Assessment of the current and emerging criteria for the histopathological classification of lung neuroendocrine tumours in the lungNENomics project.

BACKGROUND: Six thoracic pathologists reviewed 259 lung neuroendocrine tumours (LNETs) from the lungNENomics project, with 171 of them having associated survival data. This cohort presents a unique opportunity to assess the strengths and limitations of current World Health Organization (WHO) classification criteria and to evaluate the utility of emerging markers. PATIENTS AND METHODS: Patients were diagnosed based on the 2021 WHO criteria, with atypical carcinoids (ACs) defined by the presence of focal necrosis and/or 2-10 mitoses per 2 mm2. We investigated two markers of tumour proliferation: the Ki-67 index and phospho-histone H3 (PHH3) protein expression, quantified by pathologists and automatically via deep learning. Additionally, an unsupervised deep learning algorithm was trained to uncover previously unnoticed morphological features with diagnostic value. RESULTS: The accuracy in distinguishing typical from ACs is hampered by interobserver variability in mitotic counting and the limitations of morphological criteria in identifying aggressive cases. Our study reveals that different Ki-67 cut-offs can categorise LNETs similarly to current WHO criteria. Counting mitoses in PHH3+ areas does not improve diagnosis, while providing a similar prognostic value to the current criteria. With the advantage of being time efficient, automated assessment of these markers leads to similar conclusions. Lastly, state-of-the-art deep learning modelling does not uncover undisclosed morphological features with diagnostic value. CONCLUSIONS: This study suggests that the mitotic criteria can be complemented by manual or automated assessment of Ki-67 or PHH3 protein expression, but these markers do not significantly improve the prognostic value of the current classification, as the AC group remains highly unspecific for aggressive cases. Therefore, we may have exhausted the potential of morphological features in classifying and prognosticating LNETs. Our study suggests that it might be time to shift the research focus towards investigating molecular markers that could contribute to a more clinically relevant morpho-molecular classification.

Pulmonary neuroendocrine neoplasms with well differentiated morphology and high proliferative activity: illustrated by a case series and review of the literature.

OBJECTIVES: Pulmonary neuroendocrine neoplasms (NENs) are subdivided in carcinoids and neuroendocrine carcinomas (small cell lung carcinoma and large cell neuroendocrine carcinoma (LCNEC)), based on the presence of necrosis and mitotic index (MI). However, it is unclear if tumors with well differentiated morphology but high proliferation rate should be regarded as LCNEC or as high grade carcinoids. In previous case series, a longer overall survival then expected in LCNEC has been suggested. We describe 7 of those cases analyzed for pRb expression and overall survival. MATERIAL AND METHODS: Cases with well differentiated morphology, but MI > 10/2mm2 and/or Ki-67 proliferation index >20% were selected based on pathology reports of consecutive NENs in our university medical center (Maastricht UMC+, 2007-2018) and confirmed by pathological review. Immunohistochemistry was performed to assess pRb expression. RESULTS: Seven stage IV cases were included in this study. Median overall survival was 8 months (95% confidence interval 5-11 months). Cases with well differentiated morphology and preserved pRb expression (4/7) had a median overall survival of 45 months. CONCLUSION: A subgroup of pulmonary NENs with well differentiated morphology but high proliferation rate likely exists. pRb staining might be helpful to predict prognosis, but clinical relevance remains to be studied.

Exploring imaging features of molecular subtypes of large cell neuroendocrine carcinoma (LCNEC).

