scholarly journals Study of body fluid samples using flow cytometry: Six years of experience at the Hospital Universitario San Ignacio - Pontificia Universidad Javeriana Bogota - Colombia

2017 ◽  
Vol 22 (2) ◽  
pp. 123
Author(s):  
Alba Campos ◽  
Laura Trujillo ◽  
Derly López ◽  
Lina Beltrán ◽  
Eliana Arias ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Cerebrospinal Fluid ◽  
Bronchoalveolar Lavage ◽  
Solid Tumors ◽  
Tumor Cells ◽  
Body Fluid ◽  
Hematological Malignancies ◽  
Pericardial Fluid ◽  
Malignant Diseases

Flow cytometry (FCM) was implemented in 2008 at the Pontificia Universidad Javeriana and later at the Hospital Universitario San Ignacio to examine special samples of patients with hematological malignancies and solid tumors other than bone marrow and peripheral blood for diagnosis and monitoring. This study describes the main findings of special sample evaluation over a six-year period. In all, 1070 samples of body fluids from patients with benign and malignant diseases were examined by FCM. These samples were stabilized with TransFixTM and stained with six-color immunophenotyping panels. Samples included cerebrospinal fluid, bronchoalveolar lavage, pleural fluid, pericardial fluid and ascite fluid from patients with acute and chronic leukemia, myelodysplastic syndromes, lymphomas, myeloma, autoimmune diseases, immunodeficiencies and solid tumors, among others. Flow cytometry provided important information for the classification and detection of minimal numbers of tumor cells in leukemia and lymphoma cases. This work represents the first national report describing FCM implementation in special samples for diagnosis and clinical monitoring of patients with malignant and benign pathologies.

Flow Cytometric Examination of Non-Hematic Body Fluids in Hematologic Malignancies: A Comparison with Cytology.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4237-4237
Author(s):  
Clara Cesana ◽  
Barbara Scarpati ◽  
Bruno Brando ◽  
Claudia Barba ◽  
Ursula Ferri ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
B Cell ◽  
Tumor Cells ◽  
Body Fluid ◽  
Hematologic Malignancies ◽  
Hematologic Neoplasms ◽  
Becton Dickinson ◽  
Neoplastic Cells ◽  
Disease Specific ◽  
Large B Cell

Abstract Introduction: Flow cytometry is well known to detect malignant cells in peripheral blood and bone marrow of patients with hematologic malignancies. However, its role in evaluating non-hematic body fluid contamination by tumor cells is largely unexplored. Patients and Methods: Data detected by flow cytometry in non-hematic body fluid samples drawn between 2002 and 2007 from patients with hematologic neoplasms were retrospectively compared with morphological findings obtained from cytospin slides. Immunophenotyping was carried out by using disease-specific multicolor panels of quadruple monoclonal antibodies, conjugated with the fluorochromes FITC, PE, PerCP, and APC, respectively. Acquisition of information on 1x104 to 1x105 stained cells depending on the whole sample cellularity was assessed on a dual-laser FACSCalibur flow cytometer using the CellQUEST software (Becton Dickinson, San José, CA, USA). Results: Fourty-five samples (bronchoalveolar fluid, n=5; ascites, n=2; hydrocele, n=2; pleural effusion, n=8; and cerebrospinal fluid, n=28) from 32 patients were available for comparison. Diagnoses were as follows: chronic myelomonocitic leukemia (n=1), acute promyelocytic leukaemia (n=2), acute myelomonocytic leukaemia (n=1), B-chronic lymphocitic leukemia (n=4), follicular lymphoma (n=1), acute lymphoblastic leukaemia (n=6), lymphoblastic lymphoma (n=1), high grade non-Hodgkin’s lymphoma (NHL), Burkitt-like (n=3), diffuse large B-cell NHL (n=6), peripheral T-cell NHL (n=3), and NHL, unspecified (n=4). Flow cytometry detected neoplastic cells in 24 cases. Of these cases, only 17 were positive also by morphology. In 7 cases, in which tumor cells were detected by flow cytometry but not by morphology, clinical data confirmed the presence of the disease. Flow cytometry did not show neoplastic cells in 21 cases. Of these cases, only 18 were negative also by morphology. In the remaining 3, the suggestion of diffuse large B-cell NHL contamination by morphology was not confirmed by flow cytometry, demonstrating T-reactive lymphocytes that were clearly negative for disease-specific markers. Conclusions: Our data suggest that flow cytometry is a useful tool complementary to morphology for the screening of non-hematic body fluid contamination in patients with hematologic neoplasms.

