WO2023274342A1 - 特异性结合baff和il-12/23的抗原结合分子及用途 - Google Patents

特异性结合baff和il-12/23的抗原结合分子及用途 Download PDF

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WO2023274342A1
WO2023274342A1 PCT/CN2022/102621 CN2022102621W WO2023274342A1 WO 2023274342 A1 WO2023274342 A1 WO 2023274342A1 CN 2022102621 W CN2022102621 W CN 2022102621W WO 2023274342 A1 WO2023274342 A1 WO 2023274342A1
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seq
amino acid
acid sequence
hcdr1
lcdr1
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French (fr)
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毛浪勇
应华
赖炜明
陶维康
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Shanghai Hengrui Pharmaceutical Co Ltd
Jiangsu Hengrui Pharmaceutical Co Ltd
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Shanghai Hengrui Pharmaceutical Co Ltd
Jiangsu Hengrui Pharmaceutical Co Ltd
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/395—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C07K16/08—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from viruses
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N7/00—Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof

Definitions

  • the present disclosure belongs to the field of biotechnology, and more specifically, the present disclosure relates to an antigen-binding molecule specifically binding to BAFF and IL-12/23 and its application.
  • BAFF B cell activating factor
  • BAFF is a cell activating factor belonging to the TNF family. BAFF is mainly expressed on the surface of the bone marrow cell membrane and exists in the form of trimers. The BAFF on the surface of the cell membrane will be hydrolyzed by protease to form soluble BAFF and enter the blood circulation system. The soluble BAFF has the characteristics of multimerization and can form up to 60-mer . In addition, BAFF can also interact with another protein of the same family, APRIL, to form a heterologous trimer. It is currently known that there are three BAFF receptors on the surface of B cells, namely BAFF-R, BCMA and TACI.
  • BAFF interacts with these three receptors and participates in the differentiation, maturation, survival and regulation of B cells.
  • APRIL and BAFF have two common receptors, namely BCMA and TACI, and APRIL interacts with these two receptors to participate in the survival and regulation of B cells (Samy, E., et al., Int Rev Immunol, 2017.36: p. 3-19; Kamal, A. and M. Khamashta, Autoimmun Rev, 2014.13: p.1094-1101).
  • BAFF is important for maintaining B cell homeostasis, and overactivation of BAFF signaling leads to the survival of self-reactive B cells and the production of autoantibodies that promote autoimmune responses (Cancro, M.P., D.P.D'Cruz, and M.A. Khamashta, J Clin Invest, 2009.119: p.1066-73).
  • IL-12 and IL-23 are two cytokines belonging to the same family. Both IL-12 and IL-23 are heterodimeric proteins composed of two subunits (Moschen, A.R., H. Tilg, and T. Raine, Nat Rev Gastroenterol Hepatol, 2019.16: p.185-196; Frieder , J., et al., Clin Pharmacol Ther, 2018.103: p.88-101). IL-12 is composed of two subunits, p35 and p40, while IL-23 is composed of two subunits, p19 and p40.
  • p40 is a subunit shared by IL-12 and IL-23 (IL-12/23 P40), so antibodies targeting p40 can simultaneously inhibit both IL-12 and IL-23 signaling pathways.
  • the interaction between IL-12 and receptors mainly activates the differentiation of Th1 cells, and at the same time participates in stimulating the secretion of interferon- ⁇ and TNF by various immune cells.
  • IL-23 and receptors mainly activates the differentiation of Th17 cells, and stimulates a variety of cells to secrete cytokines such as IL-17, IL-22 and TNF (Lee, E.B., et al., Cutis, 2018.101: p.5-9 ; Floss, D.M., et al., Cytokine Growth Factor Rev, 2015.26: p.569-578).
  • cytokines produced by the activation of IL-12/23 signaling pathway further participate in the development of diseases such as systemic lupus erythematosus through their own pathways.
  • the present disclosure constructs an antigen-binding molecule that specifically binds BAFF, and constructs an antigen-binding molecule that specifically binds BAFF and IL-12 and/or IL-23.
  • the present disclosure provides an antigen-binding molecule comprising an antigen-binding module 1 that specifically binds BAFF and an antigen-binding module 2 that specifically binds IL-12 and/or IL-23, wherein the antigen-binding module 1 Comprising heavy chain variable region B-VH and light chain variable region B-VL, said B-VH comprises B-HCDR1, B-HCDR2 and B-HCDR3, said B-VL comprises B-LCDR1, B-LCDR2 and B-LCDR3, wherein: said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 63, and said B-LCDR1 , B-LCDR2 and B-LCDR3 respectively comprise the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 64, wherein, SEQ ID NO: 63 is
  • SEQ ID NO: 64 is:
  • X1 is selected from S, A , V or G
  • X2 is selected from A , I, M or S
  • X3 is selected from T or G
  • X4 is selected from Q
  • X5 is selected from N
  • X 6 is selected from L or P
  • X 7 is selected from H
  • X 8 is selected from H or D
  • X 9 is selected from A or G
  • X 10 is selected from S or L
  • X 11 selected from P or S
  • X 12 selected from Q or H
  • X 13 selected from D
  • a or N X 14 selected from S or I
  • X 15 selected from K, R or T
  • X 16 selected from S, W, T or D
  • X 17 is selected from H, Y or S
  • X 18 is selected from Y or R
  • X 19 is selected from G or S
  • X 20 is selected from A or D
  • X 21 is selected from E or S
  • B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2 and B-LCDR3 do not contain the following CDR combination: B-HCDR1, B-HCDR2 and B-HCDR3 respectively contain SEQ ID NO: The amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in 1, and B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprise Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 2 amino acid sequence.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the same numbering convention selected from Kabat, IMGT, Chothia, AbM and Contact.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Kabat numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the IMGT numbering rules.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Chothia numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the AbM numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Contact Numbering Rules.
