D. Lowe

Promoter methylation of P16, RARβ, E-cadherin, cyclin A1 and cytoglobin in oral cancer: quantitative evaluation using pyrosequencing

Methylation profiling of cancer tissues has identified this mechanism as an important component of carcinogenesis. Epigenetic silencing of tumour suppressor genes through promoter methylation has been investigated by a variety of means, the most recent of which is pyrosequencing. We have investigated quantitative methylation status in oral squamous cell carcinoma patients. Fresh tumour tissue and normal control tissue from resection margin was obtained from 79 consecutive patients undergoing resection of oral squamous cell carcinoma. DNA was extracted and bisulphite treated. PCR primers were designed to amplify 75–200 bp regions of the CpG rich gene promoters of p16, RARβ, E-cadherin, cytoglobin and cyclinA1. Methylation status of 4-5 CpG sites per gene was determined by pyrosequencing. Significant CpG methylation of gene promoters within tumour specimens was found in 28% for p16, 73% for RARβ, 42% for E-cadherin, 65% for cytoglobin and 53% for cyclinA1. Promoter methylation was significantly elevated in tumours compared to normal tissue for p16 (P=0.048), cytoglobin (P=0.002) and cyclin A1 (P=0.001) but not in RARβ (P=0.088) or E-cadherin (P=0.347). Concordant methylation was demonstrated in this tumour series (P=0.03). Significant differences in degree of methylation of individual CpG sites were noted for all genes except RARβ and these differences were in a characteristic pattern that was reproduced between tumour samples. Cyclin A1 promoter methylation showed an inverse trend with histological grade. Promoter methylation analysis using pyrosequencing reveals valuable quantitative data from several CpG sites. In contrast to qualitative data generated from methylation specific PCR, our data demonstrated p16 promoter methylation in a highly tumour specific pattern. Significant tumour specific methylation of cyclin A1 promoter was also seen. Cytoglobin is a novel candidate tumour suppressor gene highly methylated in upper aero-digestive tract squamous cancer.

Issues of intimacy and sexual dysfunction following major head and neck cancer treatment

Problems with sexuality and intimacy in head and neck cancer are under reported. This study set out to describe patients self-reporting of intimacy and sexual dysfunction following treatment and to explore associations with patient characteristics. Patients treated for primary head and neck squamous cell carcinoma, alive and disease free, were identified from the University Hospital Aintree Head and Neck Cancer database, January 2000 to December 2006. A postal survey with two questions from the European Organisation for Research and Treatment of Cancer Quality of Life questionnaire-Head and Neck 35 (EORTC H&N35) regarding sexuality, a self-designed question about intimacy and the University of Washington quality of life questionnaire Version 4 (UW-QOL v4) was sent to patients in March 2007. There was a 68% response (350/518). One-third of those answering the intimacy and sexuality questions reported substantial problems with sexual interest and enjoyment, and one-quarter reported problems with intimacy. Intimacy problems declined with age from 36% if aged under 55 years to 6% for those 75 years and older. Intimacy and sexuality issues were largely unrelated to site, stage of disease, treatment modality and time since surgery. Gender and having a spouse or partner were related to answering the questions but not to having problems. With one-quarter of responders reporting intimacy problems it is surprising how little information and support regarding intimacy and sexual dysfunction is offered to patients and their carers. There is a need to explore these issues more extensively with further research.

The patients' account of outcome following primary surgery for oral and oropharyngeal cancer using a ‘quality of life’ questionnaire

The purpose of this study was to collate all the University of Washington quality of life (UW-QOL) data collected in the Unit since 1995 and to summarize it in a patient-friendly format that can be used when discussing treatment outcomes. The sample of cancer patients consisted of 561 consecutive patients undergoing primary surgery for previously untreated oral and oropharyngeal squamous cell carcinoma presenting to the Regional Maxillofacial Unit Liverpool, between the years 1995 and 2004. Follow-up was on May 2006. Information sheets were constructed based on the UW-QOL, summarizing the patients at around 2 years (median 28 months). The response to each statement in all 12 domains, domains as important, and the general health-related QOL/QOL questions are given. Data has been summarized for six common groups based on the clinical presentation and treatment. The data show big differences by tumour size, free-flap surgery and adjuvant radiotherapy. Information presented in this format is potentially extremely useful when counselling patients and their families regarding the likely outcomes of treatment.

