Rauser, G. et al. Rapid generation of combined CMV-specific CD4+ and CD8+ T-cell lines for adoptive transfer into recipients of allogeneic stem cell transplants. Blood 103, 3565-3572

Medizinische Klinik II, Eberhard-Karls-Universität, Tuebingen, Germany.
Blood (Impact Factor: 10.45). 06/2004; 103(9):3565-72. DOI: 10.1182/blood-2003-09-3056
Source: PubMed


Adoptive transfer of cytomegalovirus (CMV)-specific T cells can restore long-lasting, virus-specific immunity and clear CMV viremia in recipients of allogeneic stem cell transplants if CD4(+) and CD8(+) CMV-specific T cells are detected in the recipient after transfer. Current protocols for generating virus-specific T cells use live virus, require leukapheresis of the donor, and are time consuming. To circumvent these limitations, a clinical-scale protocol was developed to generate CMV-specific T cells by using autologous cellular and serum components derived from a single 500-mL blood draw. CMV-specific T cells were stimulated simultaneously with CMV-specific major histocompatibility complex class I (MHC I)- restricted peptides and CMV antigen. Activated T cells were isolated with the interferon-gamma (IFN-gamma) secretion assay and expanded for 10 days. In 8 randomly selected, CMV-seropositive donors, 1.34 x 10(8) combined CD4(+) and CD8(+) CMV-specific T cells, on average, were generated, as determined by antigen-triggered IFN-gamma production. CMV-infected fibroblasts were efficiently lysed by the generated T cells, and CMV-specific CD4(+) and CD8(+) T cells expanded if they were stimulated with natural processed antigen. On the other hand, CD4(+) and CD8(+) T cell-mediated alloreactivity of generated CMV-specific T-cell lines was reduced compared with that of the starting population. In conclusion, the culture system developed allowed the rapid generation of allodepleted, highly enriched, combined CD4(+) and CD8(+) CMV-specific T cells under conditions mimicking good manufacturing practice.

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Available from: Georg Rauser, Feb 05, 2014
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    • "Briefly, CD4+ T cells were enriched by depletion of non-CD4+ T cells, CD2− APC were enriched by depletion of CD2+ cells and monocytes were positive selected by enrichment of CD14+ cells. Monocyte-derived DCs were generated as described previously.42 "
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    ABSTRACT: Unlike induced Foxp3(+) regulatory T cells (Foxp3(+) iTreg) that have been shown to play an essential role in the development of protective immunity to the ubiquitous mold Aspergillus fumigatus, type-(1)-regulatory T cells (Tr1) cells have, thus far, not been implicated in this process. Here, we evaluated the role of Tr1 cells specific for an epitope derived from the cell wall glucanase Crf-1 of A. fumigatus (Crf-1/p41) in antifungal immunity. We identified Crf-1/p41-specific latent-associated peptide(+) Tr1 cells in healthy humans and mice after vaccination with Crf-1/p41+zymosan. These cells produced high amounts of interleukin (IL)-10 and suppressed the expansion of antigen-specific T cells in vitro and in vivo. In mice, in vivo differentiation of Tr1 cells was dependent on the presence of the aryl hydrocarbon receptor, c-Maf and IL-27. Moreover, in comparison to Tr1 cells, Foxp3(+) iTreg that recognize the same epitope were induced in an interferon gamma-type inflammatory environment and more potently suppressed innate immune cell activities. Overall, our data show that Tr1 cells are involved in the maintenance of antifungal immune homeostasis, and most likely play a distinct, yet complementary, role compared with Foxp3(+) iTreg.Immunology and Cell Biology advance online publication, 13 May 2014; doi:10.1038/icb.2014.34.
    Immunology and Cell Biology 05/2014; 92(8). DOI:10.1038/icb.2014.34 · 4.15 Impact Factor
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    • "During EBV and CMV infections percentages of virus specific CD8 + T-cells binding to tetramers as high as 35% have been reported, depending on the viral antigen and HLA-type of the donor [41] [42] [43] [44]. After adoptive transfer of HAdV-specific T-cells induced by culturing PBMCs with HAdV lysate, the cells were not further expanded in vitro and even low numbers of transferred cells were able to expand in vivo and control the infection [37]. "
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    ABSTRACT: Adenoviruses (HAdV) can cause life threatening infections, especially in paediatric patients after allogeneic stem cell transplantation (SCT). Yet, no effective antiviral medication is available. One treatment option is adoptive transfer of HAdV-specific T-cells from the graft donor into the patient. Especially CD4+ T-cells are critical to control HAdV infection. To allow for applicability of CD4+ T-cells in adoptive therapy, sufficient numbers of HAdV-specific T-cells with low levels of residual alloreactive T-cells are required. In this study, we explored the possibility to selectively expand and isolate functional HAdV-specific T-cells from PBMCs in response to 15-mer peptides using artificial antigen-presenting cells (aAPCs), composed of liposomes harbouring HAdV-peptide/HLA-Class-II complexes. HAdV-specific T-cells generated using this method produce mainly pro-inflammatory cytokines, express perforin and granzyme B, kill HAdV-infected cells effectively and are not alloreactive. Thus, the generation and isolation of HAdV-specific CD4+ T-cells seem a critical step towards specific adoptive therapy for HAdV infections after allogeneic SCT.
    Clinical Immunology 03/2013; 146(3):228-39. DOI:10.1016/j.clim.2013.01.002 · 3.67 Impact Factor
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    • "It avoids safety and regulatory issues associated with prolonged T-cell culture and potential viral biohazards. Cytokine secretion assays using recombinant proteins or synthetic peptide pools consisting of overlapping peptides spanning an entire immunodominant protein are not restricted by HLA variations, and they enable the generation of CD4+ and/or CD8+ T-cell responses to multiple epitopes (Rauser et al., 2004). "
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    ABSTRACT: Infection with and reactivation of human cytomegalovirus (CMV), Epstein-Barr virus (EBV), and adenovirus (ADV) are frequent and severe complications in immunocompromised recipients after hematopoietic stem cell transplantation (HSCT) or solid organ transplantation (SOT). These serious adverse events are associated with significant morbidity and mortality. Donor lymphocyte infusions (DLIs) are often used to treat both viral infections and leukemia relapses after transplantation but are associated with potentially life-threatening graft-versus-host disease (GvHD). Adoptive immunotherapy with virus-specific cytotoxic effector T cells (CTLs) derived from seropositive donors can rapidly reconstitute antiviral immunity after HSCT and organ transplantation. Therefore, it can effectively prevent the clinical manifestation of these viruses with no significant acute toxicity or increased risk of GvHD. In conditions, where patients receiving an allogeneic cord blood transplant or a transplant from a virus-seronegative donor and since donor blood is generally not available for solid organ recipients, allogeneic third party T-cell donors would offer an alternative option. Recent studies showed that during granulocyte colony-stimulating factor (G-CSF) mobilization, the functional activity of antiviral memory T cells is impaired for a long period. This finding suggests that even stem cell donors may not be the best source of T cells. Under these circumstances, partially human leukocyte antigen (HLA)-matched virus-specific CTLs from healthy seropositive individuals may be a promising option. Therefore frequency assessments of virus-specific memory T cells in HLA-typed healthy donors as well as in HSCT/SOT donors using a high throughput T-cell assay were performed over a period of 4 years at Hannover Medical School. This chapter will address the relevance and potential of a third-party T-cell donor registry and will discuss its clinical implication for adoptive T-cell immunotherapy
    Frontiers in Immunology 01/2012; 3:410. DOI:10.3389/fimmu.2012.00410
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