Augmentation of Post-Transplant Immunity: Antigen Encounter at Time of Hematopoietic Stem Cell Transplantation Enhances Antigen-Specific Donor T Cell Responses in the Post-Transplant Repertoire.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2246-2246
Author(s):  
Craig A. Mullen ◽  
Ulker Kocak ◽  
Joanne L. Shaw ◽  
Shahram Mori
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Function ◽  
T Cell Responses ◽  
Spleen Cells ◽  
Hematopoietic Stem ◽  
Cd8 Cells ◽  
Post Transplant ◽  
Cell Responses ◽  
Donor T Cells

Abstract After transplant the immune system is reconstituted by cells derived from both hematopoietic stem cells and peripheral expansion of differentiated donor T cells. Immune function is poor despite transplantation of mature lymphocytes from immune competent donors. We tested the hypothesis that early antigen encounter at the time of cell transplant would enhance desired donor T cell responses in the post-transplant repertoire. 2 independent models of peptide-specific T cell responses were studied. Model 1 : The model for CD4 cells employed T cells from transgenic DO11.11 mice that constitutively express the T cell receptor for the class II restricted ovalbumin (OVA) peptide 323–339. Fig 1: Early exposure to OVA antigen enhances clonal expansion of OVA specific transgenic T-cells following syngeneic BMT. Lethally irradiated BALB/c mice were injected with 300 μg of OVA peptide in CFA or CFA alone subcutaneously one day before transplantation (D-1). The transplanted mice received 2x106 transgenic OVA specific T-cells and 6x106 non-transgenic naive BALB/c bone marrow cells. At 2 days (A) and 7weeks (B) following BMT, draining lymph nodes were isolated and examined for the presence of OVA-specific T-cells using FITC-labeled KJ-126 antibody and PE-labeled anti mouse CD4 antibody. Naïve BALB/c animals were used as negative controls (C). The absolute number of antigen-specific T-cells was determined by multiplying the total cells recovered with the percentage of OVA-specific CD4+ T-cells identified by flow. Figure Figure Model 2: The model for CD8 cells employed nontransgenic H2-Db-restricted T cell responses to the influenza nucleoprotein peptide 366–374. Fig 2: Antigen specific CD8+ cells in antigen-exposed animals are functionally active. Donor SW mice were immunized three times by ip injection of virus-infected spleen cells. Recipient C57BL/6 animals underwent BMT using influenza-immune donors spleen cells and bone marrow (10x106 and 4x106 respectively). Some transplant recipients were exposed to influenza virus on D-1. Ten days following BMT, the animals were sacrificed and spleens were isolated and stimulated in vitro with 2 μg of NP peptide. After two rounds of stimulation, the splenocytes were assayed by intracellular cytokine assay for the secretion of IFNg by staining with PE-anti IFNγ and FITC-anti-CD8 antibodies. The results are representative of three experiments (total number n=4/experimental group). Figure Figure Encounter with specific antigen at the time of T cell transplantation led to clonal expansion of donor T cells and preservation of donor T cell function in the post-transplant immune environment. Antigen-specific donor T cell function was poor if antigen encounter was delayed or omitted. Severe parent>F1 graft versus host reactions blocked the effect of early antigen exposure.

scholarly journals HCoV- and SARS-CoV-2 Cross-Reactive T Cells in CVID Patients

2020 ◽  
Vol 11 ◽  
Author(s):  
Sophie Steiner ◽  
Franziska Sotzny ◽  
Sandra Bauer ◽  
Il-Kang Na ◽  
Michael Schmueck-Henneresse ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd4 T Cells ◽  
T Cell Responses ◽  
Cross Reactivity ◽  
T Cell Immunity ◽  
Cell Reactivity ◽  
Cd8 Cells ◽  
Cell Responses ◽  
Cell Immunity

