Expression and prognostic significance of a panel of tissue hypoxia markers in head-and-neck squamous cell carcinomas

Purpose: To investigate the expression pattern of hypoxia-induced proteins identified as being involved in malignant progression of head-and-neck squamous cell carcinoma (HNSCC) and to determine their relationship to tumor pO 2 and prognosis. Methods and Materials: We performed immunohistochemical staining of hypoxia-induced proteins (carbonic anhydrase IX [CA IX], BNIP3L, connective tissue growth factor, osteopontin, ephrin A1, hypoxia inducible gene-2, dihydrofolate reductase, galectin-1, IκB kinase β, and lysyl oxidase) on tumor tissue arrays of 101 HNSCC patients with pretreatment pO 2 measurements. Analysis of variance and Fisher's exact tests were used to evaluate the relationship between marker expression, tumor pO 2, and CA IX staining. Cox proportional hazard model and log-rank tests were used to determine the relationship between markers and prognosis. Results: Osteopontin expression correlated with tumor pO 2 (Eppendorf measurements) (p = 0.04). However, there was a strong correlation between lysyl oxidase, ephrin A1, and galectin-1 and CA IX staining. These markers also predicted for cancer-specific survival and overall survival on univariate analysis. A hypoxia score of 0-5 was assigned to each patient, on the basis of the presence of strong staining for these markers, whereby a higher score signifies increased marker expression. On multivariate analysis, increasing hypoxia score was an independent prognostic factor for cancer-specific survival (p = 0.015) and was borderline significant for overall survival (p = 0.057) when adjusted for other independent predictors of outcomes (hemoglobin and age). Conclusions: We identified a panel of hypoxia-related tissue markers that correlates with treatment outcomes in HNSCC. Validation of these markers will be needed to determine their utility in identifying patients for hypoxia-targeted therapy. © 2007 Elsevier Inc. All rights reserved.

Head and neck squamous-cell cancer and its association with polymorphic enzymes of xenobiotic metabolism and repair

Tobacco smoking, alcohol drinking, and occupational exposures to polycyclic aromatic hydrocarbons are the major proven risk factors for human head and neck squamous-cell cancer (HNSCC). Major research focus on gene-environment interactions concerning HNSCC has been on genes encoding enzymes of metabolism for tobacco smoke constituents and repair enzymes. To investigate the role of genetically determined individual predispositions in enzymes of xenobiotic metabolism and in repair enzymes under the exogenous risk factor tobacco smoke in the carcinogenesis of HNSCC, we conducted a case-control study on 312 cases and 300 noncancer controls. We focused on the impact of 22 sequence variations in CYP1A1, CYP1B1, CYP2E1, ERCC2/XPD, GSTM1, GSTP1, GSTT1, NAT2, NQO1, and XRCC1. To assess relevant main and interactive effects of polymorphic genes on the susceptibility to HNSCC we used statistical models such as logic regression and a Bayesian version of logic regression. In subgroup analysis of nonsmokers, main effects in ERCC2 (Lys751Gln) C/C genotype and combined ERCC2 (Arg156Arg) C/A and A/A genotypes were predominant. When stratifying for smokers, the data revealed main effects on combined CYP1B1 (Leu432Val) C/G and G/G genotypes, followed by CYP1B1 (Leu432Val) G/G genotype and CYP2E1 (-70G>T) G/T genotype. When fitting logistic regression models including relevant main effects and interactions in smokers, we found relevant associations of CYP1B1 (Leu432Val) C/G genotype and CYP2E1 (-70G>T) G/T genotype (OR, 10.84; 95% CI, 1.64-71.53) as well as CYP1B1 (Leu432Val) G/G genotype and GSTM1 null/null genotype (OR, 11.79; 95% CI, 2.18-63.77) with HNSCC. The findings underline the relevance of genotypes of polymorphic CYP1B1 combined with exposures to tobacco smoke.