scholarly journals Bone marrow as a metastatic niche for disseminated tumor cells from solid tumors

2015 ◽  
Vol 4 ◽  
Author(s):  
Yusuke Shiozawa ◽  
Matthew R Eber ◽  
Janice E Berry ◽  
Russell S Taichman
Keyword(s):  
Bone Marrow ◽  
Solid Tumors ◽  
Tumor Cells ◽  
Disseminated Tumor Cells ◽  
Metastatic Niche

Plasma levels of tumor M2-pyruvate kinase should not be used as a tumor marker for hematological malignancies and solid tumors

2006 ◽  
Vol 44 (1) ◽  
Author(s):  
Peter Staib ◽  
Melanie Hoffmann ◽  
Timo Schinköthe
Keyword(s):  
Tumor Marker ◽  
Solid Tumors ◽  
Pyruvate Kinase ◽  
Plasma Levels ◽  
Hematological Malignancies ◽  
Malignant Diseases ◽  
Healthy Individuals ◽  
The Mean ◽  
Circulating Levels ◽  
Systemic Therapies

AbstractIt has been reported that the dimeric isoform of the enzyme pyruvate kinase M2 was overexpressed in various solid tumor cells. Hence, it was suggested that circulating levels of the so-called tumor M2-pyruvate kinase (Tu M2-PK) could be used as a tumor marker for monitoring systemic therapies of various solid tumors. We analyzed its validity as a tumor marker by comparing plasma levels of Tu M2-PK in patients with different non-malignant diseases to levels in healthy individuals and in patients with hematological diseases. Plasma levels of Tu M2-PK were measured using an ELISA assay in a total of 284 patients. The mean Tu M2-PK concentration of 32 U/mL was significantly higher in the group of patients with hematological malignancies (n=121) (p<0.001). However, 37% of healthy individuals (n=63) and 44% of patients with non-malignant diseases (n=100), especially patients with an acute inflammatory reaction (67%), were found to have elevated levels of Tu M2-PK using a cutoff level of 15U/mL. The specificity was 59% and the sensitivity was 51%. There was no significant correlation between the prevalence of a hematological malignancy and positive Tu M2-PK result. Thus, our data imply that Tu M2-PK is not a useful tumor marker for hematological malignancies and solid tumors, as a significant number of false positive results were detected in healthy individuals and patients with non-malignant diseases.

scholarly journals Malignancies and MDS in Patients with Diamond-Blackfan Anemia (DBA) from the Czech National DBA Registry

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5017-5017
Author(s):  
Dagmar Pospisilova ◽  
Jaroslav Cermak ◽  
Monika Belickova ◽  
Monika Horvathova ◽  
Jana Volejnikova
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Solid Tumors ◽  
Cancer Incidence ◽  
B Cell Lymphoma ◽  
Erythroid Cells ◽  
Diamond Blackfan Anemia ◽  
Risk Of Cancer ◽  
Malignant Condition

