Incorporation of a Molecular Prognostic Classifier Improves Conventional Non-Small Cell Lung Cancer Staging.

INTRODUCTION: Despite adoption of molecular biomarkers in the management of NSCLC, the recently adopted eighth edition of the TNM staging system utilized only clinicopathologic characteristics and validated improvement in risk stratification of early-stage disease has remained elusive. We therefore evaluated the integration of a clinically validated molecular prognostic classifier into conventional staging. METHODS: A novel staging system, the TNMB (with the B denoting biology) system, which integrates a 14-gene molecular prognostic classifier into the eighth edition of the TNM staging system, was developed by using data from 321 patients with NSCLC at the University of California, San Francisco. The TNMB staging system was subsequently validated in an independent, multicenter cohort of 1373 patients, and its implementation was compared with adoption of the seventh and eighth edition staging systems utilizing metrics of reclassification. RESULTS: Compared with staging according to the eighth edition of the TNM system, the TNMB staging system enhanced the identification of high-risk patients, with a net reclassification improvement of 0.33 (95% confidence interval [CI]: 0.24-0.41). It better predicted differences in survival, with a relative integrated discrimination improvement of 22.1% (95% CI: 8.8%-35.3%), and it improved agreement between observed and predicted survival, with a decrease in the reclassification calibration statistic of from 39 to 21. The seventh and eighth editions failed to change the net reclassification improvement (0.01 [95% CI: -0.04 to 0.03] and 0.03 [95% CI: 0.00 to 0.06], respectively) or relative integrated discrimination improvement (2.1% [95% CI: -5.8 to 9.9] and -2.5% [95% CI: -17.6 to 12.4], respectively); in addition, the eighth edition worsened calibration, with an increase in the reclassification calibration statistic from 23 to 25. CONCLUSIONS: Incorporation of a molecular prognostic classifier significantly improved identification of high-risk patients and survival predictions compared with when conventional staging is used. The TNMB staging system may lead to improved survival of early-stage disease through more effective application of adjuvant therapy.

The Landscape of Actionable Molecular Alterations in Immunomarker-Defined Large-Cell Carcinoma of the Lung.

INTRODUCTION: Patients with pulmonary large-cell carcinoma (LCC) have poor prognosis and limited treatment options. The identification of clinically actionable molecular alterations helps to guide personalized cancer treatment decisions. PATIENTS AND METHODS: A consecutive cohort of 789 resected NSCLC cases were reviewed. Fifty-nine NSCLC cases lacking morphologic differentiation, accounting for 7.5% of all resected NSCLCs, were identified and further characterized by immunohistochemistry according to the 2015 WHO lung tumor classification. Molecular alterations were investigated by multiple technologies including target capture sequencing, immunohistochemistry, and fluorescence in situ hybridizations. RESULTS: Of 59 NSCLC cases lacking morphologic differentiation, 20 (33.9%) were reclassified as adenocarcinoma (LCC-AD), 14 (23.7%) as squamous cell carcinoma (LCC-SqCC), and 25 (42.4%) as LCC-Null. Approximately 92% of LCC-Null, 95% of LCC-AD, and 86% of LCC-SqCC harbored clinically relevant alterations. Alterations characteristic of adenocarcinoma (EGFR, KRAS, ALK receptor tyrosine kinase [ALK], ROS1, and serine/threonine kinase 11 [STK11]) were detected in the LCC-AD subgroup but not in LCC-SqCC, whereas squamous-lineage alterations (phosphatidylinositol-4,5-biphosphate 3-kinase catalytic subunit alpha [PIK3CA], SRY-box 2 [SOX2], fibroblast growth factor receptor 1 [FGFR1], and AKT1) were detected in the LCC-SqCC subgroup but not in the LCC-AD group. Although some LCC-Null tumors displayed a genetic profile similar to either adenocarcinoma or squamous-cell carcinoma, more than half of the LCC-Null group were completely devoid of recognizable lineage-specific genetic profiles. High programmed death ligand 1 expression and high frequency of cell cycle regulatory gene alterations were found in the LCC-Null group offering alternative options of targeted therapy. CONCLUSIONS: This comprehensive molecular study provided further insight into the genetic architecture of LCC. The presence of clinically actionable alterations in a majority of the tumors allowed personalized treatment to emerge.

