Defective T Cell Migration in Chronic Lymphocytic Leukemia Is Repaired by Lenalidomide

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3117-3117
Author(s):  
Alan G. Ramsay ◽  
Lena Svensson ◽  
Nancy Hogg ◽  
John G. Gribben
Keyword(s):  
T Cells ◽  
Cell Migration ◽  
T Cell ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Cd8 T Cells ◽  
Direct Contact ◽  
Immune Surveillance ◽  
Lymphocytic Leukemia ◽  
T Cell Migration

Abstract We have previously demonstrated that multiple gene expression abnormalities are induced in T cells from chronic lymphocytic leukemia (CLL) patients including defects within the actin cytoskeleton signaling pathways that control immune recognition and motility (Gullu et al. JCI, 2005). T cell immune surveillance requires rapid migratory responses and LFA-1 (CD11a/CD18; αLβ2) is a promigratory receptor that engages the cytoskeleton to control migration. We hypothesized that CLL T cells may exhibit dysfunctional migration in response to ICAM-1, the principal ligand for LFA-1. Using time lapse microscopy, we observed significantly reduced chemokine SDF-1 (CXCL12) induced migration on ICAM-1 of CLL CD4 and CD8 T cells compared to age-matched healthy donor T cells. Healthy T cells tracked for 45 min displayed a random course of migration with an average speed of ~ 8 μm/min, whereas CLL T cells were slower ~ 5 μm/min (n=14, ~ 30% reduction, p<0.01). We further postulated that direct contact of CLL tumor cells with healthy T cells would induce this migratory defect. Healthy CD4 or CD8 T cells were cocultured with either allogeneic CLL B cells or allogeneic healthy B cells and subsequently used in migration assays. Co-culture with CLL cells resulted in significantly reduced T cell migration compared with co-culture with healthy B cells (~ 44% reduction in migration, n=6, p<0.01). Evidence that direct contact was required to induce this migratory defect was shown when no effect was observed when cell-cell adhesion was prevented by pretreatment of CLL cells with anti-ICAM-1 blocking antibody prior to primary co-culture with healthy T cells. This cancer-induced migratory defect was repaired when CLL T cells were pretreated with the immunomodulatory drug Lenalidomide (1μM for 1hr). Treatment with this agent enhanced the migratory potential of CLL T cells to a speed comparable to untreated and treated healthy T cells. The finding that lenalidomide can restore rapid migration in patient T cells provides evidence that this agent may increase immune surveillance in CLL patients.

Leukemia-associated monoclonal and oligoclonal TCR-BV use in patients with B-cell chronic lymphocytic leukemia

Blood ◽  
2003 ◽  
Vol 101 (3) ◽  
pp. 1063-1070 ◽  
Author(s):  
Mohammad-Reza Rezvany ◽  
Mahmood Jeddi-Tehrani ◽  
Hans Wigzell ◽  
Anders Österborg ◽  
Håkan Mellstedt
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Cd8 T Cells ◽  
Leukemia Cell ◽  
Lymphocytic Leukemia ◽  
Cell Chronic Lymphocytic Leukemia

Abstract T-cell receptor–B-variable (TCR-BV) gene usage and the CDR3 size distribution pattern were analyzed by reverse transcription–polymerase chain reaction (RT-PCR) in patients with B-cell chronic lymphocytic leukemia (B-CLL) to assess the T-cell repertoire. The use of TCR-BV families in CD4 and CD8 T cells stimulated with autologous activated leukemic cells was compared with that of freshly obtained blood T cells. Overexpression of individual TCR-BV families was found in freshly isolated CD4 and CD8 T cells. Polyclonal, oligoclonal, and monoclonal TCR-CDR3 patterns were seen within such overexpressed native CD4 and CD8 TCR-BV families. In nonoverexpressed TCR-BV families, monoclonal and oligoclonal populations were noted only within the CD8 subset. After in vitro stimulation of T cells with autologous leukemic B cells, analyses of the CDR3 length patterns showed that in expanded TCR-BV populations, polyclonal patterns frequently shifted toward a monoclonal/oligoclonal profile, whereas largely monoclonal patterns in native overexpressed TCR-BV subsets remained monoclonal. Seventy-five percent of CD8 expansions found in freshly obtained CD8 T cells further expanded on in vitro stimulation with autologous leukemic B cells. This suggests a memory status of such cells. In contrast, the unusually high frequency of CD4 T-cell expansions found in freshly isolated peripheral blood cells did not correlate positively to in vitro stimulation as only 1 of 9 expansions continued to expand. Our data suggest that leukemia cell–specific memory CD4 and CD8 T cells are present in vivo of patients with CLL and that several leukemia cell–associated antigens/epitopes are recognized by the patients' immune system, indicating that whole leukemia cells might be of preference for vaccine development.

