A Novel Asymmetrical Anti-CLL-1×CD3 Bispecific Antibody, ABL602, Induces Potent CLL1-Specific Antitumor Activity with Minimized Sensitization of Pro-Inflammatory Cytokines

Abstract Acute myeloid leukemia (AML) is a disease with high incidence of relapse that is originated and maintained from leukemia stem cells (LSCs). Expression of C-type lectin-like molecule-1 (CLL-1; also known as CLEC12A, c-type lectin domain family 12 member A) is mainly restricted to LSCs but absent in normal hematopoietic stem cells (HSCs), suggesting CLL-1 as an excellent therapeutic target for AML. This unique expression pattern paves the way to develop therapies that potentially eliminate CLL1-positive LSC while preserving CLL1-negative HSC. To re-direct T cells to AML cells, we generated IgG-based asymmetric (2+1, ABL602) bispecific antibody (BsAb) targeting CLL-1 and CD3. As a 2+1 format BsAb, ABL602 has bivalent binding to CLL-1 for target arm and monovalent binding to CD3. ABL602 exhibited higher binding activity to CLL-1-expressing AML cell lines and greater tumor-killing efficacy than 1+1 format BsAb and benchmark antibody MCLA-117 (Merus; CLEC12AxCD3 bispecific antibody). ABL602 induced potent cytotoxic activities on CLL1-expressing AML cell lines (EC 50 of 0.04~3.05pM and 0.97~16.64pM for U937 and HL-60, respectively) with concomitant T cell activation (EC 50 of 0.10~3.54pM and 0.94~4.92pM for U937 and HL-60, respectively) and cytokine/granzyme B release. Despite strong tumor-killing activity, ABL602 did not kill CLL1-negative cancer cell lines, suggesting that ABL602 induces CLL-1-dependent cytotoxicity. Moreover, ABL602 did not or minimally induce TNF-α and IL-6 in PBMC in the absence of AML cell lines, while MCLA-117 triggered high level of expression of those cytokines. In established orthotopic AML mouse model using HL-60 Luc, ABL602 demonstrated statistically significant anti-tumor activity in a dose-dependent manner. Proportions of bone marrow CD33 + AML blasts diminished in a dose-dependent manner, while CD3 + T cells more infiltrated to the bone marrow. Overall, our results indicate that ABL602, appropriately engineered 2+1 asymmetric BsAb, promotes T-cell activity specifically against CLL1-expressing AML cells and is a promising treatment strategy for AML patients by achieving the desired balance between antitumor activity and safety. Disclosures No relevant conflicts of interest to declare.

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Dose and Timing Limitations in the Use of Bone Marrow-Derived Mesenchymal Stem Cells for the Treatment of Experimental Arthritis.

Blood ◽

10.1182/blood.v112.11.2326.2326 ◽

2008 ◽

Vol 112 (11) ◽

pp. 2326-2326

Author(s):

