Use of agents capable of inhibiting activation of MAIT cells (mucosal-associated invariant T cells) for the treatment of rheumatoid arthritis

Abstract

Rheumatoid arthritis (RA) is the most common form of inflammatory rheumatism, affecting small joints, where swelling and pain reside, accompanied by structural damage that can lead to functional disability if no treatment is offered. The present inventors have shown that circulating MAIT cells are reduced and exhibit an activated and anti-apoptotic phenotype in rheumatoid arthritis patients compared with healthy controls. MAIT cell levels were also found to be increased in synovial fluid compared with peripheral blood, suggesting that the deficiency of circulating MAIT cells is due to migration of MAIT cells into joints. The severity of arthritis induced by methylated bovine serum albumin was reduced in mice from which MAIT cells had been ablated. This data suggests that MAIT cells are responsible for the exacerbation of arthritis. Thus, the present invention relates to the use of agents capable of inhibiting MAIT cell activation for the treatment of rheumatoid arthritis.

Description

[Technical Field]

[0001] Field of the invention: The present invention is in the field of medicine, particularly rheumatology.

[0002] Background of the invention: Rheumatoid arthritis (RA) is the most common form of inflammatory rheumatism, affecting small joints, the locus of swelling and pain, accompanied by structural damage, which will cause functional disability if no treatment is proposed. In order to avoid joint destruction and disability, treatment should aim to suppress systemic and joint inflammation as quickly as possible and reach goals such as remission or low disease activity in all patients. According to US, European, and French recommendations for the management of rheumatoid arthritis, methotrexate (MTX) should be initiated first as soon as possible after the diagnosis of rheumatoid arthritis is made (C. Gaujoux-Viala, L. Gossec, A. Cantagrel, M. Dougados, B. Fautrel, X. Mariette, H. Nataf, A. Saraux, S. Trope, B. Combe, Recommendations of the French Society for Rheumatology for managing rheumatoid arthritis. Joint Bone Spine 81, 287-297 (2014)). However, despite methotrexate's potent anti-inflammatory effects, methotrexate, with or without corticosteroids, induces low disease activity in 25-50% of patients with early-stage rheumatoid arthritis and induces remission in only 10-20% of patients over 6-12 months (EB Lee, R. Fleischmann, S. Hall, B. Wilkinson, JD Bradley, D. Gruben, T. Koncz, S. Krishnaswami, GV Wallenstein, C. Zang, SH Zwillich, RF van Vollenhoven, Tofacitinib versus methotrexate in rheumatoid arthritis. N Engl J Med 370, 2377-2386 (2014)). Our advances over the past several decades in understanding the pathophysiology of rheumatoid arthritis have led to the development of novel treatments designed to act against precise therapeutic targets.These new molecules, called "biologics," include tumor necrosis factor alpha (TNFα) blockers, such as adalimumab, certolizumab, etanercept, golimumab, and infliximab; interleukin-1 receptor antagonists (IL1-Ra, anakinra); inhibitors of costimulatory pathways involved in T lymphocyte activation [CTLA4-Ig or abatacept (ABA)]; monoclonal antibodies (mAbs) that bind to CD20 expressed on B cells (rituximab); and monoclonal antibodies that bind to the IL-6 receptor (tocilizumab) (J.S. Smolen, D. Aletaha, Rheumatoid arthritis therapy reappraisal: strategies, opportunities, and challenges. Nat Rev Rheumatol 11, 276-289 (2015)). All of these drugs, in conjunction with the core drug methotrexate, have proven effective in reducing joint inflammation, thereby reducing pain and limiting or halting joint destruction. However, approximately 30% of rheumatoid arthritis patients do not respond to these treatments at all, and the response to all of these drug therapies varies greatly from patient to patient. This is probably because rheumatoid arthritis is a syndrome grouped into several entities with different pathophysiological mechanisms, ranging from the most "inflammatory" form, which is more likely to respond to immunotherapy directed against proinflammatory cytokines (anti-TNFα antibodies, anti-IL6 antibodies, etc.), to the most "autoimmune" form, which responds better to anti-cellular therapy (rituximab and abscisic acid). Therefore, there is a need to identify new targets for the treatment of rheumatoid arthritis.