scholarly journals Lenalidomide and Dexamethasone Combination in Patients with Chronic Lymphocytic Leukemia (CLL) Relapsing or Resistant to Treatment (LENDEX-LLC-09): A Gene Expression Profiling Study

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4675-4675
Author(s):  
Anna Puiggros ◽  
Pau Abrisqueta ◽  
Lara Nonell ◽  
Marta Bodalo ◽  
Eulalia Puigdecanet ◽  
...  
Keyword(s):  
Gene Expression ◽  
Multiple Myeloma ◽  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Cd8 T Cells ◽  
Lymphocytic Leukemia ◽  
Down Regulation ◽  
Tnf Α

Abstract Background. Chronic lymphocytic leukemia (CLL) is a highly heterogeneous disease in which immune evasion of tumoral cells, as well as, an impaired CD4 and CD8 T-cell function have been described. Immunomodulatory drugs, such as lenalidomide, alone or in combination with other treatments are promising strategies for those patients with refractory disease. The combination of lenalidomide with dexamethasone has been investigated in multiple myeloma and has revealed as a highly efficient treatment. Nonetheless, the efficacy and mechanisms of action of this combination in CLL have not been elucidated. Aim. To assess the effect of lenalidomide and dexamethasone combination in gene expression of CLL B cells, as well as CD4+ and CD8+ T cells from CLL patients enrolled in LENDEX-LLC-09 trial. Methods. Four patients included in the LENDEX-LLC-09 trial (NCT01246557) were studied (2M/2F, med age 72). All presented with advanced CLL (2 B and 2 C Binet stages), and were previously treated by a minimum of two chemo-immunotherapy regimens. Peripheral blood samples were taken at the recruitment and the 7th day of the first cycle of lenalidomide (2.5mg/day) and dexamethasone (20mg/day, 4 days). Total RNA was extracted from CLL B cells (CD5+ CD19+) and T cells (CD4+ or CD8+) positively selected by immunomagnetic methods (Miltenyi Biotec). Good quality RNA (RIN>7) was hybridized to Human Gene 2.0 ST array (Affymetrix). Differences between gene expression of pretreated and treated samples were assessed for each cell type using linear models for microarrays. Genes with a |logFC|>1 were considered as potentially relevant. Functional analysis was performed using Ingenuity Pathway Analysis (IPA). Results and discussion. The major effect in the gene expression due to treatment was observed in CD4+ T cells, which presented 290 up-regulated genes and 103 down-regulated. CLL cells showed up-regulation of 189 and down-regulation of 53 genes, while increase and decrease in the expression of 112 and 37 genes, respectively, were found in CD8+ T cells. Globally, the most important involved networks were related to cell-to-cell signaling, cellular growth and proliferation, cell death and survival, as well as inflammatory response and immune cell trafficking. Regarding CLL B cells, TNF-α was the most up-regulated gene, as previously described in lenalidomide treated B cells. Contrarily, we did not observe significant differences in genes involved in the immunologic synapse, as CD80, CD86, CD200, PD-L1, CD276 and CD270, which have been reported as key regulators in lenalidomide mechanism of action. Of note, a general increase of genes associated with binding to cells (CD68, CTLA4, ADAM28, ITGAX, LY96) was detected. In contrast to previous studies that demonstrated a growth arrest and induction of apoptosis by lenalidomide or dexamethasone in monotherapy (Baptista et al, 2012; Fecteau et al, 2014), a global inhibition of the apoptosis (up-regulation of BTK and CD79B and inhibition of SMAD7, among others) were observed when both drugs were combined. Considering CD8+ T cells gene expression, an up-regulation of genes involved in leukocyte activation and cell-to-cell binding was detected. The most remarkable changes were found in TNF-α and IFN-γ induction, as well as in ADAM28, LY96 and CD68. In contrast to CD8+ T cells, an inhibition of CD4+ T cell proliferation was observed after the combined treatment (up-regulation of VSIG4, LILRB4 and down-regulation of ICOS). This observation suggests that dexamethasone administration inhibits the CD4+ activation promoted by lenalidomide, as has been described in multiple myeloma (Hsu et al, 2011). Regarding response to treatment, two patients initially presented a complete response with positive minimal residual disease. However, all patients finally progressed after treatment and one died due to disease progression. No significant differences in gene expression patterns were found among patients. Conclusions. Our results suggest that lenalidomide and dexamethasone combination leads to an anti-tumoral activity displayed by an activation of CD8+ T cells against the tumor, rather than an increase of apoptosis in CLL cells. More studies are needed to confirm these preliminary findings of the combined effect of lenalidomide and dexamethasone in refractory CLL patients. Acknowledgments. This work was funded by Celgene, and supported by PI11/1621, 14SGR585 and Fundació LaCaixa. Disclosures Off Label Use: Lenalidomide and dexamethasone combination in CLL.

