Matt Lechner

Human Papillomavirus Drives Tumor Development Throughout the Head and Neck: Improved Prognosis Is Associated With an Immune Response Largely Restricted to the Oropharynx

Purpose In squamous cell carcinomas of the head and neck (HNSCC), the increasing incidence of oropharyngeal squamous cell carcinomas (OPSCCs) is attributable to human papillomavirus (HPV) infection. Despite commonly presenting at late stage, HPV-driven OPSCCs are associated with improved prognosis compared with HPV-negative disease. HPV DNA is also detectable in nonoropharyngeal (non-OPSCC), but its pathogenic role and clinical significance are unclear. The objectives of this study were to determine whether HPV plays a causal role in non-OPSCC and to investigate whether HPV confers a survival benefit in these tumors. Methods Meta-analysis was used to build a cross-tissue gene-expression signature for HPV-driven cancer. Classifiers trained by machine-learning approaches were used to predict the HPV status of 520 HNSCCs profiled by The Cancer Genome Atlas project. DNA methylation data were similarly used to classify 464 HNSCCs and these analyses were integrated with genomic, histopathology, and survival data to permit a comprehensive comparison of HPV transcript-positive OPSCC and non-OPSCC. Results HPV-driven tumors accounted for 4.1% of non-OPSCCs. Regardless of anatomic site, HPV+ HNSCCs shared highly similar gene expression and DNA methylation profiles; nonkeratinizing, basaloid histopathological features; and lack of TP53 or CDKN2A alterations. Improved overall survival, however, was largely restricted to HPV-driven OPSCCs, which were associated with increased levels of tumor-infiltrating lymphocytes compared with HPV-driven non-OPSCCs. Conclusion Our analysis identified a causal role for HPV in transcript-positive non-OPSCCs throughout the head and neck. Notably, however, HPV-driven non-OPSCCs display a distinct immune microenvironment and clinical behavior compared with HPV-driven OPSCCs.

Deregulation of SYCP2 predicts early stage human papillomavirus-positive oropharyngeal carcinoma: A prospective whole transcriptome analysis.

This study was designed to identify significant differences in gene expression profiles of human papillomavirus (HPV)‐positive and HPV‐negative oropharyngeal squamous cell carcinomas (OPSCC) and to better understand the functional and biological effects of HPV infection in the premalignant pathway. Twenty‐four consecutive patients with locally advanced primary OPSCC were included in a prospective clinical trial. Fresh tissue samples (tumor vs. matched normal epithelium) were subjected to whole transcriptome analysis and the results validated on the same cohort with RT–quantitative real‐time PCR. In a separate retrospective cohort of 27 OPSCC patients, laser capture microdissection of formalin‐fixed, paraffin‐embedded tissue allowed RNA extraction from adjacent regions of normal epithelium, carcinoma in situ (premalignant) and invasive SCC tissue. The majority of patients showed evidence of high‐risk HPV16 positivity (80.4%). Predictable fold changes of RNA expression in HPV‐associated disease included multiple transcripts within the p53 oncogenic pathway (e.g. CDKN2A/CCND1). Other candidate transcripts found to have altered levels of expression in this study have not previously been established (SFRP1, CRCT1, DLG2, SYCP2, and CRNN). Of these, SYCP2 showed the most consistent fold change from baseline in premalignant tissue; aberrant expression of this protein may contribute to genetic instability during HPV‐associated cancer development. If further corroborated, this data may contribute to the development of a non‐invasive screening tool. This study is registered with the UK Clinical Research Network (ref.: 11945).

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.

Exploring the implications of HPV infection for head and neck cancer

Over recent years, human papillomavirus (HPV) has been shown to be a major risk factor for head and neck squamous cell cancer (HNSCC), and particularly oropharyngeal squamous cell carcinoma.1 ,2 In 2007, the International Agency for Research on Cancer recognised HPV as a carcinogen associated with malignant transformation in this subset of head and neck cancers. It is now well established that HPV-induced oropharyngeal cancers and those caused by other factors (such as smoking and alcohol abuse—a combination of heavy smoking and drinking leads to an almost 50-fold increased risk of oral and pharyngeal cancer3) are two separate entities, with distinct aetiologies, clinical characteristics, prognoses and a different epidemiology and molecular basis.4 The incidence of HPV-associated HNSCC has risen rapidly in the Western world over the past 40 years.5–8 For example, there has been an estimated threefold increase in tonsillar cancer during this period,9 and the overall estimated population-level incidence of HPV-positive oropharyngeal cancer, which was 2.6 per 100 000 in the USA in 2004,6 is set to triple again in the next 20 years.10 Interestingly, the overall incidence of HNSCC is falling at a time when the incidence of HPV-induced cancer has risen, with the result that the proportion of HPV-positive tonsillar cases has risen from <25% in the 1970s to 93% of cases by 2007 in parts of the developed world.7 …

