Adoptive Cellular Transfer Research Articles

Adoptive transfer of immunity against red mouth disease in rainbow trout (Oncorhynchus mykiss)

Enteric Red Mouth Disease (ERM) caused by Yersinia ruckeri is among the bacterial diseases in rainbow trout aquaculture that can be controlled by registered vaccines and part of these vaccines can be delivered by immersion which is the most convenient way of application inducing good antibody responses and ensuring decreased handling costs. However, the mechanisms by which fish are protected against Yersinia ruckeri are unclear. To shed light on this question we have bath vaccinated clonal rainbow trout donors by using a commercially available vaccine, and leukocytes have been isolated from vaccinated donors followed by adoptive cell transfer to naive fish recipients of the same trout clone. Control recipients were injected with same numbers of leukocytes from non-vaccinated clonal donors. Transfer between genetically identical donors and recipients ensured that immune cells were not rejected by the recipients. After transfer, both groups of recipients were bath infected with a virulent field strain of Yersinia ruckeri. While rainbow trout that have received leukocytes from vaccinated donors showed no mortality, less than 40% of control recipients survived the challenge. Flow cytometry using leukocytes from recipient survivors revealed that the percentage of IgM B cells was considerably higher in recipients that were injected with leukocytes from vaccinated donors when compared with control recipients, while no differences in the percentages of CD8 cells among groups were recorded. Being the first report on adoptive cellular transfer of immunity in a bacterial infection in fish these data suggest that immunity was rather transferred by IgM B cells, but not by CD8 cytotoxic T cells. . It can be further suggested that bath vaccination has caused an effective systemic immune response since protection could be transferred by non-mucosal leukocytes.

Trafficking of T cells into tumors.

T cells are a crucial component of the immune response to infection and cancer. In addition to coordinating immunity in lymphoid tissue, T cells play a vital role at the disease site, which relies on their efficient and specific trafficking capabilities. The process of T-cell trafficking is highly dynamic, involving a series of distinct processes, which include rolling, adhesion, extravasation, and chemotaxis. Trafficking of T cells to the tumor microenvironment is critical for the success of cancer immunotherapies such as adoptive cellular transfer. Although this approach has achieved some remarkable responses in patients with advanced melanoma and hematologic malignancy, the success against solid cancers has been more moderate. One of the major challenges for adoptive immunotherapy is to be able to effectively target a higher frequency of T cells to the tumor microenvironment, overcoming hurdles associated with immunosuppression and aberrant vasculature. This review summarizes recent advances in our understanding of T-cell migration in solid cancer and immunotherapy based on the adoptive transfer of natural or genetically engineered tumor-specific T cells and discusses new strategies that may enhance the trafficking of these cells, leading to effective eradication of solid cancer and metastases.

T-Cell Therapy for Multiple Myeloma Using NY-ESO-1+ T-Cell Antigen Presenting Cells (T-APC) Combined with Adoptive Cellular Transfer (ACT) to Augment Immunotherapy

T cells have been previously genetically manipulated to act as a cellular vaccine to present viral antigens such as MP1 from influenza A. We have now genetically modified T cells as antigen presenting cells (T-APC) to express the tumor-associated antigen (TAA) NY-ESO-1 (Figure 1). This TAA is reportedly found in the majority of patients with high risk multiple myeloma, as well as other malignancies, but expression is absent from normal tissues. Therefore, immunologic targeting of NY-ESO-1 may be a potential treatment strategy for myeloma that is resistant to conventional therapies. One approach to improve therapeutic outcome is by adoptive transfer of T cells rendered specific for a TAA preferentially expressed on tumor cells. T-cell specificity can be redirected to intracellular TAAs by enforced expression of a characterized T-cell receptor (TCR) or chimeric antigen receptor (CAR) that recognizes processed antigen in the context of restricting human leukocyte antigen (HLA). However, persistence of these infused genetically modified cells, which is directly related to a favorable clinical response, may be variable. Therefore, T-APC may have a dual function of increasing persistence of adoptively transferred CAR+ and/or TCR+ T cells as well as inducing long term immunity through direct and cross priming of endogenous immune cells. To enhance these presentation functions, a new co-stimulatory molecule that tethers IL-15 to the cell surface (membrane-bound IL-15, mIL15) was expressed on NY-ESO-1+T-APC using the Sleeping Beauty (SB) gene transfer system. NY-ESO-1-specific HLA A2+ T cells expressing TCR and CAR could be selectively propagated ex vivo on autologous mIL15+NY-ESO-1+T-APC. The T-APC were also able to selectively propagate NY-ESO-1-specific T cells from autologous peripheral blood which can be described by binding of specific pentamer (Figure 2). Our data demonstrate that it is feasible to generate T-APC that can activate T cells engineered to have a defined specificity for NY-ESO-1 as well as grow out NY-ESO-1-specific T cells. The SB system is in place for genetic modification of clinical-grade T cells to express CAR, TCR, NY-ESO-1, and mIL15. Thus, we are proceeding to a human trial to infuse T-APC with and without genetically modified T cells in patients with NY-ESO-1+ tumors such as myeloma. [Display omitted] [Display omitted] DisclosuresCooper:InCellerate: Equity Ownership; Sangamo: Patents & Royalties; Targazyme: Consultancy; GE Healthcare: Consultancy; Ferring Pharmaceuticals: Consultancy; Fate Therapeutics: Consultancy; Janssen Pharma: Consultancy; BMS: Consultancy; Miltenyi: Honoraria.