The inability of patients with CVID to mount specific antibody responses to pathogens has raised concerns on the risk and severity of SARS-CoV-2 infection, but there might be a role for protective T cells in these patients. SARS-CoV-2 reactive T cells have been reported for SARS-CoV-2 unexposed healthy individuals. Until now, there is no data on T cell immunity to SARS-CoV-2 infection in CVID. This study aimed to evaluate reactive T cells to human endemic corona viruses (HCoV) and to study pre-existing SARS-CoV-2 reactive T cells in unexposed CVID patients. We evaluated SARS-CoV-2- and HCoV-229E and –OC43 reactive T cells in response to seven peptide pools, including spike and nucleocapsid (NCAP) proteins, in 11 unexposed CVID, 12 unexposed and 11 post COVID-19 healthy controls (HC). We further characterized reactive T cells by IFNγ, TNFα and IL-2 profiles. SARS-CoV-2 spike-reactive CD4+ T cells were detected in 7 of 11 unexposed CVID patients, albeit with fewer multifunctional (IFNγ/TNFα/IL-2) cells than unexposed HC. CVID patients had no SARS-CoV-2 NCAP reactive CD4+ T cells and less reactive CD8+ cells compared to unexposed HC. We observed a correlation between T cell reactivity against spike of SARS-CoV-2 and HCoVs in unexposed, but not post COVID-19 HC, suggesting cross-reactivity. T cell responses in post COVID-19 HC could be distinguished from unexposed HC by higher frequencies of triple-positive NCAP reactive CD4+ T cells. Taken together, SARS-CoV-2 reactive T cells are detectable in unexposed CVID patients albeit with lower recognition frequencies and polyfunctional potential. Frequencies of triple-functional reactive CD4+ cells might provide a marker to distinguish HCoV cross-reactive from SARS-CoV-2 specific T cell responses. Our data provides evidence, that anti-viral T cell immunity is not relevantly impaired in most CVID patients.

Simulataneous Vaccination and Delayed Lymphocyte Infusion for Enhancement of Immune Reactivity Against Acute Lymphoblastic Leukemia or Lymphoma.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5248-5248
Author(s):  
Craig A. Mullen ◽  
Olga Sevastianova
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
T Cell ◽  
Lymphoblastic Leukemia ◽  
T Cell Responses ◽  
Myelogenous Leukemia ◽  
Lymphoid Tissues ◽  
Spleen Cells ◽  
Hematopoietic Stem ◽  
Allogeneic Hsct ◽  
Cell Responses

Abstract BACKGROUND: Delayed lymphocyte infusion (DLI) can be employed as an antileukemia immune therapy when patients with leukemia relapse after allogeneic hematopoietic stem cell transplantation (HSCT). While effective in patients with chronic myelogenous leukemia, it is generally ineffective with relapse of acute lymphoblastic leukemia (ALL). This ineffectiveness may be multifactorial and possible reasons include rapid leukemia cell growth in ALL, reduced propensity to apoptosis signals, and poor presentation of antigens present on lymphoblasts resulting in failure to generate a significant immune response. If the failure of malignant lymphoblasts in marrow and lymphoid tissues to elicit meaningful T cell responses plays a role in the ineffectiveness of DLI, it is possible that therapeutic vaccines may be able to ameliorate this barrier. We have recently demonstrated that vaccination immediately prior to HSCT enhances antigen specific T cell responses in the post-transplant immune repertoire (Bone Marrow Transplantation, 35:793–801, 2005.) HYPOTHESIS: Vaccination with leukemia related antigens at the time of DLI will increase donor T cell responses to leukemia related antigens without exacerbation of GVHD responses to ubiquitous host minor histocompatibility antigens (mHA). METHODS: We employ a murine MHC matched allogeneic HSCT model in which 6 antigens are molecularly characterized. C3.SW mice are donors and C57BL/6 mice are recipients. In this model the H7 mHA is the immunodominant antigen, while alloresponses to H3 and H13 are also present. By using spontaneously arising lymphoblastic leukemia/lymphoma cells from Ig-cmyc transgenic male C57BL/6 as the model malignancy in a female to female transplant the male HY associated antigens Uty, Dby and Smcy can be used as antigens restricted to the lymphoid malignancy. Four weeks after allogeneic HSCT mice were vaccinated with 107 cells expressing the HY antigens and the C57BL/6 H7, H3 and H13 minor antigens. The cells used for pre-DLI vaccination were either irradiated male C57BL/6 spleen cells or irradiated male malignant lymphoblasts. One day later mice received iv infusion of 107 spleen cells from C3.SW female donor mice previously primed against HY antigens by immunization with male C3.SW cells. Ten days interferon gamma Elispot assays were performed on HSCT recipient spleen cells using the peptide-defined antigens. RESULTS: Vaccination of HSCT recipients with male spleen cells 1 day prior to DLI from HY immune donors significantly increased T cell responses to HY peptide epitopes. Recipients of DLI alone harbored 40 interferon secreting cells per 106, while combination of vaccination and DLI yielded 94 per 106(p < 0.05). In contrast, there was no change in the number of T cells responding to the H7, H3 and H13 mHA. DLI only exhibited 26 per 106 while DLI plus vaccine exhibited 27 per 106 (p < 0.05). Whole cell irradiated lymphoblasts were not as effective as irradiated splenocytes in augmenting the HY responses, although they were equally effective in producing T cell responses in normal, nontransplanted mice. CONCLUSIONS: Simultaneous vaccination and DLI can lead to significant expansion of donor cells potentially reactive with antigens present on malignant lymphoblasts without exacerbation of T cell responses to ubiquitous mHA associated with GVHD. Current work is exploring methods by which indirect presentation of antigens from lymphoblasts can be enhanced, since it is unlikely that vaccines relying on direct antigen presentation by lymphoblasts will be effective.