Advancing the grading for drug-induced sleep endoscopy—a useful modification of the Croft-Pringle Grading System

Dear Editor, Drug-induced sleep endoscopy (DISE) has become a primary tool for the adjunctive evaluation and management of obstructive sleep apnoea (OSA) [1]. The anatomical cause for OSA is heterogeneous, and meticulous assessment of the airway during DISE helps to identify areas and levels of collapse, thereby allowing the appropriate choice of treatment. Croft and Pringle have pioneered this field and presented the first grading system for DISE in 1993, based on 90 cases [2]. Now many years later and based on our original retrospective audit [3] and further more than 5000 cases, we would like to share useful modifications of the Croft and Pringle grading system which we believe will help the surgeon to make a more informed choice on available medical and surgical management options—in line with the global need to further advance the field of DISE [4]. Firstly, the Kotecha/Lechner modification describes three different anatomical levels for grade 3 (herewith referred to as multisegmental collapse on inspiration), i.e. at the level of the nasopharynx, oropharynx and tongue base. If any of these levels contribute more than 20% to the overall collapse/ obstruction (overall obstruction being 100%), then the respective level needs to be addressed specifically, e.g. if surgery is planned. This is of particular importance for the assessment of the oropharynx and the surgeon must distinguish between lateral oropharyngeal wall collapse, circumferential collapse and antero-posterior collapse, as this also influences the surgical management, e.g. the consideration of expansion sphincter pharyngoplasty (ESP) for lateral wall collapse [5] or upper airway stimulation surgery [6]. With regard to Croft and Pringle grade 5, we suggest two further elaborations: firstly, document evidence of epiglottic retraction as a result of bulky base of tongue abutting and pushing the epiglottis towards the posterior pharyngeal wall and obscuring the glosso-epiglottic fold, and secondly, the so-called trap door phenomenon (posterior displacement of the epiglottis as a result of negative intra-thoracic pressure towards the posterior pharyngeal wall and obstruction at this level during inspiration ± expiration) which we regard to be only significant, if this is observedmore than ten times on the roll. In this latter case, the respective patient may be a candidate for laser epiglottic wedge resection ± radiofrequency interstitial thermal therapy to the tongue base. Prominence at the level of the tongue base is often caused by lymphoid hyperplasia, leading to or worsening the obstruction described above. The degree of prominence is also noted to guide the aggressiveness of the surgical technique required. In addition to the above, we believe that the most useful modification or adjunct to the Croft and Pringle grading system is the jaw-lift manoeuvre. Gentle pressure (approximately 4–5-mm protrusion) is applied to the chin from postero-inferiorly, thereby achieving mouth closure with forward movement of the lower jaw/mandible. During this manoeuvre the flexible fibre-optic nasoendoscope is sited in the nasopharynx, assessing the upper airway for both improvement of patency of the airway and decrease of snoring. We found this to be an excellent indicator for whether the respective patient will gain benefit from a trial with a mandibular advancement splint (MAS) or not. It is important not to apply too much pressure/forward movement of the mandible during the jaw-lift manoeuvre, as the assessment will stimulate the patient and then may not be representative for a mandibular advancement splint selection as a treatment option. The patient also needs to be assessed for nasal pathologies during DISE, including septal deviation, hypertrophy of the inferior Electronic supplementary material The online version of this article (https://doi.org/10.1007/s11325-017-1611-5) contains supplementary material, which is available to authorized users.

Pan-cancer deconvolution of cellular composition identifies molecular correlates of antitumour immunity and checkpoint blockade response

The nature and extent of immune cell infiltration into solid tumours are key determinants of therapeutic response. Here, using a novel DNA methylation-based approach to tumour cell fraction deconvolution, we report the integrated analysis of tumour composition and genomics across a wide spectrum of solid cancers. Initially studying head and neck squamous cell carcinoma, we identify two distinct tumour subgroups: ‘immune hot’ and ‘immune cold’, which display differing prognosis, mutation burden, cytokine signalling, cytolytic activity, and oncogenic driver events. We demonstrate the existence of such tumour subgroups pan-cancer, link clonal-neoantigen burden to hot tumours, and show that transcriptional signatures of hot tumours are selectively engaged in immunotherapy responders. We also find that treatment-naive hot tumours are markedly enriched for known immune-resistance genomic alterations and define a catalogue of novel and known mediators of active antitumour immunity, deriving biomarkers and potential targets for precision immunotherapy.

A review on drug-induced sedation endoscopy – Technique, grading systems and controversies

Sleep disordered breathing (SDB) comprises a spectrum of disorders, ranging from simple snoring to severe obstructive sleep apnoea (OSA), with a significant burden to health care systems in high income countries. If left untreated, OSA has significant cumulative, long-term health consequences. In the 1990s drug induced sedation endoscopy (DISE) has been developed to become a primary tool in the diagnosis and management of OSA. It allows meticulous endoscopic evaluation of the airway and identifies areas of collapse, thereby informing both on the selection of surgical techniques, where efficacy depends entirely on success at relieving obstruction at a certain level and on the usefulness of conservative measures, such as mandibular advancement splints. This article provides a review of the literature on DISE, covering different grading systems and techniques, explaining different rationales and discussing controversies.