Association of CYP1B1 codon 432 mutant allele in head and neck squamous cell cancer is reflected by somatic mutations of p53 in tumor tissue

Tobacco use is causally associated with head and neck squamous cell cancer (HNSCC). Here, we present the results of a case-control study that investigated the effects that the genetic variants of the cytochrome (CYP)1A1, CYP1B1, glutathione-S-transferase (GST)M1, GSTT1, and GSTP1 genes have on modifying the risk of smoking-related HNSCC. Allelisms of the CYP1A1, GSTT1, GSTM1, and GSTT1 genes alone were not associated with an increased risk. CYP1B1 codon 432 polymorphism was found to be a putative susceptibility factor in smoking-related HNSCC. The frequency of CYP1B1 polymorphism was significantly higher (P < 0.001) in the group of smoking cases when compared with smoking controls. Additionally, an odds ratio (OR) of 4.53 (2.62-7.98) was discovered when investigating smoking and nonsmoking cases for the susceptible genotype CYP1B1*2/*2, when compared with the presence of the genotype wild type. In combination with polymorphic variants of the GST genes, a synergistic-effect OR was observed. The calculated OR for the combined genotype CYP1B1*2/*2 and GSTM1*2/*2 was 12.8 (4.09-49.7). The calculated OR for the combined genotype was 13.4 (2.92-97.7) for CYP1B1*2/*2 and GSTT1*2/*2, and 24.1 (9.36-70.5) for the combination of CYP1B1*2/*2 and GSTT1-expressors. The impact of the polymorphic variants of the CYP1B1 gene on HNSCC risk is reflected by the strong association with the frequency of somatic mutations of the p53 gene. Smokers with susceptible genotype CYP1B1*2/*2 were 20 times more likely to show evidence of p53 mutations than were those with CYP1B1 wild type. Combined genotype analysis of CYP1B1 and GSTM1 or GSTT1 revealed interactive effects on the occurrence of p53 gene mutations. The results of the present study indicate that polymorphic variants of CYP1B1 relate significantly to the individual susceptibility of smokers to HNSCC.

Cytochrome p450 1b1, a new keystone in gene-environment interactions related to human head and neck cancer?

Alcohol consumption and tobacco smoking are major causes of head and neck cancers, and regional differences point to the importance of research into gene-environment interactions. Much interest has been focused on polymorphisms of CYP1A1 and of GSTM1 and GSTT1, but a number of studies have not demonstrated significant effects. This has mostly been ascribed to small sample sizes. In general, the impact of polymorphisms of metabolic enzymes appears inconsistent, with some reports of weak-to-moderate associations, and with others of no elevation of risks. The classical cytochrome P450 isoenzyme considered for metabolic activation of polycyclic aromatic hydrocarbons (PAH) is CYP1A1. A new member of the CYP1 family, CYP1B1, was cloned in 1994, currently representing the only member of the CYP1B subfamily. A number of single nucleotide polymorphisms of the CYP1B1 gene have been reported. The amino acid substitutions Val432Leu (CYP1B1*3) and Asn453Ser (CYP1B1*4), located in the heme binding domain of CYP1B1, appear as likely candidates to be linked with biological effects. CYP1B1 activates a wide range of PAH, aromatic and heterocyclic amines. Very recently, the CYP1B1 codon 432 polymorphism (CYP1B1*3) has been identified as a susceptibility factor in smoking-related head-and-neck squamous cell cancer. The impact of this polymorphic variant of CYP1B1 on cancer risk was also reflected by an association with the frequency of somatic mutations of the p53 gene. Combined genotype analysis of CYP1B1 and the glutathione transferases GSTM1 or GSTT1 has pointed to interactive effects. This provides new molecular evidence that tobacco smoke-specific compounds relevant to head and neck carcinogenesis are metabolically activated through CYP1B1 and is consistent with a major pathogenetic relevance of PAH as ingredients of tobacco smoke.

