Antibodies comprising ion-concentration-dependent antigen-binding domains, fc region variants, il-8-binding antibodies, and uses thereof

By modifying the charge of amino acid residues on the antibody surface and developing pH-dependent anti-IL-8 antibodies, the problems of short antibody retention time in plasma and binding to pre-existing antibodies were solved, achieving a longer half-life and lower immunogenicity, while improving IL-8 neutralization capacity.

CN107108729BActive Publication Date: 2026-07-31CHUGAI PHARMA CO LTD
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
CHUGAI PHARMA CO LTD
Filing Date
2016-02-04
Publication Date
2026-07-31

AI Technical Summary

Technical Problem

Existing therapeutic antibodies have a short retention time in plasma and may bind to pre-existing anti-drug antibodies, leading to increased side effects. They are also difficult to effectively remove soluble antigens such as IL-8, and current technologies have failed to effectively improve the pharmacokinetics and immunogenicity of antibodies.

Method used

By modifying the charge of amino acid residues on the antibody surface to change its isoelectric point (pI), the binding to the extracellular matrix is ​​increased, and the binding to FcRn is enhanced under acidic pH conditions, while the binding to pre-existing ADA is reduced. At the same time, pH-dependent anti-IL-8 antibodies are developed to improve IL-8 neutralization capacity.

Benefits of technology

It prolongs the half-life of the antibody in plasma, reduces binding to pre-existing ADA, enhances IL-8 neutralization capacity, and reduces immunogenicity, thus achieving rapid removal of IL-8.

✦ Generated by Eureka AI based on patent content.

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Abstract

One non-exclusive aspect provides molecules that are further improved from antibodies capable of binding antigens in an ion concentration-dependent manner. An alternative non-exclusive aspect provides safer and more favorable Fc region variants with reduced binding to pre-existing ADA. An alternative non-exclusive aspect provides novel IL-8 antibodies that are excellent as pharmaceuticals.
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Claims

1. A method for screening antibodies, wherein the antibody has an improved ability to clear antigens from plasma compared to a reference antibody, and the antigen-binding activity of the antibody is retained compared to the reference antibody, the reference antibody comprising a heavy chain consisting of the amino acid sequence of SEQ ID NO: 38 and a light chain consisting of the amino acid sequence of SEQ ID NO: 39, or comprising a first heavy chain consisting of the amino acid sequence of SEQ ID NO: 229, a second heavy chain consisting of the amino acid sequence of SEQ ID NO: 230, and a light chain consisting of the amino acid sequence of SEQ ID NO:

231. The antibody contains an antigen-binding domain, the antigen-binding activity of which varies depending on ion concentration conditions. The method includes: (a) Modifying the amino acid residues of the reference antibody to prepare a modified antibody, wherein When the reference antibody comprises a heavy chain consisting of the amino acid sequence of SEQ ID NO: 38 and a light chain consisting of the amino acid sequence of SEQ ID NO: 39, the amino acid residues modifying the reference antibody are selected from the group consisting of: (1) Based on the Kabat number in the heavy chain, replace Thr with Arg at position 77, replace Asp with Asn at position 82a, and replace Ser with Arg at position 82b. (2) Based on the Kabat number in the heavy chain, replace Thr with Arg at position 77, replace Asp with Gly at position 82a, and replace Ser with Arg at position 82b. (3) Based on the Kabat number in the heavy chain, replace Thr with Arg at position 77, replace Asp with Ser at position 82a, and replace Ser with Arg at position 82b. (4) Based on the Kabat number in the heavy chain, replace Thr with Arg at position 77 and replace Glu with Gly at position 85. (5) According to the Kabat number in the light chain, replace Gly with Lys at position 16, replace Gln with Arg at position 24, and replace Glu with Arg at position 27. (6) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, and replace Gln with Arg at position 37. (7) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Gly with Arg at position 41, and replace Gln with Lys at position 42. (8) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Ser with Lys at position 52, and replace Ser with Lys at position 56. (9) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Ser with Arg at position 65, and replace Thr with Arg at position 69. (10) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (11) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (12) According to the Kabat number in the light chain, replace Ala with Arg at position 25, replace Ser with Arg at position 26, and replace Gln with Arg at position 37. (13) According to the Kabat number in the light chain, replace Ala with Arg at position 25, replace Ser with Arg at position 26, replace Ser with Arg at position 52, and replace Ser with Arg at position 56. (14) According to the Kabat number in the light chain, replace Ala with Arg at position 25, replace Ser with Arg at position 26, replace Ser with Lys at position 52, and replace Ser with Lys at position 56. (15) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Gly with Arg at position 41, replace Gln with Lys at position 42, replace Ser with Arg at position 65, and replace Thr with Arg at position 69. (16) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Ser with Arg at position 65, and replace Thr with Arg at position 69. (17) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (18) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (19) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Gly with Arg at position 41, replace Gln with Lys at position 42, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (20) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (21) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Gly with Arg at position 41, replace Gln with Lys at position 42, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (22) According to the Kabat number in the light chain, replace Gln with Arg at position 24, replace Glu with Arg at position 27, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. When the reference antibody comprises a first heavy chain consisting of the amino acid sequence of SEQ ID NO: 229, a second heavy chain consisting of the amino acid sequence of SEQ ID NO: 230, and a light chain consisting of the amino acid sequence of SEQ ID NO: 231, the amino acid residues modifying the reference antibody are selected from the group consisting of: (1) According to the Kabat number in the light chain, replace Thr with Lys at position 74 and replace Ser with Arg at position 77. (2) Based on the Kabat number in the light chain, replace Ser with Arg at position 76 and replace Gln with Lys at position 79. (3) According to the Kabat number in the light chain, replace Gly with Arg at position 41, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (4) According to the Kabat number in the light chain, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (5) According to the Kabat number in the light chain, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (6) According to the Kabat number in the light chain, replace Gln with Arg at position 27, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (7) According to the Kabat number in the light chain, replace Gly with Arg at position 41, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (8) According to the Kabat number in the light chain, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (9) According to the Kabat number in the light chain, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (10) According to the Kabat number in the light chain, replace Gln with Arg at position 27, replace Gly with Arg at position 41, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (11) According to the Kabat number in the light chain, replace Gly with Arg at position 41, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (12) According to the Kabat number in the light chain, replace Gly with Arg at position 41, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (13) According to the Kabat number in the light chain, replace Gln with Arg at position 27, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (14) According to the Kabat number in the light chain, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (15) According to the Kabat number in the light chain, replace Gln with Arg at position 27, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Thr with Lys at position 74, and replace Ser with Arg at position 77. (16) According to the Kabat number in the light chain, replace Gln with Arg at position 27, replace Gly with Arg at position 41, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (17) According to the Kabat number in the light chain, replace Gly with Arg at position 41, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (18) According to the Kabat number in the light chain, replace Gly with Arg at position 41, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (19) According to the Kabat number in the light chain, replace Gln with Arg at position 27, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (20) According to the Kabat number in the light chain, replace Ser with Arg at position 52, replace Ser with Arg at position 56, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (21) According to the Kabat number in the light chain, replace Gln with Arg at position 27, replace Ser with Arg at position 65, replace Thr with Arg at position 69, replace Ser with Arg at position 76, and replace Gln with Lys at position 79. (22) According to the Kabat number in the light chain, replace Ser with Arg at position 76, and (23) In the light chain according to the Kabat number, replace Ser with Arg at position 65 and replace Gln with Lys at position 79; (b) Confirm the isoelectric point (pI) of the modified antibody and the reference antibody, and then select the modified antibody whose isoelectric point (pI) is higher than that of the reference antibody; (c) Determine the antigen-binding activity of the modified antibody and the reference antibody, and (d) Select a modified antibody that retains the antigen-binding activity compared to the reference antibody.

