APHERESE COLUMN FOR THE TREATMENT OF RHEUMATOID AND POLYARTHRITIS

Description

Technical field of the invention

[0001] The present invention relates to an apheresis column having a solid support comprising a composition of citrullinated peptides and said composition immobilized in the apheresis column for its use in a method of treating an autoimmune disease with anti-citrullinated protein autoantibodies. Technical background

[0002] Rheumatoid arthritis (RA) is the most common human autoimmune disease and also the most common chronic inflammatory rheumatic disease. It affects 0.5 to 1% of the population in developed countries and is characterized by chronic and destructive inflammation of the joints. It is accompanied by the production of anti-immunoglobulin antibodies, called "rheumatoid factors," as well as anti-citrullinated peptide / protein antibodies (PCAs). Antiphospholipid antibodies (APAs) often appear years before the onset of the disease (Rantapää-Dahlqvist S, et al. Arthritis Rheum. 2003 Oct;48(10):2741-9; Nielen MM, et al. Arthritis Rheum. 2004 Feb;50(2):380-6) and are present in 70 to 80% of established rheumatoid arthritis (RA) cases (De Rycke L, et al. Ann. Rheum. Dis. 2004 63:12, 1587-1593, 2004). Due to their high specificity and their presence at the beginning of the disease, they have significant diagnostic value. Often associated with a more severe course, they also have some prognostic value.Finally, they also have predictive value, as they can predict the progression of undifferentiated arthritis to rheumatoid arthritis (RA). Anti-PCA antibodies can be detected using various ELISA (Enzyme-Linked Immunosorbent Assay) tests. The tests commonly used in medical practice are commercial anti-CCP (anti-Cyclic citrullinated peptide) tests. The test developed and used by the inventors is the AhFibA (Anti-human Fibrinogen Antibodies) test, which detects antibodies that recognize citrullinated fibrinogen. The results of the AhFibA and anti-CCP tests are largely overlapping but, however, discordant in 5 to 10% of patients.

[0003] As soon as the disease is diagnosed, various treatments, some more specific than others, are prescribed to reduce the risk of progression to irreversible joint damage. After corticosteroids, immunosuppressant drugs have become the first-line treatment prescribed to the majority of patients. In cases resistant to these treatments, various biologic therapies such as anti-TNF-alpha, anti-IL6R, and anti-CD20 are initiated, sometimes successively. All these treatments must be continued chronically, are not always effective, or are only effective for a limited time, and, moreover, all present side effects of varying severity. Finally, in some patients, despite the implementation of this therapeutic arsenal, the disease persists and progresses.

[0004] It therefore remains necessary to find new ways to treat RA that are still effective and do not cause debilitating side effects for the patient.

[0005] In recent years, the role of citrullinated aminoglycosides (CIAGs) in triggering and maintaining arthritis, through the formation of immune complexes with citrullinated fibrinogen present in patients' joints, as demonstrated by the inventors, has been widely recognized by the international community. These immune complexes induce, via various effector mechanisms, the secretion of pro-inflammatory cytokines, notably TNF-alpha, which cause and maintain joint inflammation and CIAG synthesis. FR2908134 discloses citrullinated peptides and their use as a drug for the treatment of rheumatoid arthritis. Summary of the invention

[0006] The inventors identified immunodominant citrullinated peptides from citrullinated human fibrin. These peptides are preferential targets of anti-citrullinated proteins (ACPs) and therefore capable of binding to them specifically. The inventors conceived the idea of ​​using these peptides to rid patients' bodies of pathogenic ACPs. They found that by using a column in which specific citrullinated peptides were immobilized, they achieved ACP clearance rates from patient plasma approaching 100%. They then designed a new device for treating autoimmune diseases with anti-citrullinated protein autoantibodies, such as rheumatoid arthritis.

[0007] An object of the present invention therefore relates to an apheresis column loaded with a solid support comprising a composition which includes at least one peptide selected from the group consisting of: The peptide, designated α171-185 Cit, has the amino acid sequence VDIDIKIX 1 SCX 2 GSCS (SEQ ID NO: 8) where X1 and X2 each represent a citrullyl residue; the peptide, designated α621-635 Cit, has the amino acid sequence X1 GHAKSX 2 PVX 3 GIHTS (SEQ ID NO: 12) where X1, X2, and X3 each represent a citrullyl residue; the peptide, designated β60-74 Cit-NH2, has the amino acid sequence X1 PAPPPISGGGYX 2 AX 3 (SEQ ID NO: 15) where X1 and X2 each represent a citrullyl residue and X3 represents a citrullyl derivative with a carboxamide group in place of the carboxyl group; and the peptide, designated Ac -α36-50 Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 6) where the N-terminal G residue is acetylated and where X 1 and X 2 each represent a citrullyl residue and / or the peptide, called α36-50 Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 5) where X 1 and X 2 each represent a citrullyl residue, whose peptide(s) are immobilized directly or indirectly on the support.

[0008] The present invention also relates to an apheresis column loaded with a solid support comprising a composition which includes at least two peptides selected from the group consisting of: The peptide, designated α171-185 Cit, has the amino acid sequence VDIDIKIX 1 SCX 2 GSCS (SEQ ID NO: 8) where X1 and X2 each represent a citrullyl residue; the peptide, designated α621-635 Cit, has the amino acid sequence X1 GHAKSX 2 PVX 3 GIHTS (SEQ ID NO: 12) where X1, X2, and X3 each represent a citrullyl residue; the peptide, designated β60-74 Cit-NH2, has the amino acid sequence X1 PAPPPISGGGYX 2 AX 3 (SEQ ID NO: 15) where X1 and X2 each represent a citrullyl residue and X3 represents a citrullyl derivative with a carboxamide group in place of the carboxyl group; and the peptide, designated Ac -α36-50 Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 6) where the N-terminal G residue is acetylated and where X 1 and X 2 each represent a citrullyl residue and / or the peptide, called α36-50 Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 5) where X 1 and X 2 each represent a citrullyl residue.

