Unilateral Choanal Atresia: Indications of Long-Term Olfactory Deficits and Volumetric Brain Changes Postsurgically.

BACKGROUND: Very few studies have investigated whether unilateral choanal atresia is associated with permanent olfactory deficits. OBJECTIVE: This study aimed to evaluate the olfactory performance of patients with unilateral choanal atresia postsurgically. METHODS: Three patients with unilateral atresia were examined in terms of olfactory performance with the Sniffin' Sticks test (odor identification, threshold, and discrimination), size of the olfactory bulb, and volumetric brain changes. RESULTS: All patients demonstrated significantly lower olfactory performance in terms of odor threshold on the same side with the choanal atresia. Grey matter reductions were found ipsilaterally in the hippocampus. CONCLUSIONS: This pilot study indicates that persistent olfactory deficits and volumetric brain changes are present in patients with unilateral choanal atresia.

Metabolomics, machine learning and immunohistochemistry to predict succinate dehydrogenase mutational status in phaeochromocytomas and paragangliomas.

Phaeochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine tumours with a hereditary background in over one-third of patients. Mutations in succinate dehydrogenase (SDH) genes increase the risk for PPGLs and several other tumours. Mutations in subunit B (SDHB) in particular are a risk factor for metastatic disease, further highlighting the importance of identifying SDHx mutations for patient management. Genetic variants of unknown significance, where implications for the patient and family members are unclear, are a problem for interpretation. For such cases, reliable methods for evaluating protein functionality are required. Immunohistochemistry for SDHB (SDHB-IHC) is the method of choice but does not assess functionality at the enzymatic level. Liquid chromatography-mass spectrometry-based measurements of metabolite precursors and products of enzymatic reactions provide an alternative method. Here, we compare SDHB-IHC with metabolite profiling in 189 tumours from 187 PPGL patients. Besides evaluating succinate:fumarate ratios (SFRs), machine learning algorithms were developed to establish predictive models for interpreting metabolite data. Metabolite profiling showed higher diagnostic specificity compared to SDHB-IHC (99.2% versus 92.5%, p = 0.021), whereas sensitivity was comparable. Application of machine learning algorithms to metabolite profiles improved predictive ability over that of the SFR, in particular for hard-to-interpret cases of head and neck paragangliomas (AUC 0.9821 versus 0.9613, p = 0.044). Importantly, the combination of metabolite profiling with SDHB-IHC has complementary utility, as SDHB-IHC correctly classified all but one of the false negatives from metabolite profiling strategies, while metabolite profiling correctly classified all but one of the false negatives/positives from SDHB-IHC. From 186 tumours with confirmed status of SDHx variant pathogenicity, the combination of the two methods resulted in 185 correct predictions, highlighting the benefits of both strategies for patient management. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Gustatory pathway in humans: A review of models of taste perception and their potential lateralization.

The focus of this review is to interpret recent advances in human gustatory pathways with respect to the laterality of gustatory responses. Psychophysical, neuroimaging, and clinical anatomical studies published in peer reviewed scientific journals were examined. From the anatomical and neuroimaging studies a total of six models are outlined and discussed in the light of some recent psychophysical and clinical results. In the specific of the salt condition and right preferred hand the outcomes have revealed a predominant left ipsilateral pathway with evidences of ipsilateral projection from the left primary gustatory cortex (PGC) to the orbitofrontal cortex, while a bilateral projection from the right oral cavity to the left and right insula seems to be more consistent. Also, the right side predominance of the chemosensory perception is objected. Additionally, the gustatory response appears to be dependent on the taste quality, supporting the idea of a chemotopical organization of the PGC as well as the Labelled-Line Model theory of peripheral taste quality encoding. However, where the fibers branch along the ascending pathway is not unequivocally established. Interestingly, factors like handedness appear to be remarkable when studying the lateralization of brain functions. Finally we suggest that further studies must include handedness and taste quality as distinctive factors that can help to interpret the results in a unique way.

Metabolome-guided genomics to identify pathogenic variants in isocitrate dehydrogenase, fumarate hydratase, and succinate dehydrogenase genes in pheochromocytoma and paraganglioma.