2. The method according to claim 1, wherein the antigen is a soluble antigen.

3. The method according to claim 1 or 2, wherein the antigen-binding domain is a domain whose antigen-binding activity is higher under low ion concentration conditions than under high ion concentration conditions.

4. The method according to claim 1 or 2, wherein the ion concentration is the hydrogen ion concentration.

5. The method of claim 4, wherein its ratio of KD at acidic pH range to that at neutral pH range for the antigen is 2 or higher.

5. The method of claim 4, wherein its ratio of KD at acidic pH range to that at neutral pH range for the antigen is 2 or higher. 酸性pH范围 / KD 中性pH范围 is 2 or higher.

6. The method of claim 5, further comprising modifying amino acid residues of the modified antibody, wherein the amino acid residues modifying the modified antibody are selected from the group consisting of: (1) According to the EU designation, replace Asp with Lys at position 413. (2) According to the EU designation, replace Gln with Arg at position 311. (3) According to the EU designation, replace Pro with Arg at position 343. (4) According to the EU designation, replace Asp with Arg at position 401. (5) Based on the EU designation, replace Asp with Lys at position 401, and (6) Replace Gly with Arg at position 402 according to the EU number.

7. The method according to claim 1 or 2, wherein the constant region of the modified antibody has Fcγ receptor (FcγR)-binding activity, and wherein the FcγR-binding activity is enhanced under neutral pH conditions compared to a reference antibody containing a constant region of natural IgG.

8. The method of claim 7, wherein the FcγR is FcγRIIb.

9. The method according to claim 1 or 2, wherein the constant region of the modified antibody has binding activity to one or more activating FcγRs selected from the group consisting of FcγRIa, FcγRIb, FcγRIc, FcγRIIIa, FcγRIIIb and FcγRIIa, and to FcγRIIb, and the FcγRIIb binding activity is maintained or enhanced and the binding activity to activating FcγR is reduced compared to a reference antibody that is distinguished only by its constant region being the constant region of natural IgG.

10. The method according to claim 1 or 2, wherein the constant region of the modified antibody has FcRn-binding activity, and wherein the FcRn-binding activity of the constant region is enhanced under neutral pH conditions compared to a reference antibody that differs only in that its constant region is a constant region of natural IgG.

11. The method according to claim 1 or 2, wherein the method further comprises: (e) Obtain the nucleic acid encoding the modified antibody selected in claim 1(d), (f) Culturing host cells to express the nucleic acid of (e); and (g) Collect the antibody from the host cell culture.

12. A pharmaceutical composition comprising an antibody produced by the method of claim 1 or 2.