scholarly journals Dynamic Clonal Hematopoiesis and Functional T-cell Immunity in a Super-centenarian

2019 ◽  
Author(s):  
Erik B. van den Akker ◽  
Stavros Makrodimitris ◽  
Marc Hulsman ◽  
Martijn H. Brugman ◽  
Tanja Nikolic ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Immunity ◽  
List Type ◽  
Hematopoietic Stem ◽  
Blood Samples ◽  
Clonal Hematopoiesis ◽  
Cell Immunity ◽  
Increased Risk

AbstractThe aged hematopoietic system is characterized by decreased immuno-competence and by a reduced number of hematopoietic stem cells (HSCs) that actively generates new blood cell (age-related clonal hematopoiesis, ARCH). While both aspects are commonly associated with an increased risk of aging-related diseases, it is currently unknown to what extent these aspects co-occur during exceptional longevity. Here, we investigated these aspects in blood cells of an immuno-hematopoietically normal female who reached 111 years. Blood samples were collected across a 9-year period at ages 103, 110 and 111 years. We applied several genetic sequencing approaches to investigate clonality in peripheral blood samples and sorted cell subsets. Immuno-competence was characterized using flow cytometry, T-cell receptor excision circle (TREC) assays, and in vitro proliferation assays. We identified a single DNMT3A-mutated HSC clone with a complex subclonal architecture and observed ongoing subclonal dynamics within the 9-year timeframe of our sampling. The mutated HSC generated 78-87% myeloid cells, 6-7% of the B-cells, 6% of CD8+ T-cells, and notably 22% of the CD4+ T-cells. Intriguingly, we found that T-cells were capable of robust proliferation when challenged in vitro. Moreover, we observed a surprisingly high TREC content, indicative of recent generation of naive T-cells. Concluding, we observed long-term stability of extreme ARCH with ongoing clonal dynamics combined with functional T-cell immunity. Our results indicate that extreme ARCH does not compromise immuno-competence and that a clonally expanded CD4+ T-cell subset may serve as a potential hallmark of the supercentenarian immune system.Key pointsLongitudinal blood sampling from a female aged 103-111 revealed a dynamic clonal hematopoiesis contributing to myeloid and lymphoid subsetsDespite the highly advanced age and extreme clonal hematopoiesis we observed functional T-cell immunity

Berkeley Sickle Mice Demonstrate CD8- and NK1.1-Dependent Increased Rejection of Allogeneic Hematopoietic Stem Cell Transplants.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2332-2332
Author(s):  
Leslie Kean ◽  
Kelly Hamby ◽  
Jennifer Perry ◽  
Christian Larsen ◽  
David Archerq
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Sickle Cell Disease ◽  
Hematopoietic Stem Cell ◽  
Sickle Cell ◽  
Cell Disease ◽  
Hematopoietic Stem ◽  
Cell Immunity ◽  
Increased Risk

Abstract While hematopoietic stem cell transplantation (HSCT) represents the only curative therapy for sickle cell disease, sickle patients undergoing HSCT face many complications, including an increased risk of graft rejection compared to non-sickle patients. We have used the Berkeley sickle mouse model to study the potential mechanisms underlying this increased risk of rejection. Using a CD28/CD40 costimulation-blockade-based non-myeloablative HSCT regimen, we transplanted Berkeley sickle mice with fully allogeneic SJL bone marrow. While the vast majority (>85%, n=25) of control C57BL/6 animals became stably chimeric and immunologically donor-tolerant with this transplant regimen, sickle mice were much more prone to reject the transplant (~20% graft acceptance, n=25). Both CD8+ cells and NK1.1+ cells were found to contribute to this rejection, as depletion of either of these cell populations led to a marked increase in the percent of engrafted mice (>85% graft acceptance, n=15–25), while depletion of CD4+ cells led to the opposite effect, with 0% (n=25) animals engrafted in this depletion cohort. The increased propensity of HSCT rejection in the Berkeley sickle mice may, in part, be explained by the presence of increased numbers of donor-reactive T cells (5–10-fold compared to C57BL/6 controls) in naïve sickle mice, despite their lack of exposure to donor antigens, and their housing in a Specific-Pathogen-Free environment. We speculate that these increased numbers of anti-donor T cells may occur as a result of heightened inflammation in the context of active sickle cell disease, which could lead to increased expansion and persistence of a T cell repertoire containing anti-donor heterologous T cell immunity. This heterologous immunity may have a profound effect on the success of HSCT for sickle cell disease, especially when non-myeloablative regimens are employed.