Frequent HPV-independent p16/INK4A overexpression in head and neck cancer

OBJECTIVES p16INK4A (p16) is the most widely used clinical biomarker for Human Papillomavirus (HPV) in head and neck squamous cell cancer (HNSCC). HPV is a favourable prognostic marker in HNSCC and is used for patient stratification. While p16 is a relatively accurate marker for HPV within the oropharynx, recent reports suggest it may be unsuitable for use in other HNSCC subsites, where a smaller proportion of tumors are HPV-driven. MATERIALS AND METHODS We integrated reverse phase protein array (RPPA) data for p16 with HPV status based on detection of viral transcripts by RNA-seq in a set of 210 HNSCCs profiled by The Cancer Genome Atlas project. Samples were queried for alterations in CDKN2A, and other pathway genes to investigate possible drivers of p16 expression. RESULTS While p16 levels as measured by RPPA were significantly different by HPV status, there were multiple HPV (-) samples with similar expression levels of p16 to HPV (+) samples, particularly at non-oropharyngeal subsites. In many cases, p16 overexpression in HPV (-) tumors could not be explained by mutation or amplification of CDKN2A or by RB1 mutation. Instead, we observed enrichment for inactivating mutations in the histone H3 lysine 36 methyltransferase, NSD1 in HPV (-)/p16-high tumors. CONCLUSIONS RPPA data suggest high p16 protein expression in many HPV (-) non-oropharyngeal HNSCCs, limiting its potential utility as an HPV biomarker outside of the oropharynx. HPV-independent overexpression of wild-type p16 in non-oropharyngeal HNSCC may be linked to global deregulation of chromatin state by inactivating mutations in NSD1.

Pan-cancer deconvolution of tumour composition using DNA methylation

The nature and extent of immune cell infiltration into solid tumours are key determinants of therapeutic response. Here, using a DNA methylation-based approach to tumour cell fraction deconvolution, we report the integrated analysis of tumour composition and genomics across a wide spectrum of solid cancers. Initially studying head and neck squamous cell carcinoma, we identify two distinct tumour subgroups: ‘immune hot’ and ‘immune cold’, which display differing prognosis, mutation burden, cytokine signalling, cytolytic activity and oncogenic driver events. We demonstrate the existence of such tumour subgroups pan-cancer, link clonal-neoantigen burden to cytotoxic T-lymphocyte infiltration, and show that transcriptional signatures of hot tumours are selectively engaged in immunotherapy responders. We also find that treatment-naive hot tumours are markedly enriched for known immune-resistance genomic alterations, potentially explaining the heterogeneity of immunotherapy response and prognosis seen within this group. Finally, we define a catalogue of mediators of active antitumour immunity, deriving candidate biomarkers and potential targets for precision immunotherapy.Determining the extent of immune cell infiltration into solid tumours is essential for adequate therapeutic response. Here the authors develop a DNA methylation-based approach for tumour cell fraction deconvolution and analyse tumour composition and genomics across a wide spectrum of solid cancers.

A cross-sectional survey of awareness of human papillomavirus-associated oropharyngeal cancers among general practitioners in the UK

Objectives To examine the level of awareness of the link between human papillomavirus (HPV) and oropharyngeal cancer (OPC) and epidemiological trends in HPV-related OPC among general practitioners (GPs) in the UK. Design Cross-sectional survey. Participants 384 GPs from England, Scotland, Wales and Northern Ireland. Setting The survey was administered at GP training courses and via email to lists of training course attendees. Primary and secondary outcome measures Proportion of respondents aware of the link between HPV and OPC; respondents’ self-rated knowledge of OPC; proportion of participants aware of the epidemiological trends in HPV-associated OPC. Results 384 questionnaires were completed with an overall response rate of 72.9%. 74.0% of participants recognised HPV as a risk factor for OPC, which was lower than knowledge about the role of smoking, chewing tobacco and alcohol consumption (all >90% recognition). Overall, 19.4% rated their knowledge of OPC as very good or good, 62.7% as average and 17.7% as poor or very poor. The majority (71.9%) were aware that rates of HPV-associated OPC have increased over the last two decades. Fewer than half (41.5%) of the participants correctly identified being male as a risk factor of HPV-associated OPC, while 58.8% were aware that patients with HPV-associated OPC tend to be younger than those with non-HPV-associated disease. Conclusions The association of HPV infection with OPC is a relatively recent discovery. Although the level of awareness of HPV and OPC among GPs was high, the characteristics of HPV-associated OPC were less well recognised, indicating the need for further education.