Esophageal and head and neck cancer

Clinical Nutrition for Oncology Patients provides clinicians with the information they need to help cancer survivors and patients make informed choices about their nutrition and improve their short-term and long-term health. This comprehensive resource outlines nutritional management recommendations for care prior to, during, and after treatment and addresses specific nutritional needs and complementary therapies that may be of help to a patient. This book is written by a variety of clinicians who not only care for cancer survivors and their caregivers but are also experts in the field of nutritional oncology.

Long term survival following the detection of circulating tumour cells in head and neck squamous cell carcinoma

Background Techniques for detecting circulating tumor cells in the peripheral blood of patients with head and neck cancers may identify individuals likely to benefit from early systemic treatment. Methods Reconstruction experiments were used to optimise immunomagnetic enrichment and RT-PCR detection of circulating tumor cells using four markers (ELF3, CK19, EGFR and EphB4). This method was then tested in a pilot study using samples from 16 patients with advanced head and neck carcinomas. Results Seven patients were positive for circulating tumour cells both prior to and after surgery, 4 patients were positive prior to but not after surgery, 3 patients were positive after but not prior to surgery and 2 patients were negative. Two patients tested positive for circulating cells but there was no other evidence of tumor spread. Given this patient cohort had mostly advanced disease, as expected the detection of circulating tumour cells was not associated with significant differences in overall or disease free survival. Conclusion For the first time, we show that almost all patients with advanced head and neck cancers have circulating cells at the time of surgery. The clinical application of techniques for detection of spreading disease, such as the immunomagnetic enrichment RT-PCR analysis used in this study, should be explored further.

Quality-of-life among head and neck cancer survivors at one year after treatment – A systematic review

Abstract Background: The importance of quality-of-life (QoL) research has been recognised over the past two decades in patients with head and neck (H&N) cancer. The aims of this systematic review are to evaluate the QoL status of H&N cancer survivors one year after treatment and to identify the determinants affecting their QoL. Methods: Pubmed, Medline, Scopus, Sciencedirect and CINAHL (2000–2011) were searched for relevant studies, and two of the present authors assessed their methodological quality. The characteristics and main findings of the studies were extracted and reported. Results: Thirty-seven studies met the inclusion criteria, and the methodological quality of the majority was moderate to high. While patients of the group in question recover their global QoL by 12 months after treatment, a number of outstanding issues persist – deterioration in physical functioning, fatigue, xerostomia and sticky saliva. Age, cancer site, stage of disease, social support, smoking, feeding tube placement and alcohol consumption are the significant determinants of QoL at 12 months, while gender has little or no influence. Conclusions: Regular assessments should be carried out to monitor physical functioning,degree of fatigue, xerostomia and sticky saliva. Further research is required to develop appropriate and effective interventions to deal with these issues, and thus to promote the patients’ QoL.

Monte Carlo evaluation of collapsed-cone convolution calculations in head and neck radiotherapy treatment plans

The presence of air and bone interfaces makes the dose distribution for head and neck cancer treatments difficult to accurately predict. This study compared planning system dose calculations using the collapsed-cone convolution algorithm with EGSnrcMonte Carlo simulation results obtained using the Monte Carlo DICOMToolKit software, for one oropharynx, two paranasal sinus and three nodal treatment plans. The difference between median doses obtained from the treatment planning and Monte Carlo calculations was found to be greatest in two bilateral treatments: 4.8%for a retropharyngeal node irradiation and 6.7% for an ethmoid paranasal sinus treatment. These deviations in median dose were smaller for two unilateral treatments: 0.8% for an infraclavicular node irradiation and 2.8% for a cervical node treatment. Examination of isodose distributions indicated that the largest deviations between Monte Carlo simulation and collapsed-cone convolution calculations were seen in the bilateral treatments, where the increase in calculated dose beyond air cavities was most significant.

