Evaluation and application of a broad-spectrum polymerase chain reaction assay for human papillomaviruses in the screening of squamous cell tumours of the head and neck

OBJECTIVE: The results of studies analysing the prevalence of human papillomaviruses (HPVs) in squamous cell carcinomas of the head and neck region often vary depending on the different molecular biological methods applied. Focusing on the histomorphological criteria of HPV infections, the percentage of HPV-positive cancers should be higher than has been found in most studies. The aim of this study was to increase the detection spectrum of HPV polymerase chain reaction (PCR) screening using degenerate consensus primers. MATERIAL AND METHODS: To increase the sensitivity of the assay to one copy of HPV DNA per cell, a two-step PCR was carried out. The products were directly sequenced by means of a cycle sequencing approach. Seventy biopsies from squamous cell carcinomas of the upper aerodigestive tract were screened using the primer system. RESULTS: According to the PCR results, 0/2 carcinomas of the floor of the mouth or tongue, 7/16 biopsies of oropharyngeal cancers, 3/13 hypopharyngeal cancers, 13/34 squamous cell carcinomas of the larynx, 4/4 biopsies from carcinomas of the nasal cavity or paranasal sinuses and 1/1 squamous cell carcinoma of the parotid gland were positive. CONCLUSION: This broad-spectrum PCR can effectively detect HPV in head and neck cancers.     

The changing demographics of head and neck squamous cell carcinoma in the United States

OBJECTIVES/HYPOTHESIS: Head and neck squamous cell carcinoma (HNSCCA) has declined in the United States since the late 1970s. During this time, substantial immigration from other countries has occurred, and the average lifespan has increased. We tested the hypothesis that these trends have altered the HNSCCA patient population. STUDY DESIGN: Retrospective analysis was made of population-based data from the SEER database, a national registry capturing roughly 10% of all U.S. cancer diagnoses. METHODS: We examined all unique diagnoses of HNSCCA in the database from 1976 to 1999 and determined the breakdown of cases by age, sex, and race. RESULTS: The absolute number of new HNSCCA diagnoses per year declined overall by 5% during the time period of the study, whereas new diagnoses in patients older than 74 years of age increased by more than 20%. The rate of HNSCCA per 100,000 person-years in elderly women did not change, and the rate in elderly men decreased, indicating that the observed increase in cases is explained by a growing population of elderly persons at risk. An increase in the absolute number of cases, but not the incidence rate, was also seen among persons younger than 50 years of age. Although both the absolute number of new cases and the incidence rates of HNSCCA in white male patients declined substantially, the percentage of HNSCCA patients classified as minorities increased from 14.5% to more than 20% of all cases. During the time period of the study, the overall number of HNSCCA cases in nonwhite and Hispanic patients increased by 36%. CONCLUSION: Increasing numbers of elderly and minority patients with HNSCCA are likely to alter patterns of disease and utilization of health care resources.     

Motility-related proteins as markers for head and neck squamous cell cancer

HYPOTHESIS: Increased cell motility is a hallmark of cancer cells. Proteins involved in cell motility may be used as molecular markers to characterize the malignant potential of tumors. METHODS: Molecular biology and immunohistochemistry techniques were used to investigate the expression of a selected panel of motility-related proteins (Rho A, Rac 2, Cdc42, PI3K, 2E4, and Arp2) in normal, premalignant, and squamous cell cancer cell lines of human head and neck origin. To assess the clinical potential of these proteins as molecular markers for cancer, immunohistochemistry was performed on paraffin-fixed head and neck cancer specimens (n = 15). RESULTS: All six motility-associated proteins were overexpressed in the premalignant and squamous cell cancer cell lines relative to normal keratinocytes. Immunohistochemistry with Rho A and Rac 2 showed increased staining in areas of cancer but not in normal tissue. CONCLUSION: Proteins involved in cell motility can be used as markers for head and neck squamous cell carcinoma. The head and neck cell lines used in this study may be used as a model to further investigate cell motility. Molecular markers of motility could have a significant impact on the diagnosis and staging of cancers originating from differentiated non-motile cells.     

Human papillomavirus type 16 DNA detected by the polymerase chain reaction in non-cancer tissues of the head and neck

Cancer-free tissues from various anatomical subsites in the head and neck were examined by the polymerase chain reaction (PCR) for the incidence of human papillomavirus (HPV) types 16 and 18. We detected HPV 16 DNA in 9 of 103 samples (8.7%), including specimens from the paranasal sinuses, tonsil, hypopharynx and larynx. However, no HPV-16/18 DNA was detected by Southern hybridization in these 9 samples. The significance of the presence of HPV 16 DNA in non-cancer tissues is still unknown, but PCR detection only of high-risk HPV DNA in head and neck cancer should be evaluated cautiously because of its ubiquity in this region.     

