Assessment of a New ROS1 Immunohistochemistry Clone (SP384) for the Identification of ROS1 Rearrangements in Patients with Non–Small Cell Lung Carcinoma: the ROSING Study

Introduction: The ROS1 gene rearrangement has become an important biomarker in NSCLC. The College of American Pathologists/International Association for the Study of Lung Cancer/Association for Molecular Pathology testing guidelines support the use of ROS1 immunohistochemistry (IHC) as a screening test, followed by confirmation with fluorescence in situ hybridization (FISH) or a molecular test in all positive results. We have evaluated a novel anti-ROS1 IHC antibody (SP384) in a large multicenter series to obtain real-world data. Methods: A total of 43 ROS1 FISH-positive and 193 ROS1 FISH-negative NSCLC samples were studied. All specimens were screened by using two antibodies (clone D4D6 from Cell Signaling Technology and clone SP384 from Ventana Medical Systems), and the different interpretation criteria were compared with break-apart FISH (Vysis). FISH-positive samples were also analyzed with next-generation sequencing (Oncomine Dx Target Test Panel, Thermo Fisher Scientific). Results: An H-score of 150 or higher or the presence of at least 70% of tumor cells with an intensity of staining of 2+ or higher by the SP384 clone was the optimal cutoff value (both with 93% sensitivity and 100% specificity). The D4D6 clone showed similar results, with an H-score of at least 100 (91% sensitivity and 100% specificity). ROS1 expression in normal lung was more frequent with use of the SP384 clone (p < 0.0001). The ezrin gene (EZR)-ROS1 variant was associated with membranous staining and an isolated green signal FISH pattern (p = 0.001 and p = 0.017, respectively). Conclusions: The new SP384 ROS1 IHC clone showed excellent sensitivity without compromising specificity, so it is another excellent analytical option for the proposed testing algorithm

Agreement on endoscopic ultrasonography-guided tissue specimens: Comparing a 20-G fine-needle biopsy to a 25-G fine-needle aspiration needle among academic and non-academic pathologists

Background and Aim: A recently carried out randomized controlled trial showed the benefit of a novel 20-G fine-needle biopsy (FNB) over a 25-G fine-needle aspiration (FNA) needle. The current study evaluated the reproducibility of these findings among expert academic and non-academic pathologists. Methods: This study was a side-study of the ASPRO (ASpiration versus PROcore) study. Five centers retrieved 74 (59%) consecutive FNB and 51 (41%) FNA samples from the ASPRO study according to randomization; 64 (51%) pancreatic and 61 (49%) lymph node specimens. Samples were re-reviewed by five expert academic and five non-academic pathologists and rated in terms of sample quality and diagnosis. Ratings were compared between needles, expert academic and

Comparison of pancreatic histology specimens obtained by EUS 19G versus 22G core biopsy needles: A prospective multicentre study among experienced pathologists

Background and aim: Scanty data about inter-observer agreement (IOA) among pathologists in the evaluation of pancreatic samples acquired with EUS histology needle are available. The aim of this study was to determine IOA on adequacy of pancreatic histology specimens obtained with a 22G needle by a panel of experienced pathologist, in comparison with the 19G needle. Methods: This multicentre prospective study involved 73 pancreatic specimens prepared using histology needles of different calibres. Five pathologists independently reviewed all the samples, assessing the presence of a core, specimen adequacy and the possibility to perform additional analyses. IOA determined by Fleiss’ Kappa statistic was used as the primary outcome measure. Secondary outcome was to compare 22G versus 19G needle results. Results: A core was present in 57% of pancreatic specimens obtained by 22G needle. The specimens were considered adequate in 72% of cases, with poor agreement among pathologists (p = 0.02, Fleiss’ κ = 0.26). The possibility to perform further analyses was rated as ‘positive’ in 66% of cases without significant difference among observers (p = 0.80). When comparing the results, the presence of a core and the adequacy of tissue slides were significantly better for the 19G needle (57% vs. 84% p = 0.002; 72% vs. 83% p = 0.004, respectively). Reproducibility in the assessment of pancreatic sample adequacy was significantly better with the 19G needle (κ = 0.26 for 22G samples vs. κ = 0.81 for 19G samples). Conclusions: Our results suggest that histology sampling of pancreatic masses should be performed with a 19G histology needle, since is able to provide a core in the majority of cases, with 83% of adequate specimens and excellent results in term of reproducibility among pathologists

Comparison of pancreatic histology specimens obtained by EUS 19G versus 22G core biopsy needles: A prospective multicentre study among experienced pathologists