OBJECTIVES: Radiological characteristics and radiomics signatures can aid in differentiation between small cell lung carcinoma (SCLC) and non-small cell lung carcinoma (NSCLC). We investigated whether molecular subtypes of large cell neuroendocrine carcinoma (LCNEC), i.e. SCLC-like (with pRb loss) vs. NSCLC-like (with pRb expression), can be distinguished by imaging based on (1) imaging interpretation, (2) semantic features, and/or (3) a radiomics signature, designed to differentiate between SCLC and NSCLC. MATERIALS AND METHODS: Pulmonary oncologists and chest radiologists assessed chest CT-scans of 44 LCNEC patients for 'small cell-like' or 'non-small cell-like' appearance. The radiologists also scored semantic features of 50 LCNEC scans. Finally, a radiomics signature was trained on a dataset containing 48 SCLC and 76 NSCLC scans and validated on an external set of 58 SCLC and 40 NSCLC scans. This signature was applied on scans of 28 SCLC-like and 8 NSCLC-like LCNEC patients. RESULTS: Pulmonary oncologists and radiologists were unable to differentiate between molecular subtypes of LCNEC and no significant differences in semantic features were found. The area under the receiver operating characteristics curve of the radiomics signature in the validation set (SCLC vs. NSCLC) was 0.84 (95% confidence interval (CI) 0.77-0.92) and 0.58 (95% CI 0.29-0.86) in the LCNEC dataset (SCLC-like vs. NSCLC-like). CONCLUSION: LCNEC appears to have radiological characteristics of both SCLC and NSCLC, irrespective of pRb loss, compatible with the SCLC-like subtype. Imaging interpretation, semantic features and our radiomics signature designed to differentiate between SCLC and NSCLC were unable to separate molecular LCNEC subtypes, which underscores that LCNEC is a unique disease.

Large cell neuroendocrine carcinoma with a solitary brain metastasis and low Ki-67: a unique subtype.

INTRODUCTION: Stage IV large cell neuroendocrine carcinoma (LCNEC) of the lung generally presents as disseminated and aggressive disease with a Ki-67 proliferation index (PI) 40-80%. LCNEC can be subdivided in two main subtypes: the first harboring TP53/RB1 mutations (small-cell lung carcinoma (SCLC)-like), the second with mutations in TP53 and STK11/KEAP1 (non-small-cell lung carcinoma (NSCLC)-like). Here we evaluated 11 LCNEC patients with only a solitary brain metastasis and evaluate phenotype, genotype and follow-up. METHODS: Eleven LCNEC patients with solitary brain metastases were analyzed. Clinical characteristics and survival data were retrieved from medical records. Pathological analysis included histomorphological analysis, immunohistochemistry (pRB and Ki-67 PI) and next-generation sequencing (TP53, RB1, STK11, KEAP1 and MEN1). RESULTS: All patients had N0 or N1 disease. Median overall survival (OS) was 12 months (95% confidence interval (CI) 5.5-18.5 months). Mean Ki-67 PI was 59% (range 15-100%). In 6/11 LCNEC Ki-67 PI was ≤40%. OS was longer for Ki-67 ≤40% compared to >40% (17 months (95% CI 11-23 months) vs 5 months (95% CI 0.7-9 months), P = 0.007). Two patients were still alive at follow-up after 86 and 103 months, both had Ki-67 ≤40%. 8/11 patients could be subclassified, and both SCLC-like (n = 6) and NSCLC-like (n = 2) subtypes were present. No MEN1 mutation was found. CONCLUSION: Stage IV LCNEC with a solitary brain metastasis and N0/N1 disease show in the majority of cases Ki-67 PI ≤40% and prolonged survival, distinguishing them from general LCNEC. This unique subgroup can be both of the SCLC-like and NSCLC-like subtype.

DLL3 expression in large cell neuroendocrine carcinoma (LCNEC) and association with molecular subtypes and neuroendocrine profile.

