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Aims: Patients with recurrent or metastatic head and neck squamous cell carcinoma (HNSCC) are eligible for first‐line immune checkpoint inhibition if their tumour is positive for programmed death ligand 1 (PD‐L1) determined by the combined positive score (CPS). This nationwide study, using real‐world data, investigated the developing PD‐L1 testing landscape in the first 3 years after introduction of the test in HNSCC and examined interlaboratory variation in PD‐L1 positivity rates. Methods: Pathology reports of HNSCC patients mentioning PD‐L1 were extracted from the Dutch Pathology Registry (Palga). Tumour and PD‐L1 testing characteristics were analysed per year and interlaboratory variation in PD‐L1 positivity rates was assessed using funnel plots with 95% confidence limits around the overall mean. Results: A total of 817 PD‐L1 tests were reported in 702 patients among 19 laboratories; 85.2% of the tests on histological material were stated to be positive. The national PD‐L1 positivity rate differed significantly per year during the study period (79.7–89.9%). The use of the recommended 22C3 antibody increased from 59.9 to 74.3%. A total of 673 PD‐L1 tests on histological material from 12 laboratories were analysed to investigate interlaboratory variation. Four (33%) deviated significantly from the national mean of PD‐L1‐positive cases using CPS ≥ 1 cut‐off, while two (17%) deviated significantly for CPS ≥ 20 cut‐off. Conclusion: In the first 3 years of PD‐L1 assessment in HNSCC, the testing landscape became more uniform. However, interlaboratory variation in PD‐L1 positivity rates between Dutch laboratories was substantial. This implies that there is a need for further test standardisation to reduce this variation. [ABSTRACT FROM AUTHOR]

Aims: Programmed death‐ligand 1 (PD‐L1) immunostaining is used to predict which non‐small‐cell lung cancer (NSCLC) patients will respond best to treatment with programmed cell death protein 1/PD‐L1 inhibitors. PD‐L1 immunostaining is sometimes performed on alcohol‐fixed cytological specimens instead of on formalin‐fixed paraffin‐embedded (FFPE) biopsies or resections. We studied whether ethanol prefixation of clots from endobronchial ultrasound‐guided transbronchial needle aspiration (EBUS‐TBNA) results in diminished PD‐L1 immunostaining as compared with formalin fixation. Methods and results: FFPE cell blocks from EBUS‐TBNA specimens of 54 NSCLC patients were identified. For each case, paired samples were available, consisting of clots directly immersed in formalin and clots prefixed in Fixcyt (50% ethanol). Serial sections were immunostained for PD‐L1 by use of the standardised SP263 assay and the 22C3 antibody as a laboratory‐developed test (LDT). PD‐L1 positivity was determined with two cut‐offs (1% and 50%). Concordance of PD‐L1 positivity between the formalin‐fixed (gold standard) and ethanol‐prefixed material was assessed. When the 22C3 LDT was used, 30% and 36% of the ethanol‐prefixed specimens showed false‐negative results at the 1% and 50% cut‐offs, respectively (kappa 0.64 and 0.68). When SP263 was used, 22% of the ethanol‐prefixed specimens showed false‐negative results at the 1% cut‐off (kappa 0.67). At the 50% cut‐off, concordance was higher (kappa 0.91), with 12% of the ethanol‐prefixed specimens showing false‐negative results. Conclusion: Ethanol fixation of EBUS‐TBNA specimens prior to formalin fixation can result in a considerable number of false‐negative PD‐L1 immunostaining results when a 1% cut‐off is used and immunostaining is performed with SP263 or the 22C3 LDT. The same applies to use of the 50% cut‐off when immunostaining is performed with the 22C3 LDT. [ABSTRACT FROM AUTHOR]

Background: Immunohistochemical staining of programmed death‐ligand 1 (PD‐L1) is used to determine which patients with non–small cell lung cancer (NSCLC) may benefit most from immunotherapy. Therapeutic management of many patients with NSCLC is based on cytology instead of histology. In this study, concordance of PD‐L1 immunostaining between cytology cell blocks and their histologic counterparts was analyzed. Furthermore, the effect of various fixatives and fixation times on PD‐L1 immunoreactivity was studied. Methods: Paired histologic and cytologic samples from 67 patients with NSCLC were collected by performing fine‐needle aspiration on pneumonectomy/lobectomy specimens. Formalin‐fixed, agar‐based or CytoLyt/PreservCyt‐fixed Cellient cell blocks were prepared. Sections from cell blocks and tissue blocks were stained with SP263 (standardized assay) and 22C3 (laboratory‐developed test) antibodies. PD‐L1 scores were compared between histology and cytology. In addition, immunostaining was compared between PD‐L1–expressing human cell lines fixed in various fixatives at increasing increments in fixation duration. Results: Agar cell blocks and tissue blocks showed substantial agreement (κ = 0.70 and κ = 0.67, respectively), whereas fair‐to‐moderate agreement was found between Cellient cell blocks and histology (κ = 0.28 and κ = 0.49, respectively). Cell lines fixed in various alcohol‐based fixatives showed less PD‐L1 immunoreactivity compared with those fixed in formalin. In contrast to SP263, additional formalin fixation after alcohol fixation resulted in preserved staining intensity using the 22C3 laboratory‐developed test and the 22C3 pharmDx assay. Conclusions: Performing PD‐L1 staining on cytologic specimens fixed in alcohol‐based fixatives could result in false‐negative immunostaining results, whereas fixation in formalin leads to higher and more histology‐concordant PD‐L1 immunostaining. The deleterious effect of alcohol fixation could be reversed to some degree by postfixation in formalin. Fixation of cytologic specimens in alcohol‐based fixatives results in negative effects on programmed death‐ligand 1 (PD‐L1) immunostaining when using PD‐L1 immunohistochemistry protocols validated on formalin‐fixed, paraffin‐embedded material. The use of formalin results in more histology‐concordant PD‐L1 immunostaining, and the deleterious effect of alcohol fixation potentially may be reversed to some degree by postfixation in formalin. [ABSTRACT FROM AUTHOR]