Clinicopathological analysis of Large Cell Lung Carcinomas definitely diagnosed according to the New World Health Organization Criteria.

OBJECTIVE: The definition of large cell lung carcinoma (LCC) has undergone an extensive modification in the World Health Organization (WHO) Classification (2015). Present study aimed to investigate the clinicopathological characteristics of patients diagnosed as LCC according to current WHO criteria. METHODS: LCCs diagnosed based on the previous WHO classification were reevaluated, and 17 cases of LCC were finally identified at Peking Union Medical College Hospital and Beijing Chest Hospital between 2009 and 2015. The clinicopathologic features were examined and EGFR and KRAS mutations were tested. Survival of the patients was analyzed by Kaplan-Meier method. RESULTS: The median age of the patients was 64 years (range: 40-78). Most patients were male (64.7%) and about half of the patients were at TNM stage III (47.1%). Morphologically, most cases (70.6%) were classic LCC. All patients were treated by lobectomy plus lymph node dissection, 2 with bi-lobectomy and 1 with complex lobectomy, and the other 2 patients were further treated by partial pericardiotomy. Ten patients received postoperative chemotherapy, while only 3 patients were treated with radiotherapy after surgery. Molecular analysis showed two cases of EGFR mutation (L858R) but without non-overlapping KRAS mutation. The 3-year overall survival rate was 48.4 ±â€¯15.1%. Chemotherapy was the only predictive factor that is associated with the prognosis of the patients (P = 0.003). CONCLUSION: The clinicopathological characteristics of 17 cases of stringently diagnosed LCC were retrospectively analyzed. LCC in our study showed aggressive behavior with high recurrence and metastasis and poor prognosis. Chemotherapy was only predictive factor that is significantly associated with the prognosis of the patients. Future studies based on a larger series and long term follow-up are still needed to characterize it further.

Pulmonary large cell neuroendocrine carcinoma with adenocarcinoma-like features: napsin A expression and genomic alterations.

Pulmonary large cell neuroendocrine carcinoma (LCNEC) is a highly aggressive malignancy, which was recently found to comprise three major genomic subsets: small cell carcinoma-like, non-small cell carcinoma (predominantly adenocarcinoma)-like, and carcinoid-like. To further characterize adenocarcinoma-like subset, here we analyzed the expression of exocrine marker napsin A, along with TTF-1, in a large series of LCNECs (n=112), and performed detailed clinicopathologic and genomic analysis of napsin A-positive cases. For comparison, we analyzed napsin A expression in other lung neuroendocrine neoplasms (177 carcinoids, 37 small cell carcinomas) and 60 lung adenocarcinomas. We found that napsin A was expressed in 15% of LCNEC (17/112), whereas all carcinoids and small cell carcinomas were consistently negative. Napsin A reactivity in LCNEC was focal in 12/17 cases, and weak or moderate in intensity in all cases, which was significantly lower in the extent and intensity than seen in adenocarcinomas (P<0.0001). The combination of TTF-1-diffuse/napsin A-negative or focal was typical of LCNEC but was rare in adenocarcinoma, and could thus serve as a helpful diagnostic clue. The diagnosis of napsin A-positive LCNECs was confirmed by classic morphology, diffuse labeling for at least one neuroendocrine marker, most consistently synaptophysin, and the lack of distinct adenocarcinoma component. Genomic analysis of 14 napsin A-positive LCNECs revealed the presence of mutations typical of lung adenocarcinoma (KRAS and/or STK11) in 11 cases. In conclusion, LCNECs are unique among lung neuroendocrine neoplasms in that some of these tumors exhibit low-level expression of exocrine marker napsin A, and harbor genomic alterations typical of adenocarcinoma. Despite the apparent close biological relationship, designation of adeno-like LCNEC as a separate entity from adenocarcinoma is supported by their distinctive morphology, typically diffuse expression of neuroendocrine marker(s) and aggressive behavior. Further studies are warranted to assess the clinical utility and optimal method of identifying adenocarcinoma-like and other subsets of LCNEC in routine practice.