OBJECTIVES: For stage IV pulmonary large cell neuroendocrine carcinoma (LCNEC), the only therapeutic option is palliative chemotherapy. DLL3 is a new therapeutic target, which seems to be often expressed in SCLC and LCNEC. It has recently been reported that DLL3 mRNA expression is particularly upregulated in the LCNEC subgroup with STK11/KEAP1 and TP53 co-mutations, in contrast to lower expression levels in RB1 and TP53 co-mutated LCNEC. Our aim was to investigate DLL3 protein expression in stage IV LCNEC and correlate data with mutational profiles (i.e.STK11/KEAP1/RB1), immunostaining results (pRb, NE markers) and clinical characteristics. MATERIALS AND METHODS: Immunohistochemical analysis for DLL3 (SC16.65) and ASCL1 (SC72.201) was performed on 94 and 51 FFPE tissue sections, respectively, of pathologically reviewed stage IV LCNEC. DLL3 and ASCL1 were scored positive if ≥1% of the tumor cells showed cytoplasmic/membranous or dotlike (DLL3) or nuclear (ASCL1) immunostaining. Data were correlated with available sequencing (TP53, RB1, STK11, KEAP1), immunostaining (pRb, NE markers) and clinical data. RESULTS: DLL3 was expressed in 70/94 (74%) LCNEC, 56 (80%) of which showed cytoplasmic/membranous staining. Median H-score was 55 (interquartile range 0-160). DLL3 staining was not different in pRb immunohistochemistry negative and positive patients (DLL3+ in 53/70 (76%) vs. 14/21 (67%), p = 0.409) or RB1 mutated and wildtype patients (DLL3+ in 27/34 (79%) vs. 23/33 (70%), p = 0.361). Nevertheless, 6/6 (100%) STK11 mutated, 10/11 (91%) KEAP1 mutated and 9/9 (100%) TP53 wildtype tumors were DLL3+ . Furthermore, DLL3 expression was associated with expression of ASCL1 and at least 2 out of 3 neuroendocrine markers. CONCLUSION: The high percentage (74%) of DLL3 expression in stage IV LCNEC denotes the potential of DLL3 targeted therapy in this patient group.

ALK immunohistochemistry positive, FISH negative NSCLC is infrequent, but associated with impaired survival following treatment with crizotinib.

OBJECTIVE: Metastasized non-small cell lung cancer (NSCLC) with an anaplastic lymphoma kinase (ALK) rearrangement is usually sensitive to a range of ALK-tyrosine kinase inhibitors. ALK-positive NSCLC have been identified in pivotal phase III trials with fluorescence in situ hybridization (ALK FISH+). These tumors are also expressing the fusion product (ALK immunohistochemistry (IHC)+). However, discrepant cases occur, including ALK IHC + FISH-. The aim of this study was to collect ALK IHC + cases and compare within this group response to crizotinib treatment of ALK FISH + cases with ALK FISH- cases. MATERIALS AND METHODS: In this European prospective multicenter research study patients with Stage IV ALK IHC + NSCLC treated with crizotinib were enrolled. Tumor slides were validated centrally for ALK IHC and ALK FISH. RESULTS: Registration of 3523 ALK IHC tests revealed a prevalence of 2.7% (n = 94) ALK IHC + cases. Local ALK FISH analysis resulted in 48 concordant (ALK IHC+/FISH+) and 16 discordant (ALK IHC+/FISH-) cases. Central validation revealed 37 concordant and 7 discordant cases, 5 of which had follow-up. Validation was hampered by limited amount of tissue in biopsy samples. The PFS at 1 year for ALK concordant and discordant was 58% and 20%, respectively (HR = 2.4; 95% CI: 0.78-7.3; p = 0.11). Overall survival was significantly better for concordant cases than discordant cases after central validation (HR=4.5; 95% CI= 1.2-15.9; p=0.010. CONCLUSION: ALK IHC + FISH- NSCLC is infrequent and associated with a worse outcome on personalized treatment. A suitable predictive testing strategy may be to screen first with IHC and then confirm with FISH instead of considering ALK IHC equivalent to ALK FISH according to the current guidelines.

Prevalence and prognostic value of PD-L1 expression in molecular subtypes of metastatic large cell neuroendocrine carcinoma (LCNEC).

