Late diversification in the clonal composition of human cytomegalovirus-specific CD8+ T cells following allogeneic hemopoietic stem cell transplantation

Blood ◽  
2003 ◽  
Vol 102 (9) ◽  
pp. 3427-3438 ◽  
Author(s):  
Maher K. Gandhi ◽  
Mark R. Wills ◽  
Georgina Okecha ◽  
Elizabeth K. Day ◽  
Ray Hicks ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
Hemopoietic Stem Cell ◽  
Transplant Recipients ◽  
Hemopoietic Stem Cell Transplantation ◽  
Clonal Composition

Abstract To investigate the mechanisms of human T-cell reconstitution following allogeneic hemopoietic stem cell transplantation (alloSCT), we analyzed the clonal composition of human cytomegalovirus (HCMV)-specific or Epstein-Barr virus (EBV)-specific CD8+ T cells in 10 alloSC transplant recipients and their donors. All virus-specific CD8+ T-cell clones isolated from recipients after alloSCT contained DNA of donor origin. In all 6 D+/R+ sibling alloSCTs from seropositive donors into seropositive recipients, donor virus-specific clones transferred in the allograft underwent early expansion and were maintained long term in the recipient. In contrast, in 2 of 3 HCMV D+/R- alloSC transplant recipients in whom there was no detectable HCMV infection, donor HCMV-specific clones were undetectable, whereas donor EBV-specific clones were maintained in the same EBV-seropositive recipients, suggesting that transferred clones require antigen for their maintenance. Following D-/R+ transplantation from 3 seronegative donors into seropositive recipients, a delayed primary virus-specific CD8+ T-cell response was observed, in which the T cells contained donor DNA, suggesting that new antigen-specific T cells arose in the recipient from donor-derived progenitors. In 2 of 4 HCMV D+/R+ sibling allograft recipients the clonal composition underwent diversification as compared with their donors, with delayed persistent expansion of HCMV-specific clones that were undetectable in the donor or in the recipient during the early months after transplantation; this diversification may represent expansion of new clones generated from donor-derived progenitors. We conclude that, following alloSCT, late diversification of the HCMV-specific CD8+ T-cell clonal repertoire can occur in response to persistent viral antigen.

scholarly journals Donor T Cells Maintain Myeloma-Immune Equilibrium after Autologous Stem Cell Transplantation and Concurrent Immunotherapy Promotes Cure

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 2031-2031
Author(s):  
Simone A Minnie ◽  
David Smith ◽  
Kate H Gartlan ◽  
Thomas S Watkins ◽  
Kate A Markey ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Autologous Stem Cell Transplantation ◽  
Median Survival ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
Post Transplant

