TORPEdO: A phase III trial of intensity-modulated proton beam therapy versus intensity-modulated radiotherapy for multi-toxicity reduction in oropharyngeal cancer.

•There is a lack of prospective level I evidence for the use of PBT for most adult cancers including oropharyngeal squamous cell carcinoma (OPSCC).•TORPEdO is the UK's first PBT clinical trial and aims to determine the benefits of PBT for OPSCC.•Training and support has been provided before and during the trial to reduce variations of contouring and radiotherapy planning.•There is a strong translational component within TORPEdO. Imaging and physics data along with blood, tissue collection will inform future studies in refining patient selection for IMPT.

Pre-treatment tumour perfusion parameters and initial RECIST response do not predict long-term survival outcomes for patients with head and neck squamous cell carcinoma treated with induction chemotherapy.

OBJECTIVES: Previously, we showed that pre-treatment tumour plasma perfusion (Fp) predicts RECIST response to induction chemotherapy (ICT) in locoregionally advanced head and neck squamous cell carcinoma (HNSCC). The aim here was to determine whether the pre-treatment tumour Fp estimate, changes in tumour Fp or RECIST response post 2 cycles of ICT were prognostic for long-term survival outcomes. METHODS: A prospective study enrolled patients with high stage HNSCC treated with docetaxel (T), cisplatin (P) and 5-fluorouracil (F) (ICT) followed by synchronous cisplatin and intensity modulated radiotherapy. Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) before and after two cycles of ICT was used to measure Fp and RECIST response. RESULTS: Forty-two patients were recruited and 37 underwent two scans. The median follow-up was 36 (range 23-49) months. Pre-treatment tumour Fp (stratified by median) was not prognostic for overall survival (p = 0.42), disease specific survival (p = 0.20) and locoregional control (p = 0.64). Neither change in tumour Fp nor RECIST response post two cycles of ICT was prognostic for any outcome (p>0.21). CONCLUSION: DCE-MRI parameters do not predict long-term survival outcomes following ICT and RECIST response to ICT may not be an appropriate endpoint to determine early efficacy of a treatment in HNSCC patients.

Taxane, platinum and 5-FU prior to chemoradiotherapy benefits patients with stage IV neck node-positive head and neck cancer and a good performance status.

PURPOSE: The benefit of adding docetaxel, cisplatin and 5-fluorouracil (TPF) induction chemotherapy to chemoradiotherapy (CRT) in head and neck squamous cell carcinoma (HNSCC) remains uncertain. We aimed to investigate whether ICT is well tolerated when given with prophylactic treatment against predicted adverse effects and which patients benefit most. METHODS: A single-centre audit identified 132 HNSCC patients with stage IVa/b neck node-positive disease, prescribed TPF followed by CRT. TPF involved three cycles of docetaxel (75 mg/m2 IV) and cisplatin (75 mg/m2 IV) on day 1 plus 5-FU (750 mg/m2 IV) on days 2-5. Planned CRT was 66 Gy in 30 fractions of intensity-modulated radiotherapy with concurrent cisplatin (100 mg/m2 IV) at the beginning of week 1 and 4 (days 1 and 22). All patients received prophylactic antibiotics and granulocyte colony-stimulating factor. RESULTS: Median follow-up was 39.5 months. 92.4% of patients completed three cycles of TPF; 95.5% of patients started chemoradiotherapy. Grade 3/4 adverse events were low (febrile neutropenia 3.0%), with no toxicity-related deaths. 3-year overall survival was 67.2%; disease-specific survival was 78.7%; locoregional control was 78.3%. Distant metastases rate was 9.8% (3.0% in those without locoregional recurrence). Good performance status (p = 0.002) and poor tumour differentiation (p = 0.018) were associated with improved overall survival on multivariate analysis. CONCLUSION: With prophylactic antibiotics and granulocyte colony-stimulating factor TPF was well tolerated with good survival outcomes. TPF should remain a treatment option for stage IV neck node-positive patients with a good performance status. The use of tumour grade to aid patient selection for TPF warrants investigation.