Background and aim: Scanty data about inter-observer agreement (IOA) among pathologists in the evaluation of pancreatic samples acquired with EUS histology needle are available. The aim of this study was to determine IOA on adequacy of pancreatic histology specimens obtained with a 22G needle by a panel of experienced pathologist, in comparison with the 19G needle. Methods: This multicentre prospective study involved 73 pancreatic specimens prepared using histology needles of different calibres. Five pathologists independently reviewed all the samples, assessing the presence of a core, specimen adequacy and the possibility to perform additional analyses. IOA determined by Fleiss’ Kappa statistic was used as the primary outcome measure. Secondary outcome was to compare 22G versus 19G needle results. Results: A core was present in 57% of pancreatic specimens obtained by 22G needle. The specimens were considered adequate in 72% of cases, with poor agreement among pathologists (p = 0.02, Fleiss’ κ = 0.26). The possibility to perform further analyses was rated as ‘positive’ in 66% of cases without significant difference among observers (p = 0.80). When comparing the results, the presence of a core and the adequacy of tissue slides were significantly better for the 19G needle (57% vs. 84% p = 0.002; 72% vs. 83% p = 0.004, respectively). Reproducibility in the assessment of pancreatic sample adequacy was significantly better with the 19G needle (κ = 0.26 for 22G samples vs. κ = 0.81 for 19G samples). Conclusions: Our results suggest that histology sampling of pancreatic masses should be performed with a 19G histology needle, since is able to provide a core in the majority of cases, with 83% of adequate specimens and excellent results in term of reproducibility among pathologists

Assessment of a New ROS1 Immunohistochemistry Clone (SP384) for the Identification of ROS1 Rearrangements in Patients with Non–Small Cell Lung Carcinoma: the ROSING Study

Introduction: The ROS1 gene rearrangement has become an important biomarker in NSCLC. The College of American Pathologists/International Association for the Study of Lung Cancer/Association for Molecular Pathology testing guidelines support the use of ROS1 immunohistochemistry (IHC) as a screening test, followed by confirmation with fluorescence in situ hybridization (FISH) or a molecular test in all positive results. We have evaluated a novel anti-ROS1 IHC antibody (SP384) in a large multicenter series to obtain real-world data. Methods: A total of 43 ROS1 FISH-positive and 193 ROS1 FISH-negative NSCLC samples were studied. All specimens were screened by using two antibodies (clone D4D6 from Cell Signaling Technology and clone SP384 from Ventana Medical Systems), and the different interpretation criteria were compared with break-apart FISH (Vysis). FISH-positive samples were also analyzed with next-generation sequencing (Oncomine Dx Target Test Panel, Thermo Fisher Scientific). Results: An H-score of 150 or higher or the presence of at least 70% of tumor cells with an intensity of staining of 2+ or higher by the SP384 clone was the optimal cutoff value (both with 93% sensitivity and 100% specificity). The D4D6 clone showed similar results, with an H-score of at least 100 (91% sensitivity and 100% specificity). ROS1 expression in normal lung was more frequent with use of the SP384 clone (p < 0.0001). The ezrin gene (EZR)-ROS1 variant was associated with membranous staining and an isolated green signal FISH pattern (p = 0.001 and p = 0.017, respectively). Conclusions: The new SP384 ROS1 IHC clone showed excellent sensitivity without compromising specificity, so it is another excellent analytical option for the proposed testing algorithm

Accurate Identification of ALK Positive Lung Carcinoma Patients: Novel FDA-Cleared Automated Fluorescence In Situ Hybridization Scanning System and Ultrasensitive Immunohistochemistry

BackgroundBased on the excellent results of the clinical trials with ALK-inhibitors, the importance of accurately identifying ALK positive lung cancer has never been greater. However, there are increasing number of recent publications addressing discordances between FISH and IHC. The controversy is further fuelled by the different regulatory approvals. This situation prompted us to investigate two ALK IHC antibodies (using a novel ultrasensitive detection-amplification kit) and an automated ALK FISH scanning system (FDA-cleared) in a series of non-small cell lung cancer tumor samples.MethodsForty-seven ALK FISH-positive and 56 ALK FISH-negative NSCLC samples were studied. All specimens were screened for ALK expression by two IHC antibodies (clone 5A4 from Novocastra and clone D5F3 from Ventana) and for ALK rearrangement by FISH (Vysis ALK FISH break-apart kit), which was automatically captured and scored by using Bioview's automated scanning system.ResultsAll positive cases with the IHC antibodies were FISH-positive. There was only one IHC-negative case with both antibodies which showed a FISH-positive result. The overall sensitivity and specificity of the IHC in comparison with FISH were 98% and 100%, respectively.ConclusionsThe specificity of these ultrasensitive IHC assays may obviate the need for FISH confirmation in positive IHC cases. However, the likelihood of false negative IHC results strengthens the case for FISH testing, at least in some situations