Dendritic Cell-Induced T Cell Non-Responsiveness Is Mediated by FOXP3+ and TGF-beta+IL-10+ CD4+ T Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3891-3891
Author(s):  
Zwi N. Berneman ◽  
Nathalie Cools ◽  
Viggo F.I. Van Tendeloo ◽  
Marc Lenjou ◽  
Griet Nijs ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
T Cell Immunity ◽  
Tgf Beta ◽  
Cell Immunity

Abstract Dendritic cells (DC), the professional antigen presenting cells of the immune system, exert important functions both in induction of T cell immunity as well as of tolerance. Previously, it was accepted that the main function of immature DC (iDC) in their in vivo steady state condition is to maintain peripheral tolerance to self-antigens and that these iDC mature upon encounter of so-called danger signals and subsequently promote T cell immunity. However, a growing body of experimental evidence now indicates that traditional DC maturation can no longer be used to distinguish between tolerogenic and immunogenic properties of DC. In this study, we compared the in vitro stimulatory capacity of immature DC (iDC), cytokine cocktail-matured DC (CC-mDC) and poly I:C-matured DC (pIC-mDC) in the absence and presence of antigen. All investigated DC types could induce at least 2 subsets of regulatory T cells. We observed a significant increase in both the number of functionally suppressive transforming growth factor (TGF)-beta+ interleukin (IL)-10+ T cells as well as of CD4+CD25+FOXP3+ T cells within DC/T cell co-cultures as compared to T cell cultures without DC. The induction of these regulatory T cells correlates with in vitro T cell non-responsiveness after co-culture with iDC and CC-mDC, while stimulation with pIC-mDC resulted in reproducible cytomegalovirus pp65 or influenza M1 matrix peptide-specific T cell activation as compared to control cultures in the absence of DC. In addition, the T cell non-responsiveness after stimulation with iDC was shown to be mediated by TGF-beta and IL-10. Moreover, the suppressive capacity of CD4+ T cells activated by iDC and CC-mDC was shown to be transferable when these CD4+ T cells were added to an established T cell response. In contrast, addition of CD4+ T cells stimulated by pIC-mDC made responder T cells refractory to their suppressive activity. In conclusion, we hypothesize that DC have a complementary role in inducing both regulatory T cells and effector T cells, where the final result of antigen-specific T cell activation will depend on the activation state of the DC. This emphasizes the need for proper DC activation when T cell immunity is the desired effect, especially when used in clinical trials.

The Molecular Characteristics in CDR3 of TCR Vα and Vβ Genes Associated with cGVHD in Patients after Allogeneic Hematopoietic Stem Cell Transplantation.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3247-3247
Author(s):  
Xiuli Wu ◽  
Yangqiu Li ◽  
Kanger Zhu ◽  
Xin Du ◽  
Shaohua Chen ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
T Cell Immunity ◽  
Molecular Characteristics ◽  
Hematopoietic Stem ◽  
T Cell Clones ◽  
New Name ◽  
Cell Immunity ◽  
Cell Clones

