Prolonged CD4 depletion after sequential autologous peripheral blood progenitor cell infusions in children and young adults

Blood ◽  
2000 ◽  
Vol 96 (2) ◽  
pp. 754-762 ◽  
Author(s):  
Crystal L. Mackall ◽  
Dagmar Stein ◽  
Thomas A. Fleisher ◽  
Margaret R. Brown ◽  
Frances T. Hakim ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Immune Reconstitution ◽  
Residual Disease ◽  
High Dose Chemotherapy ◽  
High Dose ◽  
Cell Dose ◽  
Age Related ◽  
Large Numbers

Administration of mobilized peripheral blood progenitor cells (PBPCs) after high-dose chemotherapy rapidly restores multilineage hematopoiesis, but the ability of such products to restore lymphocyte populations remains unclear. In this report, we evaluated immune reconstitution in a series of patients treated with sequential cycles of high-dose chemotherapy, followed by autologous PBPC infusions (median CD34+ cell dose 7.2 × 106 cells/kg [range 2-29.3]). Although patients experienced rapid reconstitution of B cells and CD8+ T cells, we observed CD4 depletion and diminished immune responsiveness in all patients for several months after completion of therapy. Mature CD4+ T cells contained within the grafts did not appear to contribute substantially to immune reconstitution because CD4 counts did not differ between recipients of unmanipulated T-cell replete infusions versus CD34 selected, T-cell–depleted infusions. Rather, at 12 months after therapy, total CD4 count was inversely proportional to age (ρ = −0.78,P = .04), but showed no relationship to CD34 cell dose (ρ = −0.42, P = .26), suggesting that age-related changes within the host are largely responsible for the limited immune reconstitution observed. These results demonstrate that in the autologous setting, the infusion of large numbers of PBPCs is not sufficient to restore T-cell immune competence and emphasize that specific approaches to enhance immune reconstitution are necessary if immune-based therapy is to be used to eradicate minimal residual disease after autologous PBPC transplantation.

Prolonged CD4 depletion after sequential autologous peripheral blood progenitor cell infusions in children and young adults

Blood ◽  
2000 ◽  
Vol 96 (2) ◽  
pp. 754-762 ◽  
Author(s):  
Crystal L. Mackall ◽  
Dagmar Stein ◽  
Thomas A. Fleisher ◽  
Margaret R. Brown ◽  
Frances T. Hakim ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Immune Reconstitution ◽  
Residual Disease ◽  
High Dose Chemotherapy ◽  
High Dose ◽  
Cell Dose ◽  
Age Related ◽  
Large Numbers

Abstract Administration of mobilized peripheral blood progenitor cells (PBPCs) after high-dose chemotherapy rapidly restores multilineage hematopoiesis, but the ability of such products to restore lymphocyte populations remains unclear. In this report, we evaluated immune reconstitution in a series of patients treated with sequential cycles of high-dose chemotherapy, followed by autologous PBPC infusions (median CD34+ cell dose 7.2 × 106 cells/kg [range 2-29.3]). Although patients experienced rapid reconstitution of B cells and CD8+ T cells, we observed CD4 depletion and diminished immune responsiveness in all patients for several months after completion of therapy. Mature CD4+ T cells contained within the grafts did not appear to contribute substantially to immune reconstitution because CD4 counts did not differ between recipients of unmanipulated T-cell replete infusions versus CD34 selected, T-cell–depleted infusions. Rather, at 12 months after therapy, total CD4 count was inversely proportional to age (ρ = −0.78,P = .04), but showed no relationship to CD34 cell dose (ρ = −0.42, P = .26), suggesting that age-related changes within the host are largely responsible for the limited immune reconstitution observed. These results demonstrate that in the autologous setting, the infusion of large numbers of PBPCs is not sufficient to restore T-cell immune competence and emphasize that specific approaches to enhance immune reconstitution are necessary if immune-based therapy is to be used to eradicate minimal residual disease after autologous PBPC transplantation.

scholarly journals Very Early Adoptive Transfer of Ex Vivo Generated Multi-Virus Specific T Cells Is a Safe Strategy for Prevention of Viral Infection after Allogeneic T Cell Depleted Stem Cell Transplantation

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 812-812 ◽  
Author(s):  
Pawel Muranski ◽  
Sarah I Davies ◽  
Sawa Ito ◽  
Eleftheria Koklanaris ◽  
Jeanine Superata ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Immune Reconstitution ◽  
Mononuclear Cells ◽  
Ex Vivo ◽  
Viral Reactivation ◽  
High Dose ◽  
Cdr3 Sequencing ◽  
Cmv Reactivation

