Radioresistant laryngeal cancers upregulate type 1 IGF receptor and exhibit increased cellular dependence on IGF and EGF signalling.

OBJECTIVES: Patients failing radiotherapy for laryngeal squamous cell carcinoma (LSCC) often require salvage total laryngectomy which has major functional consequences, highlighting a need for biomarkers of radiotherapy resistance. In other tumour types, radioresistance has been linked to epidermal growth factor receptor (EGFR) and type 1 insulin-like growth factor receptor (IGF-1R). Here, we evaluated IGF-1R and EGFR as predictors and mediators of LSCC radioresistance. DESIGN: We compared IGF-1R and EGFR immunohistochemical scores in patients with LSCC achieving long-term remission post-radiotherapy (n = 23), patients treated with primary laryngectomy (n = 22) or salvage laryngectomy following radiotherapy recurrence (n = 18). To model radioresistance in vitro, two LSCC cell lines underwent clinically relevant irradiation to 55 Gy in 2.75 Gy fractions. RESULTS: Type 1 insulin-like growth factor receptor expression was higher in pre-treatment biopsies of radiotherapy failures compared with those in long-term remission and was upregulated post-radiotherapy. Patients undergoing primary laryngectomy had more advanced T/N stage and greater tumour IGF-1R content than those achieving long-term remission. Pre-treatment EGFR did not associate with radiotherapy outcomes but showed a trend to upregulation post-irradiation. In vitro, radiosensitivity was enhanced by inhibition of EGFR but not IGF. Repeated irradiation upregulated IGF-1R in BICR18 and SQ20B cells and EGFR in SQ20B, and enhanced SQ20B radioresistance. Repeatedly irradiated SQ20B_55 cells were not radiosensitised by inhibition of IGF and/or EGFR, but IGF-1R:EGFR co-inhibition suppressed baseline cell survival more effectively than blockade of either pathway alone, and more effectively than in parental cells. CONCLUSIONS: Radiation upregulates IGF-1R and may enhance IGF/EGFR dependence, suggesting that IGF/EGFR blockade may have activity in LSCCs that recur post-radiotherapy.

Current and future techniques for human papilloma virus (HPV) testing in oropharyngeal squamous cell carcinoma.

Despite a reduction in smoking and alcohol consumption, the incidence of oropharyngeal squamous cell carcinoma (OPSCC) is rising. This is attributed to human papilloma virus (HPV) infection and screening for HPV is now recommended in all cases of OPSCC. Despite a variety of clinically available tests and new non-invasive test strategies there is no consensus on which technique is best. This review reports on current techniques for HPV detection in OPSCC and the clinical applicability of emerging techniques. Literature searches of Medline, Embase and clinicaltrials.gov using the search terms 'head and neck neoplasms', 'squamous cell carcinoma' and 'HPV testing' were performed. 45 studies were identified and included. p16 immunohistochemistry (IHC), HPV DNA in situ hybridization (ISH) and HPV polymerase chain reaction (PCR) are the commonest tests to determine HPV status. p16 IHC and HPV DNA PCR are highly sensitive whilst HPV DNA ISH is more specific, these techniques conventionally utilize surgical biopsies. New tests using PCR to screen fine needle aspirates, saliva, brush cytology and serum for HPV are promising but have variable sensitivity and specificity. These non-invasive samples avoid the morbidity of surgical biopsies and need for tissue blocks; their clinical role in screening and surveillance remains largely untested. Further work is needed to validate these tests and define their role.

IGF-1R expression is associated with HPV-negative status and adverse survival in head and neck squamous cell cancer.