[Clinicopathologic features and genetic profile of the redefined large cell lung carcinoma].

Objective: To investigate the clinicopathologic features and genetic profile of large cell lung carcinoma (LCC) redefined by new classification. Methods: Basing on 2015 WHO classification criteria in redefining large cell lung carcinoma, the expression of specific markers (TTF1, Napsin A, p40, CK5/6, CK, vimentin and ZEB1) was detected by immunohistochemistry and D-PAS staining in 303 surgically-removed lung specimens previously diagnosed as large cell lung carcinoma. The clinicopathologic and genetic characteristics (including EGFR, KRAS, BRAF, ALK and ROS1 gene mutation) were analyzed. Results: Based on the new definition of LCC, 116 cases (116/303, 38.3%) of LCC formerly diagnosed were reclassified as solid adenocarcinoma, 49 cases (49/303, 16.2%) as squamous cell carcinoma, 6 cases (6/303, 2.0%) as adenosquamous carcinoma, 22 cases (22/303, 7.3%) as spindle cell carcinoma and only 110 cases (110/303, 36.3%) as large cell carcinoma. Redefined LCCs were characterized as middle-age (range 40-80), male (102/110, 92.7%) and smoking patients (64/110, 58.2%) with intermediate-advanced stage. Among 110 cases, 9 cases with EGFR mutation and 10 cases with KRAS mutation and 1 case with ALK fusion were found. No BRAF and ROS1 alterations were identified. Conclusions: According to the new classification, LCCs formerly diagnosed are mostly reclassified as adenocarcinoma and non-keratinizing squamous cell carcinoma. The newly defined LCC may significantly benefit from clinical therapy.

[Grading of lung cancer]. / Grading von Lungenkarzinomen.

In comparison with other tumor entities there is no common generally accepted grading system for lung cancer with clearly defined criteria and clinical relevance. In the recent fourth edition of the World Health Organization (WHO) classification from 2015 of tumors of the lungs, pleura, thymus and heart, there is no generally applicable grading for pulmonary adenocarcinomas, squamous cell carcinomas or rarer forms of carcinoma. Since the new IASLC/ATS/ERS classification of adenocarcinomas published in 2011, 5 different subtypes with significantly different prognosis are proposed. This results in an architectural (histologic) grading, which is usually applied to resection specimens. For squamous cell carcinoma the number of different histological subtypes in the new WHO classification was reduced compared to earlier versions but without a common grading system. In recent publications nesting and budding were proposed as the main (histologic) criteria for a grading of squamous cell carcinomas. The grading of neuroendocrine tumors (NET) of the lungs in comparison with NET in other organs is presented in a separate article in this issue. Certain rare tumor types are high grade per definition: small cell, large cell and pleomorphic carcinomas, carcinosarcomas and pulmonary blastomas. In the future it is to be expected that these developments will be further refined, e. g. by adding further subtypes for adenocarcinomas and cytologic and/or nuclear criteria for adenocarcinoma and/or squamous cell carcinomas.

[Large-cell lung carcinoma by the 2015 WHO classification: a clinicopathologic analysis of 93 cases].