Background: Hypoxia is a negative prognostic factor in head and neck squamous cell carcinomas. Under hypoxia, the hypoxia‐inducible factor (HIF)‐1a transcription factor is overexpressed. We investigated whether there were site differences in HIF‐1a expression and its effect on patient outcomes per subsite. Design/Method: A total of 941 patients with HNSCC in the squamous cell carcinoma of the oropharynx (OPSCC, n = 302), oral cavity (OSCC, n = 391), or larynx (LSCC, n = 248) were included. Expression of HIF‐1a in tissue samples was investigated using immunohistochemistry. Overall survival (OS), disease‐free survival (DFS), and locoregional control (LRC) were analyzed. Results: HIF‐1a expression was higher in OSCC than in LSCC and OPSCC. High HIF‐1a expression led to worse prognosis in OPSCC (OS P =.029, DFS P =.085) and LSCC (OS P =.041, DFS P =.011) and better in OSCC (OS P =.055, DFS P =.012). There was no association between HIF‐1a and LRC. Conclusions: High HIF‐1a expression is related to poor outcome in OPSCC and LSCC and better outcome in OSCC. [ABSTRACT FROM AUTHOR]

Programmed cell death ligand 1 (PD‐L1) immunohistochemistry is used to determine which patients with advanced non‐small‐cell lung cancer (NSCLC) respond best to treatment with PD‐L1 inhibitors. For each inhibitor, a unique immunohistochemical assay was developed. This systematic review gives an up‐to‐date insight into the comparability of standardised immunohistochemical assays and laboratory‐developed tests (LDTs), focusing specifically on tumour cell (TC) staining and scoring. A systematic search was performed identifying publications that assessed interassay, interobserver and/or interlaboratory concordance of PD‐L1 assays and LDTs in tissue of NSCLC patients. Of 4294 publications identified through the systematic search, 27 fulfilled the inclusion criteria and were of sufficient methodological quality. Studies assessing interassay concordance found high agreement between assays 22C3, 28‐8 and SP263 and properly validated LDTs, and lower concordance for comparisons involving SP142. A decrease in concordance, however, is seen with use of cut‐offs, which hampers interchangeability of PD‐L1 immunohistochemistry assays and LDTs. Studies assessing interobserver concordance found high agreement for all assays and LDTs, but lower agreement with use of a 1% cut‐off. This may be problematic in clinical practice, as discordance between pathologists at this cut‐off may result in some patients being denied valuable treatment options. Finally, five studies assessed interlaboratory concordance and found moderate to high agreement levels for various assays and LDTs. However, to assess the actual existence of interlaboratory variation in PD‐L1 testing and PD‐L1 positivity in clinical practice, studies using real‐world clinical pathology data are needed. [ABSTRACT FROM AUTHOR]

