p16 Overexpression in Sinonasal Squamous Cell Carcinoma: Association with Human Papillomavirus and Prediction of Survival Outcomes.

p16 overexpression is often used as a surrogate marker for human papillomavirus (HPV) infection in oropharyngeal squamous cell carcinoma but remains an uncertain diagnostic tool for HPV-related sinonasal squamous cell carcinoma (SNSCC). Our study involved 79 consecutive SNSCC patients who were treated at a tertiary referral university hospital during 2006-2021. We retrospectively examined their clinical characteristics and conducted p16 immunohistochemistry and HPV detection. We found that 12.7% of the patients exhibited p16 overexpression, which was significantly more common in the nasal cavity and increased from 2015 onward. The HPV was a high-risk type and viral loads ranged from 4.2 to 1.6 × 106 copies/ng DNA with genome integration. Five-year overall survival (OS) and five-year relapse-free survival (RFS) rates were 74.6% and 69.9%, respectively. Our multivariate analysis showed that T category (T1-4a) and hemoglobin levels (≥13.7) were significant favorable prognostic factors for OS, while T category, performance status, and p16 overexpression were significantly associated with RFS. In patients with p16 overexpression, OS was 100% and RFS was 90%. Our findings suggest that p16 overexpression is a reliable surrogate marker for transcriptionally active HPV infection and predicts a favorable prognosis.

Biomarkers for Predicting Anti-Programmed Cell Death-1 Antibody Treatment Effects in Head and Neck Cancer.

In recurrent or metastatic head and neck squamous cell carcinoma (R/M-HNSCC), survival outcomes are significantly better in patients who receive anti-programmed cell death-1 (PD-1) monoclonal antibody therapy than in those who receive standard therapy. However, there is no established biomarker that can predict the anti-PD-1 antibody treatment effect and immune-related adverse events (irAEs) in these patients. This study investigated the inflammatory and nutritional status in 42 patients with R/M-HNSCC and programmed cell death ligand-1 (PD-L1) polymorphisms (rs4143815 and rs2282055) in 35 of the 42 patients. The 1- and 2-year overall survival was 59.5% and 28.6%, respectively; the 1- and 2-year first progression-free survival was 19.0% and 9.5%, respectively, and the respective second progression-free survival was 50% and 27.8%. Performance status and inflammatory and nutritional status (assessed by the geriatric nutritional risk index, modified Glasgow prognostic score, and prognostic nutritional index) were identified as significant indicators of survival outcomes in multivariate analysis. Patients with ancestral alleles in PD-L1 polymorphisms had less frequent irAEs. Performance status and inflammatory and nutritional status before treatment were closely related to survival outcomes after PD-1 therapy. These indicators can be calculated using routine laboratory data. PD-L1 polymorphisms may be biomarkers for predicting irAEs in patients receiving anti-PD-1 therapy.

Human Papillomavirus Infection and EGFR Exon 20 Insertions in Sinonasal Inverted Papilloma and Squamous Cell Carcinoma.

This study aimed to clarify the roles of high-risk human papillomavirus (HR-HPV) infection and epidermal growth factor receptor (EGFR) exon 20 mutations in sinonasal inverted papilloma (IP) and sinonasal squamous cell carcinoma (SNSCC). Samples were collected from 20 cases with IP, 7 with IP and squamous cell carcinoma (IP-SCC), and 20 with SNSCC and examined for HPV infection and EGFR exon 20 mutations. Low- or high-risk HPV DNA was observed in 25% of IP, 57.1% of IP-SCC, and 35% of SNSCC cases. Transcriptionally active HR-HPV infections in IP-SCC and SNSCC, accompanied by p16 overexpression, were observed in 28.5% and 25% of cases, respectively. Heterozygous EGFR exon 20 amino acid insertions (ex20ins), located between amino acids 768-774, were observed in 45% of IP, 28.5% of IP-SCC, and 0% of SNSCC and chronic sinusitis cases. EGFR phosphorylation sites were located at tyrosine (Y) 845, Y1068, Y1086, and Y1197 and induced PI3K/AKT/mTOR activation. The phosphorylation pattern of EGFR with ex20ins resembled that of HPV-related SNSCC and oropharyngeal cancer. The transcriptionally active HR-HPV infection and ex20ins might be responsible for the pathogenesis of IP-SCC cases with different fashions. Since IP-SCC might be a multifactorial disease, further investigation is needed to understand IP-SCC etiology.

