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US-12622946-B2 - HLA-DR/CII peptide complexes for treating arthritis

US12622946B2US 12622946 B2US12622946 B2US 12622946B2US-12622946-B2

Abstract

The present invention relates to HLA-DR/CII peptide complexes comprising a chondroitin-binding peptide at the C-terminal end of the polypeptide comprising the HLA-DR/CII alpha chain and/or the HLA-DR/CII beta chain, wherein the CII peptide is fused to the N-terminus of the HLA-DR/CII alpha chain or the HLA-DR/CII beta chain by a linker peptide, for use in treating chronic inflammatory disease, such as arthritis, in human patients. The lysines in the CII peptide, particularly the first lysine in the CII peptide, may be post-translationally modified. The invention further relates to methods of producing said HLA-DR/CII peptide complexes in mammalian cells.

Inventors

  • Nhu-Nguyen DO
  • Vilma URBONAVICIUTE
  • Sylvia CIENCIALA
  • Rikard Holmdahl
  • Harald Burkhardt

Assignees

  • Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e.V.

Dates

Publication Date
20260512
Application Date
20200807
Priority Date
20190809

Claims (17)

  1. 1 . A method of treating a chronic inflammatory disease in a human patient comprising administering to the patient a composition comprising recombinant HLA-DR/CII peptide complexes comprising (a) an extracellular region of an HLA-DR alpha chain comprising at least an alpha 1 domain; (b) an extracellular region of an HLA-DR beta chain comprising at least a beta 1 domain; (c) a collagen II peptide (CII peptide), optionally fused to the N-terminus of the HLA-DR alpha chain or the HLA-DR beta chain by a linker peptide; and (d) a chondroitin-binding peptide at the C-terminal end of the polypeptide comprising the HLA-DR alpha chain and/or the HLA-DR beta chain, wherein the CII peptide comprises the amino acid sequence selected from the group consisting of AGFKGEQGPKG (SEQ ID NO: 1), AGFKGEQGPXG (SEQ ID NO: 2), AGFKGEXGPKG (SEQ ID NO: 3), AGFKGXQGPKG (SEQ ID NO: 4), AGFKXEQGPKG (SEQ ID NO: 5), AGFKGEXGPXG (SEQ ID NO: 6), AGFKGXQGPXG (SEQ ID NO: 7), and AGFKXEQGPXG (SEQ ID NO: 8), and wherein at least the alpha 1 domain is from DRA*0101, and at least the beta 1 domain is from an HLA-DR allele selected from the group consisting of DRB1*0401, DRB1*0404, DRB1*0405, DRB1*0408, DRB1*0409, DRB1*0101, DRB1*0102, DRB1*1001, DRB1*1402, and DRB1*1303.
  2. 2 . The method according to claim 1 , wherein (a) the chondroitin-binding peptide is in its free form; (b) the recombinant HLA-DR/CII peptide complexes are not multimerized via the chondroitin-binding peptide in the composition; and/or (c) the recombinant HLA-DR/CII peptide complexes are not bound to a further molecule via the chondroitin-binding peptide in the composition.
  3. 3 . The method according to claim 1 , wherein (a) the extracellular region of the HLA-DR alpha chain comprises an alpha 1 domain and an alpha 2 domain; and/or (b) the extracellular region of the HLA-DR beta chain comprises a beta 1 domain and a beta 2 domain.
  4. 4 . The method according to claim 1 , wherein at least the alpha 1 domain is from DRA*0101 and at least the beta 1 domain is from DRB1*0401.
  5. 5 . The method according to claim 1 , wherein the CII peptide is fused to the N-terminus of the HLA-DR alpha chain or the HLA-DR beta chain by a linker peptide.
  6. 6 . The method of claim 5 , wherein the CII peptide is fused to the N-terminus of the HLA-DR beta chain by a linker peptide.
  7. 7 . The method according to claim 1 , wherein the CII peptide comprises the amino acid sequence of AGFKGEQGPKG (SEQ ID NO: 1).
  8. 8 . The method according to claim 1 , wherein the chondroitin-binding peptide comprises 5 to 20 amino acids.
