Cytoplasmic Expression of CD3ε Heterodimers by Flow Cytometry Rapidly Distinguishes Between Mature T-Cell and Natural Killer–Cell Neoplasms

2020 ◽  
Vol 154 (5) ◽  
pp. 683-691
Author(s):  
Min Shi ◽  
Phuong Nguyen ◽  
Michael M Timm ◽  
Gregory E Otteson ◽  
Pedro Horna ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cell ◽  
Natural Killer ◽  
Gene Rearrangement ◽  
Nk Cell ◽  
Molecular Genetic ◽  
Killer Cell ◽  
Cell Receptor ◽  
Molecular Study ◽  
Cytoplasmic Staining

Abstract Objectives Distinguishing between T-cell and natural killer (NK)–cell neoplasms could be difficult given their overlapping immunophenotype. In this study, we investigated whether a flow cytometry assay with cytoplasmic staining for CD3 could be used for this purpose. Methods Flow cytometry immunophenotyping was performed on 19 surface CD3 (sCD3)–negative mature T-cell neoplasms, 10 sCD3-positive mature T-cell neoplasms, 13 mature NK-cell neoplasms, and 19 normal controls. In addition to routine antibody panels (CD2, sCD3, CD4, CD5, CD7, CD8, CD16, CD45, CD56, CD57, CD94, CD158a, CD158b, CD158e, NKG2A TCRγ/δ), cytoplasmic staining for a monoclonal CD3 antibody (clone SK7/Leu-4) was assessed in all cases. A molecular study for T-cell receptor (TCR) gene rearrangement and an immunohistochemical study for TCRβ were performed. Results Our data showed all T-cell neoplasms were uniformly positive for cytoplasmic CD3 (cCD3) regardless of sCD3 expression, whereas 85% of NK-cell neoplasms completely lacked cCD3 expression. The 2 cases with classic NK-cell immunophenotype but partial cCD3 expression showed no molecular genetic features of T-cell lineage by TCR gene rearrangement studies. Conclusions Uniform cCD3 positivity and homogeneous cCD3 negativity highly suggest T-cell and NK lineage, respectively. When partial cCD3 expression is encountered, additional confirmatory studies should be pursued for the most accurate lineage assignment.

Natural Killer–Cell Lymphoma Involving the Gynecologic Tract

2000 ◽  
Vol 124 (10) ◽  
pp. 1510-1513 ◽  
Author(s):  
Paulette Mhawech ◽  
L. Jeffrey Medeiros ◽  
Carlos Bueso-Ramos ◽  
Donna M. Coffey ◽  
Alfredo F. Gei ◽  
...  
Keyword(s):  
T Cell ◽  
Natural Killer ◽  
T Cell Receptor ◽  
Nk Cell ◽  
Cell Lymphoma ◽  
Killer Cell ◽  
Cell Receptor ◽  
Lymphoid Cells ◽  
Gene Rearrangements ◽  
Neoplastic Cells

Abstract Non-Hodgkin lymphomas (NHL) can involve the gynecologic tract, most often as a manifestation of systemic involvement, and most cases reported have been of B-cell lineage. We describe 2 women with natural killer (NK)-cell lymphoma involving the gynecologic tract that initially presented with vaginal bleeding. In case 1, the patient had a stage IE nasal-type NK-cell lymphoma involving the cervix. The tumor was composed of medium-sized, irregular lymphoid cells with angioinvasion and necrosis. In case 2, the patient had a stage IV blastoid NK-cell lymphoma/leukemia infiltrating all organs in a hysterectomy and bilateral salpingo-oophorectomy specimen. Subsequent biopsy specimens revealed that the bone marrow and lymph nodes were also involved. The neoplasm was composed of small to medium lymphoid cells with fine nuclear chromatin. Case 1 was assessed immunohistochemically and the neoplastic cells were positive for CD3, CD56, and TIA-1. Case 2 was analyzed using both immunohistochemical and flow cytometry methods. The neoplastic cells were positive for cytoplasmic CD3, CD4, CD7, CD43, CD45, and CD56 and were negative for surface CD3. Both cases were negative for Epstein-Barr virus (EBV) ribonucleic acid (RNA) and molecular studies showed no evidence of T-cell receptor γ chain gene rearrangements. The immunophenotype and absence of T-cell receptor gene rearrangements support NK-cell origin. We report these cases to illustrate that NK-cell lymphomas can involve, and rarely arise in, the gynecologic tract.