PURPOSE: Metabolic aberrations have been described in neoplasms with pathogenic variants (PV) in the Krebs cycle genes encoding succinate dehydrogenase (SDH), fumarate hydratase (FH) and isocitrate dehydrogenase (IDH). In turn, accumulation of oncometabolites succinate, fumarate, and 2-hydroxyglutarate can be employed to identify tumors with those PV . Additionally, such metabolic readouts may aid in genetic variant interpretation and improve diagnostics. METHODS: Using liquid chromatography-mass spectrometry, 395 pheochromocytomas and paragangliomas (PPGLs) from 391 patients were screened for metabolites to indicate Krebs cycle aberrations. Multigene panel sequencing was applied to detect driver PV in cases with indicative metabolite profiles but undetermined genetic drivers. RESULTS: Aberrant Krebs cycle metabolomes identified rare cases of PPGLs with germline PV in FH and somatic PV in IDHx and SDHx, including the first case of a somatic IDH2 PV in PPGL. Metabolomics also reliably identified PPGLs with SDHx loss-of-function (LOF) PV. Therefore we utilized tumor metabolite profiles to further classify variants of unknown significance in SDHx, thereby enabling missense variants associated with SDHx LOF to be distinguished from benign variants. CONCLUSION: We propose incorporation of metabolome data into the diagnostics algorithm in PPGLs to guide genetic testing and variant interpretation and to help identify rare cases with PV in FH and IDHx.

Correction to: DNA methylation-based reclassification of olfactory neuroblastoma.

In the original publication, the second name of the twentieth author was incorrect. It should read as 'Miguel Sáinz-Jaspeado'. The original publication of the article has been updated to reflect the change. This correction was authored by Ulrich Schüller on behalf of all authors of the original publication.

DNA methylation-based reclassification of olfactory neuroblastoma.

Olfactory neuroblastoma/esthesioneuroblastoma (ONB) is an uncommon neuroectodermal neoplasm thought to arise from the olfactory epithelium. Little is known about its molecular pathogenesis. For this study, a retrospective cohort of n = 66 tumor samples with the institutional diagnosis of ONB was analyzed by immunohistochemistry, genome-wide DNA methylation profiling, copy number analysis, and in a subset, next-generation panel sequencing of 560 tumor-associated genes. DNA methylation profiles were compared to those of relevant differential diagnoses of ONB. Unsupervised hierarchical clustering analysis of DNA methylation data revealed four subgroups among institutionally diagnosed ONB. The largest group (n = 42, 64%, Core ONB) presented with classical ONB histology and no overlap with other classes upon methylation profiling-based t-distributed stochastic neighbor embedding (t-SNE) analysis. A second DNA methylation group (n = 7, 11%) with CpG island methylator phenotype (CIMP) consisted of cases with strong expression of cytokeratin, no or scarce chromogranin A expression and IDH2 hotspot mutation in all cases. T-SNE analysis clustered these cases together with sinonasal carcinoma with IDH2 mutation. Four cases (6%) formed a small group characterized by an overall high level of DNA methylation, but without CIMP. The fourth group consisted of 13 cases that had heterogeneous DNA methylation profiles and strong cytokeratin expression in most cases. In t-SNE analysis, these cases mostly grouped among sinonasal adenocarcinoma, squamous cell carcinoma, and undifferentiated carcinoma. Copy number analysis indicated highly recurrent chromosomal changes among Core ONB with a high frequency of combined loss of chromosome 1-4, 8-10, and 12. NGS sequencing did not reveal highly recurrent mutations in ONB, with the only recurrently mutated genes being TP53 and DNMT3A. In conclusion, we demonstrate that institutionally diagnosed ONB are a heterogeneous group of tumors. Expression of cytokeratin, chromogranin A, the mutational status of IDH2 as well as DNA methylation patterns may greatly aid in the precise classification of ONB.

Testicular cancer patients undergoing cisplatin based chemotherapy exhibit temporary olfactory threshold scores changes.

