New Interleukin-15 Superagonist (IL-15 SA) Significantly Enhances Graft-Versus-Tumor Activity After Allogeneic Hematopoietic Stem Cell Transplantation

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3254-3254
Author(s):  
Cavan P Bailey ◽  
Christopher Sauter ◽  
Michelle M Panis ◽  
Tulin Budak-Alpdogan ◽  
Hing Wong ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Cd8 T Cells ◽  
T Cell Proliferation ◽  
Hematopoietic Stem ◽  
Interleukin 15 ◽  
And Function

Abstract Interleukin-15 (IL-15) is a pleiotropic cytokine, which plays various roles in the innate and adaptive immune system, including the development, activation, homing and survival of immune effector cells. IL-15 has been previously shown to increase CD8+ T and NK cells number and function in normal mice and recipients of stem cell transplantation. However, obstacles remain in using IL-15 therapeutically, specifically its low potency and short in vivo half-life. To overcome this, a new IL-15 mutant (IL-15N72D, J. Immunol, 2009; 183:3598) has been developed, with increased biological activity. Co-expressing IL-15N72D, in conjunction with IL-15RαSu/Fc produced a biologically active and highly potent IL-15 superagonist complex (IL-15SA, also known as ALT-803, Cytokine, 2011; 56:804). We evaluated the effects of IL-15-SA on immune reconstitution and graft-versus-tumor (GVT) activity in recipients of allogeneic hematopoietic stem cell transplantation (HSCT). Lethally irradiated BALB/c recipients were transplanted with T-cell depleted (TCD) bone marrow (BM) cells from B6 mice. IL-15 SA was administered via IP injection in two doses on days +17 and +24 after transplant. Animals were sacrificed at day 28. Administration of IL-15 significantly increased the numbers of CD8+ T cells and NK cells. IL-15 SA also augmented interferon-γ secretion from CD8+ T cells. We observed similar activity in B6CBA→CB6F1 transplant model. Interestingly IL-15 SA upregulates NKG2D and CD107a expression on CD8+ T cells. IL-15 SA administration also specifically increased slow-proliferative CD8+ T-cell proliferation in conjunction with robust IFN-γ and TNF-α secretion in CD8+ T cells in recipients of CFSE (carboxyfluorescein succinimidyl ester) labeled-T-cell infusion, whereas there was no effect on CD4+ T-cell proliferation. We then tested the anti-tumor activity of IL-15 SA in three different tumor models; murine mastocytoma (P815), murine B cell lymphoma (A20) and murine renal cell carcinoma (Renca). We found that IL-15 SA administration enhanced GVT activity against P815 and A20 in recipients of allogeneic HSCT though this activity required a low-dose T cell infusion with HSCT. Interestingly, augmented GVT activity against to Renca after IL-15 SA administration in recipients of allogeneic HSCT did not require T cell infusion. We conclude that IL-15 SA is a very potent cytokine complex for enhancing CD8+ and NK cell reconstitution and function after HSCT, which would be a candidate for post-transplant immunotherapy. Disclosures: Wong: Altor Bioscience: Employment.

scholarly journals Immunosenescent Characteristics of T Cells in Young Patients Following Haploidentical Stem Cell Transplantation from Parental Donors

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3283-3283
Author(s):  
Ga Hye Lee ◽  
Kyung Taek Hong ◽  
Jung Yoon Choi ◽  
Hee Young Shin ◽  
Won-Woo Lee ◽  
...  
Keyword(s):  
Dna Damage ◽  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Hematopoietic Stem Cell Transplantation ◽  
Stem Cell Transplantation ◽  
Telomere Length ◽  
Cell Transplantation ◽  
Cd8 T Cells ◽  
Hematopoietic Stem