Flagellin, a TLR5 Agonist, Reduces GvHD in Allogeneic HSCT Recipients While Enhancing Anti-Viral Immunity: A Novel Therapeutic Approach

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 144-144
Author(s):  
Mohammad S Hossain ◽  
David L Jaye ◽  
Brian P Pollack ◽  
Alton B Farr ◽  
John Roback ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Immune Responses ◽  
Cd8 T Cells ◽  
Viral Immunity ◽  
Hematopoietic Stem ◽  
Allogeneic Hsct ◽  
Post Transplant ◽  
Radiation Chimeras ◽  
Donor T Cells

Abstract Abstract 144 In MHC-mismatched allogeneic hematopoietic stem cell transplantation (allo-HSCT), host antigen specific donor T cells mediate acute and chronic graft-versus-host disease (GvHD). Based upon the radio-protective effects of flagellin, a TLR5 agonist protein (∼50 kDa) extracted from bacterial flagella, we reasoned that flagellin might modulate donor T cells immune responses toward host antigens, reduce GvHD, and improve immune responses to CMV infection in experimental models of allogeneic HSCT. Two 50mg/mouse i.p doses of highly purified flagellin were administered 3 hrs before irradiation and 24 hrs after allo-HSCT in H-2b ^ CB6F1 and H-2k ^ B6 models. GvHD scores were obtained with weekly clinical examination and with histological scoring of intestine, colon, liver and skin at necropsy. Flagellin treatment successfully protected allo-HSCT recipients from acute and chronic GvHDs after transplantation of 5×106 splenocytes and 5×106 T cell depleted (TCD) BM, and significantly increased survival compared to PBS-treated control recipients. Reduced acute GvHD was associated with significant reduction of a) early post-transplant proliferation of donor CD4+ and CD8+ T cells measured by Ki67 and CFSE staining, b) fewer CD62L+, CD69+, CD25+, ICOS-1+ and PD-1+ donor CD4+ and CD8+ T cells compared with the PBS-treated control recipients. Decreased numbers of activated and proliferating donor T cells were associated with significantly reduced pro-inflammatory serum IFN-g, TNF-a, and IL-6 on days 4–10 post transplant in flagellin-treated recipients compared with the PBS-treated recipients. Interestingly, both flagellin-treated recipients and PBS-treated recipients had over 99% donor T cell chimerism at 2 months post transplant. Moreover, MCMV infection on 100+ days post-transplant flagellin-treated mice significantly enhanced anti-viral immunity, including more donor MCMV-peptide-tetramer+ CD8+ T cells in the blood (p<0.05), and less MCMV in the liver on day 10 post infection (p<0.02) compared with the PBS-treated control recipients. Overall immune reconstitution after flagellin-treatment was robust and associated with larger numbers of CD4+CD25+foxp3+ regulatory T cells in the thymus. To further define the role of flagellin-TLR5 agonistic interactions in the reduction of GvHD, we next generated B6 ^ TLR5 KO (KO) and KOB^6 radiation chimeras by transplanting 10 × 106 BM cells from wild-type (WT) B6 or TLR5 KO donors into the congenic CD45.1+ B6 or KO recipients conditioned with 11Gy (5.5Gyx2) TBI. The radiation chimeras were irradiated again with 9.0Gy (4.5Gy × 2) on 60 days after the first transplant and transplanted with 3 × 106 splenocytes and 5 × 106 TCD BM from H-2K congenic donors. Two 50mg doses of flagellin were administered 3 hrs before irradiation and 24 hrs after HSCT. All flagellin-treated B6 ^ B6 radiation chimeras survived with only 12% weight-loss by 80 days post transplant compared with 50% survival among recipients of flagellin-treated B6 ^ KO and 40% survival among KO ^ B6 radiation chimeras. All flagellin-treated KO^ KO and PBS-treated radiation chimeras died within 65 days post transplant. These data suggested that interaction of flagellin with the TLR5 expressing host gut epithelium and donor hematopoietic cells are both required for the maximum protective effect of this TLR5 agonist on GvHD in allogeneic HSCT recipients. Together our data demonstrate that peritransplant administration of flagellin effectively controls acute and chronic GvHD while preserving enhanced post-transplant donor anti-opportunistic immunity. Since flagellin has been found to be safe for use in humans as vaccine adjuvant in a number of clinical trials, the clinical use of flagellin in the setting of allogeneic HSCT is of interest. Disclosures: No relevant conflicts of interest to declare.