  • the antigen-binding molecules according to any one of the above, said B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3 are defined according to the Kabat numbering rules, wherein, B-HCDR1 comprises the amino acid sequence NNAIN (SEQ ID NO: 18), B-HCDR2 comprises the amino acid sequence X 1 IX 2 PMFGX 3 AKYSX 4 X 5 FQG (SEQ ID NO: 65), and B-HCDR3 comprises the amino acid sequence SRDX 6 LLFPX 7 X 8 X 9 LX 10 X 11 (SEQ ID NO: 66), B-LCDR1 comprises the amino acid sequence X 12 GX 13 X 14 LX 15 X 16 X 17 X 18 AS (SEQ ID NO: 67), B-LCDR2 Comprising the amino acid sequence GKNNRPS (SEQ ID NO: 32) and B-LCDR3 comprising the amino acid sequence X 19 SRX 20 X 21
  • the antigen binding molecule of any of the above wherein:
  • said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 47
  • said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprising the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 48, or
  • said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 5
  • said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprising the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 12, or
  • said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 6, and said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprising the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 13, or
  • said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 7
  • said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprising the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 14, or
  • said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 8
  • said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprising the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 15, or
  • said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 9
  • said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprising the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 16, or
  • said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequence of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 10
  • said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprising the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 17, or
  • said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequence of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 11
  • said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprise the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO:2.
  • the antigen-binding molecule of any one of the above, said B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2 , Bv-HCDR3, Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 are defined according to the Kabat numbering rules, where,
  • said B-HCDR1 comprises the amino acid sequence of SEQ ID NO: 18
  • said B-HCDR2 comprises the amino acid sequence of SEQ ID NO: 28
  • said B-HCDR3 comprises the amino acid sequence of SEQ ID NO: 29
  • said Said B-LCDR1 comprises the amino acid sequence of SEQ ID NO: 43
  • said B-LCDR2 comprises the amino acid sequence of SEQ ID NO: 32
  • said B-LCDR3 comprises the amino acid sequence of SEQ ID NO: 44;
  • the B-HCDR1 comprises the amino acid sequence of SEQ ID NO: 18
  • the B-HCDR2 comprises the amino acid sequence of SEQ ID NO: 21
  • the B-HCDR3 comprises the amino acid sequence of SEQ ID NO: 20
  • the Said B-LCDR1 comprises the amino acid sequence of SEQ ID NO: 34
  • said B-LCDR2 comprises the amino acid sequence of SEQ ID NO: 32
  • said B-LCDR3 comprises the amino acid sequence of SEQ ID NO: 35;
  • the B-HCDR1 comprises the amino acid sequence of SEQ ID NO: 18
  • the B-HCDR2 comprises the amino acid sequence of SEQ ID NO: 22
  • the B-HCDR3 comprises the amino acid sequence of SEQ ID NO: 23
  • the Said B-LCDR1 comprises the amino acid sequence of SEQ ID NO: 31
  • said B-LCDR2 comprises the amino acid sequence of SEQ ID NO: 32
  • said B-LCDR3 comprises the amino acid sequence of SEQ ID NO: 36;
  • said B-HCDR1 comprises the amino acid sequence of SEQ ID NO: 18
  • said B-HCDR2 comprises the amino acid sequence of SEQ ID NO: 24
  • said B-HCDR3 comprises the amino acid sequence of SEQ ID NO: 20
  • said Said B-LCDR1 comprises the amino acid sequence of SEQ ID NO: 37
  • said B-LCDR2 comprises the amino acid sequence of SEQ ID NO: 32
  • said B-LCDR3 comprises the amino acid sequence of SEQ ID NO: 38;
  • said B-HCDR1 comprises the amino acid sequence of SEQ ID NO: 18
  • said B-HCDR2 comprises the amino acid sequence of SEQ ID NO: 25
  • said B-HCDR3 comprises the amino acid sequence of SEQ ID NO: 20
  • said Said B-LCDR1 comprises the amino acid sequence of SEQ ID NO: 39
  • said B-LCDR2 comprises the amino acid sequence of SEQ ID NO: 32
  • said B-LCDR3 comprises the amino acid sequence of SEQ ID NO: 40;
  • said B-HCDR1 comprises the amino acid sequence of SEQ ID NO: 18
  • said B-HCDR2 comprises the amino acid sequence of SEQ ID NO: 26
  • said B-HCDR3 comprises the amino acid sequence of SEQ ID NO: 27
  • said Said B-LCDR1 comprises the amino acid sequence of SEQ ID NO: 41
  • said B-LCDR2 comprises the amino acid sequence of SEQ ID NO: 32
  • said B-LCDR3 comprises the amino acid sequence of SEQ ID NO: 42;
  • said B-HCDR1 comprises the amino acid sequence of SEQ ID NO: 18
  • said B-HCDR2 comprises the amino acid sequence of SEQ ID NO: 30
  • said B-HCDR3 comprises the amino acid sequence of SEQ ID NO: 20
  • said The B-LCDR1 comprises the amino acid sequence of SEQ ID NO: 31
  • the B-LCDR2 comprises the amino acid sequence of SEQ ID NO: 32
  • the B-LCDR3 comprises the amino acid sequence of SEQ ID NO: 33.
