Malignant Peritoneal Effusions Research Articles

Abstract 229: Molecular-genomic characterization of metastatic gastric cancer organoids and establishment of organoid-immune cell co-culture model

Abstract Advanced gastric cancer (AGC) showed marked molecular heterogeneity with aggressive behavior and treatment resistance, especially AGC with ascites from peritoneal metastasis. Tumor organoids are valuable in vitro model for mimicking organ characteristics and are used for developing therapeutic strategies. Therefore, we aimed to understand the characteristics of organoids established from AGC patient-derived fluids. Also, we are in progress of setting up an organoid-immune cell co-culture model for drug screening of immune checkpoint inhibitors (ICI) for AGC. All established organoids from peritoneal and pleural fluid of AGC patients were authenticated through short tandem repeat (STR) profiling, comparing with their matched PBMCs. We compared the morphological characteristics using Operetta High Content Imaging (Perkin Elmer). Also, the in house targeted sequencing and RNA sequencing data were obtained from the genome database of the Song-Dang Institute for Cancer Research and it was verified using molecular methods. Then, we co-cultured organoids with healthy donor PBMCs that were pre-activated using anti-human CD3/CD28 Dynabeads (Gibco). 3D spheroids, rather than single cells, and the PBMCs were co-cultured in matrigel (Corning Inc.) for more than 3 days. The overall success rate of the organoid establishment was 53.8% (78/145), with different sources of peritoneal (54%, 61/113) and pleural fluids (53.1%, 17/32). The organoid success rate was various as in EBV positive (66.7%, 2/3) and dMMR (0%, 0/4). Based on the analysis of 91 patients eligible for histopathology, the success rate of HER2-negative (57.5%, 46/80) is higher than that of HER2-positive (36.4%, 4/11). The organoids were observed in a variety of morphologies categorized in 3 groups such as grape-like, compact and glandular structure. Based on genomic data of 31 organoids, we could subgrouping for TCGA subtypes and EMT subtype. RTKs were amplified in 10 organoids, including the amplification of ERBB2 (n=2), MET (n=4) and FGFR (n=4). And KRAS alterations including amplification (n=2) and G12A/D (n=2) mutation were also detected. Then, we established the condition for organoid-immune co-culture model with PBMCs with Dynabead activation for 3-10 days. We confirmed that higher dose of PBMCs killed the tumor cells with reduction of organoid size and partial disintegration. We established the proper PBMC dose and characterization to maintain the organoids co-culture for further drug (ICI) sensitivity testing. In conclusion, we evaluated phenotypic and molecular characteristics of gastric cancer organoids established from malignant peritoneal and pleural effusion. The established organoid-immune cell co-culture models can be utilized to evaluate a variety of drug screenings, including ICI and various combinations, helping in the selection of drugs for individual patient. Citation Format: Han Byeol Mun, juin Park, Jinsoo Jang, Jingmin Che, Tae Soo Kim, Woo Sun Kwon, Hyun Cheol Chung, Sun Young Rha. Molecular-genomic characterization of metastatic gastric cancer organoids and establishment of organoid-immune cell co-culture model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 229.

Abstract 3832: Fluid-overload associated large B-cell lymphoma and determinants of outcomes: Analysis of a pooled database

Abstract Background: Fluid overload-associated large B-cell lymphoma (FOLBCL) is a new WHO lymphoma entity previously known as PEL-like lymphoma or HHV8-unrelated PEL-like lymphoma. It is a rare lymphoma that arises within fluid-filled body cavities without a solid component. FOLBCL occurs in the absence of immunodeficiency and is commonly associated with conditions associated with fluid overload. It expresses mature B-cell phenotype and occasional EBV infection markers. We conducted this study to delineate key clinicopathological characteristics, prognostic indicators, and treatment modalities that affect outcomes in this rare and newly defined lymphoma subtype. Materials & Methods: To study the demographic characteristics, molecular and immunohistochemical signatures, therapeutic interventions, survival, and prognostic factors, we compiled a pooled database of 115 cases that fit the diagnostic criteria for FOLBCL. Kaplan-Meier survival curves were constructed. Cox proportional hazards model and Log-rank tests were used to assess the influence of demographic and clinicopathologic factors on overall survival (OS). Results: A total of 115 patients with confirmed FOLBCL were identified. The median age was 74, with a peak incidence between 72 and 82 years. There was a male preponderance with M:F of 2. The pleural, pericardial, and peritoneal cavities were involved in 75%, 34, and 27%. The median duration of symptoms before diagnosis was 1 month. Patients presented with constitutional symptoms and hepatosplenomegaly in 13% and 1%, respectively—no patients presented with lymph node and bone marrow involvement. The median OS of the whole group was 16 months. Patients younger than 70 had better median OS than 70 and older (24 vs. 10 months, p=0.008). The presence of any malignant peritoneal effusion was associated with worse OS when compared to effusion limited to the thoracic cavity (7 vs. 24 months, p=0.005). LDH>500 was also associated with worse OS (p=0.02). Sex, the presence of fluid overload-causing conditions, and serum IL2R levels did not seem to impact OS. MYC re-arrangement had numerically worse OS when present but did not reach statistical significance. Chemotherapy demonstrated superior OS compared to no treatment (24 vs. 7 months, p=0.009). Patients who attained CR as their best response had a superior median OS (p<0.0001). Conclusion: This study presents updated clinicopathologic data from a pooled cohort of patients with FOLBCL. It identifies the age, the primary location of cavitary disease, LDH, active therapy, and quality of response to treatment as key determinants of OS. Citation Format: Philip A. Haddad, Supriya Gupta, Christopher Graham. Fluid-overload associated large B-cell lymphoma and determinants of outcomes: Analysis of a pooled database [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3832.

