Artificially modified immunoglobulins

Abstract

This invention provides an effective method for killing tumor cells by recruiting neutrophils to enhance antibody-dependent cell-mediated cytotoxicity (ADCC). [Solution] The present invention provides artificially engineered immunoglobulins or fragments thereof, as well as methods for preparing and using them. The artificially engineered immunoglobulins are derived from human IgG1 and have been artificially engineered to confer Fc-alpha receptor binding ability. Furthermore, the artificially engineered IgG1 immunoglobulins or fragments thereof may retain binding to Fc-gamma receptors and / or FcRn.

Claims

1. Artificially engineered human IgG1 immunoglobulin containing Fc regions including the first and second Fc domains A molecule or a fragment thereof, wherein the first FC domain has at least one amino acid modification The first Fc domain is the Fc domain of wild-type IgG1 (amino of SEQ ID NO: 1). The acids CH2-1.6 to CH3-125 (IMGT numbering for C-domains) are few. They both have 65% identical amino acid sequences and bind to and activate human FcαRI. Processed human IgG1 immunoglobulin or its fragments.

2. The above-mentioned modification of at least one amino acid in the first Fc domain is the Fc domain of IgA1. Main (SEQ ID NO: 254), wild-type IgA2 Fc domain (amino acid CH of SEQ ID NO: 2) 2-1.2~CH3-125 (IMGT numbering for C-domain), parent IgA2 Fc domain (amino acids CH2-1.2 to CH3-125 of SEQ ID NO: 3 (for C-domain) IMGT numbering) or affinity matured variant Fc of IgA1 or IgA2 The artificially modified immunoglobulin according to claim 1, which is a substitution corresponding to the amino acid in the main component. It's a fragment of it.

3. As a result of the modification of at least one amino acid in the first Fc domain, human F An artificially manipulated immunoglobulin or fragment thereof according to claim 1 or 2, which can bind to cRn.

4. The artificial cytoplasm according to claim 3, which can bind to human FcRn with an affinity equivalent to that of wild-type IgG1. Immunoglobulin or its fragments.

5. The first Fc domain is the artificially modified immunoglobulin or a fragment thereof which is human FcγR Claims 1 to 4 further include at least one additional amino acid modification that can bind to An artificially modified immunoglobulin or fragment thereof as described in any one of the items.

6. It is a homodimer, and the second Fc domain is derived from the Fc domain of wild-type IgG1 ( Any one of claims 1 to 5, having an amino acid sequence that is at least 65% identical to SEQ ID NO: 1 Artificially modified immunoglobulin or fragment thereof as described in item 1.

7. It is a heterodimer, and the second Fc domain is derived from the Fc domain of wild-type IgG1. It has at least 70% identical amino acid sequence to (SEQ ID NO: 1), and binds to human FcγR. An artificially manipulated immunoglobulin or fragment thereof according to claim 1 or 2, which activates it.

8. The second Fc domain binds to FcRn, as described in claim 7 for the artificially engineered immunoglobulin. Blin or a fragment thereof.

9. Claim 7 or 8, wherein the FcγR is human FcγRIa and human FcγRIIIIa. Artificially modified immunoglobulins or fragments thereof as described above.

10. The first Fc domain is located in sequence number 122, sequence number 148, or sequence number 214. The amino acid sequence contained herein is the second Fc domain, which is sequence number 122, sequence number 1 Claim 1 or 2, each having an amino acid sequence contained in 48 or SEQ ID NO: 214 The artificially modified immunoglobulin or fragment thereof as described above.

11. The first Fc domains mentioned above are sequence numbers 188-193, 234-242, 205-20 It has an amino acid sequence contained within 9 or 223, or SEQ ID NOs: 194-199, 24 A claim having an amino acid sequence contained within 4-251, 210-214, or 224. Artificially modified immunoglobulin or fragment thereof as described in 3 or 4.

12. The first Fc domain is included in sequence number 150, 151, 152, or 153. An artificially manipulated immunoglobulin or fragment thereof according to claim 5, having an amino acid sequence.

