Purpose Three programmed death-1/programmed death-ligand 1 (PD-L1) inhibitors are currently approved for treatment of non-small-cell lung cancer (NSCLC). Treatment with pembrolizumab in NSCLC requires PD-L1 immunohistochemistry (IHC) testing. Nivolumab and atezolizumab are approved without PD-L1 testing, though US Food and Drug Administration-cleared complementary PD-L1 tests are available for both. PD-L1 IHC assays used to assess PD-L1 expression in patients treated with programmed death-1/PD-L1 inhibitors in clinical trials include PD-L1 IHC 28-8 pharmDx (28-8), PD-L1 IHC 22C3 pharmDx (22C3), Ventana PD-L1 SP142 (SP142), and Ventana PD-L1 SP263 (SP263). Differences in antibodies and IHC platforms have raised questions about comparability among these assays and their diagnostic use. This review provides practical information to help physicians and pathologists understand analytical features and comparability of various PD-L1 IHC assays and their diagnostic use. Methods We reviewed and summarized published or otherwise reported studies (January 2016 to January 2017) on clinical trial and laboratory-developed PD-L1 IHC assays (LDAs). Studies assessing the effect of diagnostic methods on PD-L1 expression levels were analyzed to address practical issues related to tissue samples used for testing. Results High concordance and interobserver reproducibility were observed with the 28-8, 22C3, and SP263 clinical trial assays for PD-L1 expression on tumor cell membranes, whereas lower PD-L1 expression was detected with SP142. Immune-cell PD-L1 expression was variable and interobserver concordance was poor. Inter- and intratumoral heterogeneity had variable effects on PD-L1 expression. Concordance among LDAs was variable. Conclusion High concordance among 28-8, 22C3, and SP263 when assessing PD-L1 expression on tumor cell membranes suggests possible interchangeability of their clinical use for NSCLC but not for assessment of PD-L1 expression on immune cells. Development of LDAs requires stringent standardization before their recommendation for routine clinical use.

Background: NTRK1, NTRK2 and NTRK3 fusions are present in a plethora of malignancies across different histologies. These fusions represent the most frequent mechanism of oncogenic activation of these receptor tyrosine kinases, and biomarkers for the use of TRK small molecule inhibitors. Given the varying frequency of NTRK1/2/3 fusions, crucial to the administration of NTRK inhibitors is the development of optimal approaches for the detection of human cancers harbouring activating NTRK1/2/3 fusion genes. Materials and methods: Experts from several Institutions were recruited by the European Society for Medical Oncology (ESMO) Translational Research and Precision Medicine Working Group (TR and PM WG) to review the available methods for the detection of NTRK gene fusions, their potential applications, and strategies for the implementation of a rational approach for the detection of NTRK1/2/3 fusion genes in human malignancies. A consensus on the most reasonable strategy to adopt when screening for NTRK fusions in oncologic patients was sought, and further reviewed and approved by the ESMO TR and PM WG and the ESMO leadership. Results: The main techniques employed for NTRK fusion gene detection include immunohistochemistry, fluorescence in situ hybridization (FISH), RT-PCR, and both RNA-based and DNA-based next generation sequencing (NGS). Each technique has advantages and limitations, and the choice of assays for screening and final diagnosis should also take into account the resources and clinical context. Conclusion: In tumours where NTRK fusions are highly recurrent, FISH, RT-PCR or RNA-based sequencing panels can be used as confirmatory techniques, whereas in the scenario of testing an unselected population where NTRK1/2/3 fusions are uncommon, either front-line sequencing (preferentially RNA-sequencing) or screening by immunohistochemistry followed by sequencing of positive cases should be pursued.

Introduction: A grading system for pulmonary adenocarcinoma has not been established. The International Association for the Study of Lung Cancer pathology panel evaluated a set of histologic criteria associated with prognosis aimed at establishing a grading system for invasive pulmonary adenocarcinoma. Conclusions: A grading system based on the predominant and high-grade patterns is practical and prognostic for invasive pulmonary adenocarcinoma.

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In non-small cell lung cancer, epidermal growth factor receptor gene mutations and anaplastic lymphoma kinase (ALK) gene rearrangements have a major impact upon the level of response to treatment with specific tyrosine kinase inhibitors. This review describes the molecular basis of ALK inhibition, summarizes current data on the effectiveness and safety of ALK inhibition therapy, describes the different testing methodologies with their advantages and disadvantages, provides a suggested testing algorithm and puts forward a proposal for an external quality assessment program in ALK testing. Electronic supplementary material The online version of this article (doi:10.1007/s00428-012-1281-4) contains supplementary material, which is available to authorized users.

