Mechanisms Of Tolerance Induction Research Articles

Antigenic strength controls the generation of antigen‐specific IL‐10‐secreting T regulatory cells

Administration of peptides i.n. induces peripheral tolerance in Tg4 myelin basic protein-specific TCR-Tg mice. This is characterized by the generation of anergic, IL-10-secreting CD4+ T cells with regulatory function (IL-10 Treg). Myelin basic protein Ac1-9 peptide analogs, displaying a hierarchy of affinities for H-2 A(u) (Ac1-9[4K]<<[4A]<[4Y]), were used to investigate the mechanisms of tolerance induction, focusing on IL-10 Treg generation. Repeated i.n. administration of the highest affinity peptide, Ac1-9[4Y], provided complete protection against EAE, while i.n. Ac1-9[4A] and Ac1-9[4K] treatment resulted in only partial protection. Ac1-9[4Y] was also the most potent stimulus for IL-10 Treg generation. Although i.n. treatment with Ac1-9[4A] gave rise to IL-10-secreting CD4+ T cells, the population as a whole was also capable of secreting IFN-gamma after an in vitro recall response to Ac1-9[4A] or [4Y]. In addition to IL-10 production, other facets of tolerance, namely, anergy and suppression (both in vitro and in vivo), were affinity dependent, with i.n. Ac1-9[4Y]-, [4A]- or [4K]-treated CD4+ T cells being the most, intermediate and least anergic/suppressive, respectively. These findings demonstrate that the generation of IL-10 Treg in vivo is driven by high signal strength.

Cow's milk protein sensitisation in neonatal necrotising enterocolitis is replaced by increased transforming growth factor-beta1 response after recovery: a mechanism for tolerance induction

<h3>Background</h3> The authors have previously made a novel observation of cow9s milk protein (CMP) sensitisation of the peripheral blood mononuclear cells (PBMCs) in acute necrotising enterocolitis (NEC).1 These findings may be of clinical relevance in view of the association between NEC and formula feeding. The role of regulatory/tolerogenic cytokines has not been investigated, neither the fate of this phenomenon following recovery form NEC. <h3>Objective</h3> Evidence of cow9s milk protein specific, effector and regulatory, cytokine response is investigated in preterm infants in acute NEC, and at term compared to healthy preterm controls. <h3>Methods</h3> 28 preterm infants, 14 stage II or III modified Bell9s classification NEC babies (median postconceptional age (PCA); 32.5 (range 29–36 weeks)) and 14 PCA-matched healthy controls (median PCA; 32.5 (range 29–36) weeks) were recruited. Unstimulated, and antigen (casein and β-lactoglobulin (β-lg))- and mitogen-stimulated interferon (IFN)-γ, interleukin (IL)-4, IL-10, and transforming growth factor (TGF)-beta1-secreting cells in PBMCs were counted by the single-cell enzyme linked immunospot (ELISPOT) assay. <h3>Results</h3> During the acute-stage disease, preterm NEC babies showed a general pattern of higher responses to PHA compared to healthy preterm controls (10 fold increase) for IFN-γ, IL-4, IL-10 (p=0.002) and threefold for TGF-β1, strong responses to β-lg (10-fold increase) for IFN-γ, IL-4 and IL-10 (p=0.002) and 3 fold for TGF-beta1 and a smaller (fivefold) casein responses (p=0.002) and threefold for TGF-beta1. At term, following clinical recovery in NEC cases, the IFN-γ, IL-4, IL-10 response to β-lg and casein declined (between 3 and 10-fold), whereas TGF-beta1 secretion response had increased by eight and sixfold (p=0.002). <h3>Conclusion</h3> The change in cytokine response profile to dietary antigens (fall in IFN-γ, IL-4, IL-10, increase in TGF-beta1) from acute disease to term (recovered from NEC) may represent a mechanism by which infants become tolerant to CMP, following initial sensitisation.