Cytoglobin is upregulated by tumour hypoxia and silenced by promoter hypermethylation in head and neck cancer

Background:Cytoglobin (Cygb) was first described in 2002 as an intracellular globin of unknown function. We have previously shown the downregulation of cytoglobin as a key event in a familial cancer syndrome of the upper aerodigestive tract.Methods:Cytoglobin expression and promoter methylation were investigated in sporadic head and neck squamous cell carcinoma (HNSCC) using a cross-section of clinical samples. Additionally, the putative mechanisms of Cygb expression in cancer were explored by subjecting HNSCC cell lines to hypoxic culture conditions and 5-aza-2-deoxycitidine treatment.Results:In clinically derived HNSCC samples, CYGB mRNA expression showed a striking correlation with tumour hypoxia (measured by HIF1A mRNA expression P=0.013) and consistent associations with histopathological measures of tumour aggression. CYGB expression also showed a marked negative correlation with promoter methylation (P=0.018). In the HNSCC cell lines cultured under hypoxic conditions, a trend of increasing expression of both CYGB and HIF1A with progressive hypoxia was observed. Treatment with 5-aza-2-deoxycitidine dramatically increased CYGB expression in those cell lines with greater baseline promoter methylation.ConclusionWe conclude that the CYGB gene is regulated by both promoter methylation and tumour hypoxia in HNSCC and that increased expression of this gene correlates with clincopathological measures of a tumours biological aggression.

Assessment of problems with appearance, following surgery for oral and oro-pharyngeal cancer using the University of Washington appearance domain and the Derriford appearance scale☆

The first aim of this study was to describe appearance issues in patients following surgery for oral and oro-pharyngeal squamous cell carcinoma using the Derriford appearance scale (DAS24) and the University of Washington Quality of Life Questionnaire Version 4 appearance item (UWQOL v4). Another aim was to compare these two questionnaires and justify a cut-off in the UWQOL. Also the study compares the cancer group to reference data and explores associations with clinical factors. 383 alive and disease free patients and treated between 1992 and 2005 were sent the survey of which 252 (66%) responded. Age (younger patients), T stage 3 and 4 and adjuvant radiotherapy were key factors in patients reporting problems with appearance. There was excellent correlation between the two questionnaires. A cut-off of less than 75 in the UWQOLv4 appearance item captured most of the problems raised in the DAS24. The cancer cohort had similar DAS24 scores to the general population sample and had more positive scores than the clinical reference group (burns etc patient). In conclusion the UWQOL appearance domain appears to be a suitable means of screening for appearance issues in this cohort and could act as a trigger for further assessment and treatment.

MARGINAL MANDIBULAR NERVE INJURY DURING NECK DISSECTION AND ITS IMPACT ON PATIENT PERCEPTION OF APPEARANCE

Neck dissection to remove cervical lymph nodes is common practice in head and neck cancer management. The marginal mandibular nerve may be injured during neck dissection, particularly of level 1. The rate of injury to this nerve is underreported in the literature and its impact on patients is not well defined.

Quantitative promoter methylation differentiates carcinoma ex pleomorphic adenoma from pleomorphic salivary adenoma