BACKGROUND: Pulmonary large cell neuroendocrine carcinoma (LCNEC) is a rare tumor with high mutational burden. Two subtypes of LCNEC are recognized, the co-mutated TP53 and RB1 group and the TP53 and STK11/KEAP1 group. We investigated PD-L1 and CD8 expression in a well characterized stage IV LCNEC cohort and compared expression in the two subtypes. METHODS: Immunohistochemical (IHC) analysis for PD-L1 and CD8 was performed on pathological reviewed pretreatment tumor samples for 148 stage IV LCNEC. Data about targeted next generation sequencing (TNGS) (TP53, RB1, STK11, KEAP1) and IHC for RB1 were available for most tumors. IHC staining for PD-L1 (DAKO 28-8) was performed and scored positive if tumors showed ≥1% membranous staining. CD8 was scored for intra-tumor T-cells and stromal cells. RESULTS: PD-L1 IHC expression data could be generated in 98/148 confirmed LCNEC samples along with RB1 IHC (n = 97) of which 77 passed quality control for TNGS. PD-L1 expression was positive in 16/98 cases (16%); 5 (5%) with ≥50%. PD-L1 expression was equal in RB1 mutated and RB1 wildtype tumors. None of STK11 mutated tumors (n = 7) expressed PD-L1. PD-L1 expression was correlated with superior overall survival (OS), hazard ratio 0.55 ((95% Confidence Interval 0.31-0.96), p = 0.038). Intra-tumor CD8 was associated with PD-L1 expression (p = 0.021) and stromal and intra-tumor CD8 were correlated with improved OS (p = 0.037 and p = 0.026 respectively). CONCLUSIONS: PD-L1 expression was positive in 16% of stage IV LCNEC tumors. This was independent of molecular subtype but associated with CD8 expression. In LCNEC patients with PD-L1 and/or CD8 expression superior OS was observed.

Methylation of L1RE1, RARB, and RASSF1 function as possible biomarkers for the differential diagnosis of lung cancer.

BACKGROUND: Lung cancer is the major cause of cancer-related deaths worldwide. Differential diagnosis can be difficult, especially when only small samples are available. Epigenetic changes are frequently tissue-specific events in carcinogenesis and hence may serve as diagnostic biomarkers. MATERIAL AND METHODS: 138 representative formalin-fixed, paraffin-embedded (FFPE) tissues (116 lung cancer cases and 22 benign controls) were used for targeted DNA methylation analysis via pyrosequencing of ten literature-derived methylation markers (APC, CDH1, CDKN2A, EFEMP1, FHIT, L1RE1, MGMT, PTEN, RARB, and RASSF1). Methylation levels were analyzed with the Classification and Regression Tree Algorithm (CART), Conditional Interference Trees (ctree) and ROC. Validation was performed with additional 27 lung cancer cases and 38 benign controls. TCGA data for 282 lung cancer cases was included in the analysis. RESULTS: CART and ctree analysis identified the combination of L1RE1 and RARB as well as L1RE1 and RASSF1 as independent methylation markers with high discriminative power between tumor and benign tissue (for each combination, 91% specificity and 100% sensitivity). L1RE1 methylation associated significantly with tumor type and grade (p<0.001) with highest methylation in the control group. The opposite was found for RARB (p<0.001). RASSF1 methylation increased with tumor type and grade (p<0.001) with strongest methylation in neuroendocrine tumors (NET). CONCLUSION: Hypomethylation of L1RE1 is frequent in tumors compared to benign controls and associates with higher grade, whereas increasing methylation of RARB is an independent marker for tumors and higher grade. RASSF1 hypermethylation was frequent in tumors and most prominent in NET making it an auxiliary marker for separation of NSCLC and NET. L1RE1 in combination with either RARB or RASSF1 could function as biomarkers for separating lung cancer and non-cancerous tissue and could be useful for samples of limited size such as biopsies.

Prevalence and clinical association of gene mutations through multiplex mutation testing in patients with NSCLC: results from the ETOP Lungscape Project.