Testicular cancer is the most frequent malignant disease in young males between 15 and 35 years. Platinum based chemotherapy regimen is the therapy of choice in advanced disease. This treatment has also adverse effects caused by the cytostatic active substances, such as olfactory dysfunctions. The aim of this study was, therefore, to monitor olfactory function of testicular cancer patients during and 6 months after chemotherapy. A total of 17 patients (mean age 31.06 ± 10.26 years), which underwent chemotherapy (mean 2.47 cycles ± 0.5) were enrolled in this study. Odor threshold, discrimination and identification were assessed by means of the "Sniffin' Sticks" prior to and on day 42, 90 and 180 after chemotherapy has been completed. Furthermore, patients' ratings of olfactory function and depressive symptoms were evaluated. Threshold scores were significantly lower on day 90 (8.0 ± 2.51) compared to baseline (10.4 ± 2.20) (p = 0.014) and recovered almost completely on day 180 (9.65 ± 3.26). Odor discrimination and identification did not show significant changes during therapy. The decrease of the olfactory function during/immediately after chemotherapy was underlined by the subjectively perceived impaired olfactory function during this time. In addition almost every fourth patient presented with a depressed mood at the beginning of chemotherapy. Patients should be informed about possible transient olfactory impairment during/immediately after chemotherapy.

The administration of nasal drops in the "Kaiteki" position allows for delivery of the drug to the olfactory cleft: a pilot study in healthy subjects.

Systemic treatment with corticosteroids shows therapeutic effects, few patients benefit from intranasal topical drug application, probably due to limited access of the drug to the olfactory epithelium. The aim of the present study was to investigate how drops distribute within the nasal cavity when the "Kaiteki" position is performed. Thirteen healthy volunteers participated. Subjects were lying on the side with the head tilted and the chin turned upward. Blue liquid was used to visualize the intranasal distribution of the nasal drops. The investigation was carried out using photo documentation thorough nasal endoscopy; the intranasal distribution of the dye was judged by two independent observers in both a decongested state and a natural state where no decongestants had been used. With regard to the main criterion of this study, using the "Kaiteki" position, nasal drops reached the olfactory cleft in 96 % of the decongested cases and 75 % of the cases who had not been decongested. However, this difference was not statistically different. Because the "Kaiteki" maneuver is not too difficult to perform, it is more likely that topical steroids can be helpful in cases of olfactory loss.

Spatial distribution of FMISO in head and neck squamous cell carcinomas during radio-chemotherapy and its correlation to pattern of failure.

BACKGROUND: Tumour hypoxia can be measured by FMISO-PET and negatively impacts local tumour control in patients with head and neck squamous cell carcinoma (HNSCC) undergoing radiotherapy. The aim of this post hoc analysis of a prospective clinical trial was to investigate the spatial variability of FMISO hypoxic subvolumes during radio-chemotherapy and the co-localisation of these volumes with later recurrences as a basis for individualised dose prescription trials with dose escalation defined by FMISO-PET. METHODS: Sequential FMISO scans of 12 (of 25) patients presenting residual hypoxia taken before (FMISOpre) and during (FMISOw1-FMISOw5) radio-chemotherapy were analysed regarding the stability of the FMISO subvolumes and, in case of local failure, their correlation to local relapse. RESULTS: Consecutive FMISO-PET positive volumes could be classified as moderately stable with Dice conformity indices of 62% and 58% up to the second week of treatment. Substantial volumetric variation during treatment was observed, with more than 20% geographic miss in all patients and more than 40% in half of the patients. The localisation of the maximum standardised uptake value (SUVmax) differed with a mean distance of 7.0 mm and 13.5 mm between the pre-therapeutic and first or second FMISO-PET during treatment. A stable hypoxic consensual volume (i.e. overlap of pre-therapeutic FMISO and intra-treatment FMISO subvolumes up to week two, generated by different contouring methods) was determined for six patients with imaging information of local recurrence. Three of these six local recurrences were located within this consensual volume. CONCLUSIONS: Our data suggest that selective dose painting to hypoxic tumour subvolumes requires adaptation during treatment and sufficient margins. An alternative strategy is to escalate the dose to the gross tumour volume, accepting lesser escalation of dose outside hypoxic areas if indicated by constraints for organs at risk.

