Claire W. Michael

Background: This invited review for the special issue is a celebration of the continued growth of knowledge in the field of serous fluid cytopathology. Summary: The review begins by addressing the value of cytological examination of the fluid samples in conjunction with clinical, radiological and biochemical data interpreted by the treating physician. It then covers the advances in cytopreparatory techniques with special reference to existing and emerging ancillary techniques available to obtain maximal diagnostic, prognostic & therapeutic information for the practice of precision medicine. The concept of mesothelioma as a cytological diagnosis has been made possible in recent years through clinical research and advocacy by experts and this has added value to serous fluid samples especially when this is the only sampling method available for a patient. Advances in the recommended repertoire of immunostains and a standardised approach through the use of diagnostic categories has been made possible by the International system for reporting serous fluid cytopathology. The reporting system has found application to other types of fluid samples, most recently, cerebrospinal fluid cytology which will be included in the forthcoming edition of the terminology system and is introduced in this special issue. Key messages. The review charts a journey commemorating important landmarks in our current understanding of serous fluids in diagnostic cytopathology.

Background: Mesothelial proliferations range from reactive lesions to benign and malignant tumors, bearing witness to the plasticity of these cells. Their diagnosis often requires combination of morphological assessment, immunohistochemistry, and molecular testing. Summary: Considerable progress has been made in recent years in all the above aspects. The separation of reactive from neoplastic mesothelial proliferations has become more robust. Mesothelioma in situ is now recognized as precursor of invasive mesothelioma and better tools exist for its differentiation from benign mimics. Differentiating mesothelial tumors from other malignancies, including metastatic carcinomas, sarcomas, and other tumors, is now more easily achieved. Genetic conditions which predispose to development of mesothelioma at a young age have been identified and characterized. These issues are discussed in this review. Key messages: Mesothelial pathology is an evolving field and more knowledge of these processes is likely to be gained in coming years. The diagnosis of these tumors requires experts with subspecialty in cytology, thoracic, soft tissue/bone, and gynecologic pathology, cooperating with surgeons, radiologists, and oncologists at specialized centers.

Introduction: Lung adenocarcinoma commonly causes malignant pleural effusion (MPE), a condition with poor prognosis and limited treatment options. Pleural effusion specimens offer a minimally invasive source for diagnosis and molecular testing. This study evaluates cytologic adequacy for molecular analysis, characterizes mutational and PD‐L1 profiles, and correlates findings with survival outcomes. Methods: Over 10 years, a review of 2400 MPE identified 94 lung adenocarcinoma cases that underwent next generation sequencing (NGS). Patient demographics, cytological characteristics, molecular features, and survival outcome were integrated for analysis. Results: The mean effusion volume submitted for cytological and molecular testing was 589.8 mL (range 4–2790 mL) and mean tumor cellularity was 40% (range 10%–90%). NGS identified mutations in 86/94 cases (91%). TP53 was the most common alteration (42/94, 44.6%). FDA‐approved biomarkers were detected: KRAS (28/94, 29.7%); EGFR (15/94, 15.9%); BRAF (8/94, 8.5%); ALK (3/94, 4.2%), ROS1 (2/94, 2.1%), RET fusions (1/94, 1%); and MET amplification (2/94, 2.1%). Smoking was associated with bloody effusions ( p = 0.049), frequent TP53 mutations ( p = 0.018), and fewer EGFR mutations ( p = 0.012). Patients with effusions as primary presentation was associated with higher mortality ( p = 0.002), frequent TTF‐1 positivity ( p = < 0.001) and frequent BRAF mutations ( p = 0.049). TTF‐1 positivity was associated with higher PD‐L1 scores ( p = 0.035). Survival was shorter in male patients ( p = 0.045), smokers ( p = 0.025), concurrent pericardial effusions ( p = 0.027), and with TP53 mutation ( p = 0.002). Conclusions: Molecular testing of effusion samples is feasible even at low fluid volumes, provided tumor cellularity is adequate. Prioritizing tumor fraction over volume ensures a higher likelihood of successful molecular analysis. Effusion‐based NGS reliably identifies actionable alterations with diagnostic and prognostic relevance. Male patients, smokers, concurrent or primary presentation with MPE, and those with TP53 mutations had poorer outcomes.

There are numerous benign cellular changes that occur in the lung due to diverse underlying diseases that can be encountered in exfoliative samples and transbronchial and CT‐guided samples. Entities such as bronchial cell hyperplasia, pneumocyte type II proliferation, repair, etc. should be recognised and diagnosed. Yet, many pathologists encounter difficulties in their interpretation and frequently either diagnose them as ‘Atypical’ resulting in more invasive procedures or misdiagnose them as ‘Malignant’ with drastic consequences to the patient. Other entities may be dismissed as benign lung elements, resulting in misclassification as non‐diagnostic/non‐representative, resulting in repeated or more invasive procedures. In this review, we will present an overview of many of these diagnostic pitfalls and mimickers and describe clues to their diagnosis and how to differentiate them from malignant conditions. As in all cytologic diagnoses, both false positive and false negative errors can occur and both types of errors can have grave consequences to patient management, resulting in increased unnecessary morbidity such as pneumonectomy and, at times, litigation.

Introduction: The diagnosis of mesothelioma in situ (MIS) is now accepted by the WHO as a pre‐invasive neoplastic mesothelial proliferation and considered a diagnosis based on histologic evaluation only. Although the definition of MIS includes recurrent effusions, little is known about the cytologic features of such effusions. Since mesothelioma is usually diagnosed at an advanced stage and has a poor prognosis, early detection of a neoplastic mesothelial population in such effusions can potentially have a positive impact on the management of such a dire disease.Materials and Methods: We reviewed a total of 18 pleural effusions from nine patients with recurrent effusions. Of these, five patients had follow‐up biopsies diagnosed as MIS and the remaining four cases had negative radiology and malignant cytology proven by molecular markers (BAP1, MTAP or CDKN2A deletion) and at least 1 year follow‐up with no overt mass identified by radiology.Results: Initial effusions may mimic reactive mesothelial hyperplasia or exhibit atypia. As effusions recur, the cellularity and atypia increase and the mesothelial proliferation becomes morphologically indistinguishable from mesothelioma. Molecular alterations diagnostic of mesothelioma can be detected in these effusions, even in the initial‐benign/reactive appearing ones. The cellularity and atypia detected in such effusions surpassed those noted on the biopsies, raising questions regarding the cause of such discrepancy.Conclusion: The diagnosis of MIS can be suspected based on malignant effusion cytology supported by molecular alterations. We propose that the proliferation of neoplastic mesothelial clones represent a clinically silent “liquid phase MIS stage” corresponding to in situ stage in other organs.