scholarly journals Chronic natural killer cell lymphocytosis: a descriptive clinical study

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2721-2725 ◽  
Author(s):  
A Tefferi ◽  
CY Li ◽  
TE Witzig ◽  
MV Dhodapkar ◽  
SH Okuno ◽  
...  
Keyword(s):  
T Cell ◽  
Natural Killer ◽  
Disease Duration ◽  
Nk Cell ◽  
Receptor Gene ◽  
Killer Cell ◽  
Clinical Manifestations ◽  
Cell Receptor ◽  
Neutropenic Sepsis ◽  
Lgl Leukemia

Abstract We review the clinical manifestations and long-term outlook of patients with chronic natural killer (NK) cell lymphocytosis. After reviewing more than 1,500 peripheral blood lymphoid flow cytometry reports and molecular genetics data from patients with suspected large granular lymphocyte (LGL) proliferation, we identified 10 patients (median age at diagnosis, 60 years; range, 35 to 76 years; male:female ratio, 3:2) with persistent (greater than 6 months) increase in phenotypically determined NK cells (CD3-CD16+). Southern blot analysis performed on 9 patients showed no clonal T-cell receptor gene rearrangements. Disease duration was measured from time of initial recognition of LGL or NK cell excess (greater than 40% of the lymphocyte fraction). Clinical data from these 10 patients were compared with those from 68 patients with T-cell LGL (T-LGL) leukemia. Currently, all patients are alive (median disease duration, 5 years; range, 0.8 to 8 years). Associated disease manifestations included pure red blood cell aplasia, recurrent neutropenia, recurrent neutropenic sepsis, and vasculitic syndromes, all of which were responsive to immunosuppressive therapy. No patient had palpable lymphadenopathy or splenomegaly. Compared with the patients with T-LGL leukemia, patients with chronic NK cell leukemia has similar lymphocyte counts, associated conditions, treatment responses, and survival but had less neutropenia and anemia.

Lymphomatoid gastropathy: a distinct clinicopathologic entity of self-limited pseudomalignant NK-cell proliferation

Blood ◽  
2010 ◽  
Vol 116 (25) ◽  
pp. 5631-5637 ◽  
Author(s):  
Kengo Takeuchi ◽  
Masahiro Yokoyama ◽  
Shin Ishizawa ◽  
Yasuhito Terui ◽  
Kimie Nomura ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cell ◽  
Natural Killer ◽  
T Cell Receptor ◽  
Nk Cell ◽  
Killer Cell ◽  
Diagnostic Errors ◽  
Cell Receptor ◽  
Clinical Consequences ◽  
Epstein Barr

Abstract Diagnostic errors in distinguishing between malignant and reactive processes can cause serious clinical consequences. We report 10 cases of unrecognized self-limited natural killer–cell proliferation in the stomach, designated as lymphomatoid gastropathy (LyGa). This study included 5 men and 5 women (age, 46-75 years) without any gastric symptoms. Gastroscopy showed elevated lesion(s) (diameter, ∼ 1 cm). Histologically, medium-sized to large atypical cells diffusely infiltrated the lamina propria and, occasionally, the glandular epithelium. The cells were CD2+/−, sCD3−, cCD3+, CD4−, CD5−, CD7+, CD8−, CD16−, CD20−, CD45+, CD56+, CD117−, CD158a−, CD161−, T cell–restricted intracellular antigen-1+, granzyme B+, perforin+, Epstein-Barr early RNA−, T-cell receptor αβ−, and T-cell receptor γδ−. Analysis of the 16 specimens biopsied from 10 patients led to a diagnosis of lymphoma or suspected lymphoma in 11 specimens, gastritis for 1 specimen, adenocarcinoma for 1 specimen, and LyGa or suspected LyGa for 3 specimens. Most lesions underwent self-regression. Three cases relapsed, but none of the patients died. According to conventional histopathologic criteria, LyGa is probably diagnosed as lymphoma, especially as extranodal natural killer/T-cell lymphoma, nasal type. However, LyGa is recognized as a pseudomalignant process because of its clinical characteristics. The concept of LyGa should be well recognized.

scholarly journals Chronic natural killer cell lymphocytosis: a descriptive clinical study

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2721-2725 ◽  
Author(s):  
A Tefferi ◽  
CY Li ◽  
TE Witzig ◽  
MV Dhodapkar ◽  
SH Okuno ◽  
...  
Keyword(s):  
T Cell ◽  
Natural Killer ◽  
Disease Duration ◽  
Nk Cell ◽  
Receptor Gene ◽  
Killer Cell ◽  
Clinical Manifestations ◽  
Cell Receptor ◽  
Neutropenic Sepsis ◽  
Lgl Leukemia