Accurate, consistent and reproducible grading by pathologists is of key‐importance for identification of individual patients with invasive breast cancer (IBC) that will or will not benefit from adjuvant systemic treatment. We studied the laboratory‐specific grading variation using nationwide real‐life data to create insight and awareness in grading variation. Synoptic pathology reports of all IBC resection‐specimens, obtained between 2013 and 2016, were retrieved from the nationwide Dutch Pathology Registry (PALGA). Absolute differences in laboratory‐proportions of Grades I–III were compared to the national reference. Multivariable logistic regression provided laboratory‐specific odds ratios (ORs) for high‐ vs. low‐grade IBC. 33,792 IBC pathology reports of 33,043 patients from 39 laboratories were included, of which 28.1% were reported as Grade I (range between laboratories 16.3–43.3%), 47.6% as Grade II (38.4–57.8%), and 24.3% as Grade III (15.5–34.3%). Based on national guidelines, the indication for adjuvant chemotherapy was dependent on histologic grade in 29.9% of patients. After case‐mix correction, 20 laboratories (51.3%) showed a significantly deviant OR. Significant grading differences were also observed among pathologists within laboratories. In this cohort of 33,043 breast cancer patients, we observed substantial inter‐ and intra‐laboratory variation in histologic grading. It can be anticipated that this has influenced outcome including exposure to unnecessary toxicity, since choice of adjuvant chemotherapy was dependent on grade in nearly a third of patients. Better standardization and training seems warranted. What's new? Histologic grade serves a critical prognostic role in invasive breast cancer (IBC) and is used to guide therapeutic decisions. Evidence indicates, however, that IBC grading varies considerably. Here, grading variation in clinical practice was evaluated using real‐life data from laboratories in the nationwide Dutch Pathology Registry. Laboratories varied in IBC grade I, II, and III reporting. Among grading components, nuclear polymorphism showed the greatest difference between laboratories. Within laboratories, one‐third of pathologists deviated significantly from national proportions for IBC grade I. Despite deployment of uniform guidelines across laboratories, IBC histologic grading is not necessarily performed in a consistent manner. [ABSTRACT FROM AUTHOR]

Objective: Literature analysis on the prognostic factor of the nodal yield (NY) in neck dissections (NDs), which in general surgical oncology is a strong prognosticator and quality‐of‐care marker. Methods: We performed a systematic review of all PubMed and Embase publications until June 30, 2018 screening for data on NY as prognosticator and overall survival (OS) as outcome in patients with head and neck squamous cell carcinoma (HNSCC). Risk for bias was asserted by application of the Quality In Prognosis Studies tool. Results: Of the 823 screened publications, 15 were included in this analysis. Five out of seven that compared NY ≥18 vs <18 as prognosticator, showed significantly improved survival if NY ≥18. Six studies used other cutoffs and three reported improved survival with each additionally harvested lymph node. Conclusion: Increased NY in ND specimen for HNSCC, most commonly described as ≥18 lymph nodes, is associated with improved OS and could be used as a prognosticator and quality‐of‐care marker. [ABSTRACT FROM AUTHOR]

Background: To investigate if depth of invasion (DOI) can predict occult nodal disease in patients with cT1‐2N0 (7th TNM) oral squamous cell carcinoma (OSCC) staged by sentinel lymph node biopsy (SLNB). Methods: In 199 OSCC patients, DOI measurements and SLNB were performed. Results: Metastases were found in 64 of 199 patients (32%). Of these 64 patients, the mean DOI was 6.6 mm compared to 4.7 mm in patients without metastases (P = .003). The ROC‐curve showed an area under the curve of 0.65 with a most optimal cutoff point of 3.4 mm DOI (sensitivity 83% and specificity 47%). Regional metastases were found in 15% of patients with DOI ≤ 3.4 mm. Conclusion: DOI seems to be a poor predictor for regional metastasis in patients with cT1‐2N0 OSCC. Therefore, staging of the neck using SLNB in patients with early stage oral cancer should also be performed in tumors with limited DOI and probably in T3 (8th TNM) OSCC ≤4 cm diameter. [ABSTRACT FROM AUTHOR]

Objective: Most studies that report on salvage surgery after primary radiotherapy for head and neck squamous cell carcinoma (HNSCC) are small and heterogeneous. Subsequently, some relevant questions remain unanswered. We specifically focused on (1) difference in prognosis per tumor subsite, corrected for disease stage, and (2) differences in prognosis after salvage surgery for local, regional, and locoregional recurrences.Study Design: Retrospective analysis.Setting: Single-center study (2000-2016).Subjects and Methods: Patients treated with salvage surgery for HNSCC recurrence after (chemo)radiotherapy.Results: In total, 189 patients were included. Five-year overall survival (OS) was 33%, and median OS was 18 (95% confidence interval [CI], 11-26) months. Treatment-related mortality was 2%. Larynx carcinoma was associated with more favorable local (adjusted hazard ratio [HR] = 4.02; 95% CI, 1.46-11.10; P = .007) and locoregional control (adjusted HR = 5.34; 95% CI, 1.83-15.61; P = .002) than pharyngeal carcinoma. American Society of Anesthesiologists (ASA) score (≥3 vs 1-2: adjusted HR = 3.04; 95% CI, 1.17-7.91; P = .023), pT stage (3-4 vs 1-2: adjusted HR = 4.41; 95% CI, 1.65-11.82; P = .003), and salvage surgery for locoregional recurrences (locoregional vs local: adjusted HR = 3.81; 95% CI, 1.13-11.82; P = .021) were independent predictors for disease-free survival (DFS).Conclusion: Salvage surgery for larynx carcinoma, regardless of disease stage and other prognostic factors, results in more favorable loco(regional) control but not favorable DFS than pharyngeal carcinoma. The observed difference in DFS between salvage surgery for local and regional recurrences was not significant after correction for confounders. However, survival following salvage surgery for locoregional disease is significantly worse. For this subgroup, we propose to consider T status and comorbidity for clinical decision making, as high pT stage and ASA score are independent predictors for worse DFS. [ABSTRACT FROM AUTHOR]

