Derrick P. McCarthy

Microparticles bearing encephalitogenic peptides induce T-cell tolerance and ameliorate experimental autoimmune encephalomyelitis

Aberrant T-cell activation underlies many autoimmune disorders, yet most attempts to induce T-cell tolerance have failed. Building on previous strategies for tolerance induction that exploited natural mechanisms for clearing apoptotic debris, we show that antigen-decorated microparticles (500-nm diameter) induce long-term T-cell tolerance in mice with relapsing experimental autoimmune encephalomyelitis. Specifically, intravenous infusion of either polystyrene or biodegradable poly(lactide-co-glycolide) microparticles bearing encephalitogenic peptides prevents the onset and modifies the course of the disease. These beneficial effects require microparticle uptake by marginal zone macrophages expressing the scavenger receptor MARCO and are mediated in part by the activity of regulatory T cells, abortive T-cell activation and T-cell anergy. Together these data highlight the potential for using microparticles to target natural apoptotic clearance pathways to inactivate pathogenic T cells and halt the disease process in autoimmunity.

Therapeutic Inflammatory Monocyte Modulation Using Immune-Modifying Microparticles

Negatively charged immune-modifying microparticles bind to the scavenger receptor MARCO on inflammatory monocytes, resulting in their apoptosis and reduced inflammatory damage in a range of diseases. A New Frontier in Immune Modulation Inflammatory monocytes markedly potentiate the immune pathology observed in many diseases, yet no therapy exists that specifically inhibits these cells. The therapeutic accessibility of monocytes in the bloodstream and their inherent propensity to engulf particulate material suggest that highly negatively charged microparticles might provide a readily translatable solution to this problem. These microparticles, referred to as immune-modifying microparticles (IMPs), may be derived from numerous compounds, including the biodegradable polymer poly(lactic-co-glycolic acid) (PLGA-IMP), already used in humans for inter alia dissolvable sutures. Getts et al. now show that upon infusion, IMPs bind to a receptor with a positive domain on inflammatory monocytes, resulting in monocyte sequestration in the spleen and apoptosis through a similar pathway observed for senescing leukocytes. This safe monocyte clearance pathway culminated in substantially reduced inflammatory tissue damage in mouse models of West Nile virus encephalitis, experimental autoimmune encephalomyelitis, peritonitis, colitis, and myocardial infarction. Together, the data suggest that IMPs could transform the treatment of acute inflammation. Indeed, phase 1/2 testing is planned to begin in 2014, with rapid translation supported by the availability of clinical-grade PLGA. Inflammatory monocyte-derived effector cells play an important role in the pathogenesis of numerous inflammatory diseases. However, no treatment option exists that is capable of modulating these cells specifically. We show that infused negatively charged, immune-modifying microparticles (IMPs), derived from polystyrene, microdiamonds, or biodegradable poly(lactic-co-glycolic) acid, were taken up by inflammatory monocytes, in an opsonin-independent fashion, via the macrophage receptor with collagenous structure (MARCO). Subsequently, these monocytes no longer trafficked to sites of inflammation; rather, IMP infusion caused their sequestration in the spleen through apoptotic cell clearance mechanisms and, ultimately, caspase-3–mediated apoptosis. Administration of IMPs in mouse models of myocardial infarction, experimental autoimmune encephalomyelitis, dextran sodium sulfate–induced colitis, thioglycollate-induced peritonitis, and lethal flavivirus encephalitis markedly reduced monocyte accumulation at inflammatory foci, reduced disease symptoms, and promoted tissue repair. Together, these data highlight the intricate interplay between scavenger receptors, the spleen, and inflammatory monocyte function and support the translation of IMPs for therapeutic use in diseases caused or potentiated by inflammatory monocytes.

Mouse models of multiple sclerosis: experimental autoimmune encephalomyelitis and Theiler's virus-induced demyelinating disease.

Experimental autoimmune encephalomyelitis (EAE) and Theilers Murine Encephalitis Virus-Induced Demyelinating Disease (TMEV-IDD) are two clinically relevant murine models of multiple sclerosis (MS). Like MS, both are characterized by mononuclear cell infiltration into the CNS and demyelination. EAE is induced by either the administration of myelin protein or peptide in adjuvant or by the adoptive transfer of encephalitogenic T cell blasts into naïve recipients. The relative merits of each of these protocols are compared. Depending on the type of question being asked, different mouse strains and peptides are used. Different disease courses are observed with different strains and different peptides in active EAE. These variations are also addressed. Additionally, issues relevant to clinical grading of EAE in mice are discussed. In addition to EAE induction, useful references for other disease indicators such as DTH, in vitro proliferation, and immunohistochemistry are provided. TMEV-IDD is a useful model for understanding the possible viral etiology of MS. This section provides detailed information on the preparation of viral stocks and subsequent intracerebral infection of mice. Additionally, virus plaque assay and clinical disease assessment are discussed. Recently, recombinant TMEV strains have been created for the study of molecular mimicry which incorporate various 30 amino acid myelin epitopes within the leader region of TMEV.