We review the clinical manifestations and long-term outlook of patients with chronic natural killer (NK) cell lymphocytosis. After reviewing more than 1,500 peripheral blood lymphoid flow cytometry reports and molecular genetics data from patients with suspected large granular lymphocyte (LGL) proliferation, we identified 10 patients (median age at diagnosis, 60 years; range, 35 to 76 years; male:female ratio, 3:2) with persistent (greater than 6 months) increase in phenotypically determined NK cells (CD3-CD16+). Southern blot analysis performed on 9 patients showed no clonal T-cell receptor gene rearrangements. Disease duration was measured from time of initial recognition of LGL or NK cell excess (greater than 40% of the lymphocyte fraction). Clinical data from these 10 patients were compared with those from 68 patients with T-cell LGL (T-LGL) leukemia. Currently, all patients are alive (median disease duration, 5 years; range, 0.8 to 8 years). Associated disease manifestations included pure red blood cell aplasia, recurrent neutropenia, recurrent neutropenic sepsis, and vasculitic syndromes, all of which were responsive to immunosuppressive therapy. No patient had palpable lymphadenopathy or splenomegaly. Compared with the patients with T-LGL leukemia, patients with chronic NK cell leukemia has similar lymphocyte counts, associated conditions, treatment responses, and survival but had less neutropenia and anemia.

KLRL1, a novel killer cell lectinlike receptor, inhibits natural killer cell cytotoxicity

Blood ◽  
2004 ◽  
Vol 104 (9) ◽  
pp. 2858-2866 ◽  
Author(s):  
Yanmei Han ◽  
Minghui Zhang ◽  
Nan Li ◽  
Taoyong Chen ◽  
Yi Zhang ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Killer Cell ◽  
Cell Receptor ◽  
Sh2 Domain ◽  
Target Cells ◽  
Lymphoid Tissues ◽  
Nk Cell Receptor ◽  
Human And Mouse

Abstract Natural killer (NK) cell inhibitory receptors play important roles in the regulation of target susceptibility to natural killing. Here, we report the molecular cloning and functional characterization of a novel NK cell receptor, KLRL1, from human and mouse dendritic cells. KLRL1 is a type II transmembrane protein with an immunoreceptor tyrosine-based inhibitory motif and a C-type lectinlike domain. The KLRL1 gene is located in the central region of the NK gene complex in both humans and mice, on human chromosome 12p13 and mouse chromosome 6F3, adjacent to the other KLR genes. KLRL1 is preferentially expressed in lymphoid tissues and immune cells, including NK cells, T cells, dendritic cells, and monocytes or macrophages. Western blot and fluorescence confocal microscopy analyses indicated that KLRL1 is a membrane-associated glycoprotein, which forms a heterodimer with an as yet unidentified partner. Human and mouse KLRL1 are both predicted to contain putative immunoreceptor tyrosine-based inhibitory motifs (ITIMs), and immunoprecipitation experiments demonstrated that KLRL1 associates with the tyrosine phosphatases SHP-1 (SH2-domain-containing protein tyrosine phosphatase 1) and SHP-2. Consistent with its potential inhibitory function, pretreatment of target cells with human KLRL1-Fc fusion protein enhances NK-mediated cytotoxicity. Taken together, our results demonstrate that KLRL1 belongs to the KLR family and is a novel inhibitory NK cell receptor.

10 A multiparameter flow cytometry assay to monitor natural killer cell proliferation and activation in immuno-oncology clinical trials

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A10-A10
Author(s):  
Jennifer Tsau ◽  
Brittney Atzmiller ◽  
David Quinn ◽  
Tanya Mulvey ◽  
Sema Kurtulus ◽  
...  
Keyword(s):  
Clinical Trial ◽  
Flow Cytometry ◽  
Cell Proliferation ◽  
T Cell ◽  
Natural Killer ◽  
Nk Cell ◽  
Multiparameter Flow Cytometry ◽  
Parent Population ◽  
Flow Cytometry Assay ◽  
Cd Markers