Cascaded elasto-inertial separation of malignant tumor cells from untreated malignant pleural and peritoneal effusions.

Separation of malignant tumor cells (MTCs) from large background cells in untreated malignant pleural and peritoneal effusions (MPPEs) is critical for improving the sensitivity and efficiency of cytological diagnosis. Herein, we proposed a cascaded elasto-inertial cell separation (CEICS) device integrating an interfacial elasto-inertial microfluidic channel with a symmetric contraction expansion array (CEA) channel for pretreatment-free, high-recovery-ratio, and high-purity separation of MTCs from clinical MPPEs. First, the effects of flow-rate ratio, cell concentration, and cell size on separation performances in two single-stage channels were investigated. Then, the performances of the integrated CEICS device were characterized using blood cells spiked with three different tumor cells (MCF-7, MDA-MB-231, and A549 cells) at a high total throughput of 240 μL min-1. An average recovery ratio of ∼95% and an average purity of ∼61% for the three tumor cells were achieved. Finally, we successfully applied the CEICS device for the pretreatment-free separation of MTCs from clinical MPPEs of different cancers. Our CEICS device may provide a preparation tool for improving the sensitivity and efficiency of cytological examination.

Fluid-Overload Associated Large B-Cell Lymphoma Clinicopathologic Features and Determinants of Outcomes: Analysis of a Pooled Database

Background: Fluid overload-associated large B-cell lymphoma (FOLBCL) is a new WHO lymphoma entity previously known as PEL-like lymphoma or HHV8-unrelated PEL-like lymphoma. It is a rare lymphoma that arises within fluid-filled body cavities without a solid component. FOLBCL occurs in the absence of immunodeficiency and is commonly associated with conditions associated with fluid overload. It expresses mature B-cell phenotype and occasional EBV infection markers. We conducted this study to delineate key clinicopathological characteristics, prognostic indicators, and treatment modalities that affect outcomes in this rare and newly-defined lymphoma subtype. Methods: To study the demographic characteristics, molecular and immunohistochemical signatures, therapeutic interventions, survival, and prognostic factors, we compiled a pooled database of 115 cases that fit the diagnostic criteria for FOLBCL. Kaplan-Meier survival curves were constructed. Cox proportional hazards model and Log-rank tests were used to assess the influence of demographic and clinicopathologic factors on overall survival (OS). Results: A total of 115 patients with confirmed FOLBCL were identified. The median age was 74, with a peak incidence between 72 and 82 years. There was a male preponderance with M:F of 2. The pleural, pericardial, and peritoneal cavities were involved in 75%, 34%, and 27%, respectively. The median duration of symptoms before diagnosis was 1 month. Patients presented with constitutional symptoms and hepatosplenomegaly in 13% and 1%, respectively-no patients presented with lymph node and bone marrow involvement. The median OS of the whole group was 16 months. Patients younger than 70 had better median OS than 70 and older (24 vs. 10 months, p=0.008). The presence of any malignant peritoneal effusion was associated with worse OS when compared to effusion limited to the thoracic cavity (7 vs. 24 months, p=0.005). LDH≥500 was also associated with worse OS (p=0.02). Sex, the presence of fluid overload-causing conditions, and serum IL2R levels did not seem to impact OS. Compared to no treatment, chemotherapy had a numerically superior OS but did not reach statistical significance (19 vs. 10 months, p=0.11). However, patients that attained CR as their best response had a superior median OS (p<0.0001). Conclusions: This study presents updated clinicopathologic data from a pooled cohort of patients with FOLBCL. It identifies the age, the primary location of cavitary disease, LDH, active therapy, and quality of response to treatment as key determinants of OS.