Role of 18-Fludeoxyglucose positron emission tomography-computed tomography and subsequent panendoscopy in head and neck squamous cell carcinoma of unknown primary.

OBJECTIVES/HYPOTHESIS: Head and neck squamous cell carcinomas of unknown primary (HNSCCUP) accounts for up to 10% of presenting head and neck squamous cell carcinomas. Identification of the primary site allows for directed therapy. Where initial investigations have failed to locate the primary site, 18-fludeoxyglucose positron emission tomography-computed tomography (PET/CT) has emerged as a useful tool with improved sensitivity over positron emission tomography alone. Following PET/CT scan, the role of subsequent panendoscopy ± biopsy has not been fully assessed. We aim to evaluate and quantify the role of PET/CT and subsequent panendoscopy in HNSCCUP. STUDY DESIGN: Retrospective cohort study. METHODS: Patients with HNSCCUP presenting between January 2005 and December 2010 at a regional oncology referral center were studied. All patients who presented with a metastatic neck node and unknown primary who had undergone PET/CT prior to panendoscopy were included. The accuracy of PET/CT was calculated and compared with panendoscopy and histopathological findings. RESULTS: Fifty-two patients were included. Twenty-seven PET/CT scans suggested a primary site. Calculated diagnostic parameters were 83% sensitivity, 87% specificity, positive predictive value 89%, and negative predictive value 80%. Three false-positive PET/CT scans were noted after panendoscopy and normal histology. Importantly, three confirmed tongue base tumors were found on panendoscopy, which were undetected on PET/CT. CONCLUSIONS: PET/CT is a valuable resource for locating tumors in HNSCCUP. It helps direct biopsy and aids in the detection of local and distant metastases along with synchronous primary tumors. Importantly, due to both false-positive and false-negative PET/CT rates, panendoscopy and biopsy remains an essential adjunct investigation irrespective of PET/CT results. LEVEL OF EVIDENCE: 4 Laryngoscope, 126:1354-1358, 2016.

Tumor plasma flow determined by dynamic contrast-enhanced MRI predicts response to induction chemotherapy in head and neck cancer.

OBJECTIVES: Non-response to induction chemotherapy (IC) occurs in 30% of head and neck squamous cell carcinoma (HNSCC) and has been predicted by tumor plasma flow (Fp) derived by perfusion computed tomography. The present study was designed to test whether baseline tumor Fp determined by dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) would predict IC response. MATERIALS AND METHODS: A prospective open study powered to test the relationship between tumor Fp and response to IC (docetaxel, cisplatin, 5-fluorouracil) enrolled 50 patients with stage IV HNSCC. Response after two IC cycles was measured by MRI using Response Evaluation Criteria in Solid Tumors in 37 patients. Tumor Fp (primary end point) and multiple parameters in tumors and lymph nodes (secondary end points) were generated at baseline. Differences in baseline DCE-MRI parameters according to IC response were assessed by the Mann-Whitney U test, and predictive value by receiver operating characteristic (ROC) analysis. RESULTS: Median baseline tumor Fp was 53.2ml/100ml/min in 25 responders and 23.9 in 12 non-responders (U 82; P=0.027; area under ROC curve (AUC) 0.73). Median baseline Fp in lymph nodes was 25.8ml/100ml/min for 37 nodes in 25 responders and 17.1 for 15 nodes in 12 non-responders (U 186, P=0.066; AUC 0.67). Frequency of IC response in 37 patients was 68% overall, 83% for tumor Fp above the median (40.6ml/100ml/min) and 45% below the median. Other DCE-MRI parameters were not associated with IC response. CONCLUSION: Pre-treatment tumor Fp determined by DCE-MRI predicts IC response in HNSCC.

Prognostic value of hypoxia-associated markers in advanced larynx and hypopharynx squamous cell carcinoma.