OBJECTIVE: To analyze the clinicopathologic characteristics of large-cell lung carcinoma (LCC) subtypes according to the 2015 WHO classification. METHODS: A total of 93 cases of LCC were identified among 1 634 cases of morphologically undifferentiated non-small cell lung carcinoma. Clinicopathologic and immunohistochemical characteristics were retrospectively reviewed. RESULTS: Among 93 cases of LCC, 50 cases were solid adenocarcinoma (53.8%), 21 cases were non-keratinzing squamous cell carcinoma (22.6%), 3 cases were adenosquamous carcinoma (3.2%), 8 cases were large cell carcinoma with unclear immunohistochemical features (8.6%), and 11 cases were large cell carcinoma with null immunohistochemical features (11.8%). No significant differences in clinical features were found among various immunophenotypes. The overall survival of 1, 3 and 5 years of all cases was 51.0%, 5.4% and 1.1%, respectively and the mean survival was (15.2±1.2)months. Notably the null type had the lowest survival rate of 18% in one year, significantly worse than those of solid adenocarcinoma and non-keratinzing squamous cell carcinoma. CONCLUSIONS: The 2015 WHO classification based on immunohistochemical phenotype replaces the morphological approach in separating poorly differentiated non-small cell lung carcinoma from the former large cell lung carcinoma. The null immunohistochemical subtype may represent a distinct subtype of undifferentiated carcinoma with the worst prognosis.

The Impact of the Fourth Edition of the WHO Classification of Lung Tumours on Histological Classification of Resected Pulmonary NSCCs.

INTRODUCTION: Histopathological classification of lung cancer is of central importance in the diagnostic routine, and it guides therapy in most patients. The fourth edition of the World Health Organization (WHO) Classification of Lung Tumours was recently published and includes changes to the diagnostic procedure for non-small cell carcinomas (NSCCs), with more emphasis on immunohistochemical (IHC) staining. METHODS: A total of 656 unselective cases of resected pulmonary NSCC were diagnosed according to the 2004 WHO classification. After IHC staining with cytokeratin 5, p40, p63, thyroid transcription factor 1 (clones 8G7G3/1 and SPT24), and napsin A, the diagnoses were revised in accordance with the new fourth edition of the WHO classification. RESULTS: Reclassification led to a new histological annotation in 36 of the 656 cases (5%). Most notable was the decrease in cases previously classified as large cell carcinomas (56 versus 12 cases). This was partially due to the exclusion of 21 neuroendocrine tumors from this group, with 20 cases ascribed to the adenocarcinoma group on the basis of IHC markers. Only seven cases of adenocarcinoma or squamous cell carcinoma were reclassified after the addition of IHC staining. There was a substantial overlap in staining properties between different markers of squamous and adenocarcinomatous differentiation, respectively, but in 17 to 31 cases (3%-5%), the diagnosis depended on the choice of markers. CONCLUSIONS: The fourth edition of the WHO Classification of Lung Tumours leads to changes in histological type in 5% of resected NSCCs. The incorporation of IHC staining into NSCC diagnostics demands awareness that the choice of ancillary stains has an effect on diagnosis.

[Large cell carcinoma, lymphoepithelioma-like carcinoma, NUT carcinoma]. / Carcinome à grandes cellules, carcinome lymphoepithelioma-like, carcinome NUT.

The diagnosis of large cell carcinoma can only be made on a sampled resected tumor and should not be applied to biopsies or cytology. In the 2015 WHO classification, the definition of large cell carcinoma is restricted to carcinomas both lacking morphological signs of glandular, squamous or neuroendocrine differentiation and exhibiting a null or unclear phenotype (TTF1-/p40 ou p63 ou CK5/6+ focally). These carcinomas have an adenocarcinoma molecular profile because they harbor a significant number of KRAS and BRAF mutations, a profile that is more similar to adenocarcinoma than squamous cell carcinoma. They also have a worse prognosis than the other types of non-small cell lung carcinoma. Many large cell carcinomas previously classified on morphological data alone are now reclassified in the adenocarcinoma and squamous cell carcinoma types, including immunohistochemical features. The other large cell carcinoma subtypes from the 2004 WHO classification, i.e. large cell neuroendocrine carcinoma and basaloid carcinoma, are grouped respectively with the other neuroendocrine tumors and squamous cell carcinomas. Clear cell and rhabdoid features are now considered as cytological variants that can occur in any histopathological subtype and not as distinct subtypes. Lymphoepithelioma-like carcinoma is moved to the group of other and unclassified carcinomas as NUT carcinoma.