Abstract Background: Reactivation of latent viruses post-allogeneic stem cell transplant (SCT) negatively affects outcomes and increases non-relapse mortality. We have reported over 80% early CMV reactivation rate and significant additional costs in recipients of T cell depleted (TCD) SCT. Ex vivo generated multi-virus specific T cells (MVSTs) are effective as a therapy of active infection, but have not been evaluated as prophylaxis early after transplant. In a Phase I study (NIH 14-H-0182) we transferred MVSTs targeting immunodominant viral proteins of CMV, Epstein-Barr virus (EBV), BK and adenovirus (Ad) immediately post SCT as a novel approach to prevent viral reactivation post-SCT. Methods: All subjects were enrolled in HLA-matched T cell depleted (TCD) transplant protocol (NIH 13-H-0144). MVSTs cells were manufactured from SCT sibling donors by stimulation of elutriated lymphocytes for 14 days with seven overlapping peptide libraries (pepmixes) pulsed onto autologous dendritic cells (DCs) in presence of IL-7, IL-15 and IL-2. MVSTs were infused as early as possible (day 0 to +60) post SCT. A Phase I 3+3 dose escalation design was used as follows: Cohort 1 - 1x10e5 total nucleated cells (TNC)/kg, Cohort 2 - 5x10e5 TNC/kg, Cohort 3 - 1x10e6 TNC/kg. Three additional subjects received MVST cells manufactured using pepmix-pulsed mononuclear cells as stimulators (Cohort 3A; 1x10e6TNC/kg) under an amended protocol. The primary safety endpoint at day 42 post infusion was the occurrence of dose limiting toxicity (DLT) defined as Grade IV GVHD or any other severe adverse even (SAE) deemed to be at least "probably" or "definitely" related to the MVST infusion. Patients were followed to day +100 post SCT for secondary outcomes of efficacy, immune reconstitution and GVHD biomarkers (ST2, REG3). CDR3 sequencing (ImmunoSEQ) was performed on selected MVST products and peripheral blood samples post MVST and compared to control SCT recipients. GVHD biomarkers were analyzed pre- and post-treatment. Results: Twelve subjects were treated: nine received MVSTs generated using DCs and three subjects using mononuclear cells (cohort 3A). Median time from SCT to MVST administration was 13 days (range D +2 to +52 post-SCT). Median time to MVST for subjects in Cohort 3A was 3 days (range 2-7). There were no immediate infusion-related adverse events or DLT at the highest dose level. De novo grade II-III aGVHD post-MVST infusion was seen in three subjects (one in Cohort 1 and two in Cohort 3A), but GVHD biomarker elevation predated MVST infusion. CMV reactivation post-MVST occurred in 6 out of 12 subjects (50%) vs. 45 out of 52 patients (50% vs 87%) in a historical group of recipients of T cell depleted SCT. In all cases CMV reactivation occurred in the context of high dose steroids and in two subjects MVSTs were derived from CMV seronegative donors with minimal anti-CMV activity. One subject experienced rapidly rising AdV viremia (asymptomatic) and received an additional infusion of MVSTs. We saw self-limiting low level EBV replication in 8 cases and one BK viremia, but no disease. CDR3 sequencing of MVST products and serial peripheral blood from subjects revealed a robust contribution of ex vivo expanded cells to the overall repertoire, in contrast to untreated controls where the repertoire of (sham) MVST cell products generated from the transplant donors did not significantly overlap with the immune repertoire in peripheral blood of TCD-SCT recipients in the early post SCT period. Only at day +180 some convergence of repertoires became visible indicating spontaneous immune reconstitution (Figure). Detailed CDR3 analysis of cytokine-captured CMV pp65 and IE-1 specific CD4+ and CD8+ T cells was performed in a representative subject clearly demonstrating the detrimental effect of high-dose steroids on frequency of anti-viral T cells, precipitating CMV reactivation. Conclusions: This is the first report demonstrating safety and feasibility of using MVST immediately post-SCT to rapidly reconstitute anti-viral immunity and ameliorate the detrimental impact of the early viral reactivation in SCT recipient. No DLTs were seen and a minimal risk of aGVHD was observed, as there was no correlation with GVHD biomarkers. As revealed by serial CDR3 sequencing, MVSTs robustly contributed to the T cell repertoire. Our results suggest efficacy of this strategy in reducing viral reactivation. A Phase II study is warranted. Disclosures No relevant conflicts of interest to declare.