Abstract Successful allogeneic hematopoietic stem cell transplantation (allo-HSCT) requires reconstitution of normal T-cell immunity. Chronic graft-versus-host disease (cGVHD) is one of the major complications following allo-HSCT. The poor reconstitution of T-cell immunity (including the reconstitution of recent thymic output function and T-cell receptor (TCR) repertoire) was associated with cGVHD. In the previous study, we found that cGVHD predicted low TCR rearrangement excision circles (TRECs) levels and slow naïve T-cell recovery. Because GVHD displayed as clonal proliferation of special T-cells clones, which was triggered by donor T cells to recognize the host’s allogene antigen, in the present study we analyzed the TCR Vα and Vβ repertoire and cloanlity in patients with cGVHD, in order to find the special T-cell clones associated with cGVHD and evaluate the molecular characteristics of the CDR3 of TCR Vα and Vβ repertoire of GVHD-associated T-cell clones. Peripheral blood mononuclear cells (PBMNCs) were obtained from 5 leukemia patients with cGVHD after allo-HSCT. The expression and cloanlity analysis of TCR Vα and Vβ repertoire were detected by RT-PCR and genescan technique. Six donors served as controls. Almost all of TCR Vα and Vβ repertoire with polyclonal pattern were identified in normal controls. However, the skew expression pattern of TCR Vα and Vβ repertoire could be detected in patients with cGVHD even more than 4 year after allo-HSCT. Among 29 Vα and 24 Vβ subfamilies, there were only 4∼12 Vα and 4∼11 Vβ subfamilies expressed in patients with cGVHD. Oligoclonal or monoclonal expanded T cells were identified in TCR Vα 2, 3, 6, 10, 12, 14, 15, 25, 26 and TCR Vβ 1, 3, 7∼9, 13, 17, 19, 20 subfamilies respectively. The CDR3 sequences were further analyzed and all the sequences were blasted by internet (http://www.ncbi.nlm.nih.gov) and confirmed that it belonged to specific TCR Vα or Vβ gene rearrangement. The lengths of CDR3 were ranged from 12 to 15 amino acids. The molecular characteristics of the CDR3 of TCR Vα and Vβ genes rearrangement were TCRVα 3 (new name: Vα 17*01)-N-Jα 48*01-Cα (motif: CATEVDFGNEKLIF), TCRVα 2 (new name: Vα 12–2*01)-N-Jα 20*01-Cα (motif: CAVNLNDYKLIF), TCRVβ 1 (new name: Vβ 9*01)-N-Dβ 2*01-N-Jβ 2–1*01-Cβ 2 (motif: CASSDPPETYNEQFF), TCRVβ 7 (new name: Vβ 4–3*01)-N-Dβ 1*01-Jβ 1–1*01-Cβ 1 (motif: CASSHESGNTEAFF). Some TCR subfamily genes shared similarity in CDR3 amino acid motif. The role of specific sequences of CDR3 of TCR Vα and Vβ repertoire and T-cell clones will be confirmed in vivo by animal models.

scholarly journals Interleukin-1R Signaling Is Essential for Induction of Proapoptotic CD8 T Cells, Viral Clearance, and Pathology during Lymphocytic Choriomeningitis Virus Infection in Mice

2012 ◽  
Vol 86 (16) ◽  
pp. 8713-8719 ◽  
Author(s):  
Lars T. Joeckel ◽  
Reinhard Wallich ◽  
Sunil S. Metkar ◽  
Christopher J. Froelich ◽  
Markus M. Simon ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Interleukin 1 ◽  
Lymphocytic Choriomeningitis ◽  
Lymphocytic Choriomeningitis Virus ◽  
Viral Clearance ◽  
T Cell Immunity ◽  
Cell Immunity ◽  
Deficient Mice

The T cell granule exocytosis pathway is essential to control hepatotropic lymphocytic choriomeningitis virus strain WE (LCMV-WE) but also contributes to the observed pathology in mice. Although effective antiviral T cell immunity and development of viral hepatitis are strictly dependent on perforin and granzymes, the molecular basis underlying induction of functionally competent virus-immune T cells, including participation of the innate immune system, is far from being resolved. We demonstrate here that LCMV-immune T cells of interleukin-1 receptor (IL-1R)-deficient mice readily express transcripts for perforin and granzymes but only translate perforin, resulting in the lack of proapoptotic potentialin vitro. LCMV is not cleared in IL-1R-deficient mice, and yet the infected mice develop neither splenomegaly nor hepatitis. These results demonstrate that IL-1R signaling is central to the induction of proapoptotic CD8 T cell immunity, including viral clearance and associated tissue injuries in LCMV infection.

scholarly journals Assessment of T-cell immunity to SARS-CoV-2 in COVID-19 convalescents and vaccinated subjects, using TigraTest® SARS-CoV-2 ELISPOT kit

2021 ◽  
Vol 21 (3) ◽  
pp. 178-192
Author(s):  
D. A. Poteryaev ◽  
S. G. Abbasova ◽  
P. E. Ignatyeva ◽  
O. M. Strizhakova ◽  
S. V. Kolesnik ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Cell Response ◽  
Human Peripheral Blood ◽  
T Cell Response ◽  
T Cell Immunity ◽  
Cell Immunity