OBJECTIVES/HYPOTHESIS: To determine the prognostic value of hypoxia-associated markers carbonic anhydrase-9 (CA-9) and hypoxia-inducible factor-1α (HIF-1α) in advanced larynx and hypopharynx squamous cell carcinoma (SCCa) treated by organ preservation strategies. STUDY DESIGN: Retrospective cohort study. METHODS: Pretreatment CA-9 and HIF-1α expression, clinicopathologic data, and tumor volume were analyzed in a series of 114 patients with T3-4 SCCa larynx or hypopharynx treated by (chemo)radiation. RESULTS: Adverse prognostic factors for locoregional control were T4 classification (P = 0.008), and for disease-specific survival were CA-9 positivity (P = 0.039), T4 classification (P = 0.001), larger tumor volume (P = 0.004), N1-3 classification (P = 0.002), and pretreatment hemoglobin < 13.0 g/dl (P = 0.014). With increasing CA-9 expression, there was a trend to increasing tumor recurrence (P trend = 0.009) and decreasing survival (P trend = 0.002). On multivariate analysis, independent variables were T4 classification (hazard ratio [HR] 13.54, P = 0.01) for locoregional failure, and CA-9 positivity (HR = 8.02, P = 0.042) and higher tumor volume (HR = 3.33, P = 0.007) for disease-specific mortality. CONCLUSION: This is the first study to look specifically at T3 and T4 SCCa larynx and hypopharynx for a relationship between hypoxia-associated marker expression and clinical outcome. Pretreatment immunohistochemical CA-9 expression is an adverse prognostic factor for disease-specific survival, indicating that CA-9 expression may confer a more aggressive tumor phenotype.

Dynamic contrast-enhanced magnetic resonance imaging biomarkers in head and neck cancer: potential to guide treatment? A systematic review.

Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) generates microvascular parameters from the tracer kinetic analysis of a series of MRI images obtained in under 15 min. DCE-MRI parameters are associated with tumour hypoxia, which is well-established as a cause of treatment failure in head and neck squamous cell carcinoma (HNSCC). A systematic review was conducted of prospective DCE-MRI parameter studies in HNSCC in the English language literature. Exclusion criteria were case reports and retrospective series. Six DCE-MRI marker studies in HNSCC met the inclusion criteria. Four studies contained 21-74 patients and two studies recruited 13 and 14 patients. In studies measuring the transfer coefficient (K(trans)), higher overall K(trans) or lower skewness of K(trans) were predictive of a good outcome following chemoradiation. DCE-MRI parameters have the potential to guide treatment in HNSCC. Progress in the field requires standardisation of methods, data sharing and large multi-centre collaborative validation studies.

A 26-gene hypoxia signature predicts benefit from hypoxia-modifying therapy in laryngeal cancer but not bladder cancer.

PURPOSE: Tumor hypoxia is associated with a poor prognosis, hypoxia modification improves outcome, and hypoxic status predicts benefit from treatment. Yet, there is no universal measure of clinical hypoxia. The aim of this study was to investigate whether a 26-gene hypoxia signature predicted benefit from hypoxia-modifying treatment in both cancer types. EXPERIMENTAL DESIGN: Samples were available from 157 T2-T4 laryngeal cancer and 185 T1-T4a bladder cancer patients enrolled on the accelerated radiotherapy with carbogen and nicotinamide (ARCON) and bladder carbogen nicotinamide (BCON) phase III randomized trials of radiotherapy alone or with carbogen and nicotinamide (CON) respectively. Customized TaqMan low density arrays (TLDA) were used to assess expression of the 26-gene signature using quantitative real-time PCR. The median expression of the 26 genes was used to derive a hypoxia score (HS). Patients were categorized as TLDA-HS low (≤median) or TLDA-HS high (>median). The primary outcome measures were regional control (RC; ARCON) and overall survival (BCON). RESULTS: Laryngeal tumors categorized as TLDA-HS high showed greater benefit from ARCON than TLDA-HS low tumors. Five-year RC was 81% (radiotherapy alone) versus 100% (CON) for TLDA-HS high (P=0.009). For TLDA-HS low, 5-year RC was 91% (radiotherapy alone) versus 90% (CON; P=0.90). TLDA-HS did not predict benefit from CON in bladder cancer. CONCLUSION: The 26-gene hypoxia signature predicts benefit from hypoxia-modifying treatment in laryngeal cancer. These findings will be evaluated in a prospective clinical trial.