Age-related increase of frequency of a new, phenotypically distinct subpopulation of human peripheral blood T cells expressing lowered levels of CD4

Blood ◽  
2001 ◽  
Vol 98 (4) ◽  
pp. 1100-1107 ◽  
Author(s):  
Ewa Bryl ◽  
Magdalena Gazda ◽  
Jerzy Foerster ◽  
Jacek M. Witkowski
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
The Elderly ◽  
Cell Subpopulation ◽  
Cell Phenotype ◽  
Receptor Complexes ◽  
Age Related

Aging is associated with modifications of T-cell phenotype and function, leading to impaired activation in response to both new and recall antigens. It is not known if T-cell activation results in elimination of a number of the CD4 molecules from the cell surface, as is the case with CD3/T-cell receptor complexes, or how aging influences the process. The T cells of young and elderly donors with reduced expression of CD4 were examined to see whether these cells exhibit other phenotypic features suggesting their active state. It was found that T lymphocytes expressing CD4 can be divided into 2 semidiscrete subpopulations: the major (CD4+) population, in which the level of expression of CD4 is constant and high, and a minor population (CD4lo), in which the expression of CD4 can be up to an order of magnitude lower than on the CD4+ cells. The proportion of CD4locells is age dependent and highly variable in the apparently healthy human population, with the expression of CD4 ranging from around 10% of all peripheral blood lymphocytes in the young to more than 30% in the elderly. Lowered expression of CD4 is correlated with a reduced expression of CD3, as well as with a decreased amount of CD28 and CD95Fas. Activation of CD4lo cells is suggested by their expression of CD25 and increased amounts of HLA-DR. Phenotypic characteristics of the CD4lo T-cell subpopulation suggest that it might be formed by (perhaps chronically) activated, temporarily apoptosis-resistant cells, possibly accumulating in the elderly.

Donor DC Subsets Polarize Donor T-Cell Immune Responses in Allogeneic BMT.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 573-573
Author(s):  
Jian-Ming Li ◽  
Cynthia Giver ◽  
Doug McMillan ◽  
Wayne Harris ◽  
David L. Jaye ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
T Cell ◽  
Immune Responses ◽  
Peripheral Blood ◽  
Immune Reconstitution ◽  
Hematopoietic Cell ◽  
Post Transplant ◽  
Donor T Cells ◽  
Ifn Γ