Head and neck squamous cell carcinomas (HNSCC) are treated with surgery, radiotherapy and cisplatin-based chemotherapy, but survival from locally-advanced disease remains poor, particularly in patients whose tumors are negative for Human papillomavirus (HPV). Type 1 IGF receptor (IGF-1R) is known to promote tumorigenesis and resistance to cancer therapeutics. Here, we assessed IGF-1R immunohistochemistry on tissue microarrays containing 852 cores from 346 HNSCC patients with primary tumors in the oropharynx (n = 231), larynx (85), hypopharynx (28), oral cavity (2). Of these, 236 (68%) were HPV-negative, 110 (32%) positive. IGF-1R was detected in the cell membrane of 36% and cytoplasm of 92% of HNSCCs; in 64 cases with matched normal tonsillar epithelium, IGF-1R was overexpressed in the HNSCCs (P < 0.001). Overall survival (OS) and disease-specific survival (DSS) were reduced in patients whose tumors contained high membrane IGF-1R [OS: hazard ratio (HR) = 1.63, P = 0.006; DSS: HR = 1.63, P = 0.016], cytoplasmic IGF-1R (OS: HR = 1.58, P = 0.009; DSS: HR = 1.58, P = 0.024) and total IGF-1R (OS: HR = 2.02, P < 0.001; DSS: HR = 2.2, P < 0.001). High tumor IGF-1R showed significant association with high-tumor T-stage (P < 0.001) and HPV-negativity (P < 0.001), and was associated with shorter OS when considering patients with HPV-positive (P = 0.01) and negative (P = 0.006) tumors separately. IGF-1R was independently associated with survival in multivariate analysis including HPV, but not when lymphovascular invasion, perineural spread and T-stage were included. Of these factors, only IGF-1R can be manipulated; the association of IGF-1R with aggressive disease supports experimental incorporation of anti-IGF-1R agents into multimodality treatment programs for HPV-negative and high IGF-1R HPV-positive HNSCC.

Clear cell odontogenic carcinoma: a rare neoplasm of the maxillary bone.

Clear cell odontogenic carcinoma (CCOC) is an extremely rare neoplasm, with only 74 cases in the English-language literature. It displays a propensity for the mandible, most commonly presenting in the fifth to seventh decades. Histopathologically, CCOC is characterized by sheets and islands of vacuolated and clear cells. The aggressive nature of CCOC was noted in its first description in 1985, although it was not formally classified as malignant by the World Health Organization until 2005. This report describes a case of CCOC presenting atypically in a young patient and at an uncommon site. The authors review the details of this case, outlining management strategies referencing their experience and that described in the other limited cases in the literature.

Developing and Validating a Multivariable Prognostic-Predictive Classifier for Treatment Escalation of Oropharyngeal Squamous Cell Carcinoma: The PREDICTR-OPC Study.

PURPOSE: While there are several prognostic classifiers, to date, there are no validated predictive models that inform treatment selection for oropharyngeal squamous cell carcinoma (OPSCC).Our aim was to develop clinical and/or biomarker predictive models for patient outcome and treatment escalation for OPSCC. EXPERIMENTAL DESIGN: We retrospectively collated clinical data and samples from a consecutive cohort of OPSCC cases treated with curative intent at ten secondary care centers in United Kingdom and Poland between 1999 and 2012. We constructed tissue microarrays, which were stained and scored for 10 biomarkers. We then undertook multivariable regression of eight clinical parameters and 10 biomarkers on a development cohort of 600 patients. Models were validated on an independent, retrospectively collected, 385-patient cohort. RESULTS: A total of 985 subjects (median follow-up 5.03 years, range: 4.73-5.21 years) were included. The final biomarker classifier, comprising p16 and survivin immunohistochemistry, high-risk human papillomavirus (HPV) DNA in situ hybridization, and tumor-infiltrating lymphocytes, predicted benefit from combined surgery + adjuvant chemo/radiotherapy over primary chemoradiotherapy in the high-risk group [3-year overall survival (OS) 63.1% vs. 41.1%, respectively, HR = 0.32; 95% confidence interval (CI), 0.16-0.65; P = 0.002], but not in the low-risk group (HR = 0.4; 95% CI, 0.14-1.24; P = 0.114). On further adjustment by propensity scores, the adjusted HR in the high-risk group was 0.34, 95% CI = 0.17-0.67, P = 0.002, and in the low-risk group HR was 0.5, 95% CI = 0.1-2.38, P = 0.384. The concordance index was 0.73. CONCLUSIONS: We have developed a prognostic classifier, which also appears to demonstrate moderate predictive ability. External validation in a prospective setting is now underway to confirm this and prepare for clinical adoption.

Department Wide Validation in Digital Pathology-Experience from an Academic Teaching Hospital Using the UK Royal College of Pathologists' Guidance.