Evangelia Yannaki ◽

Anastasia Papadopoulou ◽

Minas Yiangou ◽

Evangelia Athanasiou ◽

Argyrw Paraskeva ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

T Cells ◽

Mesenchymal Stem Cells ◽

T Cell ◽

Broad Spectrum ◽

Synovial Membrane ◽

Cell Contact ◽

Dependent Manner

Abstract The recently recognized potential of mesenchymal stem cells (MSCs) to differentiate into a broad spectrum of tissues and to act as immune regulators beyond the barriers of embryonic germ layers and major histocombatibility comlex (MHC) restriction, has emerged intense research interest on their possible use in a broad spectrum of clinical entities. Although the immunoregulatory potential of MSCs has been shown to effectively control GvHD in several preclinical and clinical studies, their role in autoimmune diseases has not been extensively explored in animal models. The goal of this study was to investigate the in vitro effect of rat bone marrow-derived MSCs on cultured fibrobIast-like synoviocytes (FLS) and T-cells from the spleen after induction of adjuvant arthritis (AA) by FCA as well as their in vivo effect in a rat model of AA resembling human rheumatoid arthritis. MSCs were isolated from bone marrow and were characterized by CD45 negativity and CD54, CD29 positivity in FCM analysis. Differentiation assays were performed to confirm their adipogenic, osteogenic and chondrogenic potential. Culture of AA-FLS in the presence of supernatant from syngeneic (syng) or allogeneic (allo) MSCs at passage 2–3, reduced the AA-FLS (p<0.022) and the ConA-stimulated AA-T-cell (p=0.04) proliferation in a dose-dependent manner, as compared to AA-FLS or AA-T-cell proliferation in the absence of supernatant. Cell-to-cell contact by coculture of activated T-cells with syng or allo MSCs produced a stronger inhibition over the supernatant (p<0.0001), in all tested MSCs dilutions and even at the lowest MSCs :T-cell ratio of 0.05:1. The inhibitory effect of allo as compared to syng MSCs in activated AA T-cells, was stronger both by secreted agents (p=0.017) or by cell to cell contact (p=0.0001). In vivo, low doses of syng MSCs (0.5-5x10^5cell/recipient) administered iv, intrasplenic or intrabone marrow, at single or multiple infusions, didn’t significantly reduce the disease score of MSC-treated as compared to control rats. In contrast, repeated, higher dose (6x10^6cell/recipient), iv infusions of syng or allogeneic MSCs from male donors (Y+MSCs) to female recipients, before the onset of AA (d4 and d9 post AA induction) resulted in significantly lower arthritic scores when compared to control animals. MSC-treated animals preserved a rather normal joint architecture with focal synovial hyperplasia, limited pannus formation and without bone destruction or chondroplasia. In contrast, the joints of arthritic control rats, appeared with a thickened synovial membrane, erosive pannus and dense inflammatory cell infiltration, chondroplasia and osteoplasia. Reduced presence of CD3+, CD11b+, NF-kb+ cells and less intense angiogenesis (FVIII+cells) was demonstrated by immunohistochemistry in the synovium of transplanted rats as compared to the control group. No Y+MSCs were detected in the spleen, bone marrow or in cultured FLS from the synovial membrane at day30 post AA induction, by PCR (sry gene), immunohistochemistry (sry protein) or FISH (Y chromosome), suggesting that the observed benefit was mostly a result of immunomodulation not derived by MSCs homing to target tissues, or migration of MSCs to target tissues may have occured earlier. On the other hand, when the same cell dose was injected after the onset of arthritis (d13 and d20 post AA induction) no clinical benefit could be observed. Our data suggest that MSCs may represent a new therapeutic approach for autoimmune arthritis, however, due to dose and timing limitations in their use, further studies are needed to clinically exploit this potential.

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Cure of Murine Medulloblastoma by Adoptive Transfer of Tumor-Specific T Cells.

Blood ◽

10.1182/blood.v110.11.4904.4904 ◽

2007 ◽

Vol 110 (11) ◽

pp. 4904-4904

Author(s):

Cameron L. Nicholson ◽

Gregory E. Plautz

Keyword(s):

T Cells ◽

T Cell ◽

Cell Lines ◽

Adoptive Transfer ◽

Ex Vivo ◽

Neurological Dysfunction ◽

Dependent Manner ◽

Medulloblastoma Cell ◽

Dose Dependent ◽

Medulloblastoma Cell Lines

Abstract Medulloblastoma is the most common malignant brain tumor of childhood. Although the current overall survival has increased due to more aggressive therapy, the involved surgery, radiation and in some diseases high-dose chemotherapy have significant acute and long-term toxicities. T cell immunotherapy is a promising adjuvant for medulloblastoma because the immune response is specific for targeted antigens and is highly anatomically focused, occurring at the level of cell-to-cell interactions, thereby sparing adjacent normal brain tissue. Adoptive transfer of tumor-sensitized T cells has demonstrated therapeutic efficacy in animal tumor models, and clinical trials involving transfer of T lymphocytes are ongoing. Previous strategies have employed cultured human medulloblastoma cell lines as therapeutic targets, however priming of T cells using a spontaneously-arising medulloblastoma cell line in an immunocompetent mice model may be more representative of human cancer and more applicable to clinical practice. Spontaneous Murine medulloblastoma cell lines were created from knock-out Ptc+/−p53−/− mice and passaged in vivo. Brain tumors were established by transcranial inoculation of a single-cell suspension of murine medulloblastoma tumor cells in immunocompetent, C57BL/6N mice. Murine Medulloblastoma cell lines were rapidly tumorigenic and consistently fatal following intracranial inoculation in a dose-dependent manner. T-cells were obtained surgically from tumor draining lymph nodes (TDLN) in syngeneic mice and after initial purification, antigen-stimulated CD62Llow T cells were isolated by depletion of CD62Lhigh cells using MACS beads. T-cells were further activated ex vivo by incubation with anti-CD3 monoclonal antibody and exposure to media containing Interleukin-2 and Interleukin-7. Mice carrying 3-day established tumors receive systemic transfer of T cells by injection through the tail vein after 10 days of ex vivo culture. Mice typically receive sub-lethal total body irradiation (5 Gy) from a Cesium irradiator for lympho-depletion several hours before adoptive transfer. Mice were followed for survival and sacrificed when neurological symptoms were apparent as per Institutional Animal Care and Use Committee established guidelines. The activated T cells adoptively transferred to irradiated hosts with 3-day established intracranial MM1 tumors resulting in complete regression of established tumors (see figure). In addition, the above therapy demonstrated a dose-dependent effect, and the mice were cured without any evidence of neurological dysfunction caused by the anti-tumor response. This research suggests anti-medulloblastoma T cell vaccines may be promising for subsequent clinical testing. Figure 4. Active Immunology of 3-day MMI medulloblastoma Figure 4. Active Immunology of 3-day MMI medulloblastoma

