The ATR inhibitor berzosertib acts as a radio- and chemosensitizer in head and neck squamous cell carcinoma cell lines.

Alterations in the DNA damage response play a crucial role in radio- and chemoresistance of neoplastic cells. Activation of the Ataxia telangiectasia and Rad3-related (ATR) pathway is an important DNA damage response mechanism in head and neck squamous cell carcinoma (HNSCC). Berzosertib, a selective ATR inhibitor, shows promising radio- and chemosensitizing effects in preclinical studies and is well tolerated in clinical studies. The aim of this study was to elucidate the effect of berzosertib treatment in combination with radiation and cisplatin in HNSCC. The HNSCC cell lines Cal-27 and FaDu were treated with berzosertib alone and in combination with radiation or cisplatin. Cell viability and clonogenic survival were evaluated. The effect of combination treatment was evaluated with the SynergyFinder or combination index. Apoptosis was assessed via measurement of caspase 3/7 activation and migration was evaluated using a wound healing assay. Berzosertib treatment decreased cell viability in a dose-dependent manner and increased apoptosis. The IC50 of berzosertib treatment after 72 h was 0.25-0.29 µM. Combination with irradiation treatment led to a synergistic increase in radiosensitivity and a synergistic or additive decrease in colony formation. The combination of berzosertib and cisplatin decreased cell viability in a synergistic manner. Additionally, berzosertib inhibited migration at high doses. Berzosertib displays a cytotoxic effect in HNSCC at clinically relevant doses. Further evaluation of combination treatment with irradiation and cisplatin is strongly recommended in HNSCC patients as it may hold the potential to overcome treatment resistance, reduce treatment doses and thus mitigate adverse events.

Early on-treatment C-reactive protein and its kinetics predict survival and response in recurrent and/or metastatic head and neck cancer patients receiving first-line pembrolizumab.

PURPOSE: First-line immune checkpoint blockade has improved the prognosis of recurrent and/or metastatic head and neck squamous cell carcinoma (R/M HNSCC), but response rates remain low. In this study, we aimed to investigate the prognostic value of CRP and its early kinetics to predict response and survival in R/M HNSCC. METHODS: A total of 87 patients who received first-line pembrolizumab for R/M HNSCC were analyzed. Three-fold cross-validation was used to estimate cut-off points of CRP at baseline and on-treatment (day 40 ± 10). Treatment response and survival were analyzed according to early CRP kinetics. The neutrophil-to-lymphocyte ratio (NLR) was used as a benchmark for the prognostic performance of CRP. RESULTS: On-treatment CRP below 2 mg/dl, 4x the upper limit of normal (ULN), was associated with increased overall survival (OS), while on-treatment CRP below 3 mg/dl (6x ULN) was correlated with a higher disease control rate (DCR) and increased progression-free survival (PFS). CRP flare-responders and CRP responders showed a higher DCR and longer PFS than CRP non-responders. An NLR above 6 was a negative prognosticator for progression. In multivariable analysis, on-treatment CRP prevailed as the only significant prognosticator for OS (HR: 4.97, CI95%: 2.18-11.32, p < 0.001) and PFS (HR: 2.07, CI95%: 1.07-3.99, p = 0.030). CONCLUSION: On-treatment CRP was identified as a prognostic biomarker for objective response and survival in R/M HNSCC patients receiving first-line pembrolizumab and could be easily incorporated into clinical practice as a widely available and cost-effective biomarker.

Low free triiodothyronine and immune-related hyperthyroidism are associated with overall and progression-free survival in head and neck squamous cell carcinoma treated with pembrolizumab.