Abstract Autologous stem cell transplantation (ASCT) remains an important consolidation treatment for multiple myeloma (MM) patients, even in the era of novel agents. The prolongation of plateau-phase induced by ASCT is generally attributed to intensive cytoreduction. However, ASCT generates inflammation and profound lymphodepletion, which may result in hitherto unexpected immunological effects. To investigate potential immunological contributions to myeloma control after ASCT, we developed preclinical models of transplantation for MM using Vk*MYC myeloma that generates bony lytic lesions, a serum M band and marrow plasmacytosis that are hallmarks of clinical disease. Myeloma-bearing B6 recipients underwent myeloablative conditioning and were transplanted with naïve B6 bone marrow (BM) grafts with or without T cells from donors that were myeloma-naïve (SCT) or had low M bands at the time of harvest to mimic ASCT. Surprisingly, we demonstrate the broad induction of T cell-dependent myeloma control with enhanced median survival in recipients of grafts containing T cells compared to T cell depleted (TCD) BM alone (SCT= 91 days and ASCT > 100 days post-transplant vs TCD BM alone= 44 days; p<0.0001). Myeloma was most efficiently controlled when recipients were transplanted with memory T cells (CD44+) from autologous grafts (median survival: ASCT-CD44+ T cells >90 days post-transplant vs. CD44─ T cells = 50 days; p = 0.0006). Importantly, T cells adoptively transferred from recipients surviving > 120 days (MM-primed) protected secondary recipients compared to T cells from naïve donors (median survival: MM-primed > 120 days post-transplant vs 65 days naïve T cells; p = 0.0003). Furthermore, MM-primed CD8 T cells were restricted in TCR repertoire and provided protection in a myeloma clone-specific fashion, indicative of a tumor-specific T cell response. Despite this immune-mediated control of myeloma after SCT, progression still occurred in the majority of recipients. We phenotyped CD8+ T cells from the BM of MM-relapsed, MM-controlled and MM-free (that had never seen myeloma) mice 8 weeks after SCT. Expression of the inhibitory receptors, programmed cell death protein 1 (PD-1) and T cell immunoreceptor with Ig and ITIM domains (TIGIT) on BM CD8+ T-cells strongly correlated with myeloma cell number (r = 0.729, p<0.0001 and r = 0.796, p<0.0001 respectively). Additionally, the co-stimulatory/adhesion receptor CD226 (DNAM-1) was markedly downregulated as myeloma progressed (r = - 0.865, p<0.0001), as was interferon-γ secretion (r = - 0.76, p = 0.0022). t-SNE analysis confirmed an irreversible exhaustion signature at myeloma progression, characterized by the absence of DNAM-1 and co-expression of PD-1, TIM-3, TIGIT together with CD101 and CD38. Immune-checkpoint inhibition (CPI) early post-SCT, using antibodies against PD-1 or TIGIT facilitated long-term myeloma control (median survival in both treatment arms > 120 days post-SCT vs. 60 and 68 days respectively; p <0.05). Furthermore, TIGIT blockade limited CD8+ T cell exhaustion, increased CD107a and IFNγ secretion and expanded a memory CD8+ T cell population in the BM. Genetic deletion of either IFNγ or the IFNγ receptor from the donor graft resulted in dramatic myeloma progression after SCT. Consequently, treatment with a CD137 (4-IBB) agonist early after SCT profoundly augmented CD8+IFNγ+GranzymeB+ T-cell expansion in the BM, such that majority of treated animals eliminated myeloma and survived long-term. These data provide insights into an unappreciated mechanism of action of ASCT whereby myeloma immune-equilibrium is established and suggest that combination with immunotherapeutic strategies is a rational approach to generate long term disease control. Disclosures Smyth: Bristol Myers Squibb: Other: Research agreement; Tizona Therapeutics: Research Funding.

Immune Reconstitution Following Myeloablative Allogeneic Hematopoietic Stem Cell Transplantation: The Impact of Expanding CD28negCD8+ T-Cells on Relapse.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1146-1146
Author(s):  
Ibrahim Yakoub-Agha ◽  
Pasquine Saule ◽  
Julia Salleron ◽  
Pascale Cracco ◽  
Valerie Coiteux ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
Immune Recovery ◽  
Clinical Complication ◽  
The Impact

Abstract Allogeneic stem cell transplantation has become standard therapy for haematological malignancies through the positive immunologic graft-versus-leukaemia effect. Initial immune recovery relies on peripheral expansion of infused T-cells which switch to a memory-like phenotype. This study prospectively investigated whether changes in subset composition precedes late complications after myeloablative HLA-matched transplantation. Of 80 recipients, 51 experienced neither early infection nor acute graft-versus-host disease (GVHD), of whom 18 were still free of clinical complication throughout 395 – 1564 days of follow-up. Compared with this complication-free subgroup, patients who developed chronic GVHD as the only event recovered similar numbers of circulating T-cells with predominance of CD8+ T-cells lacking CC-chemokine receptor-7 and CD28 expression. Conversely, poor CD8+ T-cell recovery with diminished numbers of CD28neg CD8+ T-cells (~1/4th of that of relapse-free patients) preceded occurrence of relapse. In multivariate analysis, lower CD28neg CD8+ T-cell counts by day 60 were associated with greater risk of subsequent relapse (HR 0.33; 95% CI 0.14 - 0.76; P = 0.01). Enumeration of CD28neg CD8+ T-cells in patients without early clinical complication could assist in predicting risk of relapse and help build an algorithm for accelerating the immune recovery by reducing the immunosuppressive regimen and considering the introduction of prophylactic donor lymphocyte infusions.

