Association between clonogenic cell growth and clinical risk group in B- cell chronic lymphocytic leukemia

Abstract Chronic lymphocytic leukemia of B-cell origin (B-CLL) is a disease with a variable clinical course, despite the fact that the neoplastic cells in this disorder are homogeneous with respect to morphology, immunophenotype, and cell cycle stage. To further investigate the heterogeneity observed in the clinical behavior of B-CLL, we determined the phenotype and growth requirements of clonogenic cells from 28 patients with B-CLL from low-, intermediate-, and high-risk groups as defined by the Rai staging system. Using methyl-cellulose as a semi- solid media with feeder cells and/or growth factors, colonies were observed with one or more of the culture conditions tested in 25 of 28 CLLs. Phenotypic analysis of colonies demonstrated that the clonogenic cells uniformly expressed la, CD19, CD20, CD5, and the identical light chain as the original CLL cell cultured. However, heterogeneity was observed in clonogenic B-CLL cell growth among the three different CLL risk groups. Clonogenic cells from patients with low-risk CLL required either irradiated unstimulated T cells, with or without conditioned media (CM) or irradiated activated T cells alone for colony formation. Both the number of colonies (227 +/- 15) as well as the number of cells per colony (220 +/- 82) were large, with a mean cloning efficiency of 0.39%. In contrast, clonogenic cells from patients with intermediate- and high-risk CLL required the combination of both irradiated activated T cells and CM. As compared with the low-risk CLLs, both the number and size of the colonies formed by the intermediate- (74 +/- 17, 70 +/- 39) and high- (83 +/- 28, 40 +/- 14) risk groups were significantly lower (P less than .0001). Similarly, the mean cloning efficiency was significantly reduced to 0.15% and 0.14%, respectively. None of the recombinant cytokines (interleukin 1 [IL-1] to IL-7, tumor necrosis factor, alpha and gamma-interferon, B-cell growth factor, and granulocyte macrophage colony-stimulating factor) alone or in combination with each other could entirely replace the stimulatory effect of the activated T cells. These data suggest that clinical progression of B-CLL is associated with a loss of clonogenic potential in the circulating pool of neoplastic cells, which require as yet undefined factors provided by activated T cells and CM.

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Association between clonogenic cell growth and clinical risk group in B- cell chronic lymphocytic leukemia

Blood ◽

10.1182/blood.v76.1.142.bloodjournal761142 ◽

1990 ◽

Vol 76 (1) ◽

pp. 142-149

Author(s):

R Dadmarz ◽

SN Rabinowe ◽

SA Cannistra ◽

JW Andersen ◽

AS Freedman ◽

...

Keyword(s):

T Cells ◽

High Risk ◽

Cell Growth ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Risk Groups ◽

Lymphocytic Leukemia ◽

Cloning Efficiency ◽

Activated T Cells ◽

Clonogenic Cells

Chronic lymphocytic leukemia of B-cell origin (B-CLL) is a disease with a variable clinical course, despite the fact that the neoplastic cells in this disorder are homogeneous with respect to morphology, immunophenotype, and cell cycle stage. To further investigate the heterogeneity observed in the clinical behavior of B-CLL, we determined the phenotype and growth requirements of clonogenic cells from 28 patients with B-CLL from low-, intermediate-, and high-risk groups as defined by the Rai staging system. Using methyl-cellulose as a semi- solid media with feeder cells and/or growth factors, colonies were observed with one or more of the culture conditions tested in 25 of 28 CLLs. Phenotypic analysis of colonies demonstrated that the clonogenic cells uniformly expressed la, CD19, CD20, CD5, and the identical light chain as the original CLL cell cultured. However, heterogeneity was observed in clonogenic B-CLL cell growth among the three different CLL risk groups. Clonogenic cells from patients with low-risk CLL required either irradiated unstimulated T cells, with or without conditioned media (CM) or irradiated activated T cells alone for colony formation. Both the number of colonies (227 +/- 15) as well as the number of cells per colony (220 +/- 82) were large, with a mean cloning efficiency of 0.39%. In contrast, clonogenic cells from patients with intermediate- and high-risk CLL required the combination of both irradiated activated T cells and CM. As compared with the low-risk CLLs, both the number and size of the colonies formed by the intermediate- (74 +/- 17, 70 +/- 39) and high- (83 +/- 28, 40 +/- 14) risk groups were significantly lower (P less than .0001). Similarly, the mean cloning efficiency was significantly reduced to 0.15% and 0.14%, respectively. None of the recombinant cytokines (interleukin 1 [IL-1] to IL-7, tumor necrosis factor, alpha and gamma-interferon, B-cell growth factor, and granulocyte macrophage colony-stimulating factor) alone or in combination with each other could entirely replace the stimulatory effect of the activated T cells. These data suggest that clinical progression of B-CLL is associated with a loss of clonogenic potential in the circulating pool of neoplastic cells, which require as yet undefined factors provided by activated T cells and CM.

