Chimeric protein in the treatment of amyloidosis

Abstract

The present invention relates to a chimeric protein comprising at least one human amyloid P component and at least one fragment of an Fc region of a human antibody, the human amyloid P component and the fragment of an Fc region with which it is associated being bound to each other by means of a hinge region.

1 .- 29 . (canceled) 
     
     
         30 . A chimeric protein comprising at least one human amyloid P component and at least one fragment of an Fc region of a human antibody, wherein the human amyloid P component and the fragment of an Fc region are bonded to each other by a hinge region. 
     
     
         31 . The chimeric protein as claimed in  claim 30 , wherein the human amyloid P component comprises an amino acid sequence having at least 80% identity with SEQ ID NO: 1. 
     
     
         32 . The chimeric protein as claimed in  claim 30 , wherein the Fc region of the antibody is the Fc region of a human immunoglobulin, preferably the Fc region of an IgG, more preferably the Fc region of an IgG1 or of an IgG2, and even more preferably the Fc region of an IgG1. 
     
     
         33 . The chimeric protein as claimed in  claim 30 , wherein the fragment of an Fc region of a human antibody comprises at least one amino acid sequence having at least 80% identity with SEQ ID NO: 3. 
     
     
         34 . The chimeric protein as claimed in  claim 30 , wherein said Fc region comprises at least two modifications in the amino acid sequence, namely:
 (i) a modification in the amino acid sequence selected from the group consisting of 378V, 378T, 434Y and 434S, and   (ii) at least one modification in the amino acid sequence selected from the group consisting of 2260, 230S, 230T, 230L, 241 L, 264E, 307P, 315D, 330V, 362R, 378V, 378T, 389T, 389K, 434Y and 434S,   wherein the numbering of the amino acids of the Fc region is that of the EU index proposed by Kabat, and wherein the modification (i) does not occur at the same amino acid position as the modification (ii).   
     
     
         35 . The chimeric protein as claimed in  claim 30 , wherein the hinge region comprises at least one amino acid sequence having at least 60% identity, preferably at least 80% identity, with a sequence selected from the SEQ ID NOs: 13 and 15 to 18, preferably SEQ ID NO: 13. 
     
     
         36 . The chimeric protein as claimed in  claim 30 , wherein said chimeric protein further comprises, between the hinge region and the amyloid P component, at least one non-structuring peptide sequence, preferably SEQ ID NO: 5. 
     
     
         37 . The chimeric protein as claimed in  claim 30 , wherein said chimeric protein comprises at least two units, each unit comprising at least one human amyloid P component and at least one fragment of an Fc region of a human antibody bonded to each other by a hinge region, wherein the two units are bonded to each other by at least one disulfide bridge, and wherein the fragments of an Fc region of a human antibody are identical or different. 
     
     
         38 . The chimeric protein as claimed in  claim 30 , wherein said chimeric protein comprises at least one human amyloid P component and at least two fragments of an Fc region of a human antibody, which may be identical or different, wherein said human amyloid P component and the first fragment are bonded to each other by a first hinge region, wherein the first and second fragments of an Fc region are bonded to each other covalently by means of a bond formed from a spacer chain and a second hinge region, wherein the first hinge region and the second hinge region are identical or different, and wherein the first and second fragments of an Fc region form a single polypeptide chain constituting a functional dimeric Fc region. 
     
     
         39 . The chimeric protein as claimed in  claim 38 , wherein the spacer chain is represented by at least one amino acid sequence SEQ ID NO: 5. 
     
     
         40 . A nucleic acid comprising at least one polynucleotide sequence encoding at least one chimeric protein as defined in  claim 30 . 
     
     
         41 . A vector comprising a nucleic acid as defined in  claim 40 . 
     
     
         42 . A host cell transfected with a vector as defined in  claim 41 . 
     
     
         43 . A process for producing a chimeric protein as defined in  claim 30 , said process comprising:
 a) transfecting a host cell with a vector;   b) culturing said host cell under conditions such that the chimeric protein is produced; and   c) collecting said chimeric protein produced by said host cell in step b).   
     