scholarly journals Targeting CD38 is lethal to Breg-like chronic lymphocytic leukemia cells and Tregs, but restores CD8+ T-cell responses

2020 ◽  
Vol 4 (10) ◽  
pp. 2143-2157 ◽  
Author(s):  
Alak Manna ◽  
Timothy Kellett ◽  
Sonikpreet Aulakh ◽  
Laura J. Lewis-Tuffin ◽  
Navnita Dutta ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Chronic Lymphocytic Leukemia ◽  
Cd8 T Cells ◽  
Mononuclear Cells ◽  
Transforming Growth Factor ◽  
Ex Vivo ◽  
Xenograft Model ◽  
Lymphocytic Leukemia ◽  
Dependent Manner

Abstract Patients with chronic lymphocytic leukemia (CLL) are characterized by monoclonal expansion of CD5+CD23+CD27+CD19+κ/λ+ B lymphocytes and are clinically noted to have profound immune suppression. In these patients, it has been recently shown that a subset of B cells possesses regulatory functions and secretes high levels of interleukin 10 (IL-10). Our investigation identified that CLL cells with a CD19+CD24+CD38hi immunophenotype (B regulatory cell [Breg]–like CLL cells) produce high amounts of IL-10 and transforming growth factor β (TGF-β) and are capable of transforming naive T helper cells into CD4+CD25+FoxP3+ T regulatory cells (Tregs) in an IL-10/TGF-β-dependent manner. A strong correlation between the percentage of CD38+ CLL cells and Tregs was observed. CD38hi Tregs comprised more than 50% of Tregs in peripheral blood mononuclear cells (PBMCs) in patients with CLL. Anti-CD38 targeting agents resulted in lethality of both Breg-like CLL and Treg cells via apoptosis. Ex vivo, use of anti-CD38 monoclonal antibody (mAb) therapy was associated with a reduction in IL-10 and CLL patient-derived Tregs, but an increase in interferon-γ and proliferation of cytotoxic CD8+ T cells with an activated phenotype, which showed an improved ability to lyse patient-autologous CLL cells. Finally, effects of anti-CD38 mAb therapy were validated in a CLL–patient-derived xenograft model in vivo, which showed decreased percentage of Bregs, Tregs, and PD1+CD38hiCD8+ T cells, but increased Th17 and CD8+ T cells (vs vehicle). Altogether, our results demonstrate that targeting CD38 in CLL can modulate the tumor microenvironment; skewing T-cell populations from an immunosuppressive to immune-reactive milieu, thus promoting immune reconstitution for enhanced anti-CLL response.

scholarly journals Immunomodulatory effects of different intravenous immunoglobulin preparations in chronic lymphocytic leukemia

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ana Colado ◽  
Esteban Enrique Elías ◽  
Valeria Judith Sarapura Martínez ◽  
Gregorio Cordini ◽  
Pablo Morande ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
T Cell Activation ◽  
Cell Activation ◽  
Lymphocytic Leukemia ◽  
Immunomodulatory Effects ◽  
Immunoglobulin Preparations