Training program impact on thermoplastic immobilization for head and neck radiation therapy

Purpose: To determine whether uniform guidelines and training in the stabilization and formation of thermoplastic shells can improve the reproducibility of set-up for Head and Neck cancer patients. Methods and materials: Image based measurements of the planning and treatment positions for 35 head and neck cancer patients undergoing radical radiotherapy were analysed to provide a baseline of the reproducibility of thermoplastic immobilization. Radiation therapists (RT) were surveyed to establish a perception of their confidence in thermoplastic procedures. An evidence based staff training program was created and implemented. Set-up reproduction and staff perception were reviewed to measure the impact of the training program. Results: The mean (SD) 3D vectors of anatomical displacement, measured on the patient images, improved from 4.64 (2.03) for the baseline group compared to 3.02 (1.65) following training (p < 0.01). The proportion of 3D displacements of patient data exceeding 5 mm 3D vector was decreased from 37.1% to 5.7% (p < 0.001) and the 3 mm vector from 85.7% to 42.9% (p < 0.001). The post-training survey scores demonstrated improved confidence in reproducibility of set-up for head and neck patients. Conclusion: The Thermoplastic Shells Training Program has been found to improve the treatment reproducibility for head and neck radiation therapy patients. Uniform guidelines have increased RT confidence in thermoplastic procedures.

Adaptive radiotherapy for virally mediated head and neck cancer : predicting which patients may benefit most

Background: Recent clinical studies have demonstrated an emerging subgroup of head and neck cancers that are virally mediated. This disease appears to be a distinct clinical entity with patients presenting younger and with more advanced nodal disease, having lower tobacco and alcohol exposure and highly radiosensitive tumours. This means they are living longer, often with the debilitating functional side effects of treatment. The primary objective of this study was to determine how virally mediated nasopharyngeal and oropharyngeal cancers respond to radiation therapy treatment. The aim was to determine risk categories and corresponding adaptive treatment management strategies to proactively manage these patients. Method/Results: 121 patients with virally mediated, node positive nasopharyngeal or oropharyngeal cancer who received radiotherapy treatment with curative intent between 2005 and 2010 were studied. Relevant patient demographics including age, gender, diagnosis, TNM stage, pre-treatment nodal size and dose delivered was recorded. Each patient’s treatment plan was reviewed to determine if another computed tomography (re-CT) scan was performed and at what time point (dose/fraction) this occurred. The justification for this re-CT was determined using four categories: tumour and/or nodal regression, weight loss, both or other. Patients who underwent a re-CT were further investigated to determine whether a new plan was calculated. If a re-plan was performed, the dosimetric effect was quantified by comparing dose volume histograms of planning target volumes and critical structures from the actual treatment delivered and the original treatment plan. Preliminary results demonstrated that 25/121 (20.7%) patients required a re-CT and that these re-CTs were performed between fractions 20 to 25 of treatment. The justification for these re-CTs consisted of a combination of tumour and/or nodal regression and weight loss. 16/25 (13.2%) patients had a replan calculated. 9 (7.4%) of these replans were implemented clinically due to the resultant dosimetric effect calculated. The data collected from this assessment was statistically analysed to identify the major determining factors for patients to undergo a re-CT and/or replan. Specific factors identified included nodal size and timing of the required intervention (i.e. how when a plan is to be adapted). This data was used to generate specific risk profiles that will form the basis of a biologically guided adaptive treatment management strategy for virally mediated head and neck cancer. Conclusion: Preliminary data indicates that virally mediated head and neck cancers respond significantly during radiation treatment (tumour and/or nodal regression and weight loss). Implications of this response are the potential underdosing or overdosing of tumour and/or surrounding critical structures. This could lead to sub-optimal patient outcomes and compromised quality of life. Consequently, the development of adaptive treatment strategies that improve organ sparing for this patient group is important to ensure delivery of the prescribed dose to the tumour volume whilst minimizing the dose received to surrounding critical structures. This could reduce side effects and improve overall patient quality of life. The risk profiles and associated adaptive treatment approaches developed in this study will be tested prospectively in the clinical setting in Phase 2 of this investigation.

