In vivo reprogramming of immune cells: Technologies for induction of antigen-specific tolerance

Abstract

Technologies that induce antigen-specific immune tolerance by mimicking naturally occurring mechanisms have the potential to revolutionize the treatment of many immune-mediated pathologies such as autoimmunity, allograft rejection, and allergy. The immune system intrinsically has central and peripheral tolerance pathways for eliminating or modulating antigen-specific responses, which are being exploited through emerging technologies. Antigen-specific tolerogenic responses have been achieved through the functional reprogramming of antigen-presenting cells or lymphocytes. Alternatively, immune privileged sites have been mimicked using biomaterial scaffolds to locally suppress immune responses and promote long-term allograft survival. This review describes natural mechanisms of peripheral tolerance induction and the various technologies being developed to achieve antigen-specific immune tolerance in vivo. As currently approved therapies are non-specific and carry significant associated risks, these therapies offer significant progress towards replacing systemic immune suppression with antigen-specific therapies to curb aberrant immune responses.

Keywords: Allergy; Autoimmune disease; Drug delivery; Immune tolerance; Nanoparticle; Regulatory T cells; Transplantation.

Figure 1

Highlighted approaches of technologies implemented for antigen-specific tolerance induction. Most antigen-specific tolerance strategies result in reprogramming lymphocytes through antigen presenting cells (APCs), however, there are platforms that target T cells and specifically recognize their autoreactive T cell receptors. Inspired by the natural clearance of apoptotic cells which results in peripheral tolerance maintenance, antigen has been delivered by various platforms including antigen-coupled splenocytes (Ag-SP), erythrocyte-targeted peptides (Ag-RBC), and antigen-loaded synthetic particles. These carriers are internalized, processed by APCs, and induce tolerogenic costimulation and soluble signaling pathways that direct T cell phenotypes away from immunogenic effector T cell activation and toward regulatory T cells (Tregs), anergy, or deletion. Direct interaction of particle-bound peptide-major histocompatibility complexes (pMHC-NPs) with antigen-experienced T cells can induce a tolerogenic regulatory-like T_R1 phenotype that can mitigate immune-mediated disease progression.

Figure 2

Mode of antigen-association with particles affects the risk of anaphylaxis following intravenous administration in individuals with prior antigen sensitization. Antigen is associated with particles by surface-coupling (NP-Ag) or by encapsulation (NP(Ag)) methods. In vivo, granulocyte activation occurs when NP-Ag or soluble antigen is recognized by circulating IgE antibodies or binds to pre-bound IgE on granulocytes. Cross-linking of IgE on granulocytes triggers degranulation and subsequent release of histamine and other inflammatory mediators that cause increased permeability, distension of blood capillaries, and anaphylaxis. (i) Binding of antigen-specific IgE to NP-Ag can trigger granulocyte activation. (ii) Binding of antigen-specific IgG to NP-Ag reduces potential granulocyte activation but can result in off-targeted biodistribution and reduce tolerance induction. (iii) Pre-mature antigen release from NP(Ag) can result in granulocyte activation but to a lesser extent than NP-Ag. (iv) NP(Ag) with negligible rate of antigen release reduces the risk of granulocyte activation and enables unaffected distribution to the liver and spleen to induce tolerogenic responses.

Figure 3

Erythrocyte-binding TER119 scFv antibodies fused with autoantigen (p31) specifically target red blood cells in situ after intravenous administration and induce antigen-specific tolerance in a type 1 diabetes model. (A) TER119-p31 induces tolerance and results in normoglycemia. Normoglycemic NOD/ShiLtJ mice received adoptive transfer of diabetogenic BDC2.5 CD4⁺ T cells and 3 intravenous treatments of saline, p31, or TER119-p31 (n =8, n = 9, and n = 9, respectively) over the first week. ***P < 0.0001. (B) Immunohistochemistry of pancreatic islets excised 4 days after treatment and stained for CD3ε T cells (green), insulin (red), and nuclei (blue). Saline-treated and untargeted autoantigen mimetope p31 resulted in T cell infiltration and islet destruction in contrast to mice treated with red blood cell-binding autoantigen (TER119-p31) which prevented T cell infiltration and preserved insulin production. (Scale bar = 100 μm). Reproduced from [48] with permission.