Transfer of Specific T Cell Clones with Graft-Versus-Leukemia (GVL) Activity from Donor to Recipient in Stem Cell Transplantation: Identification, Quantification and Characterization.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2958-2958 ◽  
Author(s):  
Katayoun Rezvani ◽  
David A. Price ◽  
Jason Brenchley ◽  
Yasemin Kilical ◽  
Emma Gostick ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
T Cell Clones ◽  
Memory Phenotype ◽  
Cell Clones ◽  
Healthy Donors

Abstract The self-antigens PR1 and WT1 that are aberrantly expressed on malignant cells may be important target antigens for GVL effects from donor-derived anti-leukemia T cells. It is now clear that T cells recognizing these antigens circulate in transplant recipients and can be detected in small numbers in healthy individuals. To determine whether the same T cell clones in the donor are transferred to the recipient and induce GVL effects we sought for presence of leukemia-reactive T cell clones in healthy donors and their transfer into the patient after transplant and following DLI. We identified CD8+ T cell clones specific for PR1 and WT1 from 2 healthy donors. The HLA-A2/PR1-binding and HLA-A2/WT1-binding CD8+ T cells were purified by flow cytometric cell sorting and analyzed for their T cell receptor (TCR) usage by template switch anchored RT-PCR. This showed an oligoclonal population of WT1-specific CD8+ T cells and a polyclonal population of PR1-specific CD8+ T cells. In addition, using a fluorescent peptide/MHC class I multimeric complex incorporating mutations in the a3 domain that abrogate binding to the CD8 coreceptor, we selectively isolated WT1-specific CD8+ T cells of high functional avidity and demonstrated that high avidity T cells comprise a single clonotype. One patient with CML received an alloSCT from the donor in whom PR1-specific CD8+ T cell clones were detected. Using quantitative real-time PCR for IFN-g production and HLA-A2/PR1 tetrameric complexes, we showed the emergence of PR1-specific CD8+ T cells in the blood of the recipient 10 weeks after SCT and again 8 weeks post-DLI given to treat a molecular relapse of CML. HLA-A2/PR1 tetramer-positive CD8+ T cells were sorted by flow cytometry post-alloSCT and again following DLI. By comparing TCRb CDR3 sequences, we confirmed direct transfer and expansion of PR1-specific CD8+ T cell clones from the donor into the recipient and the reemergence of the same PR1-specific clones following DLI. The appearance of these HLA-A2/PR1 tetramer-positive CD8+ T cells was followed by complete molecular remission of CML by sensitive PCR for BCR/ABL The PR1-specific CD8+ T cells in the donor were of memory phenotype and expanded in the recipient after both alloSCT and DLI. During the early phase post-transfer in the recipient, the majority of PR1-specific CD8+ T cells had an effector memory phenotype (CD45RO+ CD57+). There was a shift towards a central memory phenotype (CD45 RO+ CD57−) during the course of the GVL effect. This is the first direct demonstration of the transfer of leukaemia-reactive specific T cell clones from a healthy donor to a patient with leukemia. Further, the identification and monitoring of T cell clones that mediate the GVL effect as described here can be undertaken before stem cell transplantation and could aid donor selection.