Salivary duct carcinoma (SDC) is a subtype of salivary gland cancer with a dismal prognosis and a need for better prognostication and novel treatments. The aim of this national cohort study was to investigate clinical outcome, prognostic factors, androgen receptor (AR) and human epidermal growth factor receptor 2 (HER2) expression. SDC patients diagnosed between 1990 and 2014 were identified by the Nationwide Network and Registry of Histo‐ and Cytopathology in the Netherlands (PALGA). Subsequently, medical records were evaluated and pathological diagnoses reviewed. Data were analyzed for overall survival (OS), disease‐free survival (DFS), distant metastasis‐free survival (DMFS) and prognostic factors. AR was evaluated by immunohistochemistry (IHC), HER2 by IHC and fluorescent in‐situ hybridization. A total of 177 patients were included. The median age was 65 years, 75% were male. At diagnosis, 68% presented with lymph node metastases and 6% with distant metastases. Median OS, DFS and DMFS were 51, 23 and 26 months, respectively. In patients presenting without distant metastases, the absolute number of positive lymph nodes was associated with poor OS and DMFS in a multivariable analysis. AR and HER2 were positive in 161/168 (96%) and 44/153 (29%) tumors, respectively, and were not prognostic factors. SDC has a dismal prognosis with primary lymph node involvement in the majority of patients. The absolute number of lymph node metastases was found to be the only prognostic factor for DMFS and OS. AR expression and—to a lesser extent—HER2 expression hold promise for systemic treatment in the metastatic and eventually adjuvant setting. [ABSTRACT FROM AUTHOR]

Objective Staging of laryngeal cancer largely depends on cartilage invasion. Presence of cartilage invasion affects treatment choice and prognosis. On MRI and contrast-enhanced CT (CECT) it may be challenging to differentiate cartilage invasion from inflammation. The purpose of this study is to compare the diagnostic properties of dynamic contrast-enhanced CT (DCECT) and CECT for visual detection of cartilage invasion in laryngeal cancer. Study Design Prospective cohort study. Methods Patients with T3 or T4 laryngeal squamous cell carcinoma treated with total laryngectomy were evaluated using 0.625 mm slice CT. DCECT derived permeability and blood volume maps and CECT images were visually evaluated for the presence of invasion of the cartilaginous T-stage subsites of laryngeal cancer, by detecting continuity with the tumor-bulk of increased permeability, increased blood volume, and enhancement. Histological evaluation of the surgical total laryngectomy specimen served as the gold standard. Sensitivity, specificity, negative predictive value, and positive predictive value were calculated and compared using the McNemar and Chi-squared test. Results From 14 included patients, a total of 462 subsites were available for T-stage analys is, of which 84 were cartilage. The median time between CT imaging and total laryngectomy was 1 day (range 1-34 days). There was no significant difference in the detection of cartilage invasion between DCECT and CECT. The sensitivity of CECT was better for all subsites combined (0.85 vs. 0.75; p < 0.01). Conclusion DCECT does not improve visual detection of cartilage invasion in T3 and T4 laryngeal cancer compared to CECT. Level of Evidence 2b, individual cohort study. [ABSTRACT FROM AUTHOR]

In current molecular cancer diagnostics, using blood samples of cancer patients for the detection of genetic alterations in plasma (cell-free) circulating tumor DNA (ct DNA) is an emerging practice. Since ct DNA levels in blood are low, highly sensitive Droplet Digital PCR (dd PCR) can be used for detecting rare mutational targets. In order to perform dd PCR on blood samples, a standardized procedure for processing and analyzing blood samples is necessary to facilitate implementation into clinical practice. Therefore, we assessed the technical sample workup procedure for dd PCR on blood plasma samples. Blood samples from healthy individuals, as well as lung cancer patients were analyzed. We compared different methods and protocols for sample collection, storage, centrifugation, isolation, and quantification. Cell-free DNA (cf DNA) concentrations of several wild-type targets and BRAF and EGFR-mutant ct DNA concentrations quantified by dd PCR were primary outcome measurements. Highest cf DNA concentrations were measured in blood collected in serum tubes. No significant differences in cf DNA concentrations were detected between various time points of up to 24 h until centrifugation. Highest cf DNA concentrations were detected after DNA isolation with the Quick cf DNA Serum & Plasma Kit, while plasma isolation using the QIAamp Circulating Nucleic Acid Kit yielded the most consistent results. Dd PCR results on cf DNA are highly dependent on multiple factors during preanalytical sample workup, which need to be addressed during the development of this diagnostic tool for cancer diagnostics in the future. [ABSTRACT FROM AUTHOR]