With the onset of the COVID-19 pandemic, a number of molecular-based tests have been developed to diagnose SARS-CoV-2 infection. However, numerous available serological tests lack sufficient sensitivity or specificity. They do not detect specific antibodies in a significant proportion of patients with PCR-confirmed COVID-19. There is evidence that some convalescents have a relatively short-lived humoral immunity. In contrast, a number of publications have shown that T-cell response to human coronaviruses, including SARS-CoV-1, MERS, and SARS-CoV-2, can be strong and long-term. Assessment of T-cell immunity to SARS-CoV-2 is important not only for stratification of risks and identification of potentially protected populations with immunity acquired as a result of previous infection, but also for determining immunogenicity and potential efficacy of vaccines under development. The existing methods of quantitative or semi-quantitative assessment of specific T-cell response are mainly used in scientific research and are not standardised. The aim of the study was to develop and verify experimentally a test kit to be used in a standardised procedure for in vitro determination of T-cells specific to SARS-CoV-2 antigens, in human peripheral blood. Materials and methods: the TigraTest® SARS-CoV-2 kit developed by GENERIUM, which determines the number of T-cells secreting interferon gamma in vitro, was tested in the study. Samples of venous blood of volunteers from three different groups were analysed in the study: presumably healthy volunteers; COVID-19 convalescents; individuals vaccinated against SARS-CoV-2. Results: the authors developed the TigraTest® SARS-CoV-2 kit for in vitro determination of T-cells specific to SARS-CoV-2 antigens in human peripheral blood, demonstrated its specificity and performed preliminary assessment of its sensitivity. The study analysed the range and magnitude of the T-cell response in convalescent and vaccinated individuals. A pronounced T-cell response was also shown in some individuals with no symptoms or with unconfirmed diagnosis. It was discovered that the mean T-cell response to peptides of the spike protein (S-protein) was higher in the vaccinated individuals than in the convalescent patients. A correlation was determined between the severity of the disease and the level of T-cell response. Specific contributions of various groups of antigens to the T-cell response after COVID-19 infection were also determined. Conclusions: the TigraTest® SARS-CoV-2 kit is a specific and sensitive tool for the assessment of T-cell immunity to the SARS-CoV-2 virus, which can also be used for vaccinated individuals. The kit may be used in clinical practice for comprehensive assessment of immunity to SARS-CoV-2.

T Cell-Mediated Control of HCMV Infection in Pediatric Patients Receiving Hematopoietic Stem Cell Transplantation.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 5087-5087
Author(s):  
Franco Locatelli ◽  
Daniele Lilleri ◽  
Laura Lozza ◽  
Giovanna Giorgiani ◽  
Piero De Stefano ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Count ◽  
Hcmv Infection ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
Lymphoproliferative Response ◽  
Hematopoietic Stem ◽  
Cell Immunity