Abstract Introduction: Impaired or inappropriate immune reconstitution after allogeneic bone marrow transplantation (BMT) can lead to infection, graft-versus-host disease (GvHD) and leukemia relapse. We have previously reported that BM contains two populations of dendritic cell (DC) subsets, CD11b+ DC and CD11b− DC, and that CD11b depleted donor BM promoted increased donor T-cell chimerism and increased graft-versus-leukemia (GvL) activity in C57BL/6 → B10BR transplants [BBMT, 2004, 10: 540]. To explore the mechanism by which CD11b-depletion improved allo-reactivity, we performed allogeneic hematopoietic cell transplants using defined populations of donor stem cells, DCs, and T-cells in a MHC mis-matched BMT model. Methods: We transplanted FACS purified populations of 50,000 GFP+ CD11b- DC or CD11b+ DC in combination with 5,000 FACS purified Lin- Sca-1+ c-kit+ hematopoietic stem cells (HSC) and 300,000 or 1,000,000 congenic spleen T-cells from C57BL/6 donors into C57BL/6[H-2Kb], B10BR[H-2Kk] and PL/J[H-2Ku] recipients. Proliferation of CFSE stained donor T-cells was measured at 72 hours post-transplant. FACS cytometric bead array and intracellular cytokine staining measured serum and intracellular cytokines in donor T-cells. Results: The initial proliferation and Ki-67 expression of CFSE labeled donor T-cells in allogeneic recipients were much higher than in syngeneic recipients (homeostatic proliferation). Confocal microscopy showed co-localization of donor DC subsets with donor T-cells in the recipient spleens at 3 and 10 days post-transplant. In the allogeneic transplant settings, donor T-cells co-transplanted with CD11b- DC showed increased IFN-γ synthesis at 3 and 10 days post-transplant compared to donor T-cells co-transplanted with HSC plus CD11b+ DC or HSC alone. Increased proliferation of donor T-cells led to increased donor T-cell chimerism at day 10, 30, 60, and day105 post-transplant among recipients of CD11b- DC compared to recipients of HSC alone or HSC plus CD11b+ DC (Figure 1). Transplantation of spleen T-cells and CD11b- DC did not increase GvHD, but was associated with full donor chimerism. In contrast, transplantation of allogeneic CD11b+ DC led to persistence and expansion of residual host T-cells (Figure 2), increased numbers of donor CD4+CD25++Foxp3+ T-cells, and higher serum level of IL-10 supporting early post-transplant expansion of donor T regulatory cells (Treg). Conclusions: Donor CD11b- DC promoted immune reconstitution by polarizing donor T-cells to Th1 immune responses associated with increased IFN-γ synthesis and donor T-cell proliferation, while donor CD11b+ DC suppressed immune reconstitution by inhibiting donor T-cell allogeneic immune responses. These data support a novel paradigm for the regulation of post-transplant immunity and suggest clinical methods to test the hypothesis that manipulation of the DC content of a hematopoietic cell allograft regulates post transplant immunity in the clinical setting. Figure 1. Donor Spleen Derived T-cells in Peripheral Blood [* p<0.05, v.s. recipients of HSC plus CD11b(+)DC and spleen T-cells] Figure 1. Donor Spleen Derived T-cells in Peripheral Blood [* p<0.05, v.s. recipients of HSC plus CD11b(+)DC and spleen T-cells] Figure 2. Host Derived T-cells in Peripheral Blood [* p<0.05, v.s. recipients of HSC plus CD11b(-)DC and spleen T-cells] Figure 2. Host Derived T-cells in Peripheral Blood [* p<0.05, v.s. recipients of HSC plus CD11b(-)DC and spleen T-cells]

Cellular Immune Responses To Vector In a Gene Therapy Trial For Hemophilia B Using An AAV8 Self-Complementary Factor IX Vector

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 717-717
Author(s):  
Etiena Basner-Tschakarjan ◽  
Federico Mingozzi ◽  
Yifeng Chen ◽  
Amit Nathwani ◽  
Edward Tuddenham ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Board Of Directors ◽  
Peripheral Blood ◽  
Factor Ix ◽  
High Dose ◽  
Advisory Committees ◽  
Ifn Γ ◽  
Cellular Immune