Analysis of TREC (T Cell Receptor Excision Circles) Levels in CD94 Expressing CD8 T Cells in Chronic GVHD.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5367-5367
Author(s):  
Noriaki Iwao ◽  
Junji Tanaka ◽  
Naoko Kato ◽  
Takeshi Kondo ◽  
Yoko Miura ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
T Cell Receptor ◽  
Cd8 T Cells ◽  
Chronic Gvhd ◽  
Cd8 T Cell ◽  
Cell Receptor ◽  
Excision Circles

Abstract CD94 is one of the C-type lectin family members, forms a heterodimer with NKG2 gene family, and CD94 /NKG2A are inhibitory receptors. Not only NK cells but a subset of T cells express CD94/NKG2A, and previously we revealed the proportion of CD94/NKG2A expressing CD8 T cells were higher in patients with chronic graft versus host disease (GVHD), and CD94 expressing T cells have suppressive effects on mixed lymphocyte culture(MLC). We focus on CD94 positive T cell during T cell reconstitution after allogeneic hematopietic stem cell transplantation (allo-HSCT). T cell receptor excision circles (TREC) are suggested to be a useful marker of recent thymic output. In this study, we attempt to study TREC-containing CD8 T cell subset expressing CD94, and to examine the relation of TREC DNA level in CD94 expressing CD8 T cell and GVHD. We analyzed peripheral blood mononuclear cells (PBMCs) isolated from 24 patients (82 samples) undergone allo-HSCT including 15 patients with bone marrow transplantation and 9 patients with non-myeloablative stem cell transplantation. Informed consent was obtained from all patients. CD4 positive T cells were separated from PBMCs by magnetic cell sorting, and CD4 negative cell population was divided into CD94 positive CD8 T cells and CD94 negative CD8 T cells by fluorescence activated cell sorter. Genomic DNA was extracted from these separated T cell subsets. TREC DNA copy numbers per 105 isolated T cells (TREC level) were quantified by real time PCR. We investigated TREC levels in clinical status with pre-allo-HSCT, no episodes of GVHD or before manifestation of GVHD (No GVHD), chronic GVHD on disease (C-GVHD), and no symptoms and remission status of GVHD after immunosuppressive therapy (R-GVHD). Statical analyses were carried out by Mann-Whitney U test. There were no significant differences in TREC level of sorted CD4 positive T cells in C-GVHD compared with No GVHD (p=0.75) and R-GVHD (p=0.61), and also CD94 negative CD8 T cells in C-GVHD compared with No GVHD (p=0.79) and R-GVHD (p=0.20). On the other hand, TREC level of CD94 positive CD8 T cells in C-GVHD decreased in comparison with No GVHD (p=0.015) and R-GVHD (p=0.0019). The reduction of TREC level is thought to be induced not only by low thymic output but also by dilution of TREC concentration due to peripheral T cell expansion without duplications of TREC. These results may suggest that CD94 positive T cells play a role in modulation of GVHD, and proliferate during chronic GVHD with dilution of TREC in CD94 positive CD8 T cells. It is suggested that TREC level of CD94 expressing CD8 T cells may be useful markers of chronic GVHD.

Streptamer Technology for the Assessment of CMVpp65 Specific CD8+ T Cell Frequencies and for the Adoptive T Cell Transfer to Post-Transplant Patients.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1964-1964
Author(s):  
Anita Schmitt ◽  
Junxia Yao ◽  
Hermann Einsele ◽  
Ulrich Grigoleit ◽  
Dirk Busch ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
Cell Transfer ◽  
Adoptive T Cell Transfer ◽  
T Cell Transfer ◽  
Cmv Antigenemia