The impact and prospect of traumatic brain injury on olfactory function: a cross-sectional and prospective study.

Traumatic brain injury (TBI) can cause olfactory loss. The aim of this cross-sectional and prospective study was to determine the prevalence of olfactory loss among 110 patients with TBI within 3 months after the trauma. In 81 patients ("cross-sectional"-group), olfactory function could be measured using the validated "Sniffin' Sticks" test for odor threshold and odor identification. In addition, the prospective change of olfactory function was studied in 36 patients ("follow-up"-group) by means of a validated odor threshold, discrimination and identification test. Olfactory function was significantly better in patients with TBI I° compared to individuals with TBI II° and III°. Clinically significant improvement of olfactory function was found in 36% of the patients, most frequently during the first 6 months after the injury, in a median follow-up interval of 21 months. TBI I° has in general no major effect on olfaction. In contrast, patients with TBI II° and III° exhibit smell loss in 57%. Chances for olfactory recovery were highest within the first 6 months after the trauma.

The value of subcutaneous xenografts for individualised radiotherapy in HNSCC: Robust gene signature correlates with radiotherapy outcome in patients and xenografts.

PURPOSE: To assess the robustness of prognostic biomarkers and molecular tumour subtypes developed for patients with head and neck squamous cell carcinoma (HNSCC) on cell-line derived HNSCC xenograft models, and to develop a novel biomarker signature by combining xenograft and patient datasets. MATERIALS AND METHODS: Mice bearing xenografts (n = 59) of ten HNSCC cell lines and a retrospective, multicentre patient cohort (n = 242) of the German Cancer Consortium-Radiation Oncology Group (DKTK-ROG) were included. All patients received postoperative radiochemotherapy (PORT-C). Gene expression analysis was conducted using GeneChip Human Transcriptome Arrays. Xenografts were stratified based on their molecular subtypes and previously established gene classifiers. The dose to control 50 % of tumours (TCD50) was compared between these groups. Using differential gene expression analyses combining xenograft and patient data, a gene signature was developed to define risk groups for the primary endpoint loco-regional control (LRC). RESULTS: Tumours of mesenchymal subtype were characterized by a higher TCD50 (xenografts, p < 0.001) and lower LRC (patients, p < 0.001) compared to the other subtypes. Similar to previously published patient data, hypoxia- and radioresistance-related gene signatures were associated with high TCD50 values. A 2-gene signature (FN1, SERPINE1) was developed that was prognostic for TCD50 (xenografts, p < 0.001) and for patient outcome in independent validation (LRC: p = 0.007). CONCLUSION: Genetic prognosticators of outcome for patients after PORT-C and subcutaneous xenografts after primary clinically relevant irradiation show similarity. The identified robust 2-gene signature may help to guide patient stratification, after prospective validation. Thus, xenografts remain a valuable resource for translational research towards the development of individualized radiotherapy.

PET/MRI in head and neck cancer: initial experience.

PURPOSE: To evaluate the feasibility of PET/MRI (positron emission tomography/magnetic resonance imaging) with FDG ((18)F-fluorodeoxyglucose) for initial staging of head and neck cancer. METHODS: The study group comprised 20 patients (16 men, 4 women) aged between 52 and 81 years (median 64 years) with histologically proven squamous cell carcinoma of the head and neck region. The patients underwent a PET scan on a conventional scanner and a subsequent PET/MRI examination on a whole-body hybrid system. FDG was administered intravenously prior to the conventional PET scan (267-395 MBq FDG, 348 MBq on average). The maximum standardized uptake values (SUV(max)) of the tumour and of both cerebellar hemispheres were determined for both PET datasets. The numbers of lymph nodes with increased FDG uptake were compared between the two PET datasets. RESULTS: No MRI-induced artefacts where observed in the PET images. The tumour was detected by PET/MRI in 17 of the 20 patients, by PET in 16 and by MRI in 14. The PET/MRI examination yielded significantly higher SUV(max) than the conventional PET scanner for both the tumour (p < 0.0001) and the cerebellum (p = 0.0009). The number of lymph nodes with increased FDG uptake detected using the PET dataset from the PET/MRI system was significantly higher the number detected by the stand-alone PET system (64 vs. 39, p = 0.001). CONCLUSION: The current study demonstrated that PET/MRI of the whole head and neck region is feasible with a whole-body PET/MRI system without impairment of PET or MR image quality.