[0009] The present invention also relates to the use of a composition comprising at least one citrullinated peptide selected from the group consisting of peptides having sequences SEQ ID NO: 8, SEQ ID NO: 12, SEQ ID NO: 15, SEQ ID NO: 6 and SEQ ID NO: 5 in the manufacture of an apheresis column, preferably an apheresis column intended for the treatment of an autoimmune disease with anti-citrullinated protein autoantibodies such as rheumatoid arthritis.

[0010] The present invention also relates to a composition comprising at least one citrullinated peptide selected from the group consisting of peptides having sequences SEQ ID NO: 8, SEQ ID NO: 12, SEQ ID NO: 15, SEQ ID NO: 6 and SEQ ID NO: 5 for its use in an autoimmune disease with anti-citrullinated protein autoantibodies and in particular rheumatoid arthritis, where the peptide(s) of the composition are immobilized on a solid support loaded in an apheresis column. Detailed description of the invention Composition

[0011] The present invention relates to an apheresis column loaded with a solid support comprising a composition including at least one peptide selected from the group consisting of: The peptide, called α171-185 Cit, has the amino acid sequence VDIDIKIX 1 SCX 2 GSCS (SEQ ID NO: 8) where X1 and X2 each represent a citrullyl residue; the peptide, called α621-635 Cit, has the amino acid sequence X1 GHAKSX 2 PVX 3 GIHTS (SEQ ID NO: 12) where X1, X2, and X3 each represent a citrullyl residue; the peptide, called β60-74 Cit-NH2, has the amino acid sequence X1 PAPPPISGGGYX 2 AX 3 (SEQ ID NO: 15) where X1 and X2 each represent a citrullyl residue and X3 represents a citrullyl derivative with a carboxamide group in place of the carboxyl group; and the peptide, called Ac -α36-50Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 6) where the N-terminal G residue is acetylated and where X 1 and X 2 each represent a citrullyl residue and / or the peptide, called α36-50 Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 5) where X 1 and X 2 each represent a citrullyl residue.

[0012] The different sequences are summarized in Table 1 below. [Table 1] Protein / peptide Amino acid sequence SEQ ID NO Human fibrinogen α chain (NP_068657) (AAI01936) 1 Human fibrinogen beta chain (AAA18024) 2 α36-50 GPRWVERHQSACKDS 3 Ac -α36-50 GPRVVERHQSACKDS where G in the N-terminus is acetylated 4 α36-50 Cit GPX 1 VVEX 2 HQSACKDS where X 1 and X 2 each represent a citrullyl residue 5 Ac- α36-50Cit GPX 1 VVEX 2 HQSACKDS where G at the N-terminus is acetylated and where X 1 and X 2 each represent a citrullyl residue, 6 α171-185 VDIDIKIRSCRGSCS 7 α171-185 Cit VDIDIKIX 1 SCX 2 GSCS where X1 and X2 each represent a citrullyl residue 8 α501-515 SGIGTLDGFRHRHPD 9 α501-515 Cit SGIGTLDGFX 1 HX 2 HPD where X 1 and X 2 each represent a citrullyl residue 10 α621-635 RGHAKSRPVRGIHTS 11 α621-635 Cit X1 GHAKSX2 PVX3 GIHTS where X1, X2 and X3 each represent a citrullyl residue 12 β60-74 RPAPPPISGGGYRAR 13 β60-74 Cit X1 PAPPPISGGGYX2 AX3 where X1, X2 and X3 each represent a citrullyl residue 14 β60-74 Cit-NH2 X 1 PAPPPISGGGYX 2 AX 3 where X 1 and X 2 each represent a citrullyl residue and X 3 represents a citrullyl derivative with a carboxamide group (CONH 2 ) in place of the terminal carboxyl group (COOH). 15

[0013] The composition also includes derivatives or fragments of the peptides defined above. These peptide derivatives may, for example, bear modifications designed to facilitate their synthesis and / or improve their stability.

[0014] Examples of such derivatives include peptides containing amino acids whose carboxyl groups are esterified or transformed into amide groups. In the case of the β60-74 Cit-NH2 peptide, a derivative of this peptide could be the so-called β60-74 Cit peptide with amino acid sequences SEQ ID NO: 14, in which the C-terminal citrullyl residue is not amidated.

[0015] Peptide derivatives may also include peptides including amino acids in which an amino group is, for example, methylated, homocitrullinated, carbamylated or acetylated.

[0016] An example is the peptide Ac-α36-50 Cit, which is an N-terminal acetylated derivative of the peptide α36-50 Cit. The peptides α36-50 Cit and Ac-α36-50 Cit can be used as variants of each other in the present invention. However, the peptide Ac-α36-50 Cit is preferred.

[0017] The composition may include at least 1, 2, 3, 4 or 5 peptides selected from the group consisting of α171-185 Cit, α621-635 Cit, β60-74 Cit-NH2 and Ac-α36-50Cit and / or α36-50 Cit, or derivatives or fragments thereof.

[0018] Preferably, the composition according to the invention may comprise at least 1, 2, 3 or 4 peptides selected from the group consisting of Ac-α36-50Cit, α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2 or derivatives or fragments thereof.

[0019] The composition may include 1, 2, 3, 4 or 5 peptides selected from the group consisting of α171-185 Cit, α621-635 Cit, β60-74 Cit-NH2 and α36-50 Cit and / or Ac-α36-50 Cit, or derivatives or fragments thereof.

[0020] Preferably, the composition comprises 1, 2, 3, 4 or 5 peptides selected from the group consisting of Ac-α36-50 Cit, α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2 or derivatives or fragments thereof.

[0021] Preferably, the composition includes the 4 peptides: Ac-α36-50 Cit, α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2.

[0022] Even more preferably, the composition consists of Ac-α36-50 Cit, α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2.

[0023] The composition may also consist of 1, 2, 3 or 4 peptides chosen from the group consisting of Ac-α36-50 Cit, α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2 or derivatives or fragments thereof.

[0024] The composition may further include the peptide, referred to as α501-515 Cit, having the amino acid sequence SGIGTLDGFX 1 HX 2 HPD (SEQ ID NO: 10) where X1 and X2 each represent a citrullyl residue or derivatives or fragments thereof. Also, in one embodiment, the composition comprises and / or consists of at least 1, 2, 3, 4, 5, or 6 peptides, or 1, 2, 3, 4, 5, or 6 peptides selected from the group consisting of Ac-α36-50 Cit and / or α36-50Cit, α171-185 Cit, α621-635 Ci, β60-74 Cit-NH2, and α501-515 Cit, or derivatives or fragments thereof.