Background Intraoperative analysis of lymph nodes during a sentinel lymph node biopsy (SLNB) procedure could result in one-step surgery for early oral squamous cell carcinoma (OSCC) with an occult nodal metastasis. One-step nucleic acid amplification rapidly detects cytokeratin 19 ( CK19) RNA with high accuracy. Sensitivity and specificity of CK19 expression in OSCC was evaluated. Methods Immunohistochemical CK19 expression was done in 207 patients with OSCC with 65 cases of paired nodal metastases. Results CK19 was expressed in 65% of all OSCC and even less in early OSCC (56%), with poor correlation between primary tumor and (occult) nodal metastasis. Furthermore, ectopic glandular tissue in close proximity of lymph nodes showed strong CK19 expression. Conclusion CK19 lacks both sensitivity and specificity as a biomarker for nodal metastasis in OSCC, which questions the suitability for CK19-based one-step nucleic acid amplification in SLNB procedures. Therefore, future studies should focus on other biomarkers, because the concept of fast intraoperative diagnostics during SLNB remains attractive. [ABSTRACT FROM AUTHOR]

Background Accurate nodal staging is pivotal for treatment planning in early (stage I-II) oral cancer. Unfortunately, current imaging modalities lack sensitivity to detect occult nodal metastases. Chromosomal region 11q13, including genes CCND1, Fas-associated death domain ( FADD), and CTTN, is often amplified in oral cancer with nodal metastases. However, evidence in predicting occult nodal metastases is limited. Methods In 158 patients with early tongue and floor of mouth (FOM) squamous cell carcinomas, both CCND1 amplification and cyclin D1, FADD, and cortactin protein expression were correlated with occult nodal metastases. Results CCND1 amplification and cyclin D1 expression correlated with occult nodal metastases. Cyclin D1 expression was validated in an independent multicenter cohort, confirming the correlation with occult nodal metastases in early FOM cancers. Conclusion Cyclin D1 is a predictive biomarker for occult nodal metastases in early FOM cancers. Prospective research on biopsy material should confirm these results before implementing its use in routine clinical practice. © 2016 Wiley Periodicals, Inc. Head Neck 39: 326-333, 2017 [ABSTRACT FROM AUTHOR]

Background There is an ongoing debate about the value of (neo-)adjuvant chemotherapy in high- and intermediate-grade osteosarcoma of the head and neck. Methods All records of patients older than 16 years diagnosed with osteosarcoma of the head and neck in the Netherlands between 1993 and 2013 were reviewed. Results We identified a total of 77 patients with an osteosarcoma of the head and neck; the 5-year overall survival (OS) was 55%. In 50 patients with surgically resected high- or intermediate-grade osteosarcoma of the head and neck younger than 75 years, univariate and multivariable analysis, adjusting for age and resection margins, showed that patients who had not received chemotherapy had a significantly higher risk of local recurrence (hazard ratio [HR] = 3.78 and 3.66, respectively). Conclusion In patients younger than 75 years of age with surgically resected high- and intermediate-grade osteosarcoma of the head and neck, treatment with (neo-)adjuvant chemotherapy resulted in a significantly smaller risk of local recurrence. Therefore, we suggest (neo-)adjuvant chemotherapy in patients amenable to chemotherapy. © 2016 The Authors Head & Neck Published by Wiley Periodicals, Inc. Head Neck 39: 140-146, 2017 [ABSTRACT FROM AUTHOR]

Aims Although high-grade dysplasia ( HGD) is a risk factor for malignant transformation and the future development of adenomas/carcinomas, grade is not incorporated in the Dutch guidelines for colonoscopy surveillance, partly because of presumed interobserver variability. The aim of this study was to analyse, in a nationwide cohort of colorectal adenomas, the interlaboratory variability in the grading of dysplasia in daily practice. Methods and results From the Dutch Pathology Registry, all synoptically reported classic adenomas in The Netherlands in 2013 were identified. The proportion of adenomas with HGD was determined for biopsies and polypectomies, and compared between 37 laboratories by the use of multivariable logistic regression analyses. In total, 21 030 colonoscopies of 20 270 patients were included. HGD was reported in 530 (3.6%) of 14 866 adenomas diagnosed on biopsies (range between laboratories: 0-13.6%) and in 983 (11.8%) of 8346 adenomas diagnosed on polypectomies (range: 3.1-42.9%). After adjustment for case mix, 13 (35%) laboratories reported a significantly lower or higher frequency of HGD than average. Conclusions We observed considerable interlaboratory variation in the grading of dysplasia in colorectal adenomas, which could be only partly explained by differences in case mix. Therefore, better standardization of grading criteria is needed before grade of dysplasia can usefully be incorporated in colonoscopy surveillance guidelines. [ABSTRACT FROM AUTHOR]