AbstractHypogammaglobulinemia is the most frequently observed immune defect in chronic lymphocytic leukemia (CLL). Although CLL patients usually have low serum levels of all isotypes (IgG, IgM and IgA), standard immunoglobulin (Ig) preparations for replacement therapy administrated to these patients contain more than 95% of IgG. Pentaglobin is an Ig preparation of intravenous application (IVIg) enriched with IgM and IgA (IVIgGMA), with the potential benefit to restore the Ig levels of all isotypes. Because IVIg preparations at high doses have well-documented anti-inflammatory and immunomodulatory effects, we aimed to evaluate the capacity of Pentaglobin and a standard IVIg preparation to affect leukemic and T cells from CLL patients. In contrast to standard IVIg, we found that IVIgGMA did not modify T cell activation and had a lower inhibitory effect on T cell proliferation. Regarding the activation of leukemic B cells through BCR, it was similarly reduced by both IVIgGMA and IVIgG. None of these IVIg preparations modified spontaneous apoptosis of T or leukemic B cells. However, the addition of IVIgGMA on in vitro cultures decreased the apoptosis of T cells induced by the BCL-2 inhibitor, venetoclax. Importantly, IVIgGMA did not impair venetoclax-induced apoptosis of leukemic B cells. Overall, our results add new data on the effects of different preparations of IVIg in CLL, and show that the IgM/IgA enriched preparation not only affects relevant mechanisms involved in CLL pathogenesis but also has a particular profile of immunomodulatory effects on T cells that deserves further investigation.

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

Blood ◽  
1988 ◽  
Vol 71 (4) ◽  
pp. 1012-1020 ◽  
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.

Defective LFA-1 Mediated T Cell Motility In Chronic Lymphocytic Leukemia Is Mediated by Defects In the Rho GTPase Signaling Pathway

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 914-914
Author(s):  
Alan G. Ramsay ◽  
Rachel Evans ◽  
Lena Svensson ◽  
Shahryar Kiaii ◽  
Nancy Hogg ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Migration ◽  
Cell Adhesion ◽  
T Cell ◽  
Chronic Lymphocytic Leukemia ◽  
Cell Motility ◽  
Healthy Donor ◽  
Rho Gtpase ◽  
Lymphocytic Leukemia ◽  
Donor T Cells