[0025] The reactivity profile of AAPCs differs between patients. Thus, the inventors showed that most of the plasmas tested were reactive to the peptide(s) β60-74 Cit-NH2 and / or Ac-α36-50 Cit.

[0026] Also, according to one embodiment, the composition includes at least the peptides β60-74 Cit-NH2 and / or Ac-α36-50 Cit (or alternatively α36-50 Cit).

[0027] However, some plasmas are not reactive to these peptides but are reactive to others. Therefore, it is preferable to combine as many peptides as possible to cover the widest range of plasma reactivity profiles.

[0028] Preferably, the composition therefore includes at least the peptides α171-185 Cit, α621-635 Cit, β60-74 Cit-NH2 and Ac-α36-50 Cit (or alternatively α36-50 Cit). More preferably it includes the peptides α171-185 Cit, α621-635 Cit, β60-74 Cit-NH2 and Ac-α36-50 Cit (or alternatively α36-50 Cit) and optionally the peptide α501-515 Cit.

[0029] The present invention relates to the use of the composition as defined above in the manufacture of an apheresis column, preferably an apheresis column intended for the treatment or prevention of any autoimmune disease with anti-citrullinated protein autoantibodies. The autoimmune disease with anti-citrullinated protein autoantibodies may be selected from the group consisting of Sjögren's syndrome, juvenile idiopathic arthritis, and rheumatoid arthritis. The autoimmune disease with anti-citrullinated protein autoantibodies is preferably rheumatoid arthritis. Apheresis column

[0030] The present invention also relates to an apheresis column loaded with a solid support comprising the composition as defined above, of which the peptide(s) are immobilized directly or indirectly on the support.

[0031] The term "loaded" means that the column carries or contains the solid support in such a way that a liquid such as blood or a blood product can flow through the column in contact with the solid support.

[0032] Generally speaking, apheresis is a technique in which a liquid is circulated ex vivo(Extracorporeally) blood or a blood product from a subject is modified by a medical device through the addition, removal, and / or replacement of component(s) before being reinjected into the subject. More specifically, this is called plasmapheresis when the product treated by apheresis is plasma. Plasmapheresis has notably been described to desensitize ABO-incompatible kidney transplant candidates; these candidates possessed anti-ABO antibodies that compromised the tolerance of the ABO-incompatible graft (L. Rostaing et al., Treatment of large plasma volumes using specific immunoadsorption to desensitize ABO-incompatible kidney transplant candidates, J. Nephropathology. 2016;5(3):90-97).

[0033] Thus, the apheresis column according to the invention makes it possible to remove AAPCs from the blood or a blood product such as plasma from a subject.

[0034] In a conventional apheresis method, blood is drawn directly from a vein or artery of the patient. In some embodiments, the blood is separated into one or more blood products (for example, a solid fraction comprising red and white blood cells and platelets, and a liquid fraction such as plasma). A component (for example, autoantibodies) is removed from one of the blood products (for example, the plasma). Optionally, the blood products are combined. The blood can then be reinjected into the patient's vein or artery.

[0035] The present invention relates in particular to an apheresis column loaded with a solid support comprising at least 1, at least 2, at least 3, at least 4 or at least 5 or 1, 2, 3, 4 or 5 peptide(s) selected from the group consisting of: The peptide, called α171-185 Cit, has the amino acid sequence VDIDIKIX 1 SCX 2 GSCS (SEQ ID NO: 8) where X1 and X2 each represent a citrullyl residue; the peptide, called α621-635 Cit, has the amino acid sequence X1 GHAKSX 2 PVX 3 GIHTS (SEQ ID NO: 12) where X1, X2, and X3 each represent a citrullyl residue; the peptide, called β60-74 Cit-NH2, has the amino acid sequence X1 PAPPPISGGGYX 2 AX 3 (SEQ ID NO: 15) where X1 and X2 each represent a citrullyl residue and X3 represents a citrullyl derivative with a carboxamide group in place of the carboxyl group; and the peptide, called Ac -α36-50 Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 6) where the N-terminal G residue is acetylated and where X 1 and X 2 each represent a citrullyl residue and / or the peptide, called α36-50 Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 5) where X 1 and X 2 each represent a citrullyl residue, or derivatives or fragments thereof, of which the peptide(s) are immobilized directly or indirectly on the support.

[0036] The solid support of the apheresis column may also include the peptide, called α501-515 Cit, having the amino acid sequence SGIGTLDGFX 1 HX 2 HPD (SEQ ID NO: 10) where X 1 and X 2 each represent a citrullyl residue or derivatives or fragments thereof.

[0037] Preferably, the solid support of the apheresis column includes the 4 peptides: Ac-α36-50 Cit, α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2.

[0038] The present invention also relates to an assembly of columns comprising at least 1, at least 2, at least 3, at least 4 or at least 5, or 1, 2, 3, 4 or 5 apheresis columns, each apheresis column loaded with a solid support comprising a peptide selected from the group consisting of: The peptide, called α171-185 Cit, has the amino acid sequence VDIDIKIX 1 SCX 2 GSCS (SEQ ID NO: 8) where X1 and X2 each represent a citrullyl residue; the peptide, called α621-635 Cit, has the amino acid sequence X1 GHAKSX 2 PVX 3 GIHTS (SEQ ID NO: 12) where X1, X2, and X3 each represent a citrullyl residue; the peptide, called β60-74 Cit-NH2, has the amino acid sequence X1 PAPPPISGGGYX 2 AX 3 (SEQ ID NO: 15) where X1 and X2 each represent a citrullyl residue and X3 represents a citrullyl derivative with a carboxamide group in place of the carboxyl group; and the peptide, called Ac -α36-50Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 6) where the N-terminal G residue is acetylated and where X1 and X2 each represent a citrullyl residue and / or -the peptide, called α36-50 Cit, having the amino acid sequence GPX 1 VVEX 2 HQSACKDS (SEQ ID NO: 5) where X1 and X2 each represent a citrullyl residue or derivatives or fragments thereof, of which the peptide(s) are immobilized directly or indirectly on the support.