Background: Reported prevalence of driver gene mutations in non-small-cell lung cancer (NSCLC) is highly variable and clinical correlations are emerging. Using NSCLC biomaterial and clinical data from the European Thoracic Oncology Platform Lungscape iBiobank, we explore the epidemiology of mutations and association to clinicopathologic features and patient outcome (relapse-free survival, time-to-relapse, overall survival). Methods: Clinically annotated, resected stage I-III NSCLC FFPE tissue was assessed for gene mutation using a microfluidics-based multiplex PCR platform. Mutant-allele detection sensitivity is >1% for most of the ∼150 (13 genes) mutations covered in the multiplex test. Results: Multiplex testing has been carried out in 2063 (76.2%) of the 2709 Lungscape cases (median follow-up 4.8 years). FFPE samples mostly date from 2005 to 2008, yet recently extracted DNA quality and quantity was generally good. Average DNA yield/case was 2.63 µg; 38 cases (1.4%) failed QC and were excluded from study; 95.1% of included cases allowed the complete panel of mutations to be tested. Most common were KRAS, MET, EGFR and PIK3CA mutations with overall prevalence of 23.0%, 6.8%, 5.4% and 4.9%, respectively. KRAS and EGFR mutations were significantly more frequent in adenocarcinomas: PIK3CA in squamous cell carcinomas. MET mutation prevalence did not differ between histology groups. EGFR mutations were found predominantly in never smokers; KRAS in current/former smokers. For all the above mutations, there was no difference in outcome between mutated and non-mutated cases. Conclusion: Archival FFPE NSCLC material is adequate for multiplex mutation analysis. In this large, predominantly European, clinically annotated stage I-III NSCLC cohort, none of the mutations characterized showed prognostic significance.

Female urothelial cell carcinoma in a failed kidney graft of a male recipient.

We present a case of a male kidney transplant patient harbouring two kidney grafts of which one is functional. In the failed graft, he developed urothelial cell carcinoma with cells containing XX-chromosome, and female tumour cells were also found in the bladder. The patient underwent donor nephrectomy, was treated with epirubicin bladder instillations, and immunosuppression was tapered. Less than a year before re-transplantation a CT scan showed no abnormalities of the first graft. Transplantectomy before a second kidney transplantation is debated.

Reviewing the current evidence supporting early B-cells as the cellular origin of Merkel cell carcinoma.

Merkel cell carcinoma (MCC) is a highly malignant skin cancer characterized by early metastases and poor survival. Although MCC is a rare malignancy, its incidence is rapidly increasing in the U.S. and Europe. The discovery of the Merkel cell polyomavirus (MCPyV) has enormously impacted our understanding of its etiopathogenesis and biology. MCCs are characterized by trilinear differentiation, comprising the expression of neuroendocrine, epithelial and B-lymphoid lineage markers. To date, it is generally accepted that the initial assumption of MCC originating from Merkel cells (MCs) is unlikely. This is owed to their post-mitotic character, absence of MCPyV in MCs and discrepant protein expression pattern in comparison to MCC. Evidence from mouse models suggests that epidermal/dermal stem cells might be of cellular origin in MCC. The recently formulated hypothesis of MCC originating from early B-cells is based on morphology, the consistent expression of early B-cell lineage markers and the finding of clonal immunoglobulin chain rearrangement in MCC cells. In this review we elaborate on the cellular ancestry of MCC, the identification of which could pave the way for novel and more effective therapeutic regimens.

Dichotomous ALK-IHC Is a Better Predictor for ALK Inhibition Outcome than Traditional ALK-FISH in Advanced Non-Small Cell Lung Cancer.