Prospective technical validation and assessment of intra-tumour heterogeneity of a low density array hypoxia gene profile in head and neck squamous cell carcinoma.

BACKGROUND AND PURPOSE: Tumour hypoxia is associated with a poor prognosis in head and neck squamous cell carcinoma (HNSCC), however there is no accepted method for assessing hypoxia clinically. We aimed to conduct a technical validation of a hypoxia gene expression signature using the TaqMan Low Density Array (TLDA) platform to investigate if this approach reliably identified hypoxic tumours. MATERIALS AND METHODS: Tumour samples (n=201) from 80 HNSCC patients were collected prospectively from two centres. Fifty-three patients received pimonidazole prior to surgery. TaqMan Low Density Array-Hypoxia Scores (TLDA-HS) were obtained by quantitative real-time PCR (qPCR) using a 25-gene signature and customised TLDA cards. Assay performance was assessed as coefficient of variation (CoV). RESULTS: The assay was sensitive with linear reaction efficiencies across a 4 log(10) range of inputted cDNA (0.001-10 ng/µl). Intra- (CoV=6.9%) and inter- (CoV=2.0%) assay reproducibility were excellent. Intra-tumour heterogeneity was lower for TLDA-HS (23.2%) than for pimonidazole (67.2%) or single gene measurements of CA9 (62.2%), VEGFA (45.0%) or HIG2 (39.4%). TLDA-HS in HNSCC cell lines increased with decreasing pO(2). TLDA-HS correlated with Affymetrix U133 Plus 2.0 microarray HS (p<0.01) and positive pimonidazole scores (p=0.005). CONCLUSIONS: Gene expression measurements of hypoxia using a 25-gene signature and TLDA cards are sensitive, reproducible and associated with lower intra-tumour heterogeneity than assaying individual genes or pimonidazole binding. The approach is suitable for further assessment of prognostic and predictive capability in clinical trial material.

Molecular and cellular processes underlying the hallmarks of head and neck cancer.

The hallmarks of cancer were updated by Hanahan and Weinberg in 2011. Here we discuss the updated hallmarks in relation to what is known of the molecular and cellular processes underlying the development of head and neck squamous cell carcinoma (HNSCC). Several mechanisms are described, and recent surveys of HNSCC suggest a limited number of mutations, from which more mechanisms may emerge. There are also epigenetic changes to the control of normal processes. More than one mechanism underlies each hallmark. Processes essential to the development of HNSCC need not be essential to the proliferation of the fully developed tumour. Attention is paid to the emerging hallmarks, deregulation of cellular energy metabolism and evasion of immune destruction, and enabling characteristics, genome instability and mutation and tumour-promoting inflammation. HNSCC may adapt to hypoxia, suppress HLA expression, and express Toll-like receptors to facilitate inflammation, which support the proliferation of the tumour.

Perfusion estimated with rapid dynamic contrast-enhanced magnetic resonance imaging correlates inversely with vascular endothelial growth factor expression and pimonidazole staining in head-and-neck cancer: a pilot study.

PURPOSE: To analyze, in a pilot study, rapidly acquired dynamic contrast-enhanced (DCE)-MRI data with a general two-compartment exchange tracer kinetic model and correlate parameters obtained with measurements of hypoxia and vascular endothelial growth factor (VEGF) expression in patients with squamous cell carcinoma of the head and neck. METHODS AND MATERIALS: Eight patients were scanned before surgery. The DCE-MRI data were acquired with 1.5-s temporal resolution and analyzed using the two-compartment exchange tracer kinetic model to obtain estimates of parameters including perfusion and permeability surface area. Twelve to 16 h before surgery, patients received an intravenous injection of pimonidazole. Samples taken during surgery were used to determine the level of pimonidazole staining using immunohistochemistry and VEGF expression using quantitative real-time polymerase chain reaction. Correlations between the biological and imaging data were examined. RESULTS: Of the seven tumors fully analyzed, those that were poorly perfused tended to have high levels of pimonidazole staining (r = -0.79, p = 0.03) and VEGF expression (r = -0.82, p = 0.02). Tumors with low permeability surface area also tended to have high levels of hypoxia (r = -0.75, p = 0.05). Hypoxic tumors also expressed higher levels of VEGF (r = 0.82, p = 0.02). CONCLUSIONS: Estimates of perfusion obtained with rapid DCE-MRI data in patients with head-and-neck cancer correlate inversely with pimonidazole staining and VEGF expression.