[0039] In one embodiment, the apheresis column assembly comprises four apheresis columns, each loaded with a solid support comprising, respectively, the Ac-α36-50 Cit peptide, the α171-185 Cit peptide, the α621-635 Cit peptide, and the β60-74 Cit-NH2 peptide. In one embodiment, the apheresis column assembly may further comprise the α501-515 Cit peptide.

[0040] The peptides according to the invention are preferably synthetic peptides.

[0041] The peptides according to the invention can be biotinylated by methods known in the art. In specific embodiments, the peptide(s) are biotinylated via a spacer group. The spacer separates the citrullinated peptide from the solid support, facilitating its recognition by the aminoglycosides. Any suitable spacer that maintains the binding properties of the peptide according to the invention to the aminoglycosides can be used in the invention. In specific embodiments, the spacer is polyethylene glycol (PEG) or aminohexanoic acid (AHX).

[0042] The solid support for immobilizing the peptides of the invention is known in the art. The solid support is preferably made of a material that does not activate blood cells. It is preferable to use a support treated with an anticoagulant agent, for example, a heparinized support. Alternatively, the patient's blood can be treated with an anticoagulant such as heparin before application to the support.

[0043] The solid support can be made of polymers such as polysaccharides, preferably with a high molecular weight, for example, 100 kDa or more, such as agarose or cellulose. The polysaccharide in the support can be cross-linked or non-cross-linked. Other polymers such as carboxylated polystyrene can also be used. Solid supports can be in the form of magnetic beads or glass.

[0044] The solid support can be porous or non-porous. It can be in the form of particles that may be spherical or irregular. The average particle size can range, for example, from 10 µm to 2 mm, preferably from 30 µm to 100 µm.

[0045] The support can be treated with an anticoagulant agent such as heparin.

[0046] Methods for immobilizing peptides on a solid support are known. Thus, a peptide according to the invention can be immobilized on the support directly or indirectly. Advantageously, the peptide(s) selected from the group consisting of α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2, and optionally the peptide α501-515 Cit, are immobilized directly or indirectly on the solid support by their N-terminal end and / or the peptide(s) Ac-α36-50Cit and / or α36-50 Cit are immobilized directly or indirectly to the solid support by their C-terminal end.

[0047] Preferably, the peptides in α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2, and optionally the peptide α501-515 Cit, are immobilized directly or indirectly on the solid support by their N-terminal end and the peptide Ac-α36-50 Cit (or alternatively α36-50 Cit) is immobilized directly or indirectly on the solid support by its C-terminal end.

[0048] Indirect immobilization can be achieved using a suitable compound, referred to as an intermediate compound. A preferred method for the indirect immobilization of a peptide relies on the interaction between biotin and an intermediate compound such as avidin or streptavidin. Thus, biotinylation of the peptide and the use of avidin or streptavidin immobilized on the solid support allow for reliable attachment of the peptides to the solid support. Specifically, the method may include supplying the peptide according to the invention in biotinylated form, supplying a solid support having streptavidin (or alternatively avidin) immobilized on its surface, contacting the support with an aqueous solution of the biotinylated peptide(s) according to the invention, and rinsing the support with an aqueous solvent.

[0049] In a preferred embodiment, the solid support comprises streptavidin or avidin immobilized directly onto it. The peptides Ac-α36-50 Cit (or alternatively α36-50 Cit), α171-185 Cit, α621-635 Cit, and β60-74 Cit-NH2, and optionally the peptide α501-515 Cit, are biotinylated and linked to the streptavidin (or avidin). The citrullinated peptides may be biotinylated at the C-terminus or the N-terminus. Preferably, the peptides α171-185 Cit, α621-635 Cit, and β60-74 Cit-NH2, and optionally the peptide α501-515 Cit, are N-terminally biotinylated, and the peptide Ac-α36-50 Cit (or alternatively α36-50 Cit) is C-terminally biotinylated. The peptides Ac-α36-50 Cit (or alternatively α36-50 Cit), α171-185 Cit, α621-635 Cit, and β60-74 Cit-NH2, and optionally the peptide α501-515 Cit, may be biotinylated via a spacer group, for example, aminohexanoic acid (AHX) or alternatively PEG.Besides the affinity between biotin and avidin or streptavidin, antibody-antigen interactions can also be used for the indirect immobilization of peptides on a support.

[0050] Alternatively, peptides can be immobilized directly onto a solid support using bioconjugation techniques, such as direct immobilization of peptides onto cyanogen bromide-activated solid supports via amino functionalities in the peptide's primary sequence, or direct immobilization of peptides onto solid supports containing epoxides activated via carboxyl functionalities in the peptide's primary sequence. Alternatively, "click" chemistry can be used to immobilize peptides onto solid supports, with both the peptide and the support modified with appropriate mutually reactive chemical functionalities (azides and alkynes). In other embodiments, Staudinger ligation chemistry can be used to immobilize appropriately modified peptides onto suitably derived solid supports. Treatment method

[0051] The present invention also relates to a composition as defined above for use in an apheresis column, preferably an apheresis column intended for the treatment or prevention of an autoimmune disease with anti-citrullinated protein autoantibodies.

[0052] An autoimmune disease with anti-citrullinated protein autoantibodies is defined as a disease in which an individual's body develops antibodies against citrullinated proteins of the same individual. Autoimmune diseases with anti-citrullinated protein autoantibodies can be selected, in particular, from the group consisting of Sjögren's syndrome, juvenile idiopathic arthritis, and rheumatoid arthritis. Preferably, rheumatoid arthritis is the autoimmune disease with anti-citrullinated protein autoantibodies.

[0053] The present invention also relates to a composition as defined above for use in the treatment or prevention, preferably by apheresis, of an autoimmune disease with anti-citrullinated protein autoantibodies, preferably rheumatoid arthritis, where the peptide(s) of the composition are immobilized on a solid support.

[0054] Preferably, the solid support is loaded onto an apheresis column.