scholarly journals NLRP6 in Donor T Cells Separately Regulates CD4 and CD8 Mediated Graft-Versus-Host Disease in Experimental Murine BMT

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1926-1926
Author(s):  
Masahiro Suto ◽  
Eri Matsuki ◽  
Erika Sekiguchi ◽  
Hiroya Tamaki ◽  
Isao Tawara ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Research Funding ◽  
Pharmaceutical Research ◽  
T Cell Responses ◽  
Inflammasome Activation ◽  
Activated T Cells ◽  
Cell Responses ◽  
Donor T Cells

NLRP6 (NOD-like receptor family pyrin domain containing 6) is an important inflammasome component and is highly expressed in intestinal epithelial and in immune cells. NLRP6 mediated inflammasome activation plays a critical role in response to intestinal infection and preventing dysbiosis of gut microbiota through the secretion of IL-18 and mucus. However, we recently found that NLRP6 plays a pathogenic role in GVHD that is independent of microbial dysbiosis, which is in contrast to its well-appreciated microbiome-dependent protective role in intestinal colitis and tumorigenesis. Interestingly, we also found that activated T cells increased NLRP6 expression, but the T cell autonomousrole of NLRP6 in regulating T cell responses is unknown. Because NLRP6 is an important regulator of GVH responses, we tested the hypothesis that NLRP6 deficiency in donor T cells would ameliorate GVHD. To test our hypothesis, we first performed adetailed phenotypic analysis of various T cell subsets and activation markers in naïve NLRP6-/-and wild-type (WT) B6 animals and found a similar distribution of naïve, memory, effector and regulatory T cells. In order to examine whether the absence of NLRP6 in donors affects GVHD, WT-BALB/canimals were lethally irradiated (700cGy) and transplanted on day 0 with 5x106bone marrow and 1.0x106 splenic CD90+T cells from either syngeneic WT-BALB/c, allogeneic MHC-mismatched WT-B6 or NLRP6-/-animals. Contrary to our hypothesis, the recipients receiving donor T cells from NLRP6-/-animals showed a significantly worse survival compared to allogeneic WT-B6 animals (p<0.05). GVHD mortality and severity were also increased in an MHC mismatched B6 into B10.BR model, and in an MHC mismatched haploidentical B6 into F1model (p<0.05). In contrast, GVHD severity and mortality were similar in an MHC matched multiple minor antigen mismatched B6 into C3H.sw model. We hypothesized that GVHD severity and mortality was similar in the B6 into C3H.sw model because NLRP6 regulates CD4+ and CD8+ T cell responses, differently. In order to test this, we transplanted C3H.sw recipients as above except we infused either 1x106CD4+ or CD8+ T cells from B6-WT or NLRP6-/-animals. GVHD severity and mortality (P<0.05) were enhanced only when NLRP6-/-CD4+ T cells transplanted. We confirmed enhanced GVHD mortality and severity mediated by donor NLRP6-/-CD4+ T cells in a second MHC-mismatched GVHD model, B6 into BALB/c (p<0.05). To explore how NLRP6 effects T cell responses independent ofinflammasome activation, we tested naïve T cell proliferation in vitro after allogeneic or non-specific TCR stimulation by anti-CD3 and CD28 antibody and found that NLRP6-/-CD4+ but not CD8+T cells proliferated more than WT-B6 CD4+ or CD8+ T cells, respectively, following either stimulus. Furthermore, allogeneicNLRP6-/-T cells also caused greater mortality compared to WT allogenic T cells in a non-irradiated B6 into F1 model, which lacks inflammasome activation associated with conditioning induced DAMPs and PAMPs. Microarray analysis of activated T cells from NLRP6-/-animals showed higher expression of IL-2 and IFN-γ than WT B6 T cells, and we observed no effect of NLRP6 in a Treg suppression assay. These data suggest that NLRP6 regulates CD4+ T cell- mediated immune responses and that NLRP6 in donor T cells is critical for controlling CD4+ T cell mediated GVHD. The effect of NLRP6 on T cell mediated GVL is currently under investigation. Disclosures Tawara: Kyowa Hakko Kirin: Honoraria, Research Funding; Ono Pharmaceutical: Research Funding; Astellas Pharma: Research Funding. Ishizawa:Otsuka Pharmaceutical: Research Funding; Pfizer: Research Funding; Novartis: Speakers Bureau; Bristol-Myers Squibb: Speakers Bureau.

scholarly journals Glycolysis Inhibition Induces Functional and Metabolic Exhaustion of CD4+ T Cells in Type 1 Diabetes