  9. 9 . The method according to claim 1 , wherein the chondroitin-binding peptide is a polyhistidine tag.
  10. 10 . The method of claim 9 , wherein the chondroitin-binding peptide is a hexahistidine tag.
  11. 11 . The method according to claim 1 , wherein the extracellular region of the HLA-DR alpha chain comprising at least an alpha 1 domain and the extracellular region of the HLA-DR beta chain comprising at least a beta 1 domain are expressed as a single fusion polypeptide.
  12. 12 . The method according to claim 1 , wherein the recombinant HLA-DR/CII peptide complexes comprise (a) a first polypeptide comprising the extracellular region of the HLA-DR alpha chain comprising at least an alpha 1 domain; and (b) a second polypeptide comprising the extracellular region of the HLA-DR beta chain comprising at least a beta 1 domain, wherein the HLA-DR alpha chain is fused at its C-terminal end to a first functional domain of a leucine zipper heterodimerisation motif, and the HLA-DR beta chain is fused at its C-terminal end to a second complementary functional domain of a leucine zipper heterodimerisation motif.
  13. 13 . The method according to claim 12 , wherein the first functional domain and the second complementary functional domain are (a) an acidic and a basic leucine zipper heterodimerisation domain; and/or (b) a jun-fos leucine zipper motif.
  14. 14 . The method according to claim 1 , wherein the recombinant HLA-DR/CII peptide complexes comprise a mixture of CII peptides with unmodified and/or one or more post-translationally modified lysine residue(s); wherein (a) the mixture of CII peptides consists of CII peptides with unmodified lysine residues; (b) the mixture of CII peptides consists of CII peptides with the first lysine being hydroxylysine (Hyl); (c) the mixture of CII peptides consists of CII peptides with the first lysine being galactosyl-hydroxylysine; (d) the mixture of CII peptides consists of CII peptides with unmodified lysine residues and CII peptides with the first lysine being galactosyl-hydroxylysine; (e) the mixture of CII peptides comprises CII peptides with the first lysine being galactosyl-hydroxylysine; (f) the mixture of CII peptides comprises CII peptides with unmodified lysine residues and CII peptides with the first lysine being galactosyl-hydroxylysine; (g) the mixture of CII peptides comprises CII peptides with unmodified lysine residues and CII peptides with the first lysine being galactosyl-hydroxylysine and/or hydroxylysine (Hyl); or (h) the mixture of CII peptides comprises CII peptides with unmodified lysine residues and CII peptides with the first lysine being O-glycosylated hydroxylysine and/or hydroxylysine (Hyl); and wherein the optional second lysine, if present, in the post-translationally modified CII peptide is unmodified, hydroxylysine, galactose-hydroxylysine and/or glucosyl-galactosyl-hydroxylysine.
  15. 15 . The method of claim 14 , wherein the optional second lysine, if present, in the post-translationally modified CII peptide is (a) unmodified, hydroxylysine and/or galactose hydroxylysine, or (b) unmodified.
  16. 16 . The method according to claim 14 , wherein the composition does not contain HLA-DR/CII peptide complexes comprising a mixture of CII peptides with a glucosyl-galactosyl-hydroxylysine modification.
  17. 17 . The method according to claim 1 , wherein the chronic inflammatory disease is selected from the group consisting of rheumatoid arthritis, osteoarthritis, psoriatic arthritis, non-radiographic axial spondyloarthritis, ankylosing spondylitis, juvenile idiopathic arthritis, relapsing polychondritis, systemic lupus erythematosus, Lyme disease, Meniere diseases, autoimmune inner ear disease (AIED), and Still's disease.