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Dual PI3K and mTOR Inhibition in Waldenstrom Macroglobulinemia

Blood ◽

10.1182/blood.v112.11.4986.4986 ◽

2008 ◽

Vol 112 (11) ◽

pp. 4986-4986

Author(s):

Emanuel N. Husu ◽

Aldo M. Roccaro ◽

Antonio Sacco ◽

Molly R. Melhem ◽

Abdel kareem Azab ◽

...

Keyword(s):

Cell Cycle ◽

Bone Marrow ◽

Antitumor Activity ◽

Dna Synthesis ◽

Cell Lines ◽

Parp Cleavage ◽

Dependent Manner ◽

Secreting Cell ◽

Mtor Inhibition ◽

Dose Dependent

Abstract Background. We have previously showed that PI3K/Akt is constitutively active in Waldenström Macroglobulinemia (WM) malignancies, mediating growth, survival, cell cycle regulation, and migration in primary tumor cells. Once activated, Akt phosphorylates downstream targets, including mammalian target of rapamycin (mTOR). Both PI3K/Akt and mTOR represent valid targets for antitumor therapeutic strategies. We therefore evaluated the antitumor activity of NVP-BEZ235 (Novartis, MA) in WM. Methods: WM cell lines (BCWM.1) and IgM secreting cell lines (MEK1, Namalwa) were used. Bone marrow primary CD19+ malignant cells and bone marrow stromal cells (BMSC) were obtained from WM patients. Cytotoxicity, DNA synthesis, and cell cycle were measured using MTT assay, [3H]-thymidine uptake, PI staining/flow cytometry, respectively. Effects of NVP-BEZ235 on cell signaling cascades were determined using immunoblotting and immunofluorescence. Adhesion on fibronectin has been evaluated in WM cells in the presence of NVP-BEZ235. Results: NVP-BEZ235 induced cytotoxicity and inhibited DNA synthesis with an IC50 of 20–25nM in BCWM.1 at 48 hours. Similar effects were demonstrated in all IgM secreting cell lines and in primary CD19+ WM cells, with an IC50 between 20nM and 50nM. No cytotoxicity was observed on peripheral blood mononuclear cells, indicating selective toxicity of the compound on the malignant lymphoplasmacytic clone. We observed that NVP-BEZ235 inhibited Akt (but not ERK phosphorylation) in a dose-dependent manner in BCWM.1 cells at 6 hours. Phosphorylation of GSK3α/β and ribosomal protein-S6, downstream target proteins of Akt, were also markedly inhibited. NVP-BEZ235-inhibited Akt phosphorylation was further confirmed by immunofluorescence. NVP-BEZ235 induced caspase-9, PARP cleavage and increased the release of Smac/DIABLO from the mitochondria to the cytosol, suggesting an induction of apoptosis in a caspase-dependent and –independent manner. We showed that NVP-BEZ235 inhibited adhesion of BCWM.1 cells to fibronectin in a dose-dependent fashion. Lastly, adherence to BMSCs did not confer protection to WM cells against NVP-BEZ235- induced cytotoxicity. Conclusions. These data indicate that NVP-BEZ235 has significant antitumor activity in WM, thus providing the framework for clinical trials in this disease.