Introduction: Pediatric and adolescent patients in need of allogeneic hematopoietic stem cell transplantation generally receive stem cells from older, unrelated or parental donors when a sibling donor is not available. Despite encouraging clinical outcomes, it has been suggested that immune reconstitution accompanied by increased replicative stress and a large difference between donor and recipient age may worsen immunosenescence in pediatric recipients. Therefore, in this study paired samples were collected at the same time from donors and recipients of haploidentical hematopoietic stem cell transplantation (HaploSCT). Methods: We conducted flow cytometry-based phenotypic and functional analyses and telomere length measurements of 21 paired T-cell sets from parental donors and children who received T cell-replete HaploSCT with post-transplant cyclophosphamide (PTCy) at Seoul National University Children's Hospital between February 2014 and January 2017. The conditioning regimen was comprised of targeted busulfan (total target area under the curve, 75,000 mg•h/L) with intensive pharmacokinetic monitoring, fludarabine and cyclophosphamide. Results: Fourteen pediatric, adolescent, and young adult patients with malignant disease and seven with nonmalignant disease were included with a median post-transplantation period of 16.9 months (range, 12.4-38.8). Senescent T cells, CD28- or CD57+ subsets of both CD4+ and CD8+ T cells, were significantly expanded in patients compared with parental donors. Further, not only CD4+CD28- T cells, but also CD4+CD28+ T cells showed reduced cytokine production capacity and impaired polyfunctionality compared with parental donors, whereas their TCR mediated proliferation capacity was comparable. Of note, the telomere length in patient T cells was preserved, or even slightly longer, in senescent T cells compared with donor cells. We also found that the patients had a higher level of γ-H2AX-expressing CD28- senescent T cells compared with the donors, which is used as a DNA damage marker. Regression analysis showed that senescent features of CD4+ and CD8+ T cells in patients were influenced by donor age and the frequency of CD28- cells, respectively. Conclusions: Our data suggest that T cells undergo premature immunosenescent changes and exhibit functional defects in pediatric HaploSCT recipients. Further, there is an increased level of DNA damage in patient CD4+ T cells compared to those of parental donors. Therefore, long-term, comprehensive immune monitoring of these patients is necessary. Disclosures No relevant conflicts of interest to declare.

Development of Novel Stem Cell Transplantation and Gene-Immunotherapy Using WT1-Specific T-Cell Receptor Gene.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3028-3028
Author(s):  
Toshiki Ochi ◽  
Hiroshi Fujiwara ◽  
Kozo Nagai ◽  
Toshiaki Shirakata ◽  
Kiyotaka Kuzushima ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Hematopoietic Stem Cells ◽  
Cell Transplantation ◽  
Clinical Efficacy ◽  
Cd8 T Cells ◽  
Hematopoietic Stem