[0003] Mucosal-associated invariant T (MAIT) cells are a subset of innate-like immune cells found in peripheral blood, intestinal mucosa, and abundantly in the human liver. In humans, MAIT cells express the α chain of the invariant T cell receptor, i.e., the Vα7.2-Jα33 chain. MAIT cells can produce interferon-γ, granzyme B (GrB), and IL-17; are restricted by the major histocompatibility complex class I-related molecule MR1 (MHC class I-related-1 molecule); and are activated by cells infected with various microorganisms. Vitamin B2 (riboflavin) metabolites produced by bacteria and yeast are required to generate MAIT cell-activating ligands. Recently, MAIT cells have also been shown to have various harmful and protective functions in autoimmune, inflammatory, and metabolic diseases. The involvement of MAIT cells in a wide range of pathological conditions makes them attractive targets for promising therapeutic approaches (Toubal A, Nel I, Lotersztajn S, Lehuen A. Mucosal-associated invariant T cells and disease. Nat Rev Immunol. 2019 Oct;19(10):643-657).

[0004] Summary of the Invention: The invention is defined by the claims. In particular, the invention relates to the use of agents capable of inhibiting the activation of MAIT cells for the treatment of rheumatoid arthritis.

[0005] Detailed description of the invention: A first object of the present invention relates to a method for treating rheumatoid arthritis in a patient in need thereof, comprising administering to the patient a therapeutically effective amount of an agent capable of inhibiting the activation of MAIT cells.

[0006] As used herein, the term "rheumatoid arthritis" or "RA" has its common meaning in the art and refers to a systemic, autoimmune, inflammatory condition characterized by persistent synovial inflammation of the joints, progressive joint destruction, and varying degrees of deformity and functional disability. The process begins with the intervention of humoral and cellular factors, which generate inflammation-mediating molecules, attract and activate peripheral blood cells, and cause the proliferation and activation of synovial cells, which invade and destroy articular cartilage, subchondral bone, tendons, and ligaments. Examples of physiological indicators of rheumatoid arthritis include symmetric joint swelling and pain during passive movement, which are characteristic but not invariable features of rheumatoid arthritis. In rheumatoid arthritis, disease activity can be measured according to standard criteria accepted in the art. The "Disease Activity Score" or "DAS" is a measure of rheumatoid arthritis activity. In Europe, the Disease Activity Score is an accepted standard for research and clinical practice. The following parameters are included in the calculation (Van Gestel AM, Prevoo MLL, van't Hof MA, et al. Development and validation of the European League Against Rheumatism response criteria for RA. Arthritis Rheum 1996; 39:34-40): number of tender joints by palpation (TEN), number of swollen joints (SW), erythrocyte sedimentation rate (ESR), and patient assessment of disease activity (VAS; mm).

[0007] As used herein, the term "patient" refers to a mammal. In some embodiments, the patient refers to any patient (preferably a human) suffering from rheumatoid arthritis. In some embodiments, the patient is resistant to methotrexate or to a biologic selected from the group consisting of tumor necrosis factor alpha (TNFα) blockers, such as adalimumab, certolizumab, etanercept, golimumab, and infliximab; interleukin-1 receptor antagonists (IL1-Ra, anakinra); costimulatory pathway inhibitors involved in T lymphocyte activation [CTLA4-Ig or abatacept (ABA)]; monoclonal antibodies (mAbs) that bind to CD20 expressed on B cells (rituximab); and monoclonal antibodies that bind to IL-6 receptors (tocilizumab).