Mucosal melanoma of the head and neck: radiotherapy or surgery?

INTRODUCTION: Head and neck mucosal melanoma (MuM) is rare, comprising < 1% of all melanomas in Western Europe. METHODS: A retrospective analysis of case records of patients treated between 1965 and 2001 was carried out. (Survival outcomes were obtained from the case notes and cancer registry.) The median age of the 68 patients was 63 years (range 29-86 years). Thirty-nine percent were male, and 61% were female. (The minimum follow-up time was 15 months.) The two most common primary sites were the sinonasal complex (65%) and oral cavity (19%). Twenty-one percent of patients presented with metastases (nodal or distant). Fifty-five patients were treated with curative intent: 30 patients with primary radiotherapy and 25 patients with surgery +/- postoperative radiotherapy. RESULTS: The overall survival was 22% at 5 years, and the cancer-specific survival was 32% at 5 years. CONCLUSION: MuM has a poor overall prognosis. Poor prognostic indicators are site at presentation and presentation with metastasis. This series is unique in that a significant proportion of patients were given primary radiotherapy as definitive treatment. Surgery may have advantages, particularly for oral cavity MuM. In contrast to previous reports, definitive radiotherapy is worthy of consideration as curative treatment.

Effect of temporal bone resection on temporomandibular joint function: a quality of life study.

OBJECTIVES: Temporal bone resection for carcinoma may affect quality of life (QOL) and result in temporomandibular joint (TMJ) disorders. The aims of this study were to 1) assess TMJ function after temporal bone resection, and 2) assess the impact of TMJ dysfunction on QOL. STUDY DESIGN: Chart and patient review and QOL study. SETTING: Tertiary referral center (Manchester Royal Infirmary). SUBJECTS AND METHODS: Thirty patients who had undergone temporal bone resection were identified. Thirteen patients were alive and were included in the study. All patients were submitted to a clinical examination to identify TMJ disorders and determine facial nerve function, and they all answered the University of Washington QOL (UW-QOL) questionnaire (version 4). RESULTS: Eight patients had TMJ disorders with reduced mobility in either direction and/or significant pain. Main factors affecting QOL were pain (P = 0.001), appearance (P = 0.001), and anxiety (P = 0.000). Neither facial nerve palsy nor TMJ disorders affected QOL. The responses to the UW-QOL questionnaire showed that 69 percent of our patients had a good QOL. A total of 61.53 percent of our patients had TMJ dysfunction presenting as restriction of jaw mobility with or without pain. CONCLUSION: TMJ dysfunction is present in a significant number of patients after temporal bone resection, resulting in longstanding problems, which should be addressed accordingly. Poor QOL results from ongoing pain and psycho-social disturbance.

Exon array analysis of head and neck cancers identifies a hypoxia related splice variant of LAMA3 associated with a poor prognosis.

The identification of alternatively spliced transcript variants specific to particular biological processes in tumours should increase our understanding of cancer. Hypoxia is an important factor in cancer biology, and associated splice variants may present new markers to help with planning treatment. A method was developed to analyse alternative splicing in exon array data, using probeset multiplicity to identify genes with changes in expression across their loci, and a combination of the splicing index and a new metric based on the variation of reliability weighted fold changes to detect changes in the splicing patterns. The approach was validated on a cancer/normal sample dataset in which alternative splicing events had been confirmed using RT-PCR. We then analysed ten head and neck squamous cell carcinomas using exon arrays and identified differentially expressed splice variants in five samples with high versus five with low levels of hypoxia-associated genes. The analysis identified a splice variant of LAMA3 (Laminin alpha 3), LAMA3-A, known to be involved in tumour cell invasion and progression. The full-length transcript of the gene (LAMA3-B) did not appear to be hypoxia-associated. The results were confirmed using qualitative RT-PCR. In a series of 59 prospectively collected head and neck tumours, expression of LAMA3-A had prognostic significance whereas LAMA3-B did not. This work illustrates the potential for alternatively spliced transcripts to act as biomarkers of disease prognosis with improved specificity for particular tissues or conditions over assays which do not discriminate between splice variants.