Abstract Abstract 914 T lymphocytes have an essential role in adaptive immunity and rely on tightly regulated signaling through integrin lymphocyte function-associated antigen (LFA)-1 to migrate into lymph nodes and interact with antigen-presenting cells. Malignant cells modify their immune microenvironment to prevent effective host anti-tumor responses, promote tumor progression, and suppress the therapeutic benefit of immunotherapy treatments. Here we assessed LFA-1-mediated cell migration of highly purified T cells from treatment naïve chronic lymphocytic leukemia (CLL) patients compared to age-matched healthy donor T cells using CXCL12 stimulation and immobilized ICAM-1, the principal integrin ligand. Video microscopy with motility tracking analysis identified that both CD4 and CD8 T cells from CLL patients (n=14) exhibited significantly reduced migration rates (P < .01) compared to healthy donor T cells (5.5 ± 0.3 (SEM) μm/min and 4.4 ± 0.2 μm/min compared to 8.2 ± 0.3 μm/min and 7.5 ± 0.3 μm/min respectively). We further identified that direct CLL cell contact, and not soluble factors alone, induced similar T cell motility dysfunction in previously healthy CD3 T cells. Primary co-culture of healthy donor T cells with CLL cells caused a significant decrease in the speed of migration on ICAM-1 compared to coculture with control healthy B cells (6.2 ± 0.3 μm/min versus 9.5 ± 0.6 μm/min) (n=9) (P < .05). Next we sought to repair this T cell defect in CLL using a clinically relevant agent. We identify that treatment of CLL patient T cells (n=9) with lenalidomide restores rapid LFA-1 mediated migration on ICAM-1. Ex vivo treatment of CLL T cells with lenalidomide (1μM for 24 hours) significantly increased the speed of T cell migration compared to untreated patient T cells (7 ± 0.4 μm/min versus 2.5 ± 0.7 μm/min) (P < .05) and the rescued T cell migratory function of lenalidomide exposed patient T cells was comparable to healthy donor T cells treated with or without drug. Interference reflection microscopy (IRM) examining the contact zone between migrating T cells and ICAM-1 identified a significant CLL patient T cell adhesion defect (P < .05) with reduced spreading area and strength of adhesive contacts (pixel density) compared to healthy donor T cells. IRM was further utilized with pharmacological inhibitors to demonstrate that exposure to lenalidomide rescued CLL T cell adhesion by acting on the Rho family GTPases that are dysregulated in cancer patient T cells. Lenalidomide significantly increased (P < .05) levels of active RhoA in CLL patient T cells compared to untreated cells. In addition, untreated CLL patient T cells adhering to ICAM-1 exhibited significantly reduced expression levels of phosphorylated myosin light chain (MLC) compared to healthy donor T cells (P < .05) and this defect was repaired following lenalidomide treatment. MLC is normally phosphorylated by MLC kinase at the T cell leading edge and by the RhoA target, ROCK at the trailing edge, and is an important downstream signaling molecule during LFA-1-mediated T cell motility. Further expression analysis identified that lenalidomide significantly increased (P < .01) ICAM-1-engaged high-affinity LFA-1 in CLL patient T cells to levels comparable to healthy donor T cells. Overall, our results show that T cells in CLL patients have dysfunctional tumor-induced cytoskeletal signaling via the Rho GTPase signaling pathway, and this is reversed by lenalidomide, rescuing dynamic LFA-1 mediated outside-in signalling and migration. Lenalidomide's immunomodulatory activity was highly cancer T cell specific: rescuing defective LFA-1 migration and signaling in CLL T cells, but with no detectable effects on healthy donor T cells. These findings provide important mechanistic insight into the action of lenalidomide, and highlight the potential clinical utility of immunomodulatory drugs to rescue normal immune function in cancer. Disclosures: Gribben: Roche: Consultancy; Celgene: Consultancy; GSK: Honoraria; Napp: Honoraria.

Expansion of CMV-specific CD8+CD45RA+CD27- T cells in B-cell chronic lymphocytic leukemia

Blood ◽  
2003 ◽  
Vol 102 (3) ◽  
pp. 1057-1063 ◽  
Author(s):  
Wendelina J. M. Mackus ◽  
Florine N. J. Frakking ◽  
Annette Grummels ◽  
Laila E. Gamadia ◽  
Godelieve J. de Bree ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Chronic Lymphocytic Leukemia ◽  
Immune Responses ◽  
B Cell ◽  
Cd8 T Cells ◽  
Cytotoxic T Cells ◽  
Lymphocytic Leukemia ◽  
The Absolute ◽  
Cell Chronic Lymphocytic Leukemia

Abstract In patients with B-cell chronic lymphocytic leukemia (B-CLL), the absolute number of T cells is increased. Although it has been suggested that these T cells might be tumor specific, concrete evidence for this hypothesis is lacking. We performed a detailed immunophenotypic analysis of the T-cell compartment in the peripheral blood of 28 patients with B-CLL (Rai 0, n = 12; Rai I-II, n = 10; Rai III-IV, n = 6) and 12 healthy age-matched controls and measured the ability of these patients to mount specific immune responses. In all Rai stages a significant increase in the absolute numbers of CD3+ cells was observed. Whereas the number of CD4+ cells was not different from controls, patients with B-CLL showed significantly increased relative and absolute numbers of CD8+ cells, which exhibited a CD45RA+CD27- cytotoxic phenotype. Analysis of specific immune responses with tetrameric cytomegalovirus (CMV)–peptide complexes showed that patients with B-CLL had significantly increased numbers of tetramer-binding CMV-specific CD8+ T cells. The rise in the total number of CD8+ cytotoxic T cells was evident only in CMV-seropositive B-CLL patients. Thus, our data suggest that in patients with B-CLL the composition of T cells is shifted toward a CD8+ cytotoxic cell type in an effort to control infections with persistent viruses such as CMV. Moreover, they offer an explanation for the high incidence of CMV reactivation in CLL patients treated with T cell–depleting agents, such as the monoclonal antibody (mAb) alemtuzumab (Campath; α-CD52 mAb). Furthermore, because in CMV-seronegative patients no increase in cytotoxic CD8+ T cells is found, our studies do not support the hypothesis that tumor-specific T cells account for T-cell expansion in B-CLL.

