Diabetes Volume 70, June 2021 1247 The SAgA of Antigen-Specific Immunotherapy for Type 1 Diabetes Roberto Mallone1,2 and Sylvaine You1 Diabetes 2021;70:1247–1249 | https://doi.org/10.2337/dbi21-0011 1 Islet antigen-specific strategies are the holy grail of genic BDC2.5 CD4 T cells (8), or with its more potent immunotherapy for type 1 diabetes (T1D) (1), as they se- p79 mimotope. Contrary to free peptides, these SAgAs ef- lectively target the autoimmune responses involved in ficiently prevented diabetes in NOD mice only when used b-cell destruction (2). The idea is to induce a response op- in combination. While this may reflect the need to posite to that of a conventional vaccine. The administra- achieve a critical quantitative threshold of T cells targeted, tion of antigen(s) in the absence of inflammation (e.g., a synergistic functional effect is plausible. Indeed, 2.5HIP- adjuvants) should potentiate the outcomes of physiological reactive and p79-reactive T cells, which were, surprisingly, immune homeostasis, i.e., anergy, regulatory polarization, distinct populations, underwent different outcomes upon 1 and, to a lesser extent, deletion. Such antigens can be de- treatment with their cognate SAgA (Fig. 1). IL-10 Treg- livered as proteins, peptides, bacteria engineered to secrete ulatory (Tr)1 cells were induced by the more potent p79 1 these products, DNA plasmids, or nanoparticles coated ligand, while Foxp3 Tregs were amplified, but not de with antigens (either alone or preloaded on MHC mole- novo induced, by the weaker (and less soluble) 2.5HIP. COMMENTARY cules). Peptides are easy to synthesize by amino acid These effects were dose-dependent, as SAgAs induced Tr1 1 chemistry, but they are variably water-soluble and short- cells at higher dose and Foxp3 Tregs at lower dose. lived in vivo (in the order of minutes), which makes it dif- SAgAs promoted more effective and persistent engage- ficult to achieve the steady concentrations more suitable ment and expansion of autoreactive T cells than free pep- for tolerance induction. Nonetheless, peptide-based immu- tides in vivo, with no deletional effect observed. Critically, notherapies have proven safe and, in some cases, docu- SAgAs were stronger inducers of tolerance-associated mented encouraging immune and clinical outcomes (3). markers, including decreased CD44 expression and in- In this issue of Diabetes, Firdessa-Fite et al. (4) give creased CD73/FR4, Lag-3, PD-1, and KLRG-1, suggestive peptides a boost by mounting them on a hyaluronan of anergic/regulatory/exhausted phenotypes. The cSAgA backbone to obtain a soluble antigen array (SAgA). First, formulation proved superior to hSAgA at inducing CD73/ this backbone accounts for their superior solubility. The FR4, PD-1, and IL-10, at preventing diabetes, and at linker used was either stable (“click” chemistry, cSAgA) or avoiding anaphylaxis (likely as a result of decreased shed- hydrolysable (hSAgA), to facilitate peptide release upon ding of free peptides). Although NOD mice are particu- uptake by antigen-presenting cells (APCs). Second, SAgAs larly prone to peptide-induced anaphylaxis (9), this issue deliver multiple peptide copies to individual APCs, thus is critical in the allergy setting but probably less so in hu- providing a multivalent (high avidity) engagement of T man T1D. Originally described after insulin B9-23 peptide cells that is more efficient for signal transduction. This immunization in NOD mice (10), such reactions were not may be critical for the low-affinity T cells involved in au- observed, despite Th2 polarization, in a small-scale hu- toimmunity. A gradient of biodistribution was observed man trial with a similar peptide (11), and in other pep- in draining lymph nodes proximal or distal to the inter- tide-based intervention trials. scapular subcutaneous immunization site, which raises Some questions are open for further investigation, in- the question of whether sites closer to pancreatic lymph cluding whether the therapeutic outcome is IL-10–depen- 1 nodes (5) may achieve superior effects. Thymic delivery dent, the role of B-cell and CD8 T-cell tolerance, the may also deserve attention (6,7). need for continuous treatment, the efficacy at inducing SAgAs were loaded with either the hybrid peptide diabetes remission, and human T-cell studies in vitro 2.5HIP, which is the natural antigen ligand of diabeto- and/or in humanized mice. Although the peptides to load 1Universite de Paris, Institut Cochin, CNRS, INSERM, Paris, France © 2021 by the American Diabetes Association. Readers may use this article 2Assistance Publique H^opitaux de Paris, Cochin Hospital, Service de as long as the work is properly cited, the use is educational and not for Diabetologie et Immunologie Clinique, Paris, France profit, and the work is not altered. More information is available at https:// Corresponding author: Roberto Mallone, [email protected] www.diabetesjournals.org/content/license. See accompanying article, p. XXXX. Diabetes Publish Ahead of