Abstract In a clinical study of gene transfer for hemophilia B an adeno-associated virus vector serotype 8 (AAV8) expressing a self-complementary liver-specific expression cassette for the factor IX (FIX) transgene was administered intravenously in ten affected subjects. The results of the first part of the study have been published (NEJM 365:2357-65, 2011). In this abstract we present the immunomonitoring data, using Interferon-gamma (IFN-γ) ELISpot and polyfunctional T cell analysis of peripheral blood mononuclear cells (PBMCs) to monitor cellular immune responses to vector capsid and to Factor IX. We have previously shown that the cellular immune response was directed solely towards AAV capsid epitopes, not FIX, and that the response was dose-dependent. Out of six subjects infused in the high dose cohort (2x1012vg/kg), 4/6 manifested a minor rise in liver enzyme levels and detection of capsid-specific T cell reactivitiy in the ELISpot assay at ∼7-10 weeks post vector infusion. Maximum results on IFN- γ ELISpots ranged from 200-500 sfu/million cells. In two of these cases a modest decline in FIX level also occurred. Prompt initiation of prednisolone reversed these effects and rescued FIX levels. The remaining two subjects infused at the high dose, showed no rise in liver enzyme levels at any time point. However capsid reactive T cells were detectable in one subject as early as one to two weeks after vector infusion in peripheral blood by IFN-γ ELISpot assay, while no activation at all was detected in the other subject, possibly due to low cell recovery and viability of the cells. A similar immune response profile, with early detection of activated T cells but no rise in liver enzymes, was also observed in both subjects in the intermediate dose cohort in the first part of this study. Polyfunctional T cell analysis revealed concurrent Interleukin-2, Tumor necrosis factor-alpha and CD107a positivity in activated T cells at the peak of activation. Furthermore it showed that capsid-specific early T cell responses were detectable in the CD4+ T cell and later in the CD8+T cell compartment. Long-term immune monitoring of all subjects is ongoing. Importantly in one of the first two subjects treated at the high dose, capsid reactive T cells were detected by ELISpot 1.5 years after gene transfer; these cells were not detected in the other subject in whom long-term follow-up samples are available. Of note, capsid-reactive T cells were also seen at late time points (>1 year after infusion) in a middle dose subject and a low dose subject. Despite detectable T cell reactivity towards the AAV capsid in the peripheral blood FIX expression remained stable, suggesting that there is a short window of time during which transduced hepatocytes present a target for cytotoxic T cells, and that T cell positivity after this window is without any clinical consequences. In conclusion, for this scAAV8 vector there appears to be a critical threshold vector dose for a clinically detectable immune response, starting at 2x1012 vg/kg. The clinically detectable response occurred in four out of six subjects so far, and was manifest within a critical time interval of 7-10 weeks post infusion. The capsid-specific response was polyfunctional and detected in CD4+ and CD8+T cells in peripheral blood. It is important to note that not all subjects treated at the high dose developed an immune response. However, given the limited dataset, it is not yet possible to define predictive parameters, e.g. HLA type of a subject, for an immune response. Continued monitoring and future studies with more subjects will be necessary to confirm the presented findings, in particular time and rate of occurrence of a cellular response as well as successful treatment with a short course of Prednisolon. Disclosures: Tuddenham: Pfizer: Consultancy. Reiss:Hemophilia of Georgia: Honoraria. High:BristolMyersSquibb: Consultancy, membership on a Data Safety and Monitoring Board, membership on a Data Safety and Monitoring Board Other; Elsevier, Inc.: royalties from textbook, royalties from textbook Patents & Royalties; Genzyme, Inc.: Membership on an entity’s Board of Directors or advisory committees; Intrexon: Consultancy; Novo Nordisk: Consultancy, Member of a grant review committee, Member of a grant review committee Other; Shire : Consultancy; Benitec: Consultancy; bluebirdbio, Inc.: Consultancy, Equity Ownership, Membership on an entity’s Board of Directors or advisory committees; BioMarin: Consultancy; Alnylam Pharmaceuticals: Consultancy, Membership on an entity’s Board of Directors or advisory committees.

Favorable Immune Reconstitution Profile after Allogeneic Hematopoietic Stem Cell Transplantation with Post-Transplant Cyclophosphamide

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2236-2236
Author(s):  
Omer Hassan Jamy ◽  
Ayman Saad ◽  
Rachael Orlandella ◽  
Samantha B Langford ◽  
Ravi K. Paluri ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Peripheral Blood ◽  
Research Funding ◽  
Immune Reconstitution ◽  
First Year ◽  
Gvhd Prophylaxis ◽  
One Year

Abstract Background: The administration of post-transplant high-dose cyclophosphamide (PTCy) has been shown to be an effective strategy for GvHD prophylaxis following allogeneic peripheral blood stem cell transplantation(PBSCT) from alternative donors. PTCy is toxic to allogeneic activated proliferating T lymphocytes, such as effector T cells. Conversely, it may not materially affect memory T cells. Methods: We evaluated immune reconstitution profile and transplant outcome in patients who received PBSCT with and without PTCy. PTCy was given on day +3 and +4 following haploidentical transplant (HAPLO), or only on day +3 following HLA-matched unrelated donor (MUD) transplant. No PTCy was given to patients with HLA-matched related donors (MRD). All patients received GvHD prophylaxis as tacrolimus (day +5 to +180) and MMF (day +5 to +35). Preparative regimens were myeloablative regimens (fludarabine/busulfan, fludarabine/TBI 12 Gy, or CY/TBI 12) in all patients except 4 patients (received fludarabine/melphalan). Immune reconstitution profile (IRP) was tested via serial flow cytometry analysis of peripheral blood lymphocytes after transplant were done on days +30, +100, and +180. Results: Data of 70 patients who underwent allogeneic PBSCT in our institution were analyzed in 3 groups; MRD (n=22), MUD (n=35), and HAPLO (n=13). The total cohort had 33 males (47%), and had median age of 52 years (range 20-70). All patients had hematological malignancy except one patient with HLH. The median duration of follow up was 6 months (range 1-17). The median day of neutrophil and platelet engraftment were 13, 12, 17 and 18, 15, 22 days for MRD, MUD and HAPLO groups respectively. The one-year overall survival of the whole group was 67% (95% confidence interval: 48-80) with no difference in OS among the 3 cohorts (log rank P value 0.4) (Figure 1). Lymphocyte and lymphocyte subset (T, B, NK) count recovery for MUD and HAPLO was significantly less (p<0.05) than MRD during the first month post-HSCT but these differences were statistically insignificant by day +60 and remained so through day +365. Recovery of both CD4+ and CD8+ naïve T cell (CD45RA+CD27+CD197+) population was generally slower for HAPLO patients during the first year and significantly less through day+ 180 for CD4+ T cells. As predicted, central memory (CD45RA-CD27+CD197+) CD4+ and CD8+ T cells remained proportionately equivalent at 40% and 28% respectively for all groups during the first year. The effector memory (CD45RA-CD27+CD197-) population was also proportionately consistent at 25% of total for both CD4+ and CD8+ subsets. Interestingly, the effector T cell population (CD45RA+CD27-CD197-) trended higher for all three recipient groups at each time point for both CD4+ and CD8+ populations increasing from 20% at one month to over 40% at one year. Conclusion: Post-PBSCT survival was not significantly different from alternative donor graft recipients and those that received MRD PBSCT. Lymphocyte recovery was impaired for the PTCy groups in the immediate post-PBSCT period but quickly recovered to that seen in MRD recipients. Figure 1 Figure 1. Disclosures Saad: Spectrum: Honoraria; American Porphyria foundation: Research Funding; Astellas: Research Funding; Alexion: Honoraria. Lamb:Incysus, Ltd: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Research Funding.