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795 APVO603: a dual 4-1BB and OX40 bispecific approach utilizing ADAPTIRTM technology designed to deliver a conditional T cell/NK response against solid tumors

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2021-sitc2021.795 ◽

2021 ◽

Vol 9 (Suppl 3) ◽

pp. A830-A830

Author(s):

Michelle Nelson ◽

Ashly Lucas ◽

Rebecca Gottschalk ◽

Catherine McMahan ◽

Jane Gross ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Solid Tumors ◽

Tumor Cells ◽

Bispecific Antibody ◽

Expression Profiles ◽

T Cell Subsets ◽

Dependent Manner ◽

Dose Dependent

BackgroundAPVO603 is a dual targeting bispecific antibody for 4-1BB (CD137) and OX40 (CD134), engineered with Aptevo's ADAPTIRTM technology. We have previously shown that the distinct characteristics of APVO603 may enable conditional agonism of 4-1BB and OX40 only when cross-linked through engagement of the other receptor via cis and/or trans cellular interactions. Thus, APVO603 is designed with the potential to overcome both the on-target toxicity and limited efficacy observed with 4-1BB and OX40 monoclonal antibody treatment in the clinic.MethodsGenevestigator Software was used to analyze curated transcriptomic data for the expression profiles of OX40 and 4-1BB across select human heme and solid cancer patient sample data sets, as well as, non diseased tissue. Primary inducible Treg (iTreg) cells were sub-optimally stimulated with an anti-CD3/CD28 antibody and cell proliferation was assessed using CFSE-labelled. Cytokines were measured using intracellular flow-based methods. For in vitro tumor lysis studies, activated T cells were co-cultured with Nuclight-labelled tumor cells expressing a tumor-associated antigen (TAA) and activated with TAA x CD3 bispecific protein. Live tumor cells were continually assessed using the Incucyte Live-Cell Analysis System and Cell-By-Cell Software Module.ResultsOX40 and 4-1BB displayed distinct tumor expression profiles, however, several tumor indications were identified with high co-expression and may aid in identifying indications for the clinical development of APVO603. In vitro, APVO603 favored activation of effector T cell subsets and had minimal impact in augmenting iTreg cells proliferation, cytokine production or expression of effector-related molecules, despite the fact that a portion of the iTreg cells expressed OX40 and 4-1BB. The mechanistic activity of APVO603 resulted in dose-dependent control of in vitro tumor growth when paired with a T-cell activating TAA x CD3 bispecific under standard conditions or those leading to T cell exhaustion. In preclinical assays using PBMCs sub-optimally stimulated with TAA x CD3, APVO603 enhanced TAA-expressing tumor cell lysis when compared to TAA x CD3 alone.ConclusionsAPVO603 is a dual-agonistic bispecific antibody that augments the effector function of activated CD4+ and CD8+ T cells and NK cells, but not iTreg cells, in a dose-dependent manner and reduces growth of tumors in vitro and in vivo. Further, mechanistic evaluation supports the ability of APVO603 to pair with T-cell modulating IO approaches to support a more fit T cell response and favorable TME. This preclinical data supports further development of APVO603, a promising immuno-oncology therapeutic with potential for benefit in hematologic and solid tumors.

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A comparison of murine T-cell–depleted adult bone marrow and full-term fetal blood cells in hematopoietic engraftment and immune reconstitution

Blood ◽

10.1182/blood.v99.1.364 ◽

2002 ◽

Vol 99 (1) ◽

pp. 364-371 ◽

Cited By ~ 16

Author(s):

