Granular Lymphocyte Proliferation Research Articles

Large Natural Killer Cell Lymphoma Arising From an Indolent Natural Killer Cell Large Granular Lymphocyte Proliferation

Natural killer cell large granular lymphocyte proliferation is a relatively rare disorder that typically runs a chronic, indolent course. We present a patient with a 3 1/2-year history of natural killer cell large granular lymphocyte proliferation terminating in large cell lymphoma with natural killer cell features. The diagnosis of natural killer cell large granular lymphocyte proliferation was based on flow cytometric demonstration of an expanded population of CD3- CD16+/CD56+ lymphocytes in the peripheral blood. The patient experienced various rheumatologic symptoms, but was hematologically stable for 3 1/2 years. He then developed fevers, night sweats, weight loss, and a left lower lobe lung mass. Resection of the mass showed a large cell lymphoma with immunohistochemical positivity for CD2, CD7, CD56, and T-cell intracellular antigen-1, compatible with natural killer cell origin. In situ hybridization for Epstein-Barr virus and polymerase chain reaction analysis for T-cell receptor gene rearrangement were negative. To our knowledge, this is the second documented report of chronic natural killer cell large granular lymphocyte proliferation terminating in an aggressive large natural killer cell lymphoma.

Clonal T- and natural killer-cell large granular lymphocyte proliferations in a single patient established by array-based comparative genomic hybridization analysis

Clonal T- and natural killer-cell large granular lymphocyte proliferations in a single patient established by array-based comparative genomic hybridization analysis

T-cell large granular lymphocytic (T-LGL) leukemia: Experience in a single institution over 8 years

T-cell large granular lymphocytic (T-LGL) leukemia is characterized by cytopenia and clonal proliferation of large granular lymphocytes. We identified 26 patients with T-LGL leukemia seen at our institution over a period of 8 years. The majority of the patients were asymptomatic at diagnosis. Nine patients were treated with cyclosporine; one achieved a complete remission, and four had a hematological response. Other treatment modalities included single agent alemtuzumab, alemtuzumab combined with pentostatin, fludarabine, and combination of fludarabine and cyclophosphamide. Significant responses were not seen with any of these treatment regimens. We conclude that cyclosporine therapy may be beneficial for T-LGL leukemia patients. New treatment modalities are needed for these patients.

Phenotypic and functional analyses of dendritic cells in patients with lymphoproliferative disease of granular lymphocytes (LDGL)

We investigated whether dendritic cells (DCs) play a role in favoring granular lymphocyte (GL) proliferation in patients with lymphoproliferative disease of granular lymphocytes (LDGL). The presence of in vivo circulating DCs was studied in 11 patients (5 CD3+ and 6 CD3- LDGL). Autologous immature (iDCs) and mature (mDCs) DCs generated in vitro were studied for stimulatory activity on cell proliferation of CD3+ and CD3- GLs. The topographic organization of GLs and DCs was also studied in bone marrow (BM) biopsies. Peripheral blood (PB) CD3- GLs from patients showed significant proliferative activity in the presence of iDCs and mDCs. Conversely, monoclonal CD3+ GLs were unresponsive to autologous and allogeneic PB DCs. Analysis of BM biopsies demonstrated a topographic distribution of DCs and GLs that indicates contact between the 2 cell types. On functional assays, DCs obtained from BM were more efficient than PB DCs in stimulating CD3- GLs, and surprisingly, a low but definite stimulatory effect was demonstrated also on CD3+ GLs. The putative contact between DCs and GLs in the BM and, more crucial, the proliferative response of discrete GL populations to DC stimulation suggest the presence of a specific antigen within BM DCs, providing evidence for a role of DCs in the pathogenesis of LDGL.

Clinical and Pathological Presentation of T-Cell Large Granular Lymphocyte Proliferations (T-LGL): A Single Institution Experience.