Topical antibiotics prophylaxis for infections of indwelling pleural/peritoneal catheters (TAP-IPC): A pilot study.

Indwelling pleural catheter (IPC) and indwelling peritoneal catheter (IPeC) have established roles in the management of malignant pleural and peritoneal effusions but catheter-related infections remain a major concern. Topical mupirocin prophylaxis has been shown to reduce peritoneal dialysis catheter infections. This study aimed to assess the (i) compatibility of IPC with mupirocin and (ii) feasibility, tolerability and compliance of topical mupirocin prophylaxis in patients with an IPC or IPeC. (i) Three preparations of mupirocin were applied onto segments of IPC thrice weekly and examined with scanning electron microscope (SEM) at different time intervals. (ii) Consecutive patients fitted with IPC or IPeC were given topical mupirocin prophylaxis to apply to the catheter exit-site following every drainage/dressing change (at least twice weekly) and followed up for 6 months. (i) No detectable structural catheter damage was found with mupirocin applied for up to 6 months. (ii) Fifty indwelling catheters were inserted in 48 patients for malignant pleural (n = 41) and peritoneal (n = 9) effusions. Median follow-up was 121 [median, IQR 19-181] days. All patients tolerated mupirocin well; one patient reported short-term local tenderness. Compliance was excellent with 95.8% of the 989 scheduled doses delivered. Six patients developed catheter-related pleural (n = 3), concurrent peritoneal/local (n = 1) and skin/tract (n = 2) infections from Streptococcus mitis (with Bacillus species or anaerobes), Staphylococcus aureus, Klebsiella pneumoniae and Pseudomonas aeruginosa. This first study of long-term prevention of IPC- or IPeC-related infections found topical mupirocin prophylaxis feasible and well tolerated. Its efficacy warrants future randomized studies.

Hyperthermic intrathoracic/intraperitoneal chemotherapy versus conventional intrapleural/intraperitoneal chemotherapy for the malignant effusion: a multi-center randomized clinical trial

Objective To compare the efficacy and safety of hyperthermic intrathoracic/intraperitoneal chemotherapy versus conventional intrapleural/intraperitoneal chemotherapy in the treatment of malignant pleural or peritoneal effusion. Methods A randomized clinical trial was carried out in 8 cancer centers across China. Patients with malignant pleural or peritoneal effusion were randomly assigned to the study group or control group. Patients in the study group were treated with cisplatin-based hyperthermic intrathoracic chemotherapy (HITHOC) or hyperthermic intraperitoneal chemotherapy (HIPEC), while the control group was treated with conventional intrapleural or intraperitoneal chemotherapy using same chemotherapeutic regime as the study group. The objective response rate (ORR) was analyzed as primary outcome. Quality-of-life (QOL) score was recorded as secondary outcome using the questionnaire 30 (QLQ-C30) of the European Organization for Research and Treatment of Cancer (EORTC). The efficacy and safety of the two treatments were compared. Results Total 135 patients were recruited and randomized in this study, with 67 patients in the study group and 68 patients in the control group. The ORR in the study group (80.70%) was significantly higher than that in the control group (31.03%, p < 0.001). However, neither changes of QOL scores, nor incidence rates of adverse events were significantly different between the two groups (p = 0.076 and 0.197, respectively). Conclusion Efficacy of HITHOC or HIPEC is superior to that of conventional modality for the treatment of malignant effusion with comparable side effects.

Diagnostic Accuracy of Cell Block and Immunohistochemistry in Effusion Cytology.