INTRODUCTION: Thyroid function is frequently impaired in recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC). In patients treated with pembrolizumab, immune-related adverse events (irAEs) of the thyroid are common. However, the prognostic significance of baseline and on-treatment thyroid dysfunction is currently unclear. METHODS: This study included 95 patients who received pembrolizumab for R/M HNSCC between 2016 and 2022. Baseline thyroid status, according to serum hormone levels, and irAEs were assessed. Univariable and multivariable Cox regression analyses were performed for overall survival (OS) and progression-free survival (PFS). Furthermore, the best overall response according to the prognostic groups was examined. RESULTS: Low fT3 (HR: 2.52, p = 0.006), immune-related hyperthyroidism (HR: 0.11, p = 0.038), ECOG performance status ≥2 (HR: 3.72, p = 0.002), and platinum-refractory disease (HR: 3.29, p = 0.020) were independently associated with OS. Furthermore, immune-related hyperthyroidism was associated with longer PFS (HR: 0.13, p = 0.007), a higher objective response rate (83% vs. 31%, p = 0.018), and a higher disease control rate (100% vs. 43%, p = 0.008). Thyroid-related autoantibodies were elevated in 40% of thyroid irAEs cases with available measurements. Out of 16 thyroid irAEs, 15 occurred in patients with fT3 above the lower limit of normal. CONCLUSION: Low fT3 was associated with worse OS. Immune-related hyperthyroidism was correlated with both improved OS and PFS. Baseline fT3 assessment and close on-treatment monitoring of serum thyroid levels may be valuable for risk stratification in R/M HNSCC patients receiving pembrolizumab.

High USP4 mRNA is associated with an HPV-positive status in head and neck squamous cell carcinoma patients.

INTRODUCTION: Head and neck squamous cell carcinoma (HNSCC) is among the most common cancers in the world with a low survival rate and common diagnosis at late stages. Deubiquitination of proteins is involved in tumor growth, metastasis, apoptosis, and immunosuppressive pathways. The impact of the ubiquitin-specific protease (USP4) on survival was only scarcely investigated so far. The goal of our research was to analyze the association of USP4 expression with prognosis and clinicopathological features in HNSCC. METHODS: USP4 mRNA levels were derived from The Cancer Genome Atlas (TCGA) for a cohort of 510 patients. Protein expression of USP4 was analyzed by immunohistochemistry in a second cohort of 113 patients. Associations between USP4 levels and overall survival, disease-free survival and clinicopathological data were analyzed. RESULTS: High levels of USP4 mRNA were associated with prolonged overall survival in univariable analysis. There was no more association with survival after correction for the confounders HPV, stage and smoker status. High USP4 mRNA levels were linked to a lower T-stage, the patient's age at diagnosis, and a positive HPV status. USP4 protein levels were not associated with prognosis or other features. CONCLUSION: Since high USP4 mRNA was not an independent prognostic marker, we assume that the association is a result of the correlation of high USP4 mRNA with an HPV-positive status. Therefore, further investigation of USP4 mRNA and its association with the HPV status of HNSCC patients is warranted.

Targeting TGF beta receptor 1 in head and neck squamous cell carcinoma.

OBJECTIVES: The transforming growth factor-Beta (TGF-ß) pathway may be involved in the radioresistance of head and neck squamous cell carcinoma (HNSCC). This study analyzed TGF-ß receptor 1 (TGFBR1) expression in HNSCC patients and evaluated the antineoplastic and radiosensitizing effects of vactosertib, a novel TGFBR1 inhibitor, in vitro. MATERIALS AND METHODS: TGFBR1 expression was examined in HNSCC patients at the mRNA level in silico and the protein level by immunohistochemistry, including surgical specimens of primary tumors, matched lymph node metastasis, and recurrent disease. Furthermore, a novel small molecule TGFBR1 inhibitor was evaluated in HNSCC cell lines. Finally, an indirect coculture model using patient-derived cancer-associated fibroblasts was applied to mimic the tumor microenvironment. RESULTS: Patients with high TGFBR1 mRNA levels showed significantly worse overall survival in silico (OS, p = 0.024). At the protein level, an association between TGFBR1+ tumor and OS was observed for the subgroup with TGFBR1-stroma (p = 0.001). Those results prevailed in multivariable analysis. Inhibition of TGFBR1 showed antineoplastic effects in vitro. In combination with radiation, vactosertib showed synergistic effects. CONCLUSION: Our results indicate a high risk of death in tumorTGFBR1+ |stromaTGFBR1- expressing patients. In vitro data suggest a potential radiosensitizing effect of TGFBR1 inhibition by vactosertib.