Description

CROSS REFERENCE TO RELATED APPLICATIONS This application is a national stage application under 35 U.S.C. 371 and claims the benefit of PCT Application No. PCT/EP2020/072280 having an international filing date of 7 Aug. 2020, which designated the United States, and which PCT application claimed the benefit of Europe patent application Ser. No. 19/191,077.7 filed 9 Aug. 2019, the contents of each of which are incorporated herein by reference in their entireties. TECHNICAL FIELD The present invention relates to HLA-DR/CII peptide complexes comprising a chondroitin-binding peptide at the C-terminal end of the polypeptide comprising the HLA-DR alpha chain and/or the HLA-DR beta chain, wherein the CII peptide is fused to the N-terminus of the HLA-DR alpha chain or the HLA-DR beta chain by a linker peptide, for use in treating chronic inflammatory diseases, such as arthritis, in human patients. The lysines in the CII peptide, particularly the first lysine in the CII peptide, may be post-translationally modified. The invention further relates to methods of producing said HLA-DR/CII peptide complexes in mammalian cells. SEQUENCE LISTING The instant application contains a Sequence Listing, which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. Said XML copy, created on Apr. 9, 2025, is named Updated_SL_9Apr2025 and is 37,233 bytes in size. TECHNOLOGICAL BACKGROUND Rheumatoid arthritis (RA) is a common, severe disease representing a major health concern with 4-7 million affected people in Europe. It is caused by an aberrant autoimmune inflammation of joints associated with pain, progressive cartilage and bone destruction leading to functional disability and ultimately immobility/invalidity if not adequately treated. Today's pharmaceutical treatment is initiated immediately upon establishment of the clinical diagnosis and is effective in 60-70% of the cases, but does not achieve cure from the disease. Drug treatment predominantly targets common effector pathways of inflammation thereby causing broad immunosuppressive effects associated with an increased risk for infection. The immunogenetics of RA suggests a key role for aberrant pathways of T-cell activation in the initiation and/or perpetuation of disease. In the T-cell activation process, CD4+ T-cells are engaged by antigenic peptide fragments complexed with human major histocompatibility complex (MHC) class II (such as human leukocyte antigen-DR isotype (HLA-DR)), leading to their activation in the context of co-stimulatory signals provided by professional antigen presenting cells. The strongest evidence supporting a role for CD4+ T cells in disease pathogenesis is the genetic association between RA and certain alleles of the HLA-DRB1 locus coding for an amino acid consensus motif Q/R R/K R A A on the beta-chain of the peptide binding pocket of the MHC class II molecule HLA-DR (amino acid position 70-74, the so called “shared epitope”) (Gregersen P K et al., Arthritis Rheum. 1987; 30:1205-1213). Compelling evidence for a pathogenic role of T cells in RA is further provided by their frequent detectability in inflammatory synovial infiltrates of moderate to severe disease indicating their collaboration with B cells in local immune reactions to promote the maturation of specific autoantibody responses. Moreover, an impaired CD4+CD25 (hi) regulatory T cell (Treg) function has been suggested to be involved in the pathogenesis of RA. Accordingly, the dysregulated chronically activated T cell compartment in RA represents a key target for therapeutic immunomodulatory intervention. RA is today believed to start many years before clinical onset. RA as polygenetic disease with the above mentioned shared-epitope encoding alleles at the HLA-DRB1 locus as the strongest risk factor, develops in respectively predisposed individuals. However, yet ill-defined environmental and/or life style factors (smoking) are also involved in triggering an autoimmune response associated with the generation of antibodies to IgG (rheumatoid factors) and to citrullinated proteins (ACPA) that can persist in arthritis prone but still healthy individuals for a preclinical period of up to two decades. Around clinical onset, an immune response to type II collagen (CII) and to citrullinated CII is detectable (Burkhardt H et al., Eur J Immunol. 2005; 35:1643-52). CII is the major protein component in joint cartilage. RA patients that carry the DRB1*0401 allele (50% of Caucasian RA-patients) have been demonstrated to harbor T cells in their repertoire that specifically respond to a major CII epitope corresponding to the amino acid sequence 259-273 of the triple helical CII region. The T cell determinant critical for activation of the T cell receptor (TCR) has been described to be dependent on the physiologically galactosylated hydroxylysine residue at position 264 (Baecklund J. et al., Proc Natl Acad Sci USA. 2002; 99:9960-5). However, in humans this d