Abstract We are studying the development of HCMV-specific CD4+ and CD8+ T cell response after allogeneic hematopoietic stem cell transplantation (HSCT) in pediatric patients. A new technique was developed to simultaneously detect HCMV-specific CD4+ and CD8+ effector T cells using HCMV-infected autologous dendritic cells as stimulators and intracellular staining of IFN-γ production by T cells. This prospective study is based on monthly determination of both HCMV-specific T cell number and in vitro lymphoproliferative response to crude HCMV antigen. Patients are routinely monitored for HCMV infection/reactivation in blood (by determination of either antigenemia or quantitation of viral DNA) and treated according to a strategy of pre-emptive therapy. So far, of 41 patients receiving HSCT from an HLA-identical related donor (n=18), unrelated donor (n=15) or a T cell-depleted HSCT from a haploidentical relative (n=8), 25 patients have reached day +180, while 16 patients completed a follow-up of 90 days. Among the 28 HCMV-seropositive HSCT recipients, 25 developed HCMV-specific CD4+ and CD8+ T-cell response within the first 60 days after transplantation. In these patients, absolute CD4+ T cell count increased over time, but remained lower than that of healthy controls also at later time points. By contrast, CD8+ T cells reached and maintained absolute levels comparable to those of controls already from day +60. At this time, HCMV-specific CD4+ T cell count was comparable to that of controls, while HCMV-specific CD8+ T cell count was higher than that of controls, with no significant change thereafter. On the other hand, in vitro lymphoproliferative response to HCMV antigen was detectable only in about one half of these patients, even at day +180. HCMV infection was detected in blood of 22 of the 25 patients in whom HCMV-specific T cells were present. It was either self-limiting (n=14) or in 8 patients required shorter ganciclovir course (median 7 days, range 5-14) than in the 3 HCMV seropositive patients who developed HCMV infection in the absence of specific immunity (median 67 days, range 42–82, p<0.001). No patient developed HCMV disease or late viral infections. Conversely, HCMV-specific response was detected in only 3/13 HCMV seronegative recipients (none of whom developing detectable HCMV infection in blood). In these patients, both absolute and HCMV-specific T cell counts were lower than those of both controls and HCMV-seropositive HSCT recipients. Our data suggest that effective HCMV-specific T cell immunity can promptly develop after HSCT (regardless of donor type or T-cell depletion of the graft), particularly in seropositive recipients in whom latent virus may be a major antigenic drive for rapid reconstitution of T cell compartment, especially of CD8+ lymphocytes. On the other hand, transfer of memory T cell immunity from seropositive donors to seronegative recipients does not appear to be always sufficient to permit detection of virus-specific lymphocytes in patient’s peripheral blood in the early period after the allograft, possibly also due to the lower chance of in vivo antigen stimulation. The frequent dissociation between IFN-γ production and lymphoproliferative response remains to be explained. Future studies could address modulation of antiviral intervention on the reconstitution of HCMV-specific T cell immune response.

Antibody Targeting of Activated Dendritic Cells Prevents AGVHD and Preserves T Cell Immunity.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3240-3240
Author(s):  
Hannah Cullup ◽  
John Wilson ◽  
Alison M. Rice ◽  
Anne M. Dickinson ◽  
David J. Munster ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
T Cell Immunity ◽  
Mouse Serum ◽  
Human Pbmc ◽  
Cell Immunity ◽  
Life Threatening

Abstract AGVHD is a life threatening complication of allogeneic HSCT, initiated by host and donor DC stimulation of donor T lymphocytes. Current AGVHD prophylaxis targets T lymphocytes, compromising anti-viral and therapeutic anti-leukemic responses. The presence of activated blood DC predicts for clinical AGVHD and we predict that a new strategy targeting therapy to activated DC should prevent alloimmune induction of AGVHD but preserve protective T cell immune responses. CD83 is a cell surface molecule expressed by activated DC. Having shown that a rabbit polyclonal antibody to human CD83 (RA83) depletes activated DC and suppresses alloimmune reactions in vitro, we tested the effect of RA83 treatment on T cell immunity in vitro and its ability to prevent human PBMC induced xenogeneic AGVHD in vivo. The effect of RA83 treatment on T cell numbers, proliferation and cytokine secretion in allogeneic MLR was compared with appropriate controls, including Campath-1H. Allogeneic responses in vitro were mirrored in a DC dependent in vivo model of xenogeneic AGVHD, in which 50×106 human PBMC were injected into an irradiated SCID mouse. Human cytokine levels were measured in MLR tissue culture supernatant (TCSN) and mouse serum at the time of sacrifice. Cytotoxic T cell responses to viral antigens (CMV and FMP) were analysed by specific pentamer analysis and Cr51 release assays, prior to and following HLA restricted peptide antigen specific expansion. RA83 was shown to have NK-mediated ADCC capacity. Cellular proliferation in the allogeneic MLR was reduced by both RA83 and Campath-1H treatment (p= 0.004 and 0.01 respectively vs controls) and both antibodies improved mouse survival in the human xenogenic AGVHD model (RA83: 93%, Campath-1H: 100%, p<0.0001). IFN-g was significantly reduced in the TCSN from MLR treated with RA83 (p=0.0391) and in sera taken from RA83 (p=0.0002) and Campath-1H (p=0.0051) treated mice. Serum IL-4 levels were maintained in RA83 and Campath-1H treated mice. The serum levels of IL-5, IL-8 and TNF in mice treated with RA83 were markedly reduced compared to controls (p=0.0256, 0.0025, 0.025, respectively), and the reductions were similar to those seen in Campath-1H treated mice. Similar numbers of T cells were recovered from RA83 treated and control MLR, and both CMV and FMP specific CD8+ T cells were retained. These cells were readily expanded by peptide pulsing and autologous restimulation and had specific cytotoxic activity comparable to control cultures (see figure). In contrast, Campath-1H treatment removed specific anti-viral responses (vs controls: CMV: p<0.00001 and FMP: p=0.0051). Specific antibody to CD83 depletes activated DC in vitro and prevents xenogeneic human DC dependent AGVHD in vivo. This was accompanied by a Th1 to Th2 skewing of the cytokine response. RA83, but not Campath-1H treatment retained normal numbers of T cells and maintained normal cytotoxic responses to common post-transplant viral infections. Depletion of activated DC may be an effective means of AGVHD control, which maintains T cell immunity to life threatening infections and potentially anti-leukaemia responses. RA83 treatment of allo-MLR preserves T cell anti-CMV immunity RA83 treatment of allo-MLR preserves T cell anti-CMV immunity