Background Identification of prognostic patient characteristics in head and neck squamous cell carcinoma (HNSCC) is of great importance. Human papillomavirus (HPV)-positive HNSCCs have favorable response to (chemo)radiotherapy. Apparent diffusion coefficient, derived from diffusion-weighted MRI, has also shown to predict treatment response. The purpose of this study was to evaluate the correlation between HPV status and apparent diffusion coefficient. Methods Seventy-three patients with histologically proven HNSCC were retrospectively analyzed. Mean pretreatment apparent diffusion coefficient was calculated by delineation of total tumor volume on diffusion-weighted MRI. HPV status was analyzed and correlated to apparent diffusion coefficient. Results Six HNSCCs were HPV-positive. HPV-positive HNSCC showed significantly lower apparent diffusion coefficient compared to HPV-negative. This correlation was independent of other patient characteristics. Conclusion In HNSCC, positive HPV status correlates with low mean apparent diffusion coefficient. The favorable prognostic value of low pretreatment apparent diffusion coefficient might be partially attributed to patients with a positive HPV status. © 2015 Wiley Periodicals, Inc. Head Neck 38: E613-E618, 2016 [ABSTRACT FROM AUTHOR]

Fibroblast growth factor receptor 3 ( FGFR3) is a member of the fibroblast growth factor receptor tyrosine kinase family. It has been identified as a promising therapeutic target in multiple types of cancer. We have investigated FGFR3 protein expression and FGFR3 gene copy-numbers in a single well-documented cohort of oral and oropharyngeal squamous cell carcinoma. Tissue microarray sets containing 452 formalin-fixed paraffin-embedded tissues were immunohistochemically stained with an anti- FGFR3 antibody and hybridized with a FGFR3 fluorescence in situ hybridization probe. FGFR3 protein expression was correlated with clinicopathological and survival data, which were retrieved from electronic medical records. FGFR3 mRNA data of 522 head and neck squamous cell carcinoma ( HNSCC) were retrieved from The Cancer Genome Atlas ( TCGA). Fibroblast growth factor receptor 3 ( FGFR3) protein was overexpressed in 48% (89/185) of oral and 59% (124/211) of oropharyngeal squamous cell carcinoma. Overexpression of FGFR3 protein was not related to overall survival or disease-free survival in oral ( HR[hazard ratio]: 0.94; 95% CI: 0.64-1.39; P = 0.77, HR: 0.94; 95% CI: 0.65-1.36; P = 0.75) and oropharyngeal squamous cell carcinoma ( HR: 1.21; 95% CI: 0.81-1.80; P = 0.36, HR: 0.42; 95% CI: 0.79-1.77; P = 0.42). FGFR3 mRNA was upregulated in 3% (18/522) of HNSCC from the TCGA. The FGFR3 gene was gained in 0.6% (1/179) of oral squamous cell carcinoma but no amplification was found in oral and oropharyngeal squamous cell carcinoma. In conclusion, FGFR3 protein is frequently overexpressed in oral and oropharyngeal squamous cell carcinoma. Therefore, it may serve as a potential therapeutic target for FGFR3-directed therapies in oral and oropharyngeal squamous cell carcinoma. [ABSTRACT FROM AUTHOR]

Current conventional treatment modalities in head and neck squamous cell carcinoma ( HNSCC) are nonselective and have shown to cause serious side effects. Unraveling the molecular profiles of head and neck cancer may enable promising clinical applications that pave the road for personalized cancer treatment. We examined copy number status in 36 common oncogenes and tumor suppressor genes in a cohort of 191 oropharyngeal squamous cell carcinomas ( OPSCC) and 164 oral cavity squamous cell carcinomas ( OSCC) using multiplex ligation probe amplification. Copy number status was correlated with human papillomavirus ( HPV) status in OPSCC, with occult lymph node status in OSCC and with patient survival. The 11q13 region showed gain or amplifications in 59% of HPV-negative OPSCC, whereas this amplification was almost absent in HPV-positive OPSCC. Additionally, in clinically lymph node-negative OSCC (Stage I-II), gain of the 11q13 region was significantly correlated with occult lymph node metastases with a negative predictive value of 81%. Multivariate survival analysis revealed a significantly decreased disease-free survival in both HPV-negative and HPV-positive OPSCC with a gain of Wnt-induced secreted protein-1. Gain of CCND1 showed to be an independent predictor for worse survival in OSCC. These results show that copy number aberrations, mainly of the 11q13 region, may be important predictors and prognosticators which allow for stratifying patients for personalized treatment of HNSCC. [ABSTRACT FROM AUTHOR]

ABSTRACT Background Gene expression profiling revealed a strong signature predicting lymph node metastases in oral squamous cell carcinoma (OSCC). Four of the most predictive genes are secretory leukocyte protease inhibitor (SLPI), lipocalin-2 (LCN2), thrombospondin-2 (THBS2), and tumor-associated calcium signal transducer 2 (TACSTD2). This study correlates their protein expression with lymph node metastases, overall survival (OS), and disease-specific survival (DSS). Methods Two hundred twelve patients with OSCC were included for protein expression analysis by immunohistochemistry. Results SLPI expression correlates with lymph node metastases in the whole cohort, not in a subgroup of cT1 to 2N0. SLPI expression correlates with OS (hazard ratio [HR] = 0.61) and DSS (HR = 0.47) in multivariate analysis. LCN2, THBS2, and TACSTD2 show no correlation with lymph node metastases, OS, or DSS. Conclusion Although SLPI expression correlates with lymph node metastases, it has no additional value in determining lymph node metastases in early oral cancer. However, it is an independent predictor for both OS and DSS and therefore a relevant prognostic biomarker in OSCC. © 2014 Wiley Periodicals, Inc. Head Neck 37: 1130-1136, 2015 [ABSTRACT FROM AUTHOR]