Brain Activity in Patients With Unilateral Vocal Fold Paralysis Detected by Functional Magnetic Resonance Imaging.

OBJECTIVES: Understanding brain activity in response to unilateral vocal fold paralysis is essential to determine the neural compensatory mechanism underlying adaptation to voice disorders and to develop novel and improved rehabilitation programs for these disorders. We aimed to clarify brain activity during phonation (prolonged vowel, |i:|) in patients with chronic left vocal fold paralysis (LVFP) and compare with that in normal controls. STUDY DESIGN: Case-control study. METHODS: This functional magnetic resonance imaging (fMRI) study of an event-related task comprised 12 individuals with LVFP of more than 6 months duration and 12 healthy controls. The experimental task alternated phonation (prolonged vowel, |i:|) and no phonation (rest) conditions. The functional images obtained were single-shot gradient-echo echo-planar imaging. The volumes were acquired parallel to the anterior-posterior commissure plane and were sensitive to BOLD contrast. Data sets were processed and statistically analyzed using Statistical Parametric Mapping 8 software. Within-group analyses were conducted by applying the one-sample t test (P < 0.001, uncorrected). A random-effects analysis was used for group comparison. RESULTS: The LVFP group showed significantly higher brain activity in the right premotor areas, left parietal lobule, right primary somatosensory areas, and bilateral supplementary motor area and lower brain activity in the auditory-related areas of the superior temporal gyrus. There were no significant correlations of the percent signal change on fMRI with disease duration, maximum phonation time, or age. CONCLUSION: Patients with chronic unilateral vocal fold paralysis have stronger activity during voluntary phonation in various central networks. More detailed information on the central nervous system regions related to voluntary phonation from early to chronic phase is needed to understand the compensatory mechanisms in vocal fold paralysis and to establish an effective rehabilitation program. This is the first report to investigate brain activity in chronic unilateral vocal fold paralysis.

Development of Antibodies against HPV-6 and HPV-11 for the Study of Laryngeal Papilloma.

Laryngeal papilloma (LP), which is associated with infection by human papillomavirus (HPV)-6 or -11, displays aggressive growth. The precise molecular mechanism underlying the tumorigenesis of LP has yet to be uncovered. Building on our earlier research into HPV-6, in this study, the viral gene expression of HPV-11 was investigated by quantitative PCR and DNA/RNA in situ hybridization. Additionally, newly developed antibodies against the E4 protein of HPV-6 and HPV-11 were evaluated by immunohistochemistry. The average viral load of HPV-11 in LP was 1.95 ± 0.66 × 105 copies/ng DNA, and 88% of HPV mRNA expression was found to be E4, E5a, and E5b mRNAs. According to RNA in situ hybridization, E4 and E5b mRNAs were expressed from the middle to upper part of the epithelium. E4 immunohistochemistry revealed a wide positive reaction in the upper cell layer in line with E4 mRNA expression. Other head and neck lesions with HPV-11 infection also showed a positive reaction in E4 immunohistochemistry. The distribution pattern of HPV DNA, viral mRNA, and E4 protein in LP with HPV-11 infection was quite similar to that of HPV-6. Therefore, it might be possible to apply these E4-specific antibodies in other functional studies as well as clinical applications, including targeted molecular therapies in patients with HPV-6 and HPV-11 infection.

Coordinated Expression of HPV-6 Genes with Predominant E4 and E5 Expression in Laryngeal Papilloma.