Next-Generation Sequencing of a Cohort of Pulmonary Large Cell Carcinomas Reclassified by World Health Organization 2015 Criteria.

CONTEXT: The classification of pulmonary large cell carcinoma has undergone a major revision with the recent World Health Organization (WHO) 2015 Classification. Many large cell carcinomas are now reassigned to either adenocarcinoma with solid pattern or nonkeratinizing squamous cell carcinoma based on immunopositivity for adenocarcinoma markers or squamous cell carcinoma markers, respectively. Large cell carcinomas that are negative for adenocarcinoma and squamous cell carcinoma immunomarkers are now classified as large cell carcinoma with null immunohistochemical features (LCC-N). Although a few studies investigated the mutation profile of large cell carcinomas grouped by immunostain profile before the publication of the new WHO classification, investigation of tumors previously diagnosed as large cell carcinoma and reclassified according to the 2015 WHO classification has not, to our knowledge, been reported. OBJECTIVE: To determine the mutation profiles of pulmonary large cell carcinomas reclassified by WHO 2015 criteria. DESIGN: Archival cases of non-small cell lung carcinoma with large cell carcinoma morphology (n = 17) were reclassified according to 2015 WHO criteria. To determine mutation profile, we employed Ion Torrent (Life Technologies, Carlsbad, California)-based next-generation sequencing (50 genes; more than 2800 mutations) in addition to real-time quantitative reverse transcription polymerase chain reaction for ALK translocation detection. RESULTS: Two of 17 cases (12%) were reclassified as LCC-N, and both had mutations-BRAF D594N in one case and KRAS G12C in the other case. Seven of 17 cases (41%) were reclassified in the adenocarcinoma with solid pattern group, which showed one KRAS G12C and one EGFR E709K + G719C double mutation in addition to mutations in TP53. Eight of 17 cases (47%) were reclassified in the nonkeratinizing squamous cell carcinoma group, which showed mutations in PIK3CA, CDKN2A, and TP53. No ALK translocations or amplifications were detected. CONCLUSIONS: The adenocarcinoma with solid pattern group showed mutations typical of adenocarcinoma, whereas the nonkeratinizing squamous cell carcinoma group showed mutations typical of squamous cell carcinoma. Both LCC-N cases had mutations associated with adenocarcinoma, supporting the hypothesis that LCC-N is related to adenocarcinoma.

Treatment outcomes of patients with different subtypes of large cell carcinoma of the lung.

BACKGROUND: Although large cell neuroendocrine carcinoma (LCNEC) and lymphoepithelioma-like carcinoma (LELC) are the variants of large cell carcinoma (LCC) of lung, there are few studies comparing them. The aim of this study was to compare the clinical characteristic and treatment outcomes of LCNEC, LELC, and classic LCC. METHODS: Patients with LCNEC, LELC, or classic LCC were identified in a prospectively collected database, and their data were analyzed. RESULTS: A total of 46 patients with classic LCC, 30 with LCNEC, and 18 with LELC, who received surgical resection with curative intent, were identified and included in the analysis. Patients with LELC were younger, and the frequency of nonsmokers was greater than in patients with classic LCC or LCNEC. In patients with LCNEC or LELC, most lesions were located on the left side. There were 5 surgical deaths, and the median follow-up time of the surviving patients was 44.1 months. The 5-year disease free survival among the three subgroups was similar (p = 0.601), but patients with LELC had a significantly better overall survival than the other two subgroups (LELC vs classic LCC, p = 0.009; LELC vs LCNEC, p = 0.002). Multivariate analysis showed tumor location site, tumor stage, and LELC were independent prognostic factors of overall survival. CONCLUSIONS: The clinical manifestations and treatment outcomes of LCNEC, LELC, and classic LCC are different. LCNEC has a poor survival, and survival is not different than that of classic LCC. LELC is associated with younger age and a higher frequency of nonsmokers, and the treatment outcomes are better than those of other subtypes.