Proliferations of T-cell large granular lymphocytes (T-LGL) are a rare and heterogeneous disease entity characterized by a chronic course, cytopenias and circulating cytotoxic T lymphocytes. We present the results of a single institution retrospective review of the clinical presentation of T-LGL proliferations. A total of 23 patients evaluated for cytopenias were identified as having T-LGL by flow cytometry, ten male and thirteen female, with a median age of 67 years at diagnosis. Twelve patients had a prior malignancy (non-Hodgkins lymphoma=4, MDS/AML=4, breast cancer=2, squamous cell cancer (SCC) of the skin =2, lung SCC=1, vocal cord SCC=1, uterine cancer=1) and five patients had a history of autoimmune disease (rheumatoid arthritis=2, vasculitis=1, polyarthritis=1, Wegener's granulomatosis=1), while seven patients had no previous history of autoimmune disease or malignancy. Four patients with prior malignancy had active disease and this led to death in three. Of the twelve patients informative for T-cell receptor gene rearrangements, clonal populations were found in eight. All patients who had cytopenias requiring treatment were initially treated with cyclosporine monotherapy, with the further addition of growth factors if necessary.Multiparameter flow cytometric analysis of blood or bone marrow aspirates revealed that patients with a prior history of systemic malignancy had the common immunophenotype of CD3+(h), CD8+(h), CD2+, CD5+, CD7(h), CD56(h), CD57+, CD38+, CD16−. In contrast, patients who did not have a prior history of malignancy were CD3+, CD8+, CD2+, CD5+, CD7+, CD56+/−, CD57+, CD38+/(h), CD16+/− (h=heterogeneous). Thus patients with a prior malignancy had more heterogeneous expression of the T-cell markers CD3, CD8 and CD7 and were less likely to express CD16. We conclude that prior malignancy is more common in patients with T-LGL proliferations than previously recognized, may be twice as frequent as autoimmune disease in association with T-LGL and may have a distinct immunophenotype. Our data suggest that malignant cells may induce proliferations of T-LGL.

Natural killer receptors in patients with lymphoproliferative diseases of granular lymphocytes

The most relevant criterion for recognizing chronic granular lymphocyte (GL) proliferations, defined as lymphoproliferative disease of GL (LDGL), has been historically based on the number of the proliferating cells displaying typical GL morphology. With the extensive development of immunological and molecular techniques, two major groups of LDGL have been recognized, one belonging to the T-cell and the other to the NK cell lineage. The recent definition of a series of receptors of NK cells (NKR) and the identification of the specific targets recognized has expanded our knowledge of the properties of these cells and the discrimination between functional reactive and pathological proliferations. Some of these receptors are expressed by GL of T-cell lineage, suggesting a possible involvement in the genesis of GL proliferation. Following an extensive description of NKR in humans, this review will summarize the recent data on phenotypic and functional characteristics of NKR expressed by proliferating GL in patients with LDGL, discussing their role in this disorder.

Clinical features of large granular lymphocyte leukemia

The spectrum of large granular lymphocyte (LGL) proliferations consists of four distinct entities: reactive/transient LGL expansion, chronic LGL lymphocytosis, classical indolent LGL leukemia, and aggressive LGL leukemia. LGL leukemias are classified as lymphoid malignancies. They are divided into CD3+/T-cell LGL (85% of cases) and CD3−/natural killer (NK) cell LGL leukemia (15% of cases). Recent progress in the comprehension of the leukemogenesis has shown a dysregulation of survival signals in leukemic cells. Identification of LGL expansion has been improved using T-cell receptor (TCR)β/γ polymerase chain reaction (PCR) analysis and a combination of Vβ and killer cell immunoglobulin-like receptor (KIR)-specific monoclonal antibodies. LGL leukemias are characterized by a clonal LGL infiltration of the bone marrow, spleen, and liver. Monoclonality is recognized by phenotypic, molecular, and karyotypic analysis. T-LGL leukemias affect the elderly and display a relatively indolent behavior. Approximately 60% to 70% of patients are symptomatic: recurrent infections secondary to chronic neutropenia, anemia, and autoimmune disease such as rheumatoid arthritis are the main clinical manifestations. Long-lasting remission can be obtained with low-dose methotrexate, cyclosporine A, or cyclophosphamide. Conversely, NK LGL leukemias behave aggressively, and most patients do not respond to chemotherapy.

Cytomegalovirus infection associated with clonal proliferation of T-cell large granular lymphocytes: causal or casual?