2021 ◽  
Vol 12 ◽  
Author(s):  
Christina P. Martins ◽  
Lee A. New ◽  
Erin C. O’Connor ◽  
Dana M. Previte ◽  
Kasey R. Cargill ◽  
...  
Keyword(s):  
T Cells ◽  
Type 1 Diabetes ◽  
T Cell ◽  
Cell Function ◽  
T Cell Responses ◽  
Metabolic Reprogramming ◽  
Cell Responses ◽  
T Cell Function

In Type 1 Diabetes (T1D), CD4+ T cells initiate autoimmune attack of pancreatic islet β cells. Importantly, bioenergetic programs dictate T cell function, with specific pathways required for progression through the T cell lifecycle. During activation, CD4+ T cells undergo metabolic reprogramming to the less efficient aerobic glycolysis, similarly to highly proliferative cancer cells. In an effort to limit tumor growth in cancer, use of glycolytic inhibitors have been successfully employed in preclinical and clinical studies. This strategy has also been utilized to suppress T cell responses in autoimmune diseases like Systemic Lupus Erythematosus (SLE), Multiple Sclerosis (MS), and Rheumatoid Arthritis (RA). However, modulating T cell metabolism in the context of T1D has remained an understudied therapeutic opportunity. In this study, we utilized the small molecule PFK15, a competitive inhibitor of the rate limiting glycolysis enzyme 6-phosphofructo-2-kinase/fructose-2,6- biphosphatase 3 (PFKFB3). Our results confirmed PFK15 inhibited glycolysis utilization by diabetogenic CD4+ T cells and reduced T cell responses to β cell antigen in vitro. In an adoptive transfer model of T1D, PFK15 treatment delayed diabetes onset, with 57% of animals remaining euglycemic at the end of the study period. Protection was due to induction of a hyporesponsive T cell phenotype, characterized by increased and sustained expression of the checkpoint molecules PD-1 and LAG-3 and downstream functional and metabolic exhaustion. Glycolysis inhibition terminally exhausted diabetogenic CD4+ T cells, which was irreversible through restimulation or checkpoint blockade in vitro and in vivo. In sum, our results demonstrate a novel therapeutic strategy to control aberrant T cell responses by exploiting the metabolic reprogramming of these cells during T1D. Moreover, the data presented here highlight a key role for nutrient availability in fueling T cell function and has implications in our understanding of T cell biology in chronic infection, cancer, and autoimmunity.

scholarly journals Thymic stromal lymphopoietin limits primary and recall CD8+ T-cell anti-viral responses

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Risa Ebina-Shibuya ◽  
Erin E West ◽  
Rosanne Spolski ◽  
Peng Li ◽  
Jangsuk Oh ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Viral Infection ◽  
Cell Function ◽  
Influenza Infection ◽  
T Cell Responses ◽  
T Cell Response ◽  
Thymic Stromal Lymphopoietin ◽  
Direct Role ◽  
Cell Responses

Thymic stromal lymphopoietin (TSLP) is a cytokine that acts directly on CD4+ T cells and dendritic cells to promote progression of asthma, atopic dermatitis, and allergic inflammation. However, a direct role for TSLP in CD8+ T-cell primary responses remains controversial and its role in memory CD8+ T cell responses to secondary viral infection is unknown. Here, we investigate the role of TSLP in both primary and recall responses in mice using two different viral systems. Interestingly, TSLP limited the primary CD8+ T-cell response to influenza but did not affect T cell function nor significantly alter the number of memory CD8+ T cells generated after influenza infection. However, TSLP inhibited memory CD8+ T-cell responses to secondary viral infection with influenza or acute systemic LCMV infection. These data reveal a previously unappreciated role for TSLP on recall CD8+ T-cell responses in response to viral infection, findings with potential translational implications.

Spectratype Analysis of In Vitro Donor Anti-Host T Cell Repertoire Responses Predict Those of In Vivo Post-BMT.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2163-2163
Author(s):  
Thea M. Friedman ◽  
Kira Goldgirsh ◽  
Jenny Zilberberg ◽  
Stephanie A. Berger ◽  
Joanne Filicko-O’Hara ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Responses ◽  
T Cell Repertoire ◽  
Cell Repertoire ◽  
Post Transplant ◽  
Cell Responses ◽  
Repertoire Analysis ◽  
Alloreactive T Cell