Abstract Abstract 3028 Poster Board II-1004 Purpose The Wilms' tumor 1 (WT1) is one of the zinc-finger transcriptional regulators, and its expression level is very low in most tissues of adults. In contrast, various kinds of leukemia and solid tumors express WT1 abundantly, and high expression level of WT1 is correlated with disease aggressiveness and poor prognosis. These findings indicate that WT1 is a promising target antigen for anti-cancer cellular immunotherapy. Following identification of immunogenic epitopes derived from WT1 which are recognized by HLA class I-restricted and HLA class II-restricted T cells, phase I/II WT1 peptide vaccination trials have been conducted. Although the positive correlation between the clinical efficacy and vaccine-induced WT1-specific T-cell response has been reported, the clinical efficacy is not satisfactory. Adoptive transfer of WT1-specific T cells seems to be the promising approach to achieve marked improvement in clinical efficacy of WT1-targeting immunotherapy, however, it still remains difficult to expand WT1-specific T cells sufficiently ex vivo. To overcome these problems, we attempted to establish gene-immunotherapy targeting WT1 using T-cell receptor (TCR) gene isolated from the WT1-specific T-cell clone. We also verified the feasibility of novel stem cell transplantation by transducing WT1-specific TCR gene into hematopoietic stem cells. Methods We cloned the full length TCR-αa and -β genes from a WT1235-243-specific and HLA-A*2402-restricted cytotoxic T lymphocyte (CTL) clone. The WT1-specific TCR gene-repressing retroviral and lentiviral vectors were constructed. Retroviral vector was transduced to human peripheral T cells in retronectin-coated plate. WT1-specific functions of TCR gene-transduced CD8+ T cells and CD4+ T cells were examined by evaluating WT1 peptide-specific cytotoxicity by 51Cr-release assay and WT1 peptide-specific Th1 cytokine production, respectively. To improve the efficacy of WT1-specific TCR expression, we developed the novel retroviral vector which can inhibit selectively intrinsic TCR expression (si-TCR vector). Finally, we transduced the WT1-specific TCR lentiviral vector into human cord blood CD34+ cells, and transplanted them to NOD/SCID/common-γnull mice. Then, we examined whether WT1-specific human mature T cells can differentiate in mice. The presence of WT1-specific human T cells in mice was determined by tetramer assay and IFN-γ production in response to stimulation with WT1 peptide. Results Following transfer of WT1-specific TCR gene into peripheral blood lymphocytes, WT1 peptide-specific CD8+ and CD4+ T cells could be expanded easily in vitro. TCR gene-transduced CD8+ T cells exerted cytotoxicity against WT1 peptide-pulsed target cells and human leukemia cells in an HLA-A*2402-restricted manner. Similarly, TCR gene-transduced CD4+ T cells showed WT1-specific Th1 cytokine production in response to stimulation with human leukemia cells in HLA-A*2402-restricted fashion depending on the interaction of CD4 and HLA class II molecules. The newly developed si-TCR vector appeared to inhibit expression of endogenous TCR efficiently and improved the efficacy of WT1-specific TCR expression 3 to 5-fold higher as compared to the conventional vector. Three months after transplantation of WT1-specific TCR gene-transduced human hematopoietic stem cells in NOD/SCID/common-γnull mice, differentiation of WT1-specific human T cells in murine spleen was evaluated. Tetramer assay revealed that human mature T cells expressing WT1-specific TCR on their cell surface were clearly detected. Furthermore, these WT1-specific CD8+ T cells appeared to produce IFN-γ in response to stimulation with WT1 peptide-loaded HLA-A*2402-positive cells. Conclusion The adoptive gene-immunotherpay using WT1-specific TCR gene against leukemia seems to be promising. Moreover, the novel stem cell transplantation using WT1-specific TCR gene-transduced hematopoietic stem cells might open the door to induce long-lasting anti-leukemic cellular immunity in patients with leukemia. Disclosures No relevant conflicts of interest to declare.

scholarly journals New Interleukin-15 Superagonist (IL-15 SA) Combined with Donor Lymphocyte Infusion (DLI) Significantly Enhances Graft-Versus-Tumor Activity after Allogeneic Hematopoietic Stem Cell Transplantation

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1118-1118
Author(s):  
Cavan Bailey ◽  
Michelle M Panis ◽  
Cihangir Buyukgoz ◽  
Tulin Budak-Alpdogan ◽  
Neal Flomenberg ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Cd8 T Cells ◽  
Tumor Burden ◽  
Donor Lymphocyte Infusion ◽  
Hematopoietic Stem