  • SEQ ID NO: 63 is:
  • SEQ ID NO: 64 is:
  • the antigen binding molecule of any one of the above, wherein,
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 47
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 48, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 5
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 12, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 6
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 13, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 7
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 14, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 8
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 15, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 9
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 16, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 10
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 17, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 11
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 2;
  • the B-VH comprises the amino acid sequence of SEQ ID NO: 5, and the B-VL comprises the amino acid sequence of SEQ ID NO: 12; or the B-VH comprises the amino acid sequence of SEQ ID NO: 6 Amino acid sequence, and the B-VL comprises the amino acid sequence of SEQ ID NO: 13; or the B-VH comprises the amino acid sequence of SEQ ID NO: 7, and the B-VL comprises the amino acid sequence of SEQ ID NO: 14 or the B-VH comprises the amino acid sequence of SEQ ID NO: 8, and the B-VL comprises the amino acid sequence of SEQ ID NO: 15; or the B-VH comprises the amino acid sequence of SEQ ID NO: 9, and The B-VL comprises the amino acid sequence of SEQ ID NO: 16; or the
  • the antigen-binding molecule according to any one of the above, wherein the antigen-binding moiety 1 that specifically binds to BAFF is scFv.
  • the scFv molecule has the following structure from N-terminus to C-terminus: VL-linker-VH or VH-linker-VL.
  • the linker is a peptide linker having the structure "L 1 -(GGGGS)nL 2 ", wherein L 1 is a bond, A, GS, GGS or GGGS, and n is 0, 1, 2 , 3, 4, 5, 6, 7, 8, 9 or 10, L2 is a bond, G, GG, GGG or GGGG, and the peptide linker is not a bond.
  • the peptide linker is 3-15 amino acid residues in length.
  • the linker structure is: (G x S) y , wherein x is selected from an integer of 1-5 (eg, 1, 2, 3, 4 or 5), and y is selected from an integer of 1-6 (eg, 1, 2, 3, 4, 5, or 6).
  • the linker is GGGGSGGGGSGGGGS (SEQ ID NO: 70).
  • the P-HCDR1, P-HCDR2, P-HCDR3, P-LCDR1, P-LCDR2, P-LCDR3, Pv-HCDR1, Pv-HCDR2, Pv-HCDR3, Pv-LCDR1, Pv- LCDR2 and Pv-LCDR3 are defined according to the Kabat numbering convention.
  • the P-HCDR1, P-HCDR2, P-HCDR3, P-LCDR1, P-LCDR2, P-LCDR3, Pv-HCDR1, Pv-HCDR2, Pv-HCDR3, Pv-LCDR1, Pv- LCDR2 and PV-LCDR3 are defined according to the IMGT numbering rules.
  • the P-HCDR1, P-HCDR2, P-HCDR3, P-LCDR1, P-LCDR2, P-LCDR3, Pv-HCDR1, Pv-HCDR2, Pv-HCDR3, Pv-LCDR1, Pv- LCDR2 and Pv-LCDR3 are defined according to the Chothia numbering convention.
  • the P-HCDR1, P-HCDR2, P-HCDR3, P-LCDR1, P-LCDR2, P-LCDR3, Pv-HCDR1, Pv-HCDR2, Pv-HCDR3, Pv-LCDR1, Pv- LCDR2 and Pv-LCDR3 are defined according to the AbM numbering convention.
  • the P-HCDR1, P-HCDR2, P-HCDR3, P-LCDR1, P-LCDR2, P-LCDR3, Pv-HCDR1, Pv-HCDR2, Pv-HCDR3, Pv-LCDR1, Pv- LCDR2 and PV-LCDR3 are defined according to the Contact Numbering Rules.
  • the antigen-binding molecule according to any one of the above, said P-HCDR1, P-HCDR2, P-HCDR3, P-LCDR1, P-LCDR2, P-LCDR3 are defined according to the Kabat numbering rules,
  • the P-HCDR1 comprises the amino acid sequence of SEQ ID NO: 53
  • the P-HCDR2 comprises the amino acid sequence of SEQ ID NO: 54
  • the P-HCDR3 comprises the amino acid sequence of SEQ ID NO: 55
  • the P-LCDR1 Comprising the amino acid sequence of SEQ ID NO: 56
  • the P-LCDR2 comprising the amino acid sequence of SEQ ID NO: 57
  • the P-LCDR3 comprising the amino acid sequence of SEQ ID NO: 58.
  • said heavy chain variable region P-VH comprises at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity of the amino acid sequence of the light chain variable region P-VL comprising at least 90% (such as at least 90%, 95%, 96%, 97%) with SEQ ID NO: 52 %, 98%, or 99%) sequence identity of amino acid sequences.
  • the antigen-binding molecule according to any one of the above, the heavy chain variable region P-VH comprises the amino acid sequence of SEQ ID NO: 51, and the light chain variable region P-VL comprises SEQ ID NO: Amino acid sequence of 52.
  • the antigen-binding molecule of any one of the above which comprises a heavy chain constant region CH and a light chain constant region CL.
  • the heavy chain constant region CH is a human IgG heavy chain constant region
  • the light chain constant region CL is a human lambda or kappa light chain constant region.
  • the heavy chain constant region CH is a human IgG1 heavy chain constant region
  • the light chain constant region CL is a human lambda light chain constant region.
  • the heavy chain constant region CH is a human IgG1 heavy chain constant region
  • the light chain constant region CL is a human lambda light chain constant region.
