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1
Huisman, Wesley, Didier A. T. Leboux, Lieve E. van der Maarel, Lois Hageman, Derk Amsen, J. H. Frederik Falkenburg, and Inge Jedema. "The Scope of Allo-HLA Cross-Reactivity By (Third Party) Virus Specific T Cells Is Surprisingly Affected By HLA Restriction Rather Than Virus Specificity." Blood 132, Supplement 1 (November 29, 2018): 2048. http://dx.doi.org/10.1182/blood-2018-99-116028.
Abstract:
Abstract Reactivations of cytomegalovirus (CMV), Epstein Bar virus (EBV) and adenovirus (AdV) are frequently seen in immune compromised patients after allogeneic stem cell transplantation (alloSCT), and are associated with high morbidity and mortality. T cell immunity is essential for anti-viral protection, but a fully competent T cell repertoire generally does not develop until 3-6 months after transplantation. Especially patients transplanted with a virus non- experienced donor are at risk of developing severe complications. Adoptive transfer of partially HLA-matched virus specific T cells from healthy third party donors is a potential strategy to temporarily provide anti-viral immunity to these patients. However, these partially HLA-matched T cells harbor a risk of mediating allo-HLA cross-reactivity. Here, we investigated whether virus specificity and HLA restriction of the virus specific T cells influence the risk of allo-HLA cross-reactivity, and thus the development of GVHD. To determine the occurrence and diversity of allo-HLA cross-reactivity, virus specific CD8 T cells from homozygous HLA-A*01:01/B*08:01 and HLA-A*02:01/B*07:02 donors were isolated by cell sorting using tetramers for various peptides from CMV, EBV and AdV. Allo-HLA cross-reactivity was tested using an allogeneic EBV-LCL panel covering 116 different HLA molecules and confirmed using K562 cells retrovirally transduced with single HLA alleles of interest. A significant proportion of the virus specific T cell populations (n=174; 20 specificities) isolated from 27 healthy donors exerted allo-HLA cross-reactivity, as measured by recognition of 1 or more HLA mismatched EBV-LCLs from the panel. Similar frequencies were found for the various viral specificities showing 30% of the CMV, 46% of the EBV and 36% of the AdV-specific T cell populations to be allo-HLA cross-reactive. However, for some specificities (e.g. HLA-A*0201-restricted EBV-LMP2-FLY) allo-HLA cross-reactivity was infrequent (n=1/11), whereas for other specificities (e.g. HLA-B*08:01-restricted EBV-BZLF1-RAK) the majority of the T cell populations (n=9/13) was allo-HLA reactive. Surprisingly, a much larger fraction of HLA-B*08:01 restricted virus specific T cell populations showed allo-HLA cross-reactivity (72%, 36 out of 50 T cell lines), compared to the other HLA restricted virus specific T cell populations (29% of HLA-A*01:01, 30% of HLA-A*02:01 and 26% of HLA-B*07:02 restricted virus specific T cell lines). HLA-B*08:01 restricted virus specific T cells also exhibited the broadest allo-HLA reactivity, reacting to a median of 5 allo EBV-LCLs (range 1-17). In contrast, HLA-A*01:01, HLA-A*02:01 and HLA-B*07:02 restricted virus specific T cells reacted to a median of 1, 2 and 3 (ranges 1-7) allo EBV-LCLs, respectively. Dissection of the diversity/specificity of the allo-HLA reactivity using the panel of 40 different single HLA transduced K562 cells further illustrated the extensive allo-HLA cross-reactivity for HLA-B*08:01 restricted T cells isolated from homozygous HLA-A*01/B*08 donors compared to virus specific T cells restricted by other HLA alleles. These data show that allo-HLA cross-reactivity by virus specific T cells is highly influenced by the HLA restriction and not by the viral specificity of the T cell populations. Of the HLA-A*01, A*02, B*07 and B*08-restricted virus specific T cell populations isolated from homozygous donors, HLA-B*08:01 restricted virus specific T cells showed the highest frequency and diversity of allo-HLA cross-reactivity. Our results indicate that selection of virus specific T cells with specific HLA restrictions may decrease the risk of developing GVHD after infusion of third-party virus specific T cells to patients with uncontrolled viral reactivation after alloSCT. Disclosures No relevant conflicts of interest to declare.
2
Webb, S. R., J. H. Li, I. MacNeil, P. Marrack, J. Sprent, and D. B. Wilson. "T cell receptors for responses to Mls determinants and allo-H-2 determinants appear to be encoded on different chromosomes." Journal of Experimental Medicine 161, no. 1 (January 1, 1985): 269–74. http://dx.doi.org/10.1084/jem.161.1.269.
Abstract:
Previous studies have shown that T cell clones specific for strong Mlsa,d determinants concomitantly display apparently random reactivity to allo-H-2 determinants. One explanation for this finding is that T cell recognition of Mlsa,d and allo-H-2 determinants is controlled by separate sets of receptors. If these receptors were chromosomally unlinked, karyotypically unstable T cell hybrids with dual reactivity for Mlsa,d and particular allo-H-2 determinants would be expected, occasionally, to lose reactivity for one set of determinants, but not the other. The results presented here provide direct support for this prediction.
3
Amir, Avital L., Lloyd J. A. D'Orsogna, Dave L. Roelen, Marleen M. van Loenen, Renate S. Hagedoorn, Renate de Boer, Menno A. W. G. van der Hoorn, et al. "Allo-HLA reactivity of virus-specific memory T cells is common." Blood 115, no. 15 (April 15, 2010): 3146–57. http://dx.doi.org/10.1182/blood-2009-07-234906.
Abstract:
Abstract Graft-versus-host disease and graft rejection are major complications of allogeneic HLA-mismatched stem cell transplantation or organ transplantation that are caused by alloreactive T cells. Because a range of acute viral infections have been linked to initiating these complications, we hypothesized that the cross-reactive potential of virus-specific memory T cells to allogeneic (allo) HLA molecules may be able to mediate these complications. To analyze the allo-HLA reactivity, T cells specific for Epstein-Barr virus, cytomegalovirus, varicella zoster virus, and influenza virus were tested against a panel of HLA-typed target cells, and target cells transduced with single HLA molecules. Eighty percent of T-cell lines and 45% of virus-specific T-cell clones were shown to cross-react against allo-HLA molecules. The cross-reactivity of the CD8 and CD4 T-cell clones was directed primarily against HLA class I and II, respectively. However, a restricted number of CD8 T cells exhibited cross-reactivity to HLA class II. T-cell receptor (TCR) gene transfer confirmed that allo-HLA reactivity and virus specificity were mediated via the same TCR. These results demonstrate that a substantial proportion of virus-specific T cells exert allo-HLA reactivity, which may have important clinical implications in transplantation settings as well as adoptive transfer of third-party virus-specific T cells.
4
Huisman, Wesley, Didier A. T. Leboux, Lieve E. van der Maarel, Lois Hageman, Derk Amsen, Fred Falkenburg, and Inge Jedema. "Off-Target HLA Cross-Reactivity By (Third Party) Virus-Specific T Cells Is Surprisingly Affected By HLA Restriction and HLA Background but Not By Virus Specificity." Blood 134, Supplement_1 (November 13, 2019): 4440. http://dx.doi.org/10.1182/blood-2019-124785.
Abstract:
Reactivations of cytomegalovirus (CMV), Epstein Barr virus (EBV) and adenovirus (AdV) occur frequently in immune compromised patients after allogeneic stem cell transplantation (alloSCT) and cause high morbidity and mortality. T-cell immunity is essential for anti-viral protection, but a fully competent T-cell repertoire generally does not develop until 3-6 months after transplantation. Especially patients transplanted with a graft from a virus non-experienced donor are at risk. Adoptive transfer of partially HLA-matched virus-specific T cells from healthy third party donors is a potential strategy to temporarily provide anti-viral immunity to these patients. However, such T cells harbor a risk of mediating off-target toxicity due to allo-HLA cross-reactivity. It is not currently known whether the degree of allo-HLA cross-reactivity is random or whether rules exist that might allow prediction of specific T-cell populations. Here, we investigated whether virus specificity, HLA type of the donor or HLA restriction of the virus-specific T cells influence the risk of allo-HLA cross-reactivity. Through cell sorting using tetramers for various peptides from CMV, EBV and AdV, 164 CD8 T-cell populations (21 specificities) were isolated from peripheral blood of 24 healthy donors, homozygous for HLA-A*01:01/B*08:01 and HLA-A*02:01/B*07:02. Allo-HLA cross-reactivity was tested using an allogeneic EBV-LCL panel covering 116 different HLA molecules and confirmed using K562 cells retrovirally transduced with single HLA alleles of interest. Forty percent of all virus-specific T-cell populations exerted allo-HLA cross-reactivity. Similar frequencies were found for the various viral specificities showing 33% of the CMV, 43% of the EBV and 38% of the AdV-specific T-cell populations to be allo-HLA cross-reactive. Surprisingly, a much larger fraction of the HLA-B*08:01-restricted virus-specific T-cell populations exhibited allo-HLA cross-reactivity (77%) than from those restricted by the other HLAs (32% of HLA-A*01:01, 38% of HLA-A*02:01 and 26% of HLA-B*07:02-restricted virus-specific T-cell populations). HLA-B*08:01-restricted virus-specific T cells also exhibited the broadest allo-HLA reactivity, reacting to a median of 5 different allogeneic EBV-LCLs (range 1-17). In contrast, HLA-A*01:01, HLA-A*02:01 and HLA-B*07:02-restricted virus-specific T cells reacted to a median of 1, 2 and 3 (range 1-7) different allogeneic EBV-LCLs, respectively. Dissection of the diversity/specificity of the allo-HLA reactivities using a panel of 40 different single HLA-A, B, or C-transduced K562 cells further illustrated recurrent recognition of a restricted group of allogeneic HLA-B molecules by HLA-B*08:01-restricted T-cell populations, mediated by single T-cell clones. Heterozygosity for recurrently recognized allo-HLA-B molecules led to a significant decrease in the broadness of allo-HLA cross-reactivity by HLA-B*08:01-restricted T-cell populations, presumably due to negative thymic selection. In contrast, heterozygosity HLA-B molecules that were not part of the restricted group of cross-recognized alleles did not significantly decrease allo-HLA cross-reactivity. These data show that allo-HLA cross-reactivity by virus-specific T cells is highly influenced by their HLA restriction and the HLA background of the donors, but not by their virus specificity. Of the HLA-A*01, A*02, B*07 and B*08-restricted virus-specific T-cell populations isolated from homozygous donors, HLA-B*08:01-restricted virus-specific T cells showed the highest frequency and diversity of allo-HLA cross-reactivity with recurrent recognition of groups of specific mismatched allogeneic HLA-B alleles. Our results indicate that selection of virus-specific T cells with specific HLA restrictions and HLA backgrounds may decrease the risk of off-target toxicity after infusion of third-party virus-specific T cells to patients with uncontrolled viral reactivation after alloSCT. Disclosures No relevant conflicts of interest to declare.
5
Hasenkamp, Justin, Andrea Borgerding, Bjoern Chapuy, Gerald Wulf, Inga Missal, Wolfram Jung, Lorenz Truemper, and Bertram Glass. "Allo-Reactive NK Cells after HLA-Matched Allogeneic Hematopoietic Stem Cell Transplantation." Blood 108, no. 11 (November 16, 2006): 2904. http://dx.doi.org/10.1182/blood.v108.11.2904.2904.
Abstract:
Abstract Allo-reactive natural killer (NK) cells frequently occur early after haplo-mismatch hematopoietic stem cell transplantation (HSCT) with killer cell immunoglobuline-like receptor (KIR)-ligand mismatch in graft versus host (GvH) direction. Clinical data and experiments in mice indicate a beneficial influence on relapse rates, graft acceptance and Graft-versus-Host disease (GvHD). We determined the incidence of allo-reactive donor type NK cells after HLA A-, B-, DR-, DQ-matched allogeneic HSCT on a functional level. Clinical course, chimerism (PCR), immune-reconstitution (FACS) and frequencies of functional active and allo-reactive NK cells (ELISpot) were longitudinal determined in 19 patients so far. Patients (pts) suffered for high risk AML (7 pts), CML failing cytogenetic response to imatinib (3 pts), poor risk ALL (2 pts), relapse/refractory high-grade NHL (6 pts) and Multiple Myeloma (13q-) (1 pt). All patients received myeloablative conditioning regimens and GvHD-prophylaxis with cyclosporine A or tacrolimus and short course mycophenolat mofetil without in vivo or ex vivo T cell depletion. Chimerism analyses ensured hematopoietic reconstitution from donor type in 19/19 patients. In 3/19 patients NK cell activity was absent even against HLA class I negative control target cells. Absence of functional active NK cells correlates with severe acute GvHD accompanied by high doses of glucocorticosteroid medication. In all other patients we detected at least once functional active NK cells in peripheral blood. In 4/19 cases we detected allo-reactive NK cells after HSCT at days (d) +28, +68, +128 (case 19), d +56 (case 8), d +355 (case 1) and d +379 (case 13). Two cases were transplanted in KIR-ligand mismatch in GvH direction (donors HLA-CAsn80 and -CLys80, recipients missing HLA-CLys80). Allo-reactive NK cells were absent in all patients with known complete KIR-ligand match. Flow cytometry data on reconstitution of NK cell repertoire showed individual heterogeneous results. After median observation time post HSCT of 268 d (31–902) 3 patients died due to relapse. None of the patients with NK cell allo-reactivity experienced relapse. This is the first proof of circulating functionally active, allo-reactive NK cells after HLA-A, -B, -DR and -DQ matched HSCT. We detected NK cell allo-reactivity in all donor-recipient pairs with KIR-ligand (HLA-C) mismatch in GvH direction. After haplo-mismatch HSCT and T cell depletion NK cell allo-reactivity is restricted early after transplantation (within 3 months). In contrast, we detected late onset (>1 year) of NK cell allo-reactivity after one-locus (HLA-C) mismatch HSCT without T cell depletion of the grafts.