Abstract Immunotherapeutic strategies have gained recognition as viable alternatives to more conventional modalities for the treatment of cancer. In this regard, adoptive T cell therapy through allogeneic blood and marrow transplantation (BMT) has provided the strongest evidence that anti-tumor effects could be achieved against hematological malignancies. However, the major complications of BMT still include graft failure, opportunistic infections, disease relapse and graft-versus-host disease (GVHD). The presence of mature donor T cells in the transplant inoculum reduces the incidence of the first three complications, while unfortunately increasing the risk of GVHD, which can be directed against either HLA or minor histocompatibilty antigen (miHA) disparities. Thus, a major objective in the field has been to develop tactics that could facilitate the separation of graft-versus-tumor (GVT) effects from the deleterious effects of GVHD. One such approach would be to selectively deplete donor alloreactive T cells in the donor inoculum while allowing residual T cells to provide some protection against infection and to support a tumor-specific GVT response. For a more targeted approach, delayed donor lymphocyte infusion (DLI) of positively-selected donor GVT-reactive T cells could be used weeks to months post-transplant, if these elements were identifiable. In this regard, TCR Vβ repertoire analysis by CDR3-size spectratyping can be a powerful tool for the characterization of alloreactive T cell responses. Theoretically, molecular analysis of T cell responses in vitro, given the high sensitivity of the PCR-based spectratyping technique, should identify the most potentially critical Vβ families involved in the later development of GVHD and GVT effects in patients. To this end, we tested the hypothesis that T cell repertoire analysis of HLA-matched sibling (SIB) or matched unrelated donors (URD) from in vitro, host-stimulated, mixed lymphocyte cultures (MLC) would be predictive of the TCR-Vβ spectratype analysis of the T cell repertoire in the patient following BMT. In this study, we examined 17 patient pairs and report that for the resolvable Vβ families, we observed overall 71.2 ± 11.9% (mean ± SD.; range 40%–85%) of the in vitro anti-host T cell responses were predictive of those in the patient post-transplant. Of the 28.8% non-predictive Vβ families, 6.9 ± 6.3% (range 0%–27%) exhibited skewing in the MLC but no skewing in the patient post-transplant repertoire, 9.3 ± 6.3% (range 0%–18.8%) exhibited skewing in different peaks within the same Vβ family, and 12.5 ± 10.8% (range 0%–40%) showed skewing in the patient post-transplant and none in the MLC. Taken together, these results suggest that the in vitro MLC T cell responses show good consistency with post-transplant patient responses. Thus, in vitro spectratyping may be useful for predicting the alloreactive T cell responses involved in GVHD and could be used to guide custom-designed select Vβ family T cell-depleted transplants to improve patient outcomes. The additional advantage of this approach is that minimization of GVHD risk can be obtained without any direct knowledge of the specific miHA involved in the individual donor-patient pair.

Autologous Whole Tumor Cell Vaccination Early After Allogeneic Stem Cell Transplantation Elicits Anti-Tumor T Cell Responses in Patients with Advanced Chronic Lymphocytic Leukemia (CLL)

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1892-1892
Author(s):  
Ute E. Burkhardt ◽  
Ursula Hainz ◽  
Kristen E. Stevenson ◽  
Di Wu ◽  
Vincent T. Ho ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Cd8 T Cells ◽  
Research Funding ◽  
T Cell Responses ◽  
Lymphocytic Leukemia ◽  
Autologous Tumor ◽  
Post Transplant ◽  
Cell Responses