Abstract Donor lymphocyte infusion (DLI) has been successfully used clinically to augment the graft-versus-tumor (GVT) effect following hematopoietic stem cell transplantation (HSCT) in relapsed patients. However, improvements can still be made in enhancing anti-tumor activity, reducing graft-versus-host disease (GVHD) and decreasing complications from opportunistic infections. Our studies present clear evidence of increased tumor clearance via cytokine therapy in combination with DLI as a way to “boost” the infused cells function. Interleukin-15 (IL-15) is a potent cytokine that increases CD8+ T and NK cells number and function in normal mice and recipients of stem cell transplantation. Despite this, obstacles remain for use of IL-15 therapeutically, specifically its low potency and short in vivo half-life. To overcome this, a new IL-15 superagonist (IL-15 SA-(ALT-803)) has been developed with a longer half-life and increased potency. Administration of IL-15 SA to recipients of CFSE labeled T cells increases proliferation of CD8+T cells and IFN-γ and TNF-α secretion from CD8+T cells. We developed a murine DLI model by titrating the dose of infused T cells in a parent-F1 model, and then combined IL-15 SA administration with DLI in murine recipients of allogeneic HSCT. In this model, lethally irradiated CB6F1 (H2Kb/d) mice were transplanted with T- cell depleted bone marrow cells from C57BL6 mice (H2Kb). All recipients of HSCT were also co-injected A20 B-cell lymphoma cells transfected with a luciferase-producing gene, which allows bioluminescent imaging and tracking of tumor progress in vivo. Mice receiving DLI (2.5 X 105 T cells) with IL-15 SA injections given at 1μg/mouse on days 17 and 24 post-BMT show less tumor burden and increased overall survival (p = 0.04) and decreased tumor growth (p = 0.02) (Figure 1). The IL-15 SA treated group had a significantly less weight loss than the control group (p = 0.007). No GVHD symptoms were noted via weekly clinical scoring, highlighting both the efficacy and overall safety of the IL-15 SA therapy. Furthermore, we evaluated T- cell exhaustion markers on CD8+ T cells in surviving mice. We found increased programmed death-1 (PD-1) expression on T cells even when the tumor burden is cleared. Treatment with IL-15 SA reduced PD-1 expression on donor CD8+ T-cells in mice surviving more than 120 days post-transplant. We conclude that IL-15 SA enhances CD8+ T cell function by increasing cytokine secretion and proliferation of T cells whereas could also prevent T cell exhaustion. We suggest that IL-15 SA is a long-waited lymphoid growth factor and has the potential to use in combination with DLI for the treatment of recurrent disease after HSCT. Disclosures No relevant conflicts of interest to declare.

Generation of Human T/NK Cell Precursors for Adoptive Transfer To Enhance Immune Reconstitution in Allogeneic Hematopoietic Stem Cell Transplantation Recipients.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3209-3209
Author(s):  
Sonali Chaudhury ◽  
Johannes Zakarzewski ◽  
Jae-Hung Shieh ◽  
Marcel van der Brink ◽  
Malcolm A.S. Moore
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Hematopoietic Stem Cell Transplantation ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Hematopoietic Stem ◽  
Serum Free ◽  
Cd34 Cells

Abstract Allogeneic hematopoietic stem cell transplantation (HSCT) is associated with significant post-transplant immunoincompetence which affects in particular the T cell lineage and results in an increased susceptibility to infections. Novel strategies to enhance immune recovery after HSCT could prevent malignant relapse and immune deficiency and improve the overall outcome of this therapy. We have established a serum free culture system using murine bone marrow stroma expressing the Notch ligand Delta-like 1 (DL1) to obtain high numbers of human pre-T cells from CD34+ cells. Human cord blood CD34+ cells were plated on OP9 DL1 stroma transduced with adenovirus expressing thrombopoietin (ad-TPO) at an MOI of 30. Media used was QBSF-60 (Serum free media prepared by Quantity Biologicals) supplemented with Flt-3 ligand and IL-7 (10ng/ml). At 4–5 weeks we obtained a 10 5–10 7 fold expansions of cultured cells of which about 70–80% were CD5, CD7 positive pre T cells (Fig 1). We then developed an optimal system to study human lymphohematopoiesis using mouse models (NOD/SCID/IL2rϒnull and NOD/SCIDβ2null) and established an adequate pre T cell number (4 × 10 6) and radiation dose (300 Rads). We injected CD34 and pre-T cells (CD45 +, CD4−, CD5+, CD7+) derived from OP9 DL1 cultures into these mice and achieved ~50%engraftment of NK in the bone marrow and spleen of the mice at 2 weeks following transplant. The thymus from the same mice showed evidence of about 12–15% CD7+ pre T cells. We are currently studying the function of the generated NK and T cells both in vivo and in vitro studies. Figure Figure