Tolerance induction after specific immunotherapy with pollen allergoids adjuvanted by monophosphoryl lipid A in children

Specific immunotherapy (SIT) is a well-established and clinically effective treatment for allergic diseases. A pollen allergoid formulated with the T helper type 1 (Th1)-inducing adjuvant monophosphoryl lipid A (MPL) facilitates short-term SIT. Little is known about mechanisms of tolerance induction in this setting. In a prospective study, 34 patients allergic to grass pollen (25 male, nine female, median age 10.2 years) received a total of 44 SIT courses (20 in the first, 24 in the second) with MPL-adjuvanted pollen allergoids. Immunogenicity was measured by levels of specific immunoglobulin G (IgG(grass)) and IgG4(grass) by antibody blocking properties on basophil activation, and by induction of CD4(+), CD25(+) and forkhead box P3 (FoxP3(+)) regulatory T cells (T(reg)). Specific IgG and IgG4 levels increased only slightly in the first year of SIT. In the second year these changes reached significance (P < 0.0001). In keeping with these findings, we were able to show an increase of T(reg) cells and a decreased release of leukotrienes after the second year of treatment. In the first year of treatment we found little evidence for immunological changes. A significant antibody induction was seen only after the second course of SIT. Short-course immunotherapy with pollen allergoids formulated with the Th1-inducing adjuvant MPL needs at least two courses to establish tolerance.

Oral Immunotherapy In The Sensitized Mouse: Barriers To Tolerance Induction

RATIONALE: Oral immunotherapy (OIT) with food allergens results in desensitization, but permanent tolerance induction has not yet been demonstrated. Oral tolerance can be generated in mice and humans to a neo-antigen, but not a recall antigen. Our aim was to establish a mouse model of OIT to examine mechanisms of tolerance induction.

Different mechanisms regulate CD4+ T cell independent induction of oral and nasal tolerance of CD8+ T cells

Mucosal administration of antigens is known to induce antigen specific regulatory CD4(+) T cells, but less is known about the effects on CD8(+) T cell function. Using a murine model for mucosal tolerance induction, we show that both oral and nasal OVA (ovalbumin) application reduced OVA specific CD8(+) T effector cell numbers and suppressed in vivo cytotoxicity in response to subsequent immunisation. To investigate whether CD4(+) T cells are essential for oral or nasal CD8(+) T cell tolerance, we used MHC class II deficient mice. Normal CD8(+) T cell tolerance was observed in MHC class II deficient mice, indicating that CD4(+) T cells are not required for both oral and nasal CD8(+) T cell tolerance induction. To study the direct effects of mucosal antigen application on naive CD8(+) T cells, we adoptively transferred OVA specific transgenic CD8(+) T cells and analysed their fate after mucosal antigen application. Oral OVA application reduced the numbers of OVA specific CD8(+) T cells, whereas nasal OVA application induced proliferation. However, the expanded population of OVA specific CD8(+) T cells from nasally treated mice was unable to proliferate upon re-stimulation. In conclusion, our studies point to suppressive effects of the mucosal immune system directly on CD8(+) T cells and indicate multiple mechanisms of tolerance induction. More insight in these mechanisms of tolerance induction is necessary for the development of mucosal tolerance therapies for autoimmune diseases and allergy.

Suppression of HLA expression by lentivirus-mediated gene transfer of siRNA cassettes and in vivo chemoselection to enhance hematopoietic stem cell transplantation