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Defective interleukin-2 production and responsiveness by T cells in patients with chronic lymphocytic leukemia of B cell variety

Blood ◽

10.1182/blood.v67.2.279.279 ◽

1986 ◽

Vol 67 (2) ◽

pp. 279-284 ◽

Cited By ~ 21

Author(s):

O Ayanlar-Batuman ◽

E Ebert ◽

SP Hauptman

Keyword(s):

Cell Proliferation ◽

T Cells ◽

T Cell ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Proliferative Response ◽

Interleukin 2 ◽

Lymphocytic Leukemia ◽

T Cell Proliferation ◽

Activated T Cells

Abstract The present studies were designed to investigate the mechanism(s) of the defective T cell proliferative response to various stimuli in patients with B cell chronic lymphocytic leukemia B-CLL. In 14 patients with advanced B-CLL (stage III or IV) we found the T cell response in the autologous (auto) and allogeneic (allo) mixed lymphocyte reaction (MLR) to be 35.7% and 30% of the controls, respectively. Proliferation in the MLR depends upon the production of and response to interleukin 2 (IL 2), a T cell growth factor. IL 2 production in eight B-CLL patients was 22% of the control. The response to IL 2 was measured by the increase in the T cell proliferation in the MLR with the addition of IL 2. T cell proliferation in both the auto and allo MLR of CLL patients was significantly lower than in the controls after the addition of IL 2. The proliferative response of normal T cells to stimulation by CLL B cells was 50% of the control. This latter response was increased to control levels when cultures were supplemented with exogenous IL 2, suggesting that CLL B cells could stimulate IL 2 receptor generation in normal T cells in an allo MLR, but not IL 2 production. The presence of IL 2 receptors on activated T cells was directly determined using anti- Tac, a monoclonal antibody with specificity for the IL 2 receptor. Of the mitogen- or MLR-activated T cells in CLL patients, 6% and 10%, respectively, expressed Tac antigen, whereas identically stimulated control T cells were 60% and 47% Tac+, respectively. Our findings suggest that T cells in B-CLL are defective in their recognition of self or foreign major histocompatibility antigens as demonstrated by their impaired responsiveness in the MLR. Thus, these cells are unable to produce IL 2 or generate IL 2 receptors.

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Defective interleukin-2 production and responsiveness by T cells in patients with chronic lymphocytic leukemia of B cell variety

Blood ◽

10.1182/blood.v67.2.279.bloodjournal672279 ◽

1986 ◽

Vol 67 (2) ◽

pp. 279-284 ◽

Cited By ~ 1

Author(s):

O Ayanlar-Batuman ◽

E Ebert ◽

SP Hauptman

Keyword(s):

Cell Proliferation ◽

T Cells ◽

T Cell ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Proliferative Response ◽