AIM: we describe our experience of validating departmental pathologists for digital pathology reporting, based on the UK Royal College of Pathologists (RCPath) "Best Practice Recommendations for Implementing Digital Pathology (DP)," at a large academic teaching hospital that scans 100% of its surgical workload. We focus on Stage 2 of validation (prospective experience) prior to full validation sign-off. METHODS AND RESULTS: twenty histopathologists completed Stage 1 of the validation process and subsequently completed Stage 2 validation, prospectively reporting a total of 3777 cases covering eight specialities. All cases were initially viewed on digital whole slide images (WSI) with relevant parameters checked on glass slides, and discordances were reconciled before the case was signed out. Pathologists kept an electronic log of the cases, the preferred reporting modality used, and their experiences. At the end of each validation, a summary was compiled and reviewed with a mentor. This was submitted to the DP Steering Group who assessed the scope of cases and experience before sign-off for full validation. A total of 1.3% (49/3777) of the cases had a discordance between WSI and glass slides. A total of 61% (30/49) of the discordances were categorised as a minor error in a supplementary parameter without clinical impact. The most common reasons for diagnostic discordances across specialities included identification and grading of dysplasia, assessment of tumour invasion, identification of small prognostic or diagnostic objects, interpretation of immunohistochemistry/special stains, and mitotic count assessment. Pathologists showed similar mean diagnostic confidences (on Likert scale from 0 to 7) with a mean of 6.8 on digital and 6.9 on glass slide reporting. CONCLUSION: we describe one of the first real-world experiences of a department-wide effort to implement, validate, and roll out digital pathology reporting by applying the RCPath Recommendations for Implementing DP. We have shown a very low rate of discordance between WSI and glass slides.

Mixed Olfactory Neuroblastoma and Adenocarcinoma with In Situ Neuroendocrine Hyperplasia.

Olfactory neuroblastoma (ONB) is a rare malignant neoplasm arising from the superior aspect of the nasal vault. Cases are characterised by insidious clinical presentation and high rates of recurrence despite surgical resection and adjuvant radiotherapy. There are a small number of reports showing ONB with divergent epithelial or ganglionic differentiation, and ONB has also been found to coincide with adenocarcinoma. We present a case of mixed ONB with adenocarcinoma. The clinical presentation was unusual, with a tonic-clonic seizure preceded by chronic headache and anosmia. Imaging revealed a mass extending from the olfactory recess of the left nasal cavity through the cribriform plate to the anterior cranial fossa. The pathology demonstrated intraepithelial neuroendocrine cell hyperplasia in the left olfactory groove. This finding provides a unique insight into the cellular origin of this rare tumour, and appears to confirm the theory that ONB arises from neural stem cells in the olfactory neuroepithelium. Despite radical treatment, the patient suffered a distant recurrence within 1 year of treatment, which underlines the aggressive nature of this tumour.

Relation of a hypoxia metagene derived from head and neck cancer to prognosis of multiple cancers.

Affymetrix U133plus2 GeneChips were used to profile 59 head and neck squamous cell cancers. A hypoxia metagene was obtained by analysis of genes whose in vivo expression clustered with the expression of 10 well-known hypoxia-regulated genes (e.g., CA9, GLUT1, and VEGF). To minimize random aggregation, strongly correlated up-regulated genes appearing in >50% of clusters defined a signature comprising 99 genes, of which 27% were previously known to be hypoxia associated. The median RNA expression of the 99 genes in the signature was an independent prognostic factor for recurrence-free survival in a publicly available head and neck cancer data set, outdoing the original intrinsic classifier. In a published breast cancer series, the hypoxia signature was a significant prognostic factor for overall survival independent of clinicopathologic risk factors and a trained profile. The work highlights the validity and potential of using data from analysis of in vitro stress pathways for deriving a biological metagene/gene signature in vivo.

Clinically actionable mutation profiles in patients with cancer identified by whole-genome sequencing.

Next-generation sequencing (NGS) efforts have established catalogs of mutations relevant to cancer development. However, the clinical utility of this information remains largely unexplored. Here, we present the results of the first eight patients recruited into a clinical whole-genome sequencing (WGS) program in the United Kingdom. We performed PCR-free WGS of fresh frozen tumors and germline DNA at 75× and 30×, respectively, using the HiSeq2500 HTv4. Subtracted tumor VCFs and paired germlines were subjected to comprehensive analysis of coding and noncoding regions, integration of germline with somatically acquired variants, and global mutation signatures and pathway analyses. Results were classified into tiers and presented to a multidisciplinary tumor board. WGS results helped to clarify an uncertain histopathological diagnosis in one case, led to informed or supported prognosis in two cases, leading to de-escalation of therapy in one, and indicated potential treatments in all eight. Overall 26 different tier 1 potentially clinically actionable findings were identified using WGS compared with six SNVs/indels using routine targeted NGS. These initial results demonstrate the potential of WGS to inform future diagnosis, prognosis, and treatment choice in cancer and justify the systematic evaluation of the clinical utility of WGS in larger cohorts of patients with cancer.