Intrinsic chemosensory signal recorded from the human nasal mucosa in patients with smell loss.

Physiological investigation of olfactory receptor function in hyposmic or anosmic patients is rare. Pioneers examined the electro-olfactogram in patients with olfactory disturbance. Although the electro-olfactogram is an established method to record olfactory responses from human olfactory epithelium, the response is only measured at specific sites of the olfactory mucosa. In contrast to that the response of the olfactory epithelium to chemosensory stimuli can be studied in a specific nasal area by means of intrinsic optical signal recording. Five functionally anosmic patients were included in the present study. In all patients, responses could be obtained following trigeminal stimulation with CO2. In some patients, responses could be obtained after olfactory stimulation with H2S and PEA. The present data show that in the studied patients trigeminal function seems to be preserved, while it appears that in some patients olfactory function is preserved to a certain degree.

Influence of background noise on the performance in the odor sensitivity task: effects of noise type and extraversion.

Recent research demonstrated that background noise relative to silence impaired subjects' performance in a cognitively driven odor discrimination test. The current study aimed to investigate whether the background noise can also modulate performance in an odor sensitivity task that is less cognitively loaded. Previous studies have shown that the effect of background noise on task performance can be different in relation to degree of extraversion and/or type of noise. Accordingly, we wanted to examine whether the influence of background noise on the odor sensitivity task can be altered as a function of the type of background noise (i.e., nonverbal vs. verbal noise) and the degree of extraversion (i.e., introvert vs. extrovert group). Subjects were asked to conduct an odor sensitivity task in the presence of either nonverbal noise (e.g., party sound) or verbal noise (e.g., audio book), or silence. Overall, the subjects' mean performance in the odor sensitivity task was not significantly different across three auditory conditions. However, with regard to the odor sensitivity task, a significant interaction emerged between the type of background noise and the degree of extraversion. Specifically, verbal noise relative to silence significantly impaired or improved the performance of the odor sensitivity task in the introvert or extrovert group, respectively; the differential effect of introversion/extraversion was not observed in the nonverbal noise-induced task performance. In conclusion, our findings provide new empirical evidence that type of background noise and degree of extraversion play an important role in modulating the effect of background noise on subjects' performance in an odor sensitivity task.

Improvement of chronic rhinitis under aspirin.

In a subset of aspirin-tolerant asthmatics, administration of aspirin improves respiratory symptoms. We present a patient with chronic rhinosinusitis with nasal polyps who exhibited relief of nasal obstruction and nasal discharge and improvement in the sense of smell following oral administration of 150 mg of aspirin daily. Improvement in the patency of the nasal passages was documented by nasal endoscopy and magnetic resonance imaging. Improvement of olfactory function was documented by validated psychophysical olfactory testing and by means of olfactory event-related potentials.

Head/neck paragangliomas: focus on tumor location, mutational status and plasma methoxytyramine.