[0008] As used herein, the term "treatment" or "treating" refers to both prophylactic or preventative treatment, as well as curative or disease-modifying treatment (including treatment of patients at risk of or suspected of having a disease, as well as patients who are ill or have been diagnosed with a disease or medical condition), including the suppression of clinical recurrence. Treatment may be administered to patients with a medical disorder or who are likely to eventually develop a disorder to prevent, cure, delay the onset of, reduce the severity of, or ameliorate one or more symptoms of the disorder or a recurring disorder, or to extend the patient's survival beyond that expected in the absence of such treatment. By "therapeutic regimen" is meant a pattern of disease treatment, e.g., a dosing pattern used during therapy. A therapeutic regimen can include an induction regimen and a maintenance regimen. The phrase "induction regimen" or "induction period" refers to a therapeutic regimen (or portion of a therapeutic regimen) used for the initial treatment of a disease. The general goal of an induction regimen is to provide high levels of drug to the patient during the initial period of the treatment regimen. An induction regimen may use (in part or in whole) a "loading regimen," which may involve administering a higher dose of drug than a physician would use during a maintenance regimen, administering a drug more frequently than a physician would administer a drug during a maintenance regimen, or both. The phrase "maintenance regimen" or "maintenance period" refers to a therapeutic regimen (or portion of a therapeutic regimen) used to maintain a patient during disease treatment, e.g., so that the patient remains in remission for an extended period of time (months or years). A maintenance regimen may use continuous therapy (e.g., administering a drug at regular intervals, e.g., weekly, monthly, yearly, etc.) or intermittent therapy (e.g., intermittent treatment, intermittent treatment, treatment upon relapse, or treatment upon reaching certain predetermined criteria (e.g., symptoms of disease, etc.)).

[0009] As used herein, the term "MAIT cells" or "mucosal-associated invariant T cells" refers to a population of T cells present in mammals, preferably humans, that display an invariant T cell receptor α chain containing Vα7.2-Jα33 (in humans), a defined length of CDR3, and a limited number of Vβ segments, along with an activated phenotype (CD44) (see, e.g., Lantz and Bendelac, 1994, J. Exp. Med. 180:1097-106; Tilloy et al., J. Exp. Med., 1999, 1907-1921; Treiner et al. (2003) Nature 422:164-169, the entire disclosures of each of which are incorporated herein by reference). MAIT cells are generally CD8 positive (most expressing the homodimeric form of CD8αα) or CD4 negative / CD8 negative (double negative) and are restricted by the non-classical MHC class I molecule MHC class I-associated-1 molecule. For the purposes of the present invention, any T cell expressing the invariant Vα7.2-Jα33α T cell receptor chain is considered to be a MAIT cell. Typically, the α chain is associated with an invariant CDR3 and either Vβ2 or Vβ13.

[0010] As used herein, the expression "an agent capable of inhibiting the activation of MAIT cells" refers to any molecule that can inhibit the pro-inflammatory function of MAIT cells under cellular and / or physiological conditions.

[0011] In some embodiments, the agent is a small organic molecule. MAIT cell inhibitors are known in the art and typically include those described in Corbett, AJ et al. T-cell activation by transitory neo-antigens derived from distinct microbial pathways. Nature 509, 361-365 (2014); and Keller, AN et al. Drugs and drug-like molecules can modulate the function of mucosal-associated invariant T cells Nat Immunol. 2017 Apr;18(4):402-411. Other examples include those described in International Patent Application No. WO2014005194. In some embodiments, the inhibitor is selected from the group consisting of 6-formylpterin, acetyl-6-formylpterin (Ac-6-FP), 3-formylsalicylic acid (3-F-SA), 5-formylsalicylic acid (5-F-SA), and 2-hydroxy-1-naphthaldehyde (2-OH-1-NA).