Introduction: Diamond-Blackfan anemia (DBA) is an inherited bone marrow failure syndrome characterized by macrocytic anemia, reticulocytopenia, selective deficiency of erythroid precursors, presence of congenital anomalies and an increased risk of cancer. DBA is caused by germline mutations of genes coding for ribosomal proteins (RP). Interestingly, somatic RP mutations have also been found in several malignant diseases (T-ALL, CLL, Hodgkin lymphoma, myelodysplastic syndrome (MDS) and AML). The only representative overview of malignancies in DBA was published in 2012 based on the data from the North American DBA Registry. The malignancies developed in 3% of patients (MDS and AML in 4 patients and solid tumors in 15 patients). The observed-to-expected ratio for all cancers combined was 5.4 (p<0.05). Median age at diagnosis was 41 years and cumulative incidence of solid tumors/leukemias was approximately 20% at the age of 46 years. Five years later, additional solid tumors were reported, in particular gastrointestinal tumors, and an incidence of MDS was alarming. Our aim was to evaluate cancer incidence within Czech National DBA Registry and characterize underlying molecular pathology. Patients and methods: Czech National DBA Registry currently includes 62 patients. In cooperation with 8 national centers of pediatric and adult hematology, we collected data about patients followed with pre-malignant or malignant condition including a detailed analysis of the course of the disease, treatment and type of developed malignant disorder. In patients with an evolving MDS, a classical cytogenetic analysis, flow cytometry, mutational profile (TruSight Myeloid Sequencing Panel (Illumina) containing 54 genes) and commercial TUNEL assay for detection of apoptotic changes of erythroid cells in the bone marrow were performed. Results: Eight of 62 patients from the Registry (13%) had malignant or pre-malignant condition: four females (6.5%) had solid tumors, 3 males (4.8%) had MDS and one female (1.6%) had multiple myeloma. Age of the onset of these disorders ranged between 25-70 years. Three patients harbored RPL5 mutation, 3 patients RPL11 mutation and 2 patients RPS19 mutation. Cancer incidence was significantly higher within the RPL11 and RPL5 subgroups (p=0.0056, Fisher Exact Test). Two female patients were diagnosed with triple-negative breast carcinoma, both of them during pregnancy. The first patient died despite treatment at the age of 29 years, shortly after delivery of her second child. The second patient is currently undergoing neoadjuvant chemotherapy.One patient developed diffuse large B cell lymphoma (DLBCL), underwent chemotherapy and autologous BMT and is alive in second remission.One patient underwent hemicolectomy for colorectal adenocarcinoma at 53 years of age and is in remission 6 years after the surgery.One patient succumbed to multiple myeloma which evolved from monoclonal gammopathy of unknown significance (MGUS) after 14 years of follow-up.Three male patients developed suspected MDS at the age of 25, 28 and 29 years. Two of them had RPL5 mutation and one harbored RPS19 mutation. None of them had decrease of erythroid cells in the bone marrow, but apoptosis of erythroid progenitors was significantly increased in all cases. All 3 patients had bicytopenia in peripheral blood (anemia, leukopenia) and dysplasia in 2 or 3 hematopoietic lineages in the bone marrow, thus fulfilling criteria for MDS with multilineage dysplasia (MDS-MLD). They had no abnormalities detected by flow cytometry or cytogenetics. The patient with RPS19mut harbored ASXL1 mutation; in patient with RPL5 mut, mutational screening is ongoing. Conclusion: Our results confirmed increased incidence of cancer (13%) in patients with DBA at young age. Our cases of DLBCL and MGUS in DBA are the first published to date. In cases with suspected MDS, cytopenia and dysplastic changes in the bone marrow may either reflect specific features of ribosomopathy, or represent a severe disorder of regulatory mechanisms with propensity to clonal proliferation. International collaboration is required to refine the incidence of malignancies in DBA and to issue consensus guidelines for timely detection of solid tumors, leukemias and MDS. The best approach to DBA patients who developed MDS without harboring any mutation considered prognostically significant in MDS, is another subject of discussion. Support: NV16-32105A Disclosures No relevant conflicts of interest to declare.

scholarly journals Original Article Non-Hematological Malignancies Seeding in Bone Marrow

2020 ◽  
Vol 7 (7) ◽  
pp. A341-348
Author(s):  
Gaurav PS Gahlot ◽  
Ankur Ahuja ◽  
Ajay Kumar Baranwal ◽  
Gunjan Madhukar ◽  
Bhushan Asthana ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Solid Tumors ◽  
Hematological Malignancies ◽  
Hematological Parameters ◽  
Bone Marrow Involvement ◽  
Primary Site ◽  
Research Centre ◽  
Lytic Lesion ◽  
Unknown Primary ◽  
Hematopoietic Tissue

