Peptide with reduced dimer formation

Abstract

The present invention relates to peptide which are formulated or engineered to prevent or reduce the formation of dimers.

1 . A composition comprising:
 i) at least one peptide of 9 to 25 amino acids in length wherein the peptide comprises a region comprising at least one T cell epitope; and   ii) at least one agent which inhibits peptide dimer formation which is thioglycerol or thioanisole;   wherein a minimal proportion of the peptide of the composition is present in solution as a dimer.   
     
     
         2 . A composition according to  claim 1 , wherein the proportion of peptide as defined in i) that is present as a dimer in solution in the absence of the agent is at least 0.5%; and/or wherein the epitope is an MEW Class II-binding T cell epitope 
     
     
         3 . A composition according to  claim 2 , wherein the proportion of peptide present as a dimer in solution is measured after the peptide has been in solution for at least 72 hours at about 25° C. and about 60% relative humidity. 
     
     
         4 . A composition according to  claim 1 , wherein less than 5% of the peptide is present in dimeric form in solution. 
     
     
         5 . A composition according to  claim 1 , wherein the peptide has an improved ability to induce tolerance in an individual compared to the dimer form of the peptide. 
     
     
         6 . A composition according to  claim 1 , wherein the native sequence of the region comprises at least one cysteine residue. 
     
     
         7 . A composition according to  claim 1 , wherein the native sequence of the protein from which the region derives comprises approximately 33% cysteine residues; and/or wherein the native sequence of the region comprises one, two, three or more cysteine residues up to a maximum of 25% of the total number of amino acid residues in the peptide. 
     
     
         8 . A composition according to  claim 1 , wherein the peptide does not comprise an epitope capable of cross-linking IgG expressed on the cell surface of B cells or IgE expressed on the surface of mast cells or basophils and/or wherein the region consists entirely of the minimal sequence of the T cell epitope. 
     
     
         9 . A composition according to  claim 1 , wherein the epitope derives from:
 i) an allergen selected from: a plant allergen (particularly a grass allergen), animal dander allergens, a mold or fungal allergen, a dust allergen, an antibiotic or other drug, a stinging insect venom, an environmental allergen or a food allergen; or   ii) an antigen selected from the major antigens associated with Acute disseminated encephalomyelitis (ADEM); Addison's disease; Ankylosing spondylitis; Antiphospholipid antibody syndrome (APS); Aplastic anemia; Autoimmune hepatitis; Autoimmune Oophoritis; Coeliac disease; Crohn's disease; Diabetes mellitus type 1; Gestational pemphigoid; Goodpasture's syndrome; Graves' disease; Guillain-Barré syndrome (GBS); Hashimoto's disease; Idiopathic thrombocytopenic purpura; Kawasaki's Disease; Lupus erythematosus; Multiple sclerosis; Myasthenia gravis; Narcolepsy, Opsoclonus myoclonus syndrome (OMS); Optic neuritis; Ord's thyroiditis; Pemphigus; Pernicious anaemia; Polyarthritis in dogs; Primary biliary cirrhosis; Rheumatoid arthritis; Reiter's syndrome; Sjögren's syndrome; Takayasu's arteritis; Temporal arteritis (also known as “giant cell arteritis”); Warm autoimmune hemolytic anemia; or Wegener's granulomatosis   
     
     
         10 . A composition according to  claim 1 , wherein the epitope derives from: cat dander protein Fel d1; House dust mite proteins Der P1, Der P2 and Der P7; Ragweed protein amb a 1.1, a 1.2, a1.3 or a1.4; Rye grass proteins lol p1 and lol p5; Timothy grass proteins phl p1 and phl p5; Bermuda grass protein Cyn d 5; Alternaria alternate proteins Alt a 1, Alt a 2 and Enolase (Alt a 6); Birch protein Bet v1 and P14; German Cockroach proteins Bla g 1, Bla g 2, Bla g 3, Bla g 4, Bla g 5 and Bla g 6; Mugwort protein Art v 1; Russian thistle protein Sal k 1 and Sal k 2; peanut Ara h1, Ara h2, Ara h3, Ara h4, Ara h5, Ara h6, plant profilins or lipid transfer proteins or a human leukocyte antigen. 
     
     
         11 . A composition according to  claim 1  for use in treating or preventing a disease by tolerisation of an individual to the protein from which the T cell epitope derives. 
     
     
         12 . A composition according to  claim 1  for use in treating or preventing an allergic disease, an autoimmune disease, an alloimmune response or a maternal-foetal immune response by tolerisation, or for use in tolerising an individual to a neoantigen or to a protein which is being provided to the individual in therapy. 
     