Since the 2015 WHO classification was introduced into clinical practice, immunohistochemistry (IHC) has figured prominently in lung cancer diagnosis. In addition to distinction of small cell versus non-small cell carcinoma, patients' treatment of choice is directly linked to histologic subtypes of non-small cell carcinoma, which pertains to IHC results, particularly for poorly differentiated tumors. The use of IHC has improved diagnostic accuracy in the classification of lung carcinoma, but the interpretation of IHC results remains challenging in some instances. Also, pathologists must be aware of many interpretation pitfalls, and the use of IHC should be efficient to spare the tissue for molecular testing. The International Association for the Study of Lung Cancer Pathology Committee received questions on practical application and interpretation of IHC in lung cancer diagnosis. After discussions in several International Association for the Study of Lung Cancer Pathology Committee meetings, the issues and caveats were summarized in terms of 11 key questions covering common and important diagnostic situations in a daily clinical practice with some relevant challenging queries. The questions cover topics such as the best IHC markers for distinguishing NSCLC subtypes, differences in thyroid transcription factor 1 clones, and the utility of IHC in diagnosing uncommon subtypes of lung cancer and distinguishing primary from metastatic tumors. This article provides answers and explanations for the key questions about the use of IHC in diagnosis of lung carcinoma, representing viewpoints of experts in thoracic pathology that should assist the community in the appropriate use of IHC in diagnostic pathology.

Most gene expression profiling studies of mesothelioma have been based on relatively small sample numbers, limiting their statistical power. We did Affymetrix U133A microarray analysis on 99 pleural mesotheliomas, in which multivariate analysis showed advanced-stage, sarcomatous histology and P16/CDKN2A homozygous deletion to be significant independent adverse prognostic factors. Comparison of the expression profiles of epithelioid versus sarcomatous mesotheliomas identified many genes significantly overexpressed among the former, including previously unrecognized ones, such as uroplakins and kallikrein 11, both confirmed by immunohistochemistry. Examination of the gene expression correlates of survival showed that more aggressive mesotheliomas expressed higher levels of Aurora kinases A and B and functionally related genes involved in mitosis and cell cycle control. Independent confirmation of the negative effect of Aurora kinase B was obtained by immunohistochemistry in a separate patient cohort. A role for Aurora kinases in the aggressive behavior of mesotheliomas is of potential clinical interest because of the recent development of small-molecule inhibitors. We then used our data to develop microarray-based predictors of 1 year survival; these achieved a maximal accuracy of 68% in cross-validation. However, this was inferior to prognostic prediction based on standard clinicopathologic variables and P16/CDNK2A status (accuracy, 73%), and adding the microarray model to the latter did not improve overall accuracy. Finally, we evaluated three recently published microarray-based outcome prediction models, but their accuracies ranged from 63% to 67%, consistently lower than reported. Gene expression profiling of mesotheliomas is an important discovery tool, but its power in clinical prognostication has been overestimated.

There is an enormous gap between the antiproliferative and in vivo antitumor efficacy of gemcitabine in cell linebased models and its clinical efficacy. This may be due to insensitiveness of the precursor, cancer stem cell (CSC) compartment to cytotoxic agents. The hedgehog pathway is associated with CSC signaling and control. We used a direct xenograft model of pancreatic cancer and a twostage approach was used to test the hypotheses that targeting CSC could increase the efficacy of gemcitabine. Tumors from a gemcitabine-sensitive xenograft were treated with gemcitabine first, and randomized, after tumor regression to continuing treatment with gemcitabine, a hedgehog inhibitor alone or in combination with gemcitabine. We tested markers described as associated with CSC such as CD24, CD44, ALDH, nestin, and the hedgehog pathway. After induction with gemcitabine, treated tumor showed an enrichment in CSC markers such as ALDH and CD24. Subsequently, a release from gemcitabine prompted a repopulation of proliferating cells and a decrease in such markers to equilibrate from pretreatment levels. Combined treatment with gemcitabine and cyclopamine induced tumor regression and decrease in CSC markers and hedgehog signaling. Cytoplasmic CD24 and ALDH were inversely and strongly associated with growth and were expressed in a minority of cells that we propose constitute the CSC compartment. Hedgehog inhibitors as part of a dual compartment therapeutic approach were able to further reduce tumor growth and decreased both static and dynamic markers of CSC. Direct tumor xenografts are a valid platform to test multicompartment therapeutic approaches in pancreatic cancer. [Mol Cancer Ther 2009;8(2):310 -4]