Proposed morphologic classification of prostate cancer with neuroendocrine differentiation.

On July 31, 2013, the Prostate Cancer Foundation assembled a working committee on the molecular biology and pathologic classification of neuroendocrine (NE) differentiation in prostate cancer. New clinical and molecular data emerging from prostate cancers treated by contemporary androgen deprivation therapies, as well as primary lesions, have highlighted the need for refinement of diagnostic terminology to encompass the full spectrum of NE differentiation. The classification system consists of: Usual prostate adenocarcinoma with NE differentiation; 2) Adenocarcinoma with Paneth cell NE differentiation; 3) Carcinoid tumor; 4) Small cell carcinoma; 5) Large cell NE carcinoma; and 5) Mixed NE carcinoma - acinar adenocarcinoma. The article also highlights "prostate carcinoma with overlapping features of small cell carcinoma and acinar adenocarcinoma" and "castrate-resistant prostate cancer with small cell cancer-like clinical presentation". It is envisioned that specific criteria associated with the refined diagnostic terminology will lead to clinically relevant pathologic diagnoses that will stimulate further clinical and molecular investigation and identification of appropriate targeted therapies.

Large cell carcinoma of the lung: clinically oriented classification integrating immunohistochemistry and molecular biology.

This study aimed at challenging pulmonary large cell carcinoma (LLC) as tumor entity and defining different subgroups according to immunohistochemical and molecular features. Expression of markers specific for glandular (TTF-1, napsin A, cytokeratin 7), squamous cell (p40, p63, cytokeratins 5/6, desmocollin-3), and neuroendocrine (chromogranin, synaptophysin, CD56) differentiation was studied in 121 LCC across their entire histological spectrum also using direct sequencing for epidermal growth factor receptor (EGFR) and v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations and FISH analysis for ALK gene translocation. Survival was not investigated. All 47 large cell neuroendocrine carcinomas demonstrated a true neuroendocrine cell lineage, whereas all 24 basaloid and both 2 lymphoepithelioma-like carcinomas showed squamous cell markers. Eighteen out of 22 clear cell carcinomas had glandular differentiation, with KRAS mutations being present in 39 % of cases, whereas squamous cell differentiation was present in four cases. Eighteen out of 20 large cell carcinomas, not otherwise specified, had glandular differentiation upon immunohistochemistry, with an exon 21 L858R EGFR mutation in one (5 %) tumor, an exon 2 KRAS mutation in eight (40 %) tumors, and an ALK translocation in one (5 %) tumor, whereas two tumors positive for CK7 and CK5/6 and negative for all other markers were considered adenocarcinoma. All six LCC of rhabdoid type expressed TTF-1 and/or CK7, three of which also harbored KRAS mutations. When positive and negative immunohistochemical staining for these markers was combined, three subsets of LCC emerged exhibiting glandular, squamous, and neuroendocrine differentiation. Molecular alterations were restricted to tumors classified as adenocarcinoma. Stratifying LCC into specific categories using immunohistochemistry and molecular analysis may significantly impact on the choice of therapy.

Immunohistochemical and molecular characterization of clear cell carcinoma of the lung.

The diagnostic entity of clear cell carcinoma of the lung (CCCL) is controversial, with many investigators arguing against its inclusion in the World Health Organization classification of lung carcinoma. Because of its rarity and questions regarding its histogenesis, I studied 3 groups of carcinomas with immunohistochemical and molecular assays, including 6 cases of CCCL, 7 cases of adenocarcinoma with clear cell change, and 11 cases of squamous carcinomas with prominent clear cell change. CCCL tended to be present in older individuals with an adenocarcinoma immunophenotype (cytokeratin 7 and TTF1 positivity). Molecular analysis by Sanger sequencing revealed KRAS mutations in 5 of 6 cases of CCCL, 2 of 7 adenocarcinomas with clear cell change, and 2 of 11 squamous carcinomas with clear cell change. Although perhaps not a distinct pathologic entity, in this pilot study, CCCL has an immunophenotype similar to solid-type adenocarcinoma with clear cell change and displays more frequent and unusual KRAS mutations than expected in most adenocarcinomas of the lung.