ß-CATENIN is a positive prognostic marker for HPV-positive head and neck squamous cell carcinoma.

PURPOSE: The evolutionary-conserved Wnt/ß-CATENIN (WBC) pathway has been implicated in the pathogenesis of different solid malignant tumors. We evaluated the prognostic relevance of ß-CATENIN, a pivotal mediator of WBC activation, in patients with human papillomavirus (HPV)-positive head and neck squamous cell carcinoma (HNSCC). METHODS: We analyzed if patients with HPV-positive HNSCC from the "The Cancer Genome Atlas" (TCGA cohort, n = 41) can be stratified based on their CTNNB1 mRNA expression. Moreover, in a tissue microarray (TMA) of primary tumor sections from HPV-positive HNSCC patients treated in a tertiary academic center (in-house cohort, n = 31), we evaluated the prognostic relevance of ß-CATENIN expression on protein level. RESULTS: In silico mining of CTNNB1 expression in HPV-positive HNSCC revealed that high CTNNB1 expression was linked to better overall survival (OS, p = 0.062). Moreover, high ß-CATENIN expression was significantly associated with a better OS in our in-house cohort (p = 0.035). CONCLUSION: Based on these findings, we postulate that ß-CATENIN expression could serve (potentially in conjunction with other WBC pathway members) as a marker for better survival outcomes in patients with HPV-positive HNSCC. However, it is evident that future studies on bigger cohorts are warranted.

The Geriatric Nutritional Risk Index (GNRI) as a Prognostic Biomarker for Immune Checkpoint Inhibitor Response in Recurrent and/or Metastatic Head and Neck Cancer.

Malnutrition is a frequent comorbidity in head and neck cancer patients and has been shown to impair immunotherapy response in other cancer types. The geriatric nutritional risk index (GNRI) assesses malnutrition using the patient's ideal weight, actual weight, and serum albumin. The aim of this study was to evaluate the prognostic relevance of malnutrition as determined by the GNRI for the response to immunotherapy in recurrent and/or metastatic head and neck squamous cell carcinoma (R/M HNSCC). A total of 162 patients with R/M HNSCC who received immune checkpoint inhibitors were included. The associations between the GNRI and progression-free survival (PFS), overall survival (OS), and the disease control rate (DCR) were computed. Univariable analysis showed worse PFS for GNRI ≤ 98 (p < 0.001), ECOG performance status (PS) ≥ 2 (p = 0.012), and enteral (p = 0.009) and parenteral (p = 0.015) nutritional supplementation, and worse OS for GNRI < 92 (p < 0.001), ECOG PS ≥ 2 (p < 0.001), and enteral (p = 0.008) and parenteral (p = 0.023) nutritional supplementation. In our multivariable model, GNRI ≤ 98 (p = 0.012) and ECOG PS ≥ 2 (p = 0.025) were independent prognostic factors for PFS. For OS, GNRI < 92 (p < 0.001) and ECOG PS ≥ 2 (p < 0.001) were independent prognostic factors. A GNRI ≤ 98 was significantly associated with a lower DCR compared to a GNRI > 98 (p = 0.001). In conclusion, our findings suggest that the GNRI may be an effective predictor for response to immunotherapy in R/M HNSCC.

The prognostic role of PSMD14 in head and neck squamous cell carcinoma.