  • the heavy chain constant region CH is a human IgG1 heavy chain constant region
  • the light chain constant region CL is a human kappa light chain constant region.
  • the heavy chain constant region CH comprises the amino acid sequence of SEQ ID NO:59
  • the light chain constant region CL comprises the amino acid sequence of SEQ ID NO:60.
  • the Fc region comprises a first subunit Fc1 and a second subunit Fc2 capable of associating with each other.
  • the Fc region is an IgG Fc region.
  • the Fc region is an IgG 1 Fc region.
  • the Fc region has one or more amino acid substitutions that reduce homodimerization; and/or the Fc region has one or more amino acids that reduce binding of the Fc region to an Fc receptor replace.
  • the Fc region has YTE mutations (M252Y, S254T, and T256E), L234A, L235A mutations, and/or S228P mutations, and the numbering of the mutations is according to the EU index.
  • the Fc region comprises a first subunit and a second subunit capable of associating with each other, the first subunit and/or the second subunit having one or more amino acids that reduce homodimerization replace.
  • the first subunit has a raised structure according to the pestle and socket technique
  • the second subunit has a pore structure according to the pestle and socket technique
  • the first subunit has a pore structure according to the pestle and socket technique
  • the second subunit has a raised structure according to the pestle and socket technique.
  • the first subunit has one or more amino acid substitutions at positions selected from 354, 356, 358, and 366
  • the second subunit has positions selected from 349, 356, 358, 366, One or more amino acid substitutions at positions 368 and 407.
  • the second subunit has one or more amino acid substitutions at positions selected from 354, 356, 358, and 366
  • the first subunit has positions selected from 349, 356, 358, 366, One or more amino acid substitutions at positions 368 and 407.
  • the first subunit has one or more amino acid substitutions selected from 354C, 356E, 358M, and 366W
  • the second subunit has one or more amino acid substitutions selected from 349C, 356E, 358M, 366S, 368A, and 407V One or more amino acid substitutions.
  • the second subunit has one or more amino acid substitutions selected from 354C, 356E, 358M, and 366W
  • the first subunit has one or more amino acid substitutions selected from 349C, 356E, 358M, 366S, 368A, and 407V
  • the first subunit includes amino acid substitutions of 354C, 356E, 358M, and 366W
  • the second subunit includes amino acid substitutions of 349C, 356E, 358M, 366S, 368A, and 407V.
  • the second subunit includes amino acid substitutions of 354C, 356E, 358M, and 366W and the first subunit includes amino acid substitutions of 349C, 356E, 358M, 366S, 368A, and 407V.
  • the antigen-binding molecule is a scFv
  • the antigen-binding module 2 is a scFv comprising a heavy chain variable region P-VH, a light chain variable region P- Full-length antibody of VL, heavy chain constant region CH and light chain constant region CL
  • the antigen-binding module 1 is fused directly or through a linker to the N-terminal of the variable region of the antigen-binding module 2 or the constant region of the antigen-binding module 2
  • the N-terminus of the scFv is directly or through a linker fused to the C-terminus of the heavy chain constant region CH of the antigen binding module 2; in some embodiments, the C-terminus of the scFv is directly Or fused to the N-terminus of the heavy chain variable region P-VH of the antigen-binding module 2 through a linker.
  • the antigen-binding molecule according to any one of the above, said antigen-binding molecule comprising a first chain having a structure represented by formula (a) and a second chain having a structure represented by formula (b); or Comprising a first chain having a structure shown in formula (c) and a second chain having a structure shown in formula (b), wherein,
  • linker 1 and linker 2 are the same or different peptide linkers.
  • the peptide linker independently has the structure L 1 -(GGGGS)nL 2 , wherein L 1 is a bond, A, GS, GGS or GGGS, and n is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, L2 is a bond, G, GG, GGG or GGGG, and the peptide linker is not a bond.
  • the peptide linker is 3-15 amino acid residues in length.
  • the linker 1 and the linker 2 are each independently selected from a linker of formula (d), wherein formula (d): (G x S) y , wherein x is selected from an integer of 1-5 ( For example 1, 2, 3, 4 or 5), y is selected from an integer of 1-6 (for example, 1, 2, 3, 4, 5 or 6).
  • the linker 1 is selected from: GGGS (SEQ ID NO: 69)
  • the linker 2 is selected from: GGGGSGGGGSGGGGS (SEQ ID NO: 70).
  • the antigen-binding molecule wherein the first chain comprises at least 90% (eg, at least 90%, 95%, 96%, 97%, 98%, or 99%) of SEQ ID NO: 61 %) sequence identity, and said second strand comprises at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 50 or said first strand comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 62, and The second strand comprises an amino acid sequence having at least 90% (eg, at least 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:50.
  • the antigen binding molecule has a first strand comprising the amino acid sequence of SEQ ID NO:61 and a second strand comprising the amino acid sequence of SEQ ID NO:50, or comprising the amino acids of SEQ ID NO:62 A first strand of the sequence and a second strand comprising the amino acid sequence of SEQ ID NO:50.