6
Webb, S. R., A. Okamoto, and J. Sprent. "Analysis of T hybridomas prepared from a T cell clone with three specificities. Recognition of self + X and allo-H-2 determinants segregates from recognition of Mlsa determinants." Journal of Immunology 141, no. 6 (September 15, 1988): 1828–34. http://dx.doi.org/10.4049/jimmunol.141.6.1828.
Abstract:
Abstract To see k information on T cell recognition of Mlsa determinants, hybridomas were prepared from a well-characterized F23.2+ (V beta 8.2+) T cell clone specific for three different ligands, i.e., 1) Mlsa determinants, 2) fowl gamma-globulin (F gamma G) plus self-H-2 (H-2d), and 3) allo-H-2, e.g., H-2p, determinants. Fusion of the clone to the BW5147 thymoma line produced a triple-reactive T hybridoma which generated two types of spontaneous variants. One type of variant (type I) lost Mlsa reactivity but retained reactivity to both F gamma G/H-2d and allo-H-2p. These variants, which were generated at high frequency, stained strongly with a mAb, A1.57, with idiotypic specificity for the TCR molecules of the parental clone. The second type of variant (type II) reacted to Mlsa determinants but showed no reactivity to F gamma G/H-2d or to allo-H-2p. These variants failed to express the A1.57 idiotypic determinants of the parent clone, but were F23.2+ (V beta 8.2+); nonequilibrium pH gradient electrophoresis analysis suggested that these hybrids expressed a mixed TCR heterodimer composed of the parental clone beta-chain and the BW5147 alpha-chain. Three aspects of the data are very difficult to accommodate with the view that Mlsa, F gamma G, and allo-H-2 determinants are all recognized via a common TCR molecule: 1) the independent (and frequent) segregation of Mlsa reactivity from F gamma G/H-2d and allo-H-2p reactivity, 2) the retention of Mlsa reactivity by the type II variants despite loss of the parental clone alpha-chain, and 3) the loss of Mlsa reactivity by the type I variants despite high expression of the A1.57+ TcR molecules derived from the parental clone. The data support a model in which Mlsa determinants are recognized by a separate T cell structure, which we envisage as a monomorphic accessory molecule unrelated to the TCR. Since the type II hybridoma variants invariably retained quantitatively normal TcR expression, the triggering phase of anti-Mlsa responses appears to be TCR dependent. The model we favor is that anti-Mlsa/Mlsa interaction increases TCR binding with Ia epitopes to above the threshold required for cell triggering. A key feature of this model is that Mlsa and Ia determinants are recognized as separate structures rather than as a complex.
7
Elkon, Keith B., and Dalit Ash Any. "Autoimmunity Versus Allo- and Xeno-Reactivity in SCID Mice." International Reviews of Immunology 11, no. 4 (January 1994): 283–93. http://dx.doi.org/10.3109/08830189409051175.
8
Laghmouchi, Aicha, Conny Hoogstraten, Peter Van Balen, Rick van de Water, Marian van de Meent, J. H. Frederik Falkenburg, and Inge Jedema. "The Allo-HLA-DP Restricted T Cell Repertoire Contains a Variety of Tissue-Restricted Specificities with Therapeutic Value." Blood 128, no. 22 (December 2, 2016): 3356. http://dx.doi.org/10.1182/blood.v128.22.3356.3356.
Abstract:
Abstract T cell-depleted allogeneic stem cell transplantation (alloSCT) is applied in patients with hematological malignancies to reduce the risk of graft versus host disease (GvHD), but the associated increased risk of infections and disease relapse makes scheduled donor-lymphocyte-infusion (DLI) necessary. Since HLA-DP is not taken into account in the matching procedure, stem cell grafts from unrelated donors are often only 10/10 matched, and mismatched for HLA-DP. Under non-inflammatory conditions, expression of HLA-DP is restricted to hematopoietic cells. Therefore, treatment with HLA-DP mismatched donor CD4 T cells can induce a specific graft versus leukemia (GvL) effect. However, in some cases, patients receiving HLA-DP mismatched CD4 T cells suffer from GvHD mediated by a profound allo-HLA-DP specific immune response. Adoptive transfer of in-vitro selected allo-HLA-DP restricted donor T cells directed against antigens specifically expressed on hematopoietic cells may be an elegant strategy to induce a specific GvL effect without coinciding GvHD. To investigate the feasibility of this approach, the allo-HLA-DP restricted T cell repertoire was dissected to unravel potential tissue specificities and to investigate the presence of hematopoiesis-specific CD4+ T cells with therapeutic value within this compartment. To induce allo-HLA-DP directed T cell responses, HLA-DP mismatched (10/10 matched) donor/patient pairs were selected. Donor PBMC were co-cultured with patient mature monocyte-derived dendritic cells (DC) for 14 days. The donor cells were first re-stimulated with autologous DC and depleted of activated auto-reactive CD137+ T cells using magnet cell separation (MACS). The negative fraction was then stimulated with the HLA-DP mismatched patient DC to induce activation of allo-reactive T cells. These allo-reactive CD137+ CD4+ T cells were clonally isolated using flowcytometric cell sorting, and expanded. Allo-HLA-DP restricted recognition of different hematopoietic and non-hematopoietic stimulator cells by the T cell clones was assessed using IFNγ and IL-4 ELISA. The T cells were tested against a large panel of hematopoietic cells (monocytes, DC, EBV-LCL and PHA blasts) of donor and patient origin, leukemic cell samples (AML) and non-hematopoietic cells (IFNγ pretreated, HLA-class-II expressing, skin-derived fibroblasts, and target-HLA-DPB1-transduced HELA, lung, kidney, and colon carcinoma cell lines). After re-stimulation with patient DC, flowcytometry showed frequencies of 0.5-10% of allo-DC activated (CD137+) CD4+ T cells. After cell sorting 1521 T cell clones from 4 different HLA-DP mismatched patient/donor pairs were tested against donor EBV-LCL, donor DC, patient EBV-LCL and patient DC as initial screening for allo-reactivity. 80% of the T cell clones showed allo-reactivity, as defined by recognition of patient, but not donor-derived EBV-LCL and/or DC. 14% of the tested clones showed no reactivity and 6% were auto-reactive. The HLA-DP restriction was analyzed of 400 selected allo-reactive T cell clones, using a panel consisting of donor, patient and 3rd party EBV-LCL or DC, 3rd party fibroblasts and target HLA-DPB1 transduced HELA cells. Of these 400 T cell clones, 65% were confirmed to be HLA-DP restricted. From these allo-HLA-DP restricted T cell clones 80% recognized both hematopoietic and non-hematopoietic cells expressing the target allo-HLA-DPB1. The other 20% of the allo-HLA-DP restricted T cell clones only produced cytokines when stimulated with hematopoietic cells (EBV-LCL and/or DC), and not when stimulated with non-hematopoietic cells (Fibroblasts, HELA, carcinoma cell lines). Moreover, 40% of these T cell clones showing hematopoiesis-restricted recognition only recognized DC, but not EBV-LCL expressing the target HLA-DPB1 allele. These clones also recognized primary AML blasts and proliferating CD34+ progenitor cells, illustrating a myeloid lineage restricted recognition pattern. These results illustrate that reactivity of allo-HLA-DP restricted T cells is not only determined by the expression of the target HLA-DPB1 allele, but is also dictated by cell lineage-specific gene expression causing differential peptide expression in HLA-DPB1. As a result, a significant portion of the allo-HLA-DP specific T cell repertoire harbors a specific GvL recognition pattern with high therapeutic value. Disclosures No relevant conflicts of interest to declare.
9
Noonan, Kimberly, Leo Luznik, and Ivan M. Borrello. "Enrichment of Allogeneic Tumor Antigen-Specific T Cells From Bone Marrow (BM) of Patients Treated with High-Dose Post-Transplant Cyclophoshamide (Cy) – A Novel Approach to Adoptive Immunotherapy." Blood 118, no. 21 (November 18, 2011): 647. http://dx.doi.org/10.1182/blood.v118.21.647.647.
Abstract:
Abstract Abstract 647 Background: Donor lymphocyte infusions (DLI) has limited efficacy and is associated with substantial graft vs host disease (GVHD) in treating relapse after allogeneic BMT (alloBMT) in large part due to the absence of significant tumor specificity. Thus, novel strategies are required that increase the tumor specificity with reduced alloreactivity. High-dose Cy post-BMT effectively reduces GVHD through early elimination of allo-reactive T cells and enables immune reconstitution free of ongoing pharmacologic immunosuppression. Since the BM is both the site of disease in most hematologic malignancies and a reservoir of tumor specific T cells, we hypothesized that marrow infiltrating lymphocytes (MILs) collected after alloBMT in patients treated with high-dose Cy could be a source of tumor-specific T cells for adoptive immunotherapy. Methods: BM was obtained from patients 2 –12 months after HLA-matched alloBMT on the clinical trial using high-dose post-transplant Cy as a single agent GVHD prophylaxis. The MILs were activated and expanded with CD3/CD28 antibody-coated beads. Results: Allo-MILs can be reproducibly expanded (574-fold avg expansion; 14–2000) at all time points tested. The activated allo-MILs are not anergic, exhibit anti-HLA-reactivity against third party allo-antigens but do not respond to recipient allo-antigens. Tumor-specific MILs were significantly expanded as determined by reactivity to HL60/K562 myeloid cell lysates (P<0.0001; pre- vs. post-activation). In contrast, no change was observed in cultures pulsed with irrelevant antigens, thus suggesting the specificity of the response. The ability to expand allogeneic antigen-specific MILs was also confirmed with a 15.5 fold expansion of PR-1 specific (HLA) A*0201-restricted CD8+ T-cells using tetramers. Conclusions: We can augment the tumor-specificity of MILs obtained post-transplant with minimal allo-reactivity in patients treated with post-BMT high-dose Cy. This could represent a novel approach to highly tumor-specific DLI. A phase I/II trial is planned to assess the safety and feasibility of administering ex vivo expanded post-transplant, allogeneic MILs to patients with relapsed hematologic malignancies after alloHSCT. Disclosures: No relevant conflicts of interest to declare.
10
D’Orsogna, Lloyd, Heleen van den Heuvel, Cees van Kooten, Sebastiaan Heidt, and Frans H. J. Claas. "Infectious pathogens may trigger specific allo-HLA reactivity via multiple mechanisms." Immunogenetics 69, no. 8-9 (July 17, 2017): 631–41. http://dx.doi.org/10.1007/s00251-017-0989-3.
11
Byun, Nari, Hyun-Je Kim, Il-Hee Yoon, and Oh-Sang Kwon. "Cross reactivity of allo and xenoimmune response in transplantation (TRAN2P.964)." Journal of Immunology 194, no. 1_Supplement (May 1, 2015): 209.4. http://dx.doi.org/10.4049/jimmunol.194.supp.209.4.
Abstract:
Abstract Many type I diabetes patients are suffering from hypoglycaemia unawareness. And porcine islet transplantation is one of the promising modalities to treat the problematic condition in a physiologic manner. But by transplanting porcine islet, xenoantigen sensitization is inevitable. For clinical application of porcine islet transplantation, the knowledge of the outcome and consequent effects of xenoantigen sensitization needs to be fully established. Our focus was whether xenoantigen sensitization would influence subsequent allotransplantation in any way. We confirmed that xenoantigen sensitization does not induce alloreactive antibody. And by ELISpot assay we demonstrated that xenoreactive T cells are not responsive to alloantigen. Lastly, we confirmed that xenoantigen sensitized mice have the same graft survival day compared with non sensitized mice in allogeneic islet transplantation setting. With these results put together, we conclude that porcine antigen sensitization can not induce alloantigen sensitization in mice.
12
Hueso, Thomas, Leonardo Magro, Marie Jonquel-chevalier Curt, Ibrahim Yakoub-Agha, and David Seguy. "Pretransplant Plasmatic Citrulline and Monocyte-Derived Macrophage Reactivity : Two Useful Surrogate Markers to Predict Acute GvHD." Blood 128, no. 22 (December 2, 2016): 4592. http://dx.doi.org/10.1182/blood.v128.22.4592.4592.
