# US 12,600,791 B2 · Bispecific antibody, preparation method therefor and application thereof > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 12,600,791 B2 - Document ID: US12600791B2 - Status: Granted - Kind: B2 - Application number: 17/616,043 - Filed: 2019-12-17 - Published: 2022-10-13 - Granted: 2026-04-14 - Earliest claimed: 2019-07-02 - Assignees: Excelmab, Inc. - Applicants: Excelmab Inc. - Inventors: Wenjun Zhang; Feng Li; Yanan Hua; Jiaxi Liu; Chunhua Fang - Web page: https://patentgrep.com/patent/US12600791B2 ## CPC - C07K 16/2887: {against CD20} - A61P 35/02: specific for leukemia - C07K 16/2809: {against the T-cell receptor (TcR)-CD3 complex} - A61K 2039/505: {comprising antibodies} - C07K 2317/31: multispecific - C07K 2317/33: Crossreactivity, e.g. for species or epitope, or lack of said crossreactivity - C07K 2317/52: Constant or Fc region; Isotype - C07K 2317/522: CH1 domain - C07K 2317/526: CH3 domain - C07K 2317/565: Complementarity determining region [CDR] - C07K 2317/622: Single chain antibody (scFv) - C07K 2317/732: Antibody-dependent cellular cytotoxicity [ADCC] - C07K 2317/92: Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value - C07K 2317/94: Stability, e.g. half-life, pH, temperature or enzyme-resistance - C07K 2319/30: Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) - A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) ## Abstract Provided are a bispecific antibody and a preparation method therefor. The antibody comprises structural domains capable of binding to CD20 and CD3, and a heterodimer Fc region. The structural domains capable of binding to CD20 and CD3 are each independently selected from a Fab region, a ScFv region, or an sDab region. The bispecific antibody targets CD20 and CD3, and mediates T cell-specific killing of tumor cells. ## Claims ### Claim 1 A bispecific antibody, comprising a domain capable of binding to CD20, a domain capable of binding to CD3 and a heterodimeric Fc region, wherein the domain capable of binding to CD20 is selected from the group consisting of a Fab region, a scFv region and a sDab region; wherein the domain capable of binding to CD3 comprises a first immunoglobulin Fab region, wherein the first immunoglobulin Fab region is formed by a first heavy chain and a first light chain binding to each other, wherein a variable region of the heavy chain has the amino acid sequence as set forth in SEQ ID NO: 8, and a variable region of the light chain has the amino acid sequence as set forth in SEQ ID NO: 10; and the heterodimeric Fc region is composed of two polypeptide chains, wherein each of the polypeptide chains contains opposite-charged amino acid modifications. ### Claim 2 (depends on 1) The bispecific antibody according to claim 1, wherein the domain capable of binding to CD20 comprises a second immunoglobulin Fab region, wherein the second immunoglobulin Fab region is formed by a second heavy chain and a second light chain binding to each other, and the second heavy chain and a CL portion of the second light chain both contain one non-cysteine residue substituted for a native cysteine residue and one cysteine residue substituted for a non-cysteine residue, wherein in the second heavy chain, the non-cysteine residue substituted for a native cysteine residue is C220S, and the cysteine residue substituted for a non-cysteine is L128C; and wherein in the CL portion of the second light chain, the non-cysteine residue substituted for a native cysteine residue is C214S, and the cysteine residue substituted for a non-cysteine is F118C. ### Claim 3 (depends on 2) The bispecific antibody according to claim 2, wherein the first immunoglobulin Fab region does not contain a non-cysteine residue substituted for a native cysteine residue and does not contain a cysteine residue substituted for a non-cysteine residue. ### Claim 4 (depends on 1) The bispecific antibody according to claim 1, wherein the domain capable of binding to CD20 comprises a second immunoglobulin Fab region and the second immunoglobulin Fab region is formed by a second heavy chain and a second light chain binding to each other; and the first heavy chain and the second heavy chain bind to each a other, to form a heterodimeric Fc region. ### Claim 5 (depends on 4) The bispecific antibody according to claim 4, wherein the first heavy chain and the second heavy chain comprise an Fc region selected from the group consisting of a human antibody IgG1, IgG2, IgG3 and IgG4. ### Claim 6 (depends on 4) The bispecific antibody according to claim 4, wherein the second heavy chain has the amino acid sequence as set forth in SEQ ID NO: 1, the second light chain has the amino acid sequence as set forth in SEQ ID NO: 3, the first heavy chain has the amino acid sequence as set forth in SEQ ID NO: 7, and the first light chain has the amino acid sequence as set forth in SEQ ID NO: 9. ### Claim 7 (depends on 1) The bispecific antibody according to claim 1, wherein the domain capable of binding to CD20 comprises a scFv-Fc fusion protein polypeptide chain, wherein the first heavy chain has the amino acid sequence as set forth in SEQ ID NO: 7, and the first light chain has the amino acid sequence as set forth in SEQ ID NO: 9, and the first immunoglobulin heavy chain and the scFv-Fc fusion protein a polypeptide chain bind to each other, to form a heterodimeric Fc region. ### Claim 8 (depends on 7) The bispecific antibody according to claim 7, wherein the first heavy chain has the amino acid as set forth in SEQ ID NO: 7, and a scFv region of the scFv-Fc fusion protein polypeptide chain has the amino acid as set forth in SEQ ID NO: 12, SEQ ID NO: 13 or SEQ ID NO: 14. ### Claim 9 (depends on 7) An antibody conjugate, wherein the antibody conjugate is formed by conjugating a substance to the bispecific antibody according to claim 7. ### Claim 10 (depends on 9) The antibody conjugate according to claim 9, wherein the substance is a cytotoxin, a radioisotope, a fluorescent label, a luminescent substance, a chromogenic substance or an enzyme. ### Claim 11 (depends on 7) A pharmaceutical composition, comprising the bispecific antibody according to claim 7. ### Claim 12 (depends on 1) A method for treating B cell lymphomas, rheumatoid arthritis, multiple sclerosis or systemic lupus erythematosus comprising administering to a subject in need thereof a therapeutically effective amount of the bispecific antibody according to claim 1.