Selective neck dissection for the treatment of neck metastasis from squamous cell carcinoma of the head and neck

OBJECTIVE: Our objective was to determine the proportion of patients disease free in the neck, with the primary site controlled, who have been treated with a selective neck dissection (SND) for squamous cell carcinoma (SCCa) of the upper aerodigestive tract, and who had cervical metastasis less than 3 cm. STUDY DESIGN: A cohort of patients who fit the inclusion/exclusion criteria was identified retrospectively. Then all surviving patients were followed for a minimum of 2 years. METHODS: A group of 52 patients who had 58 selective neck dissections for cervical metastases from SCCa of the upper aerodigestive tract were identified. The mean age was 56 years (range, 20-85 y), there were 40 males and 12 females, and mean follow-up was 24.5 months (range, 1-64 mo). Twenty-six patients had clinically negative (cNo) neck examinations and 26 had clinically positive neck examinations. Postoperative radiation was given for extracapsular spread, greater than 2 positive nodes, T3, T4, or recurrent disease if the patient had not received radiation before surgery. These radiation criteria excluded 18 patients from postoperative radiation treatment. RESULTS: Kaplan-Meier survival analysis showed that the regional control rate with the primary site controlled was 0.94. Six patients developed recurrent neck disease. Three of these 6 patientswere surgically salvaged. Four recurrences were in the dissected field and 2 were out of the dissected field (level V). CONCLUSIONS: With similar indications for radiation therapy, the regional control rate in this cohort is comparable to control rates obtained with modified radical neck dissection.     

Integrins in head and neck squamous cell carcinoma invasion

OBJECTIVE: To relate the invasive properties of different squamous cell cancer cell lines to the function and expression of the integrins. STUDY DESIGN: A series of in vitro and in vivo experiments were designed to assess and compare integrin expression and function in two different head and neck squamous cell carcinoma cell lines. METHODS: Invasive properties of two squamous cell carcinoma cell lines (UM-SCC-1 and JHU-022-SCC) were assessed using an in vitro artificial matrix assay as well as an in vivo system with orthotopically implanted tumor cells in mice. Whole cell and surface expression levels of integrin subunits (alpha2, alpha3, alpha5, alpha6, beta1, and beta4) were determined for each cell line using Western blot analysis and flow cytometry. We compared the ability of JHU-022-SCC and UM-SCC-1 cells to bind the extracellular matrix elements collagen IV, fibronectin, laminin 5, and laminin10 using an in vitro adhesion assay. Contributions of the different integrins to the adhesive properties were determined by selective antibody blocking of different subunits. RESULTS: The UM-SCC-1 cell line is 50% more invasive in vitro and displays a greater propensity for perineural and lymphatic invasion in vivo. The UM-SCC-1 cells exhibited greater adherence to fibronectin than JHU-022-SCC cells. Alpha6 and beta4 expression is approximately twofold greater in the JHU-022-SCC cells. Alpha2, alpha3, and beta1 expression appears to be upregulated in UM-SCC-1 cells. CONCLUSION: The UM-SCC-1 carcinoma cells are more invasive than JHU-022-SCC cells and may be related to differential expression of the integrins alpha6beta4, alpha3beta1, and alpha2beta1.     

Human papillomavirus types in cases of squamous cell carcinoma of head and neck in Colombia

Estimating the type-specific prevalence of human papillomavirus (HPV) in head and neck cancer (HNSCC) is helpful in predicting the impact of HPV immunization.ObjectiveTo estimate the overall prevalence, and gender and age-specific prevalence of HPV in HNSCC.MethodThis cross sectional retrospective study was carried out in four pathology laboratories of Medellin, Colombia. HPV testing was performed by GP5+/6+ PCR-based RLB and HPV 16 and 18 type-specific PCR.Results175 primary HNSCC cases consecutively diagnosed between 1999 and 2008 with confirmed diagnosis and amplifiable DNA were included. Overall HPV prevalence was 18.9%. HPV was found in 23.9%, 17.5% and 13.3% of the oral cavity, larynx and oropharynx cases respectively. Among HPV positive cases, 82% were HPV 16 and 18% were HPV 18. No other HPV genotypes were identified. Most patients were males. Male patients were younger that their female counterparts, particularly in oral cavity cancer cases.ConclusionHPV 16 and 18 genotypes were found in nearly 20% of HNSCC cases in Colombian patients. The impact of HPV vaccination for the prevention of HNSCC in this population deserves further evaluation.     