Background:Potential epigenetic biomarkers for malignant transformation to carcinoma ex pleomorphic adenoma (Ca ex PSA) have been sought previously with and without specific comparison with the benign variant, pleomorphic salivary adenoma (PSA). Previous analysis has been limited by a non-quantitative approach. We sought to demonstrate quantitative promoter methylation across a panel of tumour suppressor genes (TSGs) in both Ca ex PSA and PSA.Methods:Quantitative methylation-specific real-time polymerase chain reaction (qMSP) analysis of p16INK4A, CYGB, RASSF1, RARβ, human telomerase reverse transcriptase (hTERT), Wilms’ tumour 1 (WT1) and TMEFF2 gene promoters was undertaken on bisulphite-converted DNA, previously extracted from archival fixed tissue specimens of 31 Ca ex PSA and an unrelated cohort of 28 PSA. All target regions examined had formerly been shown to be hypermethylated in salivary and/or mucosal head and neck malignancies.Results:The qMSP demonstrated abnormal methylation of at least one target in 20 out of 31 (64.5%) Ca ex PSA and 2 out of 28 (7.1%) PSA samples (P<0.001). RASSF1 was the single gene promoter for which methylation is shown to be a statistically significant predictor of malignant disease (P<0.001) with a sensitivity of 51.6% and a specificity of 92.9%. RARβ, TMEFF2 and CYGB displayed no apparent methylation, while a combinatory epigenotype based on p16, hTERT, RASSF1 and WT1 was associated with a significantly higher chance of detecting malignancy in any positive sample (odds ratio: 24, 95% CI: 4.7–125, P<0.001).Conclusions:We demonstrate the successful application of qMSP to a large series of historical Ca ex PSA samples and report on a panel of TSGs with significant differences in their methylation profiles between benign and malignant variants of pleomorphic salivary adenoma. qMSP analysis could be developed as a useful clinical tool to differentiate between Ca ex PSA and its benign precursor.

A survey of 14 to 16 year olds as to their attitude toward and use of sunbeds

Objective Sunbeds pose a potential risk to health. Although not recommended for children under 16 years of age they are used by this age group. The purpose of this study was to assess the awareness of suntanning guidelines, the use of sunbeds and the attitude towards tanning in children aged between 14 and 16 years. Design and method 499 pupils aged 14 to 16 years responded to a questionnaire survey in two schools in a mixed urban part of Merseyside. Results Overall, sunbeds had been used by 43 per cent of pupils. Girls had used them much more than boys with use increasing by age for both sexes. Overall, 65 per cent said they were aware of guidelines. About half thought that guidelines advising people how often to use a sunbed were a good thing. One in 10 of those using a sunbed also said they had had problems with their eyes or skin. Most boy and girl users thought using sunbeds made them look healthy, made them more attractive and confident, that they created a base tan before a holiday and that they were a good treatment for spots. Conclusion There is considerable awareness of sunbed guidelines; however, sunbed use in the under 16 year age group remains high. This poses a public health issue and better education and enforcement strategies are required.

Quality of life in older people with oral cancer.

There is a growing elderly population presenting with, and surviving, oral cancer. Making decisions about treatment is complex, and is influenced in part by prognosis and expected outcomes for health-related quality of life (HRQoL). We used the University of Washington Head and Neck Quality of Life scale (UWQoL) to compare HRQoL in patients by age group at a time closest to one year after primary surgery. Survival was analysed using data from the Office for National Statistics. A consecutive series of 1091 patients treated curatively for oral squamous cell carcinoma (SCC) at the regional maxillofacial unit at Aintree University Hospital between 1992 and 2009 were included in the main analysis. UWQoL data for 638 patients were available from about one year after treatment. Older patients (65 years or over) reported better physical and emotional function notably in regard to appearance, speech, saliva (75 years and over), pain, shoulder, mood, and anxiety than younger patients with head and neck cancer treated by operation. In conclusion, older patients seem to cope and adjust well to treatment and this is reflected in their HRQoL scores, which are higher in many domains than those of younger patients with head and neck cancer.

The Patient Concerns Inventory: A Tool to Uncover Unmet Needs in a Cancer Outpatient Clinic

The Patient Concerns Inventory (PCI) is a holistic, patient reported tool that aids in identifying concerns and issues. it is a 55-item checklist of issues that may be grouped into categories of general function, specific head and neck function, emotional-psychological and others, which include finances, treatment related matters and social care. The PCI also gives patients an opportunity to identify from a list of 15 multidisciplinary team (MDT) members whom they would like to see or be referred on to.