Purpose: ALK rearrangement detection using FISH is the standard test to identify patients with non-small cell lung carcinoma (NSCLC) eligible for treatment with ALK inhibitors. Recently, ALK protein expression in resectable NSCLC showed predictive value. We evaluated tumor response rate and survival after crizotinib treatment of patients with advanced NSCLC with ALK activation using both dichotomous immunohistochemical (IHC) staining and FISH.Experimental Design: Patients with stage IV NSCLC treated with crizotinib were selected. Tumor response was assessed. ALK rearrangements were detected by FISH (Vysis ALK-break-apart FISH-Probe KIT) and IHC [Ventana ALK (D5F3) CDx assay]. Cohorts of patients with ALK-FISH-positive advanced NSCLC from four other hospitals were used for validation.Results: Twenty-nine consecutive patients with ALK-positive advanced NSCLC diagnosed by FISH and/or IHC on small biopsies or fine-needle aspirations (FNA) were treated with ALK inhibitors. All ALK-IHC-positive patients responded to crizotinib except three with primary resistance. No tumor response was observed in 13 ALK-FISH-positive but ALK-IHC-negative patients. This was confirmed in an external cohort of 16 patients. Receiver operator characteristic (ROC) curves for ALK-IHC and ALK-FISH compared with treatment outcome showed that dichotomous ALK-IHC outperforms ALK-FISH [tumor response area under the curve: (AUC), 0.86 vs. 0.64, P = 0.03; progression-free survival (PFS): AUC 0.86 vs. 0.36, P = 0.005; overall survival (OS): AUC, 0.78 vs. 0.41, P = 0.01, respectively].Conclusions: Dichotomous ALK-IHC is superior to ALK-FISH on small biopsies and FNA to predict tumor response and survival to crizotinib for patients with advanced NSCLC. Our data strongly suggest adapting the guidelines and using dichotomous ALK-IHC as standard companion diagnostic test to select patients with NSCLC who benefit from ALK-targeting therapy. Clin Cancer Res; 23(15); 4251-8. ©2017 AACR.

Postzygotic mosaicism in basal cell naevus syndrome.

Basal cell naevus syndrome (BCNS) is an autosomal dominant disorder most commonly caused by a germline mutation in the Drosophila homologue of patched-1 gene (PTCH1). Here we describe a patient with clinical signs of BCNS, caused by postzygotic mosaicism of a PTCH1 mutation. We performed restriction fragment length polymorphism analysis and Droplet Digital polymerase chain reaction to determine the degree of mosaicism in different tissues of this patient. Our case shows that a relatively low-grade mosaicism can lead to clinical signs reminiscent of those caused by a germline mutation. This finding has important implications for genetic counselling and therefore is pivotal to recognize for dermatologists, as well as for clinical geneticists and clinical laboratory geneticists.

Different prognostic models for different patient populations: validation of a new prognostic model for patients with oropharyngeal cancer in Western Europe.

OBJECTIVE: The presence of human papillomavirus (HPV) infection in oropharyngeal squamous cell carcinoma (OPSCC) is a major determinant in prognostic risk modelling. Recently, a prognostic model was proposed in which HPV status, comorbidity and nodal stage were the most important prognostic factors to determine high-, intermediate- and low-risk survival groups. Here, we report on the validation of this model using an independent single-institutional cohort. METHODS: A total number of 235 patients curatively treated for OPSCC in the period 2000-2011 at the MUMC (Maastricht University Medical Center, The Netherlands) were included. The presence of an oncogenic HPV infection was determined by p16 immunostaining, followed by a high-risk HPV DNA PCR on the p16-positive cases. The model variables included were HPV status, comorbidity and nodal stage. As a measure of model performance, the Harrell's Concordance index (Harrell's C-index) was used. RESULTS: The 5-year overall survival (OS) estimates were 84.6%, 54.5% and 28.7% in the low-, intermediate- and high-risk group, respectively. The difference between the survival curves was highly significant (P<0.001). The Harrell's C-index was 0.69 (95% confidence interval (CI): 0.63-0.75). CONCLUSION: In this study a previously developed prognostic risk model was validated. This model will help to personalise treatment in OPSCC patients. This model is publicly available at www.predictcancer.org.