[0012] In some embodiments, the agent is an antibody. Thus, as used herein, the term "antibody" is used to refer to any antibody-like molecule having an antigen-binding region, and this term includes any antibody-like molecule having an antigen-binding domain, such as Fab', Fab, F(ab')2, single domain antibodies (DABs), TandAbs dimers, Fv, scFv (single-chain Fv), two-chain Fv, two-chain-single-chain Fv, Fd, linear antibodies, minibodies, diabodies, bispecific antibody fragments, bibodies, tribodies (scFv-Fab fusions, bispecific or trispecific, respectively); sc-diabodies; kappa (lambda) bodies (scFv-CL fusions, ); BiTEs (bispecific T cell-inducing antibodies, scFv-scFv tandems that attract T cells); DVD-Igs (dual variable domain antibodies, bispecific formats); SIPs (small immune proteins, a type of minibody); SMIPs ("small modular immunopharmaceuticals" scFv-Fc dimers; DARTs (two-chain stabilized diabodies "dual affinity retargeting"); miniature antibody mimetics containing one or more CDRs, and the like. Techniques for preparing and using various antibody-based constructs and fragments are well known in the art (see Kabat et al., 1991, which is specifically incorporated herein by reference).

[0013] In some embodiments, the agent is an antibody that depletes MAIT cells (i.e., a "depleting antibody"). As used herein, the term "depletion" with respect to MAIT cells refers to a measurable reduction in the number of MAIT cells in a subject. The reduction can be at least about 10%, e.g., at least about 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or more. In some embodiments, the depleting antibody binds to a cell surface marker of MAIT cells, preferentially a specific cell surface marker of MAIT cells. In some embodiments, the agent is an anti-Vα7.2-Jα33 depleting antibody, such as those described in International Patent Publication No. WO2008087219. In some embodiments, the depleting antibody mediates antibody-dependent cell-mediated cytotoxicity. As used herein, the term "antibody-dependent cell-mediated cytotoxicity" or "ADCC" refers to a cell-mediated response in which nonspecific cytotoxic cells (e.g., natural killer (NK) cells, neutrophils, and macrophages) recognize bound antibodies on target cells and subsequently cause lysis of the target cells. In some embodiments, the depletion antibodies are IgG1 antibodies. In some embodiments, the depletion antibodies are IgG3 antibodies.

[0014] In some embodiments, the agent is an antibody that blocks presentation of antigenic ligands (e.g., microbial vitamin B metabolites) by MHC class I-associated-1 molecules. In some embodiments, the antibody blocks the interaction between MHC class I-associated-1 molecules and Vα7.2-Jα33 receptors. In some embodiments, the antibody binds to MHC class I-associated-1 molecules. Thus, these antibodies are referred to as "neutralizing" or "blocking" or "blocking" antibodies. In some embodiments, the agent is an anti-MHC class I-associated-1 molecule-neutralizing antibody. In some embodiments, the agent is an anti-Vα7.2-Jα33 neutralizing antibody, such as those described in International Patent Publication No. WO2008087219. Such antibodies are useful, inter alia, for reducing MAIT immune cell activity.

[0015] As used herein, the phrase "therapeutically effective amount" refers to an amount of a drug effective at a dosage and for a period of time required to achieve a desired therapeutic result. A therapeutically effective amount of a drug can vary depending on factors such as the disease state, age, sex, and weight of the individual, as well as the ability of the drug to elicit a desired response in the individual. A therapeutically effective amount is also an amount in which any toxic or adverse effects of the antibody or antibody portion are outweighed by the therapeutically beneficial effects. The effective dose and dosage regimen for a drug depends on the disease or condition to be treated and can be determined by one of ordinary skill in the art. A physician with ordinary skill in the art can easily determine and prescribe the effective amount of the pharmaceutical composition required. For example, a physician can start the dose of a drug used in a pharmaceutical composition at a level lower than required to achieve the desired therapeutic effect and gradually increase the dose until the desired effect is achieved. Generally, an appropriate dose of a composition of the present invention will be the amount of the compound that is the lowest dose effective to produce a therapeutic effect according to a particular dosage regimen. Such an effective dose will generally depend on the factors described above.