Abstract:
Abstract Acute graft versus host disease (aGvHD) represents the first complication after allogeneic stem cell transplantation (allo-HCT). During conditioning regimen (CR) intestinal damage induces microbial translocation, which primes macrophage reactivity and leads to donor-derived T cells alloreactivity. To date, little is reported on intestinal status and macrophage reactivity prior to CR. This study aimed to assess citrulline, an amino acid reflecting intestinal damage, and circulating monocytes-derived macrophage reactivity 30 days before CR. Blood samples from consecutive patients undergoing allo-HCT for hematological malignancies and from 10 healthy donors were analyzed. Citrulline levels were assessed by high performance liquid chromatography associated to spectrometry. Monocytes were isolated and stimulated with pathogen-associated molecular patterns as LPS, PAM, flagelline, MDP, curdlan and PBS as control. In each condition, cytokines (TNF-α, IL-1β, IL-6, IL-10) releasing was evaluated by multiplex fluorescent immunoassay. Citrulline and cytokine levels were analyzed relatively to aGvHD onset within 100 days post-transplant. Forty-seven patients underwent allo-HCT for different indications, including acute leukemia (n=26) and myelodysplastic syndrome (n=11). Twenty patients developed an aGvHD I-IV. Among the groups, differences were depicted for levels of citrulline, IL-1β, IL-6 and IL-10. Citrulline levels were lower in aGvHD group than in no-aGvHD group (p<0.05). Among all cytokines, only IL-6 and IL-10 were greater in aGvHD group than no-aGvHD group, especially after curdlan stimulation (p<0.05). Citrulline levels below the normal value (<20 μmol/L), IL-6 and IL-10 above thresholds (326 pg/ml and 78pg/ml after curdlan stimulation respectively) were associated with the aGvHD development (log rank test, p<0.05). Any patients with citrulline>20 µmol/L, IL-6<326 pg/ml and IL-10<78 pg/ml (n=10) declared aGvHD within the 100-days post-transplant. Our results highlight the role of intestinal damage and macrophage reactivity in aGVHD development. Pretransplant plasmatic citrulline level and monocyte-derived-macrophage reactivity are promising accessible surrogate markers suitable for identifying patients likely to develop aGvHD. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.
13
Burkhardt, Ute E., Ursula Hainz, Kristen E. Stevenson, Di Wu, Vincent T. Ho, Natalie R. Goldstein, Mildred Pasek, et al. "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 120, no. 21 (November 16, 2012): 1892. http://dx.doi.org/10.1182/blood.v120.21.1892.1892.
Abstract:
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.
14
ten Berge, Ineke, Kirstin Heutinck, Unsal Yapici, Geurt Stokman, Sandrine Florquin, Cees van Kooten, G. Garcia-Romo, et al. "The interplay between antiviral immunity and allo-immune reactivity after renal transplantation." Transplant Immunology 31, no. 4 (October 2014): 191–94. http://dx.doi.org/10.1016/j.trim.2014.09.004.
15
Karahan, Gonca E., Frans H. J. Claas, and Sebastiaan Heidt. "Heterologous Immunity of Virus-Specific T Cells Leading to Alloreactivity: Possible Implications for Solid Organ Transplantation." Viruses 13, no. 12 (November 24, 2021): 2359. http://dx.doi.org/10.3390/v13122359.
Abstract:
Exposure of the adaptive immune system to a pathogen can result in the activation and expansion of T cells capable of recognizing not only the specific antigen but also different unrelated antigens, a process which is commonly referred to as heterologous immunity. While such cross-reactivity is favourable in amplifying protective immune responses to pathogens, induction of T cell-mediated heterologous immune responses to allo-antigens in the setting of solid organ transplantation can potentially lead to allograft rejection. In this review, we provide an overview of murine and human studies investigating the incidence and functional properties of virus-specific memory T cells cross-reacting with allo-antigens and discuss their potential relevance in the context of solid organ transplantation.
16
Bondanza, Attilio, Veronica Valtolina, Zulma Magnani, Maurilio Ponzoni, Katharina Fleischhauer, Mark Bonyhadi, Catia Traversari, et al. "Suicide gene therapy of graft-versus-host disease induced by central memory human T lymphocytes." Blood 107, no. 5 (March 1, 2006): 1828–36. http://dx.doi.org/10.1182/blood-2005-09-3716.
Abstract:
In allogeneic hematopoietic cell transplantation (allo-HCT), the immune recognition of host antigens by donor T lymphocytes leads to a beneficial graft-versus-leukemia (GvL) effect as well as to life-threatening graft-versus-host disease (GvHD). Genetic modification of T lymphocytes with a retroviral vector (RV) expressing the herpes simplex virus-thymidine kinase (TK) suicide gene confers selective sensitivity to the prodrug ganciclovir (GCV). In patients, the infusion of TK+ lymphocytes and the subsequent administration of GCV resulted in a time-wise modulation of antihost reactivity for a GvL effect, while controlling GvHD. Because activation required for genetic modification with RV may reduce antihost reactivity, we investigated the requirements for maximizing the potency of human TK+ lymphocytes. Whereas T-cell receptor triggering alone led to effector memory (EM) TK+ lymphocytes, the addition of CD28 costimulation through cell-sized beads resulted in the generation of central memory (CM) TK+ lymphocytes. In a quantitative model for GvHD using nonobese diabetic/severely combined immunodeficient mice, CM TK+ lymphocytes were more potent than EM TK+ lymphocytes. GCV administration efficiently controlled GvHD induced by CM TK+ lymphocytes. These results warrant the clinical investigation of CM suicide gene-modified human T lymphocytes for safe and effective allo-HCT.
17
Amir, Avital L., Dirk M. van der Steen, Renate S. Hagedoorn, Michel G. D. Kester, Cornelis A. M. van Bergen, Jan W. Drijfhout, Arnoud H. de Ru, J. H. Frederik Falkenburg, Peter A. van Veelen, and Mirjam H. M. Heemskerk. "Allo-HLA–reactive T cells inducing graft-versus-host disease are single peptide specific." Blood 118, no. 26 (December 22, 2011): 6733–42. http://dx.doi.org/10.1182/blood-2011-05-354787.
Abstract:
Abstract T-cell alloreactivity directed against non–self-HLA molecules has been assumed to be less peptide specific than conventional T-cell reactivity. A large variation in degree of peptide specificity has previously been reported, including single peptide specificity, polyspecificity, and peptide degeneracy. Peptide polyspecificity was illustrated using synthetic peptide-loaded target cells, but in the absence of confirmation against endogenously processed peptides this may represent low-avidity T-cell reactivity. Peptide degeneracy was concluded based on recognition of Ag-processing defective cells. In addition, because most investigated alloreactive T cells were in vitro activated and expanded, the previously determined specificities may have not been representative for alloreactivity in vivo. To study the biologically relevant peptide specificity and avidity of alloreactivity, we investigated the degree of peptide specificity of 50 different allo-HLA–reactive T-cell clones which were activated and expanded in vivo during GVHD. All but one of the alloreactive T-cell clones, including those reactive against Ag-processing defective T2 cells, recognized a single peptide allo-HLA complex, unique for each clone. Down-regulation of the expression of the recognized Ags using silencing shRNAs confirmed single peptide specificity. Based on these results, we conclude that biologically relevant alloreactivity selected during in vivo immune response is peptide specific.
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Kuball, Jürgen, Wouter Scheper, Suzanne van Dorp, Sabina Kersting, Floor Pietersma, Samantha Hol, Victoria Marcu-Malina, Sabine Becke, Bodo Plachter, and Debbie van Baarle. "Multipotent Vδ2-negative γδT-cells after CMV-reactivation in allogeneic stem cell transplantation (162.36)." Journal of Immunology 188, no. 1_Supplement (May 1, 2012): 162.36. http://dx.doi.org/10.4049/jimmunol.188.supp.162.36.
Abstract:
Abstract Human cytomegalovirus (CMV) infections and relapse of disease remain major problems after allogeneic stem cell transplantation (allo-SCT). Great promise as an alternative strategy, with a potential use for very broad patient population, arises from the non-MHC-restricted target-recognition of cancer cells and virally-infected cells by γδT-cells. Therefore, we investigated the role of γδT-cells after allo-SCT and CMV infection. An increase in selectively Vδ1-positive γδT-cells was observed in CMV-reactivating patients from CMV-positive conventional donors. Even more important, γδT-cell expansions were also observed after CMV-negative cordblood transplantation. However, in contrast to expansions from CMV-positive donors, γδT-cells from cordblood donors did not only include Vδ1-positive but also Vδ3-positive cells. Expanded Vδ2-negative γδT-cells were not only able to lyse CMV-infected fibroblasts, but also to partially mature dendritic cells (DCs) and to kill primary leukemic blasts. CMV- and leukemia-reactivity were restricted to the same clonal population and separated from DC-maturation capacities of Vδ2-negative T-cells. Vδ2-negative γδTCRs selectively mediated anti-leukemia-reactivity. This makes CMV-reactive γδT-cells e.g. from cordblood donors as well as individual Vδ2-negative γδTCRs interesting candidates for adoptive immunotherapy strategies
19
Bondanza, Attilio, Veronica Valtolina, Zulma Magnani, Shin Kaneko, Maurilio Ponzoni, Francesca Sanvito, Mark Bonyhadi, et al. "Suicide Gene Therapy of Graft-Versus-Host Disease Induced by Central Memory Human T Lymphocytes." Blood 106, no. 11 (November 16, 2005): 3096. http://dx.doi.org/10.1182/blood.v106.11.3096.3096.
Abstract:
Abstract Suicide gene therapy (SGT) is a powerful approach to exploit anti-host reactivity following allogeneic hemopoietic cell transplantation (allo-HCT) for a full graft-versus-leukemia (GvL) effect, while controlling graft-versus-host disease (GvHD). This is accomplished through genetic modification of donor lymphocytes with a suicide gene. The herpes simplex thymidine kinase (TK) suicide gene converts at a cellular level the pro-drug ganciclovir (GCV) into tri-phosphate toxic derivatives, thence conferring selective sensitivity. Clinical studies as well as animal models have substantiated the concept that a time-wise GCV administration is able to actually separate the therapeutic GvL effect from life-threatening GvHD. Genetic modification of lymphocytes with TK is currently pursued through retroviral vectors (RV). In vitro RV genetic modification requires proliferation, which is easily accomplished by polyclonal stimulation. Polyclonal stimulation with anti-CD3 antibodies (aCD3) has been show to reduce anti-host reactivity of gene-modified lymphocytes. This possibly limits the impact of the suicide gene strategy in allo-HCT. In this study we tackled the rules governing anti-host reactivity of TK+ human lymphocytes. We found that TK+ lymphocytes generated with aCD3 are mainly CD45RA−CCR7− effector memory (EM) cells (84,6±6,6%). Upon re-stimulation they produce interferon-γ, perforin B and granzyme A, but fail to up-regulate CD40L. EM TK+ lymphocytes have a mixed phenotype in regard to CD28/CD27 co-expression and displayed a limited ability to engraft and cause GvHD in a xenogeneic model using conditioned NOD/scid mice (take: 11%). In sharp contrast, TK+ lymphocytes generated with novel protocols taking advantage of anti-CD3 and anti-CD28 antibodies conjugated to para-magnetic cell-sized beads are enriched for CD45RA−CCR7+ central memory (CM) cells (65,3±6,2%) that are able to produce IL-2 and to strongly up-regulate CD40L. CM TK+ lymphocytes are homogenously CD28+CD27+ (91,1,3±2,5%). When infused in conditioned NOD/scid mice CM TK+ lymphocytes persistently engrafted and caused lethal GvHD in a significant fraction of mice (take: 55%, P=0,0017 vs EM TK+ cells). GCV administration to diseased animals resulted in the elimination of TK+ cells in blood and in target organs. Treated animals were rescued with survival up to 120 days (P=0,009 vs saline-treated mice). These results demonstrate that CM TK+ lymphocytes retain significant anti-host reactivity and provide a clue to their in vitro generation. CM TK+ lymphocytes are promising candidates for safe and effective allo-HCT for the cure of hematological malignancies.
20
Karmaus, W., and V. Gangur. "Does allo-immune reactivity play a role in the prenatal programming of childhood allergy?" Clinical Experimental Allergy 35, no. 4 (April 2005): 405–7. http://dx.doi.org/10.1111/j.1365-2222.2005.02218.x.
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Markiewicz, Miroslaw, Urszula Siekiera, Tomasz Kruzel, Monika Dzierzak-Mietla, Patrycja Zielinska, and Slawomira Kyrcz-Krzemien. "DynaChip Anti-HLA Antibody Analysis In Sera of Patients Following Allo-HSCT From HLA-Mismatched Unrelated Donors." Blood 116, no. 21 (November 19, 2010): 4575. http://dx.doi.org/10.1182/blood.v116.21.4575.4575.