Hematopoietic stem cell dose correlates with the speed of immune reconstitution after stem cell transplantation

Blood ◽  
2004 ◽  
Vol 103 (11) ◽  
pp. 4344-4352 ◽  
Author(s):  
Benny J. Chen ◽  
Xiuyu Cui ◽  
Gregory D. Sempowski ◽  
Jos Domen ◽  
Nelson J. Chao
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
B Cells ◽  
Hematopoietic Stem Cells ◽  
Immune Reconstitution ◽  
Hematopoietic Stem ◽  
Time Points ◽  
Cell Dose

Abstract In the current study, we tested whether higher numbers of hematopoietic stem cells correlate with the speed of immune reconstitution in a congenic transplantation model (C57BL/Ka, CD45.1, Thy1.1→C57BL/6, CD45.2, Thy1.2) using purified hematopoietic stem cells (c-Kit+Thy1.1lowLin-/lowSca-1+). There were 3 different doses of stem cells used (400, 1000, and 5000). Phenotypic analyses in peripheral blood and spleen demonstrated that higher numbers of infused stem cells are associated with more rapid regeneration of T cells (CD4+, CD8+, naive CD4+, naive CD8+) and B cells at early time points. The numbers of T and B cells eventually became equivalent between different dose groups at late time points. Production of interleukin-2 and inter-feron-γ per T cell was similar regardless of stem cell dose even when tested at the time when there were significant differences in peripheral T-cell counts. The improved immune recovery was attributed to a more rapid regeneration of donor-type immune cells. Higher numbers of total thymocytes and signal joint T-cell receptor excision circles were observed in the higher dose stem cell recipients, suggesting that accelerated regeneration of T cells was due to enhanced thymopoiesis. (Blood. 2004;103:4344-4352)

scholarly journals Method of Mobilization: Implication on Cell Subsets in the Graft and Immune Reconstitution Post Autologous Hematopoietic Cell Transplantation (AHCT)

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1132-1132
Author(s):  
Melhem M. Solh ◽  
Rathmann Kristin ◽  
Sauvi chang-Fong ◽  
Jeremiah Oyer ◽  
Wesam B. Ahmed ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Transplantation ◽  
Peripheral Blood ◽  
Immune Reconstitution ◽  
Hematopoietic Cell Transplantation ◽  
Hematopoietic Cell ◽  
P Value ◽  
Autologous Hematopoietic Cell Transplantation ◽  
Graft Composition

