Vilma Urbonaviciute, Laura Romero-Castillo, Bingze Xu, +14, Huqiao Luo, Nadine Schneider, Sylvia Weisse, Nhu-Nguyen Do, Ana Oliveira-Coelho, Gonzalo Fernandez Lahore, Taotao Li, Pierre Sabatier, Christian M.Beusch, Johan Viljanen, Roman A.Zubarev, Jan Kihlberg, Johan Bäcklund, Harald Burkhardt, and Rikard Holmdahl admin@frankenthalerfoundation.org Authors Info & Affiliations
Edited by Shimon Sakaguchi, Osaka Daigaku, Osaka, Japan; received November 1, 2022; accepted May 8, 2023
June 12, 2023
120 (25) e2218668120
Current treatments of autoimmune diseases, like rheumatoid arthritis, is directed to inflammatory consequences of the disease process, with limitations regarding effectiveness and side effects. We have shown in vivo a highly effective tolerogenic vaccine, which consists of a complex between an antigenic glycopeptide from COL2 and a relevant MHCII molecule. The vaccine operates through binding directly to the T cell receptor on the T cell surface, leading to differentiation of the T cell into a distinct regulatory phenotype mediating a dominant tissue-specific tolerance.
A longstanding goal has been to find an antigen-specific preventive therapy, i.e., a vaccine, for autoimmune diseases. It has been difficult to find safe ways to steer the targeting of natural regulatory antigen. Here, we show that the administration of exogenous mouse major histocompatibility complex class II protein bounding a unique galactosylated collagen type II (COL2) peptide (A q–galCOL2) directly interacts with the antigen-specific TCR through a positively charged tag. This leads to expanding a VISTA-positive nonconventional regulatory T cells, resulting in a potent dominant suppressive effect and protection against arthritis in mice. The therapeutic effect is dominant and tissue specific as the suppression can be transferred with regulatory T cells, which downregulate various autoimmune arthritis models including antibody-induced arthritis. Thus, the tolerogenic approach described here may be a promising dominant antigen-specific therapy for rheumatoid arthritis, and in principle, for autoimmune diseases in general.
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Autoimmune diseases affect a large part (5 to 10%) of the population but still lack preventive and curative treatments. Rheumatoid arthritis (RA) is a common, severe, chronic disease characterized by autoimmune inflammation of peripheral joints, leading to progressive cartilage and bone destruction. Existing therapeutics predominantly target common later stages of effector pathways of inflammation, thereby increasing the risk of side effects such as infections. To improve early treatment, the antigen-specific autoimmunity regulating the onset of joint inflammation must be addressed. Before and around clinical onset, the autoimmune response expands to include citrullinated proteins (1). In addition, an immune response to joint proteins, including the major joint protein, type II collagen (COL2) and to citrullinated COL2, appears (2–4). COL2 is of particular interest, as it is the major protein component in cartilage as well as in central lymphoid organs, the thymus, and bone marrow (5–7). Indeed, immunization of COL2 with potent adjuvant induces the development of autoimmune arthritis in rodents (collagen-induced arthritis [CIA]) (8).
Experimental arthritis is dependent on major histocompatibility complex class II region (MHCII) interaction (9). In the mouse, CIA is associated with the MHCII allele A q (10) and dependent on T cell recognition of the galactosylated 259-273 COL2 epitope (galCOL2 259-273) (11, 12). Interestingly, T cells specific for the galCOL2 259-273 peptide occur in RA patients carrying the DRB1*0401 allele (13–16).
With the aim of developing a therapeutic vaccine, we have found that a recombinant A q molecule binding the galCOL2 259-273 peptide (a protein complex hereafter named A q–galCol2) could suppress the development of CIA (17, 18). Complexes of A q containing the COL2 peptide without galactosylation at position 264 had no effect. The reason for this remarkably selective regulatory effect is likely related to the fact that galactosylated COL2 is expressed only in cartilage (19), whereas nonglycosylated COL2 is expressed also in the thymus (5, 7). Thus, the T cell response to nonglycosylated COL2 is regulated by central tolerance mechanisms, whereas the T cell response to the galactosylated antigen is regulated by peripheral tolerance mechanisms.