Introduction Although the cytology of effusion fluids is considered a routine laboratory test, it has recently emerged as an essential tool in determining the primary site of origin of carcinoma of unknown primary. The sensitivity for diagnosing malignancy has enhanced with the inclusion of cytospin, cell block (CB), and immunohistochemistry (IHC) to effusion fluid cytology due to the improvement in morphological preservation and good cellular yield. The purpose of this study was to assess the diagnostic yield, sensitivity, specificity, positive predictive value, and negative predictive value of IHC and CB in effusion cytology. Methodology An institution-based cross-sectional study was conducted over a period of six months on 150 cases of effusion fluids submitted for diagnostic purposes. After the preparation of cytospin, the residual amount of centrifuged deposit was mixed with CytoLyt solution, thrombin, and plasma, and CBs were prepared. IHC was applied to the CB. Calretinin was used for mesothelial cells, and BerEP4, TTF-1, ER, WT-1, and CD-X2 were used for the confirmation and origin of malignant cells. Results The mean age of the patients was 51.75 ± 16.63 years. The male-to-female ratio was 1:1.24. Out of 150 cases, 78 were pleural effusions, 68 were peritoneal effusions, and four were pericardial effusions. Out of 150 cases, based on cytological examination alone, 66 (44%) were classified as benign, 27 (18%) as malignant, and 57 (38%) were suspicious for malignancy. When cytology was combined with CB and IHC, the diagnostic yield was increased to benign 95 (63.33%), malignant 48 (32%), and suspicious for malignancy 7 (4.67%). The most common cause of malignant pleural effusion was breast carcinoma in females and lung carcinoma in males. The most common primary tumor in malignant peritoneal effusion was ovarian carcinoma in females and colonic adenocarcinoma in males. The sensitivity and specificity of combined cytology with cell block and IHC were 92.31% and 98.95%, respectively. This combination produced significantly better results (p-value = 0.001) for detecting malignancy and reduced suspicious cases from 38% to 4%. Conclusion CB, in combination with IHC, increases the diagnostic yield and aids in detecting malignancy at an unknown primary site in effusion fluids. Both of these techniques can thus enhance the sensitivity and specificity of the diagnosis of effusion cytology. Hence, CB and IHC have advanced utility over cytological smears in effusion fluid cytological diagnosis.

Clinicopathological prognostic parameters in patients with tubo-ovarian carcinoma effusions.

ObjectiveTo analyse the predictive and prognostic role of clinicopathological parameters in patients with tubo‐ovarian carcinoma and malignant effusion.MethodsA retrospective series of 700 malignant peritoneal (n = 610) and pleural (n = 90) effusions from 558 patients was revised for histotype based on the 2014 World Health Organization criteria. The role of clinicopathological parameters in determining outcome was assessed.ResultsThe majority of specimens (597 effusions from 473 patients) were high‐grade serous carcinomas (HGSC), followed by low‐grade serous carcinoma (LGSC; 48 effusions, 37 patients), clear cell carcinoma (CCC; 23 effusions, 19 patients) and carcinosarcoma (CS; 16 effusions, 16 patients). Patients with CCC and CS had the shortest, those with HGSC intermediate, and those with LGSC longest overall and progression‐free survival (both P < 0.001). For patients with HGSC, older age (P = 0.002), more advanced FIGO stage (IV vs III; P < 0.001), delayed/no surgery (P < 0.001), larger residual disease volume (RD; P < 0.001), non‐complete response to chemotherapy at diagnosis (P < 0.001), and primary platinum resistance (P < 0.001) were associated with shorter overall survival. In Cox multivariate analysis, FIGO stage (P = 0.002) and primary platinum resistance (P < 0.001) were independent prognosticators. Significant association was additionally found for parameters analysed for progression‐free survival in HGSC (previous chemotherapy: P = 0.029; age: P = 0.046; FIGO stage, upfront therapy, RD: P < 0.001), of which previous chemotherapy, upfront therapy, and RD were independent prognosticators (all P < 0.001).ConclusionsThe vast majority of malignant effusions in patients with tubo‐ovarian carcinoma are derived from serous carcinoma or related tumours, such as CS. Histology is a powerful prognostic factor in this patient group, as are established clinical parameters.

In vitro cell culture of patient derived malignant pleural and peritoneal effusions for personalised drug screening

BackgroundMalignant serous effusion (MSE) denotes a manifestation of metastatic disease with typical high concentrations of both cancer and immune cells, making them an ideal resource for in vitro cytologic studies. Hence, the aim of the study was to investigate the features of 2D and 3D MSE culture systems as well as their feasibilities for in vitro drug screening.MethodsPleural and peritoneal effusions from 8 patients were collected and processed for 2D monolayer and 3D hanging drop cell culture into GravityPLUS™ plates. Representative markers for cell components, proliferation rate and tumour classification were investigated by immunohistochemistry, followed by absolute quantification using a digitalised image analysis approach. Further, we implemented another 3D cell culture model based on a low attachment method for in vitro drug sensitivity testing of carboplatin, pemetrexed and pembrolizumab for 5 patients.ResultsMonolayer cell culture was favourable for the growth of mesothelial cells, while hanging drop culture in GravityPLUS™ plates showed better ability for preserving cancer cells, inducing positive diagnostic markers expression and restraining the growth of mesothelial cells. For in vitro drug testing, MSE from five patients presented various drug sensitivities, and one case showed strong response to PD-1 checkpoint inhibition (pembrolizumab). For some patients, the application of combinatorial drugs had better therapeutic responses compared to monotherapy.ConclusionsDigitalised quantification of data offers a better understanding of different MSE culture models. More importantly, the proposed platforms are practical and amenable for performing in vitro chemo-/immunotherapeutic drug testing by using routine cytologic MSE in a personalised manner. Next to cell blocks, our work demonstrates the prognostic and predictive value of cytologic effusion samples.