  • the antigen binding molecule has:
  • the present disclosure provides an antigen-binding molecule that specifically binds to a BAFF antigen, comprising a heavy chain variable region B-VH and a light chain variable region B-VL, the B-VH comprising B-HCDR1, B-HCDR2 and B-HCDR3, the B-VL comprises B-LCDR1, B-LCDR2 and B-LCDR3, wherein: the B-HCDR1, B-HCDR2 and B-HCDR3 comprise SEQ ID NO:63 respectively The amino acid sequence of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3, and the B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprise the Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO:64 Amino acid sequence, wherein, SEQ ID NO: 63 is:
  • SEQ ID NO: 64 is:
  • X1 is selected from S, A , V or G
  • X2 is selected from A , I, M or S
  • X3 is selected from T or G
  • X4 is selected from Q
  • X5 is selected from N
  • X 6 is selected from L or P
  • X 7 is selected from H
  • X 8 is selected from H or D
  • X 9 is selected from A or G
  • X 10 is selected from S or L
  • X 11 selected from P or S
  • X 12 selected from Q or H
  • X 13 selected from D
  • a or N X 14 selected from S or I
  • X 15 selected from K, R or T
  • X 16 selected from S, W, T or D
  • X 17 is selected from H, Y or S
  • X 18 is selected from Y or R
  • X 19 is selected from G or S
  • X 20 is selected from A or D
  • X 21 is selected from E or S
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the same numbering convention selected from Kabat, IMGT, Chothia, AbM and Contact.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Kabat numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the IMGT numbering rules.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Chothia numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the AbM numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Contact Numbering Rules.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3 are defined according to the Kabat numbering rules, wherein the B-HCDR1 comprises the amino acid sequence NNAIN( SEQ ID NO: 18), B-HCDR2 comprises the amino acid sequence X 1 IX 2 PMFGX 3 AKYSX 4 X 5 FQG (SEQ ID NO: 65), and B-HCDR3 comprises the amino acid sequence SRDX 6 LLFPX 7 X 8 X 9 LX 10 X 11 (SEQ ID NO: 66), B-LCDR1 comprises the amino acid sequence X 12 GX 13 X 14 LX 15 X 16 X 17 X 18 AS (SEQ ID NO: 67), B-LCDR2 comprises the amino acid sequence GKNNRPS (SEQ ID NO: 32 ) and B-LCDR3 comprising the amino acid sequence X 19 SRX 20 X 21 X 22 GX 23 X 24 WX 25 (SEQ ID
  • the antigen-binding molecule that specifically binds to a BAFF antigen as described in any one of the above, wherein: (i) said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise Bv in SEQ ID NO:47 - the amino acid sequence of HCDR1, Bv-HCDR2 and Bv-HCDR3, and said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprise the amino acids of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 48 sequence, or (ii) said B-HCDR1, B-HCDR2 and B-HCDR3 respectively comprise the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 in SEQ ID NO: 5, and said B-LCDR1, B-LCDR2 and B-LCDR3 respectively comprise the amino acid sequences of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 12, or (i) said B-
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Kabat numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the IMGT numbering rules.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Chothia numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the AbM numbering convention.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv- LCDR2 and Bv-LCDR3 are defined according to the Contact Numbering Rules.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3 are defined according to the Kabat numbering rules, wherein (i) the B-HCDR1 comprising the amino acid sequence of SEQ ID NO: 18, the B-HCDR2 comprising the amino acid sequence of SEQ ID NO: 28, the B-HCDR3 comprising the amino acid sequence of SEQ ID NO: 29, and the B-LCDR1 comprising the amino acid sequence of SEQ ID NO : the amino acid sequence of 43, the B-LCDR2 comprises the amino acid sequence of SEQ ID NO: 32, and the B-LCDR3 comprises the amino acid sequence of SEQ ID NO: 44; or (ii) the B-HCDR1 comprises the amino acid sequence of SEQ ID NO : the amino acid sequence of 18, the B-HCDR2 comprises the amino acid sequence of SEQ ID NO: 21, the B-HCDR3 comprises the amino acid sequence of SEQ ID NO: 20, and the B-LCDR1 comprises the amino acid sequence
  • the antigen-binding molecule that specifically binds to a BAFF antigen according to any one of the above, wherein the B-VH comprises an antigen-binding molecule having at least 90% (e.g., at least 90%, 95%, 96%) of SEQ ID NO: 63 %, 97%, 98%, or 99%) sequence identity
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98%) sequence identity to SEQ ID NO: 64 % or 99%) sequence identity of the amino acid sequence, wherein SEQ ID NO: 63 is:
  • SEQ ID NO: 64 is:
  • the antigen-binding molecule that specifically binds to a BAFF antigen as described in any one of the above,
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 47
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 48, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 5
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 12, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 6
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 13, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 7
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 14, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 8
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 15, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 9
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 16, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 10
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 17, or
  • said B-VH comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 11
  • said B-VL comprises an amino acid sequence having at least 90% (e.g., at least 90%, 95%, 96%, 97%, 98% or 99%) sequence identity to SEQ ID NO: 2;
  • the antigen-binding molecule that specifically binds to the BAFF antigen as described in any one of the above comprises the amino acid sequence of SEQ ID NO: 47 and the B-VL comprises the amino acid of SEQ ID NO: 48 sequence, or the B-VH comprises the amino acid sequence of SEQ ID NO: 5, and the B-VL comprises the amino acid sequence of SEQ ID NO: 12; or the B-VH comprises the amino acid sequence of SEQ ID NO: 6, And the B-VL comprises the amino acid sequence of SEQ ID NO: 13; or the B-VH comprises the amino acid sequence of SEQ ID NO: 7, and the B-VL comprises the amino acid sequence of SEQ ID NO: 14; or the The B-VH comprises the amino acid sequence of SEQ ID NO:8, and the B-VL comprises the amino acid sequence of SEQ ID NO:15; or the B-VH comprises the amino acid sequence of SEQ ID NO:9, and the B-VL comprises the amino acid sequence of SEQ ID NO:9
  • the antigen-binding molecule that specifically binds to a BAFF antigen as described in any one of the above includes a heavy chain constant region CH and a light chain constant region CL.