Adaptive radiotherapy for virally mediated head and neck cancer

Introduction: Clinical investigation has revealed a subgroup of head and neck cancers that are virally mediated. The relationship between nasopharyngeal cancer and Epstein Barr Virus (EBV) has long been established and more recently, the association between oropharyngeal cancer and Human Papillomavirus (HPV) has been revealed1,2 These cancers often present with nodal involvement and generally respond well to radiation treatment, evidenced by tumour regression1. This results in the need for treatment plan adaptation or re-planning in a subset of patients. Adaptive techniques allow the target region of the radiotherapy treatment plan to be altered in accordance with treatment-induced changes to ensure that under or over dosing does not occur3. It also assists in limiting potential overdosing of surrounding critical normal tissues4. We sought to identify a high-risk group based on nodal size to be evaluated in a future prospective adaptive radiotherapy trial. Method: Between 2005-2010, 121 patients with virally mediated, node positive nasopharyngeal (EBV positive) or oropharyngeal (HPV positive) cancers, receiving curative intent radiotherapy treatment were reviewed. Patients were analysed based on maximum size of the dominant node at diagnosis with a view to grouping them in varying risk categories to determine the need of re-planning. The frequency and timing of the re-planning scans were also evaluated. Results: Sixteen nasopharyngeal and 105 oropharyngeal tumours were reviewed. Twenty-five (21%) patients underwent a re-planning CT at a median of 22 (range, 0-29) fractions with 1 patient requiring re-planning prior to the commencement of treatment. Based on the analysis, patients were subsequently placed into risk categories; ≤35mm (Group 1), 36-45mm (Group 2), ≥46mm (Group 3). Re-planning CT’s were performed in Group 1- 8/68 (11.8%), Group 2- 4/28 (14.3%), Group 3- 13/25 (52%). Conclusion: In this series, patients with virally mediated head and neck cancer and nodal size > 46mm appear to be a high-risk group for the need of re-planning during a course of curative radiotherapy. This finding will now be tested in a prospective adaptive radiotherapy study. ‘Real World’ Implications: This research identifies predictive factors for those patients with virally mediated head and neck cancer that will benefit most from treatment adaptation. This will assist in minimising the side effects experienced by these patients thereby improving their quality of life after treatment.

Adaptive radiotherapy for virally mediated head and neck cancer

Purpose: Virally mediated head and neck cancers (VMHNC) often present with nodal involvement, and are generally considered radioresponsive, resulting in the need for a re-planning CT during radiotherapy (RT) in a subset of patients. We sought to identify a high-risk group based on nodal size to be evaluated in a future prospective adaptive RT trial. Methodology: Between 2005-2010, 121 patients with virally-mediated, node positive nasopharyngeal (EBV positive) or oropharyngeal (HPV positive) cancers, receiving curative intent RT were reviewed. Patients were analysed based on maximum size of the dominant node with a view to grouping them in varying risk categories for the need of re-planning. The frequency and timing of the re-planning scans were also evaluated. Results: Sixteen nasopharyngeal and 105 oropharyngeal tumours were reviewed. Twenty-five (21%) patients underwent a re-planning CT at a median of 22 (range, 0-29) fractions with 1 patient requiring re-planning prior to the commencement of treatment. Based on the analysis, patients were subsequently placed into 3 groups; ≤35mm (Group 1), 36-45mm (Group 2), ≥46mm (Group 3). Re-planning CT’s were performed in Group 1- 8/68 (11.8%), Group 2- 4/28 (14.3%), Group 3- 13/25 (52%). Sample size did not allow statistical analysis to detect a significant difference or exclusion of a lack of difference between the 3 groups. Conclusion: In this series, patients with VMHNC and nodal size > 46mm appear to be a high-risk group for the need of re-planning during a course of definitive radiotherapy. This finding will now be tested in a prospective adaptive RT study.