Immune Reconstitution after Adoptive Infusion of BPX501 Cells (donor T cells transduced with iC9 suicide gene) in Children Given Alpha/Beta T-Cell Depleted HLA-Haploidentical Hematopoietic Stem Cell Transplantation (haplo-HSCT): Preliminary Phenotypic and Functional Results of a Phase I-II Trial

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3093-3093
Author(s):  
Pietro Merli ◽  
Giuseppina Li Pira ◽  
Valentina Bertaina ◽  
Matilde Sinibaldi ◽  
Barbarella Lucarelli ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Phase I ◽  
Prospective Trial ◽  
T Cell Immunity ◽  
Immune Recovery ◽  
Hematopoietic Stem ◽  
Cell Immunity ◽  
Over Time

Abstract Background: Immune recovery is crucial for patients treated with allogeneic HSCT and in particular of those receiving a T-cell depleted haplo-HSCT. We recently developed a novel method of graft manipulation based on physical elimination of α/β T cells and B-lymphocytes for preventing graft-versus-host disease (GvHD) and EBV-related lymphoproliferative disorders, respectively. Thanks to this approach, we successfully conducted a prospective trial in children with malignant or non-malignant disorders (ClinicalTrial.gov identifier: NCT01810120). Although patients enrolled in this trial had faster immune recovery and lower incidence of infections than those given haplo-HSCT after infusion of positively selected CD34+ cells, reconstitution of adaptive T-cell immunity remains suboptimal. We therefore designed a phase I/II trial aimed at testing the effect on post-transplant immune recovery of adoptive infusion (within 14 + 4 days after transplantation) of BPX-501 cells in children given haplo-HSCT after depletion of α/β T and B cells (ClinicalTrials.gov identifier: NCT02065869). Patients and methods: As of July 25th 2015, 23 children have been infused with BPX-501 cells. The 9 children included in the phase I portion of the study were given 2.5x105, 5x105, and 1x106 BPX-501 cells/kg, respectively, while the 14 included in the phase II received 1x106 BPX-501 cells/kg. This analysis refers to the 16 patients with a minimum follow-up of 90 days; 7 children had acute leukemia and 9 non-malignant disorders. Basic phenotype of circulating lymphocytes was assessed by flow cytometry on fresh heparinized peripheral blood samples at 10, 20, 30, 60, 90, 120 and 150 days post haplo-HSCT, respectively. The following antibodies were used: anti-TCRαβ FITC/anti-TCRγδ PE/anti-CD3 PerCP-Cy™5.5 (WT31, 11F2, SK7), anti-CD4 APC Cy7 (RPA-T4), anti-CD19 BV 510 (SJ25C1), anti-CD3 BV 421 (UCHT1), anti-CD56 PeCy7 (B159), anti-CD16 APC (B73.1), anti-CD8 APC (RPA-T8) from BD Biosciences (San Diego, CA, USA). Antigen-driven activation of peripheral mononuclear cells was evaluated by standard lymphoproliferation assay (LPA) with 3H-thymidine pulsing on day 4 and harvesting 18 hours later. Antigens included PHA or CMV, EBV and AdV whole viral lysate. Results were scored positive with stimulation indexes (SI) >10 for PHA and >3 for viral antigens. Results: None of the patients died from transplant-related complications. Chimerism analysis investigated through short tandem repeats showed that in all but 4 patients, cells were of donor origin before the infusion of BPX-501 cells. In the 4 patients, there was a reversion to complete donor chimerism after infusion of BPX-501 cells. At early time points after haplo-HSCT, gδ T cells predominated over αβ T lymphocytes; subsequently, this latter population became the more largely represented. The number of both CD3+ T lymphocytes and of BPX-501 cells is shown in Panel A of Figure 1, reconstitution of whole T cells in historical children given haplo-HSCT after depletion of α/β T cells is also shown. The number of CD3+ T lymphocytes reached greater than 0.5x109/L 2 months after infusion of BPX-501 cells. Remarkably, while usually immune recovery after transplantation is characterized by prevalence of CD8+ cells, in our patients the physiological predominance of CD4+ lymphocytes was maintained (Panel B of Figure 1. Reconstitution of natural killer cells (NK) is shown in Panel C of Figure 1. As compared to patients receiving CD34+ selected cell haplo-HSCT, children included in this study had a faster reconstitution of mature KIR+/NKG2A- NK cells. Serum levels of IgA and IgM over time are shown in Panel D of Figure 1: there was a recovery of newly synthetized Ig at 3 months. The analysis of the function of T cells showed that the proliferative response to a polyclonal mitogen or to CMV lysate was comparable to that of a healthy control in 50% of patients as early as day + 60 after haplo-HSCT and BPX-501; on day +150, all patients reached a normal SI. Response to both EBV and AdV antigens was slightly delayed, but progressively improved over time (see also Figure 2). Conclusions: Overall, these data indicate that infusion of BPX-501 cells is able to accelerate the recovery of adaptive T-cell immunity since these cells, once infused, expand in vivo and persist over time, potentially contributing to protect patients from infections. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures Moseley: Bellicum Pharmaceuticals: Employment, Equity Ownership.