Clonal proliferation of T-cell large granular lymphocytes (LGL) is an indolent disorder characterized by splenomegaly, lymphocytosis and frequent manifestations of immune disturbances. The LGL are CD3 + CD4 − CD8 + CD56 −. The clonality of the tumor cell population is often only demonstrable by T-cell receptor ( TCR) gene rearrangement study because chromosomal abnormality is distinctly rare. We describe a case of T-cell LGL leukemia that presented initially as cytomegalovirus infection. The leukemic LGL are shown to be clonal by both TCR gene rearrangement and chromosomal studies. They persist after subsidence of the cytomegalovirus infection.

Neutropenia in rheumatoid arthritis and large granular lymphocyte leucosis

Objective. Pts with chronic clonal proliferation of large granular lymphocytes (LGL leukemia) often have neutropenia, splenomegaly, and rheumatoid arthritis (RA), thereby resembling the manifestations observed in pts with Felty’s syndrome. The present study sought to indicate that pts with these disorders represent two distinct subsets. We compare clinical, hematological, immunophenotiping and immunogenetic features in Felty’s syndrome pts with and without the LGL leukemia. Material and methods 10 pts with T-LGL leukemia were studied. Surface phenotype was estimated using monoclonal antibodies CD8-PE and CD3-FITC/CD16-PE (two-color) (Caltag, USA) by the flow cytometric analysis (Partec, Daco). Analysis of TCR gene rearrangement was performed by using PCR-LIS SSCP (low ionic strength single strand conformational polymorphism). Comparison with Felty s syndrome and RA pts based on the review of literature. Results. LGL leukemia is a distinct clinicopathologic entity often associated with RA. LGL leukemia pts with RA showed the same immunogenetis associations seen in RA/Felty’s syndrome, while LGL leukemia pts without arthritis did not. Conclusion. Hematologic, immunophenotyping and molecular genetic analysis are very important and highly representative tools in differential diagnosis of neutropenia in RA, and propose that Felty’s syndrome and LGL leukemia represent different variants of broader syndrome comprising RA, neutropenia, LGL expansions, and splenomegaly.

Features of large granular lymphocytes (LGL) expansion following allogeneic stem cell transplantation: a long-term analysis.

Large granular lymphocyte (LGL) proliferation typically follows a chronic course during which major features are cytopenia and immune abnormalities. Elevated numbers of LGL were reported in a few cases following allogeneic stem cell transplantation (allo-SCT). In this report, we present a retrospective analysis of LGL cases that occurred following allo-SCT in a cohort of 201 consecutive patients transplanted over a period of 7 years. Six cases were identified and LGL expansion occurred more frequently following a reduced fludarabine and anti-T lymphocyte globulin-based preparative regimen (4 cases/49), than after a conventional myeloablative regimen (2 cases/152). Expansion of LGL was seen between 3 and 15 months following allo-SCT. Hematopoiesis, with mild to severe cytopenia, was a favored target for LGL. Autoimmune manifestations including polyarthritis and hypergammaglobulinemia were also observed. LGL proliferation was observed in the context of chronic antigenic stimulation associated with recurrent viral infections especially CMV. Moreover, five out of these six high risk patients achieved a long-term complete remission concomitant or following LGL expansion. These data suggest that LGL might be a subset of effector lymphocytes which may participate to the graft-versus-tumor effect.

A unique case of splenic marginal zone-cell lymphoma with synchronous clonal T-cell large granular lymphocyte proliferation: an immunologic, immunohistochemical and genotypic study

We describe a case of synchronous splenic marginal zone-cell lymphoma (SMZL) and T-cell large granular lymphocyte leukemia involving the spleen, liver, bone marrow and peripheral blood. The synchronous occurrence of these two processes was documented by morphological, immunophenotypical and molecular (PCR) analyses of all affected tissues. The pathogenetic mechanisms which may be responsible for the concomitant appearance of these two rather infrequent entities in the same anatomic sites are discussed.

DNA microarray analysis of T cell-type lymphoproliferative disease of granular lymphocytes.