CTL Responses Against Tumor Associated Antigens Are Part of the Graft Versus Leukemia-Effect and Protect from Relapse after Allogeneic Hematopoetic Stem Cell Transplantation.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3231-3231
Author(s):  
Markus Kapp ◽  
Stefan Stevanovic ◽  
Kerstin Fick ◽  
Juergen Loeffler ◽  
Sen Mui Tan ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Cd8 T Cells ◽  
T Cell Responses ◽  
Proteinase 3 ◽  
Post Transplantation ◽  
Cell Responses

Abstract The Graft-versus-Leukemia (GVL) effect following allogeneic hematopoetic stem cell transplantation (HSCT) is one of the most prominent examples showing the ability of the immune system to eliminate malignant diseases. This effect was a strictly clinically described phenomenon, but in the last years T-cell responses against tumor-associated antigens (TAA) could partly be set in correlation with clinical benefit. Previously, TAA such as WT1 and proteinase-3 have been proposed as the targets for T-cells to establish a GVL effect. Now, we examined in addition other TAA (MUC1 and HM1.24) as possible T-cell targets of GVL related immune responses. We have defined new peptide epitopes from the MUC1 and HM1.24 antigens by the reverse immunology approach to increase the number of patients who can be screened and to expand the repertoire of immunologic monitoring as well as therapeutic approaches. A total of 25 patients after allogeneic stem cell transplantation have been screened and we are able to detect T-cell responses to both the MUC1 and HM1.24 antigens on top of the WT1 and the proteinase-3 antigen. Interestingly, we could detect a significant relationship between relapse and the absence of a T-cell response to TAA: Only 1/10 patients (10%) with TAA-specific CTL relapsed in contrast to 8/15 patients (53.3%) without TAA-specific CTL responses (p < 0.05). Furthermore, we demonstrated MUC1 peptides presented by HLA A*6801, B*0702 and B*4402 to be specifically recognized by CD3+/CD8+ T-cells. In conclusion, CD8+ T-cell responses directed to TAA might contribute to the GVL effect and are not limited to WT1 and proteinase-3. These observations clearly highlight both the importance and the potential of immunotherapeutic approaches in allogeneic stem cell recipients. Figure 1: New defined HLA class I epitopes predicted by computer analysis are recognized by specific CTL in patients post allogeneic HSCT. IFN-γ staining of PBMC from, patient No. 17 (AML, CR), 672 days post transplantation (A), patient No. 8 (AML, CR), 1035 days post transplantation (B) Cells were stimulated with 10μg/ml of the indicated peptides. Gates were set on lymphocytes by forward/side scattering (R1) and on CD3+/CD8+ cells (R2). Percentage numbers show peptide-specific CD3+/CD8+ T-cells from all CD3+/CD8+ T-cells. Figure 1:. New defined HLA class I epitopes predicted by computer analysis are recognized by specific CTL in patients post allogeneic HSCT. . / IFN-γ staining of PBMC from, patient No. 17 (AML, CR), 672 days post transplantation (A), patient No. 8 (AML, CR), 1035 days post transplantation (B) Cells were stimulated with 10μg/ml of the indicated peptides. Gates were set on lymphocytes by forward/side scattering (R1) and on CD3+/CD8+ cells (R2). Percentage numbers show peptide-specific CD3+/CD8+ T-cells from all CD3+/CD8+ T-cells.

