Engineered antibody constant domain molecules
View Patent ↗Described herein are engineered antibody constant domain molecules, such as CH2 or CH3 domain molecules, comprising at least one mutation, or comprising at least one complementarity determining region (CDR), or a functional fragment thereof, engrafted in a loop region of the CH2 domain. The CH2 domain molecules described herein are small, stable, soluble, exhibit little to no toxicity and are capable of binding antigen.
1. A polypeptide comprising a human immunoglobulin CH2 domain of IgG, IgA or IgD, or a human immunoglobulin CH3 domain of IgE or IgM, wherein the CH2 domain or CH3 domain comprises an N-terminal truncation of 7 amino acids, and wherein Loop 1 of the CH2 or CH3 domain is mutated, wherein the polypeptide has a molecular weight of less than about 15 kD, and wherein the polypeptide specifically binds an antigen.
2. The polypeptide of claim 1 , wherein the polypeptide comprises a CH2 domain from IgG.
3. The polypeptide of claim 1 , wherein the CH2 domain or CH3 domain further comprises a mutated Loop2, a mutated Loop3, a mutated Loop A-B, a mutated Loop C-D, a mutated Loop E-F, or any combination thereof.
4. The polypeptide of claim 1 having a molecular weight of about 12 kD to about 14 kD.
5. The polypeptide of claim 1 , wherein the CH2 domain or CH3 domain further comprises a C-terminal truncation of about 1 to about 4 amino acids.
6. The polypeptide of claim 1 , wherein the CH2 domain or CH3 domain is capable of binding an Fc receptor, a complement protein, or both.
7. The polypeptide of claim 1 , wherein the antigen is from a pathogen.
8. The polypeptide of claim 7 , wherein the pathogen is a virus or bacterium.
9. The polypeptide of claim 8 , wherein the virus is human immunodeficiency virus (HIV).
10. The polypeptide of claim 1 , wherein the antigen is a cancer-specific antigen or a tumor-associated antigen.
11. The polypeptide of claim 10 , wherein the cancer is leukemia, lymphoma, multiple myeloma, malignant melanoma, breast cancer, lung cancer, prostate cancer, colon cancer or renal cell carcinoma.
12. The polypeptide of claim 1 , wherein the antigen is TNF-α.
13. A composition comprising the polypeptide of claim 1 and a pharmaceutically acceptable carrier.
14. A composition comprising the polypeptide of claim 1 conjugated to an effector molecule or a detectable label.
15. The polypeptide of claim 1 , further comprising a first amino acid substitution and a second amino acid substitution, wherein the first and second amino acid substitutions each replace the original residue with a cysteine residue, wherein the cysteine residues form a disulfide bond.
16. The polypeptide of claim 15 , wherein the first amino acid substitution is in the N-terminal A strand and the second amino acid substitution is in the C-terminal G strand.
17. The polypeptide of claim 15 comprising a CH2 domain of IgG.
18. The polypeptide of claim 17 , wherein the first amino acid substitution is L12 to C12 and the second amino acid substitution is K104 to C104 (numbered with reference to SEQ ID NO: 5).
19. The polypeptide of claim 17 , wherein the first amino acid substitution is V10 to C10 and the second amino acid substitution is K104 to C104 (numbered with reference to SEQ ID NO: 5).
20. The polypeptide of claim 1 , wherein the mutated Loop 1 comprises random substitutions of any combination of alanine, tyrosine, aspartic acid or serine residues.
21. The polypeptide of claim 20 , wherein all Loop 1 residues are replaced by alanine, tyrosine, aspartic acid or serine residues.
22. The polypeptide of claim 20 , wherein the mutated Loop 1 further comprises an extra glycine residue on the C-terminus of Loop 1.
23. The polypeptide of claim 1 , wherein the CH2 domain or CH3 domain further comprises a mutated Loop 3, and wherein the mutated Loop 3 comprises random substitutions of any combination of alanine, tyrosine, aspartic acid or serine residues.
24. The polypeptide of claim 23 , wherein all Loop 3 residues are replaced by alanine, tyrosine, aspartic acid or serine residues.
25. The polypeptide of claim 23 , wherein the mutated Loop 3 of the CH2 domain or CH3 domain further comprises an extra glycine residue on the C-terminus of Loop 3.