Diagnosis of primary site of tumour origin using immunohistochemistry on cell block in cases of malignant ascitic fluid

Introduction: Presence of malignant cells in peritoneal fluid has serious implications as most of the times, it represents a metastasis from a distant organ, along with poor prognosis. Aim: 1) To ascertain primary site of origin from malignant ascitic fluid cases using cytoblock preparation and immunohistochemistry. 2) To find out frequency of various cancers from different parts of the body in malignant peritoneal fluid. Materials and methods: This is retrospective cross-sectional observation-based study in which 200 cases of malignant peritoneal fluid were studied and analysed. Routine clinical data, radiological investigations, histopathological diagnosis, cytomorphological features and immunohistochemistry were considered and statistically analysed. Results: 1) Malignant ascitic fluid cases are more common in older age group of patients and females outnumber the male patients substantially. 2) Adenocarcinoma is the most common histological type. 3) Ovary is the most common primary site of malignant peritoneal effusion. 4) Cytoblock preparation and immunohistochemistry on cytoblock can be very helpful in finding the primary site of origin in those cases where patients first presented to the hospital with signs of malignant ascites and no symptomatology related to primary site. Conclusion: Cytological examination of serous effusions is an accurate, prompt, affordable technique having diagnostic and therapeutic implications. With the help of ancillary methods, the phenotype of cells can be identified, classify as well as confirm our diagnosis.

Clinicopathological Study of 117 Body Fluids: Comparison of Conventional Smear and Cell Block Technique.

Background. Cell block method (CB) has emerged as an invaluable tool for diagnosis of effusions. It can help overcome the problems faced by conventional smear (CS) by differentiating between reactive, inflammatory and malignant cells. The aim of the study is to compare and correlate the CB diagnosis with the CS findings of various pathological conditions including malignancy. Materials and Methods. Two years prospective cross-sectional study of 117 fluids received for routine examination and/or for cytology was conducted. CS as well as CB was simultaneously prepared from the fluid and the results were correlated and tabulated for statistical analysis. Results. Mean age of presentation was 43±21.1 years and male: female ratio was 1.3:1. Ascitic fluid (46.2%) was the most common followed by pleural (40.2%). Among malignancies, primary ovarian and lung carcinoma were the most common to present with malignant ascites (33.3%) and pleural effusion (66.7%) respectively. Six suspicious for malignancy on CS were provided a definitive diagnosis of malignancy on CB. Overall, CB increased the yield of malignancy by 8.3%. The agreement between CB and CS for malignant effusions and suspicious for malignancy were 41.7% and 14.3% respectively. Sensitivity of CS method when compared to CB, for malignant peritoneal and pleural effusions was 90% and 75% respectively while the specificity was 68% and 79% respectively. Conclusion. CB has a better diagnostic yield of malignancy and helps in providing a definitive diagnosis for cases that are suspicious for malignancy on CS. Hence, CB should be routinely employed along with CS for all effusions.

Interleukin-2 boosts local and abscopal antitumor effect of radiotherapy combined with anti-PD-1: A translational research from clinical radio-memory effect.

e14244 Background: Preclinical and clinical studies have shown that prior receipt of radiotherapy enhances antitumor immune responses, a phenomenon we call the “radio-memory effect.” However, all of the evidence regarding this effect to date comes from work with anti-PD-1 or anti-PD-L1 inhibitors. Here we explore whether this effect also occurs with other forms of immune therapy, specifically interleukin-2 (IL-2). If so, whether this triple combination therapy could produce synergetic anti-tumor effect. Methods: 1. Clinical work: we retrospectively assessed outcomes in patients with malignant peritoneal effusion (MPE) who had previously received radiotherapy for colon or rectum cancer within 18 months before the intrapleural infusion of IL-2 or cisplatin. All patients received intrapleural infusion of IL-2 or cisplatin, and most had had several cycles of standard chemotherapy for colon or rectum cancer. 2. Translational work: MC38 cell line (5×106) was subcutaneously inoculated into the right lower extremity of C57/BL mice to construct a primary tumor model. Six days later, MC38 cell line (1×106) was subcutaneously inoculated into the left lower limb to construct a second tumor model. The main research is as follows: To explore the optimal radiation dose and fraction for radiotherapy; Whether the addition of IL-2 will further enhance the efficacy of radiotherapy combined with anti-PD-1; To explore the optimal combination model of radiation, anti-PD-1 and IL-2; Whether the addition of IL-2 will increase the abscopal effect produced by radiotherapy and anti-PD-1. Results: 1. We identified 3,747 patients with MPE (median age 66 years [range 24–81)) treated at one of several institutions from August 2009 through May 2016; 412 patients had been treated with IL-2 and 592 with cisplatin and had survived for at least 3 months afterward. 2. Hypofractionated radiotherapy (12Gy × 2) combined with IL-2 is better; IL-2 will enhance the efficacy of radiotherapy combined with anti-PD-1; IL-2 and anti-PD-1 followed by radiotherapy might maximize the anti-tumor effect; IL-2 might increase the abscopal effect produced by radiotherapy combined with anti-PD-1. Conclusions: We speculate that previous radiotherapy could enhance the efficacy of IL-2 and the anti-tumor effect produced by radiotherapy combined with anti-PD-1 could be enhanced by IL-2. The “radio-memory” effect could be beneficial in future studies.

