Sites Of Antigen Encounter Research Articles

Microneedle-Mediated Delivery of Immunomodulators Restores Immune Privilege in Hair Follicles and Reverses Immune-Mediated Alopecia.

Disorders in the regulatory arm of the adaptive immune system result in autoimmune-mediated diseases. While systemic immunosuppression is the prevailing approach to manage them, it fails to achieve long-lasting remission due to concomitant suppression of the regulatory arm and carries the risk of heightened susceptibility to infections and malignancies. Alopecia areata is a condition characterized by localized hair loss due to autoimmunity. The accessibility of the skin allowslocal rather than systemic intervention to avoid broad immunosuppression. It is hypothesized that the expansion of endogenous regulatory T cells (Tregs) at the site of antigen encounter can restore the immune balance and generate a long-lasting tolerogenic response. A hydrogel microneedle (MN) patch is therefore utilized for delivery of CCL22, a Treg-chemoattractant, and IL-2, a Tregsurvival factor to amplify them. In an immune-mediated murine model of alopecia, local bolstering of Tregnumbers is shown, leading to sustained hair regrowth and attenuation of inflammatory pathways. In a humanized skin transplant mouse model, expansion of Tregs within human skin is confirmed without engendering peripheral immunosuppression. The patch offers high-loading capacity and shelf-life stability for prospective clinical translation. By harmonizing immune responses locally, the aim is to reshape the landscape of autoimmune skin disease management.

Regulatory T cells engineered with TCR signaling-responsive IL-2 nanogels suppress alloimmunity in sites of antigen encounter.

Adoptive cell transfer of ex vivo expanded regulatory T cells (Tregs) has shown immense potential in animal models of auto- and alloimmunity. However, the effective translation of such Treg therapies to the clinic has been slow. Because Treg homeostasis is known to require continuous T cell receptor (TCR) ligation and exogenous interleukin-2 (IL-2), some investigators have explored the use of low-dose IL-2 injections to increase endogenous Treg responses. Systemic IL-2 immunotherapy, however, can also lead to the activation of cytotoxic T lymphocytes and natural killer cells, causing adverse therapeutic outcomes. Here, we describe a drug delivery platform, which can be engineered to autostimulate Tregs with IL-2 in response to TCR-dependent activation, and thus activate these cells in sites of antigen encounter. To this end, protein nanogels (NGs) were synthesized with cleavable bis(N-hydroxysuccinimide) cross-linkers and IL-2/Fc fusion (IL-2) proteins to form particles that release IL-2 under reducing conditions, as found at the surface of T cells receiving stimulation through the TCR. Tregs surface-conjugated with IL-2 NGs were found to have preferential, allograft-protective effects relative to unmodified Tregs or Tregs stimulated with systemic IL-2. We demonstrate that murine and human NG-modified Tregs carrying an IL-2 cargo perform better than conventional Tregs in suppressing alloimmunity in murine and humanized mouse allotransplantation models. In all, the technology presented in this study has the potential to improve Treg transfer therapy by enabling the regulated spatiotemporal provision of IL-2 to antigen-primed Tregs.

The Lymphoma Tumor Microenvironment Influences Toxicity after CD19 CAR T Cell Therapy