PURPOSE: PSMD14 is an essential protein for proteasomal degradation. Inhibition of this protein disrupts homeostasis and inhibits cancer cell viability. Overexpression of PSMD14 was associated with advanced cancer characteristics and a worse prognosis in various carcinomas. This study aimed to analyze PSMD14 copy number variation, mRNA and protein expression in HNSCC, and its role as an independent prognostic biomarker. METHODS: PSMD14 mRNA expression and copy number variations were analyzed in "The Cancer Genome Atlas (TCGA)" in 510 patients. Protein expression was evaluated using immunohistochemistry in a second cohort including 115 patients. PSMD14 levels were analyzed for correlation with clinicopathological data, overall and disease-free survival. RESULTS: PSMD14 mRNA expression and copy number variation were high in 44 and 50% of patients, respectively. Protein expression of PSMD14 was high in 56%. In both cohorts, high PSMD14 levels were associated with advanced staging. High PSMD14 mRNA expression was additionally associated with a worse prognosis in univariable analysis. However, after correction for possible confounders, PSMD14 mRNA was not an independent prognostic marker. CONCLUSION: PSMD14 is commonly expressed in HNSCC patients and associated with advanced stages. High expression of PSMD14 mRNA was associated with a worse outcome. However, this may be a result of the association of PSMD14 with poor prognosticators. Based on our study, further evaluation of PSMD14 as a prognostic marker and potential therapeutic target is warranted.

Radiogenomic markers enable risk stratification and inference of mutational pathway states in head and neck cancer.

PURPOSE: Head and neck squamous cell carcinomas (HNSCCs) are a molecularly, histologically, and clinically heterogeneous set of tumors originating from the mucosal epithelium of the oral cavity, pharynx, and larynx. This heterogeneous nature of HNSCC is one of the main contributing factors to the lack of prognostic markers for personalized treatment. The aim of this study was to develop and identify multi-omics markers capable of improved risk stratification in this highly heterogeneous patient population. METHODS: In this retrospective study, we approached this issue by establishing radiogenomics markers to identify high-risk individuals in a cohort of 127 HNSCC patients. Hybrid in vivo imaging and whole-exome sequencing were employed to identify quantitative imaging markers as well as genetic markers on pathway-level prognostic in HNSCC. We investigated the deductibility of the prognostic genetic markers using anatomical and metabolic imaging using positron emission tomography combined with computed tomography. Moreover, we used statistical and machine learning modeling to investigate whether a multi-omics approach can be used to derive prognostic markers for HNSCC. RESULTS: Radiogenomic analysis revealed a significant influence of genetic pathway alterations on imaging markers. A highly prognostic radiogenomic marker based on cellular senescence was identified. Furthermore, the radiogenomic biomarkers designed in this study vastly outperformed the prognostic value of markers derived from genetics and imaging alone. CONCLUSION: Using the identified markers, a clinically meaningful stratification of patients is possible, guiding the identification of high-risk patients and potentially aiding in the development of effective targeted therapies.

Intratympanic Substance Distribution After Injection of Liquid and Thermosensitive Drug Carriers: An Endoscopic Study.

OBJECTIVE: In the treatment of inner ear conditions, intratympanic injection emerges as an important drug delivery method. Novel compounds designed for intratympanic injection are routinely loaded in viscous drug carriers. To date, it is unclear if they can freely distribute in the middle ear. The aims of this study were to investigate the middle ear distribution of different drug carriers during intratympanic injection and to determine an optimal injection method for thermosensitive hydrogels. METHODS: Twenty-one human temporal bones were intratympanically injected with fluid drug carriers or poloxamer-407 hydrogels at different tympanic membrane injection sites (inferior, anterior-superior) using different needle types (Whitacre, Quincke). Fluid distribution was evaluated via an endoscopic view. Injection volume, duration, backflow, and overall safety were analyzed. RESULTS: Liquid drug carriers distribute effortlessly in the middle ear, whereas an additional ventilation hole is advantageous when applying thermosensitive hydrogels. The round window is coated with required volumes between 150 and 200 µl, irrespective of the injection position. Required volumes to also coat the stapedial footplate ranged from 310 to 440 µl. Use of the Whitacre-type needle reduced backflow to the ear canal and enabled longer tympanic membrane visibility when no additional ventilation hole was placed. CONCLUSION: Intratympanic injection is a safe and reliable method for the application of thermosensitive hydrogels. The round window niche is readily filled regardless of the injected formulation and injection position. Although fluid drug carriers distribute effortlessly in the middle ear, the placement of an additional ventilation hole might facilitate the application of viscous hydrogels.