Lymphoproliferative disease of granular lymphocytes (LDGL) is characterized by the clonal proliferation of large granular lymphocytes of either T- or natural killer cell origin. To better understand the nature of T cell-type LDGL, we purified the CD4-CD8+ proliferative fractions from LDGL patients (n=4) and the surface marker-matched T cells isolated from healthy volunteers (n=4), and compared the expression profiles of 3456 genes using DNA microarray. Through this analysis, we identified a total of six genes whose expression was active in the LDGL T cells, but silent in the normal ones. Interestingly, expression of the gene for interleukin (IL) 1beta was specific to LDGL T cells, which was further confirmed by the examination of the serum level of IL-1beta protein. Given its important role in inflammatory reactions, the disease-specific expression of IL-1beta may have a causative relationship with the LDGL-associated rheumatoid arthritis. Spectratyping analysis of the T-cell receptor repertoire also proved the monoclonal or oligoclonal nature of LDGL cells. These data have shown that microarray analysis with a purified T-cell subset is an efficient approach to investigate the pathological condition of Tcell-type LDGL.

Analysis of TNF-receptor and ligand superfamily molecules in patients with lymphoproliferative disease of granular lymphocytes

In 21 patients with lymphoproliferative disease of granular lymphocytes (LDGL), we investigated the expression and the function of molecules belonging to TNF-receptor and TNF-ligand superfamilies (CD30/CD30L; CD40/CD40L; CD27/CD70; Fas [CD95]/FasL[CD95L]). Fourteen patients were characterized by a proliferation of granular lymphocytes (GLs) expressing the CD3(+)CD16(+) phenotype, whereas 7 cases showed the CD3(-)CD16(+) CD56 +/- phenotype. Our data show that both CD3(+) and CD3-GLs are preferentially equipped with CD30, CD40, CD40L, CD70, and CD95 antigens; this pattern is usually associated with the lack of CD27 and CD30L antigens expression. CD95L was demonstrated in the cytoplasm in 14 of 21 cases by flow cytometry, but a definite signal was demonstrated in all cases studied using polymerase chain reaction analysis. On functional grounds, a stimulatory activity on rIL-2 mediated redirected-cytotoxicity against Fcgamma+ P815 targets was demonstrated with anti-CD30, CD40, CD40L, CD70, CD95, and CD95L mAbs, although resting cells were unable to exhibit significant redirected-cell lysis. The addition of anti-CD30, CD30L, CD40, CD40L, CD95, and CD95L mAbs did not show any significant effect on cell proliferation at resting conditions or after rIL-2 stimulation, whereas anti-CD70 mAb mediated cell proliferation in 6 of 10 cases tested. This figure was not related to an increase in apoptotic cells, as investigated by Annexin-V expression. Our data indicate that both CD3(+) and CD3(-) GLs are equipped with different costimulatory antigens, supporting the concept that these cells are in vivo activated and suggesting that these molecules might play a role in the cytotoxic mechanisms of GLs. (Blood. 2000;96:647-654)

Analysis of TNF-receptor and ligand superfamily molecules in patients with lymphoproliferative disease of granular lymphocytes: Presented in part in abstract form at the 87th Meeting of the American Association for Cancer Research, San Diego, CA, April 1997 (abstract no. 3698).

AbstractIn 21 patients with lymphoproliferative disease of granular lymphocytes (LDGL), we investigated the expression and the function of molecules belonging to TNF-receptor and TNF-ligand superfamilies (CD30/CD30L; CD40/CD40L; CD27/CD70; Fas [CD95]/FasL[CD95L]). Fourteen patients were characterized by a proliferation of granular lymphocytes (GLs) expressing the CD3+CD16+phenotype, whereas 7 cases showed the CD3−CD16+ CD56 ± phenotype. Our data show that both CD3+ and CD3-GLs are preferentially equipped with CD30, CD40, CD40L, CD70, and CD95 antigens; this pattern is usually associated with the lack of CD27 and CD30L antigens expression. CD95L was demonstrated in the cytoplasm in 14 of 21 cases by flow cytometry, but a definite signal was demonstrated in all cases studied using polymerase chain reaction analysis. On functional grounds, a stimulatory activity on rIL-2 mediated redirected-cytotoxicity against Fcγ+ P815 targets was demonstrated with anti-CD30, CD40, CD40L, CD70, CD95, and CD95L mAbs, although resting cells were unable to exhibit significant redirected-cell lysis. The addition of anti-CD30, CD30L, CD40, CD40L, CD95, and CD95L mAbs did not show any significant effect on cell proliferation at resting conditions or after rIL-2 stimulation, whereas anti-CD70 mAb mediated cell proliferation in 6 of 10 cases tested. This figure was not related to an increase in apoptotic cells, as investigated by Annexin-V expression. Our data indicate that both CD3+ and CD3− GLs are equipped with different costimulatory antigens, supporting the concept that these cells are in vivo activated and suggesting that these molecules might play a role in the cytotoxic mechanisms of GLs.