Abstract:
Abstract Abstract 4575 Introduction: Anti-HLA antibodies constitute potentially important factor that may influence outcomes of HLA-mismatched allogeneic hematopoietic stem cell transplantation (allo-HSCT). The rationale of this study was to detect presence of anti-HLA antibodies in recipients of allo-HSCT from HLA-mismatched unrelated donors. Patients and Methods: Anti-HLA-A,B,C,DR,DQ,DP antibodies were identified in sera collected from 46 recipients of allo-HSCT from HLA-mismatched unrelated donors. Sera were collected between 1 month and 5.5 years after allo-HSCT, and additionally before allo-HSCT in 17 pts. We have used microchips spotted with purified HLA class I and HLA class II antigens to allow binding of anti-HLA antibodies present in tested sera to the surface of the microchip, pre-optimised reagents and DynaChip Processor (Dynal Invitrogen Corporation) for assay processing, data acquisition and analysis. Results: Antibodies against HLA class I, II or I and II were detected in 15%, 11% and 35% of pts whereas no antibodies were detected in 39% of patients. Antibodies were directed against HLA-A, B, C, DR and DQ in 37%, 46%, 35%, 48% and 35% of pts, respectively. Pre-transplant anti-HLA antibodies have been detected in 7 pts (41%) out of 17 tested before allo-HSCT. In this group percent of Panel Reactive Antibodies (% PRA) increased following allo-HSCT in 3 pts and decreased in 4. In 5 out of 10 remaining pts without pre-transplant antibodies, %PRA increased post-transplant. DynaChip software allowed to define specificities of HLA-A,B,C,DR and DQ antibodies on low and high resolution levels. The specificity of antigens that masked results of antibody identification has been also defined in 2 pts. At this stage we did not define exactly whether detected anti-HLA antibodies were donor-specific. Cross-reactive groups (CREG's) analysis has been also used to compare antibodies’ reactivity. Anti-HLA-DP antibodies were not detected in the examined group of transplanted patients. Conclusions: Presented preliminary study results indicate, that anti-HLA antibodies can appear post-transplant in mismatched allo-HSCT recipients. Further analysis aiming to evaluate their influence on transplant outcomes is ongoing. We intend to extend the search for anti-HLA antibodies with use of Luminex LabScreen method. Disclosures: No relevant conflicts of interest to declare.
22
Udaka, K., K. H. Wiesmüller, S. Kienle, G. Jung, and P. Walden. "Self-MHC-restricted peptides recognized by an alloreactive T lymphocyte clone." Journal of Immunology 157, no. 2 (July 15, 1996): 670–78. http://dx.doi.org/10.4049/jimmunol.157.2.670.
Abstract:
Abstract Alloreactive T lymphocytes are readily detected in unprimed animals although they have never encountered the alloantigen before. This well-established phenomenon is usually explained with the assumption that a self-MHC molecule complexed with a defined peptide resembles the allo-MHC molecule with another peptide and induces the corresponding T cell specificities. Here, for the first time and in support of this hypothesis, self-MHC-restricted peptides are described for a T cell clone that was induced with allo-MHC. The allo-MHC-specific CTL clone 2C was derived from a H-2b mouse and recognizes H-2Ld complexed with the naturally occurring endogenous peptide LSPFPFDL. H-2Kb was shown to be involved in positive selection of its TCR, and peptides associated with this MHC molecule are implicated in the process. To identify such peptides, positional scanning with random peptide libraries combined with an iterative approach was employed. Several active peptides were found and the most efficient, SIYRYYGL, was chosen for further studies. Recognition by 2C of the two MHC-peptide adducts H-2Ld + LSPFPFDL and H-2Kb + SIYRYYGL is mediated by the same TCR and appears to be similarly efficient as concluded from inhibition experiments with an Id-specific Ab. CTLs from SIYRYYGL-primed H-2b mice respond to H-2Ld + LSPFPFDL. This reciprocal cross-reactivity suggests that structural features are shared by the two MHC-peptide complexes.
23
Klobuch, Sebastian, Kathrin Hammon, Sarah Vatter-Leising, Elisabeth Neidlinger, Michael Zwerger, Annika Wandel, Laura Maria Neuber, et al. "HLA-DPB1 Reactive T Cell Receptors for Adoptive Immunotherapy in Allogeneic Stem Cell Transplantation." Cells 9, no. 5 (May 20, 2020): 1264. http://dx.doi.org/10.3390/cells9051264.
Abstract:
HLA-DPB1 antigens are mismatched in about 80% of allogeneic hematopoietic stem cell transplantations from HLA 10/10 matched unrelated donors and were shown to be associated with a decreased risk of leukemia relapse. We recently developed a reliable in vitro method to generate HLA-DPB1 mismatch-reactive CD4 T-cell clones from allogeneic donors. Here, we isolated HLA-DPB1 specific T cell receptors (TCR DP) and used them either as wild-type or genetically optimized receptors to analyze in detail the reactivity of transduced CD4 and CD8 T cells toward primary AML blasts. While both CD4 and CD8 T cells showed strong AML reactivity in vitro, only CD4 T cells were able to effectively eliminate leukemia blasts in AML engrafted NOD/SCID/IL2Rγc−/− (NSG) mice. Further analysis showed that optimized TCR DP and under some conditions wild-type TCR DP also mediated reactivity to non-hematopoietic cells like fibroblasts or tumor cell lines after HLA-DP upregulation. In conclusion, T cells engineered with selected allo-HLA-DPB1 specific TCRs might be powerful off-the-shelf reagents in allogeneic T-cell therapy of leukemia. However, because of frequent (common) cross-reactivity to non-hematopoietic cells with optimized TCR DP T cells, safety mechanisms are mandatory.
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Adam, Jacqueline, Natascha Wuillemin, Stephan Watkins, Heidi Jamin, Klara K. Eriksson, Peter Villiger, Stefano Fontana, Werner J. Pichler, and Daniel Yerly. "Abacavir Induced T Cell Reactivity from Drug Naïve Individuals Shares Features of Allo-Immune Responses." PLoS ONE 9, no. 4 (April 21, 2014): e95339. http://dx.doi.org/10.1371/journal.pone.0095339.
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Pearson, Todd, Marie King, Leonard Shultz, Jean Leif, Dale Greiner, John Mordes, Aldo Rossini, et al. "Development of Latest-generation HU-PBMC-NOD/SCID Mice to Study Human Islet Allo-reactivity." Clinical Immunology 123 (2007): S69. http://dx.doi.org/10.1016/j.clim.2007.03.374.
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Tawara, Isao, Kana Okamori, Naoyuki Katayama, Hiroshi Shiku, and Hiroaki Ikeda. "Tumor-Specific Donor Lymphocyte Infusion for Tumor Relapse after MHC-Haploidentical Hematopoietic Stem Cell Transplantation." Blood 128, no. 22 (December 2, 2016): 2157. http://dx.doi.org/10.1182/blood.v128.22.2157.2157.
Abstract:
Abstract Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative therapy for refractory hematologic malignancies such as leukemia and myelodysplastic syndrome. Tumor relapse, graft-vs host disease (GVHD) and infections are, however, major obstacles to successful allo-HSCT. Donor lymphocyte infusion (DLI) is carried out in some cases of tumor relapse and infections after allo-HSCT. DLI potentially induces or aggravates GVHD due to allo-reactivity of the lymphocytes used for infusion and the efficacy of DLI for tumor relapse is actually limited. Therefore, development of the method of DLI with enhanced anti-tumor effects without induction or aggravation of GVHD is expected to improve the results of DLI in the context of tumor relapse. Tumor antigen-specific T cell receptor (TCR)-expressing cell infusion is one of the potentially effective immunotherapies for refractory tumors. We previously reported that tumor antigen-specific TCR-gene-transduced human lymphocytes engineered with a novel retrovirus vector silencing endogenous TCRs showed reduced allo-reactivity (ASH annual meeting 2014). Utilizing this technology, we may be able to enhance anti-tumor effects without induction or aggravation of GVHD. In this study, we conducted a mouse pre-clinical model that mimics tumor-specific TCR-engineered DLI for tumor relapse after MHC-haploidentical HSCT and explored the effect of DLI on tumor growth and GVHD. We performed experimental study utilizing the following model: MHC-haploidentical BALB/c(H-2d)→CB6F1(H-2b/d) HSCT, BALB/c-derived sarcoma CMS5a and T cells from TCR-transgenic mouse DUC18. CMS5a has a unique mutated form of a mitogen-activated kinase ERK2, and a nonamer peptide, called 9m, incorporating the resulting amino acid substitution is presented on H-2Kd and recognized by the CD8+ CTL clone C18. DUC18 is a BALB/c-background TCR transgenic line that expresses the rearranged Vα10.1 and Vβ8.3 genes of the C18. CD8+ T cells of DUC18 mice can suppress CMS5a tumor growth in the syngeneic BALB/c hosts. Allo-reactivity of DUC18 T cells was reduced as compared to that of wild type BALB/c T cells. Based on the results of preliminary experiments, we set standard experimental conditions as follows: X-ray irradiation (10 Gy) for pre-conditioning, 5x106 whole bone marrow and 0.25x106 T cell transplant (BMT) for mild GVHD induction, 1x106 CMS5a inoculation for progressive tumor growth in BMT recipients, and 4x106 CD8+ T cells for observation of the effect of DLI on tumor growth and GVHD. In some experiment, CD4+ T cells were co-infused with CD8+ T cells. DLI was carried out 3 days after CMS5a inoculation at 2, 4 or 8 weeks after BMT. We first used CD8+ T cells from naïve BALB/c or DUC18 mice for DLI at 8 weeks after BMT. No GVHD aggravation was observed in either BALB/c or DUC18 CD8+ T cell recipients and, as expected, suppression of tumor growth was observed only in the DUC18 CD8+ T cell recipients. Next, DLIs using CD8+ T cells only, or both CD8+ and CD4+ (CD8+/CD4+) T cells from naïve mice were carried out at 4 or 2 weeks after BMT. No apparent effect of CD4+ T cell co-infusion on tumor growth and GVHD was observed in both types of donor T cell recipients when DLI was carried out at 4 weeks after BMT. However, GVHD was aggravated in CD8+/CD4+ T cell recipients from both types of donors when DLI was carried out at 2 weeks after BMT, and no CD4-driven additional anti-tumor effect was observed in DUC18 T cell recipients. We then performed DLI using in vitro activated/expanded T cells including both CD8+ and CD4+ as a more clinically relevant model. BALB/c and DUC18 splenocytes were activated and expanded in vitro and infused (adjusted CD8+ T cell number to 4x106/mouse) into recipients at 2 or 8 weeks after BMT. Aggravation of GVHD was observed in either BALB/c or DUC18 activated/expanded T cell recipients when DLI was carried out at 2 weeks after BMT. On the other hand, it was not observed when DLI was carried out at 8 weeks after BMT. Anti-tumor effect of DUC18 T cells was observed regardless of timings of DLI. Taken together, infusion of tumor-specific donor lymphocytes with reduced allo-reactivity for tumor relapse after MHC-haploidentical HSCT will be promising strategy. Timing of DLI and condition of recipients such as inflammatory status may affect GVHD. Multiple model studies are required for development of more effective DLI strategies without aggravation of GVHD. Disclosures Tawara: Astellas: Honoraria. Katayama:Nippon Shinyaku: Honoraria; Chugai: Honoraria, Research Funding; Alexion Pharmaceuticals: Honoraria; Dainippon Sumitomo Pharma: Honoraria; Pfizer: Honoraria; Celgene: Honoraria; Shionogi: Honoraria; Kyowa Hakko Kirin: Honoraria, Research Funding; Taisho Toyama Pharma: Honoraria; Shire: Honoraria; Astellas: Honoraria, Research Funding; Daiichi Sankyo: Honoraria; Eisai: Honoraria; Takeda: Honoraria; Bristol-Myers Squibb Japan: Honoraria. Ikeda:Ono Pharmaceutical Co., Ltd: Honoraria; Daiichi Sankyo Co., Ltd: Honoraria.
27
Jürgens, Birgit, Julia Raberger, Dietmar Fuchs, and Andreas Heitger. "Indoleamine 2,3-Dioxygenase in Human Hematopoietic Stem Cell Transplantation." International Journal of Tryptophan Research 3 (January 2010): IJTR.S4076. http://dx.doi.org/10.4137/ijtr.s4076.
Abstract:
In recent years tryptophan metabolism and its rate limiting enzyme indoleamine 2,3-dioxygenase (IDO) have attracted increasing attention for their potential to modulate immune responses including the regulation of transplantation tolerance. The focus of this review is to discuss some features of IDO activity which particularly relate to hematopoietic stem cell transplantation (HSCT). HSCT invariably involves the establishment of some degree of a donor-derived immune system in the recipient. Thus, the outstanding feature of tolerance in HSCT is that in this type of transplantation it is not rejection, which causes the most severe problems to HSCT recipients, but the reverse, graft-versus-host (GvH) directed immune responses. We will discuss the peculiar role of IDO activity and accelerated tryptophan metabolism at the interface between immune activation and immune suppression and delineate from theoretical and experimental evidence the potential significance of IDO in mediating tolerance in HSCT. Finally, we will examine therapeutic options for exploitation of IDO activity in the generation of allo-antigen-specific tolerance, i.e. avoiding allo-reactivity while maintaining immunocompetence, in HSCT.