Abstract Cytomegalovirus (CMV) reactivation constitutes a serious complication after allogeneic peripheral blood stem cell transplantation (PBSCT). The frequency of CMVpp65 specific CD8+ T cells is pivotal for the clearance of CMV. CMVpp65 specific CD8+ T cell frequencies can be measured using tetra-, penta- and streptamer technologies, streptamers can also be applied therapeutically. In donors, these frequencies might allow us to define the best available donor in addition to the mere serostatus. In the present study we investigated the specificity and sensitivity of all three methods and compared the results to the serostatus. A therapeutical application, i.e. an adoptive transfer of CMV specific CD8+ T cells selected by streptamer technology to a patient with acute lymphatic leukemia suffering from life-threatening CMV antigenemia after allogeneic PBSCT was performed. 23 samples from CMV seropositive healthy volunteers (HV) and 10 samples from CMV seropositive patients before and after allogeneic stem cell transplantation (all HLA-A2 or -B7 positive) were analyzed with tetra-, penta- or streptamer conjugated to PE by flow cytometry. Our lab took part in an inter-lab CMV multimer assay including 20 European countries in the framework of www.kimt.de. For the adoptive T cell transfer a donor leukapheresis was performed followed by an HLA-B7 CMVpp65 streptamer positive selection. The patient received 2×10E5 CMV specific CD8+ T cells per kg body weight as a single transfusion. Optimal amounts of HLA-A2 multimers to stain a pellet of 10E6 cells were 0.44 mcg tetramer, 0.15 mcg pentamer and 0.2 mcg MHC/0.3 mcg streptactin complex. Surprisingly, only in 48% (11/23) seropositive HV CD8+ multimer+ T cells could be detected. The ALL patient developed a foscarvir resistant CMV antigenemia with a maximum of 959/500,000 CMVpp65 positive cells. After a switch to ganciclovir/valganciclovir and an adoptive transfer of CMV specific T cells, the antigenemia was cleared. Valganciclovir was discontinued, but CMV antigenemia remained controlled. The frequency of CMVpp65 specific CD8+ T cells increased dramatically from 0.0% till 19.8%. All of these T cells were donor derived as demonstrated by small tandem repeat (STR) analysis. The patient did not develop signs of CMV disease at any time point. This study demonstrates the power of multimer staining to define appropriate donors for transplantation. Donors should be screened for their CMVpp65 specific CD8+ T cell frequency. All three multimer technologies can be used yielding similar results. The streptamer technology additionally offers the advantage to select CMVpp65 specific CD8+ T cells at the GMP level for adoptive T cell transfer and can induce long-lasting CD8+ T cell responses effectively clearing even a high virus load.

scholarly journals Characterization of human cytomegalovirus peptide–specific CD8+ T-cell repertoire diversity following in vitro restimulation by antigen-pulsed dendritic cells

Blood ◽  
2002 ◽  
Vol 99 (1) ◽  
pp. 213-223 ◽  
Author(s):  
Karl Peggs ◽  
Stephanie Verfuerth ◽  
Arnold Pizzey ◽  
Jenni Ainsworth ◽  
Paul Moss ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Viral Replication ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
T Cell Repertoire ◽  
Cell Repertoire ◽  
Repertoire Diversity ◽  
Clonal Composition

Under conditions of impaired T-cell immunity, human cytomegalovirus (HCMV) can reactivate from lifelong latency, resulting in potentially fatal disease. A crucial role for CD8+ T cells has been demonstrated in control of viral replication, and high levels of HCMV-specific cytotoxic T-lymphocytes are seen in immunocompetent HCMV-seropositive individuals despite very low viral loads. Elucidation of the minimum portion of the anti-HCMV T-cell repertoire that is required to suppress viral replication requires further study of clonal composition. The ability of dendritic cells to take up and process exogenous viral antigen by constitutive macropinocytosis was used to study HCMV-specific T-cell memory in the absence of viral replication. The specificity and clonal composition of the CD8+ T-cell responses were evaluated using HLA tetrameric complexes and T-cell receptor β chain (TCRBV) spectratypic analyses. There was a skewed reactivity toward the matrix protein pp65, with up to 40-fold expansion of CD8+ T cells directed toward a single peptide-MHC combination. Individual expansions detected on TCRBV spectratype analysis were HCMV-specific and composed of single or highly restricted numbers of clones. There was preferential TCRBV gene usage (BV6.1/6.2, BV8, and BV13 in HLA-A*0201+ individuals) but lack of conservation of CDR3 length and junctional motifs between donors. While there was a spectrum of TCR repertoire diversity directed toward individual MHC-peptide combinations between donors, a relatively small number of clones appeared to predominate the response in each case. These data provide further insight into the range of anti-HCMV responses and will aid the design and monitoring of adoptive immunotherapy protocols.