Potential risk factors associated with the use of cidofovir to treat benign human papillomavirus-related disease.

BACKGROUND: Cidofovir is currently being used off-licence to treat different viral infections, such as benign low-risk human papillomavirus (HPV)-related recurrent respiratory papillomatosis (RRP). There are concerns over the safety of this practice as rat studies demonstrated a high malignant transformation rate. As yet, there are no clinical reports of cidofovir-induced malignant changes in humans. METHODS: Telomerase immortalised human keratinocytes (hTert) stably expressing E6 proteins from either low-risk HPV6b or high-risk HPV16 and vector control cells were treated with either low-dose (5 microg/ml) or higher dose (30 microg/ml) cidofovir for 2 days and the effects evaluated by clonogenic survival assays. Based on these results, gene expression microarray analysis was performed on cidofovir-treated low-risk E6 and vector cells before, during and after drug treatment, and the results verified by real-time PCR. RESULTS: Both low-risk and high-risk E6-expressing cells show significantly improved long-term survival compared with vector control cells when exposed to 5 microg/ml cidofovir for 2 days, (hTert T6E6 P=0.0007, hTert T16E6 P=0.00023 and hTert vector control P=0.62). Microarray and real-time PCR analyses of low-dose cidofovir-treated low-risk E6-expressing cells revealed changes in gene expression that are known to be associated with malignant progression, which were not observed in drug-treated vector control cells. CONCLUSIONS: This is the first report that cidofovir can both increase cell survival and induce alterations in gene expression that are known to be associated with malignant transformation in cells transduced only with the E6 gene from low-risk HPV. It is our belief that these data provide cause for concern over the off-license use of this drug to treat RRP.

Shoulder morbidity after pectoralis major flap reconstruction for head and neck cancer.

BACKGROUND: The effect of pectoralis major flap (PMF) harvest on shoulder function, allowing for the effects of neck dissection, has not previously been objectively measured. METHODS: Twenty-two patients who underwent PMF reconstruction were studied. The control group comprised 35 patients with neck dissection (without PMF). Neck dissections in both groups were classified into 3 grades; grade 1: no neck dissection/selective neck dissection; grade 2: modified radical neck dissection; grade 3: radical neck dissection/extended radical neck dissection. Objective shoulder assessments were carried out using Constant score. RESULTS: Constant score deteriorated with grade of neck dissection (p < .005). The median Constant score for PMF group and neck dissection only group were 82 and 90, respectively (p = .40). Subgroup analysis within neck dissection grade did not show any significant difference, but the effect of PMF was noted to be greatest in grade 2 patients (p = .064). CONCLUSIONS: There is minimal or low shoulder morbidity, additional to neck dissection, caused by PMF reconstruction in head and neck surgery.

Cidofovir induces an increase in levels of low-risk and high-risk HPV E6.

BACKGROUND: Cidofovir is a nucleoside analogue that is used off-license to treat recurrent respiratory papillomatosis (RRP) caused by HPV6/11. However, the effect of this drug upon low-risk HPV 6/11 gene expression is unknown. METHODS: The expression of E6 was evaluated by RT-PCR in HPV-ve C33A cervical carcinoma cells stably transfected with both low- and high-risk HPV E6 cDNA's and in SiHa (HPV16+ve) cervical carcinoma cells after treatment with 2 doses and durations of exposure to cidofovir. RESULTS: Compared to the vector only transcript, E6 RNA levels showed an 8-fold increase in low-risk and 20-fold increase in high-risk E6-expressing cells. High-risk E6 protein levels were also detected by Western blot in cidofovir-treated C33A Type16 E6-transfected cells. CONCLUSION: These data may indicate a potential rationale for increased risk of genetic instability and thus transformation due to drug-induced increase in the level of E6.