Awareness increases that the tumor biology influences treatment outcome and prognosis in cancer. Tumor hypoxia is thought to decrease sensitivity to radiotherapy and some forms of chemotherapy. Presence of hypoxia may be assessed by investigating expression of endogenous markers of hypoxia ( EMH) using immunohistochemistry (IHC). In this systematic review we investigated the effect of EMH expression on local control and survival according to treatment modality in head and neck cancer (head and neck squamous cell carcinoma [ HNSCC]). A search was performed in MEDLINE and EMBASE. Studies were eligible for inclusion that described EMH expression in relation to outcome in HNSCC patients. Quality was assessed using the Quality in Prognosis Studies (QUIPS) tool. Hazard ratios for locoregional control and survival were extracted. Forty studies of adequate quality were included. HIF-1a, HIF-2a, CA- IX, GLUT-1, and OPN were identified as the best described EMHs. With exception of HIF-2a, all EMHs were significantly related to adverse outcome in multiple studies, especially in studies where patients underwent single-modality treatment. Positive expression was often correlated with adverse clinical characteristics, including disease stage and differentiation grade. In summary, EMH expression was common in HNSCC patients and negatively influenced their prognosis. Future studies should investigate the effect of hypoxia-modified treatment schedules in patients with high In summary, EMH expression. These may include ARCON, treatment with nimorazole, or novel targeted therapies directed at hypoxic tissue. Also, the feasibility of surgical removal of the hypoxic tumor volume prior to radiotherapy should be investigated. [ABSTRACT FROM AUTHOR]

Resistance to treatment is the main problem of targeted treatment for cancer. We followed ten patients during treatment with vemurafenib, by three-dimensional imaging. In all patients, only a subset of lesions progressed. Next-generation DNA sequencing was performed on sequential biopsies in four patients to uncover mechanisms of resistance. In two patients, we identified mutations that explained resistance to vemurafenib; one of these patients had a secondary BRAF L505H mutation. This is the first observation of a secondary BRAF mutation in a vemurafenib-resistant patient-derived melanoma sample, which confirms the potential importance of the BRAF L505H mutation in the development of therapy resistance. Moreover, this study hints toward an important role for tumor heterogeneity in determining the outcome of targeted treatments. [ABSTRACT FROM AUTHOR]

The multikinase inhibitor sorafenib is under clinical investigation for the treatment of many solid tumors, but in most cases, the molecular target responsible for the clinical effect is unknown. Furthermore, enhancing the effectiveness of sorafenib using combination strategies is a major clinical challenge. Here, we identify sorafenib as an activator of AMP-activated protein kinase (AMPK), in a manner that involves either upstream LKB1 or CAMKK2. We further show in a phase II clinical trial in KRAS mutant advanced non-small cell lung cancer (NSCLC) with single agent sorafenib an improved disease control rate in patients using the antidiabetic drug metformin. Consistent with this, sorafenib and metformin act synergistically in inhibiting cellular proliferation in NSCLC in vitro and in vivo. A synergistic effect of both drugs is also seen on phosphorylation of the AMPKα activation site. Our results provide a rationale for the synergistic antiproliferative effects, given that AMPK inhibits downstream mTOR signaling. These data suggest that the combination of sorafenib with AMPK activators could have beneficial effects on tumor regression by AMPK pathway activation. The combination of metformin or other AMPK activators and sorafenib could be tested in prospective clinical trials. [ABSTRACT FROM AUTHOR]

Oropharyngeal squamous cell carcinoma ( OPSCC) is associated with human papillomavirus ( HPV) in a proportion of tumors. HPV-positive OPSCC is considered a distinct molecular entity with a prognostic advantage compared to HPV-negative cases. Silencing of cancer-related genes by DNA promoter hypermethylation may play an important role in the development of OPSCC. Hence, we examined promoter methylation status in 24 common tumor suppressor genes in a group of 200 OPSCCs to determine differentially methylated genes in HPV-positive versus HPV-negative primary OPSCC. Methylation status was correlated with HPV status, clinical features, and patient survival using multivariate methods. Additionally, methylation status of 16 cervical squamous cell carcinomas ( SCC) was compared with HPV-positive OPSCC. Using methylation-specific probe amplification, HPV-positive OPSCC showed a significantly higher cumulative methylation index ( CMI) compared to HPV-negative OPSCC ( P=0.008). For the genes CDH13, DAPK1, and RARB, both HPV-positive and HPV-negative OPSCC showed promoter hypermethylation in at least 20% of the tumors. HPV status was found to be an independent predictor of promoter hypermethylation of CADM1 ( P < 0.001), CHFR ( P = 0.027), and TIMP3 ( P < 0.001). CADM1 and CHFR showed similar methylation patterns in OPSCC and cervical SCC, but TIMP3 showed no methylation in cervical SCC in contrast to OPSCC. Methylation status of neither individual gene nor CMI was associated with survival. These results suggest that HPV-positive tumors are to a greater extent driven by promotor hypermethylation in these tumor suppressor genes. Especially CADM1 and TIMP3 are significantly more frequently hypermethylated in HPV-positive OPSCC and CHFR in HPV-negative tumors. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented in response to the article regarding rearranged during transfection (RET) protein as the molecular therapeutic target in multiple types of cancer.