Head and neck paragangliomas (HNPGLs) are tumors of parasympathetic origin that occur at variable locations and are often secondary to germline mutations in succinate dehydrogenase (SDH) subunit genes. Occasionally, these tumors produce catecholamines. Here, we assessed whether different locations of HNPGLs relate to the presence of SDHx mutations, catecholamine production and other presentations. In this multicenter study, we collected clinical and biochemical data from 244 patients with HNPGLs and 71 patients without HNPGLs. We clarified that jugulotympanic HNPGLs have distinct features. In particular, 88% of jugulotympanic HNPGLs arose in women, among whom only 24% occurred due to SDHx mutations compared to 55% in men. Jugulotympanic HNPGLs were also rarely bilateral, were of a smaller size and were less often metastatic compared to carotid body and vagal HNPGLs. Furthermore, we showed that plasma concentrations of methoxytyramine (MTY) were higher (P < 0.0001) in patients with HNPGL than without HNPGL, whereas plasma normetanephrine did not differ. Only 3.7% of patients showed strong increases in plasma normetanephrine. Plasma MTY was positively related to tumor size but did not relate to the presence of SDHx mutations or tumor location. Our findings confirm that increases in plasma MTY represent the main catecholamine-related biochemical feature of patients with HNPGLs. We expect that more sensitive analytical methods will make biochemical testing of HNPGLs more practical in the future and enable more than the current 30% of patients to be identified with dopamine-producing HNPGLs. The sex-dependent differences in the development of HNPGLs may have relevance to the diagnosis, management and outcomes of these tumors.

Biomarker signatures for primary radiochemotherapy of locally advanced HNSCC - Hypothesis generation on a multicentre cohort of the DKTK-ROG.

PURPOSE: To develop prognostic biomarker signatures for patients with locally advanced head and neck squamous cell carcinoma (HNSCC) treated by primary radiochemotherapy (RCTx) based on previously published molecular analyses of the retrospective biomarker study of the German Cancer Consortium - Radiation Oncology Group (DKTK-ROG). MATERIAL AND METHODS: In previous studies on the retrospective DKTK-ROG HNSCC cohort treated with primary RCTx, the following clinical parameters and biomarkers were evaluated and found to be significantly associated with loco-regional tumour control (LRC) or overall survival (OS): tumour volume, p16 status, expression of cancer stem cell markers CD44 and SLC3A2, expressions of hypoxia-associated gene signatures, tumour mutational burden (TMB), single nucleotide polymorphisms (SNPs) in the ERCC2 gene (rs1799793, rs13181) and ERCC5 gene (rs17655) as well as the expression of CXCR4, SDF-1 and CD8. These biomarkers were combined in multivariable modelling using Cox-regression with backward variable selection. RESULTS: A baseline signature containing the widely accepted parameters tumour volume, p16 status, cancer stem cell marker expression (CD44), and hypoxia-associated gene expression has been defined, representing the main hypothesis of the study. Furthermore, the baseline signature was extended by additional prognostic biomarkers and a data-driven signature without any pre-hypothesis was generated for both endpoints. In these signatures, the SNPs rs1799793 and rs17655 as well as CXCR4, SDF-1 and SLC3A2 expression were additionally included. The signatures showed significant patient stratifications for LRC and OS. CONCLUSION: Three biomarker signatures were defined for patients with locally advanced HNSCC treated with primary RCTx for the endpoints LRC and OS. These signatures will be validated in the prospective HNprädBio study of the DKTK-ROG that recently completed recruitment, before potential application in an interventional trial.

Development and validation of a 6-gene signature for the prognosis of loco-regional control in patients with HPV-negative locally advanced HNSCC treated by postoperative radio(chemo)therapy.

PURPOSE: The aim of this study was to develop and validate a novel gene signature from full-transcriptome data using machine-learning approaches to predict loco-regional control (LRC) of patients with human papilloma virus (HPV)-negative locally advanced head and neck squamous cell carcinoma (HNSCC), who received postoperative radio(chemo)therapy (PORT-C). MATERIALS AND METHODS: Gene expression analysis was performed using Affymetrix GeneChip Human Transcriptome Array 2.0 on a multicentre retrospective training cohort of 128 patients and an independent validation cohort of 114 patients from the German Cancer Consortium - Radiation Oncology Group (DKTK-ROG). Genes were filtered based on differential gene expression analyses and Cox regression. The identified gene signature was combined with clinical parameters and with previously identified genes related to stem cells and hypoxia. Technical validation was performed using nanoString technology. RESULTS: We identified a 6-gene signature consisting of four individual genes CAV1, GPX8, IGLV3-25, TGFBI, and one metagene combining the highly correlated genes INHBA and SERPINE1. This signature was prognostic for LRC on the training data (ci = 0.84) and in validation (ci = 0.63) with a significant patient stratification into two risk groups (p = 0.005). Combining the 6-gene signature with the clinical parameters T stage and tumour localisation as well as the cancer stem cell marker CD44 and the 15-gene hypoxia-associated signature improved the validation performance (ci = 0.69, p = 0.001). CONCLUSION: We have developed and validated a novel prognostic 6-gene signature for LRC of HNSCC patients with HPV-negative tumours treated by PORT-C. After successful prospective validation the signature can be part of clinical trials on the individualization of radiotherapy.