Several different attempts have been made to develop tolerogenic strategies, but so far with limited clinical success. These approaches include whole-antigen tolerance (20), utilizing altered or unaltered peptide ligands (21–23), forming soluble peptide–MHC complexes, and utilizing various antigen-loaded particles (23). Furthermore, transferring Tregs (24) or tolerogenic dendritic cells (25) has also been studied as a potential approach toward promoting immune tolerance and reducing autoimmune disorders.
An efficient approach is by treatment with recombinant MHCII molecules bound to peptides, also called recombinant T cell receptor ligands (RTLs), which have demonstrated efficacy in several models of autoimmune diseases, such as uveitis, multiple sclerosis, and rheumatoid arthritis (21, 22, 26). The mode of action/s is however unclear. It has been demonstrated that such treatment primary target regulatory dendritic cells delivering the peptide for presentation (27–29), or coupled to particles, directly activating T cells (30). The variable results are likely dependent on the used carrier, peptide or the structure of the peptide–MHCII complex.
We previously reported that the galCOL2 259-273 peptide in complex with the soluble A q MHCII molecules induced self-tolerance in an antigen-specific manner (18). We have now found that the MHCII–peptide complex operates via direct interaction with the cognate T cell receptors, which are facilitated by the positively charged His-tag in the recombinant MHCII molecule, in combination with the galactosylated COL2 peptide. This interaction leads to the activation of distinct nonconventional regulatory T cells.
Consistent with the previous data (18), subcutaneous (s.c.) administration of A q–galCOL2 complexes using osmotic pumps significantly reduced the severity and incidence of CIA as well as considerably delayed the onset of the arthritis (Fig. 1 A). We used osmotic pumps instead of i.v. injections to prolong the availability of A q–galCOL2, based on in vitro and in vivo s tability experiments (SI Appendix, Fig. S1).
Fig. 1.
Image 1
Tolerogenic collagen peptide-based vaccine attenuates autoimmune pathologies (CIA, DTH, CAIA) in a T cell–dependent manner. (A) CIA in vaccinated mice as assessed by mean clinical score of arthritis severity (Left) and incidence of arthritis (percentage of affected mice [Right]) on different days postimmunization (DPIM). QB mice were immunized with 100 μg COL2 on day 0 and boosted on day 35 with 50 μg COL2. On day 7, mice were vaccinated s.c. with A q–peptide complexes using osmotic pumps (n = 10 for A q–galCOL2, n = 13 for A q–CLIP). (B) Histological examination of an ankle joint from A q–galCOL2-treated QB mice. (C) Histological examination of an ankle joint from A q–CLIP-treated mice demonstrating cell infiltration and cartilage/bone destruction. (D) Anti-COL2 IgG serum levels assessed at day 35 during CIA by ELISA. (E) DTH reactions (ear swelling) following treatment with A q–galCOL2 complexes. (F) Mean clinical score of arthritis severity (Left) and incidence (Right) following transfer of T cells from A q–galCOL2-treated and A q–CLIP-treated QB mice (n = 5 per group). (G) CAIA following transfer of CD4+T cells from A q–galCOL2-treated and A q–CLIP-treated BQ mice (n = 5 per group). Mice were immunized with COL2 on day 5 and vaccinated either with 100 μg A q–galCOL2 or A q–CLIP using osmotic pumps. On day 15, purified CD4+ T cells were transferred to BQ.Cia9i recipients, which were injected with the arthritogenic CAIA cocktail (M2139 and ACC1 anti-COL2 antibodies) on the same day as the T cell transfer. (H) CAIA following transfer of CD4+T cells from A q–galCOL2-treated and A q–CLIP-treated BQ mice (n = 5 per group). Mice were immunized with COL2 and on day 5 vaccinated either with 100 µg A q–galCOL2 or A q–CLIP using osmotic pumps. On day 15, purified CD4+ T cells were transferred to BQ.Fcgr2b−/− recipients, which were injected with the arthritogenic CAIA cocktail (M2139, L10D9 and 15A11 anti-COL2 antibodies) 10 d earlier than the T cell transfer. Results are expressed as mean ± SEM. DPIN, days postinjection.
The arthritis scoring data were confirmed by paw histology (Fig. 1 B and C). Anti-COL2 IgG antibody levels were reduced in the A q–galCol2-treated group as compared with the control group treated with A q–CLIP (Fig. 1 D).
Since Th1-mediated immunity is involved in RA (31, 32) and in animal models for RA (33