Exhaustion of CMV Specific T Cells with Enhanced PD-1 Expression In Persistent Cytomegalovirus Infection After Allogeneic Stem Cell Transplantation

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3912-3912 ◽  
Author(s):  
Tomonori Kato ◽  
Tetsuya Nishida ◽  
Miho Murase ◽  
Makoto Murata ◽  
Tomoki Naoe
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Cd8 T Cells ◽  
Culture Media ◽  
Cmv Infection ◽  
Control Serum ◽  
And Control

Abstract Abstract 3912 Cytomegalovirus (CMV) is one of the most common pathogens causing morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HSCT), despite preemptive treatments employing antiviral drugs. Cytotoxic T cells are indispensable to control CMV infections. Chronic viral infections with human immunodeficiency virus or hepatitis C virus were shown to be associated with exhausted T cells with high expression of the inhibitory molecule programmed death 1 (PD-1). Recently, it has been reported that PD-1 up-regulation on CMV specific T cells was associated with CMV infection after renal and liver transplantation. PD-1 expression on CMV specific T cells after HSCT has not been well examined. We evaluated the involvements of exhausted CMV specific T cells characterized by high PD-1 expression in persistent CMV infection after allogeneic HSCT. Peripheral blood mononuclear cells (PBMC) and serum were obtained from an HLA-A*2402-positive patient who had received bone marrow transplantation from an HLA-A, B, C and DR matched unrelated donor. This patient failed to eliminate CMV for more than one year after transplantation despite intermittent administration of ganciclovir and foscarnet. Control PBMC and serum were obtained from an HLA-A*2402-positive healthy volunteer because the Japan Marrow Donor Program prohibits blood collection for research use from donor. All blood was collected with written informed consent. We at first analyzed frequencies of CMV-specific CD8+ T cells in patient and control PBMC by flow cytometer using QYDPVAALF/A*2402-specific tetramer and CD8 antibodies. QYDPVAALF is derived from CMV pp65 protein and presented by the HLA-A*2402 molecule. Tetramer stained cells were detected in the patient PBMC but control PBMC (0.11% versus undetectable). Patient and control PBMC were stimulated by a synthetic peptide QYDPVAALF in culture media containing IL-2 for 14 days, and stained with QYD/A*2402-specific tetramer. Remarkably, post-stimulated patient PBMC contained only 0.54% of tetramer stained CD8+ T cells, whereas a more dramatic increase (14.1%) in control PBMC. We analyzed frequencies of IFN-g secreting CD8+ T cells in PBMC after stimulation with a peptide pool covering the whole CMV pp65 protein for 4 hours. Less patient CD8+ T cells produced IFN-g, compared with the control CD8+ T cells (0.5% versus 1.1%) These data demonstrate dysfunction of CMV-specific CD8+ T cells in the patient with persistent CMV infection. To examine the mechanism of dysfunction of CMV-specific CD8+ T cells, we analyzed the expression of PD-1 on CMV-specific CD8+ T cells 14 days after stimulation with QYDPVAALF peptide. Multiparameter flow cytometry and tetramer assay exhibited higher expression of PD-1 on CMV-specific CD8+ T cells generated from patient PBMC, compared with CMV-specific CD8+ T cells generated from control PBMC. To find out whether the engagement of PD-1 to its ligand (PD-L1) leads to T cell exhaustion, we stimulated patient PBMC with QYDPVAALF peptide for 14 days in the presence or absence of anti PD-L1 antibody which blocks PD-1/PD-L1 inhibitory pathway. Blockade of PD-1/PD-L1 pathway resulted in 3.9-fold increase in patient CMV specific T cells. These findings demonstrate that PD-1 is associated with the exhaustion of CMV specific CD8+ T cells during persistent CMV infection in this patient. To examine the effect of patient serum on CMV specific CD8+ T cells, we stimulated patient PBMC with QYDPVAALF peptide for 14 days in culture media with patient or control serum. CMV specific CD8+ T cells increased 4-fold and 55-fold in the presence of patient and control serum, respectively. Patient serum led to higher PD-1 expression on CMV specific CD8+ T cells, compared with control serum (Fig). These findings suggest that patient serum may contain what regulates PD-1 expression level of exhausted T cells. Further investigations to identify factors regulating PD-1 expression in patient serum are in progress. The identification of the factors may provide new strategies to improve exhausted T cell function. Disclosures: No relevant conflicts of interest to declare.

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