Immunohistochemistry in the differential diagnostics of primary lung cancer: an investigation within the Southern Swedish Lung Cancer Study.

OBJECTIVES: To assess immunohistochemical (IHC) stains differentially expressed between different types of lung cancer. METHODS: We evaluated 16 different IHC stains in 209 prospectively included, surgically treated primary lung cancers, including 121 adenocarcinomas, 65 squamous cell carcinomas, 15 large-cell carcinomas, 5 adenosquamous carcinomas, 2 sarcomatoid carcinomas, and 1 small-cell carcinoma, using the tissue microarray technique. RESULTS: Cytokeratin 5 (CK5) and P63 were both positive in 10% or more of the cells in 97% of the squamous cell carcinomas, with the former being positive (<10% of the cells) in only 2 non-squamous cell carcinomas. Thyroid transcription factor 1 (TTF1) and napsin A were positive in 10% or more of the cells in 88% and 87% of the adenocarcinomas, respectively, with 94% of the adenocarcinomas being positive in at least 1 marker. Fifteen percent of the adenocarcinomas were positive for estrogen receptor. CONCLUSIONS: CK5, TTF1, and napsin A are sensitive markers for squamous cell carcinoma and adenocarcinoma of the lung.

Neuroendocrine tumours--challenges in the diagnosis and classification of pulmonary neuroendocrine tumours.

Pulmonary neuroendocrine (NE) proliferations are a diverse group of disorders which share distinct cytological, architectural and biosynthetic features. Tumours composed of NE cells are dispersed among different tumour categories in the WHO classification of tumours and as such do not conform to a singular group with regards to treatment and prognosis. This is reflected by the highly variable behaviour of NE proliferations, ranging from asymptomatic, for instance in diffuse idiopathic pulmonary NE cell hyperplasia and tumourlets, to highly malignant cancers such as small cell lung cancer and large cell NE carcinoma. In this review NE proliferations are described as distinct entities ranging from low grade lesions to high grade cancers. The differential diagnoses are considered with each of the entries. Finally, mention is made of tumours which may show some NE features.

Distinct profile of driver mutations and clinical features in immunomarker-defined subsets of pulmonary large-cell carcinoma.

Pulmonary large-cell carcinoma-a diagnostically and clinically controversial entity-is defined as a non-small-cell carcinoma lacking morphologic differentiation of either adenocarcinoma or squamous cell carcinoma, but suspected to represent an end stage of poor differentiation of these tumor types. Given the recent advances in immunohistochemistry to distinguish adenocarcinoma and squamous cell carcinoma, and the recent insights that several therapeutically relevant genetic alterations are distributed differentially in these tumors, we hypothesized that immunophenotyping may stratify large-cell carcinomas into subsets with distinct profiles of targetable driver mutations. We therefore analyzed 102 large-cell carcinomas by immunohistochemistry for TTF-1 and ΔNp63/p40 as classifiers for adenocarcinoma and squamous cell carcinoma, respectively, and correlated the resulting subtypes with nine therapeutically relevant genetic alterations characteristic of adenocarcinoma (EGFR, KRAS, BRAF, MAP2K1/MEK1, NRAS, ERBB2/HER2 mutations and ALK rearrangements) or more common in squamous cell carcinoma (PIK3CA and AKT1 mutations). The immunomarkers classified large-cell carcinomas as variants of adenocarcinoma (n=62; 60%), squamous cell carcinoma (n=20; 20%) or marker-null (n=20; 20%). Genetic alterations were found in 38 cases (37%), including EGFR (n=1), KRAS (n=30), BRAF (n=2), MAP2K1 (n=1), ALK (n=3) and PIK3CA (n=1). All molecular alterations characteristic of adenocarcinoma occurred in tumors with immunoprofiles of adenocarcinoma or marker-null, but not in tumors with squamous immunoprofiles (combined mutation rate 50% vs 30% vs 0%, respectively; P<0.001), whereas the sole PIK3CA mutation occurred in a tumor with squamous profile (5%). Furthermore, marker-null large-cell carcinomas were associated with significantly inferior disease-free (P<0.001) and overall (P=0.001) survival. In conclusion, the majority (80%) of large-cell carcinomas can be classified by immunomarkers as variants of adenocarcinoma or squamous cell carcinoma, which stratifies these tumors into subsets with a distinct distribution of driver mutations and distinct prognoses. These findings have practical implications for diagnosis, predictive molecular testing and therapy selection.