scholarly journals T-cell receptor-α repertoire of CD8+ T cells following allogeneic stem cell transplantation using next-generation sequencing

Haematologica ◽  
2018 ◽  
Vol 104 (3) ◽  
pp. 622-631 ◽  
Author(s):  
Cornelia S. Link-Rachner ◽  
Anne Eugster ◽  
Elke Rücker-Braun ◽  
Falk Heidenreich ◽  
Uta Oelschlägel ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
Next Generation Sequencing ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
T Cell Receptor ◽  
Cd8 T Cells ◽  
Allogeneic Stem Cell Transplantation ◽  
Cell Receptor ◽  
Generation Sequencing

WT1-Specific CD8+ T Lymphocytes May Participate in the Elimination of Acute Lymphoblastic Leukemia Following Allogeneic Stem Cell Transplantation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3679-3679 ◽  
Author(s):  
Katayoun Rezvani ◽  
Agnes Yong ◽  
Stephan Mielke ◽  
Bipin N. Savani ◽  
David A. Price ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Cd8 T Cells ◽  
Lymphoblastic Leukemia ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
Wt1 Gene ◽  
Cell Responses

Abstract There is clinical evidence that a graft-versus-leukemia (GVL) effect occurs following allogeneic stem cell transplantation for acute lymphoblastic leukemia (ALL). However, the potency of this GVL effect is often associated with unwanted graft-versus-host-disease (GVHD) and disease relapse remains a major contributor to treatment failure. Wilms’ tumor gene 1 (WT1) is overexpressed in 70–90% of cases of ALL and has been identified as a convenient minimal residual disease (MRD) marker. WT1 is an attractive immunotherapeutic target in ALL because peptides derived from WT1 can induce CD8+ T-cell responses, and being non-allelic, WT1 would be unlikely to provoke GVHD. We investigated whether CD8+ T-cells directed against an HLA-A*0201 restricted epitope of WT1 (WT126) occur in ALL patients during the early phase of immune reconstitution post-SCT (days 30–180). We analyzed CD8+ T-cell responses against WT1 in 10 HLA-A*0201+ ALL SCT recipients and their respective donors using WT1/HLA-A*0201 tetrameric complexes and flow cytometry for intracellular IFN-gamma. We studied the kinetics WT1-specific CD8+ T-cell responses in consecutive samples obtained post-SCT. CD8+ T-cells recognizing WT1 were detected ex vivo in samples from 5 of 10 ALL patients post-SCT but not in patients pre-SCT. WT1-tetramer+ CD8+ T cells had a predominantly effector memory phenotype (CD45RO+CD27−CD57+). WT1 gene expression in pre-SCT and donor samples was assayed by quantitative real-time PCR (RQ-PCR). WT1 expression in PBMC from healthy donors was significantly lower than in patients (median 0, range 0–66 ×10−4 WT1/ABL compared to patients, median 12, range 0–2275 ×10−4 WT1/ABL) (P < 0.01). There was a strong correlation between the emergence of WT1-specific CD8+ T cells and a reduction in WT1 gene expression (P < 0.001) (as depicted below) suggesting direct anti-ALL activity post-SCT. Disappearance of WT1-specific CD8+ T-cells from the blood coincided with reappearance of WT1 gene transcripts, consistent with a molecular relapse, further supporting the direct involvement of WT1-specific CD8+ T-cells in the GVL response. These results provide evidence for the first time of spontaneous T-cell reactivity against a leukemia antigen in ALL patients. Our results support the immunogenicity of WT1 in ALL patients post-SCT and a potential application for WT1 peptides in post-transplant immunotherapy of ALL. 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.