Interleukin 2 ◽

Lymphocytic Leukemia ◽

T Cell Proliferation ◽

Activated T Cells

The present studies were designed to investigate the mechanism(s) of the defective T cell proliferative response to various stimuli in patients with B cell chronic lymphocytic leukemia B-CLL. In 14 patients with advanced B-CLL (stage III or IV) we found the T cell response in the autologous (auto) and allogeneic (allo) mixed lymphocyte reaction (MLR) to be 35.7% and 30% of the controls, respectively. Proliferation in the MLR depends upon the production of and response to interleukin 2 (IL 2), a T cell growth factor. IL 2 production in eight B-CLL patients was 22% of the control. The response to IL 2 was measured by the increase in the T cell proliferation in the MLR with the addition of IL 2. T cell proliferation in both the auto and allo MLR of CLL patients was significantly lower than in the controls after the addition of IL 2. The proliferative response of normal T cells to stimulation by CLL B cells was 50% of the control. This latter response was increased to control levels when cultures were supplemented with exogenous IL 2, suggesting that CLL B cells could stimulate IL 2 receptor generation in normal T cells in an allo MLR, but not IL 2 production. The presence of IL 2 receptors on activated T cells was directly determined using anti- Tac, a monoclonal antibody with specificity for the IL 2 receptor. Of the mitogen- or MLR-activated T cells in CLL patients, 6% and 10%, respectively, expressed Tac antigen, whereas identically stimulated control T cells were 60% and 47% Tac+, respectively. Our findings suggest that T cells in B-CLL are defective in their recognition of self or foreign major histocompatibility antigens as demonstrated by their impaired responsiveness in the MLR. Thus, these cells are unable to produce IL 2 or generate IL 2 receptors.

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ICOS Is Induced by B Cell Receptor Signalling on Chronic Lymphocytic Leukemia B Cells.

Blood ◽

10.1182/blood.v104.11.969.969 ◽

2004 ◽

Vol 104 (11) ◽

pp. 969-969 ◽

Cited By ~ 1

Author(s):

Tetsuya Fukuda ◽

Traci L. Toy ◽

Laura Z. Rassenti ◽

Kanti R. Rai ◽

Thomas J. Kipps

Keyword(s):

Flow Cytometry ◽

T Cells ◽

B Cells ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Progressive Disease ◽

Leukemia Cell ◽

Lymphocytic Leukemia ◽

Leukemia Cells ◽

Activated T Cells

Abstract Patients with chronic lymphocytic leukemia (CLL) cells that express unmutated immunoglobulin (Ig) heavy chain variable region genes (IgVH genes) generally have a more aggressive clinical course than do patients with leukemia cells that express mutated IgVH. The reason(s) accounting for this are not known. Microarray gene expression analyses revealed that CLL cells that express unmutated IgVH could be distinguished from the leukemia cells that express mutated IgVH via the differential expression of a relatively small number of genes, one of which encodes the zeta-associated chain of 70kD (ZAP-70), which generally is expressed by CLL cells that express unmutated IgVH. Although the expression of ZAP-70 is associated with expression of unmutated IgVH in CLL, this association is not absolute. This was the case for a pair of monozygotic twins who both developed CLL at age 57. Although each of the twins had leukemia cells that expressed mutated IgVH, only one of the twins had leukemia cells that lacked expression of ZAP-70 protein and has indolent, non-progressive disease (Blood100: 4609–14, 2002). We performed microarray analysis using Affymetrix HG-U133A array on the isolated leukemia cells of each twin to define the genes that were differentially expressed between the two. In addition to ZAP-70, we found that the CLL cells of the twin with progressive disease also expressed the inducible co-stimulatory molecule (ICOS), a member of the CD28/CTLA-4 family of immune accessory co-stimulatory molecules that ordinarily only is expressed by activated T cells. Expression of ICOS protein by this leukemia B cell population, but not by the CLL B cells population of the other twin, was confirmed using fluorochrome-labeled anti-ICOS mAb and flow cytometry. We examined the CLL B cells from 58 additional patients for expression of ICOS by flow cytometry and found that 16 (28%) also expressed ICOS. We found that expression of ICOS was associated with expression of ZAP-70, as assessed via flow cytometry and immunoblot analyses. Whereas 14 of the 29 ZAP-70+ cases expressed ICOS, only 2 of the 29 ZAP-70-negative cases expressed this immune co-stimulatory molecule. Nevertheless, we found that nearly all of the 56 of the 58 cases expressed B7h, the ligand for ICOS. The two cases that did not express detectable B7h expressed ZAP-70 and were ICOS+. In preliminary studies, we found that treatment of ICOS-negative, ZAP-70+ CLL cells (n = 2) with goat anti-human Ig could induce expression of ICOS, suggesting that, as on T cells, this molecule also might be inducible in some cases of B cell CLL. Culture of ICOS+ CLL cells with an anti-B7h mAb capable of blocking ICOS-B7h interactions significantly enhanced ICOS surface expression, as assess by flow cytometry, suggesting that B7h may down-modulate ICOS through paracrine/autocrine receptor-ligand interactions. Because of this we evaluated for functional expression of ICOS on CLL B cells. We found that ligation of ICOS could induce enhanced signaling via the PI3K/Akt pathway in isolated CLL B cells, resulting in enhanced phosphorylation and activation of Akt. As such, we speculate that the expression of ICOS and its ligand in B cell CLL may enhance leukemia cell survival and/or proliferation, potentially contributing to the more aggressive disease observed in some patients with this disease.