Detection of HPV-associated oropharyngeal tumours in a 16-year cohort: more than meets the eye.

BACKGROUND: Accurate assessment of the prevalence of the human papilloma virus (HPV) in oropharyngeal tumours (OpSCC) is important because HPV-positive OpSCC are consistently associated with an improved overall survival. Recently, an algorithm has become available that reliably detects clinically relevant HPV in tumour tissue, however, no complete cohorts have been tested. The aim was to determine the prevalence of active high-risk HPV infection in a complete cohort of OpSCC collected over a 16-year period. METHODS: Using a triple algorithm of p16 immunohistochemistry, HPV-BRISH and HPV-PCR, we assessed the prevalence of active HPV infection in all OpSCC diagnosed in our hospital from 1997 to 2012 (n=193) and a random selection of 200 oral tumours (OSCC). RESULTS: Forty-seven OpSCC (24%) were HPVGP PCR-positive; 42 cases were HPV16+, 1 HPV18+, 3 HPV33+ and 1 HPV35+. Brightfield in situ hybridisation did not identify additional HPV-positive cases. Human papilloma virus-associated tumour proportion increased from 13% (1997-2004) to 30% (2005-2012). Human papilloma virus-positivity was an independent predictor for longer disease-specific survival (HR=0.22; 95%CI:0.10-0.47). Only one OSCC was HPV+. CONCLUSIONS: In our cohort, the incidence of HPV-associated OpSCC is low but increasing rapidly. The strict detection algorithm, analysis of disease-specific survival and the complete cohort, including palliatively treated patients, may influence the reported prevalence and prognostic value of HPV in OpSCC.

EGFR mutated non-small cell lung cancer patients: more prone to development of bone and brain metastases?

OBJECTIVES: Both bone and brain are frequent sites of metastasis in non-small cell lung cancer (NSCLC). Conflicting data exist whether EGFR mutant (+) patients are more prone to develop brain metastases or have a better outcome with brain metastases compared to EGFR/KRAS wildtype (WT) or KRAS+ patients. For bone metastases this has not been studied. METHODS: In this retrospective case-control study all EGFR+ (exons 19 and 21) patients diagnosed at two pathology departments were selected (2004/2008 to 2012). For every EGFR+ patient a consecutive KRAS+ and WT patient with metastatic NSCLC (mNSCLC) was identified. Patients with another malignancy within 2 years of mNSCLC diagnosis were excluded. Data regarding age, gender, performance score, histology, treatment, bone/brain metastases diagnosis, skeletal related events (SRE) and subsequent survival were collected. RESULTS: 189 patients were included: 62 EGFR+, 65 KRAS+, 62 WT. 32%, 35% and 40%, respectively, had brain metastases (p=0.645). Mean time to brain metastases was 20.8 [± 12.0], 10.8 [± 9.8], 16.4 [± 10.2] months (EGFR+-KRAS+, p = 0.020, EGFR+-WT, p = 0.321). Median post brain metastases survival was 12.1 [5.0-19.1], 7.6 [1.2-14.0], 10.7 [1.5-19.8] months (p = 0.674). 60%, 52% and 50% had metastatic bone disease (p=0.528). Mean time to development of metastatic bone disease was 13.4 [± 10.6], 23.3 [± 19.4], 16.4 [± 9.6] months (p = 0.201). Median post metastatic bone disease survival was 15.0 [10.6-20.3], 9.0 [5.2-12.9], 3.2 [0.0-6.9] months (p = 0.010). Time to 1st SRE was not significantly different. CONCLUSIONS: Incidence of brain and bone metastases was not different between EGFR+, KRAS+ and WT patients. Post brain metastases survival, time from mNSCLC diagnosis to metastatic bone disease and 1st SRE did not differ either. Post metastatic bone disease survival was significantly longer in EGFR+ patients. Although prevention of SRE's is important for all patients, the latter finding calls for a separate study for SRE preventing agents in EGFR+ patients.