Introduction: Of patients receiving CD19 CAR T cell therapy for large B cell lymphoma (LBCL), approximately 1 in 10 experience severe cytokine release syndrome (CRS) and 1 in 3 experience severe neurotoxicity. While CAR T cells trigger the onset of these toxicities, CRS and neurotoxicity are thought to occur as a consequence of activated myeloid cells amplifying cytokine and catecholamine release, thereby stimulating inflammation both systemically and at the blood-brain barrier. However, patient and tumor-related factors that account for differences in the amount of toxicity remain poorly understood. Methods: Serum cytokine levels were measured on an ELLA point of care device prior to lymphodepleting chemotherapy and throughout inpatient treatment with CD19 CAR T cell therapy (axicabtagene ciloleucel) for LBCL. Catecholamine levels were measured as we have previously reported. Tumor biopsies were taken within 1 month prior to infusion of CAR T cells. RNA expression was measured by RNAseq and/or a Nanostring IO360 panel consisting of 770 genes found in the tumor microenvironment (TME) in cancer. Analysis used nSolver to identify cell types, GSEA and differential gene expression between groups. Mouse CAR T cell studies utilized mouse CD19-targeted CAR T cells derived from C57BL/6 splenocytes and cultured in vitro with myeloid cells and target cells to evaluate cytotoxicity and/or cytokine secretion. Elicited mouse macrophages were collected from peritoneal fluid 4 days after IP injection of 3% Brewer's thioglycollate medium. In vivo studies with mouse CD19-targeted CAR T cells were performed in IL2Ra-/- mice given cyclophosphamide as a pre-conditioning chemotherapy followed by adoptive transfer and analyses for CAR T cell and B cell persistence, as well as cytokines. Results: Of 58 patients undergoing CD19 CAR T cell therapy for LBCL, 8 (14%) had severe (grade 3 or higher) CRS and 16 (28%) had severe (grade 3 or higher) neurotoxicity. At baseline, peripheral blood levels of IL-6, IFN-γ, IL-15 and ferritin were significantly higher in patients who would subsequently experience severe CRS and severe neurotoxicity. Confirming our recent animal model of CRS we determined that peak serum catecholamine levels were higher in patients experiencing severe CRS. To identify if myeloid cells potentiate cytokine release we co-cultured CAR T cells with CD19 target and macrophages obtained from elicited mouse peritoneum. When these macrophages were added, IL-6 release from CAR T cells significantly increased compared to when macrophages were absent. Next, we studied the baseline TME in LBCL CAR T patients. Of 36 patients, 10 (27%) experienced severe neurotoxicity following CAR T cell therapy. By cell type score, the severe neurotoxicity group had a lower expression of genes associated with T cells overall and specifically Tregs. Also significantly lower in the severe neurotoxicity group were T cell genes including multiple subunits of CD3, CD3ζ, FOXP3, ICOS, CD62L and others. Association of increased T cell infiltration in the TME with low neurotoxicity raised the possibility that suppressive T cell subsets play a role in limiting toxicity post-CAR T cell therapy. To test this hypothesis, we injected CD19-targeted CAR T cells into an immune competent mouse model of Treg depletion (IL2Ra-/-) with established CD19+ leukemia. Treg deficient mice experienced a massive cytokine release after CAR T infusion and died prematurely due to CAR T toxicity compared to control mice with Tregs intact. Conclusions: Our observations suggest that the incidence of severe toxicity following CD19 CAR T cell therapy is influenced by baseline characteristics that are present prior to the infusion of CAR T cells. These include systemic inflammation characterized by high cytokine levels and a TME notable for a lack of infiltrating T cells. We posit a model whereby inflammation primes myeloid cells that are further activated upon CAR T cell infusion to release toxic amounts of cytokines and catecholamines. T cell subsets in the TME may modulate CAR T cells at the site of antigen encounter and prevent excessive CAR T activation. Reducing systemic inflammation or encouraging T cell infiltration into tumor prior to CAR T infusion are potential strategies for lowering the toxicity associated with CAR T therapy. Disclosures Jain: Kite/Gilead: Consultancy. Chavez:Kite Pharmaceuticals, Inc.: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Genentech: Speakers Bureau; Janssen Pharmaceuticals, Inc.: Speakers Bureau. Shah:Novartis: Honoraria; Spectrum/Astrotech: Honoraria; Celgene/Juno: Honoraria; Kite/Gilead: Honoraria; Incyte: Research Funding; Jazz Pharmaceuticals: Research Funding; Pharmacyclics: Honoraria; Adaptive Biotechnologies: Honoraria; AstraZeneca: Honoraria. Bachmeier:Kite/Gilead: Speakers Bureau. Mullinax:Iovance: Research Funding. Locke:Novartis: Other: Scientific Advisor; Cellular BioMedicine Group Inc.: Consultancy; Kite: Other: Scientific Advisor. Davila:Anixa: Consultancy; Precision Biosciences: Consultancy; Novartis: Research Funding; GlaxoSmithKline: Consultancy; Adaptive: Consultancy; Celgene: Research Funding; Atara: Research Funding; Bellicum: Consultancy.

Oral Exposure to Trypanosoma cruzi Elicits a Systemic CD8+T Cell Response and Protection against Heterotopic Challenge

Trypanosoma cruzi infects millions of people in Latin America and often leads to the development of Chagas disease. T. cruzi infection can be acquired at or near the bite site of the triatomine vector, but per os infection is also a well-documented mode of transmission, as evidenced by recent microepidemics of acute Chagas disease attributed to the consumption of parasite-contaminated foods and liquids. It would also be convenient to deliver vaccines for T. cruzi by the oral route, particularly live parasite vaccines intended for the immunization of reservoir hosts. For these reasons, we were interested in better understanding immunity to T. cruzi following oral infection or oral vaccination, knowing that the route of infection and site of antigen encounter can have substantial effects on the ensuing immune response. Here, we show that the route of infection does not alter the ability of T. cruzi to establish infection in muscle tissue nor does it impair the generation of a robust CD8(+) T cell response. Importantly, oral vaccination with attenuated parasites provides protection against wild-type (WT) T. cruzi challenge. These results strongly support the development of whole-organism-based vaccines targeting reservoir species as a means to alleviate the burden of Chagas disease in affected regions.