Current approaches for hematopoietic stem cell (HSC) and organ transplantation are limited by donor and host-mediated immune responses to allo-antigens. Application of these therapies is limited by the toxicity of preparative and post-transplant immunosuppressive regimens and a shortage of appropriate HLA-matched donors. We have been exploring two complementary approaches for genetically modifying donor cells that achieve long-term suppression of cellular proteins that elicit host immune responses to mismatched donor antigens, and provide a selective advantage to genetically engineered donor cells after transplantation. The first approach is based on recent advances that make feasible targeted down-regulation of HLA expression. Suppression of HLA expression could help to overcome limitations imposed by extensive HLA polymorphisms that restrict the availability of suitable donors. Accordingly, we have recently investigated whether knockdown of HLA by RNA interference (RNAi) enables allogeneic cells to evade immune recognition. For efficient and stable delivery of short hairpin-type RNAi constructs (shRNA), we employed lentivirus-based gene transfer vectors that integrate into genomic DNA, thereby permanently modifying transduced donor cells. Lentivirus-mediated delivery of shRNA targeting pan-Class I and allele-specific HLA achieved efficient and dose-dependent reduction in surface expression of HLA in human cells, and enhanced resistance to allo-reactive T lymphocyte-mediated cytotoxicity, while avoiding non-MHC restricted killing. Complementary strategies for genetic engineering of HSC that would provide a selective advantage for transplanted donor cells and enable successful engraftment with less toxic preparative and immunosuppressive regimens would increase the numbers of individuals to whom HLA suppression therapy could be offered. Our second strategy is to provide a mechanism for in vivo selection of genetically modified HSC and other donor cells. We have uniquely combined transplantation during the neonatal period, when tolerance may be more readily achieved, with a positive selection strategy for in vivo amplification of drug-resistant donor HSC. This model system enables the evaluation of mechanisms of tolerance induction to neo-antigens, and allogeneic stem cells during immune ontogeny. HSC are transduced ex vivo by lentivirus-mediated gene transfer of P140K-O(6)-methylguanine-methyltransferase (MGMT(P140K)). The MGMT(P140K) DNA repair enzyme confers resistance to benzylguanine, an inhibitor of endogenous MGMT, and to chloroethylating agents such as BCNU. In vivo chemoselection enables enrichment of donor cells at the stem cell level. Using complementary approaches of in vivo chemoselection and RNAi-induced silencing of HLA expression may enable the generation of histocompatibility-enhanced, and eventually, perhaps "universally" compatible cellular grafts.

Tolerance Induction by Neonatal Gene Delivery

Advances in the design and efficiency of gene delivery vectors have enabled the initiation of clinical trials in gene therapy for genetic and other disorders. However, the development of inhibitory immune responses to vector antigens and to therapeutic proteins remains an obstacle. Efforts to limit these immune responses by immunosuppressive and immuno-modulatory approaches have met with limited success. Our approach is to deliver and express viral vectors early in immune ontogeny and thereby induce immune tolerance to both vectors and therapeutic proteins. We have previously shown that in utero delivery of AAV-2 vectors produces lifelong gene expression without immune responses, and that augmented levels of gene expression are achieved with re-administration of AAV vectors. Because fetal injections are limited by technical issues, our current focus is to use a neonatal model for defining the critical period when tolerance to vector and transgene may be achieved by primary injection. We are also exploring mechanisms of tolerance induction to neo-antigens.We have delivered AAV serotypes 1 and 8 with higher transduction efficiencies than AAV-2, to assess the expression levels, duration, and tissue distribution of luciferase by semi-quantitative longitudinal in vivo bioluminescence assays. In both C57BL/6 and BALB/c strains, neonatal injection of AAV1-Luc or AAV8-Luc by either IP, IV or IT routes produces lifelong gene expression. After IP injection at day 1–2 of life, gene expression increases 10–20 fold over the next several months. Highest levels of expression were achieved by IP injection, with lowest levels observed after IV injection. Injection of AAV1-Luc achieved higher levels of luciferase expression than did injection of AAV8- Luc. In contrast to the localized distribution of AAV1 mediated luciferase expression in the injected area, widespread, systemic expression of luciferase mediated by AAV8 after neonatal delivery is observed, regardless of the route of delivery. The effect of this altered tropism on gene expression levels and tolerance induction is being examined. In both C57BL/6 and BALB/c mice, IP injection of AAV1-Luc or AAV8-Luc at 1–2 days, 1 week, 2 weeks, or 3 weeks of age produced lifelong expression of luciferase and resulted in increasing levels of antibody responses against AAV1 or AAV8 with increasing age at primary injection. Antibody titers to AAV1 or AAV8 in animals injected at day 1–2 of life were comparable to background levels in uninjected animals. In C57BL/6 mice receiving a primary injection of AAV8-Luc, secondary injection of AAV8-Luc boosted the antibody response to AAV8 in the animals first injected at 1 week, 2 weeks or 3 weeks, but not in the animals injected at 1–2 day of life. We are currently exploring whether augmented expression with re-administration of AAV vectors in adult animals is due to an active process such as tolerance, partial tolerance or anergy. Developing strategies for the induction of tolerance to gene delivery vectors and therapeutic gene products will be an important advance for gene therapy for genetic and other disorders.