Chromosome instability predicts the progression of premalignant oral lesions.

OBJECTIVES: One of the main problems in reducing the incidence of oral squamous cell carcinoma (OSCC) is the inability to appropriately deal with leukoplakia. Accurately identifying lesions which will progress to malignancy is currently not possible. The present study aims to establish the value of chromosome instability (CI) detection by DNA image cytometry and FISH analysis for prognosis and monitoring of oral leukoplakia. MATERIALS AND METHODS: For this purpose, we included from our archives 102 oral leukoplakia cases, which had been diagnosed between 1991 and 2008. Patient follow-up data were collected and the histopathological diagnosis was revised. CI assessment was carried out on paraffin-embedded tissue sections using both DNA image cytometry (ICM) and dual target FISH for chromosomes 1 and 7. RESULTS: 16 of 102 Patients developed carcinoma in situ or OSCC. Both detection methods were found to yield prognostic information independent of the histopathological diagnosis. CI was a strong individual marker of progression, with hazard ratios (HRs) of 7.2 and 6.8 for ICM and FISH respectively. Moreover, this approach seems suitable for monitoring lesions over time (especially ICM). Combining histopathology and CI enables subdivision of patients into three risk groups, with different probabilities of malignant progression. CONCLUSION: CI detection seems a reliable method for risk assessment of oral premalignancies and its application may contribute to a better risk-counselling and appropriate treatment regimen or watchfull-waiting approach of patients.

Human Papillomavirus and Oropharyngeal Squamous Cell Carcinoma: A Case-Control Study regarding Tobacco and Alcohol Consumption.

We aimed to determine the role of HPV in the pathogenesis and outcome of oropharyngeal squamous cell carcinoma (OSCC) in lifelong nonsmoking and nondrinking patients. A case-case analysis was performed to compare the presence of HPV-DNA in tumor cells of 16 nonsmoking and nondrinking with 16 matched smoking and drinking patients (matching criteria: age at incidence, gender, tumor sublocation, tumor stage). HPV was detected using 2 PCR tests, FISH analysis, and p16(INK4A) immunostaining. Nonsmoking and nondrinking patients had more HPV-positive tumors than smoking and drinking patients (n = 12; 75% versus n = 2; 13%; P < 0.001). All HPV-positive tumors showed p16(INK4A) overexpression, and 1 HPV-negative tumor had p16(INK4A) overexpression, (P < 0.001). Overall survival and disease-specific survival were higher for HPV-positive compared to HPV-negative cases (P = 0.027, P = 0.039, resp.). In conclusion, HPV is strongly associated with OSCC of nonsmoking and nondrinking patients. Specific diagnostic and therapeutic actions should be considered for these patients to achieve a better prognosis.

[Human papillomavirus and cancer of the oropharynx. Molecular interaction and clinical implications]. / Humane Papillomviren und Oropharynxkarzinome. Molekulare Interaktion und klinische Auswirkung.

One-third of the cases of oropharyngeal squamous cell carcinoma (OSCC) contain oncogenic human papillomavirus (HR-HPV). Epidemiologic and molecular evidence underlines the causal role of HR-HPV in these tumors, which can be defined as HPV-related OSCC. These tumors differ from chemical/toxin-induced OSCC in several biological aspects, including specific molecular and genetic alterations. This leads to a characteristic clinical profile of HPV-related OSCC. Sexual risk factors play a role; however, the knowledge about natural infection and the rate of persistence of HR-HPV in the oropharynx is marginal. It is shown that the distinct biological behavior of the HPV-related subset of oropharyngeal tumors results in a more favorable prognosis. This might be the result of a better response to chemotherapy and radiotherapy. However, further studies are needed to show whether it will be possible to reliably select patients for individualized therapy depended on the HPV status of their tumors. Therefore, we think it will be mandatory to consider and stratify HPV status in the design of prospective clinical trials in the future.