Die Rolle der Gewebe-Mikroumgebung bei der Kontaktallergie

Allergic contact dermatitis (ACD) is a T cell-mediated inflammatory skin disease. In order to activate the T cell response, the innate immune system has to generate an inflammation that activates skin cells and triggers allergen-loaded dendritic cells (DC) to migrate to the local lymph nodes. There, DC confer the information about their tissue of origin and the site of antigen encounter to T cells which are induced to upregulate skin-specific homing receptors. This enables the recruitment of allergen-specific effector T cells to the skin following allergen contact. Similar findings were made for the small intestine. Our recent studies in the mouse model show a crucial role for the tissue microenvironment both for the inflammatory response and for the transfer of the topographical information from the DC to the T cells. Thus, we have found that Toll-like receptors and the IL-12/IL-12R system are important for DC activation by contact allergens and imply a role for endogenous TLR ligands generated during ACD. Moreover, cells of the peripheral tissue microenvironment provide information via soluble factors and cell-cell contact which license DC emigrating from skin but also DC emigrating from the small intestine to confer topographical information to T cells by imprinting tissue-specific homing receptors on the T cells in the draining lymph nodes. Our results illustrate the crucial role of the tissue context and of the communication between tissue-specific cell types for the shaping of immune responses. These findings underline the necessity to take into account the unique tissue microenvironments upon analysis of immune responses.

Lymphocytes

Antigen-nonspecific inflammation appears to contribute to postischemic brain injury. Because there is a breach in the integrity of the blood-brain barrier after stroke, the immune system encounters novel central nervous system (CNS) antigens that allow for the development of a CNS antigen-specific autoimmune response. The nature of the immune response generated on antigen encounter is determined by the microenvironment at the site of antigen encounter. For instance, a systemic inflammatory response, such as that which would accompany an infection, could alter the microenvironment in such a way as to promote the initiation of deleterious autoimmunity. If patients who develop an infection in the immediate poststroke period are predisposed toward a CNS autoimmune response, it might help to explain why infection after stroke is associated with increased disability. We present data to support this hypothesis and to show that the breach in the blood-brain barrier can also be capitalized on to modulate the immune response to create a neuroprotective environment after stroke.

New strategies in immune tolerance induction.

Induction of tolerance in clinical organ transplantation that will obviate the use of chronic immunosuppression and preserve host immune response to other antigens remains the goal of transplant research. The thymus plays a critical role in the ability of the immune system to discriminate between self- and nonself-antigens or harmful and harmless alloantigens. We now know that multiple factors determine how the immune system responds to a self-antigen or foreign antigen. These determinants include developmental stage of the host, stage of T-cell maturity, site of antigen encounter, type and maturity of antigen-presenting cells, and presence and type of costimulatory molecules. Our understanding of the mechanisms of T-cell interactions with peptide/ major histocompatibility complex in peripheral lymphoid organs has led to experiments that translate into peripheral T-cell tolerance. The induction of high-avidity peripheral alloreactive T cells in the early phase of organ transplantation makes it difficult to achieve long-term alloantigen-specific tolerance without the use of transient perioperative immunosuppression. Therefore, protocols that induce robust tolerance in rodent and nonhuman primate models involve the use of donor antigen combined with a short course of perioperative immunosuppression. These studies suggest that the underlying mechanisms of peripheral tolerance include deletion, anergy, immune deviation, and regulatory T cells. This review focuses on recent advances in tolerance induction in experimental animal models and discusses their relevance to the development of protocols for the induction and maintenance of clinical transplant tolerance.

Cutaneous lymphocyte antigen expression on human effector B cells depends on the site and on the nature of antigen encounter.

In contrast to T cells, information on skin-homing B cells expressing the cutaneous lymphocyte antigen (CLA) is sparse. CLA expression on human B cells was investigated among circulating immunoglobulin-secreting cells (ISC) and among antigen-specific antibody-secreting cells (ASC) elicited by parenteral, oral or rectal primary immunization, or by parenteral or oral secondary immunization with Salmonella typhi Ty21a. CLA expression was examined by combining cell sorting with an enzyme-linked immunospot assay. Among all ISC, the proportion of CLA(+) cells was 13-21%. Parenteral immunization induced antigen-specific ASC of which 13% were CLA(+), while oral and rectal immunizations were followed by only 1% of CLA(+) ASC (p<0.001). Oral re-immunization was followed by an up-regulation of CLA (34-48%) regardless of the route of priming. Parenteral re-immunization elicited ASC of which 9-14% were CLA(+). In conclusion, the expression of CLA on human effector B cells depends on the site of antigen encounter: intestinal stimulation elicits cells with no CLA, while parenteral encounter elicits significant numbers of CLA(+) cells. Even though primary antigen encounter in the intestine failed to stimulate CLA expression, up-regulation of CLA was found upon intestinal antigen re-encounter. These findings may be of relevance in the pathogenesis of some cutaneous disorders.

Negative Selection of Immature B Cells by Receptor Editing or Deletion Is Determined by Site of Antigen Encounter

Immature B cells that encounter self-antigen are eliminated from the immune repertoire by negative selection. Negative selection has been proposed to take place by two distinct mechanisms: deletion by apoptosis or alteration of the antigen receptor specificity by receptor editing. While convincing evidence exists for each, the two models are inherently contradictory. In this paper, we propose a resolution to this contradiction by demonstrating that the site of first antigen encounter dictates which mechanism of negative selection is utilized. We demonstrate that the bone marrow microenvironment provides signals that block antigen-induced deletion and promote RAG reinduction. In the periphery, the absence of these signals allows the immature B cell to default to apoptosis as a result of BCR engagement.