.40). CONCLUSION: There is a significant association between prior IDA and ischemic stroke.

Patterns of recurrence and second primary tumors in oral squamous cell carcinoma treated with surgery alone.

The aim of the present study is to review the recurrence patterns and incidence of second primary tumors (SPTs) in patients with oral squamous cell carcinoma (OSCC) who underwent surgery alone without postoperative adjuvant therapy. Data on patients recorded in the head and neck cancer registry of Cathay General Hospital were reviewed. A total of 72 patients with T1-3N0 OSCC who underwent surgery alone were included. Among the 72 patients, 44 had T1 tumors, 22 had T2 tumors, and 6 had T3 tumors. The 5-year overall survival (OS) rate was 77.3%, the recurrence-free survival rate was 74.1%, and the SPT-free survival rate was 73%. Eighteen (25%) patients had disease recurrence (regional recurrence in nine patients, local recurrence in seven patients, and locoregional recurrence in two patients). Most patients with local recurrence alone (6/7 patients, 85.7%) could be salvaged with treatment. However, locoregional control was obtained in only five (45.5%) of 11 patients with neck recurrence after surgical salvage therapy. At the time of analysis, 20 patients developed SPTs, and 15 (75%) of the SPTs were in the oral cavity. The annual incidence of SPT was 5%. Neck recurrence and SPT were associated with 48.4% and 24.4% lower 5-year OS rates, respectively. Multivariate analysis revealed that neck recurrence was a significant risk factor for low OS (p = 0.008). Neck recurrence was the most important prognostic factor for OS. The incidence of SPT development was high. Regular and long-term monitoring for recurrence and development of SPTs is necessary to improve the survival rate.

Cryptococcal infection of the vocal folds.

Cryptococcosis of the larynx is rare. In this manuscript we present a case of laryngeal cryptococcosis with a clear history of pigeon exposure. Cryptococcal laryngitis may present as hoarseness without other typical symptoms, even in immunocompetent individuals.

Head and neck manifestations of spontaneous pneumomediastinum.

OBJECTIVE: Spontaneous pneumomediastinum (SPM) is a rare disease entity that often manifests localized signs in the head and neck region. The thoracic features of SPM have been well described; however, there is a paucity of information on its otolaryngological characteristics. The authors describe the clinical management among SPM patients having primarily head and neck symptoms. STUDY DESIGN: Case series with chart review. SETTING: Cathay General Hospital, Taiwan. RESULTS: The study included 13 men and 1 woman, with a mean age of 18.8 years (range, 14-29 years). The primary initial symptoms were neck swelling (11), neck pain (10), and odynophagia (9). Neck soft tissue and chest radiography was diagnostic of SPM in all patients. Conservative treatment consisted of bed rest and analgesics, which led to rapid resolution of SPM. CONCLUSIONS: SPM is a benign entity that responds well to conservative treatment. The results of our investigation highlight the importance of an ear, nose, and throat (ENT) clinical examination as a guide for diagnosing SPM because of the high percentage of ENT manifestations in the initial clinical profiles. Secondary causes of SPM must be ruled out to avoid an unfavorable outcome.

Increased prevalence of interleukin-17-producing CD4(+) tumor infiltrating lymphocytes in human oral squamous cell carcinoma.

BACKGROUND: T helper 17 (Th17) and regulatory T cells share plasticity in the expression of interleukin (IL)-17 and forkhead box P3 (FOXP3), but their mutual presence in human diseases is unclear. METHODS: IL-17 and FOXP3 were analyzed by immunohistostaining and flow cytometry. The cytokine milieu was analyzed by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: Oral squamous cell carcinoma expresses high levels of IL-1ß, IL-6, and transforming growth factor (TGF)-ß. A unique subset of FOXP3(+) IL-17-producing CD4(+) T cells was consistently identified in tumor-infiltrating lymphocytes from advanced stages of cancer, but not in the circulation, at a frequency of 0.5% to 5.5 % of total CD4(+) T and positively correlated with the frequency of IL-17(+)FOXP3(-) T cells. The IL-17(+)FOXP3(+) T cells express CCR6 and suppress the proliferation of autologous CD4(+) CD25(-) responder T-cells in vitro. CONCLUSIONS: The prevalence of IL-17-producing FOXP3(+) CD4(+) tumor infiltrating lymphocytes is increased in oral squamous cell carcinoma.

Complete unilateral maxillary sinus opacity in computed tomography.

BACKGROUND/PURPOSE: Unilateral maxillary sinus opacity can be caused by many diseases, but an exact diagnosis is difficult to make. The aim of this study was to describe the pathological conditions and clinical features of patients with unilateral maxillary sinus opacity. METHODS: From 2004 to 2008, 830 consecutive patients underwent sinus surgery or endonasal endoscopic biopsy at an academic tertiary care center. The preoperative computed tomography (CT) images for these patients were reviewed, and 11 6 patients were identified with complete unilateral maxillary sinus opacification. We then analyzed presenting symptoms, physical examinations, specific CT findings, and pathology. RESULTS: The most frequent diagnoses were as follows: chronic rhinosinusitis (52.6%), fungus ball (29.3%), antrochoanal polyp (2.6%), benign tumor (10.4%), and malignancy (5.1%). Fungus ball was the most common diagnosis (10/18, 55.6%) in the subgroup of patients with isolated maxillary sinus opacity without disease in the other sinuses. Nasal discharge and foul-smelling breath were more common in inflammatory disease than in neoplastic disease. Neoplastic disease was more likely to present as epistaxis; a polyp or mass revealed by nasal endoscopy; mass effect in the cheek, palate, or gingiva; and bony erosion on CT. Erosion of the maxillary posterolateral wall and extra-sinus extension suggested malignancy. CONCLUSION: Although unilateral maxillary sinus opacity is usually inflammatory in origin, fungal sinusitis and neoplastic disorder are also likely. A careful history-taking, a thorough head and neck examination including nasal endoscopy, and CT evaluation are all imperative for reaching a correct diagnosis.