In vivo distribution and antitumor effect of infused immune cells in a gastric cancer model

Adoptive cellular transfer has been employed for cancer immunotherapy, including patients with gastric cancer. However, little is known about the distribution of effector cells after their injection via different pathways. In this study, we used human gastric cancer cells (BGC823) tagged with enhanced green fluorescent protein (EGPF) to establish a subcutaneous gastric cancer model in nude mice. Cytokine-induced killer (CIK) cells and cytotoxic T lymphocytes (CTLs) were generated from human peripheral blood and labeled with red fluorescent PKH26. A portion of CIK cells was armed with CEA/CD3-bispecific single-chain antibody. When CIK cells were injected into nude mice with established subcutaneous gastric cancer via peritumoral (p.t.), intravenous (i.v.) and intraperitoneal (i.p.) infusion respectively, the distribution of cells was observed using a live fluorescence imaging system. We found that only a very small number of CIK cells could travel to the tumor site after i.p. or i.v. infusion, and they inhibited subcutaneous tumor growth in vivo only immediately following injection. In contrast, p.t. injection resulted in a significantly higher accumulation of CIK cells at the tumor site for 48 hours and mediated the greatest tumor inhibition compared with the other two injection methods. In addition, we compared the antitumor activity of CIK, CEA/CD3-bscAb-CIK and CTL cells in vitro and in vivo after p.t. injection. Among the three types of immune cells, CTLs demonstrated the strongest antitumor activity both in vitro and in vivo. CEA/CD3-bispecific single chain antibody could effectively link T lymphocytes and tumor cells expressing CEA, and resulted in significantly higher accumulation of CIK cells at the tumor site compared with the parental CIK cells. This study indicates that peritumoral injection of immune effector cells by minimally invasive surgical procedures represents an effective delivery method of adoptive cellular immunotherapy. Tumor-specific immune cells, such as CTLs, are a better choice of effector cells than CIKs in cellular immunotherapy. Furthermore, CD3+ immune cells armed with the CEA/CD3-bispecific single chain antibody could more effectively travel to and accumulate at the site of tumors expressing CEA, such as gastric cancer.

The Preparative Method and Clinical Application of Autologous memory T Lymphocyte.