Myeloid Chimerism Reflects Engraftment of Donor Hematopoiesis, Whereas T Cell Chimerism Reflects Survival and Expansion of Donor and Recipient Residual Mature T Cells Early After T Cell Depleted Allogeneic Stem Cell Transplantation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4475-4475
Author(s):  
Jessica C. Harskamp ◽  
Esther H.M. van Egmond ◽  
Hans L. Vos ◽  
Stijn J.M. Halkes ◽  
Roel Willemze ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cd8 T Cells ◽  
Nk Cell ◽  
Median Frequency ◽  
Cell Compartment ◽  
Hematopoietic Lineage ◽  
Chimerism Analysis

Abstract Abstract 4475 Allogeneic stem cell transplantation (alloSCT) is frequently complicated by life-threatening graft versus host disease (GVHD). Previous studies demonstrated that T cell depletion (TCD) of the graft significantly decreases the incidence and severity of GVHD, and is associated with a higher percentage of patients with mixed chimerism (MC). In most studies chimerism analysis is performed on the total bone marrow (BM) leukocyte fraction, and changes in chimerism are related to engraftment. In this study we investigated whether MC in the total BM leukocyte fraction truly reflects engraftment or if it is influenced by survival and expansion of donor and recipient residual mature T cells, and whether hematopoietic lineage specific chimerism analysis is therefore a better method to determine engraftment. It is likely that chimerism analysis of the stem cell compartment is best reflected in peripheral blood (PB) in those cells that are continuously produced and short lived, such as monocytes and granulocytes, and therefore PB myeloid chimerism primarily reflects engraftment. In contrast, previous studies have shown by T cell receptor excision circle analysis that T cell neogenesis is virtually absent in the first 6 months after alloSCT, and that predominantly memory T cells are present in PB and BM. Therefore, we hypothesize that MC of these long lived T cells merely reflects survival and expansion of recipient and donor residual T cells. Since the life span of B and NK cells is longer than myeloid cells, but shorter than T cells, we anticipate that in the first 6 months after alloSCT, B and NK cell chimerism reflects a combination of survival and neogenesis. To analyze these hypotheses we performed hematopoietic lineage specific chimerism analysis on PB cells of 22 patients (median age 52 years, range 23-73, 11 males) receiving a TCD alloSCT between June and November 2008 after a myeloablative (n=11) or non myeloablative conditioning regimen (n=11) for AML, ALL, high risk MDS, multiple myeloma, CML, CLL or NHL. At intervals of 6 weeks PB was collected, and monocytes, granulocytes, B and NK cells, CD4+ and CD8+ T cells were sorted. The total leukocyte fraction was obtained by erythrocyte lysis of BM. DNA was isolated to perform chimerism analysis using short tandem repeats - PCR. Our results show that in the BM leukocyte fraction 47% of the patients were MC at 3 months after alloSCT, with a median frequency of patient cells of 4%. However, of the patients with MC in the total leukocyte fraction, 67% was complete chimeric in the myeloid subsets and MC in the T cell compartment. In the PB myeloid subsets (monocytes and granulocytes) less than 28% of the patients were MC during the first 6 months after alloSCT with a median frequency of patient cells less than 5%. In the B and NK cell subsets, at most time points more patients were MC (7-43%) with higher frequencies of patient cells (2-14%) compared to the myeloid subsets. The CD4 and CD8 T cell subsets showed the highest frequencies of MC in numbers of patients (31-61%) as well as the highest MC frequencies of patient cells (13-80%). Phenotypic analysis of the T cell compartment showed that 98% of the CD4 and CD8 T cells were memory cells during the first 6 months after alloSCT. Preliminary data indicate that the median percentage of donor derived T cells increased during the first 6 months after alloSCT, correlating with development of mild GVHD, suggesting that T cell chimerism is influenced by immunogenic triggers. In conclusion, these results illustrate that for engraftment and neogenesis of donor hematopoiesis, myeloid chimerism analysis provides more accurate information than total BM leukocyte chimerism analysis, since the results are greatly influenced by T cell chimerism. Since almost all T cells were memory cells within the first 6 months after alloSCT, T cell chimerism analysis reflects survival and expansion of mature donor as well as recipient T cells, and can therefore not be used to measure engraftment. Disclosures: No relevant conflicts of interest to declare.

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