BackgroundNatural Killer (NK) cells have garnered increasing interest as potential cellular therapies or as targets of biotherapeutic agents due to their ability to kill tumor cells in a non-antigen dependent manner. Hence, measurement of NK cell proliferation and/or activation following treatment can serve as a useful biomarker for assessing the efficacy of immunomodulatory therapies.MethodsWe developed a novel 13-parameter flow cytometry panel incorporating cell differentiation (CD) markers important for identification of NK cell subsets (CD56, CD16), their proliferation (Ki-67), activation (CD25, CD335, NKG2D) and inhibition (CD159a) status. Additionally, CD markers that identify other cellular subsets known to be amenable to cytokine modulation (e.g., CD3 and CD14) were included for concurrent monitoring of T cell proliferation and monocyte activation. Method validation focused on analytical sensitivity, specificity and precision as key criteria of assay performance using peripheral blood mononuclear cells (PBMCs) stimulated with NK cell-activating cytokines and resting PBMCs from healthy donors.ResultsThe assay design allowed for robust quantitation of NK cell, T cell and monocyte functionalities. Lower limit of quantification (LLOQ) of target biomarker population was determined to be 1.0% of the parent population, based upon an analysis of 110 key target populations that displayed a co-efficient of variation (CV) of ≤25% and their frequencies ranged from 0.1% to 97.8% of the parent population. Additionally, ≤25% CV was observed in precision assessments, confirming the repeatability and reproducibility of the assay. Clinical trial utility of the assay was verified on cryopreserved PBMCs from patients with a variety of solid tumor malignancies. In these patients, the assay could clearly identify proliferating and activated NK cells, as well as proliferating T cells and activated monocytes, thus demonstrating its suitability for clinical trial applications.ConclusionsWe developed and validated a novel multiparameter flow cytometry assay that allows for simultaneous measurement of proliferation, activation and inhibitory status of key immune cell subsets. Thus, this assay can help shed light on the mode of efficacy of novel therapeutic agents that modulate the immune system, aimed at treatment of cancer and autoimmune diseases.

Clinical Analysis Of 120 Cases Of Natural Killer/T Cell Lymphoma, Nasal Type By Single Hospital In The North-Western China

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5111-5111
Author(s):  
Rong Liang ◽  
Xie qun Chen ◽  
Zhe Wang ◽  
Bai qin Xian ◽  
qin-Guo guo Yan ◽  
...  
Keyword(s):  
T Cell ◽  
Natural Killer ◽  
T Cell Receptor ◽  
Gene Rearrangement ◽  
Cell Lymphoma ◽  
Cell Receptor ◽  
Primary Site ◽  
Individual Therapy ◽  
T Cell Lymphoma ◽  
Ebv Dna

Abstract Objectives To improve the understanding of extranodal natural killer(NK)/ T cell lymphoma (ENKTCL) with poor prognosis and provide experiential references for individual therapy via a retrospective analysis of the clinical and pathological features. Methods 120 NKTCL cases from April 2007 to Oct 2012 in single center of Northwestern China were retrospectively analyzed on their pathologic diagnosis and clinical manifestations. Pathological examinations were mainly depended on morphology, immunohistochemisty for immunophenotype and In situ hybridizationc for epstein - barr virus (EBV) small encoded RNA£¨EBER£©. Polymerase chain reaction (PCR) for the amount of EBV DNA in whole-blood and T-cell receptor (TCR) gene rearrangement were performed. Chemotherapy and or radiotherapy were the main treatments. Complete remission(CR), 2 year(2y) overall survival (OS) and progressive free survival (PFS) according to clinical characteristics were analyzed. Results The median age of 120 NKTCL cases was 43.19+13.7 years old. 98 primary nasal ENKTCL cases accounted for 81.4%, whose 2y OS and PFS were 88.6% and 69.1% respectively. 22 non-nasal ENKTCL accounted for 18.3%£€whose 2y OS and PFS were 58.8% and 45.5% respectively, which was statistically significant comparing with primary nasal ENKTCL cases. It was found that patients with primary intestinal or Ki67 greater than 80% were dead in first year. Patients with primary liver and intestinal had higher Ki67 than patients with primary nasal. Rate of 2y OS of 43 patients with Ki67 from 60% to 80% was 60% and PFS was 36% as compared to 86.3% of rate of OS and 57.5% for PFS of 30 patients with Ki67 from 30% to 50% and 100% of rate of OS and 78% for PFS of 7 patients with Ki67 less than 30%. Basing on analysis showed that from the cell of origin, it was found that primary site was nasal and 2y OS 100%¡¢PFS 75%£€in 8 cases with CD56+,CD3+ and T-cell receptor (TCR) gene rearrangement, which was statistically significant comparing with CD56- nasal ENKTCL cases. EBER was positive in every case. Among the 13 EBV –DNA samples detected, there were 5 samples with more than 6.1X107 copies/ml with OS 60%. 2y OS and PFS of Patients with normal ferritin ¦Â-microglobulin were longer than that of higher ferritin and ¦Â-microglobuli( P). 2y OS and PFS of 10 cases with I stage who had only radiotherapy was 100% respectively. 47 cases with II-IV stage received only chemotherapy. Their 2y OS and PFS was 100% and 90% respectively for patients with II stage better than that of III and IV stage. 63 cases with treatment of chemotherapy and radiotherapy. Among them, 2y OS and PFS of local nasal and invasion nasal were different. International prognostic index, IPI was good for prognosis for OSºÍPFS. Conclusion£º It was implied that Ki67, CD56, EBER, EBV-DNA and primary site was related with the prognosis of NKTCL. It needs further and more clinical observation and verification to judge if they could be additional markers for prognostication and clinical stratification to be incorporated in clinical individual management algorithms. The chemotherapies containing asparaginase or polyehylene glycol asparaginase are more effective. Autologous stem cell transplantation could make patients long live and pretransplant CR status was a major prognostic factors. Disclosures: No relevant conflicts of interest to declare.