Benny J. Chen ◽

Xiuyu Cui ◽

Gregory D. Sempowski ◽

Maria E. Gooding ◽

Congxiao Liu ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

T Cells ◽

T Cell ◽

Cord Blood ◽

Blood Cells ◽

Immune Reconstitution ◽

Full Term ◽

Fetal Blood ◽

Adult Bone

Umbilical cord blood has been increasingly used as a source of hematopoietic stem cells. A major area of concern for the use of cord blood transplantation is the delay in myeloid and lymphoid recovery. To directly compare myeloid and lymphoid recovery using an animal model of bone marrow and cord blood as sources of stem cells, hematopoietic engraftment and immune recovery were studied following infusion of T-cell–depleted adult bone marrow or full-term fetal blood cells, as a model of cord blood in a murine allogeneic transplantation model (C57BL/6 [H-2b] → BALB/c [H-2d]). Allogeneic full-term fetal blood has poorer radioprotective capacity but greater long-term engraftment potential on a cell-to-cell basis compared with T-cell–depleted bone marrow. Allogeneic full-term fetal blood recipients had decreased absolute numbers of T, B, and dendritic cells compared with bone marrow recipients. Splenic T cells in allogeneic full-term fetal blood recipients proliferated poorly, were unable to generate cytotoxic effectors against third-party alloantigens in vitro, and failed to generate alloantigen-specific cytotoxic antibodies in vivo. In addition, reconstituting T cells in fetal blood recipients had decreased mouse T-cell receptorδ single-joint excision circles compared with bone marrow recipients. At a per-cell level, B cells from fetal blood recipients did not proliferate as well as those found in bone marrow recipients. These results suggest that full-term fetal blood can engraft allogeneic hosts across the major histocompatibility barrier with slower hematopoietic engraftment and impaired immune reconstitution.

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Nonmalignant T cells stimulate growth of T-cell lymphoma cells in the presence of bacterial toxins

Blood ◽

10.1182/blood-2006-04-017863 ◽

2006 ◽

Vol 109 (8) ◽

pp. 3325-3332 ◽

Cited By ~ 38

Author(s):

Anders Woetmann ◽

Paola Lovato ◽

Karsten W. Eriksen ◽

Thorbjørn Krejsgaard ◽

Tord Labuda ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Cell Lines ◽

Mhc Class Ii ◽

Interleukin 2 ◽

Class Ii ◽

Bacterial Toxins ◽

Dependent Manner ◽

Malignant Cells ◽

T Cell Lines

AbstractBacterial toxins including staphylococcal enterotoxins (SEs) have been implicated in the pathogenesis of cutaneous T-cell lymphomas (CTCLs). Here, we investigate SE-mediated interactions between nonmalignant T cells and malignant T-cell lines established from skin and blood of CTCL patients. The malignant CTCL cells express MHC class II molecules that are high-affinity receptors for SE. Although treatment with SE has no direct effect on the growth of the malignant CTCL cells, the SE-treated CTCL cells induce vigorous proliferation of the SE-responsive nonmalignant T cells. In turn, the nonmalignant T cells enhance proliferation of the malignant cells in an SE- and MHC class II–dependent manner. Furthermore, SE and, in addition, alloantigen presentation by malignant CTCL cells to irradiated nonmalignant CD4+ T-cell lines also enhance proliferation of the malignant cells. The growth-promoting effect depends on direct cell-cell contact and soluble factors such as interleukin-2. In conclusion, we demonstrate that SE triggers a bidirectional cross talk between nonmalignant T cells and malignant CTCL cells that promotes growth of the malignant cells. This represents a novel mechanism by which infections with SE-producing bacteria may contribute to pathogenesis of CTCL.

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Dissecting the Immune Cell Progeny of CAR-Expressing Hematopoietic Stem Cells in a Humanized Mouse Model

Blood ◽

10.1182/blood-2019-129746 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 4640-4640

Author(s):

Heng-Yi Liu ◽

Nezia Rahman ◽

Tzu-Ting Chiou ◽

Satiro N. De Oliveira

Keyword(s):