Application value of paraffin-embedded section of cell block in differential diagnosis of benign or malignant pleural and peritoneal effusion

Objective To investigate the value of paraffin-embedded section of cell block in the diagnosis of benign or malignant pleural and peritoneal effusion. Methods 105 patients suspected with malignant pleural and peritoneal effusion admitted into tour hospital from November, 2015 to December, 2017 were selected as the research objects. The specimens of pleural and peritoneal effusion of all the patients were collected. And these specimens were given paraffin-embedded section of cell block, cell smear, and exfoliative cell pathological examination. The results of exfoliative cell pathological examination were taken as the gold standard. The positive rates of malignant pleural and peritoneal effusion diagnosed by paraffin-embedded section of cell block and cell smear were evaluated. The diagnostic value of the two methods was compared. Results Among the 50 pleural effusion specimens, the results of exfoliative cell pathological examination showed that there were 36 cases of malignant pleural effusion. Among the 55 peritoneal effusion specimens, the results of exfoliative cell pathological examination showed that there were 42 cases of malignant peritoneal effusion. The positive detection rate of pleural effusion diagnosed by paraffin-embedded section of cell block (80.56%) was higher than that diagnosed by cell smear (58.33%), with a statistical difference (P<0.05). The positive detection rate of peritoneal effusion diagnosed by paraffin-embedded section of cell block (85.71%) was higher than that diagnosed by cell smear (64.29%), with a statistical difference (P<0.05). The sensitivity, specificity, and accuracy of paraffin-embedded section of cell block in the diagnosis of malignant pleural and peritoneal effusion were higher than those of cell smear, with statistical differences (all P<0.05). Conclusions Paraffin-embedded section of cell block in the differential diagnosis of benign or malignant pleural and peritoneal effusion has high sensitivity and specificity, and can increase the diagnostic accuracy and provide an important basis for clinical diagnosis and treatment of the disease. Key words: Pleural effusion; Paraffin-embedded section of cell block; Cell smear; Exfoliative cell pathological examination

Analyzing diagnostic utility of cytospin and cell block techniques in serous effusion cytology

Introduction: Fluid cytology plays a pivotal role delineating benign from malignant effusions, tumor staging and diagnosing recurrences. Objectives: To assess the diagnostic utility of cytospin and cell block methods in cytodiagnosis of serous effusions.Material and methods: A total of 240 cases inclusive of 77 pleural and 163 peritoneal effusions were analyzed by cytospin and cell block preparation following conventional smear cytology.Results: Out of 240 cases, 13% were malignant pleural effusion and 14.7% were malignant peritoneal effusion. Cell block diagnosed additional 4% and 2.4% cases of malignant origin in pleural and peritoneal fluid respectively and also aided in ascertain the primary site. Few rare cases such as effusion in synovial sarcoma and squamous cell carcinoma were also diagnosed. Conclusion: Cell block technique aids in cytodiagnosis of malignant effusions but has demerits also, hence both cytospin and cell block should be used as complement.

Comparative study of paired box gene 8, cytokeratin 7, and cytokeratin 20 in differentiating the ovarian from nonovarian carcinoma cells in peritoneal effusions