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van Bergen, Cornelis A. M., Simone A. P. Van Luxemburg-Heijs, Matthijs Eefting, Maria W. Honders, Inge Jedema, Constantijn J. M. Halkes, Marieke Griffioen, and J. H. Frederik Falkenburg. "Graft Versus Leukemia Separates From Graft Versus Host Disease By Magnitude and Avidity Of The Allo-Reactive T Cell Response After Allogeneic Stem Cell Transplantation and Donor Lymphocyte Infusion." Blood 122, no. 21 (November 15, 2013): 2014. http://dx.doi.org/10.1182/blood.v122.21.2014.2014.
Abstract:
Abstract Donor lymphocyte infusion (DLI) after allogeneic stem cell transplantation (alloSCT) can be a curative treatment for patients with hematological malignancies due to the capacity of allo-reactive donor derived T cells to mediate a curative potent graft versus leukemia (GVL) effect. However, associated acute graft versus host disease (GVHD) remains a major risk. To study the role of CD8+ T cells in GVL reactivity and GVHD, we selected patients who responded to DLI (without preceding cytoreductive treatment) for recurrent disease or incomplete donor chimerism after alloSCT. The patients were grouped according to absence (7 patients) or presence (6 patients) of GVHD. To quantify the number of circulating activated CD8+ T cells before DLI and at the time of disease regression or conversion to full donor chimerism we measured the frequencies of CD8+ HLA-DR+ T cells in peripheral blood samples by flowcytometry. Before DLI, highly variable numbers of CD8+ HLA-DR+ T cells were found (37.8 ± 42.9 x106/L), that significantly increased after DLI (309±473 x106/L, p<0.005), demonstrating involvement of CD8+ HLA-DR+ T cells in immune responses after DLI. To determine the specificity and functional avidity of the CD8+ HLA-DR+ T cells, these cells were isolated using flowcytometric cell sorting and clonally expanded. From a total of 30 samples, on average 225 T cell clones per sample were obtained and tested for recognition of patient and donor derived EBV-LCL, CD40L stimulated B cells (CD40L-B cells) and monocyte derived dendritic cells (monoDC). Surprisingly, in many samples from both patient cohorts high percentages of clones recognizing EBV-LCL derived from both patient and donor but not recognizing CD40L-B cells and monoDC were found. These T cells may be involved in anti-EBV responses irrespective of the presence of a GVL effect or GVHD. To investigate whether the magnitude of the allo-immune response was different in patients with or without GVHD coinciding the GVL effect, we compared the frequencies of allo-reactive T cell clones in samples from both patient groups. Significantly lower percentages of allo-reactive T cell clones were found in patients without GVHD as compared to patients with GVHD (5.1 ± 7.0% versus 32.5 ± 20.0% respectively, p<0.01), showing that coinciding GVHD is associated with an increased magnitude of the allo-reactive T cell response. Per patient, we determined the number of unique antigens targeted by the isolated T cell clones by characterizing the targeted MiHA using whole genome association scanning. In line with the lower total number of allo-reactive T cells, a lower number of unique MiHA was targeted in patients without GVHD (2.7±3.5) as compared to patients with GVHD (10.2±5.8, p=0.015). To determine whether occurrence of GVHD could be explained by the tissue specificity and functional avidity of the allo-reactive T cell response after DLI, we tested the T cell clones obtained from both patient cohorts for recognition of fibroblasts (FB) derived from skin biopsies of the patient. To mimic pro-inflammatory conditions, FB were pretreated for 4 days with 100 IU/ml IFN-γ. Recognition of untreated FB was exclusively mediated by T cell clones obtained from patients with GVHD, whereas recognition of IFN-γ pretreated FB was found for clones isolated from patients with or without coinciding GVHD. In addition, several T cell clones isolated from patients without GVHD were found to be directed against MiHA encoded by genes with a broad expression profile in non-hematopoietic cells comprising FB, despite absence of FB recognition under non-inflammatory conditions. This suggests that in addition to the tissue expression profile of the MiHA other factors, comprising the local inflammatory milieu, play a role in the risk of developing GVHD. In conclusion, our data show a strong correlation between the magnitude and the functional avidity of the allo-reactive CD8+ T cell response and the occurrence of GVHD after DLI. We hypothesize that the limited production of pro-inflammatory cytokines due to the moderate magnitude of the immune response in patients mounting a GVL response without coinciding GVHD reactivity may have prevented the induction of GVHD by the lower avidity allo-reactive T cells, that under pro inflammatory conditions can mediate GVHD by recognition of normal non-hematopoietic cells of the patient. Disclosures: No relevant conflicts of interest to declare.
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Mielke, Stephan, Raquel Nunes, Katayoun Rezvani, Vicki S. Fellowes, Yong Fan, Scott R. Solomon, Christian Scotto, Elizabeth J. Read, and A. John Barrett. "High Efficiency Clinical Scale Selective Depletion of Alloreacting T Cells Using Expanded T Lymphocytes as Antigen-Presenting Cells and a TH9402-Based Photodepletion Technique in HLA-Mismatched and Matched Donor-Recipient Pairs." Blood 108, no. 11 (November 16, 2006): 721. http://dx.doi.org/10.1182/blood.v108.11.721.721.
Abstract:
Abstract Selective depletion (SD) is a strategy to eliminate host-reactive donor lymphocytes from blood stem cell allografts to prevent GVHD and maintain GVL-effects. As an alternative to CD25-immunotoxin-based SD, we investigated a photodepletion (PD) process, whereby allo-activated donor cells are labeled with a photosensitizing rhodamine-based dye, 4,5-dibromorhodamide 123 (TH9402), and exposed to visible light, which preferentially eliminates allo-activated dye-retaining cells. Stimulator cells were prepared from recipient-derived leukapheresis mononuclear cells (MNCs) and cultured using anti-CD3 and 100 IU IL-2/ml. Responder cells (leukapheresis MNCs) from 3 random HLA-mismatched volunteers and 3 HLA-matched sibling donors were cocultured 1:1 with irradiated stimulators for 3 days. Cultured cells were incubated with 7.5 μM TH9402, followed by dye-extrusion and exposure to 5 Joule light energy in the PD light source (Celmed Bioscience Inc., Canada) at 5x106 cells/ml in FEP plastic bags. Flow cytometry was performed before and after PD and included surface phenotyping for CD25 and intracellular staining for forkhead protein3 (foxp3). Depletion efficacy was studied by mixed lymphocyte reactions (MLR) in mismatched pairs and by helper-T-lymphocyte precursor (HTLp) frequency assay in matched pairs. All six clinical-scale experiments provided sufficient reduction of allo-reactivity and retention of third party responses as measured against a pool of 5 donors. In mismatched pairs mean reduction of allo-reactivity was 703-fold (± 141) when compared to unmanipulated donors. Third-party responses were maintained, with a mean reduction of only 1.3 ± 0.15-fold (Figure A). In matched pairs alloreactivity was reduced below the “GVHD-threshold” of 1/100.000 whilst third party responses remained above 1/10.000 precursors (Figure B). Effective depletion was observed despite that fact that a small fraction of CD25+ T cells remained after PD. These were mainly CD4+CD25+ T cells (1.5 ± 1.1 % of CD4+) and very residual CD8+CD25+ T cells (0.3 ± 0.2 % of CD8+), and represented predominantly CD4+foxp3+ regulatory T cells (Tregs) which persisted after PD (1.4 ± 1.7 % of CD4+). This establishes a clinical scale PD process capable of highly efficient removal of alloreactive lymphocytes from mismatched and matched MLRs while maintaining desirable third party responses. As PD targets activation-based changes in MDR-1 that result in an altered dye efflux, the mechanism of action is distinct from surface-marker-based allodepletion (e.g. CD69, CD25). Thus, PD may overcome instability of activation-based surface marker expression resulting in more consistent and effective depletion. PD may further mitigate the risk of GVHD by conserving Tregs. This approach will now be tested in a clinical SD trial. Figure Figure.
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David, V., J. F. Bourge, P. Guglielmi, D. Mathieu-Mahul, L. Degos, and A. Bensussan. "Human T cell clones use a CD3-associated surface antigen recognition structure to exhibit both NK-like and allogeneic cytotoxic reactivity." Journal of Immunology 138, no. 9 (May 1, 1987): 2831–36. http://dx.doi.org/10.4049/jimmunol.138.9.2831.
Abstract:
Abstract In the present study, we developed human non-MHC-restricted CTL clones from human peripheral blood mononuclear cells activated in vitro with recombinant IL 2 and subsequently expended with PHA. The CD3/Ti+ clones were selected for their ability to exhibit non-MHC-restricted CTL reactivity by killing various tumor cell lines in culture, including the line K562 which does not express MHC antigens. We report that, at least for some of the NK-like T cell clones, it is possible to establish an allo-CTL activity, and that the CD3-associated surface antigen recognition structure might be involved in both reactivities.
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Rowntree, Louise C., Thi H. O. Nguyen, Stephanie Gras, Tom C. Kotsimbos, and Nicole A. Mifsud. "Deciphering the clinical relevance of allo-human leukocyte antigen cross-reactivity in mediating alloimmunity following transplantation." Current Opinion in Organ Transplantation 21, no. 1 (February 2016): 29–39. http://dx.doi.org/10.1097/mot.0000000000000264.
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Gamadia, Laila E., Ester B. Remmerswaal, Sugianto Surachno, Neubury M. Lardy, Pauline M. Wertheim-van Dillen, Ren?? A. W. van Lier, and Ineke J. M. ten Berge. "CROSS-REACTIVITY OF CYTOMEGALOVIRUS-SPECIFIC CD8+ T CELLS TO ALLO-MAJOR HISTOCOMPATIBILITY COMPLEX CLASS I MOLECULES." Transplantation 77, no. 12 (June 2004): 1879–85. http://dx.doi.org/10.1097/01.tp.0000131158.81346.64.
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Qureshi, Khalid M., Jou Lee, Michelle B. Paget, Clifford J. Bailey, S. John Curnow, Hilary E. Murray, and Richard Downing. "Low gravity rotational culture and the integration of immunomodulatory stem cells reduce human islet allo-reactivity." Clinical Transplantation 29, no. 1 (December 5, 2014): 90–98. http://dx.doi.org/10.1111/ctr.12488.
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Methe, Heiko, Adam Groothuis, Mohamed H. Sayegh, and Elazer R. Edelman. "Matrix adherence of endothelial cells attenuates immune reactivity: induction of hyporesponsiveness in allo‐ and xenogeneic models." FASEB Journal 21, no. 7 (January 30, 2007): 1515–26. http://dx.doi.org/10.1096/fj.06-7051com.
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D’Orsogna, Lloyd J., Frans H. J. Claas, Mina John, Coral-Ann Almeida, Paula van Miert, Sonia Fernandez, and Yvonne Zoet. "OR24 Allo-HLA reactivity by HIV-specific T-cells: A potential adjunct to HIV vaccine design?" Human Immunology 76 (October 2015): 9. http://dx.doi.org/10.1016/j.humimm.2015.07.016.
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Sommer, Cesar, Bijan Boldajipour, Julien Valton, Roman Galetto, Trevor Bentley, Janette Sutton, Yajin Ni, et al. "ALLO-715, an Allogeneic BCMA CAR T Therapy Possessing an Off-Switch for the Treatment of Multiple Myeloma." Blood 132, Supplement 1 (November 29, 2018): 591. http://dx.doi.org/10.1182/blood-2018-99-119227.