Neonatal Bone Marrow Transplantation Following Ex Vivo Gene Transfer and in Vivo Chemoselection for Hereditary Disorders

Transplantation of genetically engineered hematopoietic stem cells (HSC) holds promise for treatment of genetic and other disorders; however, several obstacles restrict the clinical application of this approach. These include leukemogenesis resulting from insertional mutagenesis, the lack of a competitive advantage for transplanted genetically engineered autologous HSCs, and the requirement for the construction and clinical approval of a gene transfer vehicle specific to each disorder. Allogenic transplantation approaches are further limited by a shortage of compatible donors, the toxicity of preparative regimens, immunosuppression, and graft versus host disease (GVHD).We have established a neonatal transplantation model to develop strategies to overcome these hurdles to HSC transplantation and gene therapy. We have uniquely combined allogenic transplantation during the neonatal period, when tolerance may be more readily achieved, with a positive selection strategy for in vivo amplification of drug-resistant donor HSC. This model system enables the evaluation of tolerance induction mechanisms to neo-antigens, and allogenic stem cell engraftment during immune ontogeny. In this model, bone marrow-derived HSC are transduced ex vivo by lentivirus-mediated gene transfer of P140K-O6-methylguanine-methyltransferase (MGMTP140K) and GFP (MAG vector). The MGMTP140K DNA repair enzyme confers resistance to benzylguanine, an inhibitor of endogenous MGMT, and to chloroethylating agents such as BCNU. This enables enrichment of donor cells at the stem cell level by in vivo chemoselection. Furthermore, in vivo administration of BG/BCNU may potentially deplete allo-reactive cells of donor and host origin, reducing the need for toxic ablative or immunosuppressive treatment.The in vivo selection strategy was first tested in neonates in the absence of MHC barriers. Syngenic whole bone marrow (WBM) was transduced with MAG and 5 × 105 cells were transplanted. Five weeks after transplantation and prior to chemoselection, 0.5% of cells expressed GFP in peripheral blood. Following two cycles of in vivo selection we observed 39.5% expression of GFP in peripheral blood. To assess the engraftment of fully MHC-mismatched HSC in neonates without in vivo selection, 107 B10.Br (H-2k) WBM was transplanted into C57BL/6 (H-2b) neonates using a non-myleoablative regimen. This resulted in 98% engraftment in adults without development of GVHD. To evaluate in vivo selection in an allogenic setting, 2 × 106 MAG transduced WBM from BALB/c (H-2d) adult mice were transplanted into C57BL/6 × BALB/c F1 (H-2b/d) neonatal mice. Despite having a graft versus host MHC disparity, we observed an increase from 4.7% (5 weeks after transplant) to 37.4% GFP expression following two cycles of in vivo chemoselection without evidence of GVHD.We have demonstrated successful engraftment and enrichment of both syngenic and allogenic HSC expressing MGMTP140K following neonatal transplantation and in vivo selection. Stable engraftment was achieved without myeloablation or post-transplant immunosuppression. No evidence of GVHD was observed after full MHC-mismatched transplantation. These results hold promise for the development of non-myeloablative allogenic transplant approaches using this in vivo selection strategy.