scholarly journals A Detailed Analysis of Parameters Supporting the Engraftment and Growth of Chronic Lymphocytic Leukemia Cells in Immune-Deficient Mice

2021 ◽  
Vol 12 ◽  
Author(s):  
Piers E. M. Patten ◽  
Gerardo Ferrer ◽  
Shih-Shih Chen ◽  
Jonathan E. Kolitz ◽  
Kanti R. Rai ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Intraperitoneal Administration ◽  
Human Condition ◽  
Lymphocytic Leukemia ◽  
Lymphoid Tissues ◽  
Quiescent State

Patient-derived xenograft models of chronic lymphocytic leukemia (CLL) can be created using highly immunodeficient animals, allowing analysis of primary tumor cells in an in vivo setting. However, unlike many other tumors, CLL B lymphocytes do not reproducibly grow in xenografts without manipulation, proliferating only when there is concomitant expansion of T cells. Here we show that in vitro pre-activation of CLL-derived T lymphocytes allows for a reliable and robust system for primary CLL cell growth within a fully autologous system that uses small numbers of cells and does not require pre-conditioning. In this system, growth of normal T and leukemic B cells follows four distinct temporal phases, each with characteristic blood and tissue findings. Phase 1 constitutes a period during which resting CLL B cells predominate, with cells aggregating at perivascular areas most often in the spleen. In Phase 2, T cells expand and provide T-cell help to promote B-cell division and expansion. Growth of CLL B and T cells persists in Phase 3, although some leukemic B cells undergo differentiation to more mature B-lineage cells (plasmablasts and plasma cells). By Phase 4, CLL B cells are for the most part lost with only T cells remaining. The required B-T cell interactions are not dependent on other human hematopoietic cells nor on murine macrophages or follicular dendritic cells, which appear to be relatively excluded from the perivascular lymphoid aggregates. Notably, the growth kinetics and degree of anatomic localization of CLL B and T cells is significantly influenced by intravenous versus intraperitoneal administration. Importantly, B cells delivered intraperitoneally either remain within the peritoneal cavity in a quiescent state, despite the presence of dividing T cells, or migrate to lymphoid tissues where they actively divide; this dichotomy mimics the human condition in that cells in primary lymphoid tissues and the blood are predominately resting, whereas those in secondary lymphoid tissues proliferate. Finally, the utility of this approach is illustrated by documenting the effects of a bispecific antibody reactive with B and T cells. Collectively, this model represents a powerful tool to evaluate CLL biology and novel therapeutics in vivo.

scholarly journals Lysis of malignant B cells from patients with B-chronic lymphocytic leukemia by autologous T cells activated with CD3 x CD19 bispecific antibodies in combination with bivalent CD28 antibodies

Blood ◽  
1993 ◽  
Vol 82 (6) ◽  
pp. 1803-1812 ◽  
Author(s):  
H Bohlen ◽  
T Hopff ◽  
O Manzke ◽  
A Engert ◽  
D Kube ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
T Cell Activation ◽  
Cell Activation ◽  
Lymphocytic Leukemia ◽  
Bispecific Antibodies ◽  
Autologous Tumor ◽  
Activation Markers

Abstract Bispecific antibodies (bi-MABs) can be used to target T cells to autologous tumor cells. It has been shown that the activation of resting human T cells requires two independent signals, namely the cross-linking of the T-cell receptor (TCR)-CD3 complex together with the CD28 homodimer. In the present study, we demonstrate the activation of T cells from patients with chronic lymphocytic leukemia (CLL) using bi-MABs against the CD3 and CD19 antigens (CD3 x CD19) in combination with monospecific, bivalent antibodies against the CD28 antigen. Mononuclear cells from patients with CLL were cultured with the bi-MAB CD3 x CD19 and monospecific CD28 antibodies. The CD3 x CD19 bi-MABs were isolated by the hybridoma-hybridoma fusion technique and purified by hydrophobic interaction chromatography. T-Cell activation as demonstrated by increased proliferation, upregulation of T-cell activation markers (CD25, CD38), and cytotoxicity against autologous CLL cells and allogeneic B cells was shown in seven of eight CLL specimens. The stimulation with CD3 x CD19 bi-MABs with CD28 antibodies preferentially induced proliferation of CD4+ T cells. The effective dose of purified antibodies required for optimal T-cell activation was 100 ng/mL in vitro, which suggests that this antibody combination may be useful for immunotherapy of patients with B-CLL.