Stem Cells ◽

T Cells ◽

T Cell ◽

Hematopoietic Stem Cells ◽

Cell Lines ◽

Immune Cells ◽

Research Funding ◽

Humanized Mice ◽

Tumor Challenge ◽

Hematopoietic Stem

Background: Chemotherapy-refractory or recurrent B-lineage leukemias and lymphomas yield less than 50% of chance of cure. Therapy with autologous T-cells expressing chimeric antigen receptors (CAR) have led to complete remissions, but the effector cells may not persist, limiting clinical efficacy. Our hypothesis is the modification of hematopoietic stem cells (HSC) with anti-CD19 CAR will lead to persistent generation of multilineage target-specific immune cells, enhancing graft-versus-cancer activity and leading to development of immunological memory. Design/Methods: We generated second-generation CD28- and 4-1BB-costimulated CD19-specific CAR constructs using third-generation lentiviral vectors for modification of human HSC for assessment in vivo in NSG mice engrafted neonatally with human CD34-positive cells. Cells were harvested from bone marrows, spleens, thymus and peripheral blood at different time points for evaluation by flow cytometry and ddPCR for vector copy numbers. Cohorts of mice received tumor challenge with subcutaneous injection of lymphoma cell lines. Results: Gene modification of HSC with CD19-specific CAR did not impair differentiation or proliferation in humanized mice, leading to CAR-expressing cell progeny in myeloid, NK and T-cells. Humanized NSG engrafted with CAR-modified HSC presented similar humanization rates to non-modified HSC, with multilineage CAR-expressing cells present in all tissues with stable levels up to 44 weeks post-transplant. No animals engrafted with CAR-modified HSC presented autoimmunity or inflammation. T-cell populations were identified at higher rates in humanized mice with CAR-modified HSC in comparison to mice engrafted with non-modified HSC. CAR-modified HSC led to development of T-cell effector memory and T-cell central memory phenotypes, confirming the development of long-lasting phenotypes due to directed antigen specificity. Mice engrafted with CAR-modified HSC successfully presented tumor growth inhibition and survival advantage at tumor challenge with lymphoma cell lines, with no difference between both constructs (62.5% survival for CD28-costimulated CAR and 66.6% for 41BB-costimulated CAR). In mice sacrificed due to tumor development, survival post-tumor injection was directly correlated with tumor infiltration by CAR T-cells. Conclusions: CAR modification of human HSC for cancer immunotherapy is feasible and continuously generates CAR-bearing cells in multiple lineages of immune cells. Targeting of different malignancies can be achieved by adjusting target specificity, and this approach can augment the anti-lymphoma activity in autologous HSC recipients. It bears decreased morbidity and mortality and offers alternative therapeutic approach for patients with no available sources for allogeneic transplantation, benefiting ethnic minorities. Disclosures De Oliveira: National Institute for Health Research Biomedical Research Centre at Great Ormond Street Hospital for Children NHS Foundation Trust and University College London: Research Funding; NIAID, NHI: Research Funding; Medical Research Council: Research Funding; CIRM: Research Funding; National Gene Vector Repository: Research Funding.

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Receptor-specific inhibition of bone marrow erythropoiesis by recombinant DNA-derived interleukin-2

Blood ◽

10.1182/blood.v69.5.1368.1368 ◽

1987 ◽

Vol 69 (5) ◽

pp. 1368-1375 ◽

Cited By ~ 19

Author(s):

SE Burdach ◽

LJ Levitt

Keyword(s):

Bone Marrow ◽

T Cells ◽

T Cell ◽

Recombinant Dna ◽

Interleukin 2 ◽

Target Cells ◽

Erythroid Progenitor ◽

Dependent Inhibition ◽

Dose Dependent Inhibition ◽

Dose Dependent

Abstract Interleukin-2 (IL-2) induces differential secretion of lymphokines by IL-2 receptor (IL-2R)-positive and IL-2R-negative T cells. We studied T cell IL-2R-specific modulation of adult bone marrow erythropoiesis by recombinant IL-2 (rIL-2). I3–2R were induced by CD3 T cell surface determinant-triggering and analyzed by cytofluorography. Bone marrow monocyte and T cell-depleted (NAB-T) target cells were assessed for early erythroid progenitor expression (BFU-E) in the presence of 0 to 10(3) U/mL of rIL-2, rIL-2 had no significant effect on BFU-E expression in the absence of T cells or in the presence of IL-2R- negative T cells. rIL-2 caused a dose-dependent inhibition (75% to 90%) of BFU-E in the presence of autologous IL-2R-positive T cells. The addition of anti-IL2-receptor antibody to cultures containing rIL-2 plus IL-2R-positive T cells entirely abrogated rIL-2-mediated inhibition of BFU-E. In the presence of rIL-2 (10(2) U/mL) production of interferon gamma (IF-gamma) by adult marrow CD3-triggered IL-2R- positive T cells was increased 37- to 125-fold compared to IL-2R- negative T cells. rIF-gamma caused a dose-dependent (88% +/- 17% at 10(3) U/mL) inhibition of adult BFU-E in the presence of CD3-triggered autologous T cells. rIL2-mediated inhibition of adult BFU-E in the presence of IL-2R-positive T cells was partially abrogated (52% +/- 16%) following addition of monospecific IF-gamma antibody. These results demonstrate (a) rIL-2 modulation of adult marrow erythropoiesis is selectively dependent upon both the presence or absence of autologous T cells and the IL-2R status of these T cells; and (b) rIL-2- induced inhibition of adult marrow erythropoiesis is mediated in part by release of IF-gamma from IL-2R-positive T cells.