Relation of erythropoietin and erythropoietin receptor expression to hypoxia and anemia in head and neck squamous cell carcinoma.

PURPOSE: The use of erythropoietin in head and neck squamous cell carcinoma (HNSCC) has been associated with poor survival. This study examines the protein and mRNA expression of erythropoietin and erythropoietin receptor in HNSCC and their relation to hypoxia, hemoglobin (Hb), and clinical outcome. EXPERIMENTAL DESIGN: The immunohistochemical expression of erythropoietin and erythropoietin receptor was assessed in 151 cases of HNSCC. Expression was compared with the hypoxia-dependent proteins hypoxia-inducible factor-1alpha (HIF-1alpha) and carbonic anhydrase-9 (CA-9) and correlated with clinical outcome. The mRNA expression of erythropoietin and erythropoietin receptor was measured in paired samples of HNSCC. RESULTS: Erythropoietin and erythropoietin receptor were expressed in 95% and 99% of tumors, respectively. Using a weighed expression score, there was a positive correlation between erythropoietin and erythropoietin receptor expression (r = 0.18, P = 0.03). HIF-1alpha (r = 0.38, P < 0.01) and CA-9 (r = 0.26, P = 0.002) correlated with erythropoietin expression, but there was no correlation with erythropoietin receptor. No correlation was found between Hb and erythropoietin (r = 0.07, P = 0.36) or erythropoietin receptor (r = -0.02, P = 0.8), and no survival difference between high and low erythropoietin or erythropoietin receptor expression (P = 0.59 and P = 0.98, respectively). The mRNA expression of erythropoietin (P = 0.03) but not erythropoietin receptor (P = 0.62) was significantly increased in 11 paired samples of HNSCC. CONCLUSION: In vivo, the HIF pathway regulates erythropoietin at the mRNA level but not erythropoietin receptor expression in HNSCC. Anemia does not seem to influence the hypoxic microenvironment of tumors sufficiently to alter the expression of erythropoietin. The effects of exogenous erythropoietin may be acting via receptors expressed on tumor cells in vivo, or on vascular cells, which also express the pathway.

γH2AX expression in cytological specimens as a biomarker of response to radiotherapy in solid malignancies.

Many anticancer treatments, including radiotherapy, act by damaging DNA and hindering cell function and proliferation. H2AX is a histone protein directly associated with DNA that is phosphorylated to produce γH2AX that accumulates in foci in an early response to DNA double-strand breaks, the most deleterious lesion caused by anticancer therapy. This study reports a γH2AX detection assay that has the potential to be used as a biomarker of response to guide cancer treatment. γH2AX immunostaining was applied to tumour cell specimens obtained using fine needle aspiration (FNA). Liquid-based cytology and direct smear cytology methods were evaluated and immunostaining protocols established using FNA samples from five cancer patients. The assay was then applied to three patients before and after radiotherapy. Results demonstrate induction of γH2AX foci following treatment, persisting for as long as one week after therapy. Immunostaining for γH2AX has been successfully applied to FNA samples, providing an opportunity to evaluate γH2AX as a treatment response marker in cancer.

Tumour depth of invasion of pT1 squamous cell carcinoma of the oral tongue and risk of pathologically detected neck metastases.

BACKGROUND: Tumour depth of invasion (TDI) is considered a predictor of pathologically detected neck metastases (PDNM) for squamous cell carcinoma (SCC), but different investigators have arrived at different cut-off of TDI. However, the relationship between TDI of pT1 SCC of the oral tongue and PDNM remains unknown. METHODS: Data was collected for patients with pT1SCC of the oral tongue. TDI, neurovascular invasion, pattern of invasion and presence of PDNM were recorded. The relationship between data was studied using logistic regression and ROC methods. RESULTS: With all other factors held constant, data showed that the odds ratio for each millimetre increase in TDI and risk of PDNM was 1.09 (95% CI: 0.95 - 1.25, p = 0.234), which was insignificant. CONCLUSION: TDI is not accurate and cannot be used as predictor of PDNM in patients with pT1 SCC of the tongue. Further, true TDI can only be assessed on resection specimens. This article is protected by copyright. All rights reserved.