Immunological Basis and Management of Food Allergy

One of the most common allergies in children involves cow's milk, which contains approximately 20 different proteins that can cause allergic reactions. It is well known that children exhibiting signs of cow's milk allergy early in life often go on to develop allergy-related respiratory diseases; thus, management of early sensitisations and symptoms of food allergies is crucial to preventing subsequent allergic complications. Constant allergen exposure and other environmental factors determine whether a sensitised individual will become chronically allergic and experience persistent symptoms. Management of food allergies in children focuses on minimising sensitisation and encouraging immune system maturation through the exposure of children to exogenous stimuli known to prime the immune system. Hypoallergenic molecules or allergen avoidance can also be used to induce tolerance in allergy-prone children. Available evidence suggests that the onset of the sensitisation phase and the degree of inflammation can be modulated by external factors such as nutrition, and guidelines outlining the most effective dietary regimen for the prevention of allergic disease have been published. The underlying mechanisms of tolerance induction and the potential benefits of prophylactic treatment for food allergies remain to be determined.

Autoimmunity in Common Variable Immunodeficiency

Autoimmunity has been increasingly recognized as a major issue in patients with common variable immunodeficiency (CVID), the most common symptomatic primary immunodeficiency in adulthood. Different authors report high prevalences of autoimmune diseases in CVID, and several mechanisms have been proposed to explain this apparent paradox. Genetic predisposition, under current surveillance, innate and adaptive immunity deficiencies leading to persistent/recurrent infections, variable degrees of immune dysregulation, and possible failure in central and peripheral mechanisms of tolerance induction or maintenance may all contribute to increased autoimmunity. Data on the clinical/immunological profile of affected patients and treatment are available mostly concerning autoimmune cytopenias, the most common autoimmune diseases in CVID. Treatment is based on conventional alternatives, in association with short experience with new agents, including rituximab and infliximab. Benefits of early immunoglobulin substitutive treatment and hypothetical premature predictors of autoimmunity are discussed as potential improvements to CVID patients' follow-up.

Early fetal gene delivery utilizes both central and peripheral mechanisms of tolerance induction

We previously reported the induction of stable immune tolerance following direct injection of retroviral vectors into preimmune fetal sheep. In the present studies, we conduct detailed analysis of the thymus of recipients of in utero gene transfer (IUGT) to delineate the mechanism of the observed immune tolerance and assess the impact of recipient age on this process. Fetal sheep at varying gestational ages received the MSCV-NeoR-RFP retroviral vector. The thymus was then collected from these animals at 27 to 30 days postinjection and analyzed for evidence of transduction of key immunoregulatory thymic cells. Our results reveal that both thymic epithelial cells (TEC), crucial for presentation of self-antigen during T-cell thymic selection, and the cells comprising the Hassall's corpuscles, which can present antigen directly and also instruct dendritic cells to induce the formation of CD4(+)CD25(+) T-regulatory cells in the thymus, were only efficiently transduced if IUGT was performed early in gestation. Our findings thus demonstrate, for the first time, that early IUGT can potentially take advantage of multiple tolerogenic avenues in the fetus, transducing both TEC, which promote central tolerance, and Hassall's corpuscles, which induce formation of T regulatory cells that could act to maintain peripheral tolerance to the transgene products.

Generation and characterization of transgenic mice expressing a T‐cell receptor specific for the tumour‐associated antigen MDM2

T-cell-based antigen-specific immunotherapy targeting tumour-associated antigens offers the potential for cancer immunotherapy. However, the majority of identified tumour-associated antigens are also expressed at low levels in normal tissues and mechanisms of tolerance induction are likely to affect the quality of T-cell responses to such antigens. In this study a T-cell receptor transgenic model was developed to determine the magnitude of T-cell tolerance to the tumour-associated antigen murine double minute-2 (MDM2), a widely expressed protein that is found at elevated levels in many tumours. The analysis of transgenic mice showed that thymic deletion was responsible for purging large numbers of MDM2-specific T cells from the repertoire. However, some T cells with specificity for MDM2 were able to escape thymic deletion and persisted in the peripheral T-cell pool. Functional analysis revealed that these T cells displayed defects in antigen-driven expansion. This functional impairment of the MDM2-specific T cells was maintained following adoptive transfer of the T cells into hosts that were unable to present the T-cell-receptor-recognized antigen. This study demonstrates that thymic deletion and the functional impairment of T cells present in the periphery both operate to establish T-cell tolerance to the tumour-associated antigen MDM2. Furthermore, the tolerant phenotype was stable and did not require continuous MDM2 peptide presentation in normal tissues.

