With a growing number of predictive biomarkers needed to manage patients with non-small cell lung cancer (NSCLC), there has been a paradigm shift in care and handling of diagnostic samples. Among the various testing methods, immunohistochemistry (IHC) is the most cost- effective and widely available. Furthermore, over the past decade immunotherapy has emerged as one of the most promising cancer treatments. In this scenario IHC is the most used testing method available for PDL-1/PD1 immunotherapy. Several monoclonal antibodies targeting programmed death 1 (PD-1)/programmed death ligand-1 (PD-L1) pathways have been integrated into standard-of-care treatments of a wide range of cancer types, once provided evidence of PD-L1 expression in tumor cells by immunohistochemistry (IHC). Since currently available PD-L1 assays have been developed on formalin-fixed paraffin embedded (FFPE) histological specimens, a growing body of research is being dedicated to confirm the feasibility of applying PDL-1 assays also to cytological samples. Albeit promising results have been reported, several important issues still need to be addressed. Among these are the type of cytological samples, pre-analytical issues, cyto-histological correlation, and inter-observer agreement. This review briefly summarizes the knowledge of the role of cytopathology in the analysis of PD-L1 by immunocytochemistry (ICC) and future directions of cytopathology in the immunotherapy setting.

Desmoplastic small round cell tumor (DSRCT) is rare and a highly aggressive neoplasm that typically involves the soft tissues of the abdomen or pelvis in children or young adults, showing a male predilection. Although it can occurs over a wide age range, the peak incidence is in the third decade of life. DSRCT usually shows widespread abdominal serosal involvement, and overall patient survival is poor. On the other hand, extra‐abdominal DSRCT is very rare. DSRCT in major salivary glands has been reported, but it is extremely rare. In the majority of reported series diagnosis is made by the histological analysis of FFPE tissues together with immunohistochemistry (IHC) and molecular analysis, particularly the demonstration of chromosomal translocation involving EWSR1. Very few cases have been diagnosed so far by Fine Needle Aspiration (FNA) cytology. Moreover ancillary studies have been performed in all reported cases in FFPE samples. There is still controversy and lack of consensus regarding the suitability of cytological samples especially smears for immunocytochemical (ICC) and fluorescence in situ hybridization (FISH), what makes its standardization difficult. We report a case of a primary DSRCT of parotid gland in a 17‐year‐old male diagnosed by FNA cytology. The cytomorphological diagnosis was coupled with ICC and FISH analysis performed on stained smears. We emphasize the feasibility and reliability of cytological smears for the application of immunocytochemical and molecular techniques.

Objective Understanding the immune environment of non‐small cell lung cancer (NSCLC) is important for designing effective anticancer immunotherapies. We describe the use of multiplex immunofluorescence (mIF) assays to enable characterisation of the tumour‐infiltrating immune cells and their interactions, both across and within immune subtypes. Methods Six cytological samples of NSCLC taken by transoesophageal ultrasound‐guided fine needle aspiration were tested with an mIF assay designed to detect the expression of key immune cell markers such as CD3, CD8, CD20, CD11b, and CD68. Pan‐cytokeratin was used to detect the NSCLC cells. Fluorescence images were acquired on a Vectra‐Polaris Automated Quantitative Pathology Imaging System (Akoya Biosciences). Results MIF assay was able to reliably detect and quantify the myeloid cell markers CD11b, CD68, CD3+ and CD8+ T cells, and CD20+ B lymphocytes on cytological samples of NSCLC. Whole‐tissue analysis and its correlation with the corresponding H&E stains allowed a better understanding of the tissue morphology and the relationship between tumour and stroma compartments. Additionally, a uniform, specific, and correct staining pattern was seen for every immune marker. Conclusion The implementation of mIF assay on cytological samples taken with minimally invasive methods seems feasible and can be used to explore the immune environment of NSCLC.

resection was defined by free resection margins, but with less rigorous lymph node evaluation than systematic dissection and/or positivity of the highest mediastinal lymph node removed. Incomplete resection was defined by the presence of gorss or microscopic residual disease. Patient follow-up was updated until Jan/2019. Overall survival was analyzed by the Kaplan-Meier method, Log rank test and Cox proportional regression. Result: A total of 663 patients were identified. Mean age was 65.64 years, 338 men(50.9%). The predominant histological type was adenocarcinoma(n ¼ 466, 70.2%), followed by squamous cell carcinoma(n ¼ 162, 24.4%). Lobectomy was the most commonly performed procedure(n ¼ 576, 86.8%), followed by segmentectomy and pneumonectomy(n ¼ 40, 6.0% and n ¼ 34, 5.1%, respectively). There was 388 patientes(59.81%) classified as stage I, 146(23.1%) stage II, 97(15.3%) stage III and 11(1.74%) stage IV. Resection was complete in 374 cases (56.4%), uncertain in 252 cases(38.0%) and incomplete in 37 cases(5.5%). Mediastinal lymphadenectomy was adequate in 421 cases (63.4%) and inadequate in 242 (36.5%). Reasons for inadequate lymphadenectomy were: no nodal station sampling (n ¼ 30, 4.5%), no station 7 sampling (n ¼ 103, 15.5%) and sampling of less than 3 mediastinal stations (n ¼ 109, 16.4%). The highest mediastinal lymph node removed was positive in 45 cases (6.7%). Surgical margins were positive in 37 cases (5.5%). The median follow-up was 19.5 months (IQR 7.4 -42.5), and 5 years follow-up was completed in in 15.5%. During follow-up, 133 (20.4%) patients had recurrence of the disease. Median disease-free survival was 64 months in the general group and 84.0, 58.6 and 31.5 months in the complete, uncertain and incomplete resection groups, respectively (log rank p ¼ 0.15). Median overall survival in the complete resection group was 98.3 months, in the uncertain resection group it was 64 months. The incomplete resection group did not reach the median. There was no statistical difference in survival between groups (log rank p ¼ 0.22). Conclusion: The analysis showed a high prevalence of uncertain resection, but comparable to other studies already published. This demonstrates that lymphadenectomy is not being performed according to IASLC recommendations. However, in this study, there was no impact on overall survival and disease-free survival at 5 years, which may be due to the small sample size and the short follow-up time of the vast majority of patients included in the PLCR.

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