[0055] Preferably, the treatment or prevention of an autoimmune disease with anti-citrullinated protein autoantibodies includes treatment in vitro blood from a patient with said autoimmune disease with an apheresis column loaded with a solid support comprising the composition as defined above.

[0056] The present invention also relates to a composition as defined above for use in an apheresis column, preferably an apheresis column intended for the treatment of rheumatoid arthritis.

[0057] The present invention also relates to the use of an apheresis column as defined above, in particular the use of this apheresis column in the treatment or prevention of an autoimmune disease with anti-citrullinated protein autoantibodies and more preferably of rheumatoid arthritis.

[0058] This application also describes a method of treatment or prevention by apheresis, in a subject requiring it, in which an apheresis column as defined above is used.

[0059] This application also describes a method for purifying anti-citrullinated peptide antibodies (ACPA) from the blood or a blood product of a subject, comprising: the provision of an apheresis column as defined above, bringing this column into contact with the subject's blood or blood product in such a way as to deplete the subject's blood or blood product of AAPCs.

[0060] The subject dealt with by the invention is preferably a mammal, more preferably a human being.

[0061] According to a preferred embodiment, the subject treated according to the invention is a subject with a severe form of rheumatoid arthritis. The subject may also be a subject with high titers of anti-citrullinated protein / peptide antibodies (ACPA). By subject with high ACPA titers, we mean subjects with an OD ≥0.9 when measuring ACPA by an AhFibA test. An OD ≥0.9 in the AhFibA test corresponds to the 40% of rheumatoid arthritis patients with the highest ACPA levels ( figure 1 ).

[0062] The invention will be further illustrated by the following figures and examples. Brief description of the figures

[0063] [ Fig. 1 ] Distribution of AhFibA titers determined from serum samples from 202 AhFibA-positive rheumatoid arthritis patients. Optical density (OD corrected defines titer) threshold values ​​separating quartiles are noted. [ Fig. 2 ]Percentage of sera reactive in ELISA with respect to the 4 peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit and α171-185 Cit, among 202 sera from patients with rheumatoid arthritis and AhFibA-positive. [ Fig. 3 ] Distribution according to their reactivity profile towards the 4 peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit and α171-185 Cit, of 202 AhFibA-positive sera from patients with rheumatoid arthritis. [ Fig. 4 ] Correlations between antibody titers directed against the four peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit, and α171-185 Cit, for the 202 sera tested. At each intersection, Spearman's rank correlation coefficient and the p-value of the correlation are noted. [ Fig. 5 ]Two types of chromatography were used to simultaneously purify the antibodies anti-β60-74 Cit-NH2, ant-Ac-α36-50 Cit, anti-α621-635 Cit and anti-α171-185 Cit: either 4 monopeptide columns linked in series (A), or a single column containing a balanced mixture of the 4 monopeptide matrices (B). [ Fig. 6 ]Chromatograms were performed on pools of serum from RA patients, either on the four columns mounted in series, loaded respectively with the peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit, and α171-185 Cit (left histograms), or on a single column containing a mixture of the four peptides (right histograms). Pool 97 consisted of 97 serum samples of the same volume from patients with increasing AAPC titers representative of those found in a RA patient population; Pool 27 consisted of 27 serum samples of the same volume from patients with high AAPC titers (OD ≥ 1.5). The pools (starting serum: SD - black bar), the unretained fractions (FNR - white bar) and the eluates (grey bar) were analyzed by AhFibA ELISA (n≥4), the results are expressed in corrected OD. [ Fig. 7 ]Method for calculating the clearance rate of serum after chromatography. The retained fraction is analyzed by AhFibA ELISA, and the resulting OD is compared to those resulting from the progressive dilution of the initial serum (reference curve). The ratio of the dilutions determines the clearance rate. In this example, 1 / 50 vs. 1 / 400: 8 times fewer antibodies in the NRF, i.e., 12.5% ​​of the antibodies present in the initial serum. Therefore, 87.5% of the antibodies were cleared. [ Fig. 8 ]Calculation of the clearance rate of aminoglycosides (AAPCs) in a serum sample (no. 41) after chromatography on four columns loaded respectively with the peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit, and α171-185 Cit, mounted in series. The retained fraction was analyzed by AhFibA ELISA, and the resulting OD was compared to those resulting from the dilution of the initial serum (reference curve). The ratio of the dilutions 1 / 50 vs. 1 / 1718 allows the clearance rate to be determined: 98% of the AAPCs were cleared. [ Fig. 9 ]Chromatography was performed using Pool 97, Pool 27, and 10 individual sera from RA patients on four columns loaded with the peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit, and α171-185 Cit, respectively, and mounted in series. For each pool and individual serum, the starting sera (SD - black bars) and the retained fractions (FNR - white bars) were analyzed by AhFibA ELISA (n≥4): their titers are expressed as corrected OD. The clearance rates (%) were calculated for all chromatographed samples (gray bars). [ Fig. 10 ]Correlation between the AAPC titer (AhFibA ELISA) of the starting serum (before chromatography) and the purification rate obtained after chromatography on the 4 columns in series, loaded respectively with the peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit and α171-185 Cit. The correlation is highly significant (p=0.014): the purification is more efficient when the sera have a high AAPC titer. Examples Materials and methods Serum samples

[0064] Serum samples were obtained from 202 patients with rheumatoid arthritis who tested positive for the "AhFibA" ELISA test, which detects and titrates AAPC using citrullinated human fibrinogen as an immunosorbent. Synthetic peptides

[0065] Peptides corresponding to residues 36 to 50, 171 to 185 and 621 to 635 of the α chain of human fibrinogen (NP_068657 - Isoform 2) and residues 60 to 74 of its β chain (AAA18024) were synthesized.

[0066] The peptides were synthesized in their citrullinated form (by systematic substitution of all arginyl residues with a citrullyl residue) or in their native non-citrullinated form (arginyl residues). The β60-74 and β60-74 Cit peptides were synthesized with a carboxamide in place of the terminal carboxyl group to obtain the β60-74 NH2 and β60-74 Cit-NH2 peptides.

[0067] The peptides α36-50, and α36-50 Cit were synthesized with an acetyl on the N-terminal side to obtain the peptides Ac-α36-50 and Ac-α36-50 Cit.