Abstract:
Abstract Autologous chimeric antigen receptor (CAR) T cells targeting B-Cell Maturation Antigen (BCMA) have demonstrated promising clinical activity, inducing durable responses in patients with relapsed/refractory multiple myeloma (MM). Development of autologous CAR T therapies is however limited by logistical challenges and the time required for manufacturing, which has to be done for each patient. In addition, manufacturing may not be feasible in some patients. An allogeneic approach that utilizes engineered cells from a healthy donor could potentially expand patient access to these therapies by providing a readily available off-the-shelf product. We have previously described the screening of a library of single chain variable fragments (scFvs) with high affinity to human BCMA and the identification of candidate BCMA CARs with potent antitumor activity. Here we sought to further characterize ALLO-715, our lead allogeneic BCMA CAR T cell product, for its specificity to human BCMA, antitumor efficacy in vitro using a long-term killing assay and in xenograft mouse models with physiologic levels of human IL-7 and IL-15, and suitability for scale-up manufacturing. Allogeneic ALLO-715 CAR T cells were generated by lentiviral transduction with a second generation CAR construct incorporating a novel scFv derived from a fully-human antibody with high affinity to BCMA (KD value ~ 5 nM, determined at 37°C) and featuring a rituximab-driven off-switch. Transduced T cells were then transfected with mRNAs encoding Transcription Activator-Like Effector Nucleases (TALEN®) designed to specifically disrupt the T cell receptor alpha chain and CD52 loci. These modifications result in a cell product with a lower risk of TCR-mediated graft-versus-host disease and resistance to the CD52 antibody alemtuzumab, a lymphodepleting agent. BCMA CAR T cells exhibited robust cell expansion, with low levels of tonic signaling that resulted in minimal differentiation (> 50% Tscm/Tcm phenotype). In in vitro assays, ALLO-715 CAR T cells displayed potent cytotoxic activity when co-cultured with the target cell lines MM.1S, Molp-8, and BCMA-REH but negligible cytotoxicity against BCMA-negative REH cells. The high proliferative potential indicated by the high frequency of memory T cells was validated in long-term killing assays, where ALLO-715 CAR T cells showed substantial expansion in the presence of MM.1S cells with no evidence of exhaustion or diminished cytolytic activity after seven days of continuous exposure to target. The potency of ALLO-715 CAR T cells was unaffected by high concentrations of soluble BCMA (>10 ug/mL), which has been shown previously to interfere with the activity of some BCMA-specific CARs. In MM xenograft mouse models, ALLO-715 CAR T cells were highly efficacious at single dose. High serum IL-15 levels have been associated with CAR T cell expansion in clinical trials. To evaluate the impact of homeostatic cytokines on CAR T cell survival and antitumor activity in our xenograft models, mice were administered adeno-associated viruses (AAV) for the expression of human IL-7 and IL-15. In the presence of physiological concentrations of these cytokines, enhanced BCMA CAR T cell expansion and anti-tumor activity were observed. To assess potential off-target interactions of ALLO-715 CAR, tissue cross-reactivity studies were carried out on standard human tissue panels using a scFv-human IgG fusion protein. Consistent with the limited expression pattern of BCMA, reactivity was seen on scattered cells in lymphoid tissues such as tonsil and abundantly on BCMA-expressing cell lines, but no appreciable staining was detected in other tissues. We examined BCMA CAR T cells manufactured following a proprietary GMP-like clinical scale process and found that cell expansion and viability, T cell phenotype and in vivo antitumor efficacy were preserved. These results demonstrate the potential of ALLO-715 as a novel allogeneic BCMA CAR T therapy for the treatment of relapsed/refractory MM and other BCMA-positive malignancies. Disclosures Sommer: Allogene Therapeutics: Employment, Equity Ownership, Patents & Royalties. Boldajipour:Pfizer Inc.: Employment, Patents & Royalties. Valton:Cellectis.Inc: Employment, Equity Ownership, Patents & Royalties. Galetto:Cellectis SA: Employment, Equity Ownership, Patents & Royalties. Bentley:Allogene Therapeutics: Employment, Equity Ownership. Sutton:Allogene Therapeutics: Employment, Equity Ownership. Ni:Allogene Therapeutics: Employment, Equity Ownership. Leonard:Allogene Therapeutics: Employment, Equity Ownership. Van Blarcom:Allogene Therapeutics: Employment, Equity Ownership. Smith:Cellectis. Inc: Employment, Patents & Royalties. Chaparro-Riggers:Pfizer Inc.: Employment, Patents & Royalties. Sasu:Allogene Therapeutics: Employment, Equity Ownership, Patents & Royalties.
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Cousens, Leslie P., and Pamela A. Davol. "Targeting Lymphoma with Bispecific Antibody-Armed T Cells and their Potential for use in an Allogeneic Setting (41.31)." Journal of Immunology 182, no. 1_Supplement (April 1, 2009): 41.31. http://dx.doi.org/10.4049/jimmunol.182.supp.41.31.
Abstract:
Abstract Patients with high-risk refractory or relapsed lymphomas face a poor prognosis and limited therapeutic options. Current treatment regimens of chemotherapy with either rituximab (anti-CD20 mAb) or with allogeneic stem cell transplant (alloSCT) yield serious toxicities and high relapse rates. In particular, benefits of alloSCT are grossly limited by life-threatening graft vs host disease (GvHD). New approaches to enhance graft vs lymphoma (GvL) effects without increased GvHD are needed. Bispecific antibodies (BiAbs) are produced by linking anti-CD3 for T cell binding to an antibody specific for a tumor antigen. Anti-CD3 activated T cells (ATC) "armed" with BiAb express tumor-specific CTL function independent of further activation, proliferation, antigen presentation or costimulation. Studies here examined allo-reactivity of BiAb-armed ATC for potential application in an allogeneic setting. Data show that ATC armed with anti-CD3 x anti-CD20 BiAb specifically target and kill B cell lymphomas. However, BiAb-armed ATC are remarkably poor stimulators of, or responders to, allo-stimlulation. These results provide important evidence of a functional separation between the GvL and GvH effects of BiAb-armed ATC.
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Morse, H. C., R. A. Yetter, C. S. Via, R. R. Hardy, A. Cerny, K. Hayakawa, A. W. Hugin, M. W. Miller, K. L. Holmes, and G. M. Shearer. "Functional and phenotypic alterations in T cell subsets during the course of MAIDS, a murine retrovirus-induced immunodeficiency syndrome." Journal of Immunology 143, no. 3 (August 1, 1989): 844–50. http://dx.doi.org/10.4049/jimmunol.143.3.844.
Abstract:
Abstract The functional and phenotypic characteristics of Ly-4(CD4)+ and Ly-2(CD8)+ T cells were studied after induction of murine AIDS with LP-BM5 murine leukemia virus. Assays of spleen cells for their ability to generate in vitro CTL responses to TNP-modified autologous cells (self + x CTL) and to alloantigens (allo CTL) showed that self + x CTL responses were greatly impaired at 3 to 4 wk postinfection and were undetectable thereafter. Allo CTL responses were normal at 3 to 4 wk, but were reduced at 8 to 9 wk and absent at 14 wk postinfection. This sequential loss of self + x and allo CTL responses was related to a selective defect in Ly-4(CD4)+ Th cell function associated with impaired production of IL-2 and deficient proliferative responses to Con A or to soluble Ag. Changes in the functional characteristics of Ly-4(CD4)+ T cells were unrelated to changes in their frequency in spleen, but did correlate with marked alterations in their distribution among four subsets defined by mAb SM3C11 and SM6C10. Assays of CTL responses generated by mixtures of spleen cells from normal and infected mice suggested that active suppression of Ly-4(CD4)+ Th function may contribute to this defect. Studies of Ly-2(CD8)+ T cells showed that infection with LP-BM5 murine leukemia virus also induced a major phenotypic shift in subpopulations defined by their reactivity with mAb 6C10. However, this phenotypic change did not appear to correlate with major functional defects.
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Raghunathan, R., T. M. Mundy, J. Faust, and S. Misra. "Allogeneic leukocyte and germ cell-induced murine immunodeficiency." Journal of Immunology 135, no. 4 (October 1, 1985): 2429–31. http://dx.doi.org/10.4049/jimmunol.135.4.2429.
Abstract:
Abstract Immune deficiency, as defined by significant decreases in lymphocyte Con A and allo-reactivity and in natural killer (NK) function, was induced in normal adult mice by i.p. injections of combinations of allogeneic testicular germ cells and splenic leukocytes over 3 wk. This immune deficiency was evident at 8 wk after initial injection, and profound by 12 wk. Neither leukocytes nor testicular cells, given alone, were able to induce similar immune deficiency. These findings suggest the possibility that allogeneic germ cells and leukocytes of semen, on repeated administration, may induce immune deficiency and may act as co-factors to viral agents in the development of clinical AIDS in humans.
40
Yao, Xin, Fenlu Zhu, Huiqing Xu, and Carolyn A. Keever-Taylor. "Allo-Reactive Potential of Multi-Virus Specific Cytotoxic T Cell Lines Recognizing Adenovirus Hexon, Cytomegalovirus pp65 and Epstein Barr Virus." Blood 114, no. 22 (November 20, 2009): 5121. http://dx.doi.org/10.1182/blood.v114.22.5121.5121.
Abstract:
Abstract Abstract 5121 Introduction Cytomegalovirus (CMV), adenovirus (Ad), and Epstein-Barr virus (EBV) are major pathogens after allogeneic hematopoietic stem cell transplant. The current antiviral therapy is associated with significant myelosuppression, impaired hematopoietic reconstitution and consequently, higher rates of other infections. Adoptive immunotherapy utilizing allogeneic donor-derived virus specific cytotoxic T cell lines (CTL) has been shown to effectively treat refractory viral infections. Although such lines carry the theoretical risk of allo-reactivity and the resultant graft versus host disease, this complication has not been widely reported in recipients of single virus specific cell lines. To improve the efficiency of this therapeutic modality we have developed an approach to generate a single cell line that recognizes all three of these clinically relevant viruses while also having greatly reduced allo-reactive potential. Methods Peripheral blood mononuclear cells (PBMC) from 3 healthy CMV seropositive donors were primed 3 times with autologous monocyte-derived dendritic cells pulsed with overlapping pools of pentadecapeptides spanning the antigen coding region of highly conserved regions of the Ad hexon protein and/or pools spanning the entire CMV pp65 protein. The lines were expanded for minimally one additional week using peptide loaded EBV transformed B cells (BLCL) as antigen presenting cells (APC). CMV, Ad and EBV anti-viral activity was monitored weekly by chromium release assay and the specificity of the lines was controlled by varying the peptide pools used for subsequent priming. The IFN-gamma ELISPOT assay was used to identify single peptides and to determine HLA restriction. The frequency and subset of single and pool peptide-reactive cells was determined using IFN-gamma intracellular cytokine production by flow cytometry. Fresh PBMC and established lines were additionally primed with fully allogeneic stimulators and response measured using intracellular cytokine production. Results Each of the 3 lines had robust proliferation and CTL activity against CMV and Ad peptide-pulsed targets. The lines consisted of 44.5-82.4% CD3+CD8+cells and 18.1-56.7% CD3+CD4+cells. CMV induced IFN-gamma response in 4.3-76.6% of CD3+CD8+cells and 0.4-47.5% of CD3+CD4+cells. Ad induced 1.4-43.9% of CD3+CD8+cells and 6.9-45.7% of CD3+CD4+cells to produce IFN-gamma. The 3 lines recognized 2-4 single CMV peptides and 2-6 single Ad peptides. The reactivity against these viral peptides was HLA restricted. At the time of testing none of the lines showed a strong EBV response. However, EBV reactivity could be detected with 1-2 additional primings, indicating that these precursors were present. Compared to fresh PBMC from the same donors, there was a 34.0±5.6 fold decrease in the frequency of alloreactive CD3+CD8+cells demonstrated in the intracellular cytokine assay. Conclusions Generation of multi-viral specific CTL in a single culture is feasible so long as antigen stimulation is controlled and this approach will greatly reduce the time, cost and number of cells required to restore viral immunity for patients receiving allogeneic stem cell transplant. The risk of allo-reactivity in these lines is greatly reduced compared to fresh PBMC. This makes adoptive cell therapy more practical, easier to implement and safer. Disclosures No relevant conflicts of interest to declare.
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Castro, Januario E., Juan S. Barajas-Gamboa, Julio A. Diaz-Perez, Lina M. Ariza-Serrano, Sue Corringham, Thomas J. Kipps, and Edward D. Ball. "Improved Outcome in Unrelated Donor Recipients After Allogeneic Hematopoietic Stem Cell Transplantation for Patients with Advanced Relapse / Refractory Chronic Lymphocytic Leukemia." Blood 118, no. 21 (November 18, 2011): 4474. http://dx.doi.org/10.1182/blood.v118.21.4474.4474.
