Rage Fusion Proteins And Methods Of Use
Disclosed are RAGE fusion proteins comprising RAGE polypeptide sequences linked to a second, non-RAGE polypeptide. The RAGE fusion protein may utilize a RAGE polypeptide domain comprising a RAGE ligand binding site and an interdomain linker directly linked to an immunoglobulin C H 2 domain. Such fusion proteins may provide specific, high affinity binding to RAGE ligands. Also disclosed is the use of the RAGE fusion proteins as therapeutics for RAGE-mediated pathologies.
1 . A nucleic acid encoding a fusion protein comprising a RAGE polypeptide directly linked to a polypeptide comprising a C H 2 domain of an immunoglobulin or a portion of a C H 2 domain of an immunoglobulin, wherein the RAGE polypeptide comprises a RAGE ligand binding site, and wherein the fusion protein does not include an immunoglobulin Fc hinge region.
2 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises a RAGE interdomain linker linked to a RAGE immunoglobulin domain such that the C-terminal amino acid of the RAGE immunoglobulin domain is linked to the N-terminal amino acid of the RAGE interdomain linker, and the C-terminal amino acid of the RAGE interdomain linker is directly linked to the N-terminal amino acid of the polypeptide comprising the C H 2 domain of the immunoglobulin or the portion of the C H 2 domain of the immunoglobulin.
3 . The nucleic acid of claim 1 , comprising the nucleic acid sequence as set forth in SEQ ID NO: 30, or the nucleic acid sequence as set forth in SEQ ID NO: 31 .
4 . An expression vector comprising the nucleic acid of claim 1 .
5 . An expression vector comprising the nucleic acid of claim 3 .
6 . A cell comprising the nucleic acid of claim 1 .
7 . The nucleic acid of claim 1 , wherein the RAGE ligand binding site comprises the amino acid sequence as set forth in SEQ ID NO: 9 or a sequence 90% identical thereto, or the amino acid sequence as set forth in SEQ ID NO: 10 or a sequence 90% identical thereto.
8 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises a RAGE interdomain linker linked to a RAGE immunoglobulin domain.
9 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises a first RAGE immunoglobulin domain and a first RAGE interdomain linker linked to a second RAGE immunoglobulin domain and a second RAGE interdomain linker, such that the N-terminal amino acid of the first interdomain linker is linked to the C-terminal amino acid of the first RAGE immunoglobulin domain, the N-terminal amino acid of the second RAGE immunoglobulin domain is linked to the C-terminal amino acid of the first interdomain linker, the N-terminal amino acid of the second interdomain linker is linked to the C-terminal amino acid of the second RAGE immunoglobulin domain, and the C-terminal amino acid of the second RAGE interdomain linker is directly linked to the N-terminal amino acid of the C H 2 domain of the immunoglobulin or the portion of the C H 2 domain of the immunoglobulin.
10 . The nucleic acid of claim 1 , wherein the fusion protein comprises the amino acid sequence as set forth in SEQ ID NO: 32, SEQ ID NO: 33 or SEQ ID NO: 34.
11 . The nucleic acid of claim 9 , wherein the second RAGE interdomain linker directly linked to the C H 2 domain of the immunoglobulin or the portion of the C H 2 domain of the immunoglobulin comprises the amino acid sequence as set forth in SEQ ID NO: 22 or a sequence 90% identical thereto, or the amino acid sequence as set forth in SEQ ID NO: 24, or a sequence 90% identical thereto.
12 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises a single RAGE immunoglobulin domain linked via a RAGE interdomain linker to the N-terminal amino acid of the polypeptide comprising the C H 2 domain of the immunoglobulin or the portion of the C H 2 domain of the immunoglobulin.
13 . The nucleic acid of claim 1 , encoding a fusion protein comprising the amino acid sequence as set forth in SEQ ID NO: 35, SEQ ID NO: 36 or SEQ ID NO: 37.
14 . The nucleic acid of claim 12 , wherein the RAGE linker is directly linked to the C H 2 domain of the immunoglobulin or the portion of the C H 2 domain of the immunoglobulin and the RAGE linker comprises the amino acid sequence as set forth in SEQ ID NO: 21 or a sequence 90% identical thereto, or the amino acid sequence as set forth in SEQ ID NO: 23 or a sequence 90% identical thereto.
15 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises the amino acid sequence as set forth in SEQ ID NO: 7 or the amino acid sequence as set forth in SEQ ID NO: 8.
16 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises the amino acid sequence as set forth in SEQ ID NO: 13 or the amino acid sequence as set forth in SEQ ID NO:14.
17 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises the amino acid sequence as set forth in SEQ ID NO: 15 or the amino acid sequence as set forth in SEQ ID NO: 16.
18 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises the amino acid sequence as set forth in SEQ ID NO: 17 or the amino acid sequence as set forth in SEQ ID NO: 18.
19 . The nucleic acid of claim 1 , wherein the RAGE polypeptide comprises the amino acid sequence as set forth in SEQ ID NO: 19 or the amino acid sequence as set forth in SEQ ID NO: 20.
20 . The nucleic acid of claim 1 , wherein the amino acid sequence of the RAGE polypeptide is as set forth in SEQ ID NO: 15 or SEQ ID NO: 16.
21 . The nucleic acid of claim 1 , wherein the amino acid sequence of the RAGE polypeptide is as set forth in SEQ ID NO: 19 or SEQ ID NO: 20.
22 . The nucleic acid of claim 1 , wherein the polypeptide is derived from a human IgA, IgD, IgE, IgG or IgM.
23 . The nucleic acid of claim 1 , wherein the polypeptide is derived from a human IgG.
24 . The nucleic acid of claim 1 , wherein the polypeptide is derived from a human IgG1.
25 . The nucleic acid of claim 1 , wherein the polypeptide further comprises a C H 3 domain of the immunoglobulin or a portion thereof.
26 . The nucleic acid of claim 1 , wherein the amino acid sequence of the polypeptide comprises the amino acid sequence as set forth in SEQ ID NO: 38.
27 . The nucleic acid of claim 1 , wherein the amino acid sequence of the polypeptide is the amino acid sequence as set forth in SEQ ID NO: 38.
28 . The nucleic acid of claim 1 , wherein the nucleic acid sequence encoding the RAGE polypeptide comprises the nucleic acid sequence as set forth in any one of SEQ ID NOS: 25, 26, 27, 28 or 29.
29 . The nucleic acid of claim 1 , comprising the nucleic acid sequence set forth in SEQ ID NO: 30.
30 . The nucleic acid of claim 1 , comprising the nucleic acid sequence set forth in SEQ ID NO: 31.
31 . A host cell comprising the expression vector of claim 4 .
32 . A host cell comprising the expression vector of claim 5 .