Tolerance induction to self-MHC antigens in fetal and neonatal mouse B cells

We have studied the mechanisms of tolerance induction to self-MHC antigens in mouse B cells during fetal development and the post-natal period. To monitor the fate of autoreactive B cell clones, we used the 3-83 micro delta B cell receptor (BCR)-transgenic (Tg) and -knock-in (KI) mouse models. These BCR-Tg and -KI B cells recognize the MHC class I molecules H-2K(k) and H-2K(b), with a high or moderate affinity, respectively. We compared the fate of BCR-Tg and -KI B cells in H-2K(b)-bearing animals and H-2K(b)-negative controls at various stages of their fetal development and post-natal life. Our data show that, in contrast to what occurs in adult B cells, anergy is the main component of tolerance induction in 3-83 micro delta BCR-Tg K(b+) autoreactive fetuses, while 3-83 BCR-KI fetuses primarily use receptor editing. Interestingly, autoreactive B cell deletion is absent or merely marginal before birth. Our results indicate that tolerance induction is effective as early as embryonic day 16.5 and that in the fetus and neonate, like in the adult, the main mechanism of B cell tolerance functioning in the 3-83 KI system is receptor editing. In contrast, in the 3-83 micro delta mice where receptor editing is hindered, adult and fetal B cells differ in their preferential use of mechanisms leading to self-tolerance (i.e. deletion versus anergy).

Dendritic Cells Break Bonds to Tolerize

Typically, dendritic cells (DCs) induce peripheral tolerance under steady state but immunity during inflammation or infection. In this issue of Immunity, Jiang et al. (2007) identify disruption of E-cadherin interactions as a unique maturation pathway by which DCs are capable of mediating tolerance.

New mechanism of tolerance induction in cancer

New mechanism of tolerance induction in cancer

New mechanism of tolerance induction in cancer

New mechanism of tolerance induction in cancer

Dendritic cell-based active specific immunotherapy for malignant glioma

Immunotherapy is an appealing therapeutic modality for malignant gliomas because of its potential to selectively target residual tumor cells that have invaded the normal brain. Most immunotherapeutic studies are designed to exploit the capacity of dendritic cells for inducing cell-mediated effects as well as immune memory responses for destroying residual tumor cells and preventing recurrence. Although initial clinical studies on dendritic cell-based immunotherapy resulted in very limited success, they have prompted many new studies on exploring strategies to induce a more robust antitumor immune response by using novel adjuvants for maturation and activation of dendritic cells. More studies have focused on the mechanisms of immune suppression by tumor cells and the role of regulatory T cells in tumor growth and progression. In this article, the authors review the evolution of dendritic cell-based immunotherapeutic strategies for adjuvant treatment of malignant gliomas. The authors also discuss how new knowledge on tumor-intrinsic mechanisms of tolerance induction and immunosuppression are likely to shape the future of immunotherapy for high-grade gliomas.