Abstract:
Abstract Abstract 4474 Chronic lymphocytic leukemia (CLL) remains incurable with standard therapies and a large number of patients become refractory to treatment or develop toxicities that prevent further treatment and contribute to their mortality. Allogeneic hematopoietic stem cell transplantation (Allo-HSCT) is the only known curative treatment for CLL. However, management and prevention of toxicities as well as patient / donor selection remain as the most significant challenges that limit the success of this treatment. In this observational study we evaluated the efficacy and tolerability of Allo-HSCT in relapse / refractory patients with CLL and correlated the clinical outcome with prognostic markers as well as the donor type. All patients were enrolled at the University of California, San Diego - Moores Cancer Center between November 1998 and July 2009. Eighteen patients with progressive relapse / refractory CLL were included (9 males [8 Caucasian, 1 Hispanic], 9 females [8 Caucasian, 1 Hispanic]), nine patients received stem cells from related donors and the other nine from unrelated donors. The median age was 50 years (age range, 26–71), median interval between diagnosis and Allo-HSCT was 83 months, and the median number of regimens prior to transplant was 5.3 (range, 2–11). Five out of eighteen patients were fludarabine-refractory and one was not evaluable (NE). All patients had a high-risk disease at the time of transplantation, with advanced stages according to Rai classification, and high prevalence of IgVH unmutated state, as well a high expression of CD38 and ZAP-70. Six patients had cytogenetic abnormalities, ten were normal and two were not evaluated. The most common pre-transplantation cytogenetic alteration was deletion 17p (33.3%), followed by deletion 13q (22.2%) and deletion 11q (22.2%). Prior to Allo-HSCT, three were in CR, ten patients were in PR and five had PD. CMV serologic reactivity was present in 67% of patients. Seventeen patients received non-myeloablative conditioning regimens and one patient was treated with a myeloablative regimen. Disease status was assessed 30 and 100 days, 1, 2 and 3 years post Allo-HSCT. 13 out of 18 patients showed response to Allo-HSCT (6 PR and 7 CR) with the remaining patients having PD. Two patients had acute GVHD and 3 patients had extensive chronic GVHD. We observed no differences in the rate of GVHD in unrelated vs. related donor recipients (p=0.1). Five patients died, three due to progression of the disease, one due to post-transplant organ failure and one for acute GVHD. Survival analysis showed a median OS of 60.1 ± 3.9 months. We observed a significant difference in terms of OS in patients who had unrelated vs. related transplants (HR=3.1 [95% CI 1.5–18.7]). We observed no difference in survival of patients discriminated by the IgVH mutational status (HR =1.8 [95% CI 0.2–17.5]) or ZAP-70 expression (HR =1.0 [95% CI 0.9–1.1]). The median progression free survival was 56 months. Five of the 18 patients were retreated with donor lymphocyte infusions or a second Allo-HSCT after relapse or lost of chimerism. Three of these patients died of PD while the other two are still alive. In conclusion, Allo-HSCT induces long-term remission and survival in high-risk relapse / refractory CLL patients including patients with multiple previous treatments and progressive disease status at the time of transplantation. The response to Allo-HSCT is independent of prognostic markers such as IgVH and ZAP-70 expression. Patients that received stem cells from unrelated donors have a better clinical outcome including OS with no evidence of increased GVHD. Disclosures: No relevant conflicts of interest to declare.
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Melenhorst, J. Joseph, Ann M. Leen, Catherine M. Bollard, Máire F. Quigley, David A. Price, Cliona M. Rooney, Malcolm K. Brenner, A. John Barrett, and Helen E. Heslop. "Allogeneic virus-specific T cells with HLA alloreactivity do not produce GVHD in human subjects." Blood 116, no. 22 (November 25, 2010): 4700–4702. http://dx.doi.org/10.1182/blood-2010-06-289991.
Abstract:
Adoptive transfer of viral antigen-specific memory T cells can reconstitute antiviral immunity, but in a recent report a majority of virus-specific cytotoxic T-lymphocyte (CTL) lines showed in vitro cross-reactivity against allo-human leukocyte antigen (HLA) molecules as measured by interferon-γ secretion. We therefore reviewed our clinical experience with adoptive transfer of allogeneic hematopoietic stem cell transplantation donor-derived virus-specific CTLs in 153 recipients, including 73 instances where there was an HLA mismatch. There was no de novo acute graft-versus-host disease after infusion, and incidence of graft-versus-host disease reactivation was low and not significantly different in recipients of matched or mismatched CTL. However, we found that virus-specific T cell lines recognized up to 10% of a panel of 44 HLA disparate targets, indicating that virus-specific T cells can have cross-reactivity with HLA-mismatched targets in vitro. These data indicate that the adoptive transfer of partially HLA-mismatched virus-specific CTL is safe despite in vitro recognition of recipient HLA molecules.
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Sommer, Cesar, Hsin-Yuan Cheng, Yik Andy Yeung, Duy Nguyen, Janette Sutton, Zea Melton, Julien Valton, et al. "Preclinical Evaluation of ALLO-819, an Allogeneic CAR T Cell Therapy Targeting FLT3 for the Treatment of Acute Myeloid Leukemia." Blood 134, Supplement_1 (November 13, 2019): 3921. http://dx.doi.org/10.1182/blood-2019-129025.
Abstract:
Autologous chimeric antigen receptor (CAR) T cells have achieved unprecedented clinical responses in patients with B-cell leukemias, lymphomas and multiple myeloma, raising interest in using CAR T cell therapies in AML. These therapies are produced using a patient's own T cells, an approach that has inherent challenges, including requiring significant time for production, complex supply chain logistics, separate GMP manufacturing for each patient, and variability in performance of patient-derived cells. Given the rapid pace of disease progression combined with limitations associated with the autologous approach and treatment-induced lymphopenia, many patients with AML may not receive treatment. Allogeneic CAR T (AlloCAR T) cell therapies, which utilize cells from healthy donors, may provide greater convenience with readily available off-the-shelf CAR T cells on-demand, reliable product consistency, and accessibility at greater scale for more patients. To create an allogeneic product, the TRAC and CD52 genes are inactivated in CAR T cells using Transcription Activator-Like Effector Nuclease (TALEN®) technology. These genetic modifications are intended to minimize the risk of graft-versus-host disease and to confer resistance to ALLO-647, an anti-CD52 antibody that can be used as part of the conditioning regimen to deplete host alloreactive immune cells potentially leading to increased persistence and efficacy of the infused allogeneic cells. We have previously described the functional screening of a library of anti-FLT3 single-chain variable fragments (scFvs) and the identification of a lead FLT3 CAR with optimal activity against AML cells and featuring an off-switch activated by rituximab. Here we characterize ALLO-819, an allogeneic FLT3 CAR T cell product, for its antitumor efficacy and expansion in orthotopic models of human AML, cytotoxicity in the presence of soluble FLT3 (sFLT3), performance compared with previously described anti-FLT3 CARs and potential for off-target binding of the scFv to normal human tissues. To produce ALLO-819, T cells derived from healthy donors were activated and transduced with a lentiviral construct for expression of the lead anti-FLT3 CAR followed by efficient knockout of TRAC and CD52. ALLO-819 manufactured from multiple donors was insensitive to ALLO-647 (100 µg/mL) in in vitro assays, suggesting that it would avoid elimination by the lymphodepletion regimen. In orthotopic models of AML (MV4-11 and EOL-1), ALLO-819 exhibited dose-dependent expansion and cytotoxic activity, with peak CAR T cell levels corresponding to maximal antitumor efficacy. Intriguingly, ALLO-819 showed earlier and more robust peak expansion in mice engrafted with MV4-11 target cells, which express lower levels of the antigen relative to EOL-1 cells (n=2 donors). To further assess the potency of ALLO-819, multiple anti-FLT3 scFvs that had been described in previous reports were cloned into lentiviral constructs that were used to generate CAR T cells following the standard protocol. In these comparative studies, the ALLO-819 CAR displayed high transduction efficiency and superior performance across different donors. Furthermore, the effector function of ALLO-819 was equivalent to that observed in FLT3 CAR T cells with normal expression of TCR and CD52, indicating no effects of TALEN® treatment on CAR T cell activity. Plasma levels of sFLT3 are frequently increased in patients with AML and correlate with tumor burden, raising the possibility that sFLT3 may act as a decoy for FLT3 CAR T cells. To rule out an inhibitory effect of sFLT3 on ALLO-819, effector and target cells were cultured overnight in the presence of increasing concentrations of recombinant sFLT3. We found that ALLO-819 retained its killing properties even in the presence of supraphysiological concentrations of sFLT3 (1 µg/mL). To investigate the potential for off-target binding of the ALLO-819 CAR to human tissues, tissue cross-reactivity studies were conducted using a recombinant protein consisting of the extracellular domain of the CAR fused to human IgG Fc. Consistent with the limited expression pattern of FLT3 and indicative of the high specificity of the lead scFv, no appreciable membrane staining was detected in any of the 36 normal tissues tested (n=3 donors). Taken together, our results support clinical development of ALLO-819 as a novel and effective CAR T cell therapy for the treatment of AML. Disclosures Sommer: Allogene Therapeutics, Inc.: Employment, Equity Ownership. Cheng:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Yeung:Pfizer Inc.: Employment, Equity Ownership. Nguyen:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Sutton:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Melton:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Valton:Cellectis, Inc.: Employment, Equity Ownership. Poulsen:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Djuretic:Pfizer, Inc.: Employment, Equity Ownership. Van Blarcom:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Chaparro-Riggers:Pfizer, Inc.: Employment, Equity Ownership. Sasu:Allogene Therapeutics, Inc.: Employment, Equity Ownership.
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Blomberg, B. B., S. Hussini, H. Fainman, J. M. Mathew, A. Hernandez, M. Carreno, H. J. Hnatyszyn, et al. "Retroviral transduction of an allogeneic class II gene into human bone marrow down regulates allo-immune reactivity." Human Immunology 64, no. 10 (October 2003): S128. http://dx.doi.org/10.1016/j.humimm.2003.08.240.
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Amir, Avital, Marieke Griffioen, Michel D. G. Kester, M. Willy Honders, Roelof Willemze, J. H. Frederik Falkenburg, and Mirjam Heemskerk. "Allo-HLA Reactive CD8 T-Cells May Recognize Tissue Specific Peptides Explaining Tissue Restricted GVHD after HLA Mismatched SCT." Blood 110, no. 11 (November 16, 2007): 72. http://dx.doi.org/10.1182/blood.v110.11.72.72.
Abstract:
Abstract HLA disparity between patient and donor increases the risk of GVHD after allogeneic stem cell transplantation (SCT). However, similar to fully HLA matched SCT, the clinical manifestation of GVHD after HLA mismatched SCT is frequently restricted to skin, gut and liver. Based on the frequency of allo-HLA reactive T-cells, which is about a 1000-fold higher than the frequency of minor histocompatibility antigen specific T-cells, the immune response after HLA mismatched SCT is expected to be mediated by allo-HLA reactive T-cells. Theoretically allo-HLA reactive T-cells can exert three different types of recognition. They may directly recognize the mismatched HLA molecule, independent of the peptide it presents. Alternatively they could be peptide dependent but promiscuous in their peptide recognition, or peptide specific like in normal T-cell recognition. Previous studies all showed peptide independent recognition by at least part of the studied T-cells. Since HLA class I is expressed by all nucleated cells, these data would imply that after HLA mismatched SCT alloreactive T-cells would be able to equally damage all tissues. Whereas previous reports mainly studied in vitro generated allo-HLA reactive T-cells, we characterized the pattern of peptide recognition of a large number of different T-cells retrieved from a patient suffering from GVHD after HLA-A2 mismatched donor lymphocyte infusion. Activated CD8+ T-cells were single cell sorted based on HLA-DR expression, expanded and tested for alloreactivity and HLA restriction. 46 of the 56 generated CD8 T-cells clones were alloreactive and allo-HLA-A2 restricted. The alloreactive CD8 clones showed usage of different T-cell receptor Vβ chains with different CDR3 sequences, illustrating polyclonality. To evaluate the tissue recognition, the clones were tested against different HLA-A2 positive target cells derived from various hematopoietic and non-hematopoietic tissues. The clones showed major differential recognition of the different tissue target cells, varying from lack of recognition to strong recognition of the same target. To analyze the peptide recognition pattern, the clones were tested against TAP deficient T2 cells loaded with HPLC fractions of peptides eluted from HLA-A2. The different clones recognized different HPLC fractions, but each clone recognized of only one single fraction, indicating peptide specificity. Three clones recognized non-pulsed T2 cells, which was interpreted in previous studies to demonstrate peptide independence. T2 cells, however, do present peptides which are independent on TAP to enter the ER. By testing reactivity against TAP independent peptides, one clone showed recognition of a peptide derived from the signal sequence of TRAPα, which is positioned at the ER membrane, and is therefore TAP independent. This indicates that recognition of T2 cells by the allo-HLA reactive T-cells was also based on peptide specific recognition. Finally, two clones were screened against COS cells expressing HLA-A2 transfected with a c-DNA library constructed from an EBV-LCL. Each clone showed recognition of only one (different) gene out of 40.000 c-DNAs. These results show that all allo-HLA reactive T-cells exerted peptide specific recognition. Differential expression of tissue specific peptides that can be recognized in the context of allo-HLA molecules may explain tissue specific peptides as a cause for tissue restriction in GVHD after HLA mismatched SCT.
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Stevanovic, Sanja, Bart Nijmeijer, Marianke LJ Van Schie, Roelof Willemze, Marieke Griffioen, and J. H. Frederik Falkenburg. "Human T Cells with Distinct Specificities Mediate Graft-Versus-Leukemia Reactivity and Xenogeneic Graft-Versus-Host Disease in a NOD/Scid Mouse Model for Human Acute Leukemia." Blood 114, no. 22 (November 20, 2009): 1330. http://dx.doi.org/10.1182/blood.v114.22.1330.1330.