Print, published online May 20, 2021 1248 Commentary Diabetes Volume 70, June 2021 p79 mimotope IL-10 DCs cSAgA Myeloid cells B cells? p79-reactive Natural Tr1 CD4+ T cells 2.5HIP FR4 peptide Anergy/exhaustion CD73 IL-10+ Tr1 differentiation PD-1 Lag-3 Memory? Th1 differentiation KLRG-1 Naïve? Foxp3+ Treg expansion Treg 2.5HIP-reactive CD4+ T cells Figure 1—Tolerogenic effects of p79 and 2.5HIP cSAgAs on their cognate CD41 T cells. DCs, dendritic cells. on SAgAs will need to be adapted to the HLA haplotype transplantation (19,20). Thus, states of T-cell unresponsive- of a given individual, the overrepresentation of HLA- ness may be a common feature of tolerance restoration and, DQ2/DR3 and -DQ8/DR4 in patients with T1D eases the possibly, of different degrees of “benign,” quiescent autoim- burden of this task. The use of longer peptides requiring munity in healthy individuals (21) and in patients with dif- some intermediate processing by APCs may also be con- ferent rates of disease progression (22,23). The work of sidered to increase epitope, and possibly HLA, coverage. It Firdessa-Fite et al. provides a novel therapeutic tool to at- will also be interesting to define the critical features of tempt restoring this state of benign autoimmunity. the “ideal” peptides. Although peptide solubility may limit the choice (e.g., leading to the exclusion of insulin B9-23 in this report), the list of options is expanding (12–14) Funding. Work in the laboratory of R.M. and S.Y. is supported by grants and now includes several neo-epitopes similar to the from The Leona M. and Harry B. Helmsley Charitable Trust (1901-03689), the 2.5HIP used here (8). In this respect, the Tr1 induction Fondation pour la Recherche Medicale (EQU20193007831), the Agence Natio- achieved by SAgAs through endogenous APCs is reminis- nale de la Recherche (ANR-17-CE17-0004, ANR-19-CE15-0014-01), the cent of that induced through artificial APCs (peptide- European Foundation for the Study of Diabetes EFSD/JDRF/Lilly European Pro- MHC class II–coated nanoparticles) (15). Any b-cell gramme in Type 1 Diabetes Research 2019, and the EU Innovative Medicines peptide was effective with this strategy, but only T cells Initiative 2 Joint Undertaking under grant agreements 115797 INNODIA and 945268 INNODIA HARVEST. These joint undertakings receive support from the that had already been antigen-primed during the disease 1 1 European Union’s Horizon 2020 research and innovation programme, Europe- process could differentiate into Tr1 (CD49b Lag-3 IL- 1 an Federation of Pharmaceutical Industries Associations, JDRF, and The 10 ) cells. This point remains to be addressed for SAgAs, Leona M. and Harry B. Helmsley Charitable Trust. but the earlier timing of treatment (8-week-old pre- Duality of Interest. No potential conflicts of interest relevant to this diabetic stage) might suggest different requirements com- article were reported. pared with nanoparticles (administered at diabetes onset). A head-to-head comparison with nanoparticles (which References were shown to revert disease), as well as with SAgAs load- 1. Culina S, Boitard C, Mallone R. Antigen-based immune therapeutics for ed with other b-cell antigens or with irrelevant peptides, type 1 diabetes: magic bullets or ordinary blanks? Clin Dev Immunol 2011; would be critical to strengthen the therapeutic potential 2011:286248 and specificity of SAgAs. Nanoparticles also drove differ- 2. Carre A, Richardson SJ, Larger E, Mallone R. Presumption of guilt for T entiation of B regulatory cells, and previous experience cells in type 1 diabetes: lead culprits or partners in crime depending on age with SAgA treatment in experimental autoimmune en- of onset? Diabetologia 2021;64:15–25 cephalomyelitis (16) suggests that this may be another 3. Alhadj Ali M, Liu YF, Arif S, et al. Metabolic and immune effects of therapeutic mechanism for tolerance spreading. Upregula- immunotherapy with proinsulin peptide in human new-onset type 1 diabetes. tion of the ectonucleotidase CD73 may further contribute Sci Transl Med 2017;9:eaaf7779 4. Firdessa-Fite R, Johnson SN, Leon MA, et al. Soluble antigen arrays to this bystander effects by promoting a local immuno- efficiently deliver peptides and arrest spontaneous autoimmune diabetes. suppressive environment via adenosine. Indeed, SAgAs, in 1 Diabetes 2021;70: XXXX–XXXX particular cSAgAs, induced antigen-reactive CD4 Tcells 5. Phillips B, Nylander K, Harnaha J, et al. A microsphere-based vaccine exhibiting an anergic (CD73highFR4high) and/or exhausted 1 1 prevents and reverses new-onset autoimmune diabetes. Diabetes 2008;57: (PD-1 , KLRG-1 ) phenotype. Similar phenotypes have 1544–1555 1 1 also been described in CD4 (anergy) and CD8 Tcells(an- 6. Zelenay S, Bergman ML, Paiva RS, et al. Cutting edge: intrathymic ergy and exhaustion) following anti-CD3 or LFA3-Ig treat- differentiation of adaptive Foxp31 regulatory T cells upon peripheral proinflam- ment in the context of human T1D (17,18) or murine islet matory immunization.