Abstract Method of Mobilization: Implication on Cell Subsets in The Graft and Immune Reconstitution post Autologous Hematopoietic Cell Transplantation (AHCT) The optimal mobilization method for either myeloma or lymphoma patients undergoing AHCT is still debatable and strategies for graft collection vary between different institutions. Plerixafor, a CXCR4 antagonist is used for peripheral blood stem cell mobilization in multiple myeloma and non-Hodgkins lymphoma patients requiring AHCT. The effect of plerixafor on graft composition has scarce data that are based mostly on cryopreserved samples. Moreover; the effect of plerixafor on immune reconstitution and hematologic recovery post AHCT has not been well evaluated. The goal of our study was to compare graft composition, hematologic and immune reconstitution recovery among patients mobilized with plerixafor plus G-CSF to those mobilized with G-CSF alone. Methods: 49 patients eligible for AHCT were enrolled on a single arm prospective trial at a single transplant center. All patients were mobilized with G-CSF 10µg/kg/day for 4 consecutive days. A peripheral blood CD34 level of <20/µl on day 4 was used as a cutoff to use plerixafor 0.24mg/kg in addition to G-CSf on 9pm of the fourth day. Peripheral blood collection was started on day 5 and was continued till the target dose is achieved or a minimum CD 34+ cell dose of >2x106 cells/Kg was obtained after 3 collection days. Samples from the freshly collected graft and patients' peripheral blood on days +30 and +60 were analyzed by flow cytometry (BD FACSCanto II) . A single platform assay was used (Beckman-Coulter Stem kit) via a ISHAGE protocol. The antibody cocktail contained the following pre-conjugated monoclonal antibodies: CD56-PE (Miltenyi Biotech, Auburn, CA), CD3-APC, CD16-FITC, (Beckman Coulter, Brea, CA), CD19-PE-CY7 (BD Biosciences, San Jose, CA). Data were acquired using BD FACSCanto II (BD Biosciences) and analyzed with the FACSDiva software (BD Biosciences) to quantify CD3+ T cells, CD3+ CD56+ NK-like T cells, CD56+ CD16+ and CD56+ CD16- NK cells as well as CD19+ B cells. Results: 49 patients with a median age of 58 years (range 21-75) were mobilized with either G-CSF alone (N=16) or plerixafor +G-CSF (G+P)(N=33).The median number of collection days was 1.42 and 1.81 (p=0.2) and the median collected CD34+ dose was 8.28x106/kg and 5.24x106 /kg (p=022) in the G+P and G-CSF alone groups respectively. Both groups had similar times to neutrophil and platelet engraftment. The graft analysis showed a white blood count of 309x109/l and 262x109/l (p=0.38), median percentage of CD34+ cells of 0.75% and 0.73% (p=0.81), percentage of CD3+ T cells of 25.6% and 22% (p=0.6) in the G+P and G-CSF alone groups resepectively. Both groups had similar proportions of CD3+, CD4+,CD8+, NK, NKT and iNKT cells in the mobilized grafts. Peripheral blood samples at day +30 and day +60 were analyzed for T cell markers and hematologic recovery (table 1). There was no significant difference between absolute lymphocyte counts, NK cell counts, T cells and absolute neutrophil count. Conclusion: Plerixafor when combined with G-CSF helps in achieving mobilization goals in patients predicted to be poor mobilizers based on peripheral CD34 levels. The addition of plerixafor doesn't not seem to affect T cell composition of the graft and yields similar hematologic and immune recovery when compared to mobilization with G-CSF alone. Table 1: Immune Reconstitution at Day 30 and Day 60 post Autologous Transplantation Treatment Group G-CSF (N=16) Plerixafor + G-CSF (N=33) P-value G-CSF (N=16) Plerixafor + G-CSF (N=33) P-value Day 30 Day 60 WBC 5.08 5.41 0.873 4.94 5.38 0.654 HGB 10.86 11.19 0.353 11.22 11.17 0.757 HCT 32.35 33.66 0.321 33.36 33.53 0.565 PLT 119.88 161.42 0.068 166.94 173.73 0.949 Abs Lymph 1.09 1.44 0.296 1.41 1.50 0.974 % NK 26.14 30.38 0.277 11.53 20.09 0.095 Abs NK 0.31 0.35 0.186 0.17 0.21 0.470 % T cell 67 60 0.183 76.15 67.39 0.340 Abs T cell 0.72 0.96 0.717 1.35 .82 0.095 NKT%* 5.28 3.33 8.25 3.38 B cell % 2.38 1.52 0.922 2.63 5.58 0.424 Abs. Neut count 2.99 2.64 0.488 2.85 3.01 0.848 Disclosures No relevant conflicts of interest to declare.

scholarly journals Lymphoid Reconstitution After Autologous PBSC Transplantation With FACS-Sorted CD34+ Hematopoietic Progenitors