[0016] Typically, the agents of the present invention are administered to a subject in the form of a pharmaceutical composition containing a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers that can be used in these compositions include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts, or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol, and wool fat. For use in administering to a subject, the composition will be formulated for administration to a subject. The compositions of the present invention can be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, bucally, vaginally, or via an implanted reservoir. As used herein, the term "subcutaneous, intravenous, intramuscular, intra-articular, intrasynovial, intracapsular, intrathecal, intrahepatic, intralesional, and intracranial injection or infusion" includes techniques for injection or infusion. Sterile injectable forms of the compositions of the present invention may be aqueous or oily suspensions. These suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. Sterile injectable preparations may also be sterile injectable solutions or suspensions in non-toxic parenterally acceptable diluents or solvents, such as solutions in 1,3-butanediol. Acceptable vehicles and solvents that may be used include water, Ringer's solution, and isotonic sodium chloride solution. Furthermore, sterile, fixed oils are conveniently used as solvents or suspending media. For this purpose, any non-irritating, fixed oil may be used, including synthetic monoglycerides or diglycerides. Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions.These oily solutions or suspensions may also contain long-chain alcohol diluents or dispersants, such as carboxymethylcellulose or similar dispersants, commonly used in formulating pharmaceutically acceptable dosage forms, including emulsions and suspensions. Other commonly used surfactants, such as Tween, Span, and other emulsifiers or bioavailability enhancers, commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms, may also be used for formulation purposes. The compositions of the present invention may be orally administered in any orally acceptable dosage form, including, but not limited to, capsules, tablets, aqueous suspensions, or solutions. For tablets for oral use, commonly used carriers include lactose and cornstarch. Lubricants, such as magnesium stearate, are also typically added. Diluents useful for oral administration in capsule form include, for example, lactose. When aqueous suspensions are required for oral use, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweeteners, flavors, or coloring agents may also be added. Alternatively, the compositions of the present invention may be administered in the form of rectal suppositories. These can be prepared by mixing the drug with a suitable non-irritating excipient that is solid at room temperature but liquid at rectal temperature, and therefore will melt in the rectum to release the drug. Such excipients include cocoa butter, beeswax, and polyethylene glycol. The compositions of the present invention may also be administered topically, especially when the target of treatment includes areas or organs easily accessible by topical application, including diseases of the eyes, skin, or lower gastrointestinal tract. Suitable topical formulations for each of these areas or organs are easily prepared. The compositions for topical application may be formulated into a suitable ointment containing the active ingredient suspended or dissolved in one or more carriers. Carriers for topical administration of the compounds of the present invention include, but are not limited to, mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene, polyoxypropylene compounds, emulsifying wax, and water. Alternatively, the compositions can be formulated in a suitable lotion or cream containing the active compounds suspended or dissolved in one or more pharmaceutically acceptable carriers.Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol, and water. Topical application to the lower intestinal tract can be effected in a rectal suppository formulation (see above) or in a suitable enema formulation. Patches may also be used. The compositions of the present invention may also be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well known in the art of pharmaceutical formulation, and may be prepared as a solution in saline using benzyl alcohol or other suitable preservatives, absorption enhancers to enhance bioavailability, hydrogen fluoride, and / or other convenient solubilizing or dispersing agents. For example, the antibody present in the pharmaceutical composition of the present invention may be supplied at a concentration of 10 mg / mL in either 100 mg (10 mL) or 50 mg (50 mL) single-use vials. The product is formulated for intravenous administration in 9.0 mg / mL sodium chloride, 7.35 mg / mL sodium citrate dihydrate, 0.7 mg / mL polysorbate 80, and sterile water for injection. The pH is adjusted to 6.5. An exemplary suitable dose range for the antibody in the pharmaceutical composition of the invention is about 1 mg / mL. 2 to 50 mg / m 2 It will be understood, however, that these schedules are exemplary, and that optimal schedules and formulation regimens can be adapted, taking into account the affinity and tolerability of the particular antibody in the pharmaceutical composition, which must be determined in clinical trials. Pharmaceutical compositions of the present invention for injection (e.g., intramuscular, intravenous) can be prepared to contain sterile buffered water (e.g., 1 ml for intramuscular administration) and about 1 ng to about 100 mg, for example, about 50 ng to about 30 mg or more, preferably about 5 mg to about 25 mg, of the inhibitor of the present invention.