Abstract Abstract 1892 Patients with advanced hematological malignancies remain at high risk for eventual disease progression following reduced intensity conditioning (RIC) allogeneic hematopoietic stem cell transplantation (allo-HSCT). We hypothesized that vaccination with whole leukemia cells during the critical period of immune reconstitution early after transplant may enhance antitumor immunity and facilitate expansion of leukemia-reactive T cell responses. We tested this hypothesis in a prospective clinical trial, in which patients with advanced chronic lymphocytic leukemia (CLL) received up to 6 vaccine doses initiated between day 30–45 following RIC allo-HSCT. Each vaccine consisted of 1×107 irradiated autologous tumor cells admixed with 1×107 irradiated K562 bystander cells secreting GM-CSF (GM-K562). All patients received tacrolimus and mini-methotrexate as graft-versus-disease (GvHD) prophylaxis. Tacrolimus was maintained at therapeutic levels during the vaccination period without taper. Twenty-two patients were enrolled, all with advanced disease (median number of prior therapies 3; range 2–11). Many of the leukemias expressed markers associated with aggressive disease (e.g. unmutated IgVH - 68%) and displayed high-risk cytogenetic abnormalities (sole del(11q) - 41%; sole del(17p) - 23%; del(11q and 17p) - 18%). Greater than 50% (n=13) of patients had persistent marrow involvement (≥10%) at time of allo-HSCT. Eighteen of 22 subjects were vaccinated after allo-HSCT and received a median of 6 (range 1–6) vaccines. The remaining 4 patients were precluded from vaccination due to development of acute GvHD before day 45. Vaccines were generally well tolerated, but mild, transient injection site erythema was common. Only one grade 4 event (neutropenia) with a possible attribution to treatment occurred. We observed a similar incidence of grade II-IV aGvHD at 1 year in the 18 vaccinated patients (39%; 95% CI: 17–61%) and 42 control CLL patients that underwent RIC allo-HSCT at our institution from 2004–2009 (31%; 95%CI: 18–46%). At a median follow-up of 2.9 (range 1–4) years, the estimated 2-year rates of progression-free survival and overall survival of vaccinated study participants were 80% (95% CI: 54–92%) and 84% (95% CI: 58–95%). With these promising clinical results, we next focused on gaining insight into the mechanism that generated the observed clinical graft-versus-leukemia (GvL) responses. To delineate the specific contribution of vaccination to the overall GvL effect, we performed T cell assays to detect CLL-specific reactivity in serial pre- and post-HSCT samples obtained from vaccinated patients (n=9) who received median of 6 vaccines (range 3–6). In comparison, we examined T cell responses in study subjects (n=4) that developed aGvHD at a median of 44.5 days (range 26–56) after HSCT; and control CLL patients (n=4; no vaccine, no GvHD in the early post-transplant period) that were not enrolled in the study. Although early post-transplant vaccination had no impact on recovering absolute T cell numbers, reactivity of CD8+ T cells from the vaccinated patients was consistently directed against autologous tumor cells but not alloantigen bearing-recipient cells (PHA T cell blasts and fibroblasts) in IFNγ ELISpot assays. A peak response against autologous tumor cells was reached at day 60 after allo-HSCT (average 221 SFC/5×105 cells vs. 29 and 33 average SFC/5×105cells for PHA blasts and fibroblasts, respectively). CD8+ T cell clones were isolated from 4 vaccinated study subjects by limiting dilution and 17% (range 13–33%) reacted solely against CLL-associated antigens. In contrast, broad CD8+ T cell reactivity indicating an alloantigen response was observed in GvHD patients, while no increase in T cell reactivity against tumor-associated or alloantigens was seen in control patients. Tumor-reactive CD8+ T cells isolated from vaccinated patients secreted a broad profile of effector cytokines (GM-CSF, TNFα and IP10). Moreover, the amount of cytokines secreted by these CLL-specific CD8+ T cells steadily increased following early post-transplant vaccination, but not after allo-HSCT alone or in relation to GvHD. Our studies reveal that vaccination with autologous whole CLL/GM-K562 cells between days 30–100 after allo-HSCT is associated with induction of immunity against recipient CLL cells, and suggest that this is an effective strategy for promoting GvL following RIC allo-HSCT. Disclosures: Brown: Genzyme, Celgene: Research Funding; Calistoga, Celgene, Genentech, Pharmacyclics, Novartis, Avila: Consultancy. Cutler:Pfizer, inc: Research Funding; Astellas, Inc: Consultancy, Research Funding.

Donor T Cells Intrinsic Responses to Damps Regulated By Siglec-G-CD24 Axis Mitigate Gvhd but Maintain GVL in Experimental BMT Model

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 229-229 ◽  
Author(s):  
Tomomi Toubai ◽  
Corinne Rossi ◽  
Katherine Oravecz-Wilson ◽  
Nathan Mathewson ◽  
Cynthia Zajac ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Responses ◽  
Innate Immune ◽  
T Cell Subsets ◽  
Activation Markers ◽  
Cell Responses ◽  
Target Organs ◽  
Donor T Cells