Blood ◽  
1998 ◽  
Vol 91 (7) ◽  
pp. 2588-2600 ◽  
Author(s):  
Catherine Bomberger ◽  
Meeta Singh-Jairam ◽  
Glenn Rodey ◽  
Anastasia Guerriero ◽  
Andrew M. Yeager ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Peripheral Blood ◽  
Pcr Amplification ◽  
Cell Receptor ◽  
High Dose ◽  
Normal Numbers ◽  
Hematopoietic Progenitors ◽  
Cell Repertoire

Abstract T-cell and B-cell reconstitution was studied in nine patients who received fluorescence activated cell sorter (FACS)-sorted autologous CD34+ hematopoietic progenitor cells (HPC). The mean numbers of T cells (CD3+), B cells (CD19+) and CD34+ HPC administered to each patient were .004, .002, and 1.8 × 106 cells/kg, respectively. After high-dose myeloablative chemotherapy (busulfan, cyclophosphamide, etoposide) CD34+ HPC were infused and lymphoid reconstitution was monitored using flow cytometry and reverse transcriptase-polymerase chain reaction (RT-PCR) amplification of VDJ T-cell receptor (TcR) sequences. Restoration of normal numbers of peripheral blood T cells and B cells among recipients of FACS-sorted CD34+ HPC was delayed compared to recipients of non-T-cell–depleted PBSC autografts. In both patient groups, the circulating T cells were primarily CD4−, CD8+, αβ TcR+, and CD45RO+, CD45RA− during the first 2 months after transplant. Subsequent increases in the frequency of CD45RA+ CD45RO− T cells occurred at 2 to 3 months after transplant, suggesting maturation of CD34+hematopoietic progenitors to “naive” T cells. Analysis of the TcR repertoire after hematopoietic reconstitution demonstrated decreased diversity of Vβ TcR expression associated with global decreases in the absolute number of total peripheral blood T cells and most Vβ TcR+ subsets. Three of nine recipients of FACS-sorted CD34+ HPC demonstrated significant increases in the percentage of γδ+ peripheral T cells and CD5+ B cells at 3 to 9 weeks after transplantation, and all patients had transient oligoclonal expansions of T cells expressing specific Vβ TcR. Transplantation with highly purified CD34+ HPC results in reduced diversity of the peripheral T-cell repertoire during the early post-transplant period compared with patients receiving unmanipulated or MoAb-depleted transplants.

scholarly journals Interleukin-15 Increases Effector Memory CD8+ T Cells and NK Cells in Simian Immunodeficiency Virus-Infected Macaques

2005 ◽  
Vol 79 (8) ◽  
pp. 4877-4885 ◽  
Author(s):  
Yvonne M. Mueller ◽  
Constantinos Petrovas ◽  
Paul M. Bojczuk ◽  
Ioannis D. Dimitriou ◽  
Brigitte Beer ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Nk Cells ◽  
Peripheral Blood ◽  
Low Dose ◽  
Effector Memory ◽  
High Dose ◽  
Immunodeficiency Virus ◽  
Interleukin 15

ABSTRACT Interleukin-15 (IL-15) in vitro treatment of peripheral blood mononuclear cells (PBMC) from human immunodeficiency virus (HIV)-infected individuals specifically enhances the function and survival of HIV-specific CD8+ T cells, while in vivo IL-15 treatment of mice preferentially expands memory CD8+ T cells. In this study, we investigated the in vivo effect of IL-15 treatment in 9 SIVmac251-infected cynomolgus macaques (low dose of IL-15, 10 μg/kg of body weight, n = 3; high dose of IL-15, 100 μg/kg, n = 3; control [saline], n = 3; dose administered twice weekly for 4 weeks). IL-15 treatment induced a nearly threefold increase in peripheral blood CD8+CD3− NK cells. Furthermore, CD8+ T-cell numbers increased more than twofold, mainly due to an increase in the CD45RA−CD62L− and CD45RA+CD62L− effector memory CD8+ T cells. Expression of Ki-67 in the CD8+ T cells indicated expansion of CD8+ T cells and not redistribution. IL-15 did not affect CD4+ T-cell, B-cell, and CD14+ macrophage numbers. No statistically significant differences in changes from baseline in the viral load were observed when control-, low-dose-, and high-dose-treated animals were compared. No clinical adverse effects were observed in any of the animals studied. The selective expansion of effector memory CD8+ T cells and NK cells by IL-15 further supports IL-15's possible therapeutic use in viral infections such as HIV infection.

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