[0017] The present invention will be further illustrated by the following figures and examples, which, however, should not be construed as limiting the scope of the present invention in any way. [Brief explanation of the drawings]

[0018] [Figure 1] Deficiency of circulating MAIT cells in rheumatoid arthritis (RA). Peripheral blood mononuclear cells (5 × 106) were collected from healthy donors (n = 41) and rheumatoid arthritis patients (n = 43) and analyzed by flow cytometry. Circulating MAIT cells were identified as CD161highVα7.2+ T cells. MAIT cells among CD3+ cells were reduced in participants with RA compared with healthy donors (median (IQR) proportion of MAIT cells: 0.41% (0.19-0.97) in RA patients compared with 2.55% (1.5-2.6) in healthy donors, p < 0.0001). Each symbol represents a single individual, and the small horizontal line indicates the median. Test: nonparametric two-tailed Mann-Whitney test. IQR: interquartile range. MAIT cells: mucosal-associated invariant T cells. PBMCs: peripheral blood mononuclear cells. RA: Rheumatoid arthritis. [Figure 2] Correlation between Circulating MAIT Cell Deficiency and Disease Activity We next investigated whether the frequency of MAIT cells was associated with clinical variables in rheumatoid arthritis patients (n = 43). Disease activity was assessed using the Disease Activity Score 28 (DAS28), calculated based on joint swelling and tenderness, systemic inflammatory markers (CRP, C-reactive protein), and patient feedback. MAIT cells frequently negatively correlated with disease activity (r = -0.38, p = 0.011). Thus, the more active a patient's disease, the more pronounced the deficiency of circulating MAIT cells. Each symbol represents a single individual. Test: Spearman correlation test. CRP: C-reactive protein. DAS28: Disease Activity Score. IQR: Interquartile range. MAIT cells: Mucosal-associated invariant T cells. RA: Rheumatoid arthritis. [Figure 3]Accumulation of MAIT Cells in Synovial Fluid Compared to Blood in Patients with Rheumatoid Arthritis. Peripheral blood mononuclear cells (5 × 106 cells) and synovial fluid mononuclear cells (5 × 106 cells) were collected from rheumatoid arthritis patients (n = 15) and analyzed by flow cytometry. Circulating and synovial fluid MAIT cells were identified as CD161highVα7.2-positive cells. The proportion of synovial fluid MAIT cells among CD3-positive cells was higher than that of circulating MAIT cells (median (IQR) proportion of MAIT cells: 0.65% (0.2-4.7%) in synovial fluid compared with 0.23% (0.09-0.88%) in blood, p = 0.025). Each symbol represents a single individual. Test: two-tailed Wilcoxon matched-pair test. IQR: interquartile range. MAIT cells: mucosal-associated invariant T cells. PBMC: peripheral blood mononuclear cells. SF: synovial fluid. [Figure 4]Activation profile and disappearance of CD56 in synovial fluid MAIT cells compared with peripheral blood MAIT cells in rheumatoid arthritis patients. We next investigated the phenotype of circulating MAIT cells in healthy donors and rheumatoid arthritis patients, as well as the phenotype of synovial fluid MAIT cells in rheumatoid arthritis patients. In rheumatoid arthritis patients, circulating MAIT cells exhibited an activated phenotype, as indicated by a significantly increased frequency of CD69-positive MAIT cells compared with healthy donors (median (IQR) frequency of CD69-positive MAIT cells: 22.2% (11.6-30.4) compared with 9.8% (4-14.7) in healthy donors, p=0.0005, Figure 4A ), and an increased frequency of CD25-positive MAIT cells compared with healthy donors (median (IQR) frequency of CD25-positive MAIT cells: 22.2% (11.6-30.4) compared with 9.8% (4-14.7) in healthy donors, p=0.0005, Figure 4A ). The median (IQR) frequency of CD69-positive MAIT cells in synovial fluid compared with blood was 2.2% (0.3-18.5) vs. 0.57% (0.29-0.78), p = 0.03, Figure 4B. Furthermore, in patients with rheumatoid arthritis, synovial fluid MAIT cells were even more activated compared with blood, as indicated by a 75% increase in the frequency of CD69-positive MAIT cells in synovial fluid compared with blood (median (IQR) frequency of CD69-positive MAIT cells: 86.1% (74.5-94.1) in synovial fluid compared with 22.2% (11.6-30.4) in blood). , p=0.0001, Figure 4A). This was consistent with a strong trend toward an increase in CD25-positive MAIT cells, another T cell activation marker, in synovial fluid compared to blood (median (IQR) frequency of CD25-positive MAIT: 7.4% (1.2-34.5) compared