The role of human papillomavirus in head and neck cancer and the impact on radiotherapy outcome

Conclusion: HPV?+?HNSCC patients have improved Overall Survival (OS), Disease Specific Survival (DSS), Disease Free Survival (DFS), and Progression Free Survival (PFS). The radiotherapy treatment can’t improve the Survival of the HPV-negative HNSCC patients.

Objective: To investigate the role of Human papillomavirus in head and neck cancer and the impact on radiotherapy outcome.

Methods: A search in PubMed and Chinese CNKI (2000–2015) was performed. This meta-analysis was done using RevMan 5.1 software. Outcomes included OS, DSS, DFS, PFS, and Treatment responses rates (RR).

Results: A total of 2620 patients in 10 studies were included. The Positive detective rates of HPV and P16 are 32.5% (425/1309) and 42.5% (526/1239). OS and PFS were improved in HPV?+?patients compared to HPV???patients (HR?=?0.48; 95% CI?=?0.37–0.62, p?p?p?p?p?= 0.05).     

General primer-mediated polymerase chain reaction for simultaneous detection and typing of human papillomavirus DNA in laryngeal squamous cell carcinomas

The aim of this study was to investigate the prevalence of different human papillomavirus (HPV) types in laryngeal squamous cell carcinomas using general primer-mediated polymerase chain reaction (PCR). Tumour sections from 42 patients with laryngeal carcinomas were investigated. For HPV DNA amplification, consensus primers were used which were directed to the LI coding region of the HPV genome. Analysis of the PCR products was done using 2% agarose gel electrophoresis followed by restriction enzyme analysis to identify different HPV types. Amplification of the human TGF-β DNA was successfully performed in 36/42 (85.7%) of samples confirming the presence of sufficient DNA for viral amplification. HPV DNA was detected in 8/36 (22.2%) of the tumours examined (three HPV-6, two HPV-16, one HPV-11, two unknown HPV types). HPV DNA was not detected in any of the non-neoplastic laryngeal mucosa which was used as control (n = 15). Fifty per cent of women had HPV-positive tumours compared with 8% of men (x²= 5.8, P<0.05). Our data indicate that while the overall prevalence of HPV in laryngeal carcinomas is fairly high (22.2%), the frequency of high-risk types (HPV-16 & HPV-18) is low (5.5%). HPV probably acts as a promoter in the multistep process of carcinogenesis in squamous mucosal cells of the larynx.     

Sentinel node biopsy for oral and oropharyngeal squamous cell carcinoma of the head and neck

OBJECTIVES: The aims were to assess the technical feasibility of sentinel node biopsy (SNB), to validate SNB against elective neck dissection, and to report the results of the clinical application of the SNB concept for early oral and oropharyngeal squamous cell carcinoma. STUDY DESIGN: Prospective consecutive cohort analysis. METHODS: Between 2000 and 2006, a total of 79 patients were included. Lymphatic mapping consisted of preoperative lymphoscintigraphy and intraoperative use of a hand-held gammaprobe. Twenty-eight patients were assessed for feasibility and validation; the SNB was done in context with an elective neck dissection. Fifty-one patients were evaluated in an observational trial; elective neck dissection was performed only in case of positive SNB. RESULTS: Validation revealed a sentinel node detection rate by lymphoscintigraphy of 93%, with the gammaprobe of 100%. The negative predictive value of a negative SNB was 100%. During the observational trial 40% of the patients were upstaged as a result of a positive SNB. Intraoperative frozen section analysis showed a negative predictive value of 83%. Two patients (6%) with negative SNB experienced a neck recurrence, the negative predictive value of SNB was therefore 94%. Patients with positive SNB were treated successfully with elective neck dissection. CONCLUSIONS: SNB is technically feasible and reproducible with a high sentinel node detection rate. Validation against elective neck dissection revealed a negative predictive value of 100%. Application of the SNB concept in clinical practice was very successful. The recurrence rate within the neck was very low and the morbidity and cost of an elective neck dissection could be spared to 60% of the patients.     