Fine needle aspiration.

Fine needle aspiration test is a useful tool in the investigation of head and neck masses, be they of salivary gland, lymphoid, thyroid or other (branchial cyst) origin. Some practical aspects of this procedure are presented.

Epithelioid sarcoma with metastatic spread to the tongue.

Epithelioid sarcoma (ES) is a rare tumour that seldom presents to the otolaryngologist. It typically occurs in the extremities of young adolescents; however, it has the capability of metastasizing, often to the lungs or skin. The diagnosis is by histopathological examination and immunohistochemistry. We present a case of metastatic ES occuring in the tongue, a tumour not reported previously in the English literature.

A rare case of localised oral amyloid of the labial mucosa.

Amyloidosis is often a systemic process, and localised oral amyloidosis is rare. We present the case of a young woman with amyloid deposition in the labial mucosa of her lower lip. Systemic involvement was excluded by comprehensive assessment at the UK Amyloidosis Centre. Of 40 previously reported cases of localised oral amyloidosis we found only one that was limited to the labial mucosa.

Parotid cancer treatment with surgery followed by radiotherapy in Oxford over 15 years.

INTRODUCTION: Primary parotid malignancies represent a rare diagnosis, making high-quality comparative research unfeasible. There is little U.K.-based evidence to guide practice. A review was therefore undertaken of a large series of patients treated by a multidisciplinary team in a National Health Service tertiary referral centre. PATIENTS AND METHODS: Retrospective patient record review at the John Radcliffe Hospital in Oxford identified 401 patients who had undergone parotidectomy between 1995 and 2010, of whom 50 subjects were given a definitive diagnosis of primary parotid malignancy, treated with surgery and postoperative radiotherapy. Case notes, histology and imaging were reviewed by the study team. RESULTS: The median follow up for the cohort was 60 months (range: 1-108 months). Facial nerve function was preserved in all patients undergoing partial or total conservative parotidectomy. Although histology showed microscopically close or positive margins in 82% of cases, all patients underwent postoperative radiotherapy and locoregional recurrence was identified in only two (4%) patients. CONCLUSIONS: The data presented demonstrate a reasonable and practical multidisciplinary approach to a complex management problem. Facial nerve sparing surgery and postoperative radiotherapy result in good control of locoregional disease.

Ewings sarcoma of the zygoma reconstructed with a gold prosthesis: a rare tumor and unique reconstruction.

INTRODUCTION: Ewings sarcoma (ES) is a rare tumor, most commonly encountered within the long bones during the first 20 years of life. CASE REPORT: We report the sixth case of ES of the zygoma, occurring in a 31-year-old male, and presenting a unique reconstructive opportunity. Diagnosis and preoperative planning were aided by CT/MR coregistration. Surgical reconstruction using 3D reconstructed CT images to produce an anatomically correct model provided the basis for a gold prosthesis construction. DISCUSSION: Delayed definitive reconstruction provided opportunity for adequate tumor recurrence surveillance, and definitive histological diagnosis. The artefact produced on MR imaging from gold implants is minimal, permitting the unrestricted identification of potential future recurrence. CONCLUSION: This case highlights the benefits of coregistration of radiological imaging, and custom-made gold prostheses providing the advantage of artefact-free MR imaging, which should be considered in patients requiring resection of zygomatic tumors and subsequent reconstruction.

Malakoplakia presenting as an enlarging neck mass: Case presentation and review of the literature.

BACKGROUND: Malakoplakia is a very rare chronic inflammatory disorder, first described in 1902. In 75% of cases, the condition affects the genitourinary tract. Five cases of malakoplakia affecting the neck were previously reported in the literature. METHODS AND RESULTS: An 83-year-old woman presented with an enlarging mass in the posterior triangle of the neck that was histologically confirmed as malakoplakia. Presenting features are often nonspecific, and the diagnosis is dependent on histological findings. The characteristic microscopic findings are of Michaelis-Gutmann (M-G) bodies that stain positive with periodic acid-Schiff reagent, von Kossa's reaction for calcium, and Perl's ferrocyanide reaction to ferric iron. CONCLUSION: Although rare, a diagnosis of malakoplakia should be considered in patients with an enlarging mass. This may mimic the presentation of malignancy, particularly in patients in whom erosion through skin occurs, and histological confirmation is advocated.