Evaluation of the Prognostic Capacity of a Novel Survival Marker in Patients with Sinonasal Squamous Cell Carcinoma.

Sinonasal squamous cell carcinoma (SNSCC) is a malignant tumor associated with poor survival, and easily obtainable prognostic markers are of high interest. Therefore, we aimed to assess the prognostic value of a novel survival index (SI) combining prognostic values of clinical (T and N classifications and invasion across Ohngren's line), inflammatory (neutrophil-to-lymphocyte ratio), and nutritional (albumin and body-mass index) markers. All patients with primarily treated SNSCC between 2002 and 2020 (n = 51) were included. Each of the six SI components was stratified into a low- (0) and high-risk (1) categories. Subsequently, the cohort was stratified into low- (SI of 0-2) and high-risk SI groups (SI of 3-6). Overall survival (OS) and disease-free survival (DFS) were compared between patients with low- and high-risk SI. The log-rank test was used to test for statistical significance. Overall, the mortality rate was 41.2% (n = 21), and the recurrence rate was 43.1% (n = 22). We observed significantly better OS in patients with low-risk SI (n = 24/51, 47.1%, mean OS: 7.9 years, 95% confidence interval (CI): 6.3-9.6 years) than in high-risk SI (n = 27/51, 52.9%, mean OS: 3.4 years, 95% CI: 2.2-4.5 years; p = 0.013). Moreover, we also showed that patients with low-risk SI had a longer DFS than patients with high-risk SI (mean DFS: 6.4, 95% CI: 4.8-8.0 vs. mean DFS: 2.4 years, 95% CI 1.3-3.5, p = 0.012). The SI combines the prognostic capacity of well-established clinical, radiologic, inflammatory, and nutritional prognosticators and showed prognostic potential in our cohort of SNSCC patients.

Protein Expression of Folate Receptor Alpha in Adenoid Cystic Carcinoma of the Head and Neck.

Purpose: Folate receptor alpha (FRα) is overexpressed in various cancer entities while expression in normal tissue is limited. Thus, FRα is an attractive target in cancer therapy. Currently, various therapeutic and diagnostic approaches are under investigation in clinical trials. The aim of this study was to assess the expression and clinical relevance of FRα in adenoid cystic carcinoma of the head and neck. Patients and Methods: In this retrospective cohort study, 43 patients with adenoid cystic carcinoma (ACC) of the head and neck were included. FRα expression was analyzed in tumor tissue and tumor-free margin in a tissue microarray using immunohistochemical staining. Protein levels were correlated with clinical parameters. Results: FRα staining was positive in 47% of ACC patients. The tumor-free margin was positive in 22%. Patients with positive tumor tissue showed positive margin staining in 55%. FRα expression was not associated with the clinical parameters (sex, age, staging, grading, perineural invasion, lymphovascular invasion). Conclusion: FRα expression is common in ACC of the head and neck. Therefore, FRα should be further evaluated as a therapeutic target in ACC.

Radiosensitizing effect of galunisertib, a TGF-ß receptor I inhibitor, on head and neck squamous cell carcinoma in vitro.