Figure 4

Antigen-polymer conjugate nanoparticles display favorable physicochemical and biological properties for tolerance induction. (A) Schematic representation of Ag-coupled, Ag-encapsulated, and polymer-conjugate nanoparticles. (B) Release profile of NP(OVA₃₂₃-339), NP-OVA_323-339, and acNP-OVA_323-339. (C) Regulatory T cell induction is dependent on nanoparticle concentration. BMDCs were treated for 3 hr with various concentrations of acNP-OVA_323-339 (2, 8, 25, 150 μg/mg loading). Excess acNP-OVA_323-339 particles were subsequently washed from the cell surface prior to addition of OT-II T cells and 2 ng/mL of TGF-β1. (D) Schematic representation of antigen-polymer conjugate nanoparticles delivering multiple Ags. (E) Clinical scores of SJL/J mice treated with 1.25 mg of acNP-OVA_323-339 (8 μg/mg OVA_323-339), acNP-PLP_139-151 (8 μg/mg PLP_139-151), acNP-PLP1_78-191 (8 μg/mg PLP_178-191), or acNP-PLP_{139-151,178-191} (8 μg/mg PLP1_39-151 and 8 μg/mg PLP_178-191) and immunized with PLP_139-151 and PLP_178-191 in CFA to induce R-EAE 7 days later. (F) Corresponding cumulative clinical score for mice treated with particles (n = 5). Differences between disease courses of different treatment groups were analyzed for statistical significance using the Kruskal-Wallis test (one-way ANOVA non-parametric test) with Dunn's multiple comparisons test (p < 0.05) [94].

Figure 5

A short course of low dose rapamycin synergizes with PLG-donorAg (dAg) to enhance tolerance efficacy. Recipient mice (C57BL/6) receiving donor (BALB/c) PLG-dAg injections at days 7 and +1 in combination with a 4-day course (days 1, 0. +1, and +2) of low dose (0.1 mg/kg) rapamycin demonstrated significantly greater islet allograft survival (n=11) compared with mice treated with rapamycin alone (n=9), BALB/c PLG-dAg alone (n=17), or PLG particles coupled with lysate proteins from a third party donor SJL/J (n=4). **p < 0.01. Reproduced from [71] with permission from Elsevier.

Figure 6

Prophylactic treatment with PLG(OVA) inhibits Th2-induced airway inflammation. Naive female BALB/c mice (n = 5) were treated i.v. with 2.5 mg PLG(OVA) or control PLG(LYS) on days −7 and +7 relative to i.p. immunization with 10 μg of OVA in 3 mg of alum or alum alone on days 0 and +14 before aerosol challenge with 10 mg/mL OVA for 20 min on days +28–30 and sample collection on day +31. LYS, lysozyme. (A) Concentration of serum OVA-IgE was determined by sandwich ELISA. (B) Lungs were flushed with BALF, total cell counts were determined, and samples were cytospun onto slides before DiffQuik staining for differential cell counts of bronchoalveolar lavage eosinophils. (C) Lungs were fixed in formalin and stained with H&E. (D) Cytokines from BALF supernatant were analyzed by Milliplex. Results are mean ± SEM and are representative of three separate experiments. *P < 0.05; **P < 0.01. Reproduced with permission from [73].

Figure 7

pMHC-NPs suppress MOG-induced EAE in vivo. C57BL/6 mice were immunized with pMOG_35-55. (A) EAE scores of mice treated from day 14 (n=4 each). (B) EAE scores of mice treated from day 21 (n=10, 7 and 3 from top). (C) Representative microglial IBA1 stainings and relative rank scores in the cerebellum of mice from B (n=4–5). Reproduced with permission from [133]. ©2016 Macmillan Publishers Ltd.