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Measurement of Leukemic B-Cell Growth Kinetics in Patients with Chronic Lymphocytic Leukemia

Methods in Molecular Biology - Chronic Lymphocytic Leukemia ◽

10.1007/978-1-4939-8876-1_11 ◽

2018 ◽

pp. 129-151 ◽

Cited By ~ 1

Author(s):

Andrea N. Mazzarello ◽

Mark Fitch ◽

Marc K. Hellerstein ◽

Nicholas Chiorazzi

Keyword(s):

Cell Growth ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Growth Kinetics ◽

Lymphocytic Leukemia ◽

Cell Growth Kinetics

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Role of NFAT in Chronic Lymphocytic Leukemia and Other B-Cell Malignancies

Frontiers in Oncology ◽

10.3389/fonc.2021.651057 ◽

2021 ◽

Vol 11 ◽

Author(s):

Ilenia Sana ◽

Maria Elena Mantione ◽

Piera Angelillo ◽

Marta Muzio

Keyword(s):

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Therapeutic Potential ◽

Cell Receptor ◽

Distinct Pattern ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

B Cell Malignancies ◽

Activated T Cells ◽

Inflammatory Microenvironment

In recent years significant progress has been made in the clinical management of chronic lymphocytic leukemia (CLL) as well as other B-cell malignancies; targeting proximal B-cell receptor signaling molecules such as Bruton Tyrosine Kinase (BTK) and Phosphoinositide 3-kinase (PI3Kδ) has emerged as a successful treatment strategy. Unfortunately, a proportion of patients are still not cured with available therapeutic options, thus efforts devoted to studying and identifying new potential druggable targets are warranted. B-cell receptor stimulation triggers a complex cascade of signaling events that eventually drives the activation of downstream transcription factors including Nuclear Factor of Activated T cells (NFAT). In this review, we summarize the literature on the expression and function of NFAT family members in CLL where NFAT is not only overexpressed but also constitutively activated; NFAT controls B-cell anergy and targeting this molecule using specific inhibitors impacts on CLL cell viability. Next, we extend our analysis on other mature B-cell lymphomas where a distinct pattern of expression and activation of NFAT is reported. We discuss the therapeutic potential of strategies aimed at targeting NFAT in B-cell malignancies not overlooking the fact that NFAT may play additional roles regulating the inflammatory microenvironment.