Background In female patients, ovarian carcinoma is considered a common cause of malignant peritoneal effusions that shows a good response to chemotherapy, an issue that mandates quick and accurate diagnosis of those cases. PAX8 is a cell lineage factor that plays a crucial role in the morphogenesis of the kidney, thyroid gland, and Mullerian system. It is expressed in 95–100% of ovarian carcinomas in tissue sections. Recently, it was suggested as a potential marker for the diagnosis of ovarian carcinomas in effusions. Aim This study aimed at assessing the role of CK7, CK20, and PAX8 in differentiating ovarian from nonovarian carcinomas in peritoneal effusions. Patients and methods Immunocytochemical evaluation of CK7, CK20, and PAX8 expression in 40 cell blocks of malignant peritoneal effusion samples was carried out. In addition, the assessment of PAX8 level using double-antibody sandwich enzyme-linked immunosorbent assay technique in the serum of the included cases was also carried out. Results Of the studied 40 cases, 21 (52.5%) proved to be ovarian carcinomas, whereas 19 (47.5%) were nonovarian. Diffuse CK7 expression (scores 3, 4) was significantly detected more frequently in effusions from ovarian than from nonovarian carcinomas. In contrast, CK20 expression tended to be more prevalent in effusions of nonovarian than in those of ovarian origin; however, this was statistically insignificant. PAX8 expression was significantly more frequent in effusions from ovarian carcinomas than from nonovarian carcinomas. Sensitivity and specificity of PAX8 in the diagnosis of ovarian carcinoma was higher than that of CK7, while a combination of both CK7 and PAX8 immune scores using a generated regression equation was the most specific. In contrast, serum PAX8 level was statistically higher in ovarian more than in nonovarian carcinoma cases. Conclusion PAX8 is a useful immune marker for the diagnosis of ovarian carcinoma in peritoneal effusions. However, the combination of both CK7 and PAX8 using a generated regression equation increases the specificity of diagnosis. PAX8 serum level is a potential serum marker in diagnosing ovarian carcinomas.

Discerning the Primary Carcinoma in Malignant Peritoneal and Pleural Effusions Using Imaging Mass Spectrometry-A Feasibility Study.

Malignant effusions challenge diagnostic accuracy due to cytomorphologic overlaps between various malignant primaries. Workup of this material to establish a correct diagnosis is time consuming and limited by the sparsity of material. In order to circumvent these drawbacks, the use of MALDI imaging MS (IMS) as a diagnostic platform has been explored. Cytology cell blocks from malignant effusions (serous ovarian carcinoma and several non-ovarian carcinomas including gastric adenocarcinoma) containing at least 30% neoplastic cells are selected for generation of cytology microarrays (CMA). CMA sections are transferred to conductive glass slides, subjected to on-tissue tryptic digestion, and matrix application for MALDI-IMS analysis. Supervised classification analysis identifies serous ovarian carcinomas as the source of malignant effusions with a sensitivity of 85.7% when compared to samples from all other included primary sites. When compared to gastric adenocarcinoma, serous ovarian carcinoma samples can be delineated with a sensitivity of 97.3%. These preliminary results highlight that MALDI-IMS allows subtyping of malignant effusions to identify the precise origin of neoplastic cells. While achieving similar results compared to classical approaches such as immunocytology, more material is conserved that will be available for further tests.

Prognostic Significance of Blood, Serum, and Ascites Parameters in Patients with Malignant Peritoneal Mesothelioma or Peritoneal Carcinomatosis.

To determine effects of the biochemical and cytological properties of blood, serum, and ascites on survival of patients with malignant peritoneal effusion (MPeE), including malignant peritoneal mesothelioma (MPeM) and peritoneal carcinomatosis (PC), we conducted a retrospective study of patients with MPeE and healthy controls. Potential prognostic factors were identified as follows: age, sex, blood neutrophil-to-lymphocyte ratio (NLR), serum parameters, ascites parameters, serum-ascites albumin gradient, and the ascites-serum LDH ratio. Compared to those of the control group, serum albumin levels were significantly lower, and the NLR and serum LDH levels were significantly higher in the MPeE group. Overall survival (OS) was longer in patients with MPeM compared to that in patients with PC. Compared with patients in the MPeM, patients with PC had higher NLRs, ascites glucose levels, serum-ascites albumin gradients, and serum LDH levels. In contrast, their ascites albumin levels and ascites-serum LDH ratios were lower. Univariate analyses indicated that the NLR, serum LDH levels, ascites LDH levels, ascites coenocyte levels, and the ascites coenocyte-to-monocyte ratios affected the OS. Multivariate analyses identified only serum and ascites LDH levels as independent prognostic factors.

Loss of steroid hormone receptors is common in malignant pleural and peritoneal effusions of breast cancer patients treated with endocrine therapy.