[0068] The peptides were biotinylated at the C-terminus of an aminohexanoyl spacer for the Ac-α36-50Cit peptide and at the N-terminus of the same spacer, for the α171-185 Cit, α621-635 Cit and β60-74 Cit-NH2 peptides. ELISA tests

[0069] Antigens or immunosorbents (citrullinated fibrinogen or citrullinated and non-citrullinated peptides), solubilized in PBS (1.5mM KH 2 PO4 SIGMA 795488; 7mM K 2 HPO 4 SIGMA P3786; 0.15M NaCl SIGMA 31434; pH 7.2) at concentrations of 5 and 10µg / mL, are adsorbed into the wells of microtiter plates (MAXISORP NUNC 2023-09) by incubation overnight at 4°C, or used in an avidin-biotin system: avidin 5µg / mL in PBS incubated overnight at 4°C then, after washing, biotinylated peptide at 10µg / mL in PBS, incubated for 1h at 4°C. After blocking in 2% BSA PBS (Sigma A3059) for 1 hour at 4°C and washing in 0.1% Tween®< 20 PBS (SIGMA P1379), the samples to be tested are loaded, diluted 1 / 50 (1 / 100 in the Avidine-Biotin system) or equivalent corrected dilutions, into 2% BSA PBS with 2MNaCl. After washing, the secondary antibody (goat IgG against human IgG Fc fragment coupled to horseradish peroxidase (Horseradish Peroxidase; Southern Biotech 2040-05)) is incubated for 1 hour at 4°C.After washing, development was performed with a solution of orthophenylene diamine dihydrochloride (Sigma P2788), 0.03% H₂O₂ (Sigma H1009), in citrate / H₃PO₄ buffer pH 5 (0.05 M citric acid Sigma C0759; 0.1 M Na₂HPO₄ Prolabo 28026292) for 5 minutes at room temperature, then stopped with a 6N H₂SO₄ solution (Sigma 07208). The optical density (OD) was read at 492 nm using a Multiskan Fc plate reader (ThermoScientific).

[0070] Conditions for performing ELISA tests [Table 2] Concentration of immunosorbent Sample dilution Secondary antibody dilution AhFibA 5 µg / mL 1 / 50 1 / 15000 Anti-β60-74 10 µg / mL 1 / 50 1 / 2500 Anti-α36-50 10 µg / mL 1 / 50 1 / 2500 Anti-α621-635 10 µg / mL 1 / 50 1 / 2500 Anti-α171-185biot 10 µg / mL 1 / 100 1 / 2500

[0071] Expression of ELISA test results: for peptides, the change in OD between the reactivity of the citrullinated peptide and the reactivity of the non-citrullinated peptide is taken into account, each reactivity being previously subtracted from its blank (OD in the absence of a sample). This gives the following formula: ΔDO = DO cit − DO blanc cit − DO noncit − DO blanc noncit

[0072] Note that for fibrinogen, the non-citrullinated form never exhibits any reactivity, so it is not necessary to evaluate it. For the AhFibA test, the OD value is therefore simply subtracted from its blank (OD in the absence of a sample).

[0073] The ΔDO for peptides and the DO for citrullinated fibrinogen are considered to express the titer of the samples tested.

[0074] Inter-assay variations are corrected using a range of dilutions from a pool of patient sera tested in each microtiter plate. This reference pool, named P97, was created from 97 samples of the same volume from 97 patients with high AAPC titers. The OD values ​​of this internal range are compared to reference values, where each point in the range is the average of 30 determinations previously performed from the same pool. The least-squares method then yields a correction factor, which is applied to all OD values ​​obtained on the corresponding plate. This corrected OD corresponds to the titer of the sample. DO corrig é e = ∑ X ∗ Y ∗ ∑ X 2 ∗ DO

[0075] With: X internal values, Y: reference values Construction of chromatography columns

[0076] Four 1 mL HiTrap Streptavidine HP columns (GE; 29-0513-24) are independently loaded with each of the four biotinylated peptides: β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit, and α171-185 Cit. After equilibration of the column in PBS, a column volume (1 mL) of saturated biotinylated peptide solution is injected at 0.2 mL / min, incubated for 15 min, then collected from the column outlet and lyophilized.

[0077] The amount of peptide remaining from the lyophilized fraction is quantified by high-performance liquid chromatography (UPLC); this amount is subtracted from the starting amount to estimate the amount of peptide bound in the column. [Table 3] Peptide Quantity of bound peptide (µmol) β60-74 Cit-NH2 0,83 Ac-α36-50 Cit 1,1 α621-635 Cit 0,87 α171-185 Cit 1,7

[0078] Four columns were loaded with the respective amounts of peptides shown in the table above.

[0079] After use for serial purification tests, the 4 column matrices loaded with peptides β60-74 Cit-NH2, α36 Ac-50 Cit, α621-635 Cit and α171-185 Cit were recovered and mixed and then conditioned in a 10 mL empty column (GE, C10 / 10) with volume adapter (GE, AC10), in order to form a column loaded with the 4 mixed peptides with a final volume of 4 mL. Purge

[0080] Two representative pools, P97 (previously described) and P27, a pool of 27 patient sera with an AAPC titer greater than 1.5, as well as individual sera chosen based on their reactivity on the 4 peptides β60-74 Cit-NH2, α36 Ac-50 Cit, α621-635 Cit and α171-185 Cit, were diluted 1 / 4 in PBS before chromatography.