ECDI-fixed allogeneic splenocytes induce donor-specific tolerance for long-term survival of islet transplants via two distinct mechanisms

A major challenge for human allogeneic islet transplantation is the development of effective methods to induce donor-specific tolerance to obviate the need for life-long immunosuppression that is toxic to the insulin-producing β cells and detrimental to the host. We developed an efficient donor-specific tolerance therapy that utilizes infusions of ethylene carbodiimide (ECDI)-treated donor splenic antigen-presenting cells that results in indefinite survival of allogeneic islet grafts in the absence of immunosuppression. Furthermore, we show that induction of tolerance is critically dependent on synergistic effects between an intact programmed death 1 receptor–programmed death ligand 1 signaling pathway and CD4+CD25+Foxp3+ regulatory T cells. This highly efficient antigen-specific therapy with a complete avoidance of immunosuppression has significant therapeutic potential in human islet cell transplantation.

A Biodegradable Nanoparticle Platform for the Induction of Antigen-Specific Immune Tolerance for Treatment of Autoimmune Disease

Targeted immune tolerance is a coveted therapy for the treatment of a variety of autoimmune diseases, as current treatment options often involve nonspecific immunosuppression. Intravenous (iv) infusion of apoptotic syngeneic splenocytes linked with peptide or protein autoantigens using ethylene carbodiimide (ECDI) has been demonstrated to be an effective method for inducing peripheral, antigen-specific tolerance for treatment of autoimmune disease. Here, we show the ability of biodegradable poly(lactic-co-glycolic acid) (PLG) nanoparticles to function as a safe, cost-effective, and highly efficient alternative to cellular carriers for the induction of antigen-specific T cell tolerance. We describe the formulation of tolerogenic PLG particles and demonstrate that administration of myelin antigen-coupled particles both prevented and treated relapsing-remitting experimental autoimmune encephalomyelitis (R-EAE), a CD4 T cell-mediated mouse model of multiple sclerosis (MS). PLG particles made on-site with surfactant modifications surpass the efficacy of commercially available particles in their ability to couple peptide and to prevent disease induction. Most importantly, myelin antigen-coupled PLG nanoparticles are able to significantly ameliorate ongoing disease and subsequent relapses when administered at onset or at peak of acute disease, and minimize epitope spreading when administered during disease remission. Therapeutic treatment results in significantly reduced CNS infiltration of encephalitogenic Th1 (IFN-γ) and Th17 (IL-17a) cells as well as inflammatory monocytes/macrophages. Together, these data describe a platform for antigen display that is safe, low-cost, and highly effective at inducing antigen-specific T cell tolerance. The development of such a platform carries broad implications for the treatment of a variety of immune-mediated diseases.

Tolerance Induced by Apoptotic Antigen-Coupled Leukocytes Is Induced by PD-L1+ and IL-10–Producing Splenic Macrophages and Maintained by T Regulatory Cells

Ag-specific tolerance is a highly desired therapy for immune-mediated diseases. Intravenous infusion of protein/peptide Ags linked to syngeneic splenic leukocytes with ethylene carbodiimide (Ag-coupled splenocytes [Ag-SP]) has been demonstrated to be a highly efficient method for inducing peripheral, Ag-specific T cell tolerance for treatment of autoimmune disease. However, little is understood about the mechanisms underlying this therapy. In this study, we show that apoptotic Ag-SP accumulate in the splenic marginal zone, where their uptake by F4/80+ macrophages induces production of IL-10, which upregulates the expression of the immunomodulatory costimulatory molecule PD-L1 that is essential for Ag-SP tolerance induction. Ag-SP infusion also induces T regulatory cells that are dispensable for tolerance induction but required for long-term tolerance maintenance. Collectively, these results indicate that Ag-SP tolerance recapitulates how tolerance is normally maintained in the hematopoietic compartment and highlight the interplay between the innate and adaptive immune systems in the induction of Ag-SP tolerance. To our knowledge, we show for the first time that tolerance results from the synergistic effects of two distinct mechanisms, PD-L1–dependent T cell-intrinsic unresponsiveness and the activation of T regulatory cells. These findings are particularly relevant as this tolerance protocol is currently being tested in a Phase I/IIa clinical trial in new-onset relapsing-remitting multiple sclerosis.