Laryngeal papilloma (LP) associated with human papillomavirus (HPV)-6 or -11 infection shows aggressive growth. However, the detailed molecular mechanism of virus-driven tumorigenesis has not been uncovered fully. HPV-6 viral gene expression and dynamic alterations were investigated with in situ localization of viral DNA and RNA in 13 patients with HPV-6-infected laryngeal papilloma. The average viral load was 4.80 × 105 ± 1.86 × 105 copies/ng DNA. E4, E5a, and E5b mRNAs accounted for 96% of the expression of 9 mRNAs. The alteration of viral DNA load during recurrence paralleled the mRNA expression levels, and the expression of all mRNAs showed a similar curve. E4, E5a, and E5b were expressed in the middle to upper part of the epithelium and were co-expressed in the same cells. E4 immunohistochemistry demonstrated an extensively positive reaction in the upper cell layer in accordance with E4 mRNA expression. These results suggest that individual viral genes are coordinately expressed for viral replication, virus release, and immunosurveillance avoidance. The newly developed E4-specific monoclonal antibody can be applied to further functional studies and clinical applications such as targeted molecular therapies.

Tracheoesophageal Diversion and Puncture for Preserving Phonation in Intractable Aspiration: A Case Series.

OBJECTIVE/HYPOTHESIS: Tracheoesophageal diversion (TED) can prevent damage to the respiratory system in patients with swallowing disorders and/or repetitive aspiration pneumonia; however, TED may cause the loss of phonation. Our previous study demonstrated that TED with tracheoesophageal puncture (TEP) prevents aspiration while retaining phonation. In this study, we aimed to further evaluate the feeding status and phonation of patients who underwent TED with TEP to verify the reproducibility of this procedure. STUDY DESIGN: Case series study. METHODS: We retrospectively reviewed the medical records of 11 patients who underwent TED with TEP for intractable aspiration from February 2017 to August 2019 at Fukuoka Sanno Hospital. We evaluated the preoperative penetration aspiration score (PPAS), daily activities, preoperative and postoperative food intake level scale (FILS) score, nutrition route, maximum phonation time(MPT), and postoperative communication method. RESULTS: The study population included 10 men and 1 woman (mean age, 66 years; range, 44-81) with a PPAS of 6.8 ± 1.0. The FILS score changed from 2.1 ± 0.5 preoperatively to 7.5 ± 2.0 postoperatively (paired t-test, P < .05), while the MPT changed from 7.9 ± 4.1 to 10.3 ± 4.2 s (paired t-test, P = .9). Preoperatively, a gastric fistula (eight patients [73%]) was the main nutrition route, followed by a gastric tube (two patients [18%]). Postoperatively, the main nutritional route for eight patients (73%) was oral, while the remaining three patients (27%) used the oral route occasionally. All patients maintained laryngeal phonation function, and eight (73%) used only laryngeal phonation for communication. CONCLUSIONS: Patients with intractable aspiration who wish to retain phonatory function should be advised to undergo TED with TEP. LEVEL OF EVIDENCE: 4 Laryngoscope, 131:E1965-E1970, 2021.

Brain Activity in Patients With Adductor Spasmodic Dysphonia Detected by Functional Magnetic Resonance Imaging.

OBJECTIVES: Spasmodic dysphonia (SD) is considered a focal dystonia. However, the detailed pathophysiology of SD remains unclear, despite the detection of abnormal activity in several brain regions. The aim of this study was to clarify the pathophysiological background of SD. STUDY DESIGN: This is a case-control study. METHODS: Both task-related brain activity measured by functional magnetic resonance imaging by reading the five-digit numbers and resting-state functional connectivity (FC) measured by 150 T2-weighted echo planar images acquired without any task were investigated in 12 patients with adductor SD and in 16 healthy controls. RESULTS: The patients with SD showed significantly higher task-related brain activation in the left middle temporal gyrus, left thalamus, bilateral primary motor area, bilateral premotor area, bilateral cerebellum, bilateral somatosensory area, right insula, and right putamen compared with the controls. Region of interest voxel FC analysis revealed many FC changes within the cerebellum-basal ganglia-thalamus-cortex loop in the patients with SD. Of the significant connectivity changes between the patients with SD and the controls, the FC between the left thalamus and the left caudate nucleus was significantly correlated with clinical parameters in SD. CONCLUSION: The higher task-related brain activity in the insula and cerebellum was consistent with previous neuroimaging studies, suggesting that these areas are one of the unique characteristics of phonation-induced brain activity in SD. Based on FC analysis and their significant correlations with clinical parameters, the basal ganglia network plays an important role in the pathogenesis of SD.