[0081] After equilibrating the columns in PBS at a flow rate of 0.5 mL / min, a 6 mL volume of pool or diluted serum was injected onto the four columns mounted in series in the following order: the column loaded with β60-74 Cit-NH2 peptide, then α36 Ac-50 Cit, α621-635 Cit, and α171-185 Cit, and subsequently onto the column containing all four peptides. The retained fractions were collected. After washing in PBS, the antibodies specific to the four peptides were eluted with 0.2 M glycine-HCl buffer (Invitrogen; 15527-013) pH 3. The eluted fractions were collected and adjusted to pH 7 with 2 M Tris solution (Euromedex; 77-86-1). Starting sera, unretained fractions and eluates were then tested by ELISA after correction of the dilution factor induced by chromatography. Determining the percentage or rate of purification

[0082] For each treated pool or serum, the percentage of purification was calculated from the residual immunoreactivity in AAPC (AhFibA ELISA test) of the unretained, purified fractions, and a range of dilutions of the corresponding starting pool or serum. % é puration = 1 − Dilution SD correspondant à DO FNR Dilution exp é rimentale FNR ∗ 100

[0083] A fictitious example of calculating the percentage of purification is illustrated in the figure 7 To obtain an OD equivalent to that of the rejected fraction (tested at 1 / 50 after correcting for dilution due to chromatography), the starting serum must be diluted at 1 / 400. The concentration of amino acids in the rejected fraction is therefore 8 times lower than that of the starting serum. One-eighth, or 12.5%, of the amino acids in the starting serum are present in the rejected fraction and have therefore not been purified. It can be deduced that seven-eighths of the amino acids in the starting serum, or 87.5%, have been purified: this is the purification rate. Serum sample reactivity profile

[0084] The reactivity profiles of the 202 patient serum samples to citrullinated fibrinogen and the four citrullinated peptides were determined by ELISA in at least two independent assays, each performed in duplicate. For reproducibility, only microtiter plates whose internal range required a correction factor of between 0.5 and 2 relative to the reference range were considered. Results ELISA reactivity profiles of sera from patients with rheumatoid arthritis (RA), with respect to citrullinated fibrinogen (AhFibA test) and with respect to the 4 immunodominant peptides.

[0085] 202 sera from RA patients were tested for AhFibA and then ranked in order of increasing reactivity (OD or titer) ( figure 1 ). Based on their titer, patients were divided into quartiles: low titers (OD = [0.056 - 0.615]), medium titers (OD = [0.615 - 1.161]), high titers (OD = [1.161 - 1.876]) and very high titers (OD ≥ 1.876).

[0086] There figure 2shows the percentages of PR sera that are reactive to each of the 4 peptides: 76% are positive on the β60-74 Cit-NH2 peptide, confirming that it is indeed the most immunodominant.

[0087] The individual reactivity of sera to the 4 peptides is highly variable, allowing for the definition of numerous reactivity profiles. The frequency of the different profiles observed is shown in the figure 3 Of these 202 sera, 78% reacted to at least two of the four peptides, and 95% reacted with at least one of the four peptides. These results show that using a combination of the four peptides in chromatography should allow for targeting a very broad spectrum of patients.

[0088] The search for correlations between serum reactivities towards different peptides revealed significant correlations between the peptides β60-74 Cit-NH2, α621-635 Cit, and α171-185 Cit; these correlations reflect a certain degree of cross-reactivity between these peptides, however, their relative weakness shows that each peptide exhibits its own specific reactivity. In contrast, no correlation was found between α36 Ac-50 Cit and the other three peptides. The two-way table presented in figure 4 The analysis visualizes the Spearman coefficients calculated for all the compared peptide pairs and their significance. These results support the idea that to develop a clearance system targeting all AAPCs for the largest possible number of patients, it is necessary to use all four peptides together. This is therefore the final choice. Purification of serum pools from RA patients on 4 chromatography columns loaded respectively with the peptides β60-74 Cit-NH2, Ac- α36-50 Cit, α621-635 Cit and α171-185 Cit and mounted in series, or on a column containing a mixture of the 4 peptides.

[0089] Two chromatography systems, shown on the figure 5 , were devised to purify patient sera of the AAPC they contain using the 4 peptides. The first system consists of connecting in series 4 columns loaded respectively with one of the 4 peptides β60-74 Cit-NH2 , Ac -α36-50 Cit , α621-635 Cit and α171-185 Cit (A ), the second consists of mixing in equivalent quantities the 4 monopeptide matrices and loading them into a single column (B ).

[0090] The efficacy of the two systems was compared using two pools of sera from RA patients. These conditions were chosen because they are more demanding than those used to test an individual serum. Indeed, in a pool containing numerous sera with different reactivity profiles, the mixture of antigen-specific antibodies (ASAs) is highly polyclonal and composed of ASAs with varying antigenic specificities.

[0091] A first pool, P97, consists of 97 sera with an AhFibA titer distribution representative of that of the general population of RA patients. A second pool, P27, consists of 27 sera from patients with high AhFibA titers (OD >1.5).

[0092] There figure 6 This shows the results of chromatographies performed on these pools, either on the four columns connected in series, loaded respectively with the peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit, and α171-185 Cit, or on a single column containing a mixture of the four peptides. A significant proportion of the amino acids was absorbed by the columns and can be eluted, while another, unabsorbed portion remains in the fraction not retained by the columns. It appears that both chromatography systems are effective in purifying a large portion of the amino acids contained in the pools and that they give very similar results.

[0093] In addition, 10 AAPC positive sera were chosen based on their reactivity profiles towards the 4 peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit and α171-185 Cit (Table 4 below). [Table 4] Serum No. β60-74 Cit-NH2 Ac- α36-50 Cit α621-635 Cit α171-185 Cit n°1 +++ + + n°10 +++ + n°41 +++ ++ n°16 + +++ ++ n°29 + +++ n°25 ++ +++ + n°64 +++ +++ n°22 ++ ++ +++ n°6 +++ +++ + n°17 +++ + +++ +++ (+++: strong positive; ++: medium positive; +: weak positive; empty box: negative)

[0094] These sera, as well as the two pools P97 and P27, were passed through 4 columns in series loaded respectively with the 4 peptides β60-74 Cit-NH2, Ac-α36-50 Cit, α621-635 Cit and α171-185 Cit. For all samples, the AAPC titer decreased sharply after passage through the columns ( figure 9 ).

[0095] An indicator of purification efficiency was needed to compare the results between the different chromatographic methods and the different samples. Since the optical density of a sample analyzed by ELISA is not linearly correlated but exponentially correlated with the amount of AAPC contained in that sample, a simple ratio of optical densities between the titer of the starting serum and that of the fraction not retained on the columns could not be used to calculate a purification rate.

