Malignant Effusion Fluid Research Articles

Diagnostic Utility of Cell block and Immunocytochemistry in Differentiating Malignant from Non-malignant Effusions: A Cross-sectional Study

Introduction: Cytological evaluation of body cavity fluids has been widely employed in diagnosing the underlying aetiology. Differentiating reactive mesothelial cell proliferations from metastatic malignant cells based on cytomorphology alone is challenging. Therefore, the use of cell blocks in conjunction with immunocytochemistry can significantly improve diagnostic accuracy. Aim: To investigate the role of cell blocks and immunocytochemistry in distinguishing malignant from non- malignant effusion fluids. Materials and Methods: A prospective cross-sectional study was conducted on 70 serous effusion fluid samples (ascitic and pleural fluids) received in the Cytology section of the Pathology Department from October 2021 to November 2022. Relevant clinical details, including demographic information, age, gender, presenting complaints, and radiological and laboratory investigations, were documented from the patients’ medical records. All fluids underwent conventional cytology, and the remaining fluid was used for cell block preparations. Immunocytochemistry was performed using Ber-EP4 and Calretinin immuno-markers to differentiate between reactive mesothelial cells and malignant cells. The association between cytosmear/cell block and immunocytochemistry was calculated using the chi-square test. Results: Out of the 70 cases, 44 were pleural fluids, and 26 were ascitic fluids. Conventional cytology identified 11 positive cases, 20 suspicious cases, and 39 negative cases for malignancy. However, after cell block examination, the total number of positive cases reduced to 9, suspicious cases decreased to 12, and negative cases increased to 49, resulting in a 14.3% increase in diagnostic accuracy. Immunocytochemistry using Ber-EP4 showed strong positivity in 12 cases, indicating epithelial malignancy (adenocarcinoma), while 9 cases were Ber-EP4 negative. Calretinin positivity was observed in the mesothelial cells of all 21 cases where immunohistochemistry was performed, resulting in a 17.5% increase in diagnostic accuracy. Conclusion: The combination of cell block technique with conventional cytology improves the diagnostic yield and accuracy by providing better interpretation of architectural patterns and cytomorphology. Additionally, the application of immunocytochemistry using Ber-EP4 and Calretinin aids in distinguishing malignant from non-malignant serous effusions.

Short-term organoid culture for drug sensitivity testing of high-grade serous carcinoma

ObjectiveCancer patient-derived organoids (PDOs) grow as three dimensional (3D) structures in the presence of extracellular matrix and have been found to represent the original tumor's genetic complexity. In addition, PDOs can be grown and subjected to drug sensitivity testing in a shorter time course and with lesser expense than patient-derived xenograft models. Many patients with recurrent ovarian cancer develop malignant effusions that become refractory to chemotherapy. Since these same patients often present for palliative aspiration of ascites or pleural effusions, there is a potential opportunity to obtain tumor specimens in the form of multicellular spheroids (MCS) present in malignant effusion fluids. Our objective was to develop a short duration culture of MCS from ovarian cancer malignant effusions in conditions selected to support organoid growth and use them as a platform for empirical drug sensitivity testing. MethodsIn this study, malignant effusion specimens were collected from patients with high-grade serous ovarian carcinoma (HGSOC). MCS were recovered and subjected to culture conditions designed to support organoid growth. In a subset of specimens, RNA-sequencing was performed at two time points during the short-term culture to determine changes in transcriptome in response to culture conditions. Organoid induction was also characterized in these specimens using Ki67 staining and histologic analysis. Drug sensitivity testing was performed on all specimens. ResultsOur model describes organoids formed within days of primary culture, which can recapitulate the histological features of malignant ascites fluid and can be expanded for at least 6 days. RNA-seq analysis of four patient specimens showed that within 6 days of culture, there was significant up-regulation of genes related to cellular proliferation, epithelial-mesenchymal transition, and KRAS signaling pathways. Drug sensitivity testing identified several agents with therapeutic potential. ConclusionsShort duration organoid culture of MCS from HGSOC malignant effusions can be used as a platform for empiric drug sensitivity testing. These ex vivo models may be helpful in screening new or existing therapeutic agents prior to individualized treatment options.

Utility of PD-L1 immunocytochemistry using body-fluid cell blocks in patients with non-small-cell lung cancer.

The expression of programmed cell death ligand-1 (PD-L1) is a biomarker in patients with non-small-cell lung carcinoma (NSCLC). Patients with advanced-stage NSCLC receive a variety of molecular genetic tests, possibly resulting in insufficient tissue for immunoassay of PD-L1. Thus, to determine whether effusion fluid specimens are a reliable alternative to tissue specimens for PD-L1 testing, we compared the results of PD-L1 immunostaining using body-fluid cell blocks and tumor tissues. PD-L1 immunostaining was performed in 62 paired samples of cytology cell blocks (ie, immunocytochemistry) and tumor tissues (ie, immunohistochemistry) from 36 patients using the E1L3N, SP142, and SP263 anti PD-L1 antibody clones. Of the 62 cytology specimens, 50 were from malignant effusion fluid. PD-L1 expression was scored as the percentage of tumor cells with clear membranous staining. A strong positive correlation was observed between the immunostains on cytology cell blocks and tumor tissue (Pearson's correlation coefficient, R = .804, P < .001). When the score was categorized as <1%, ≥1% and <10%, ≥10% and <50%, and ≥50%, the overall concordance rate was 74.2% (46/62, Cohen's k = 0.568). After dichotomizing the cases using cutoff values of 1%, 10%, and 50%, the concordance rates were 84% to 100% for both adenocarcinoma and squamous cell carcinoma. The concordance rate was higher in patients with NSCLC with an EGFR mutation and using the SP263 rather than the E1L3N clone. The results of PD-L1 immunostaining of cell blocks, particularly from effusion fluid, reflect the PD-L1 expression status of NSCLC tissue.