13. The first Fc domain is sequence numbers 133, 135, 137, 139, 141, 14 3, 145, 155, 156, 157, 158, or 252 contains an amino acid sequence Furthermore, the second Fc domain is sequence numbers 132, 134, 136, 138, 140, 1 Included in 42, 144, 154, 159, 160, 161, 162, 163, or 164 An artificially manipulated immunoglobulin according to any one of claims 7 to 9, having the amino acid sequence Or a fragment thereof.

14. The first Fc domain has the amino acid sequence contained in Sequence ID No. 157, and the The Fc domain of 2 has the amino acid sequence contained in SEQ ID NO: 159 or 161. Artificially modified immunoglobulin or fragment thereof as described in item 13.

15. The first Fc domain has the amino acid sequence contained in SEQ ID NO: 252, and the The Fc domain of 2 has the amino acid sequence contained in SEQ ID NO: 159 or 161. Artificially modified immunoglobulin or fragment thereof as described in item 13.

16. A claim further comprising a variable heavy chain domain and a light chain, which recognizes and binds to a target antigen. An artificially modified immunoglobulin or fragment thereof as described in any one of items 1 to 15.

17. The target antigens are fibronectin EDA, HER2, EGFR, CD20, and CD30. The group consisting of EpCAM, GD2, and solid tumor antigens, as described in claim 16. Artificially modified immunoglobulin or its fragments.

18. One artificially modified immunoglobulin or fragment thereof according to any one of claims 1 to 17 A pharmaceutical composition comprising the above-mentioned pharmaceutically acceptable excipients, diluents, or carriers in combination.

19. The pharmaceutical composition according to claim 18, further comprising one or more additional therapeutic agents.

20. A person according to any one of claims 1 to 17 for use in the treatment of cell proliferation disorders Processed immunoglobulin or fragment thereof, or the pharmaceutical composition according to claim 18 or 19 thing.

21. The aforementioned proliferative disorders include fibrosis, breast cancer, neuroblastoma, lymphoma, colon cancer, epithelial carcinoma, and colorectal cancer. A selection from the group consisting of cancer, kidney cancer and mucosal tumors, for use according to claim 20 Artificially modified immunoglobulins or fragments thereof, or pharmaceutical compositions.

22. A method for treating cell proliferation disorders, wherein a therapeutically effective amount of any one of claims 1 to 17 The artificially modified immunoglobulin or fragment thereof described in claim 18 or 19 A method comprising administering a pharmaceutical composition to a subject for which it is required.

23. The aforementioned proliferative disorders include fibrosis, breast cancer, neuroblastoma, lymphoma, colon cancer, epithelial carcinoma, and colorectal cancer. The method according to claim 22, selected from the group consisting of cancer, kidney cancer, and mucosal tumors.

24. Code an artificially modified immunoglobulin or a fragment thereof according to any one of claims 1 to 17 A set of isolated nucleic acid molecules or nucleic acid molecules.

25. Claim 24, which is complementary DNA (cDNA) or messenger RNA (mRNA). The isolated nucleic acid molecule or set of nucleic acid molecules described above.

26. Cloni comprising one or more nucleic acid molecules or a set of nucleic acid molecules as described in claim 24 or 25 A stromal immunoglobulin or expression vector for recombinant production of the artificially manipulated immunoglobulin or its fragments. A cloning or expression vector suitable for the following.

27. A host cell comprising one or more cloning or expression vectors according to claim 26.

28. A method for producing artificially manipulated immunoglobulin or a fragment thereof according to any one of claims 1 to 17. A method for which the host cells described in claim 27 are subjected to the artificially manipulated immunoglobulin or the The cells are cultured under conditions sufficient to express the fragment, and then the human cells are extracted from the host cell culture. A method comprising purifying and recovering an immunoglobulin or fragment thereof.

29. Artificially modified immunoglobulin or fragment thereof according to any one of claims 1 to 17 or A kit comprising the pharmaceutical composition according to claim 18 or 19, the kit comprising instructions for use and the above Artificially modified immunoglobulins or their fragments or pharmaceutical compositions are administered to subjects where they are needed. A kit that includes further means of cooperation.

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