Selective depletion of alloreactive T cells by targeted therapy of heat shock protein 90: a novel strategy for control of graft-versus-host disease

Blood ◽  
2009 ◽  
Vol 114 (13) ◽  
pp. 2829-2836 ◽  
Author(s):  
Claudia Stuehler ◽  
Stephan Mielke ◽  
Manik Chatterjee ◽  
Johannes Duell ◽  
Sarah Lurati ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
T Cell Immunity ◽  
Hematopoietic Stem ◽  
Graft Versus Host ◽  
Alloreactive T Cells ◽  
Cell Immunity ◽  
Selective Depletion ◽  
Disease Specific

Abstract Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality in patients with hematologic malignancies undergoing allogeneic hematopoietic stem cell transplantation. Current treatment of GVHD relies on immunosuppressive regimens, considerably increasing the incidence of opportunistic infections. As T cells mediate both GVHD as well as protection against viral infections and the malignant disease, strategies to selectively target host-reactive T cells without impairing pathogen- and disease-specific immunity are highly warranted. Activation of T cells is accompanied by increased expression of the chaperone heat shock protein of 90 kDa (Hsp90), which stabilizes several key signaling pathways crucial for T-cell activation. In this study, selective targeting of Hsp90 in activated T lymphocytes with pharmacologic inhibitors already applied successfully in anticancer therapy resulted in induction of apoptosis predominantly in activated cells. Moreover, if T cells were stimulated with allogeneic dendritic cells, alloreactive T cells were selectively eliminated. In contrast, third party reactions including antiviral T-cell immunity were quantitatively and functionally fully preserved. These data suggest that Hsp90 represents a novel target for selective depletion of alloreactive T cells, and provide the rationale for application of Hsp90 inhibitors as potential approach to selectively prevent and treat GVHD in hematopoietic stem cell transplantation recipients without impairing pathogen- and disease-specific T-cell immunity.