Basaloid squamous cell carcinoma of the head and neck

OBJECTIVE/HYPOTHESIS: Basaloid squamous cell carcinoma (BSCC), an uncommon tumor with predilection for the upper aerodigestive tract, is a distinct variant of squamous carcinoma, because of its unique histological features and ominous clinical behavior. This study reviews the experience in treating BSCC from two institutions. STUDY DESIGN: Retrospective. METHODS: H&E-stained sections from 20 patients with BSCC of the head and neck were reviewed and clinical follow-up was obtained for all patients. RESULTS: The study group consisted of 14 male and 6 female patients. Their ages ranged from 43 to 85 years, with a mean age of 62 years. Sites of origin included the larynx (4), tongue (3), pyriform sinus (3), nose (2), floor of mouth (2), mastoid (1), tonsil (1), epiglottis (1), nasopharynx (1), trachea (1), and palate (1). Pain was the most common presenting symptom (5 cases), followed by hoarseness and bleeding (3 cases each). Tobacco and alcohol abuse was noted in 17 patients. Treatment modalities included surgery with or without chemotherapy or radiotherapy in 13 patients, chemotherapy with irradiation in 2, chemotherapy alone in 2, and radiotherapy alone in 3. Clinical follow-up revealed no evidence of disease in 11 patients. Four were alive with disease at the time of writing and five died of disease. CONCLUSION: BSCC is a highly aggressive malignant tumor that presents in elderly patients who have a history of abuse of tobacco or alcohol, or both. Greater number of patients must be studied and compared with age-matched and stage-matched controls of conventional squamous cell carcinoma to determine whether the poor clinical outcome is related more to high-stage presentation or to the tumor's high-grade malignant cytological features.     

Invasion of the carotid artery in tumours of the head and neck

The carotid arterial system may be invaded by a tumour of the head and neck. The incidence and management of this eventuality have been reviewed previously in full by us.¹ The purpose of the present paper is to record the incidence of this event in a personal series.     

Diagrammatic recording of head and neck tumours1

The increasing demands of clinical audit have resulted in the need for accurate data collection. The use of tumour maps allows standardization of the records of patients with head and neck cancer, which facilitates collation of data in multicentre studies and makes interdepartmental comparisons more meaningful. The aim of this study was to develop an improved standard set of tumour maps for recording the stage of head and neck tumours. A review of the existing tumour diagrams was performed to identify those anatomical areas that are not adequately represented or where ambiguity exists. The areas where improvements could be made were identified as: (1) the anterior commissure of the larynx; (2) axial and sagittal views of the larynx; (3) the pyriform fossa and cervical oesophagus; (4) the oropharynx and vallecula; (5) the nasal cavity and paranasal sinuses; and (6) cervical nodal involvement. A new set of tumour maps is presented in an attempt to correct some of the limitations of the existing diagrams.     

Racial disparities in patients with head and neck squamous cell carcinoma

OBJECTIVES/HYPOTHESIS: Black patients are reported to have a higher incidence of advanced disease and increased mortality from head and neck squamous cell carcinoma (HNSCC) but constitute the minority of patients in large-scale studies investigating the effect of race on outcome. This study sought to determine if racial disparities exist between black and white patients with HNSCC treated at a single large institution in the South with a high proportion of black patients. STUDY DESIGN: The authors conducted a nonrandomized retrospective cohort analysis. METHODS: The tumor registry was used to identify patients diagnosed with HNSCC from 1985 to 2002. The medical records of non-Hispanic white and black adult patients were retrospectively reviewed. Median household income, percentage of population below poverty level, and education level based on census tract and block information were obtained from U.S. Census 2000 data. Standard statistical analysis, including Kaplan-Meier survival curve analysis and Cox proportional hazards models, was used to analyze the effects of covariables on survival. RESULTS: A total of 1,128 patients met study criteria (478 black, 650 white). Compared with white patients, black patients were significantly younger (mean age, 53.9 vs. 56.4 years, P<.0001), male (81.2% vs. 72.3%, P=.0005), more commonly abused alcohol (88.0% vs. 74.3%, P<.0001), and were significantly less likely to have insurance (8.6% vs. 21.7%, P<.0001). There was no difference in the incidence of tobacco use (91.7%), advanced comorbidity (35.9%), or primary tumor site. Black patients had a significantly greater incidence of stage IV disease (65.7% vs. 46.6%, P<.0001) and nonoperative treatment (48.7% vs. 30.8%, P<.0001), which was performed for inoperable disease in 57.1% of black compared with 31.0% of white patients (P<.0001). Black patients resided in census block groups with significantly lower mean education level, median income, and a higher percentage of population below poverty compared with white patients. The 5-year disease-specific survival differed significantly between black (29.3%) and white (54.7%) patients (P<.0001). Cox proportional hazards models revealed that alcohol abuse, advanced TNM stage, high tumor grade, nodal disease, extracapsular spread, advanced comorbidity, and regional or distant metastatic disease were associated with poorer survival for all patients. An interaction with race was found for insurance status, nonoperative treatment, and extracapsular spread. Stepwise variable selection adjusting for patient, tumor, and treatment characteristics showed a significant effect only for race by payor status on disease-specific survival (P=.0228). CONCLUSIONS: Insurance status, treatment, and extracapsular spread differentially affected the survival of black patients compared with white patients. Only insurance status had a significant effect on survival in black patients after controlling for other variables. These data suggest that racial differences in HNSCC outcomes are primarily related to differences in access to health care.     