to 2.2% (0.3-18.5), p=0.057, Figure 4B). Finally, we observed a decrease in the adhesion molecule CD56 in synovial fluid compared to blood (median (IQR) frequency of CD56-positive MAIT cells: 2.2% (0.3-18.5) compared to 7.4% (1.2-34.5), p=0.057, Figure 4B). R: 38.9% (26.5-55.4%) compared with 62% (49.3-74%), p=0.0012 (Figure 4C). This suggests the migration of MAIT cells from the blood to the inflamed joints. Each symbol represents a single individual, and the small horizontal line indicates the median. Tests: Nonparametric two-sided Mann-Whitney test between healthy donor blood and rheumatoid arthritis blood; two-sided Wilcoxon matched-pair test between rheumatoid arthritis blood and rheumatoid arthritis synovial fluid. IQR: Interquartile range.MAIT cells: mucosal-associated invariant T cells. RA: rheumatoid arthritis. [Figure 5] Reduced Severity of Rheumatoid Arthritis in MAIT Cell-Depleted Mice. Arthritis was induced by intra-articular injection of methylated bovine serum albumin (mBSA) in MHC class I-related-1 molecule- / - (MAIT cell-depleted) and wild-type C57BL / 6 mice previously immunized with methylated bovine serum albumin. We included nine female mice, 6-8 weeks of age, in each group. Mice were evaluated daily for signs of arthritis, assessed by measuring tarsal thickness and scored on a scale of 0 to 4 (0 = no change, 1 = local swelling and redness of the digits, 2 = mild swelling and redness, 3 = severe swelling and redness, 4 = necrosis). Mice were sacrificed on day 8, and paws were isolated, fixed in 4% paraformaldehyde, and decalcified for histopathological evaluation. All mice developed arthritis, with peak inflammation occurring on day 1. Wild-type mice had higher clinical scores (A) and a significantly increased area under the curve (B) than MHC class I-related-1 molecule knockout mice (mean area under the curve of 4.7 in the MHC class I-related-1 molecule knockout group compared to 9.1 in the wild-type group, p=0.0064). Values ​​are means + / - standard error of the mean of nine mice in each group. Test: Student's T-test. AUC: area under the curve. MAIT cells: mucosal-associated invariant T cells. mBSA: methylated bovine serum albumin. KO: knockout, MHC class I-related-1 molecule: MHC class I-related-1 molecule. [Figure 6]Arthritis was induced by intra-articular injection of methylated bovine serum albumin (mBSA) into Vα19 transgenic mice (which have 10 times more MAIT cells than wild-type mice) and wild-type C57BL / 6 mice, both of which had been pre-immunized with methylated bovine serum albumin. We included seven Vα19 transgenic mice and 11 wild-type mice, all female, aged 6–14 weeks. Mice were evaluated daily as previously described. All mice developed arthritis, with inflammation peaking on day 1. Although there was no difference in clinical scores between the two groups in the first part of the experiment (mean AUC of 4.3 in the wild-type group compared to 4.6 in the Vα19 transgenic group, p=0.7), we observed a more pronounced persistence of arthritis in the Vα19 transgenic mice after day 5 (mean AUC of 1.1 in the Vα19 transgenic mice compared to 0.55, p=0.025). Values ​​are the mean values ​​+ / - standard error of the mean for each group of mice. Test: Student's T-test. AUC: area under the curve. MAIT cells: mucosal-associated invariant T cells. mBSA: methylated bovine serum albumin. [Figure 7]A strong trend toward reduced arthritis severity in mice treated with molecules that inhibit MAIT cell activation. Arthritis was induced by intra-articular injection of methylated bovine serum albumin (mBSA) in wild-type mice previously immunized with methylated bovine serum albumin and treated with acetyl-6-formylpterin (Ac6FP), a non-activating ligand for MAIT cells, or PBS. Ac6FP or PBS was injected intraperitoneally twice weekly. We included six wild-type mice, all female and 8 weeks old, in each group. Mice were evaluated once daily as previously described. All mice developed arthritis, with inflammation peaking on day 1. Mice treated with Ac6FP had lower clinical scores (A) and a reduced area under the curve (B) compared with mice treated with PBS (mean AUC of 3 for the Ac6FP-treated group vs. 5.5 for the PBS group, p=0.07). Values ​​are means + / - standard error of six mice per group. Test: Student's T-test. Ac6FP: acetyl-6-formylpterin. AUC: area under the curve. MAIT cells: mucosal-associated invariant T cells. mBSA: methylated bovine serum albumin. PBS: Dulbecco's phosphate-buffered saline.