Abstract Innate immune receptors like pattern recognition receptors (PRRs) including toll-like receptors (TLRs) and nucleotide-binding oligomerization domain (NOD) like-receptors (NLR) on immune cells play an important role in initiating inflammatory responses to damage- and pathogen- associated molecular patterns (DAMPs and PAMPs) expressed on invading pathogens or released from damaged cells. Although it is well known that DAMPs directly modulate innate immune functions, it is less clear whether DAMPs directly regulate T cell intrinsic function. Members of the sialic acid binding Ig-like lectin (Siglec) family have immunoreceptor tyrosine-based inhibitory motifs (ITIM) or ITIM-like regions in their intracellular domain that negatively regulate immune activation induced by DAMPs. Our previous data suggested that the Siglec- G-CD24 interaction in host APCs plays an important role in the negative regulation of graft-versus host (GVH) responses. However, the T cell autonomous role of Siglec-G in the regulation of T cell responses is not known. Because Siglecs are important negative regulators of immune responses, we tested the hypothesis that the deficiency of Siglec-G in donor T cells would enhance GVHD. To test our hypothesis, we first examined detailed phenotypic analysis of various T cell subsets and activation markers in naïve Siglec-G-/- and wild-type (WT) B6 animals and found similar distribution of naïve, memory, effector and regulatory T cells. In order to examine whether the absence of Siglec-G in donors affects GVHD, WT-BALB/cmice were lethally irradiated (850cGy) and transplanted on day 0 with 5x106 bone marrow and 0.5x106 splenic CD90+ T cells from either syngeneic WT-BALB/c, allogeneic MHC-mismatched WT-B6 or Siglec-G-/- animals. The recipients receiving donor T cells from Siglec-G-/- animals showed a significantly worse survival compared to allogeneic WT-B6 animals (p<0.05). This increased mortality was also associated with more severe GVHD damage in target organs and a higher expansion of activated CD69+, IFN-r+, and IL-17A+ donor T cells in the spleen and target organs. Enhanced GVHD mortality and severity was also observed in MHC mismatched haploidentical matched B6 in to F1models (p<0.05). To explore the mechanism, we tested whether Siglec-G deficiency alters the naïve T cell responses in vitro after allogeneic or non-specific TCR stimulation in the absence of exogenous DAMPs. Interestingly Siglec-G-/- T cells showed similar proliferation in vitro, when compared to WT B6 T cells. In addition, Siglec-G-/- Tregs are equally suppressive in suppression assay and Siglec-G-/- T cells showed severe GVHD even Tregs are depleted in allo-BMT. However, Siglec-G-/- T cells showed a higher proliferation after direct TCR stimulation (CD3/CD28) with addition of DAMP (HMGB-1) when compared to WT T cells in vitro, suggesting direct T cell intrinsic effects. Consistent with this result, allogeneic Siglec-G-/- T cells caused similar mortality compared to WT controls in non-irradiated B6 into F1 model due to the absence of DAMPs from conditioning. To test the critical cellular mechanisms, we examined the function of endogenous Siglec-G ligand, CD24. We utilized BALB/c CD24-/- animals as hosts in same BMT model and found that CD24-/- animals showed an enhanced GVHD mortality and severity when compared to WT animals (p<0.05). To enhance Siglec-G-CD24 axis, we utilized a novel CD24 fusion protein (CD24Fc) in same BMT model and found that CD24 Fc ameliorated GVHD severity and mortality in not only allogeneic WT-B6 animals (p<0.05) but also CD24-/- animals (p<0.05). Next we explored DAMPs regulation by Siglec-G-CD24 axis in GVL. We utilized the same model of CD24Fc treatment but added P815 at the same time of allo-BMT and found that CD24Fc treated animals showed equivalent GVL to non-treated animals, suggesting that regulation of DAMPs with CD24Fc mitigates GVHD with maintaining GVL effect. Collectively our data suggested that the expression of both Siglec-G on donor T cells and CD24 on hosts is critical for controlling GVHD in the context of DAMPs released from conditioning, and represents a novel strategy that CD24Fc can mitigates GVHD with maintaining GVL. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures No relevant conflicts of interest to declare.

scholarly journals Analysis of transgene-specific immune responses that limit the in vivo persistence of adoptively transferred HSV-TK–modified donor T cells after allogeneic hematopoietic cell transplantation

Blood ◽  
2006 ◽  
Vol 107 (6) ◽  
pp. 2294-2302 ◽  
Author(s):  
Carolina Berger ◽  
Mary E. Flowers ◽  
Edus H. Warren ◽  
Stanley R. Riddell
Keyword(s):  
T Cells ◽  
T Cell ◽  
Immune Responses ◽  
Cell Transplantation ◽  
Hematopoietic Cell Transplantation ◽  
T Cell Responses ◽  
Hematopoietic Cell ◽  
Allogeneic Hematopoietic Cell Transplantation ◽  
Cell Responses ◽  
Donor T Cells

AbstractThe introduction of an inducible suicide gene such as the herpes simplex virus thymidine kinase (HSV-TK) might allow exploitation of the antitumor activity of donor T cells after allogeneic hematopoietic cell transplantation (HCT) without graft versus host disease. However, HSV-TK is foreign, and immune responses to gene-modified T cells could lead to their premature elimination. We show that after the infusion of HSV-TK–modified donor T cells to HCT recipients, CD8+ and CD4+ T-cell responses to HSV-TK are rapidly induced and coincide with the disappearance of transferred cells. Cytokine flow cytometry using an overlapping panel of HSV-TK peptides allowed rapid detection and quantitation of HSV-TK–specific T cells in the blood and identified multiple immunogenic epitopes. Repeated infusion of modified T cells boosted the induced HSV-TK–specific T cells, which persisted as memory cells. These studies demonstrate the need for nonimmunogenic suicide genes and identify a strategy for detection of CD4+ and CD8+ T-cell responses to transgene products that should be generally applicable to monitoring patients on gene therapy trials. The potency of gene-modified T cells to elicit robust and durable immune responses imply this approach might be used for vaccination to elicit T-cell responses to viral or tumor antigens.

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