Abstract 5123 BackgroundThe term autolymphocyte therapy refers to an adoptive cellular transfer strategy based on the ex vivo activation of memory T cells. The assumption is that because patients with metastatic cancer have been previously exposed to tumor antigens, memory T cells exist which can be collected and activated ex vivo. The result in autologous cytokines that can be used to activate other T cells. The method is simpler than it sounds and basically consisted of 2 cell collections using a cell separator and 2 cell cultures. The supernatant from the first culture, which contains the desired cytokines, is used to activate the second cohort of T cells. ObjectiveTo establish a preparative method of autologous memory T lymphocyte and use this T lymphocyte to treat hematologic malignancies and solid tumor. Method14 patients of hematologic malignancies and solid tumor had autolymphocyte therapy in 4 hospitals since 2002(Plasmacytoma1,NHL1, colon cancer3,HCC3,NSCLC1, nasopharyngeal carcinoma3, breast cancer1, Ovarian cancer1). These patients, including 11 male and 3 female, with a mean age of 55 (range, 25-71), were finally confirmed by pathology and histology autologouslymphokine, anti-CD3 antibody, IL-2. Cimetidine and Indomethacin induced peripheral blood mononuclear cell (PBMC) ex vivo to memory T lymphocyte by two steps. We tested memory T lymphocyte by flow cytometry before and after culture, we observed side effect during infusion. ResultAfter culturing peripheral blood lymphocyte for 9 days, the number of cell and CD4+/CD45RO+T cell increased. The data demonstrated: PBMC before culture was 1.07-67.2×109(median was 2.16×109),the cell after culture was 2.0-6.5×109(median was 3.2×109),CD4+/CD45RO+T cell rate was 27.59-86%(median was 77.76%) before culture and 82.09-99.55%(median was 89.49%) after culture. The absolute number of CD4+/CD45RO+T cell increased. Among 14 patients after infusion, one patient who had no medicine before infusion had fever of 39°C, 4 patients had low-grade fever and the rest 9 patients had no side effect. Follow-up: 5 patients abroad had no follow-up. In 1 to 10 months after therapy, all patients were alive. 4 patients were in partial remission and 5 patients in stable stage. ConclusionAutologous memory T lymphocyte is a safe and an effective way to treat advanced-stage hematologic malignancies and solid tumor. The mechanism and long-tem effect need more research and observe. DisclosuresNo relevant conflicts of interest to declare.

Autoantigen-Specific IL-10-Transduced T Cells Suppress Chronic Arthritis by Promoting the Endogenous Regulatory IL-10 Response

Deficient T cell regulation can be mechanistically associated with development of chronic autoimmune diseases. Therefore, combining the regulatory properties of IL-10 and the specificity of autoreactive CD4(+) T cells through adoptive cellular gene transfer of IL-10 via autoantigen-specific CD4(+) T cells seems an attractive approach to correct such deficient T cell regulation that avoids the risks of nonspecific immunosuppressive drugs. In this study, we studied how cartilage proteoglycan-specific CD4(+) T cells transduced with an active IL-10 gene (T(IL-10)) may contribute to the amelioration of chronic and progressive proteoglycan-induced arthritis in BALB/c mice. TCR-transgenic proteoglycan-specific T(IL-10) cells ameliorated arthritis, whereas T(IL-10) cells with specificity for OVA had no effect, showing the impact of Ag-specific targeting of inflammation. Furthermore, proteoglycan-specific T(IL-10) cells suppressed autoreactive proinflammatory T and B cells, as T(IL-10) cells caused a reduced expression of IL-2, TNF-alpha, and IL-17 and a diminished proteoglycan-specific IgG2a Ab response. Moreover, proteoglycan-specific T(IL-10) cells promoted IL-10 expression in recipients but did not ameliorate arthritis in IL-10-deficient mice, indicating that T(IL-10) cells suppress inflammation by propagating the endogenous regulatory IL-10 response in treated recipients. This is the first demonstration that such targeted suppression of proinflammatory lymphocyte responses in chronic autoimmunity by IL-10-transduced T cells specific for a natural Ag can occur via the endogenous regulatory IL-10 response.

Persistence, Immune Specificity, and Functional Ability of Murine Mutant Ras Epitope-Specific CD4+ and CD8+ T Lymphocytes Following in Vivo Adoptive Transfer