[0019] Working Example: Circulating MAIT cells were reduced in rheumatoid arthritis patients compared with healthy controls (Figures 1 and 2) and displayed an activated phenotype (Figures 4A and 4B). MAIT cell levels were also found to be increased in synovial fluid compared with peripheral blood (Figure 3), had an activated profile (Figure 4A), and expression of the adhesion molecule CD56 was reduced (Figure 4C), suggesting that the lack of circulating MAIT cells is due to migration of MAIT cells into the joints. The severity of mBSA-induced arthritis was reduced in mice in which MAIT cells were ablated (Figure 5), but the persistence of arthritis was more pronounced in mice overexpressing MAIT cells (Figure 6). Furthermore, the severity of arthritis tended to be reduced in mice treated with molecules that inhibit MAIT cell activation (Figure 7). All these data suggest that MAIT cells contribute to the exacerbation of arthritis.

[0020] References: Throughout this application, various references describe the state of the art to which this invention pertains, the disclosures of which are hereby incorporated by reference into the present disclosure.

Claims

1. A method for treating rheumatoid arthritis in a patient in need thereof, comprising administering to the patient a therapeutically effective amount of an agent capable of inhibiting activation of mucosal-associated invariant (MAIT) cells.

2. The method of claim 1 , wherein the agent is a small organic molecule.

3. 3. The method of claim 2, wherein the agent is selected from the group consisting of 6-formylpterin, acetyl-6-formylpterin (Ac-6-FP), 3-formylsalicylic acid (3-F-SA), 5-formylsalicylic acid (5-F-SA), and 2-hydroxy-1-naphthaldehyde (2-OH-1-NA).

4. The method of claim 1 , wherein the agent is an antibody.

5. The method of claim 4, wherein the antibody is an antibody that eliminates MAIT cells.

6. The method of claim 4, wherein the antibody is an antibody that blocks presentation of antigenic ligands by MR1 (MHC class I related-1 molecule).

7. 7. The method of claim 6, wherein the antibody blocks the interaction between MR1 (MHC class I related-1 molecule) and the V7.2-J33 receptor.

8. The method of claim 7, wherein the antibody binds to MR1 (MHC class I related-1 molecule).

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