Validating Micronucleus Score in Effusion Fluids.

Background:Identifying malignant cells in effusion fluid is vital in staging and management of cancers. Differentiating reactive mesothelial cells from malignant cells in effusion fluid is a challenging task and there is an ongoing need for simpler and cost effective tool to aid the diagnosis. Micronucleus is an additional smaller nucleus in the cytoplasm, formed by chromosomes or chromosomal fragments formed during cell division.Aims:The aim of this study was to assess the significance of micronucleated cell in effusion fluids to distinguish adenocarcinomatous from reactive mesothelial effusions.Materials and Methods:Thirty cases of unequivocal malignant effusion fluids and 30 benign cases with reactive mesothelial cells as control were studied. Number of microucleated cells present per1000 well-preserved cells in Leishman-stained smears were counted.Results:Mean (±SD) micronucleated score in malignant and benign effusions were 15.77 ± 9.78 and 1.87 ± 1.78, respectively. The median scores were 13 and 2, respectively. Mann–Whitney test showed that this difference was statistically significant (P < 0.001). This study revealed that there was a significant difference in micronucleus scoring between benign and malignant effusions.Conclusions:Micronucleus score can be used as an additional biomarker in the interpretation of routinely stained cytosmears.

The natural history and molecular heterogeneity of HIV-associated primary malignant lymphomatous effusions.

Primary malignant lymphomatous effusions arising in individuals infected with the human immunodeficiency virus, type 1 (HIV-1) represent a rare subset of HIV-associated lymphomas. Previous studies have demonstrated that the malignant cells are monoclonal (as defined by rearrangement of the immunoglobulin gene), express cell surface CD38, and are infected with Epstein-Barr virus (EBV) and human herpes virus, type 8 (HHV-8). Despite these detailed molecular and immunophenotypic studies, clinical information on this disease entity is scant, prompting us to review the clinical features of eight cases seen at our institutions. All eight patients had total peripheral CD4+ lymphocytes < 200/microliter and presented with complaints related to body cavity distension. Routine laboratory values were nondiagnostic and yielded no prognostic information. Only two patients could tolerate and thus received chemotherapy with no obvious impact on their clinical course. The mean overall survival after diagnosis was 60 days (range 6-166 days). Four patients were examined at autopsy. The primary malignant lymphomatous effusion either was the immediate cause of death or contributed significantly to the death of only two. All four patients examined post mortem, however, had lymphomatous infiltration of serosal surfaces adjacent to the site of the primary malignant effusion. Molecular and immunologic studies performed on the malignant cells and effusion fluids revealed universal expression of cell surface CD38 and the presence of HHV-8 gene sequences, but in contrast with previous studies, only four had rearranged immunoglobulin genes or EBV present: IL-6 and IL-10 levels in the malignant effusion fluids were markedly elevated. In summary, this rare subset of HIV-associated lymphomas in our eight patients arose late in the course of HIV-associated disease, had a rapid clinical course, and was molecularly heterogeneous. A pathogenetic role for HHV-8 alone in this disease process is strengthened by our observation of four cases lacking EBV but containing HHV-8.

Carcinoembryonic Antigen Titers on Effusion Fluid

Carcinoembryonic antigen (CEA) has been demonstrated in tumors and body fluids, suggesting that effusion fluid (EF) CEA titer might separate benign and malignant EF. Plasma and EF CEA titers were determined on 141 patients with either benign (86) or malignant (55) EF. Although plasma CEA titers separated patients with either a benign or malignant EF 56% of the time, this separation was better accomplished (90% of the time) using EF CEA (P less than .0001). A malignant EF was associated with an EF CEA titer greater than or equal to ng/ ml or when, in the range 5.0 to 9.9 ng/ml, the EF CEA titer was greater than twice the plasma CEA. Paired plasma and EF CEA titers may aid in determining the benign or malignant origin of an EF.

Ectopic production of an alkaline phosphatase isoenzyme in patients with cancer.

An alkaline phosphatase isoenzyme (Regan isoenzyme) has been identified in the serum of 27 patients with various malignant tumors. This isoenzyme is biochemically and immunologically indistinguishable from placental alkaline phosphatase. In addition to being present in serum, it has been identified in both tumor tissue and malignant effusion fluids. Measurement of this isoenzyme has been clinically useful in monitoring progression or regression of tumor, identifying a source of serum alkaline phosphatase elevation, and identifying malignant effusions.