Background: Bone marrow is the site of origin for primary haematological malignancies and is the third common preferred site metastasized by the solid tumors. The malignant infiltration of the hematopoietic tissue alters the clinical course of disease, response of the treatment and influences the overall survival. The aim of this study was to assess pattern of bone marrow involvement by different solid tumors and their correlation with hematological parameters. Methods: In this retrospective study, 8064 bone marrow examinations from Jan 2011-Aug 2017 at tertiary health and research centre of Northern India were evaluated to access spectrum of different solid tumors infiltrating the bone marrow alongwith their clinical, hematological and histopathological findings. Result: Total 38 cases of non-hematological malignancies metastasizing to bone marrow were evaluated with main indications of lytic lesions, cytopenia and Pyrexia of Unknown Origin. The most common metastasis were adenocarcinoma of prostate and lung. In 33 cases, the clinical, cytomorphological and immunohistochemical analysis findings were correlated to know primary site, while in remaining five patients, even after complete diagnostic evaluation, the definitive origin could not be ascertained, therefore categorized as Carcinoma of Unknown Primary Site (CUPS). Conclusion: Our series showed that anemia as commonest parameter, followed by leukopenia, thrombocytopenia. Many cases were misdiagnosed as multiple myeloma due to lytic lesion, anemia and hypercalcemia. We concluded that unexplained cytopenia are strong indicators of bone marrow examination; an easy, convenient, sensitive, effective procedure of staging of tumor, monitoring the course and prognosis of solid tumors.

scholarly journals Detection of Circulating Tumor Plasma Cells in Monoclonal Gammopathies: Methods, Pathogenic Role, and Clinical Implications

Cancers ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 1499
Author(s):  
Luzalba Sanoja-Flores ◽  
Juan Flores-Montero ◽  
Martín Pérez-Andrés ◽  
Noemí Puig ◽  
Alberto Orfao
Keyword(s):  
Solid Tumors ◽  
Tumor Cells ◽  
Plasma Cell ◽  
Hematological Malignancies ◽  
Plasma Cells ◽  
Low Frequency ◽  
Strong Impact ◽  
Monoclonal Gammopathies ◽  
Plasma Cell Neoplasms ◽  
Cancer Dissemination

Cancer dissemination and distant metastasis most frequently require the release of tumor cells into the blood circulation, both in solid tumors and most hematological malignancies, including plasma cell neoplasms. However, detection of blood circulating tumor cells in solid tumors and some hematological malignancies, such as the majority of mature/peripheral B-cell lymphomas and monoclonal gammopathies, has long been a challenge due to their very low frequency. In recent years, the availability of highly-sensitive and standardized methods for the detection of circulating tumor plasma cells (CTPC) in monoclonal gammopathies, e.g., next-generation flow cytometry (NGF), demonstrated the systematic presence of CTPC in blood in virtually every smoldering (SMM) and symptomatic multiple myeloma (MM) patient studied at diagnosis, and in the majority of patients with newly-diagnosed monoclonal gammopathies of undetermined significance (MGUS). These methods set the basis for further detailed characterization of CTPC vs. their bone marrow counterpart in monoclonal gammopathies, to investigate their role in the biology of the disease, and to confirm their strong impact on patient outcome when measured both at diagnosis and after initiating therapy. Here, we review the currently available techniques for the detection of CTPC, and determine their biological features, physiopathological role and clinical significance in patients diagnosed with distinct diagnostic categories of plasma cell neoplasms.

Constitutive Expression of CD40L on Bone Marrow Mast Cells Supports the Growth of Lymphoplasmacytic Cells in Patients with Waldenstrom’s Macroglobulinemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2350-2350 ◽  
Author(s):  
Olivier Tournilhac ◽  
Daniel Ditzel Santos ◽  
Lian Xu ◽  
Jeffery Kutok ◽  
Yu Tsu Tai ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Mast Cells ◽  
Cell Surface ◽  
Tumor Cells ◽  
Thymidine Uptake ◽  
Dependent Manner ◽  
Rt Pcr ◽  
Potent Inducer ◽  
Dose Dependent