     
         13 . A composition according to  claim 12 , wherein the allergic disease or autoimmune disease comprises an immune response to an allergen or antigen as defined in i) or ii) below:
 i) an allergen selected from: a plant allergen (particularly a grass allergen), animal dander allergens, a mold or fungal allergen, a dust allergen, an antibiotic or other drug, a stinging insect venom, an environmental allergen or a food allergen; or   ii) an antigen selected from the major antigens associated with Acute disseminated encephalomyelitis (ADEM); Addison's disease; Ankylosing spondylitis; Antiphospholipid antibody syndrome (APS); Aplastic anemia; Autoimmune hepatitis; Autoimmune Oophoritis; Coeliac disease; Crohn's disease; Diabetes mellitus type 1; Gestational pemphigoid; Goodpasture's syndrome; Graves' disease; Guillain-Barré syndrome (GBS); Hashimoto's disease; Idiopathic thrombocytopenic purpura; Kawasaki's Disease; Lupus erythematosus; Multiple sclerosis; Myasthenia gravis; Narcolepsy, Opsoclonus myoclonus syndrome (OMS); Optic neuritis; Ord's thyroiditis; Pemphigus; Pernicious anaemia; Polyarthritis in dogs; Primary biliary cirrhosis; Rheumatoid arthritis; Reiter's syndrome; Sjögren's syndrome; Takayasu's arteritis; Temporal arteritis (also known as “giant cell arteritis”); Warm autoimmune hemolytic anemia; or Wegener's granulomatosis   
       or the alloimmune response is involved in transplant rejection or graft-versus-host disease, or the maternal-foetal immune response is Rhesus D Haemolytic Disease of the Newborn. 
     
     
         14 . A composition according to  claim 11  wherein the individual to be treated is from a population where the allele frequencies of the following DRB1 alleles is:
 4—at least 9% 
 7—at least 10% 
 11—at least 8%. 
 
     
     
         15 . A composition according to  claim 11  wherein the individual to be treated has or is at risk of a condition, wherein the condition is an adverse inflammatory reaction to a treatment comprising a peptide. 
     
     
         16 . A composition as defined in  claim 1  for use in an in vitro method of diagnosing the presence or absence in a subject of a T-cell immune response to the protein from which the epitope derives, the method comprising:
 i) contacting the composition with T cells in a sample taken from the subject, under conditions which allow the peptide and the T cells to interact; 
 ii) determining whether or not any of the T cells are stimulated; and 
 thereby determining whether or not a T-cell immune response is present or absent. 
 
     
     
         17 . A composition according to  claim 15 , wherein the T cells are present in a population of PBMCs isolated from a blood or serum sample taken from the subject and/or wherein step (ii) comprises measuring the production of a cytokine by the T cells. 
     
     
         18 . A composition according to  claim 17 , wherein the production of a cytokine is detected by an ELISPOT or multiplex bead array assay 
     
     
         19 . A composition according to  claim 17 , wherein the cytokine is interferon-gamma. 
     
     
         20 . A composition according to  claim 1 , wherein the at least one peptide comprises or consists of the sequence corresponding to any one of SEQ ID NOS: 1 to 72. 
     
     
         21 . A composition according to  claim 1 , comprising at least a first and a second peptide, wherein the first and second peptide each comprise or consist of a different sequence selected from the sequences of SEQ ID NO: 37 (MLA01), SEQ ID NO: 38 (MLA04), SEQ ID NO: 39 (MLA05), or SEQ ID NO: 40 (MLA12). 
     
     
         22 . A composition according to  claim 21 , wherein the first and second peptides comprise or consist of the sequences of:
 a) SEQ ID NOS: 37 (MLA01) and 38 (MLA04);   b) SEQ ID NOS: 37 (MLA01) and 39 (MLA05);   c) SEQ ID NOS: 37 (MLA01) and 40 (MLA12);   d) SEQ ID NOS: 38 (MLA04) and 39 (MLA05);   e) SEQ ID NOS: 38 (MLA04) and 40 (MLA12); or   f) SEQ ID NOS: 39 (MLA05) and 40 (MLA12), respectively.   
     
     
         23 . A composition according to  claim 20 , wherein the agent is thioglycerol. 
     
     
         24 . An antibody which binds to the peptide of the composition according to  claim 1 . 
     
     
         25 . An antibody according to  claim 24  which binds to the peptide when the peptide is associated with an MHC Class II molecule.