     
         44 . The process as claimed in  claim 43 , wherein the host cell is a eukaryotic cell, preferably a cell selected from the group consisting of the following cells: YB2/0, in particular the line deposited at the “American Type Culture Collection” under ATCC No. CRL-1662, SP2/0, YE2/0, 1R983F, Namalwa, PER.C6, CHO cell lines, in particular CHO-K-1, CHO-Lecl0, CHO-Lecl, CHO-Lecl3, CHO Pro-5, CHO dhfr-, Wil-2, Jurkat, Vero, Molt-4, COS-7, 293-HEK, BHK, KGH6, NSO, SP2/0-Ag 14, P3X63Ag8.653, C127, JC, LA7, ZR-45-30, hTERT, NM2C5, UACC-812, DG44 and DXB11, DHFR, HELA, CVI, COS, R1610, BALBC/3T3, HAK, BF1-1c1BPT, RAJI, HEK, EB66, or BHK. 
     
     
         45 . A pharmaceutical composition comprising a chimeric protein as defined in  claim 30 , and a pharmaceutically acceptable excipient. 
     
     
         46 . A process for treating amyloidosis, in particular amyloidosis of AL type, comprising at least one step of administering, to an individual in need thereof, an effective amount of at least one first active agent represented by a chimeric protein as defined in  claim 30 . 
     
     
         47 . A process for treating amyloidosis, in particular amyloidosis of AL type, comprising at least one step of administering, to an individual in need thereof, an effective amount of at least one first active agent represented by a composition as defined in  claim 45 . 
     
     
         48 . The process as claimed in  claim 46 , further comprising a step of administering, prior to, concomitantly with and/or subsequent to the step of administering the first active agent, at least one second active agent distinct from said first active agent, the second active agent preferably being chosen from the group consisting of 4-[bis(chloroethyl)amino]phenylalanine; 9-fluoro-11β,17,21-trihydroxy-16a-methylpregna-1,4-diene-3,20-dione; prednisone; dimethyl sulfoxide; N-[(7S)-5,6,7,9-tetrahydro-1,2,3,10-tetramethoxy-9-oxobenzo[a]heptalen-7-yl)acetamide]; (7S,9S)-7-[(2R,4S,5S,6S)-4-amino-5-hydroxy-6-methyloxan-2-yl]oxy-6,9,11-trihydroxy-9-(2-hydroxyacetyl)-4-methoxy-8,10-dihydro-7H-tetracene-5,12-dione; active agents capable of reacting with the amyloid P component, in particular of bis-d-proline type, and more particularly the compound (R)-1-[6-[(R)-2-carboxypyrrolidin-1-yl]-6-oxohexanoyl]pyrrolidine-2-carboxylic acid; proteasome inhibitors, for example bortezomib, carfilzomib, marizomib, ixazomib, delanzomib, ONX-912 or revlimid; and a mixture thereof. 
     
     
         49 . The process as claimed in  claim 46 , wherein the chimeric protein is administered via intravenous, subcutaneous or intramuscular route. 
     
     
         50 . The process as claimed in  claim 46 , wherein the chimeric protein is administered via oral, nasal, rectal or vaginal route. 
     
     
         51 . The process as claimed in  claim 46 , wherein the chimeric protein is administered via pulmonary route. 
     
     
         52 . The process as claimed in  claim 46 , wherein the chimeric protein is administered via non-parenteral route. 
     
     
         53 . The process as claimed in  claim 46 , wherein the chimeric protein is administered at a dose ranging from 0.1 to 100 mg/kg, advantageously from 0.1 to 1000 μg/kg. 
     
     
         54 . A method for producing a chimeric protein, comprising using a starting chimeric protein as defined in  claim 30  to form a product chimeric protein comprising at least two units, each unit comprising at least one human amyloid P component and at least one fragment of an Fc region of a human antibody bonded to each other by a hinge region, wherein the two units are bonded to each other by at least one disulfide bridge, and wherein the fragments of an Fc region of a human antibody are identical or different. 
     
     
         55 . A method for characterizing in vitro or ex vivo the presence of an amyloidosis deposit, comprising using at least one chimeric protein as defined in  claim 30 , as a tool, said chimeric protein being radiolabeled; preferably, the human amyloid P component is represented by 123I-SAP, and/or identified by means of a radiolabeled anti-Fc antibody; for example, said anti-Fc antibody is coupled to peroxidase.