BACKGROUND: Resistance to radiation therapy poses a major clinical problem for patients suffering from head and neck squamous cell carcinoma (HNSCC). Transforming growth factor ß (TGF-ß) has emerged as a potential target. This study aimed to investigate the radiosensitizing effect of galunisertib, a small molecule TGF-ß receptor kinase I inhibitor, on HNSCC cells in vitro. METHODS: Three HNSCC cell lines were treated with galunisertib alone, or in combination with radiation. Of those three cell lines, one has a known inactivating mutation of the TGF-ß pathway (Cal27), one has a TGF-ß pathway deficiency (FaDu) and one has no known alteration (SCC-25). The effect on metabolic activity was evaluated by a resazurin-based reduction assay. Cell migration was evaluated by wound-healing assay, clonogenic survival by colony formation assay and cell cycle by FACS analysis. RESULTS: Galunisertib reduced metabolic activity in FaDu, increased in SCC-25 and had no effect on CAL27. Migration was significantly reduced by galunisertib in all three cell lines and showed additive effects in combination with radiation in CAL27 and SCC-25. Colony-forming capabilities were reduced in SCC-25 by galunisertib and also showed an additive effect with adjuvant radiation treatment. Cell cycle analysis showed a reduction of cells in G1 phase in response to galunisertib treatment. CONCLUSION: Our results indicate a potential antineoplastic effect of galunisertib in HNSCC with intact TGF-ß signaling in combination with radiation.

A novel electrosurgical divider: performance in a self-controlled tonsillectomy study.

PURPOSE: Tonsillectomies are among the most common surgeries in otorhinolaryngology. A novel electrosurgical temperature-controlled instrument (device) promises rapid tonsillectomies and might reduce postoperative pain, but comparative studies to assess performance are warranted. METHODS: This randomized self-controlled clinical trial was conducted from October 2019 to October 2020 at the Department of Otorhinolaryngology, Head and Neck Surgery of the Medical University of Vienna. Forty-eight patients underwent a tonsillectomy with the device on one side and using cold-steel with localized bipolar cauterization on the other side (control). Main outcomes were the time for tonsil removal (per side) and the time to stop bleeding (per side). Secondary measurements were postoperative pain, assessed once on day 0 and five times on days 1, 3, 5, 7, and 10. Postoperative bleeding episodes and consequences were recorded. RESULTS: Device tonsillectomies were performed significantly faster than controls; the mean surgical time difference was 209 s (p < 0.001, 95% CI 129; 288). Intraoperative blood loss was significantly lower on the device side (all p < 0.05). Postoperative measurements of pain and bleeding were similar for both sides. Two return-to-theatre secondary bleeding events were recorded for the control side. CONCLUSION: The novel electrosurgical temperature-controlled divider reduced the tonsillectomy surgical time and intraoperative blood loss, with no apparent negative effects on postoperative pain or bleeding, compared to a cold-steel tonsillectomy with localized bipolar cauterization. In time-restricted settings, the device could be beneficial, particularly after familiarization with device handling. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03793816.

Zerumbone acts as a radiosensitizer in head and neck squamous cell carcinoma.

INTRODUCTION: Zerumbone is a phytochemical compound of the ginger plant Zingiber zerumbet with cytotoxic effects in various cancer cell lines. To date, zerumbone has shown an antiproliferative effect in oral squamous cell carcinoma cells lines. However, the effect of combination with radiation or cisplatin in head and neck squamous cell carcinoma (HNSCC) is unclear. The aim of this study was to investigate the effect of zerumbone alone, and in combination with irradiation and cisplatin on HNSCC cell lines. METHODS: The three HNSCC cell lines SCC25, Cal27 and FaDu were treated with zerumbone, radiation and/or cisplatin. Cell viability and clonogenic assays were performed. The interaction between zerumbone and radiation or cisplatin was evaluated using the combination index. Apoptosis was measured by flow cytometry and cell migration was assessed using a wound healing assay. RESULTS: Treatment with zerumbone resulted in a dose dependent induction of cytotoxicity and apoptosis in all three cell lines. The combination with cisplatin revealed a synergistic to additive effect in Cal27. The clonogenic assay showed a significant radiosensitizing effect in all three cell lines. The wound healing assay showed a reduction of cell migration in Cal27. CONCLUSION: The natural compound zerumbone shows a cytotoxic and proapoptotic effect on HNSCC cell lines. Furthermore, zerumbone enhances the radiation effect in all three cell lines and thus may be a suitable candidate for combination therapy in HNSCC.