Induction of Cerebral Ischemic Tolerance by Erythromycin Preconditioning Reprograms the Transcriptional Response to Ischemia and Suppresses Inflammation

A single dose of the macrolide antibiotic erythromycin can induce tolerance against cerebral ischemia in vivo (pharmacologic preconditioning). This study identified potential mechanisms of tolerance induction by assessing effects of erythromycin preconditioning on the cerebral transcriptional response to transient global cerebral ischemia. Preconditioned and nonpreconditioned rats were exposed to 15 min of global cerebral ischemia, and changes in cerebral gene expression were identified by complementary DNA expression array and quantified by real-time reverse-transcription polymerase chain reaction. Ischemia caused a widespread up-regulation of transcription in nonpreconditioned brains in this model. Tolerance induction by erythromycin preconditioning reversed this pattern and caused a net down-regulation of a majority of genes, effectively reprogramming the brain's response pattern to ischemia. The most striking change in transcriptional response found in preconditioned animals was an almost complete suppression of the otherwise profound induction of proinflammatory genes by global ischemia. In contrast, the same treatment had little effect on the expression of apoptosis-inducing genes after ischemia. These findings present a new molecular correlate for the induction of ischemic tolerance achieved by erythromycin preconditioning and will further the understanding of this clinically important new regimen of preemptive neuroprotection.

IL-2/Neuroantigen Fusion Proteins as Antigen-Specific Tolerogens in Experimental Autoimmune Encephalomyelitis (EAE): Correlation of T Cell-Mediated Antigen Presentation and Tolerance Induction

The purpose of this study was to assess whether the Ag-targeting activity of cytokine/neuroantigen (NAg) fusion proteins may be associated with mechanisms of tolerance induction. To assess this question, we expressed fusion proteins comprised of a N-terminal cytokine domain and a C-terminal NAg domain. The cytokine domain comprised either rat IL-2 or IL-4, and the NAg domain comprised the dominant encephalitogenic determinant of the guinea pig myelin basic protein. Subcutaneous administration of IL2NAg (IL-2/NAg fusion protein) into Lewis rats either before or after an encephalitogenic challenge resulted in an attenuated course of experimental autoimmune encephalomyelitis. In contrast, parallel treatment of rats with IL4NAg (IL-4/NAg fusion protein) or NAg lacked tolerogenic activity. In the presence of IL-2R(+) MHC class II(+) T cells, IL2NAg fusion proteins were at least 1,000 times more potent as an Ag than NAg alone. The tolerogenic activity of IL2NAg in vivo and the enhanced potency in vitro were both dependent upon covalent linkage of IL-2 and NAg. IL4NAg also exhibited enhanced antigenic potency. IL4NAg was approximately 100-fold more active than NAg alone in the presence of splenic APC. The enhanced potency of IL4NAg also required covalent linkage of cytokine and NAg and was blocked by soluble IL-4 or by a mAb specific for IL-4. Other control cytokine/NAg fusion proteins did not exhibit a similar enhancement of Ag potency compared with NAg alone. Thus, the IL2NAg and IL4NAg fusion proteins targeted NAg for enhanced presentation by particular subsets of APC. The activities of IL2NAg revealed a potential relationship between NAg targeting to activated T cells, T cell-mediated Ag presentation, and tolerance induction.

Cellular and Molecular Mechanisms of Tumor-Induced T-Cell Tolerance

The spontaneous interaction between tumor cells and the immune system has been shown to result in reciprocal changes leading to a less immunogenic tumor and immune cells less capable or unable to mount an effective response against a growing malignancy. Although several mechanisms have been proposed to account for the ability of tumor cells to render immune cells less efficient, one that has gained particular attention relates to the recognition of tumor antigens by T-cells, a process that unfortunately leads to the induction and establishment of antigen-specific T-cell tolerance rather than T-cell priming. Here, we present the experimental and clinical evidence that help identify this remarkable barrier that the immune system itself and more specifically its mechanisms of tolerance induction has imposed to our efforts to effectively harness the immune system against tumors. In particular, we will discuss the central role of bone marrow-derived antigen-presenting cells (APCs) in the induction of this state of T-cell unresponsiveness and the potential role of the tumor microenvironment in determining the tolerogenic properties of these APCs. Finally, we provide information on receptor-ligands and intracellular signaling pathways that given their role in influencing the inflammatory properties of APCs are being exploited as targets to revert mechanisms of T-cell unresponsiveness in cancer.