Reconstitution Of EBV-Specific Immunity In Adult Recipients Of Double Cord Blood Transplantation Depends On SCF and IL-7 Levels and Maintenance Of a Diverse TCR Repertoire

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3302-3302
Author(s):  
Ioannis Politikos ◽  
Haesook Kim ◽  
Sarah Nikiforow ◽  
Anoma Nellore ◽  
Lequn Li ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Immunity ◽  
Lytic Cycle ◽  
Hematopoietic Stem ◽  
Cell Numbers ◽  
Ebv Reactivation ◽  
Cell Immunity ◽  
Tcr Repertoire ◽  
Ebv Viremia

Abstract Umbilical cord blood (UCB) is an alternative source of hematopoietic stem cells (HSC) for allogeneic HSC transplantation but reconstitution of T cell immunity remains problematic even with double UCB transplantation (dUCBT). This is related to the small T cell numbers of the UCB graft, the naïve nature of these T cells and often the use of ATG as part of the conditioning regimen. Consequently, UCBT recipients are susceptible to viral infections that require intact T cell immunity for their containment. Epstein Barr virus (EBV) is a gammaherpes virus that is transmitted orally. EBV initially replicates in a lytic cycle in the epithelium of the oropharynx and then establishes a latent state in memory B cells. Reactivation of EBV from latent to lytic cycle is mediated by physiologic stress of the memory B cell pool and by defects in antiviral T cell immunity, and can cause aggressive lymphomas known as post transplant lymphoproliferative disorder (PTLD) in immunocompromised hosts. The specific immunological correlates of EBV reactivation in adult UCBT recipients have not been examined. In the present study we attempted to distinguish immune reconstitution profiles in UCBT recipients who developed EBV viremia from those who did not, in order to better understand the immune profile of UCBT recipients who control gammaherpes virus reactivation. Thirty-one patients with hematologic malignancies received dUCBT with melphalan, fludarabine, ATG conditioning and tacrolimus plus sirolimus for GvHD prophylaxis. EBV viral load was determined on a weekly basis after dUCBT. Immunophenotype of peripheral blood lymphocytes, serum cytokine levels and T cell receptor excision circle analysis (TREC) values, were examined prior to and at 1, 2, 3, 6 and 12 months after UCBT and were compared between EBV viremic and non-viremic patients using the Wilcoxon-rank sum test. During the first 12 months after dUCBT, 14 of 31 (45%) patients developed EBV viremia and four (13%) developed PTLD. At one month after dUCBT, patients who developed EBV viremia displayed higher numbers of CD19+ B cell numbers (p=0.04) and CD4+CD25+ T regulatory cells (p=0.03) compared with patients who never became viremic. Surprisingly, development of EBV viremia correlated with increased numbers of CD3+ (p=0.04), CD4+ (p=0.015) and CD8+ (p=0.021) T cells. This finding was counterintuitive, as one might expect better quantitative peripheral T cell reconstitution in patients with recovering immunity and no EBV reactivation. One mechanism of impaired immune reconstitution after UCBT is skewing towards a late effector memory T cell phenotype, a stage in which T cells are incapable of mounting protective immune responses. We examined whether development of EBV viremia was associated with altered naïve versus memory T cell distribution. We determined that patients who developed EBV reactivation had higher numbers of memory cell subsets (CD4+CD45RO+, p=0.0023; CD8+CD45RO+, p=0.019) at two months after dUCBT. Neither development nor control of EBV viremia correlated with TREC recovery, which occurred after the time period of EBV viremia. We were unable to identify EBV-specific T cells in any patient group due to the very low T cell numbers during the time of viremia. However, analysis of repertoire diversity by deep-sequencing on PCR-amplified CDR3 regions of the TCRb gene using the ImmunoSEQ assay showed a more diverse TCR repertoire, as determined by higher entropy (p=0.03) and lower clonality (p=0.03), among patients who did not develop EBV reactivation, compared with those who developed EBV viremia and PTLD. Assessment of serum levels of SCF (a c-kit ligand), IL-7, thrombomodulin, VEGF and angiopoetin-1 showed that patients without EBV viremia had significantly higher levels of SCF (p=0.0001) and IL-7 (p=0.05), at 1 and 2 months after dUCBT, compared with patients who developed EBV reactivation and PTLD. SCF-mediated signaling via c-kit is integral to the homing and longevity of uncommitted hematopoietic stem cells whereas IL-7 is a critical factor for survival and homeostasis of naïve T cells. Together our findings suggest that control of EBV reactivation after dUCBT might be linked to the support of naïve T cell homeostasis, which enables maintenance of a diverse TCR repertoire. Disclosures: No relevant conflicts of interest to declare.

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