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Adoptive T-Cell Therapy for B-Cell Acute Lymphoblastic Leukemia: Preclinical Studies

Blood ◽

10.1182/blood.v94.10.3531.422k14_3531_3540 ◽

1999 ◽

Vol 94 (10) ◽

pp. 3531-3540 ◽

Cited By ~ 22

Author(s):

Angelo A. Cardoso ◽

J. Pedro Veiga ◽

Paolo Ghia ◽

Hernani M. Afonso ◽

W. Nicholas Haining ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

T Cell ◽

B Cell ◽

Cell Lines ◽

Lymphoblastic Leukemia ◽

Adoptive T Cell Therapy ◽

Leukemic Cells ◽

Acute Leukemias ◽

T Cell Lines

We have previously shown that leukemia-specific cytotoxic T cells (CTL) can be generated from the bone marrow of most patients with B-cell precursor acute leukemias. If these antileukemia CTL are to be used for adoptive immunotherapy, they must have the capability to circulate, migrate through endothelium, home to the bone marrow, and, most importantly, lyse the leukemic cells in a leukemia-permissive bone marrow microenvironment. We demonstrate here that such antileukemia T-cell lines are overwhelmingly CD8+ and exhibit an activated phenotype. Using a transendothelial chemotaxis assay with human endothelial cells, we observed that these T cells can be recruited and transmigrate through vascular and bone marrow endothelium and that these transmigrated cells preserve their capacity to lyse leukemic cells. Additionally, these antileukemia T-cell lines are capable of adhering to autologous stromal cell layers. Finally, autologous antileukemia CTL specifically lyse leukemic cells even in the presence of autologous marrow stroma. Importantly, these antileukemia T-cell lines do not lyse autologous stromal cells. Thus, the capacity to generate anti–leukemia-specific T-cell lines coupled with the present findings that such cells can migrate, adhere, and function in the presence of the marrow microenvironment enable the development of clinical studies of adoptive transfer of antileukemia CTL for the treatment of ALL.

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Immunosupressive Property of Mesenchymal Stem Cells (MSCs) in Mixed Lymphocyte Reactions In Vitro.

Blood ◽

10.1182/blood.v106.11.4313.4313 ◽

2005 ◽

Vol 106 (11) ◽

pp. 4313-4313 ◽

Cited By ~ 1

Author(s):

Jianyong Li ◽

Lijuan Meng ◽

Yu Zhu ◽

Hua Lu ◽

Changgeng Ruan

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

T Cells ◽

T Cell ◽

T Cell Subsets ◽

Bone Marrow Mscs ◽

Human Bone Marrow Mscs ◽

Mixed Lymphocyte Reactions ◽

Activation Phase

Abstract Meesnchymal stem cells (MSCs) were successfully used in the prevention and treatment of graft versus host disease (GVHD) after allogeneic hematopoietic stem cell transplantation. To further explore the immunosuppressive property of human bone marrow (MSCs) in alloantigen-induced mixed lymphocyte reactions (MLRs) in vitro, human bone marrow MSCs and lymphocytes were prepared from healthy volunteers. MSCs were expanded in vitro in Mesencult serum free media. MSCs were cocultured with one-way MLRs and bidirectional MLRs, responder cells were labeled with carboxyfluorescein diacetate- succinimidyl ester (CFSE) in bidirectional MLRs. Cell Counting Kit-8(CCK-8)kit was used in cell proliferation detection, T-cell subsets were analyzed by flow cytometry (FCM). The results showed that MSCs were positive for CD105, CD73, CD13, CD90 and were negative for hematopoietic cell markers. In one-way MLRs, MSCs down-regulated alloantigen-induced lymphocyte expansion in a dose-dependent and MHC-independend manner. In two-way MLRs, MSCs suppressed proliferation of CFSE positive cells. T cell subsets were changed: Th2 and Tc2 were down-regulated. Th2 was reduced from 1.70% to 0.65%, and Tc2 reduced from 1.10% to 0.47%, while Th1 and Tc1 were unaffected. T cells that became CD69+, which was an early activation marker, were significantly up-regulated from 7.14% to 26.12% and CD4+CD25+T regulatory cells (CD4+CD25+Tr) were up-regulated from 4.04% to 6.19%, which indicating that suppression did not interfere with activation phase of T cells and might be mediated by CD4+CD25+Tr partly. We conclede that MSCs down-regulated alloantigen-induced lymphocyte expansion. The immunosupressive effect might involve in post-activation phase of T cells. CD4+CD25+Tr might contribute to the suppressory activity of MSCs.

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