Discordance in estrogen receptor alpha (ERα), progesterone receptor (PR), androgen receptor (AR) and human epidermal growth factor receptor 2 (HER2) status between primary breast cancers and solid distant metastases (“conversion”) has been reported previously. Even though metastatic spread to the peritoneal and pleural cavities occurs frequently and is associated with high mortality, the rate of receptor conversion and the prognostic implications thereof remain elusive.We therefore determined receptor conversion in 91 effusion metastases (78 pleural, 13 peritoneal effusions) of 69 patients by immunohistochemistry (IHC) and in situ hybridization. Data were coupled to clinical variables and treatment history.ERα, PR and AR receptor status converted from positive in the primary tumor to negative in the effusion metastases or vice versa in 25-30%, 30-35% and 46-51% of cases for the 1% and 10% thresholds for positivity, respectively. 19-25% of patients converted clinically relevant from “ERα+ or PR+” to ERα-/PR- and 3-4% from ERα-/PR- to “ERα+ or PR+”. For HER2, conversion was observed in 6% of cases. Importantly, receptor conversion for ERα (p = 0.058) and AR (p < 0.001) was more often seen in patients adjuvantly treated with endocrine therapy. Analogous to this observation, HER2-loss was more frequent in patients adjuvantly treated with trastuzumab (p < 0.001).Alike solid distant metastases, receptor conversion for ERα, PR, AR and HER2 is a frequent phenomenon in peritoneal and pleural effusion metastases. Adjuvant endocrine and trastuzumab therapy imposes an evolutionary selection pressure on the tumor, leading to receptor loss in effusion metastases. Determination of receptor status in malignant effusion specimens will facilitate endocrine treatment decision-making at this lethal state of the disease, and is hence recommended whenever possible.

Utility of cell block to detect malignancy in fluid cytology: Adjunct or necessity?

Cell block (CB) technique when supplemented with conventional smear, provides increased cellularity, preservation of architectural pattern with excellent morphology, and a clear background. We compare the utility of CB technique compared to conventional smear in detection of malignancy in serous effusions. An institution-based observational and analytical study was carried out over 1 year on 50 patients with effusions. The residual amount of centrifuged deposit after preparation of conventional smear was mixed with 10% alcohol-formalin solution, and CBs were prepared. Calretinin and cytokeratin 5 were used for reactive mesothelial cells and Wilms tumor 1, thyroid transcription factor 1, CDX2, and estrogen receptor were used to confirm the adenocarcinoma cells. Maximum patients belonged to the age group of 61-70 years. Male:female ratio 1:1.17. Most common cause of malignant peritoneal effusion was due to ovarian malignancies in females and adenocarcinoma of stomach in males while, in case of pleural effusion, it was breast carcinoma in females and lung carcinoma in males. Thirteen suspicious cases were subjected to immunohistochemistry (IHC). In 70% cases, CB findings were consistent with the findings of conventional smears. In 20% cases, the conventional smears were suspicious for malignancy, and malignancy was confirmed by CB technique, whereas in 10% cases, both smears and CB were suspicious for malignancy and the original nature of the lesion was confirmed by the IHC. Sensitivity and specificity of CB compared to conventional smear were 88.88% and 86.98%, respectively. CB produced significantly better results (P = 0.0271) while detecting malignant lesions and reducing suspicious results (P = 0.0226).

Diagnostic value of DNA quantitative analysis and thinprep cytologic test in diagnosis of malignant pleural effusion and ascites

Objective To discuss the application value of DNA quantitative analysis and thinprep cytologic test for the diagnosis of malignant peritoneal effusion. Methods Fifty-eight cases patients with pleural effusion and ascites in the Second People′s Hospital of Yueyang were involved in this study.DNA quantitative analysis and thinprep cytologic test were conducted for the collected ascites, than compared the sensitivity of that two methods. Results There were 39 cases patients were diagnose malignant tumor by biopsy or pathology, the positive rate of DNA quantitative analysis(92.31%, 36/39) was obviously higher than that of thinprep cytologic test(82.05%, 32/39), the results had no statistical significance(P=0.017). The consistency check kappa value of DNA quantitative analysis and Thinprep cytologic test with pathology diagnosis were 0.807 and 0.498 respectively.The AUC of the combined examination of DNA quantitative analysis and thinprep cytologic test for malignant peritoneal effusion was 0.92, higher than the AUC of DNA quantitative analysis and Thinprep cytologic test single index examinations(0.90, 0.75). The detection sensitivity of DNA quantitative analysis and thinprep cytologic test were 94.74%(36/38) and 84.21%(32/38) respectively, specificity were 85.00%(17/20) and 65.00%(13/20) respectively, the dia were 91.38%(53/58), 77.59%(45/58) respectively, the joint detection sensitivity was 94.87%(37/39), specificity was 95.00%(19/20), the diagnostic accuracy was 93.10%(54/58). Conclusion Compared with thinprep cytologic test, DNA quantitative analysis with high sensitivity and specificity, while the combination of the two methods can greatly improve the accuracy of diagnosis of malignant ascites. Key words: Ascites; DNA quantitative analysis; Thinprep cytologic test