Analyses of molecular subtypes and their association to mechanisms of radioresistance in patients with HPV-negative HNSCC treated by postoperative radiochemotherapy.

PURPOSE: To assess the relation of the previously reported classification of molecular subtypes to the outcome of patients with HNSCC treated with postoperative radio(chemo)therapy (PORT-C), and to assess the association of these subtypes with gene expressions reflecting known mechanisms of radioresistance. MATERIAL AND METHODS: Gene expression analyses were performed using the GeneChip Human Transcriptome Array 2.0 on a multicentre retrospective patient cohort (N = 128) of the German Cancer Consortium Radiation Oncology Group (DKTK-ROG) with locally advanced HNSCC treated with PORT-C. Tumours were assigned to four molecular subtypes, and correlation analyses between subtypes and clinical risk factors were performed. In addition, the classifications of eight genes or gene signatures related to mechanisms of radioresistance, which have previously shown an association with outcome of patients with HNSCC, were compared between the molecular subtypes. The endpoints loco-regional control (LRC) and overall survival (OS) were evaluated by log-rank tests and Cox regression. RESULTS: Tumours were classified into the four subtypes basal (19.5%), mesenchymal (18.8%), atypical (15.6%) and classical (14.1%). The remaining tumours could not be classified (32.0%). Tumours of the mesenchymal subtype showed a lower LRC compared to the other subtypes (p = 0.012). These tumours were associated with increased epithelial-mesenchymal transition (EMT) and overexpression of a gene signature enriched in DNA repair genes. The majority of the eight considered gene classifiers were significantly associated to LRC or OS in the whole cohort. CONCLUSION: Molecular subtypes, previously identified on HNSCC patients treated with primary radio(chemo)therapy or surgery, were related to LRC for patients treated with PORT-C, where mesenchymal tumours presented with worse prognosis. After prospective validation, subtype-based patient stratification, potentially in combination with other molecular classifiers, may be considered in future interventional studies in the context of personalised radiotherapy and may guide the development of combined treatment approaches.

A Novel 2-Metagene Signature to Identify High-Risk HNSCC Patients amongst Those Who Are Clinically at Intermediate Risk and Are Treated with PORT.

(1) Background: Patients with locally advanced head and neck squamous cell carcinoma (HNSCC) who are biologically at high risk for the development of loco−regional recurrences after postoperative radiotherapy (PORT) but at intermediate risk according to clinical risk factors may benefit from additional concurrent chemotherapy. In this matched-pair study, we aimed to identify a corresponding predictive gene signature. (2) Methods: Gene expression analysis was performed on a multicenter retrospective cohort of 221 patients that were treated with postoperative radiochemotherapy (PORT-C) and 283 patients who were treated with PORT alone. Propensity score analysis was used to identify matched patient pairs from both cohorts. From differential gene expression analysis and Cox regression, a predictive gene signature was identified. (3) Results: 108 matched patient pairs were selected. We identified a 2-metagene signature that stratified patients into risk groups in both cohorts. The comparison of the high-risk patients between the two types of treatment showed higher loco−regional control (LRC) after treatment with PORT-C (p < 0.001), which was confirmed by a significant interaction term in Cox regression (p = 0.027), i.e., the 2-metagene signature was indicative for the type of treatment. (4) Conclusion: We have identified a novel gene signature that may be helpful to identify patients with high-risk HNSCC amongst those at intermediate clinical risk treated with PORT, who may benefit from additional concurrent chemotherapy.