Adoptive T-cell transfer has been shown to be a potentially effective strategy for cellular immunotherapy in some murine models of disease. However, several issues remain unresolved regarding some of the basic features involved in effective adoptive transfer, such as the influence of specific peptide antigen (Ag) boost after T-cell transfer, the addition of IL-2 post-T-cell transfer, the trafficking of transferred T cells to lymphoid and nonlymphoid tissues, and the functional stability of recoverable CD4+ and CD8+ T cells. We investigated several of these parameters, particularly as they relate to the persistence and maintenance of effector functions of murine CD4+ and/or CD8+ T lymphocytes after adoptive cellular transfer into partially γ-irradiated syngeneic hosts. Our laboratory previously identified murine (H-2d) immunogenic CD4+ and CD8+ T-cell peptide epitopes reflecting codon 12 ras mutations as tumor-specific Ag. Therefore, the model system chosen here employed epitope-specific MHC class II-restricted CD4+ T cells and MHC class I-restricted CD8+ T cells produced from previously immunized BALB/c mice. Between 2 and 7 days after T-cell transfer, recipient mice received various combinations of peptide boosts and/or IL-2 treatments. At different times after the T-cell transfer, spleen and lung tissues were analyzed phenotypically to monitor the persistence of the immune T cells and functionally (via proliferation or cytotoxicity assays) to assess the maintenance of peptide specificity. The results showed that immune donor T lymphocytes (uncultured immune T cells or cloned T cells) were recoverable from the spleens and lungs of recipient mice after transfer. The recovery of Ag-specific T-cell responses was greatest from recipient mice that received peptide boosts and IL-2 treatment. However, mice that received a peptide boost without IL-2 treatment responded nearly as well, which suggested that including a peptide boost after T-cell transfer was more obligatory than exogenous IL-2 treatment to sustain adoptively transferred T cells in vivo. Ag-specific T-cell responses were weak in mice that either received IL-2 alone or did not receive the cognate peptide boost after T-cell transfer. The T-cell clones were also monitored by flow cytometry or RT-PCR based on expression of the T-cell receptor Vβ-chain, which was previously characterized. Ag-specific T cells were recovered from both spleens and lungs of recipient mice, demonstrating that the T-cell clones could localize to both lymphoid and nonlymphoid tissues. This study demonstrates that both uncultured and in vitro-cloned T lymphocytes can migrate to lymphoid tissues and nonlymphoid (e.g., lung) tissues in recipient hosts and that their functional activities can be maintained at these sites after transfer, if they are exposed to peptide Ag in vivo.

Adoptive immunotherapy as an in vivo model to explore antitumor mechanisms induced by a recombinant anticancer vaccine.

We have described previously the construction, generation, and in vivo biologic consequences of a recombinant vaccinia virus containing the human CEA gene (rV-CEA) in an experimental murine colon carcinoma model. Immunization of C57BL/6 mice with rV-CEA led to antigen-specific inhibition of tumor growth in both prophylactic and therapeutic settings. Although such antitumor effects were correlated with the induction of CEA-specific T-cell responses, their exact contribution in the tumor rejection mechanism remained unclear. In this study, we examined the mechanism of action of rV-CEA, with emphasis on definition of the immune cells important for such antitumor effects. To that end, a cellular adoptive transfer model was established in vivo, which allowed specific functional analysis of donor-derived immune cells in naive, sublethally irradiated, tumor-bearing recipients. Splenocytes from rV-CEA-immunized donors expressed strong antitumor activity in such tumor-bearing recipients, whereas nonimmune donor cells did not. Depletion of immune T cells before cellular transfer abolished the antitumor response. Moreover, depletion of CD8+ T cells before transfer resulted in the loss of antitumor activity, despite the presence of CD4+ T cells. In contrast, antitumor activity was demonstrable with CD8-containing, CD4-depleted effectors, although it was not as effective as with both T-cell subpopulations combined. Finally, in beta 2-microglobulin/CD8+ T-cell-deficient mice, rV-CEA immunization exerted only partial antitumor protection, compared with the immune-competent controls. Overall, we demonstrated that (a) antitumor activity induced by rV-CEA was essentially mediated by CD8+ effectors; and (b) the combination of both CD8+ and CD4+ lymphocytes led to maximal antitumor therapeutic effects, suggesting an important helper or immunoregulatory contribution of the CD4+ subset. Thus, adoptive cellular transfer strategies may have implications for both the study of recombinant anticancer vaccines and the development of potential clinical applications for cancer immunotherapy.

Preclinical studies, experimental therapeutics, and clinical management of advanced melanoma.

The preclinical and clinical investigations of therapeutic approaches to melanoma have progressed significantly in the past year. This article reviews recently published information relevant to preclinical studies of the biologic function of stromal tissue, growth factors (and receptors), interferons, cytokines, and antimetabolites as well as clinical studies of agents that may alter host-tumor interactions. Data published over the past year include new advances in the use of cytokines with and without adoptive cellular transfer, as well as tumor cell antigens for immunization and monoclonal antibodies for passive transfer. The data suggest that the age-old dogma that no medical treatment of melanoma prolongs survival may soon be a thing of the past.