[0096] We therefore used a graphical method which consists of determining, by AhFibA ELISA, the ODs of a series of dilutions of the starting serum, allowing us to establish a reference curve. The OD of the rejected fraction is determined in the same way, after correcting for the dilution factor related to chromatography. The graphical reading on the reference curve indicates which dilution of the starting serum corresponds to the OD of the rejected fraction. The calculated percentage of purification results from the ratio between the experimental dilution of the rejected fraction and the corresponding dilution of the starting serum. This graphical method for determining the purification rate is illustrated in the figure 7 An example of calculating this rate for one of the treated sera (serum no. 41) is shown in the figure 8 .

[0097] The calculated purification rates for the 2 pools and for the 10 individual sera after a single pass through the columns are all greater than 65%, and 7 out of 12, including the 2 pools, are greater than 85%. For serum #41, this rate reaches 98%.

[0098] The relationship between the AAPC titer of the starting serum and the purification rate obtained after passage through the four columns connected in series was studied. Figure 10 This shows that there is a strong and significant correlation (Spearman's r: 0.70; p = 0.014) between these two parameters. The higher the serum antibody titer, the more efficient its clearance. This counterintuitive result stems from the fact that high-titer antibody-drug enzyme (ADPE) has a high affinity and therefore binds more effectively to its antigens.

Claims

1. Apheresis column loaded with a solid support comprising a composition that includes at least one peptide selected from the group consisting of: - the peptide, called α171-185Cit, having the amino acid sequence VDIDIKIX1SCX2GSCS (SEQ ID NO: 8) where X1 and X2 each represent a citrullyl residue, - the peptide, called α621-635Cit, having the amino acid sequence X1GHAKSX2PVX3GIHTS (SEQ ID NO: 12) where X1, X2 and X3 each represent a citrullyl residue, - the peptide, called β60-74Cit-NH2, having the amino acid sequence X1PAPPPISGGGYX2AX3 (SEQ ID NO: 15) wherein X1 and X2 each represent a citrullyl residue and X3 represents a citrullyl derivative with a carboxamide group in place of the carboxyl group, and - the peptide, called Ac-α36-50Cit, having the amino acid sequence GPX1WEX2HQSACKDS (SEQ ID NO: 6) wherein the G residue at the N-terminal is acetylated and wherein X1 and X2 each represent a citrullyl residue and / or the peptide, referred to as α36-50Cit, having the amino acid sequence GPX1VVEX2HQSACKDS (SEQ ID NO: 5) wherein X1 and X2 each represent a citrullyl residue wherein the peptide(s) are immobilized directly or indirectly on the support.

2. Apheresis column according to claim 1, characterized in that it comprises at least 2 peptides selected from the group consisting of α36-50Cit, Ac-α36-50Cit, α171-185Cit, α621-635Cit and β60-74Cit-NH2.

3. Apheresis column according to claim 1 or 2, characterized in that it comprises the peptides α171-185Cit, α621-635Cit, β60-74Cit-NH2and the peptide(s) α36-50Cit and / or Ac-α36-50Cit.

4. Apheresis column according to any one of claims 1 to 3, characterized in that it further comprises the peptide α501-515Cit with the amino acid sequence SGIGTLDGFX1HX2HPD (SEQ ID NO: 10) wherein X1 and X2 each represent a citrullyl residue.

5. Apheresis column according to any one of claims 1 to 4, characterized in that: - the peptide(s) selected from the group consisting of α171-185Cit, α621-635Cit, and β60-74Cit-NH2, and optionally the peptide α501-515Cit, are immobilized directly or indirectly on the solid support by their N-terminal end and / or - the peptide(s) Ac-α36-50Cit and / or α36-50Cit are immobilized directly or indirectly on the solid support by their C-terminal end.

6. Apheresis column according to any of claims 1 to 5, characterized in that the peptide(s) are indirectly immobilized on the solid support by means of one or more intermediate compounds.

7. Apheresis column loaded with a solid support comprising a composition that includes at least two peptides selected from the group consisting of: - the peptide, called α71-185Cit, having the amino acid sequence VDIDIKIX1SCX2GSCS (SEQ ID NO: 8) where X1 and X2 each represent a citrullyl residue, - the peptide, called α621-635Cit, having the amino acid sequence X1GHAKSX2PVX3GIHTS (SEQ ID NO: 12) where X1, X2 and X3 each represent a citrullyl residue, - the peptide, called β60-74Cit-NH2, having the amino acid sequence X1PAPPPISGGGYX2AX3 (SEQ ID NO: 15) wherein X1 and X2 each represent a citrullyl residue and X3 represents a citrullyl derivative with a carboxamide group in place of the carboxyl group, and - the peptide, called Ac-α36-50Cit, having the amino acid sequence GPX1VVEX2HQSACKDS (SEQ ID NO: 6) wherein the G residue at the N-terminal is acetylated and wherein X1 and X2 each represent a citrullyl residue and / or the peptide, referred to as α36-50Cit, having the amino acid sequence GPX1VVEX2HQSACKDS (SEQ ID NO: 5) wherein X1 and X2 each represent a citrullyl residue.

8. Use of a composition comprising at least one citrullinated peptide selected from the group consisting of peptides having sequences SEQ ID NO: 8, SEQ ID NO: 12, SEQ ID NO: 15, SEQ ID NO: 6, and SEQ ID NO: 5 in the manufacture of an apheresis column according to any one of claims 1 to 7.

9. Composition comprising at least one citrullinated peptide selected from the group consisting of peptides having sequences SEQ ID NO: 8, SEQ ID NO: 12, SEQ ID NO:15, SEQ ID NO: 6, and SEQ ID NO: 5 for use in the treatment or prevention of an autoimmune disease with anti-citrullinated protein autoantibodies, wherein the peptide(s) of the composition are immobilized on a solid support loaded into an apheresis column.

10. Composition for use according to claim 9, characterized in that the autoimmune disease with anti-citrullinated protein autoantibodies is selected from the group consisting of Sjögren's syndrome, juvenile idiopathic arthritis, and rheumatoid arthritis, preferably rheumatoid arthritis.

11. Composition for use according to claims 9 or 10, characterized in that the treated subject has a high titer of anti-citrullinated protein autoantibodies (ACPA).