Have we overestimated the benefit of human(ized) antibodies

The infusion of animal-derived antibodies has been known for some time to trigger the generation of antibodies directed at the foreign protein as well as adverse events including cytokine release syndrome. These immunological phenomena drove the development of humanized and fully human monoclonal antibodies. The ability to generate human(ized) antibodies has been both a blessing and a curse. While incremental gains in the clinical efficacy and safety for some agents have been realized, a positive effect has not been observed for all human(ized) antibodies. Many human(ized) antibodies trigger the development of anti-drug antibody responses and infusion reactions. The current belief that antibodies need to be human(ized) to have enhanced therapeutic utility may slow the development of novel animal-derived monoclonal antibody therapeutics for use in clinical indications. In the case of murine antibodies, greater than 20% induce tolerable/negligible immunogenicity, suggesting that in these cases humanization may not offer significant gains in therapeutic utility. Furthermore, humanization of some murine antibodies may reduce their clinical effectiveness. The available data suggest that the utility of human(ized) antibodies needs to be evaluated on a case-by-case basis, taking a cost-benefit approach, taking both biochemical characteristics and the targeted therapeutic indication into account.

Oligodendrocyte death results in immune-mediated CNS demyelination

Although multiple sclerosis is a common neurological disorder, the origin of the autoimmune response against myelin, which is the characteristic feature of the disease, remains unclear. To investigate whether oligodendrocyte death could cause this autoimmune response, we examined the oligodendrocyte ablation Plp1-CreERT;ROSA26-eGFP-DTA (DTA) mouse model. Approximately 30 weeks after recovering from oligodendrocyte loss and demyelination, DTA mice develop a fatal secondary disease characterized by extensive myelin and axonal loss. Strikingly, late-onset disease was associated with increased numbers of T lymphocytes in the CNS and myelin oligodendrocyte glycoprotein (MOG)-specific T cells in lymphoid organs. Transfer of T cells derived from DTA mice to naive recipients resulted in neurological defects that correlated with CNS white matter inflammation. Furthermore, immune tolerization against MOG ameliorated symptoms. Overall, these data indicate that oligodendrocyte death is sufficient to trigger an adaptive autoimmune response against myelin, suggesting that a similar process can occur in the pathogenesis of multiple sclerosis.

Exploiting Apoptosis for Therapeutic Tolerance Induction

Immune tolerance remains the most promising yet elusive strategy for treating immune-mediated diseases. An experimental strategy showing promise in phase 1 clinical studies is the delivery of Ag cross-linked to apoptotic leukocytes using ethylene carbodiimide. This approach originated from demonstration of the profound tolerance-inducing ability of i.v. administered Ag-coupled splenocytes (Ag-SP) in mice, which has been demonstrated to treat T cell–mediated disorders including autoimmunity, allergy, and transplant rejection. Recent studies have defined the intricate interplay between the innate and adaptive immune systems in Ag-SP tolerance induction. Innate mechanisms include scavenger receptor–mediated uptake of Ag-SP by host APCs, Ag representation, and the required upregulation of PD-L1 expression and IL-10 production by splenic marginal zone macrophages leading to Ag-specific T cell regulation via the combined effects of cell-intrinsic anergy and regulatory T cell induction. In this paper, we discuss the history, advantages, current mechanistic understanding, and clinical potential of tolerance induction using apoptotic Ag-coupled apoptotic leukocytes.

An antigen-encapsulating nanoparticle platform for TH1/17 immune tolerance therapy

Tolerogenic nanoparticles (NPs) are rapidly being developed as specific immunotherapies to treat autoimmune disease. However, many NP-based therapies conjugate antigen (Ag) directly to the NP posing safety concerns due to antibody binding or require the co-delivery of immunosuppressants to induce tolerance. Here, we developed Ag encapsulated NPs comprised of poly(lactide-co-glycolide) [PLG(Ag)] and investigated the mechanism of action for Ag-specific tolerance induction in an autoimmune model of T helper type 1/17 dysfunction - relapse-remitting experimental autoimmune encephalomyelitis (R-EAE). PLG(Ag) completely abrogated disease induction in an organ specific manner, where the spleen was dispensable for tolerance induction. PLG(Ag) delivered intravenously distributed to the liver, associated with macrophages, and recruited Ag-specific T cells. Furthermore, programmed death ligand 1 (PD-L1) was increased on Ag presenting cells and PD-1 blockade lessened tolerance induction. The robust promotion of tolerance by PLG(Ag) without co-delivery of immunosuppressive drugs, suggests that these NPs effectively deliver antigen to endogenous tolerogenic pathways.