Multiple primary malignant tumours in patients with head and neck cancer: the implications for follow-up

The pattern of second primary cancer occurrence in 518 Scottish patients with head and neck cancer was determined by a retrospective study. The overall incidence of second cancers was 9% but the true incidence increased steadily in the years following initial diagnosis to reach a maximum of 21% at 11 years. After 4 years of follow-up patients were more likely to die from a second primary cancer than from the effects of the initial tumour. The Scottish cohort differed from previously reported, overseas, study groups in having a high incidence of second primary cancers in sites outside the upper aerodigestive tract. This potential demographic difference suggests a need for local audit prior to design and implementation of screening protocols for second primary cancers.     

Clinical value of 18FDG PET/CT in screening for distant metastases in head and neck squamous cell carcinoma

Objectives

The detection of distant metastases is of major importance in management of head and neck squamous cell carcinoma patients.

Design

All patients underwent ¹⁸FDG PET/CT for the detection of distant metastases.

Setting

Retrospective single‐centre study.

Participants

Head and neck squamous cell carcinoma patients with high‐risk factors for distant metastases.

Main outcome measures

Accuracy of ¹⁸FDG PET/CT for the detection of distant metastases using clinical development of distant metastases and a minimal follow‐up of twelve months as reference standard. Comparison of overall survival between patients diagnosed with distant metastases during initial screening and patients diagnosed with distant metastases during follow‐up.

Results

In 23 (12%) of the 190 patients, ¹⁸FDG PET/CT detected distant metastases at screening. Sensitivity and negative predictive value were 46.2% (95% CI 32.6‐59.7) and 82.6% (95% CI 76.8‐88.5). No difference in median overall survival from the time of distant metastases detection was found between patients diagnosed with DM during work‐up or during follow‐up.

Conclusions

In head and neck squamous cell carcinoma patients with high‐risk factors, ¹⁸FDG PET/CT has a high negative predictive value for the detection of distant metastases and should be used in daily clinical practice, although the sensitivity is limited when long‐term follow‐up is used as reference standard.     

Centrosome hyperamplification in head and neck squamous cell carcinoma: a potential phenotypic marker of tumor aggressiveness

OBJECTIVES/HYPOTHESIS: There is currently no single histological or genotypic marker that reliably predicts the biological behavior of head and neck squamous cell carcinoma (HNSCC). While multiple genetic mutations have been investigated, no single genotypic alteration has consistently correlated with tumor aggressiveness. Phenotypic markers may prove more predictive, because they can represent many different genetic alterations. We investigated the frequency of centrosome hyperamplification in HNSCC and examined its usefulness as a marker for tumor recurrence. STUDY DESIGN: Analysis of archived paraffin blocks using immunohistochemistry. METHODS: Eighteen patients who underwent resection of oral cavity squamous cell carcinoma were reviewed. Ten patients had cancers that recurred locally within 1 year of resection, and 8 patients were tumor free at 5 years. The amount of centrosome hyperamplification in the cancer specimens and all surgical margins was graded as follows: 0, none; 1+, rare hyperamplification; 2+, greater than 10% of cells per high-powered field; and 3 +, greater than 20% of cells per high-powered field. RESULTS: Centrosome hyperamplification was found in 17 of 18 tumors (94%). Grade 2+ or 3+ hyperamplification was found more in cancers that recurred (9 of 10) than in those that did not (3 of 8) and was more prevalent in the histologically normal margins of patients with recurrence (8 of 10) than in those without recurrent cancer (3 of 8). CONCLUSIONS: Our results demonstrate the extremely frequent occurrence of centrosome hyperamplification in HNSCC. Centrosome hyperamplification is a phenotypic marker for HNSCC and can reflect multiple genotypic changes. Its presence in histologically normal margins suggests that it may be useful for analysis of primary tumors and tumor margins.