µ-Crystallin Is Associated with Disease Outcome in Head and Neck Squamous Cell Carcinoma.

Thyroid hormone levels may be associated with disease outcome in head and neck squamous cell carcinoma (HNSCC). µ-Crystallin (CRYM), a thyroid hormone binding protein, blocks intracellular binding of the thyroid hormone T3 to its receptors. In this study, we aimed to analyze the association of CRYM levels with disease outcome in HNSCC patients. We retrospectively assessed immunohistochemical CRYM expression in 121 head and neck cancer patients. Preoperative thyrotropin levels could be extracted for 50 patients. Patients with low thyrotropin levels had a worse prognosis compared to euthyroid patients (5-year overall survival TSH low 20% vs. TSH norm 58%). We observed an association of CRYM+ patients with improved overall survival (5-year overall survival for CRYM+ 78.6% vs. CRYM- 56%). Interaction analysis between CRYM and HPV revealed that this effect was limited to HPV- patients (CRYM+|HPV- HR 0.12, 95% CI 0.01-0.87, p = 0.036). These results were replicated in an independent dataset. CRYM expression identified patients with favorable disease progression for HPV- HNSCC patients and could serve as a useful biomarker in this patient population. This study further confirms a correlation of thyroid hormone levels with adverse disease outcome in HNSCC patients, which could be potentially exploited as a therapeutic target.

Prognostic Relevance of Thyroid-Hormone-Associated Proteins in Adenoid Cystic Carcinoma of the Head and Neck.

The proteins sodium iodide symporter (NIS), µ-crystallin (CRYM), and thyroid hormone receptor beta (THRB) have been associated with prognosis in various cancer entities. While NIS and THRB may serve as possible therapeutic targets, the role of CRYM in cancer is still unclear. Protein levels of 44 patients with adenoid cystic carcinoma of the head and neck were analyzed using immunohistochemistry and correlated with clinicopathological data and outcome. NIS was positive in 72%, CRYM was positive in 55%, and THRB was positive in 39% of the patients. CRYM-positive adenoid cystic carcinomas were associated with a better cause-specific survival. Thus, our data indicate that CRYM might be a suitable positive prognostic marker in adenoid cystic carcinoma of the head and neck. Furthermore, expression of NIS was present in most patients and therefore evaluation of the use of radioiodine treatment is recommended.

PRKCA Overexpression Is Frequent in Young Oral Tongue Squamous Cell Carcinoma Patients and Is Associated with Poor Prognosis.

Oral tongue squamous cell carcinomas (OTSCCs) have an increasing incidence in young patients, and many have an aggressive course of disease. The objective of this study was to identify candidate prognostic protein markers associated with early-onset OTSCC. We performed an exploratory screening for differential protein expression in younger (≤45 years) versus older (>45 years) OTSCC patients in The Cancer Genome Atlas (TCGA) cohort (n = 97). Expression of candidate markers was then validated in an independent Austrian OTSCC patient group (n = 34) by immunohistochemistry. Kaplan-Meier survival estimates were computed, and genomic and mRNA enrichment in silico analyses were performed. Overexpression of protein kinase C alpha (PRKCA) was significantly more frequent among young patients of both the TCGA (p = 0.0001) and the Austrian cohort (p = 0.02), associated with a negative anamnesis for alcohol consumption (p = 0.009) and tobacco smoking (p = 0.02) and poorer overall survival (univariate p = 0.02, multivariate p< 0.01). Within the young subgroup, both overall and disease-free survival were significantly decreased in patients with PRKCA overexpression (both p < 0.001). TCGA mRNA enrichment analysis revealed 332 mRNAs with significant differential expression in PRKCA-upregulated versus PRKCA-downregulated OTSCC (all FDR ≤ 0.01). Our findings suggest that PRKCA overexpression may be a hallmark of a novel molecular subtype of early-onset alcohol- and tobacco-negative high-risk OTSCC. Further analysis of the molecular PRKCA interactome may decipher the underlying mechanisms of carcinogenesis and clinicopathological behavior of PRKCA-overexpressing OTSCC.