Abstract:
Abstract Abstract 1330 Poster Board I-352 Immunodeficient mice inoculated with human leukemia can be used as a model to investigate Graft-versus-Leukemia (GvL) effects of donor lymphocyte infusions (DLIs). In addition to GvL reactivity, treatment with DLI induces xenogeneic Graft-versus-Host Disease (GvHD) in mice, characterized by pancytopenia and weight loss. In patients treated with DLI for relapsed or residual leukemia after allogeneic stem cell transplantation, immune responses against non-leukemic cells may also cause GvHD. It has been suggested that GvL reactivity and GvHD, which co-develop in vivo, can be separated and that distinct T cells exist with the specific capacity to mediate GvL reactivity or GvHD. Since adoptive T cell transfer models that allow analysis of separation of GvL and GvHD are rare, we aimed to establish whether GvL reactivity and xenogeneic GvHD could be separated using our model of human leukemia-engrafted NOD/scid mouse after treatment with human donor T cells. In this study, non-conditioned NOD/scid mice engrafted with primary human acute lymphoblastic leukemic cells were treated with CD3+ DLI. Established tumors were effectively eliminated by emerging human T cells, but also induced xenogeneic GvHD. Flowcytometric analysis demonstrated that the majority of emerging CD8+ and CD4+ T cells were activated (HLA-DR+) and expressed an effector memory phenotype (CD45RA-CD45RO+CCR7-). To investigate whether GvL reactivity and xenogeneic GvHD were mediated by the same T cells showing reactivity against both human leukemic and murine cells, or displaying distinct reactivity against human leukemic and murine cells, we clonally isolated and characterized the T cells during the GvL response and xenogeneic GvHD. T cell clones were analyzed for reactivity against primary human leukemic cells and primary NOD/scid hematopoietic (BM and spleen cells) and non-hematopoietic (skin fibroblasts) cells in IFN-g ELISA. Isolated CD8+ and CD4+ T cell clones were shown to recognize either human leukemic or murine cells, indicating that GvL response and xenogeneic GvHD were mediated by different human T cells. Flowcytometric analysis demonstrated that all BM and spleen cells expressed MHC class I, whereas only 1-3 % of the cells were MHC class II +. Primary skin fibroblasts displayed low MHC class I and completely lacked MHC class II expression. Xeno-reactive CD8+ T cell clones were shown to recognize all MHC class I + target cells and xeno-reactive CD4+ T cells clones displayed reactivity only against MHC class II + target cells. To determine the MHC restriction of xeno-reactive T cell clones, NOD/scid bone marrow (BM) derived dendritic cells (DC) expressing high levels of murine MHC class I and class II were tested for T cell recognition in the presence or absence of murine MHC class I and class II monoclonal antibodies in IFN-g ELISA. Xeno-reactive CD8+ T cell clones were shown to be MHC class I (H-2Kd or H-2Db) restricted, whereas xeno-reactive CD4+ T cell clones were MHC class II (I-Ag7) restricted, indicating that xeno-reactivity reflects genuine human T cell response directed against allo-antigens present on murine cells. Despite production of high levels of IFN-gamma, xeno-reactive CD8+ and CD4+ T cell clones failed to exert cytolytic activity against murine DC, as determined in a 51Cr-release cytotoxicity assay. Absence of cytolysis by CD8+ T cell clones, which are generally considered as potent effector cells, may be explained by low avidity interaction between human T cells and murine DC, since flowcytometric analysis revealed sub-optimal activation of T cells as measured by CD137 expression and T cell receptor downregulation upon co-culture with murine DC, and therefore these results indicate that xenogeneic GvHD in this model is likely to be mediated by cytokines. In conclusion, in leukemia-engrafted NOD/scid mice treated with CD3+ DLI, we show that GvL reactivity and xenogeneic GvHD are mediated by separate human T cells with distinct specificities. All xeno-reactive T cell clones showed genuine recognition of MHC class I or class II associated allo-antigens on murine cells similar as GvHD-inducing human T cells. These data suggest that our NOD/scid mouse model of human acute leukemia may be valuable for studying the effectiveness and specificity of selectively enriched or depleted T cells for adoptive immunotherapy. Disclosures: No relevant conflicts of interest to declare.
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Firer, Michael A., Sofi Yado, Luboshits Galia, and Reuven Or. "A New Cellular Immunotherapy for the Treatment of Multiple Myeloma without Development of Graft-Versus-Host Disease." Blood 132, Supplement 1 (November 29, 2018): 3339. http://dx.doi.org/10.1182/blood-2018-99-110292.
Abstract:
Abstract Multiple myeloma (MM) is a malignant plasma cell disorder that accounts for approximately 10% of all hematological cancers. Despite recent advances, long-term survival is rare after autologous bone marrow transplantation (auto-BMT) and/or treatment with recently introduced anti-myeloma agents and disease recurs in virtually all patients. Allogeneic bone marrow transplantation (allo-BMT) is an effective treatment that can provide partial or complete remission in patients with MM. The therapeutic potential of allo-BMT is attributed to the "graft-versus-myeloma" (GvM) effect that aims to destroy residual tumor cells that survived an induction protocol of chemotherapy/radiotherapy and to maintain immune surveillance to prevent relapse. However, allo-BMT remains a controversial treatment, since the donor T cells that mediate the GvM effect are also the source of the cells that react to other tissue alloantigens and induce graft versus-host disease (GvHD), a major cause of morbidity and mortality in allo-BMT recipients. Nonetheless, allo-BMT remains the only potentially curable treatment for MM. Recently TCR Vβ CDR3-size spectratype analyses in an animal model of MM identified T cells subfamilies involved in the anti-host and anti-tumor reactivity. We have now carried this work further and tested the potential of integrating auto-BMT with a donor lymphocyte infusion (DLI) composed only of anti-MM reactive Vβ 2, 3 and 8.3 T cell subfamilies. The results demonstrate that these T cell subsets are indeed involved in the generation of a potent GvM response in MM bearing mice and is associated with enhanced survival. Importantly, the GvM response was not accompanied by the development of GvHD. Nonetheless, the GvM response was not sufficient to completely inhibit relapse. We then pre-stimulated donor T cells with MM cells in vitro in the presence of co-stimulatory factors and found that our selective DLI protocol induced a vigorous and long-lasting GvM which translated into long-term survival, again in the complete absence of GvHD. Interestingly, quite similar results were obtained by treating MM-bearing mice with repeat doses of naïve donor Vβ 2, 3 and 8.3 T cell subfamilies. The treated mice showed significantly lower serum paraprotein levels and myeloma infiltration in bone marrow and spleen. Taken together, the results suggest that a transplantation protocol involving only tumor cell-reactive donor T cell subfamilies might be devised for MM patients that results in a potent GvM with enhanced survival but without symptoms of GvHD. Disclosures No relevant conflicts of interest to declare.
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Burrack, Adam, Maki Nakayama, Laurie Landry, Marilyne Coulombe, and Ronald Gill. "Autoimmunity as an endogenous source of heterologous alloimmunity (TRAN3P.868)." Journal of Immunology 192, no. 1_Supplement (May 1, 2014): 202.7. http://dx.doi.org/10.4049/jimmunol.192.supp.202.7.
Abstract:
Abstract Curiously, although autoimmunity is restricted to self MHC molecules, spontaneously diabetic NOD mice reject MHC-unrelated islet allografts more rapidly than syngeneic (NOD) grafts. We hypothesized that the autoimmune (islet-reactive) T cell repertoire harbors a subset of TCRs that also cross-react to allogeneic MHC molecules. To test this concept, we performed high throughput TCR sequencing of T cells infiltrating MHC-mismatched (C3H, H-2k) islet allografts on the day of rejection in diabetic female NOD mice. TCRs expressed in high frequency in the allograft (>1%) were cloned and tested for islet autoreactivity and/or alloreactivity following TCR cDNA transfection into IL-2 producing hybridoma cells. Of note, three TCRs studied that were the most frequent in the graft were also detected in the endogenous pancreas. All three transfectants expressing these TCRs were islet reactive. Importantly, all three of these TCR also were directly alloreactive to C3H spleen cells. Two of the three TCRs were MHC class I reactive (anti-H-2Dk), and the other was MHC class II-reactive (anti-I-Ak). These results suggest that dual auto/allo-reactive T cells are selectively expanded in islet allografts. As such, we propose that autoimmunity is an unusual source of endogenous heterologous alloreactivity that may accelerate allograft rejection by simultaneous auto- and allo-reactivity.
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Pilunov, Artem, Dmitrii S. Romaniuk, Anton Shmelev, Savely Sheetikov, Anna N. Gabashvili, Alexandra Khmelevskaya, Dmitry Dianov, et al. "Transgenic HA-1-Specific CD8+ T-Lymphocytes Selectively Target Leukemic Cells." Cancers 15, no. 5 (March 3, 2023): 1592. http://dx.doi.org/10.3390/cancers15051592.
Abstract:
A significant share of allogeneic hematopoietic stem cell transplantations (allo-HSCT) results in the relapse of malignant disease. The T cell immune response to minor histocompatibility antigens (MiHAs) promotes a favorable graft-versus-leukemia response. The immunogenic MiHA HA-1 is a promising target for leukemia immunotherapy, as it is predominantly expressed in hematopoietic tissues and presented by the common HLA A*02:01 allele. Adoptive transfer of HA-1-specific modified CD8+ T cells could complement allo-HSCT from HA-1- donors to HA-1+ recipients. Using bioinformatic analysis and a reporter T cell line, we discovered 13 T cell receptors (TCRs) specific for HA-1. Their affinities were measured by the response of the TCR-transduced reporter cell lines to HA-1+ cells. The studied TCRs showed no cross-reactivity to the panel of donor peripheral mononuclear blood cells with 28 common HLA alleles. CD8+ T cells after endogenous TCR knock out and introduction of transgenic HA-1-specific TCR were able to lyse hematopoietic cells from HA-1+ patients with acute myeloid, T-, and B-cell lymphocytic leukemia (n = 15). No cytotoxic effect was observed on cells from HA-1- or HLA-A*02-negative donors (n = 10). The results support the use of HA-1 as a target for post-transplant T cell therapy.
50
Staba, Susan Lynn, Melissa Reese, and Joanne Kurtzberg. "Immunophenotype and Analysis of Allo-Reactivity of Umbilical Cord Blood (UCB)-Derived T-Cells Following Ex-Vivo Expansion." Blood 104, no. 11 (November 16, 2004): 403. http://dx.doi.org/10.1182/blood.v104.11.403.403.
Abstract:
Abstract Introduction: Unrelated umbilical cord blood (UCB) is an effective source of allogeneic hematopoietic stem cells for transplantation therapy in patients lacking matched bone marrow donors. The major cause of mortality post transplant is opportunistic infection, due in part to the immunologic naiveté of the UCB T-cells. We have been able to expanded UCB derived T-cells utilizing a skin stromal layer with a supplemental cytokine cocktail. To assist in determining a target population for ex-vivo immunization and to predict the likelihood of inducing GVHD by reinfusion of alloreactive T cells, we identified the immunophenotype of these cells using flow cytometry and assessed their allo-reactivity utilizing mixed lymphocyte culture. The long term goal is to provide UCB-derived adoptive immunotherapy for UCBT recipients. Methods: Patient derived skin fibroblasts were cultured in a media of IMDM, 10% fetal calf serum and 10% horse serum for 14 days, then irradiated to 5,000cGy. Cryopreserved UCB cells were thawed and cultured on this stromal layer in the same media supplemented with a cytokine cocktail of interleukin-7 (IL-7) (10ng/ml), flt-3 ligand (10ng/ml) stem cell factor (50ng/ml) for 14 days. Interleukin-2 (IL-2) (100u/ml) was then added and the UCB cells remained in culture for a total of 28 days. Cell count, viability, and extensive flow cytometric analysis were performed. In addition we utilized mixed lymphocyte culture to compare unmanipulated and expanded UCB reactivity to patient derived lyphoctyes after the first and second phase of expansion. Results: Mean fold increases of 9 in CD4+, 11 in CD8+ and 53 in CD4+/CD8+ cells were seen. Of the expanded T cells cells, 81% were CD4+, 20% CD8+ and 3% CD4+/8+. Virtually 100% were alpha-beta T cells. One percent were 15/56+, 79% CD62L(L-selectin)+ and 2.4% CD25(IL-2R)+ . In mixed lymphocyte culture with patient derived lyphocytes, preliminary data suggests that there is no difference in reactivity of the UCB after either the first or second phase of ex-vivo expansion when compared with an unmanipulated UCB fraction. Conclusions: The expansion of T-cells from unfractionated, red blood cell depleted, cryopreserved UCB can be accomplished using a cytokine cocktail of IL-7, Flt 3 ligand, SCF and IL-2 over a patient derived, irradiated skin stromal layer, with maximal expansion in the immature CD4+/CD8+ T-cell subset. The expanded T cells were alpha-beta T cells, which would allow immunization of these cells in standard fashion via co-culture with professional APCs loaded with target antigens. The absence of delta-gamma T cells as well as the small percentage expressing CD 25 is not suggestive of GVHD inducing populations. In mixed lymphocyte culture with patient derived lymphocytes, it does not appear that the expanded cord blood is more alloreactive then unmanipulated cord blood, despite the exposure in culture to patient derived skin fibroblasts and stimulation with cytokines. We hypothesize that the expanded T cells could serve as a target population for adoptive immunotherapy without inducing additional GVHD. Further testing will determine the optimal cellular target and conditions for ex vivo immunization against various viral and fungal antigens. The ex-vivo expanded and immunized T cells could then be reinfused into the patient to augment immune reconstitution and decrease infection related morbidity and mortality in the months following UCBT.