  • the heavy chain constant region CH is a human IgG heavy chain constant region
  • the light chain constant region CL is a human lambda or kappa light chain constant region.
  • the heavy chain constant region CH is a human IgG1 heavy chain constant region
  • the light chain constant region CL is a human lambda light chain constant region.
  • the heavy chain constant region CH is a human IgG1 heavy chain constant region
  • the light chain constant region CL is a human kappa light chain constant region.
  • the heavy chain constant region CH comprises the amino acid sequence of SEQ ID NO:45
  • the light chain constant region CL comprises the amino acid sequence of SEQ ID NO:46.
  • the antigen-binding molecule that specifically binds a BAFF antigen is a full-length antibody.
  • the heavy chain constant region of the antigen-binding molecule that specifically binds a BAFF antigen has one or more amino acid substitutions that reduce the binding of its Fc region to an Fc receptor.
  • the Fc region has YTE mutations (M252Y, S254T, and T256E), L234A, L235A mutations, and/or S228P mutations, and the numbering of the mutations is according to the EU index.
  • an antigen-binding molecule as described in any one of the above for example, an antigen-binding molecule comprising an antigen-binding moiety 1 that specifically binds BAFF and an antigen-binding moiety 2 that specifically binds IL-12 and/or IL-23, Or an antigen-binding molecule that specifically binds to a BAFF antigen
  • the antigen-binding molecule has at least one of the following characteristics (for example, comprising any 1, 2, 3, 4, 5, 6 or 7 characteristics of the following A-G):
  • the antigen-binding molecule can block the binding of human BAFF to human BAFF-R; preferably, the IC50 value for blocking the binding of human BAFF to human BAFF-R is less than 11 nM (for example, less than 11 nM, less than or equal to 10 nM, less than or equal to Equal to 9nM, less than or equal to 8nM, less than or equal to 7nM, less than or equal to 6nM, less than or equal to 5nM or less), the IC50 value is detected by Elisa method; in some embodiments, the IC50 value is tested according to the present disclosure Example 2 method detection;
  • the antigen-binding molecule can inhibit BAFF-induced B cell proliferation; preferably, the antigen-binding molecule can inhibit BAFF-induced B cell proliferation; , less than or equal to 0.10 nM, less than or equal to 0.09 nM, less than or equal to 0.08 nM, less than or equal to 0.07 nM, less than or equal to 0.06 nM or less) inhibits BAFF-induced B cell proliferation; in some embodiments , the IC50 value is detected according to the method of Test Example 3 of the present disclosure;
  • the antigen binding molecule can be less than 9.99E-10M (such as less than 9.99E-10M, less than or equal to 9.00E-10M, less than or equal to 8.20E-10M, less than or equal to 7.40E-10M, less than or equal to 6.60E -10M, less than or equal to 5.20E-10M, less than or equal to 4.00E-10M, less than or equal to 3.00E-10M or less) combined with human BAFF, the KD value is detected by the Biacore method; in some implementations In the method, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can be combined with cynomolgus BAFF with a KD value of less than 5.00E-10M (for example, less than or equal to 4.00E-10M, less than or equal to 3.00E-10M or less), and the KD value is passed Biacore method detection; In some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can bind to mouse BAFF with a KD value of less than 2.20E-10M (for example, less than or equal to 2.00E-10M, less than or equal to 1.00E-10M or less), and the KD value is determined by the Biacore method Detection; In some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can block the binding of human BAFF to human BCMA; preferably, the IC50 value of blocking the binding of human BAFF to human BCMA is less than 0.9nM (for example, less than or equal to 0.4nM, less than or equal to 0.35nM, less than or equal to or equal to 0.31nM or less), the IC50 value is detected by Elisa method; in some embodiments, the IC50 value is detected according to the method of Test Example 2 of the present disclosure; or
  • the antigen-binding molecule can block the binding of human BAFF to human TACI; preferably, the IC50 value of blocking the binding of human BAFF to human TACI is less than 0.6nM (for example, less than or equal to 0.55nM, less than or equal to 0.4nM, less than or equal to or equal to 0.33nM, less than or equal to 0.23nM or less), the IC50 value is detected by the Elisa method; in some embodiments, the IC50 value is detected according to the method of Test Example 2 of the present disclosure.