Increased Expression of CD152 by Normal T Lymphocytes in Untreated Patients with B Cell Chronic Lymphocytic Leukemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 963-963
Author(s):  
Marina Motta ◽  
Bobby Shelvin ◽  
Susan Lerner ◽  
Michael Keating ◽  
William G. Wierda
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Chronic Lymphocytic Leukemia ◽  
Cd8 T Cells ◽  
Lymphocytic Leukemia ◽  
Peripheral Tolerance ◽  
Regulatory Function ◽  
Cell Phenotype ◽  
Costimulatory Receptor

Abstract Patient with chronic lymphocytic leukemia (CLL) have defects in both cellular and humoral immunity. Despite reported increases in absolute T cell counts in untreated patients with CLL, abnormalities of T cell phenotype and function have been described as well as progressive hypogammaglobulinemia. Furthermore, defects are compounded by current treatments for the disease. Expansion and differentiation of normal antigen-specific T cells depends upon two signals: binding of the T cell receptor to antigen presented in the context of self MHC molecules and ligation of a costimulatory receptor. CD28 is the primary T cell surface costimulatory receptor and is constitutively expressed on almost all CD4+ and about 50% of CD8+ T cells. The ligands CD80 and CD86 bind CD28, thereby transducing the second enhancing signal for T cell proliferation and cytokine secretion. CD152 (CTLA-4) has homology to CD28 and binds to CD80 and CD86 with much higher affinity, but plays a critical role in the down regulating T cell responses and maintenance of peripheral tolerance. Surface CD152 is not normally expressed on resting T cells, but is induced upon activation. We hypothesized that in previously untreated patients with CLL, T cell anergy is the result of increased expression of CD152. Therefore, we studied the expression of surface and cytoplasmic CD152 (sCD152 and cCD152, respectively) in freshly isolated T cells from blood (N=40) and bone marrow (N=14) of previously untreated patients with CLL. Also, the activation status of these T cells was evaluated by evaluating IL-2 receptor subunit expression. CD4+ and CD8+ T cells from patients with CLL demonstrated significant increase in sCD152 and cCD152 compared to T cells from normal donors (Table 1). Table 1 Expression of CD152 by T Cells Mean % Positive T Cell Population Normal CLL P-value sCD152 N=13 N=40 CD4+ 0.8 5.0 <.01 CD4+/CD25+ 1.8 11.5 <.05 CD8+ 1.8 5.0 <.05 cCD152 N=13 N=19 CD4+ 6.9 40.4 <.01 CD4+/CD25+ 26.6 48.0 <.01 CD8+ 1.3 16.9 <.05 Furthermore, patients with CLL had an increased proportion of CD4+/CD25+/CD152+ cells. This subpopulation of T cells is known to have a regulatory function. T cells from patients with CLL (N=25) also showed an activated immunophenotype with significantly increased proportion of CD4+ and CD8+ T cells co-expressing the CD122/CD25 subunits of the IL-2 receptor compared to normal donors (N=10). No significant differences were seen in proportion or pattern of expression of these antigens between peripheral blood and bone marrow cells. These findings suggest that the T cells have been activated, however, may be primed for hyporesponsiveness and peripheral tolerance by expression of CD152. Correlations between CD152 expression and relevant clinical and biological variables were made in these previously untreated patients. The number of CD4+/CD152+ and CD4+/CD25+/CD152+ cells from patients with CLL inversely correlated with serum IgG and IgA levels. These findings suggest a further possible involvement of CD152 in the possible suppression of normal B cells in patients with CLL. The proportion of CD4+/CD25+/CD152+ cells also correlated with advanced Rai stage. In summary, T cells from patients with CLL are potentially primed for anergy by expression of CD152. Functional studies to investigate the role of CD152 and CD4+/CD25+/CD152+ cells in patients with CLL are ongoing, with the goal to develop immunotherapeutic strategies.

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