Abstract CD40 ligand (CD40L) is a potent inducer of normal and malignant B-cell proliferation through interaction with CD40. We and others have observed excess mast cells (MC) in bone marrow (BM) biopsies of WM patients, which are commonly found admixed with tumor aggregates. (Tournilhac et al, JCO 2004, 22:571S). We therefore sought to clarify the role of MC in WM. Co-culture of 0.5% paraformaldehyde fixed, or sublethally irradiated HMC-1, LAD, and KU mast or basophilic cell lines and sorted BM lymphoplasmacytic cells (LPC) from 10 WM patients resulted in MC dose-dependent tumor colony formation and/or proliferation as assessed by 3H-thymidine uptake studies. As demonstrated by immunohistochemical, multicolor flow cytometric (CD117+FceRI+) and/or RT-PCR analysis, CD40L was expressed on BM MC from 29 of 31 (94%), 11 of 13 (85%), and 7 of 9 (78%) of WM patients, respectively. In contrast, cell surface CD40L expression was not detected by immunohistochemistry (p=0.00005) and flow cytometry (p=0.003) in 5 normal donors, and only faint expression for 1 of 5 normal donors by RT-PCR (p=0.09). Moreover, by multicolor flow cytometry, CD40 was expressed on BM tumor cells from 14/17 (83%) patients. CD40 functionality was confirmed either by the G28.5 CD40 agonistic antibody which induced dose dependent proliferation or by the rh-CD40L which partly prevented serum starvation-induced-apoptosis of WM LPC from 4/4 and 3/4 patients respectively. Importantly, expansion of tumor cells from 3 of 4 patients in mixed cultures with paraformaldehyde fixed MC was blocked in a dose dependent manner by use of a CD40L blocking protein (CD40:Fc). These studies demonstrate that CD40L is constitutively expressed on the cell surface of BM MC in WM and support the growth of WM tumor cells, and therefore provide the framework for therapeutic targeting of MC and MC-WM cell interactions in WM.

Serum Cytochrome C as a Tumor Marker in Leukemia and Lymphoma.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4435-4435
Author(s):  
Akemi Osaka ◽  
Takashi Sawada ◽  
Yasuaki Yamada ◽  
Muneo Aoyama ◽  
Hiroo Hasegawa ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Death ◽  
Cytochrome C ◽  
Tumor Marker ◽  
Tumor Cells ◽  
Hematological Malignancies ◽  
Healthy Adults ◽  
Cell Counts ◽  
Ldh Activity ◽  
Cyt C

Abstract When cells become apoptotic, cytochrome c (Cyt c) is translocated from the mitochondria to the cytosol. It has been reported that Cyt c released to the cytosol is further released from the cells into the extracellular medium. Since tumor cells show an accelerated cell cycle with rapid cell growth accompanied by various grades of cell death including apoptosis, sera from patients with malignancies may contain increased levels of Cyt c. We thus investigated serum Cyt c as a tumor marker in hematological malignancies. Cyt c concentration was measured by our original electrochemiluminescence immunoassay. The measurement range of this assay was 5 to 3,000 ng/ml in serum. We confirmed that cell lines triggered by inducers such as anti-Fas/CD95 or TNF-related apoptosis-inducing ligand (TRAIL) released Cyt c into the culture medium within several hours. The serum Cyt c concentration in healthy adults showed a distribution from 14.6 to 36.1 ng/ml (median value: 25.1). Unexpectedly, there was no significant difference between healthy adults and patients with AML, CML, ALL, CLL, multiple myeloma (MM), non-Hodgkin Lymphoma (NHL) or adult T cell leukemia/lymphoma (ATLL), and the median values in these diseases were distributed between 19.7 and 42.4. Of note, however, there were some patients whose Cyt c concentrations were extremely high up to 2,533.2 ng/ml. Values over 50ng/ml were observed in three of the 10 patients with AML (30%), three of the 6 patients with ALL (50%), five of the 21 patients with ATLL (23.8%) and one of the 6 patients with NHL (16.7%). In contrast, there were no patients with CML, CLL or MM who showed values higher than 50 ng/ml. On subtype-analysis in ATLL, the five patients who showed more than 50 ng/ml all had aggressive subtypes, and none of the patients with indolent subtypes showed values higher than 50 ng/ml There was a significant positive correlation between Cyt c concentration and absolute ATLL cell counts in the peripheral blood. More interestingly, parallel analysis using serum and bone marrow specimens in AML and ALL indicated that bone marrow contained more than 10 times higher Cyt c than serum. These results indicate that certain rapidly growing tumor cells exhibit increased spontaneous apoptotic cell death, and serum and/or bone marrow Cyt c concentration can be a reliable marker to estimate the extent. Then, we examined serial serum Cyt c levels during chemotherapy to see whether they reflect the effect of chemotherapy. There was a clear correlation between serum Cyt c and the effect of chemotherapy; Cyt c increased several times soon after the start of chemotherapy, then returned to normal levels after completion. Although serum LDH activity also showed similar variation, the response of serum Cyt c was much quicker and sharper than that of LDH activity. In conclusion, serum Cyt c serves as a marker not only for spontaneous tumor cell death but also for monitoring chemotherapeutic response. Monitoring Cyt c provides new insight for understanding the pathophysiology of hematological malignancies.

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