Immunogenetic similarities between patients with Felty's syndrome and those with clonal expansions of large granular lymphocytes in rheumatoid arthritis.

Patients with chronic clonal proliferation of large granular lymphocytes (LGL leukemia) often have splenomegaly, neutropenia, and rheumatoid arthritis (RA), thereby resembling the manifestations observed in patients with Felty's syndrome. The present study sought to determine whether patients with these disorders represent 2 distinct subsets of neutropenic RA. Prospective cohort study of outpatients attending clinics in university and private hospitals and in offices of private practice physicians. Twenty-two patients with Felty's syndrome and 22 patients with LGL leukemia, 10 of whom had RA, were studied. HLA genotyping was performed on peripheral blood mononuclear leukocyte genomic DNA. Nineteen of the 22 patients with Felty's syndrome (86%) were DR4 positive. Nine of the 10 patients with LGL leukemia plus RA were also DR4 positive. In contrast, only 4 of the 12 patients with LGL leukemia without RA (33%) were DR4 positive, a frequency that was within the normal range. The finding of an equally high prevalence of DR4 in patients with Felty's syndrome and in those with LGL leukemia plus RA suggests that both disorders have a similar immunogenetic basis and are parts of a single disease process rather than 2 separate disorders.

Routine diagnosis of large granular lymphocytic leukaemia by Southern blot and polymerase chain reaction analysis of clonal T cell receptor gene rearrangement.

AIMS: To compare the polymerase chain reaction (PCR) assay with standard Southern blot (SB) hybridisation for the detection of clonal T cell receptor (TCR) gene rearrangements in large granular lymphocyte...

Interleukin-15 Triggers the Proliferation and Cytotoxicity of Granular Lymphocytes in Patients With Lymphoproliferative Disease of Granular Lymphocytes

The recently cloned cytokine interleukin-15 (IL-15) shares several functional activities with IL-2 in different cell systems. Although IL-15 does not show sequence homology with IL-2, it uses components of the IL-2 receptor (IL-2R) for binding and signal transduction, namely, p75 (β) and the p64 (γ) chains of IL-2R. To evaluate whether IL-15 is involved in the activation of granular lymphocytes (GL) in patients with lymphoproliferative disease of granular lymphocytes (LDGL), we evaluated the ability of IL-15 to stimulate GL proliferation, cytotoxic function, and the role of IL-2R β and γ molecules on relevant cells. Our results show that IL-15 stimulates cell proliferation and cytotoxic activity of GL in LDGL patients. Reverse-transcriptase polymerase chain reaction (RT-PCR) and phenotypic analyses using the anti–IL-2R γ-chain–specific TUGh4 monoclonal antibody (MoAb) indicate that both CD3+ and CD3− GL express the p64 IL-2R, a result previously unknown. IL-15 activity was inhibited by antibodies against p75 and p64 IL-2R chains, while no inhibitory effects are detectable with anti-p55 IL-2R antibody. The association of anti-p75 and anti-p64 IL-2R MoAbs resulted in a nearly complete (95%) inhibition of IL-15–induced GL proliferation. Using RT-PCR analysis, we demonstrated that highly purified CD3+ and CD3− GL did not express mRNA for IL-15 or IL-2. By contrast, a clear-cut IL-15 mRNA signal was detected by RT-PCR in patients' peripheral blood mononuclear cells, with monocytes likely accounting for the source of IL-15 in LDGL patients. However, even in concentrated supernatants from enriched monocyte populations, we could not demonstrate the presence of IL-15 protein. Using anti–IL-15 specific MoAbs, a membrane-bound form of this cytokine was demonstrated both on CD3+ and CD3− LDGL cells. By RT-PCR analysis, purified GL from these patients were found to express the message for IL-15 receptor α chain. Taken together, these results indicate that both CD3+ and CD3− GL are stimulated by IL-15 and that this cytokine mediates its activity through the β and γ chains of the IL-2R, providing further suggestions for the interpretation of the mechanisms that lead to cell expansion in patients with LDGL.