  • the antigen-binding molecule according to any one of the above, said antigen-binding molecule comprising an antigen-binding moiety 1 that specifically binds BAFF and an antigen-binding moiety 2 that specifically binds IL-12 and/or IL-23 , which has at least one of the following characteristics (for example, including any 1, 2, 3, 4 or 5 characteristics of the following H-L):
  • the antigen-binding molecule can block the binding of human IL-12/23 p40 to human IL-12R ⁇ 1; preferably, the IC50 value for blocking the binding of human IL-12/23 p40 to human IL-12R ⁇ 1 is less than 3.6nM ( For example, less than or equal to 3.0nM, less than or equal to 2.0nM, less than or equal to 1.0nM, less than or equal to 0.5nM, less than or equal to 0.23nM or less), the IC50 value is detected by the Elisa method; in some embodiments, The IC50 value is detected according to the method of Test Example 2 of the present disclosure;
  • the antigen-binding molecule can inhibit IL-12-induced IFN ⁇ secretion; preferably, the IC50 value of the antigen-binding molecule inhibiting IL-12-induced IFN ⁇ secretion is less than 3.0nM (for example, less than or equal to 2.6nM, less than or equal to 1.6nM or less) ; In some embodiments, the IC50 value is detected according to the method of Test Example 4 of the present disclosure;
  • J. Antigen-binding molecules can inhibit IL-23-induced IL-17 secretion; preferably, the antigen-binding molecules inhibit IL-23-induced IL-17 secretion with an IC50 value of less than 0.034nM (for example, less than or equal to 0.031nM, less than or equal to 0.030nM or less); In some embodiments, the IC50 value is detected according to the method of Test Example 4 of the present disclosure;
  • the antigen-binding molecule can bind to cynomolgus monkey IL-12/23 p40 with a KD value less than 2.7E-10M (for example, less than or equal to 2.0E-10M, less than or equal to 1.0E-10M or less), said The KD value is detected by the Biacore method; in some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure; or
  • the antigen-binding molecule can bind to human IL-23 with a KD value of less than 6.4E-11M (e.g., less than or equal to 6.3E-11M, less than or equal to 6.2E-11M or less), the KD value is determined by the Biacore method Detection; In some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure.
  • the present disclosure provides a pharmaceutical composition
  • a pharmaceutical composition comprising: a therapeutically effective amount of any one of the above antigen-binding molecules, and one or more pharmaceutically acceptable carriers, diluents, buffers or excipient.
  • the present disclosure provides an isolated nucleic acid encoding an antigen binding molecule according to any one of the above.
  • the present disclosure provides a host cell comprising the nucleic acid of any one of the above.
  • the present disclosure provides a method for treating a disease, the method comprising the step of administering the antigen-binding molecule or the pharmaceutical composition according to any one of the above to a subject.
  • the disease is a B cell disorder or an autoimmune disease.
  • the B cell disorder or autoimmune disease is a disease or condition associated with BAFF expression.
  • the autoimmune disease is selected from the group consisting of: systemic lupus erythematosus, myasthenia gravis, multiple sclerosis, insulin-dependent diabetes mellitus, Crohn's disease, rheumatoid arthritis, polyarticular juvenile rheumatoid Arthritis and psoriatic arthritis; said B-cell disorder is selected from the group consisting of neoplasms, chronic leukaemia, multiple myeloma, non-Hodgkin's lymphoma, post-transplantation lymphoproliferative disease, and light chain gammopathies .
  • the autoimmune disease is systemic lupus erythematosus.
  • the present disclosure also provides the use of the antigen-binding molecule or the pharmaceutical composition according to any one of the above in the preparation of a medicament for treating the aforementioned diseases.
  • the present disclosure also provides an antigen-binding molecule or a pharmaceutical composition as described above for use as a medicament.
  • Figure 1 Schematic diagram of the structure of BU-1.
  • Figure 2 Schematic diagram of the structure of BU-2.
  • Figure 3 The result of the experiment of inhibiting TNF ⁇ secretion by the bispecific antibody specifically binding to BAFF and IL-12/23.
  • Figure 4 The results of the IgA secretion inhibition experiment by the bispecific antibody specifically binding to BAFF and IL-12/23.
  • Figure 5 The result of the experiment of inhibiting IL-22 secretion by the bispecific antibody specifically binding to BAFF and IL-12/23.
  • Figure 6 The result of the experiment of inhibiting the secretion of IFN ⁇ by the bispecific antibody specifically binding to BAFF and IL-12/23.
  • cytokine is a general term for proteins/polypeptides released by a population of cells that act as intercellular mediators on other cells.
  • cytokines include lymphokines, monokines, chemokines and traditional polypeptide hormones.
  • exemplary cytokines include: IL-2, IFN- ⁇ , IL-6, TNF ⁇ , IL-17, and IL-5.
  • IL-12 and/or IL-23 or “IL-12/23” all mean to cover: “IL-12”, “IL-23” and “IL-12 and IL-23” -23" each.
  • amino acid refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to naturally occurring amino acids.
  • Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, eg, hydroxyproline, gamma-carboxyglutamic acid, and O-phosphoserine.
  • Amino acid analogs are compounds that have the same basic chemical structure (i.e., the alpha carbon bonded to a hydrogen, carboxyl, amino group, and R group) as a naturally occurring amino acid, such as homoserine, norleucine, methionine sulfoxide , Methylsulfonium methionine.
  • Such analogs have modified R groups (eg, norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid.
  • An amino acid mimetic refers to a chemical compound that has a structure that differs from the general chemical structure of an amino acid, but functions in a manner similar to a naturally occurring amino acid.
  • amino acid mutation includes amino acid substitutions (also called amino acid substitutions), deletions, insertions and modifications. Any combination of substitutions, deletions, insertions and modifications can be made to achieve the final construct so long as the final construct possesses the desired properties, such as reduced or binding to Fc receptors.
  • Amino acid sequence deletions and insertions include deletions and insertions at the amino and/or carboxyl termini of the polypeptide chain.
  • Specific amino acid mutations may be amino acid substitutions.
  • the amino acid mutation is a non-conservative amino acid substitution, that is, replacing one amino acid with another amino acid having different structural and/or chemical properties.
  • Amino acid substitutions include substitutions with non-naturally occurring amino acids or with derivatives of the 20 natural amino acids (e.g., 4-hydroxyproline, 3-methylhistidine, ornithine, homoserine, 5-hydroxylysine) .
