Single-chain multiple antigen-binding molecule, its preparation and use
The present invention relates to a single-chain, multiple antigen-binding molecule with diverse variable domains of a heavy and of a light chain of an immunoglobulin, which are connected in the form of a VH-VL construct, which are in turn connected together via a peptide, and to the preparation and use thereof as pharmaceutical or diagnostic aid.
1. A single-chain multiple antigen-binding molecule comprising:
(a) a variable domain of a heavy chain of an immunoglobulin (VH) with a first specificity (A), or functional parts thereof,
(b) a variable domain of a light chain of an immunoglobulin (VL) with a second specificity (B), or functional parts thereof,
(c) a variable domain of a heavy chain of an immunoglobulin (VH) with the specificity (B), or functional parts thereof, and
(d) a variable domain of a light chain of an immunoglobulin (VL) with the specificity (A), or functional parts thereof,
wherein the VH and VL domains are connected in the form of a VH-VL construct or VL-VH construct, and wherein the two VH-VL or VL-VH constructs are connected via a peptide (P) that connects variable domains with the same specificity, and wherein the first specificity (A) is different than the second specificity (B) and wherein the VH and VL domains are connected via a peptide linker (L) in the form of a VH-L-VL construct or VL-L-VH construct, wherein the linker (L) is about 1-20 amino acids long, and peptide (P) is 12 amino acids or longer.
2. The single-chain multiple antigen-binding molecule as claimed in claim 1 , which consists of two VH-VL constructs.
3. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the linker (L) comprises the amino acid sequence GGGGS (SEQ ID NO: 1).
4. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the peptide (P) is 12-40 amino acids long.
5. The single-chain multiple antigen-binding molecule as claimed in claim 4 , wherein the peptide (P) comprises the amino acid sequence GGGGSGGRASGGGS (SEQ ID NO: 2) or GGGGSGGRASGGGGS (SEQ ID NO: 3).
6. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein said molecule further comprises an effector (E) which effector (E) is a prodrug activating enzyme.
7. The single-chain multiple antigen-binding molecule as claimed in claim 6 , wherein the effector (E) is linked to said molecule via a connector (B).
8. The single-chain multiple antigen-binding molecule as claimed in claim 7 , wherein the connector (B) comprises a protease cleavage sequence.
9. The single-chain multiple antigen-binding molecule as claimed in claim 8 , wherein the protease cleavage sequence is selected from the group consisting of PSA, cathepsin, plasminogen and plasminogen activator cleavage sequence.
10. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the peptide (P) further comprises a fusogenic peptide.
11. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the first specificity (A) is directed against a protein.
12. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the first specificity (A) is directed against a cell membrane.
13. The single-chain multiple antigen-binding molecule as claimed in claim 12 , wherein the cell membrane is selected from the group consisting of cell membrane of lymphocytes, macrophages, monocytes, granulocytes, hematopoietic cells, endothelial cells, smooth muscle cells, striped muscle cells, epithelial cells, liver cells, kidney cells, glia cells, cells of the supporting tissue, tumor cells and leukemia cells.
14. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the first specificity (A) is directed against a target cell, and wherein the second specificity (B) is directed against a vector.
15. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the second specificity (B) is directed against a nucleic acid, a cationic peptide, a cationic lipid, a protein, a cationic polymer, a cationic porphyrin, or a viral vector.
16. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the first specificity (A) is directed against a toxin.
17. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the first specificity (A) is directed against a pharmaceutical.
18. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the first specificity (A) is directed against a pathogen.
19. The single-chain multiple antigen-binding molecule as claimed in claim 1 , wherein the second specificity (B) is directed against a tumor cell.
20. A pharmaceutical composition comprising a single-chain multiple antigen-binding molecule as claimed in claim 1 .