Oncogenic fusion genes that involve kinases have proven to be effective targets for therapy in a wide range of cancers. Unfortunately, the diagnostic approaches required to identify these events are struggling to keep pace with the diverse array of genetic alterations that occur in cancer. Diagnostic screening in solid tumours is particularly challenging, as many fusion genes occur with a low frequency. To overcome these limitations, we developed a capture enrichment strategy to enable high-throughput transcript sequencing of the human kinome. This approach provides a global overview of kinase fusion events, irrespective of the identity of the fusion partner. To demonstrate the utility of this system, we profiled 100 non-small cell lung cancers and identified numerous genetic alterations impacting fibroblast growth factor receptor 3 ( FGFR3) in lung squamous cell carcinoma and a novel ALK fusion partner in lung adenocarcinoma. © 2013 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. [ABSTRACT FROM AUTHOR]

Myxofibrosarcoma and myxoid liposarcomas are relatively common soft tissue tumours that are characterized by their so-called myxoid extracellular matrix and have to some extent overlap in histology. The exact composition and potential role of their myxoid extracellular matrix are insufficiently understood. To gain more insight into the biomolecular content of these tumours, we have studied 40 well-documented myxofibrosarcoma and myxoid liposarcoma cases using imaging mass spectrometry. This technique provides a multiplex biomolecular imaging analysis of the tissue, spanning multiple molecular domains and without a priori knowledge of the tissue's biomolecular content. We have developed experimental protocols for analysing the peptide, protein, and lipid content of myxofibrosarcoma and myxoid liposarcomas, and have detected proteins and lipids that are tumour-type and tumour-grade specific. In particular, lipid changes observed in myxoid liposarcomas could be related to pathways known to be affected during tumour progression. Unsupervised clustering of the biomolecular signatures was able to classify myxofibrosarcoma and myxoid liposarcomas according to tumour type and tumour grade. Closer examination of histologically similar regions in the tissues revealed intratumour heterogeneity, which was a consistent feature in each of the myxofibrosarcomas studied. In intermediate-grade myxofibrosarcoma, it was found that single tissue sections could contain regions with biomolecular profiles similar to high-grade and low-grade tumours, and that these regions were associated with the tumour's nodular structure, thus supporting a concept of tumour progression through clonal selection. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Cellular myxoma and grade I myxofibrosarcoma are mesenchymal tumours that are characterized by their abundant myxoid extracellular matrix (ECM). Despite their histological overlap, they differ clinically. Diagnosis is therefore difficult though important. We investigated their (cyto) genetics and ECM. GNAS1-activating mutations have been described in intramuscular myxoma, and lead to downstream activation of cFos. KRAS and TP53 mutations are commonly involved in sarcomagenesis whereby KRAS subsequently activates c-Fos. A well-documented series of intramuscular myxoma (three typical cases and seven cases of the more challenging cellular variant) and grade I myxofibrosarcoma ( n= 10) cases were karyotyped, analyzed for GNAS1, KRAS and TP53 mutations and downstream activation of c-Fos mRNA and protein expression. ECM was studied by liquid chromatography mass spectrometry and expression of proteins identified was validated by immunohistochemistry and qPCR. Grade I myxofibrosarcoma showed variable, non-specific cyto-genetic aberrations in 83,5% of cases ( n= 6) whereas karyotypes of intramuscular myxoma were all normal ( n= 7). GNAS1-activating mutations were exclusively found in 50% of intramuscular myxoma. Both tumour types showed over-expression of c-Fos mRNA and protein. No mutations in KRAS codon 12/13 or in TP53 were detected. Liquid chromatography mass spectrometry revealed structural proteins (collagen types I, VI, XII, XIV and decorin) in grade I myxofibrosarcoma lacking in intramuscular myxoma. This was confirmed by immunohistochemistry and qPCR. Intramuscular/cellular myxoma and grade I myxofibrosarcoma show different molecular genetic aberrations and different composition of their ECM that probably contribute to their diverse clinical behaviour. GNAS1 mutation analysis can be helpful to distinguish intramuscular myxoma from grade I myxofibrosarcoma in selected cases. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented reporting six cases showing the association of chewing khat leaves with liver injury.