scholarly journals IPH2101, a novel anti-inhibitory KIR antibody, and lenalidomide combine to enhance the natural killer cell versus multiple myeloma effect

Blood ◽  
2011 ◽  
Vol 118 (24) ◽  
pp. 6387-6391 ◽  
Author(s):  
Don M. Benson ◽  
Courtney E. Bakan ◽  
Shuhong Zhang ◽  
Shauna M. Collins ◽  
Jing Liang ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Natural Killer ◽  
Cell Function ◽  
Nk Cell ◽  
Killer Cell ◽  
Allogeneic Transplantation ◽  
Cell Receptor ◽  
Phase 2 Trial ◽  
Cell Versus

Abstract Multiple myeloma (MM) patients who receive killer cell Ig–like receptor (KIR) ligand–mismatched, T cell–depleted, allogeneic transplantation may have a reduced risk of relapse compared with patients who receive KIR ligand–matched grafts, suggesting the importance of this signaling axis in the natural killer (NK) cell-versus-MM effect. Expanding on this concept, IPH2101 (1-7F9), an anti-inhibitory KIR mAb, enhances NK-cell function against autologous MM cells by blocking the engagement of inhibitory KIR with cognate ligands, promoting immune complex formation and NK-cell cytotoxicity specifically against MM cell targets but not normal cells. IPH2101 prevents negative regulatory signals by inhibitory KIR, whereas lenalidomide augments NK-cell function and also appears to up-regulate ligands for activating NK-cell receptors on MM cells. Lenalidomide and a murine anti-inhibitory NK-cell receptor Ab mediate in vivo rejection of a lenalidomide-resistant tumor. These mechanistic, preclinical data support the use of a combination of IPH2101 and lenalidomide in a phase 2 trial for MM.

scholarly journals Human Herpesviridae Methods of Natural Killer Cell Evasion

2012 ◽  
Vol 2012 ◽  
pp. 1-10 ◽  
Author(s):  
Carl I. Odom ◽  
David C. Gaston ◽  
James M. Markert ◽  
Kevin A. Cassady
Keyword(s):  
Natural Killer ◽  
Immune Evasion ◽  
Nk Cell ◽  
Killer Cell ◽  
Cell Receptor ◽  
Hematologic Malignancies ◽  
Innate Immune ◽  
Considerable Morbidity ◽  
Amount Of Knowledge ◽  
Innate Immune Evasion

Human herpesviruses cause diseases of considerable morbidity and mortality, ranging from encephalitis to hematologic malignancies. As evidence emerges about the role of innate immunity and natural killer (NK) cells in the control of herpesvirus infection, evidence of viral methods of innate immune evasion grows as well. These methods include interference with the ligands on infected cell surfaces that bind NK cell activating or inhibitory receptors. This paper summarizes the most extensively studied NK cell receptor/ligand pairs and then describes the methods of NK cell evasion used by all eight herpesviruses through these receptors and ligands. Although great strides have been made in elucidating their mechanisms, there is still a disparity between viruses in the amount of knowledge regarding innate immune evasion. Further research of herpesvirus innate immune evasion can provide insight for circumventing viral mechanisms in future therapies.

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