Impact of collection and storage of lung tumor tissue on whole genome expression profiling.

Gene expression profiling could assist in revealing biomarkers of lung cancer prognosis and progression. The handling of biological samples may strongly influence global gene expression, a fact that has not been addressed in many studies. We sought to investigate the changes in gene expression that may occur as a result of sample processing time and conditions. Using Illumina Human WG-6 arrays, we quantified gene expression in lung carcinoma samples from six patients obtained at chest opening before and immediately after lung resection with storage in RNAlater [T1a((CO)) and T1b((LR))], after receipt of the sample for histopathology, placed in RNAlater [T2a((HP))]; snap frozen [T2b((HP.SF))]; or snap frozen and stored for 1 week [T2c((HP.SFA))], as well as formalin-fixed, paraffin-embedded (FFPE) block samples. Sampling immediately after resection closely represented the tissue obtained in situ, with only 1% of genes differing more than twofold [T1a((CO)) versus T1b((LR))]. Delaying tissue harvest for an average of 30 minutes from the operating theater had a significant impact on gene expression, with approximately 25% of genes differing between T1a((CO)) and T2a((HP)). Many genes previously identified as lung cancer biomarkers were altered during this period. Examination of FFPE specimens showed minimal correlation with fresh samples. This study shows that tissue collection immediately after lung resection with conservation in RNAlater is an optimal strategy for gene expression profiling.

Optimal combination of immunohistochemical markers for subclassification of non-small cell lung carcinomas: A tissue microarray study of poorly differentiated areas.

BACKGROUND: Recently, studies have been reported on the optimal immunohistochemical markers for subclassification of non-small cell lung carcinoma (NSCLC). The main pitfall in subclassification of NSCLC is small specimen with poorly differentiated area. In this study, we added newly proposed markers (e.g., napsin A, SOX2) to conventional markers (p63, TTF-1, CK5/6, and CK7) and evaluated optimal combination for subtyping of NSCLC, primarily focusing on the poorly differentiated area. METHODS: Eighty two resected NSCLCs with poorly differentiated areas were classified based on histologic findings. After histologic review, only poorly differentiated areas were selected and tissue microarrays were constructed to simulate small biopsy conditions. A total of 36 adenocarcinomas (ADCs), 38 squamous cell carcinomas (SQCCs), and 8 large cell carcinomas were included. All specimens were stained with TTF-1, napsin A, CK7, p63, CK5/6, and SOX2. RESULTS: With respect to ADC, TTF-1 was positive in 19 of 36 cases (53%) and napsin A was in 25 of 36 (69%). Both markers were specific for ADC (100% and 98%, respectively). With TTF-1 and napsin A in combination, sensitivity increased to 75%. CK7 was sensitive (92%) but not specific marker (76%) for ADC. With respect to SQCC, p63 was positive in 35 of 38 cases (92%) and CK5/6 was in 23 (61%). Both markers were specific for SQCC (both 93%). With p63 and CK5/6 in combination, sensitivity increased slightly to 95%, but specificity was lower at 91%. SOX2 was specific (100%) but not sensitive marker (53%) for SQCC. Combinations did not substantially increase the diagnostic performance. CONCLUSION: A simple panel of napsin A, TTF-1, and p63 can be sufficient to reliably subclassify poorly differentiated areas of NSCLC.