Interleukin-15 Triggers the Proliferation and Cytotoxicity of Granular Lymphocytes in Patients With Lymphoproliferative Disease of Granular Lymphocytes

AbstractThe recently cloned cytokine interleukin-15 (IL-15) shares several functional activities with IL-2 in different cell systems. Although IL-15 does not show sequence homology with IL-2, it uses components of the IL-2 receptor (IL-2R) for binding and signal transduction, namely, p75 (β) and the p64 (γ) chains of IL-2R. To evaluate whether IL-15 is involved in the activation of granular lymphocytes (GL) in patients with lymphoproliferative disease of granular lymphocytes (LDGL), we evaluated the ability of IL-15 to stimulate GL proliferation, cytotoxic function, and the role of IL-2R β and γ molecules on relevant cells. Our results show that IL-15 stimulates cell proliferation and cytotoxic activity of GL in LDGL patients. Reverse-transcriptase polymerase chain reaction (RT-PCR) and phenotypic analyses using the anti–IL-2R γ-chain–specific TUGh4 monoclonal antibody (MoAb) indicate that both CD3+ and CD3− GL express the p64 IL-2R, a result previously unknown. IL-15 activity was inhibited by antibodies against p75 and p64 IL-2R chains, while no inhibitory effects are detectable with anti-p55 IL-2R antibody. The association of anti-p75 and anti-p64 IL-2R MoAbs resulted in a nearly complete (95%) inhibition of IL-15–induced GL proliferation. Using RT-PCR analysis, we demonstrated that highly purified CD3+ and CD3− GL did not express mRNA for IL-15 or IL-2. By contrast, a clear-cut IL-15 mRNA signal was detected by RT-PCR in patients' peripheral blood mononuclear cells, with monocytes likely accounting for the source of IL-15 in LDGL patients. However, even in concentrated supernatants from enriched monocyte populations, we could not demonstrate the presence of IL-15 protein. Using anti–IL-15 specific MoAbs, a membrane-bound form of this cytokine was demonstrated both on CD3+ and CD3− LDGL cells. By RT-PCR analysis, purified GL from these patients were found to express the message for IL-15 receptor α chain. Taken together, these results indicate that both CD3+ and CD3− GL are stimulated by IL-15 and that this cytokine mediates its activity through the β and γ chains of the IL-2R, providing further suggestions for the interpretation of the mechanisms that lead to cell expansion in patients with LDGL.

The clinical relevance of fluctuations in absolute-lymphocyte counts during follow-up of large granular lymphocyte proliferations [letter

The clinical relevance of fluctuations in absolute-lymphocyte counts during follow-up of large granular lymphocyte proliferations [letter

Natural killer cell proliferation and renal disease: A functional and phenotypic study

We describe a 57-year-old woman who presented with a constitutional syndrome, glomerulonephritis, and lymphocytosis. The phenotypic study, using flow cytometry, showed an expansion of natural killer (NK) cells (CD2+, CD3−, CD16+, CD56+, and CD7+). We performed a functional study of peripheral blood mononuclear cells (PBMCs) and of purified CD16+ cells (NK cells) and CD3+ cells (normal T cells). The expanded NK cell population, CD16+, did not proliferate with phytohemagglutinin (PHA) or anti-CD3 but showed a dose-dependent proliferation with recombinant interleukin-2 (rlL-2) and also proliferated with phorbol dibutyrate. This population showed very strong NK and lymphokine-activated killer cell (LAK) activities. The patient's symptoms resolved spontaneously without treatment. Three years later, however, there is still abnormal renal function, and the expansion of NK cells persists, although with no indication of malignancy. We review the features of the different large granular lymphocyte proliferations and their seldom described relationship with renal disease. © 1996 Wiley-Liss, Inc.