Immunoglobulin chimeric monomer-dimer hybrids
View Patent ↗The invention relates to a chimeric monomer-dimer hybrid protein wherein said protein comprises a first and a second polypeptide chain, said first polypeptide chain comprising at least a portion of an immunoglobulin constant region and a biologically active molecule, and said second polypeptide chain comprising at least a portion of an immunoglobulin constant region without the biologically active molecule of the first chain. The invention also relates to methods of using and methods of making the chimeric monomer-dimer hybrid protein of the invention.
1. An isolated monomer-dimer hybrid immunoconjugate comprising
X-L a -F 1 :F 2 or F 2 :F 1 -L a -X
wherein
X is a single biologically active molecule,
L is a linker,
a is any integer or zero,
: represents a chemical association,
F 1 is at least a portion of an immunoglobulin constant region comprising an FcRn binding site and does not comprise a biologically active molecule, and
F 2 is at least a portion of an immunoglobulin constant region comprising an FcRn binding site and does not comprise a biologically active molecule or immunoglobulin variable region.
2. The monomer-dimer hybrid immunoconjugate of claim 1 , wherein F 2 further comprises an affinity tag.
3. The protein of claim 2 , wherein the affinity tag is a FLAG tag or His tag.
4. An isolated monomer-dimer hybrid immunoconjugate of the formula
X-L a -F 1 :F 2 -T or T-F 2 :F 1 -L a -X
wherein
X is a single biologically active molecule,
L is a linker,
a is any integer or zero,
T is a molecule having a molecular weight less than 2 kD,
: represents a chemical association,
F 1 is an immunoglobulin constant region or portion thereof comprising an FcRn binding site, and
F 2 is at least a portion of an immunoglobulin constant region comprising an FcRn binding site.
5. The monomer-dimer hybrid immunoconjugate of claim 4 , wherein T is attached to the C terminus of F 2 .
6. The monomer-dimer hybrid immunoconjugate of claim 4 , wherein T is attached to the N terminus of F 2 .
7. The monomer-dimer hybrid immunoconjugate of claim 4 , wherein T is an affinity tag.
8. The monomer-dimer hybrid immunoconjugate of claim 7 , wherein the affinity tag is a FLAG tag or His tag.
9. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein F 1 and F 2 are Fc fragments.
10. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein the immunoglobulin is IgG.
11. The monomer-dimer hybrid immunoconjugate of claim 10 , wherein the IgG is an IgG1 or an IgG2.
12. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein X is a polypeptide.
13. The monomer-dimer hybrid immunoconjugate of claim 12 , wherein X is a protein.
14. The monomer-dimer hybrid immunoconjugate of claim 13 , wherein the protein is a clotting factor.
15. The monomer-dimer hybrid immunoconjugate of claim 14 , wherein the clotting factor is Factor IX.
16. The monomer-dimer hybrid immunoconjugate of claim 14 , wherein the clotting factor is Factor VIII.
17. The monomer-dimer hybrid immunoconjugate of claim 14 , wherein the clotting factor is Factor VII or VIIa.
18. The monomer-dimer hybrid immunoconjugate of claim 13 , wherein the protein is follicle stimulating hormone (FSH).
19. The monomer-dimer hybrid immunoconjugate of claim 13 , wherein the protein is interferon.
20. The monomer-dimer hybrid immunoconjugate of claim 19 , wherein the protein is interferon α.
21. The monomer-dimer hybrid immunoconjugate of claim 20 , wherein L a consists of 15-25 amino acids.
22. The monomer-dimer hybrid immunoconjugate of claim 20 , wherein L a consists of 15-20 amino acids.
23. The monomer-dimer hybrid immunoconjugate of claim 19 , wherein the protein is interferon β.
24. The monomer-dimer hybrid immunoconjugate of claim 13 , wherein the protein is a growth factor.
25. The monomer-dimer hybrid immunoconjugate of claim 24 , wherein the growth factor is erythropoietin.
26. The monomer-dimer hybrid immunoconjugate of claim 12 , wherein X is a peptide.
27. The monomer-dimer hybrid immunoconjugate of claim 26 , wherein the peptide is a viral fusion inhibitor.
28. The monomer-dimer hybrid immunoconjugate of claim 27 , wherein the viral fusion inhibitor is an HIV fusion inhibitor.
29. The monomer-dimer hybrid immunoconjugate of claim 28 , wherein the HIV fusion inhibitor is T20 (SEQ ID NO:1), T21 (SEQ ID NO:2), or T1249 (SEQ ID NO:3).
30. The monomer-dimer hybrid immunoconjugate of claim 26 , wherein the peptide is leuprolide.
31. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein X is a small molecule.
32. The monomer-dimer hybrid immunoconjugate of claim 31 , wherein the small molecule is a VLA4 antagonist.
33. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein X is a nucleic acid.
34. The monomer-dimer hybrid immunoconjugate of claim 33 , wherein the nucleic acid is DNA.
35. The monomer-dimer hybrid immunoconjugate of claim 33 , wherein the nucleic acid is RNA.
36. The monomer-dimer hybrid immunoconjugate of claim 33 , wherein the nucleic acid is an antisense molecule.
37. The monomer-dimer hybrid immunoconjugate of claim 33 , wherein the nucleic acid is a ribozyme.
38. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein the chemical association between F 1 and F 2 is a non-covalent interaction.
39. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein the chemical association between F 1 and F 2 is a covalent bond.
40. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein the chemical association between F 1 and F 2 is a disulfide bond.
41. The monomer-dimer hybrid immunoconjugate of claim 1 or 4 , wherein the chemical association between F 1 and F 2 is not a disulfide bond.
42. An isolated monomer-dimer hybrid immunoconjugate of the formula
X-L a -F 1 :F 2 or F 2 :F 1 -L a -X
wherein
X is erythropoietin,
L is a linker,
a is any integer or zero,
: represents a chemical association,
F 1 is an immunoglobulin constant region or portion thereof comprising an FcRn binding site; and
F 2 is at least a portion of an immunoglobulin constant region comprising an FcRn binding site.
43. The protein monomer-dimer hybrid immunoconjugate of claim 42 , wherein L is an eight amino acid linker having the amino acid sequence EFAGAAAV (SEQ ID NO:31).
44. The of claim 42 , wherein F1 and F2 are Fc fragments having a mutation of asparagine to alanine at position 297.
45. An isolated monomer-dimer hybrid immunoconjugate of the formula
X-L a -F 1 :F 2 or F 2 :F 1 -L a -X
wherein
X is IFNβ,
L is a linker,
a is any integer or zero,
: represents a chemical association,
F 1 is an immunoglobulin constant region or portion thereof comprising an FcRn binding site; and
F 2 is at least a portion of an immunoglobulin constant region comprising an FcRn binding site.
46. The monomer-dimer hybrid immunoconjugate of claim 45 , wherein F 1 and F 2 are Fc fragments having a mutation of asparagine to alanine at position 297.
47. An isolated monomer-dimer hybrid immunoconjugate of the formula
X-L a -F 1 :F 2 or F 2 :F 1 -L a -X
wherein
X is Factor IX,
L is a linker,
a is any integer or zero,
: represents a chemical association,
F 1 is an immunoglobulin constant region or portion thereof comprising an FcRn binding site; and
F 2 is at least a portion of an immunoglobulin constant region comprising an FcRn binding site.
48. The monomer-dimer hybrid immunoconjugate of claim 47 , wherein F 1 and F 2 are Fc fragments having a mutation of asparagine to alanine at position 297.