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Defective T cell-mediated, isotype-specific immunoglobulin regulation in B cell chronic lymphocytic leukemia

Blood ◽

10.1182/blood.v71.4.1012.bloodjournal7141012 ◽

1988 ◽

Vol 71 (4) ◽

pp. 1012-1020 ◽

Cited By ~ 1

Author(s):

JS Moore ◽

MB Prystowsky ◽

RG Hoover ◽

EC Besa ◽

PC Nowell

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Cell Activity ◽

Lymphocytic Leukemia ◽

Specific Immunoglobulin ◽

Cell Chronic Lymphocytic Leukemia

The consistent occurrence of T cell abnormalities in patients with B cell chronic lymphocytic leukemia (B-CLL) suggest that the non- neoplastic host T cells may be involved in the pathogenesis of this B cell neoplasm. Because potential defects of immunoglobulin regulation are evident in B-CLL patients, we investigated one aspect of this by studying the T cell-mediated immunoglobulin isotype-specific immunoregulatory circuit in B-CLL. The existence of class-specific immunoglobulin regulatory mechanisms mediated by Fc receptor-bearing T cells (FcR + T) through soluble immunoglobulin binding factors (IgBFs) has been well established in many experimental systems. IgBFs can both suppress and enhance B cell activity in an isotype-specific manner. We investigated the apparently abnormal IgA regulation in a B-CLL patient (CLL249) whose B cells secrete primarily IgA in vitro. Enumeration of FcR + T cells showed a disproportionate increase in IgA FcR + T cells in the peripheral blood of this patient. Our studies showed that the neoplastic B cells were not intrinsically unresponsive to the suppressing component of IgABF produced from normal T cells, but rather the IgABF produced by the CLL249 host T cells was defective. CLL249 IgABF was unable to suppress IgA secretion by host or normal B cells and enhanced the in vitro proliferation of the host B cells. Size fractionation of both normal and CLL249 IgABF by gel-filtration high- performance liquid chromatography (HPLC) demonstrated differences in the ultraviolet-absorbing components of IgABF obtained from normal T cells v that from our patient with defective IgA regulation. Such T cell dysfunction may not be restricted to IgA regulation, since we have found similar expansion of isotype-specific FcR + T cells associated with expansion of the corresponding B cell clone in other patients with B-CLL. These data suggest that this T cell-mediated regulatory circuit could be significantly involved in the pathogenesis of B-CLL.

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Antigen presentation in an HLA-DR-restricted fashion by B-cell chronic lymphocytic leukemia cells

Blood ◽

10.1182/blood.v72.1.102.bloodjournal721102 ◽

1988 ◽

Vol 72 (1) ◽

pp. 102-108 ◽

Cited By ~ 7

Author(s):

M Yasukawa ◽

T Shiroguchi ◽

A Inatsuki ◽

Y Kobayashi

Keyword(s):

T Cells ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Antigen Presentation ◽

Interleukin 2 ◽

Class Ii ◽

Hla Class Ii ◽

Lymphocytic Leukemia ◽

Hla Dr ◽

Cell Chronic Lymphocytic Leukemia

The ability of B-cell chronic lymphocytic leukemia (B-CLL) cells to present antigen to antigen-specific T cells was investigated. B-CLL cells present herpes simplex virus (HSV) antigen and purified protein derivative (PPD) to HSV- and PPD-specific, interleukin-2-dependent T- cell lines in an antigen-specific manner. Treatment of B-CLL cells with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) induced markedly increased levels of HLA-DR expression. TPA-treated B-CLL cells showed substantially more effective presentation, especially at low antigen concentrations, than did untreated B-CLL cells. By coculturing different allogeneic combinations of B-CLL cells and T cells and by adding anti-HLA-DR monoclonal antibody to cultures, it was found that antigen presentation by B-CLL cells was restricted by HLA-DR in the same way as for macrophages. We concluded from these experiments that B- CLL cells have a capacity to serve as antigen-presenting cells in an HLA class II-restricted fashion and that increasing the amount of HLA class II antigen and activation of B-CLL cells resulted in effective antigen presentation.

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