Transcription factors CP2 and YY1 as prognostic markers in head and neck squamous cell carcinoma: analysis of The Cancer Genome Atlas and a second independent cohort.

PURPOSE: The transcription factors YY1 and CP2 have been associated with tumor promotion and suppression in various cancers. Recently, simultaneous expression of both markers was correlated with negative prognosis in cancer. The aim of this study was to explore the expression of YY1 and CP2 in head and neck squamous cell carcinoma (HNSCC) patients and their association with survival. METHODS: First, we analyzed mRNA expression and copy number variations (CNVs) of YY1 and CP2 using "The Cancer Genome Atlas" (TCGA) with 510 HNSCC patients. Secondly, protein expression was investigated via immunohistochemistry in 102 patients, who were treated in the Vienna General Hospital, utilizing a tissue microarray. RESULTS: The median follow-up was 2.9 years (1.8-4.6) for the TCGA cohort and 10.3 years (6.5-12.8) for the inhouse tissue micro-array (TMA) cohort. The median overall survival of the TCGA cohort was decreased for patients with a high YY1 mRNA expression (4.0 vs. 5.7 years, p = 0.030, corr. p = 0.180) and high YY1-CNV (3.53 vs. 5.4 years, p = 0.0355, corr. p = 0.213). Furthermore, patients with a combined high expression of YY1 and CP2 mRNA showed a worse survival (3.5 vs. 5.4 years, p = 0.003, corr. p = 0.018). The mortality rate of patients with co-expression of YY1 and CP2 mRNA was twice as high compared to patients with low expression of one or both (HR 1.99, 95% CI 1.11-3.58, p = 0.021). Protein expression of nuclear YY1 and CP2 showed no association with disease outcome in our inhouse cohort. CONCLUSION: Our data indicate that simultaneous expression of YY1 and CP2 mRNA is associated with shorter overall survival. Thus, combined high mRNA expression might be a suitable prognostic marker for risk stratification in HNSCC patients. However, since we could not validate this finding at genomic or protein level, we hypothesize that unknown underlying mechanisms which regulate mRNA transcription of YY1 and CP2 are the actual culprits leading to a worse survival.

The association between COVID-19 cases and deaths and web-based public inquiries.

BACKGROUND: Corona Virus Disease 2019 (COVID-19) emerged in December 2019 and rapidly spread globally. Since there is still no specific treatment available, prevention of disease spread is crucial to manage the pandemic. Adequate public information is very important. To assess the optimal timing, the aim of this study was to investigate the association between web-based interest and new cases and deaths due to COVID-19. METHODS: Web-based interest for queries related to 'coronavirus' was assessed between 1 January and 19 June 2020, using Google Trends in Australia, Brazil, Canada, Germany, Italy, South Africa, South Korea, Spain, United Kingdom, and the United States of America. Reliability analysis of the used search terms was performed using the intraclass correlation coefficient. To investigate the association between web-based interest and new COVID-19 cases or deaths, the relative search volume was analysed for correlation with new cases and deaths. RESULTS: Reliability analysis revealed excellent reliability for COVID-19 search terms in all countries. Web-based interest peaked between 23 February and 5 April 2020, which was prior to the peak of new infections and deaths in most included countries. There was a moderate to strong correlation between COVID-19 related queries and new cases or new deaths. CONCLUSION: Web-based interest in COVID-19 peaked prior to the peak of new infections and deaths in most countries included. Thus, monitoring public interest via Google Trends might be useful to select the optimal-timing of web-based disease-specific information and preventive measures.