  • Amino acid mutations can be generated using genetic or chemical methods well known in the art. Genetic methods can include site-directed mutagenesis, PCR, gene synthesis, and the like. Methods of altering amino acid side chain groups other than genetic engineering, such as chemical modification, may also be available. Various names may be used herein to refer to the same amino acid mutation.
  • the amino acid residue at a specific position can be expressed in the form of position + amino acid residue, for example, 366W means that the amino acid residue at position 366 is W. T366W means that the amino acid residue at the 366th position is mutated from the original T to W.
  • antigen-binding molecule is used in the broadest sense and covers various molecules that specifically bind to an antigen, including but not limited to antibodies, other polypeptides with antigen-binding activity, and antibody fusion proteins fused thereto.
  • the antigen-binding molecules herein are bispecific antigen-binding molecules (eg, bispecific antibodies).
  • bispecific antigen binding molecule refers to an antigen binding molecule capable of specifically binding to two different antigens or at least two different epitopes of the same antigen.
  • antibody is used in the broadest sense and encompasses various antibody structures including, but not limited to, monoclonal antibodies, polyclonal antibodies; monospecific antibodies, multispecific antibodies (e.g., bispecific antibodies); full-length antibodies and antibody Fragments (or antigen-binding fragments, or antigen-binding portions) as long as they exhibit the desired antigen-binding activity.
  • Native antibody refers to a naturally occurring immunoglobulin molecule. For example, native IgG antibodies are heterotetrameric proteins of approximately 150,000 Daltons, composed of two light chains and two heavy chains joined by disulfide bonds.
  • each heavy chain has a variable region (VH), also known as variable heavy domain, heavy chain variable region, followed by a heavy chain constant region (CH), the native IgG heavy chain constant region is usually Contains three constant domains (CH1, CH2 and CH3).
  • VH variable heavy domain
  • CH heavy chain constant region
  • each light chain has a variable region (VL, also known as variable light domain, or light chain variable domain), followed by a constant light domain (light chain constant region, CL ).
  • VH variable heavy domain
  • CH2 and CH3 heavy chain constant region
  • CL constant light domain
  • full-length antibody “intact antibody” and “whole antibody” are used interchangeably herein to refer to an antibody having a structure substantially similar to that of a native antibody or having a heavy chain with an Fc region as defined herein.
  • Natural complete antibody light chain includes light chain variable region VL and constant region CL, VL is at the amino terminal of light chain, light chain constant region includes ⁇ chain and ⁇ chain; heavy chain includes variable region VH and constant region (CH1, CH2 and CH3), VH is at the amino-terminus (also called N-terminus) of the heavy chain, and the constant region is at the carboxy-terminus (also called C-terminus), where CH3 is closest to the carboxy-terminus of the polypeptide, and the heavy chain can belong to any isotype, including IgG (including IgG1, IgG2, IgG3 and IgG4 subtypes), IgA (including IgA1 and IgA2 subtypes), IgM and IgE.
  • IgG including IgG1, IgG2, IgG3 and IgG4 subtypes
  • IgA including IgA1 and IgA2 subtypes
  • IgM and IgE IgE.
  • bispecific antibody refers to an antibody (including an antibody or an antigen-binding fragment thereof, such as a single-chain antibody) capable of specifically binding to two different antigens or two different epitopes of the same antigen.
  • Bispecific antibodies of various structures have been disclosed in the prior art. According to the integrity of the IgG molecule, it can be divided into IgG-like bispecific antibodies and antibody fragment bispecific antibodies. According to the number of antigen-binding regions, bispecific antibodies can be divided into bivalent, trivalent, tetravalent or more valent. According to whether the structure is symmetrical, it can be divided into symmetrical structure bispecific antibody and asymmetric structure bispecific antibody.
  • fragment-type bispecific antibodies such as Fab fragments lacking Fc fragments, which form bispecific antibodies by combining two or more Fab fragments in one molecule, have low immunogenicity and small molecular weight , with high tumor tissue permeability, typical antibody structures of this type such as F(ab) 2 , scFv-Fab, (scFv) 2 -Fab, etc.
  • IgG-like bispecific antibody for example, with Fc fragment
  • this type of antibody has a relatively large molecular weight, and the Fc fragment helps to purify the antibody and improve its solubility and stability.
  • the Fc part may also bind to the receptor FcRn, Increase antibody serum half-life.
  • bispecific antibodies such as KiH, CrossMAb, Triomab quadroma, Fc ⁇ Adp, ART-Ig, BiMAb, Biclonics, BEAT, DuoBody, Azymetric, XmAb, 2:1 TCBs, 1Fab-IgG TDB, FynomAb, two-in- one/DAF, scFv-Fab-IgG, DART-Fc, LP-DART, CODV-Fab-TL, HLE-BiTE, F(ab)2-CrossMAb, IgG-(scFv)2, Bs4Ab, DVD-Ig, Tetravalent - Bispecific antibodies such as DART-Fc, (scFv)4-Fc, CODV-Ig, mAb2, F(ab)4-CrossMAb (see Aran F. Labrijn et al., Nature Reviews Drug Discovery volume 18, pages 585–608 (2019 ); Chen S1 et al., J
  • variable region refers to the antigen-binding domain of an antigen-binding molecule (eg, an antibody).
  • the heavy chain variable region VH and the light chain variable region VL each comprise four conserved framework regions (FRs) and three complementarity determining regions (CDRs).
  • FRs conserved framework regions
  • CDRs complementarity determining regions
  • the heavy chain variable region in the antigen-binding module that specifically binds BAFF is marked as B-VH
  • the light chain variable region is marked as B-VL
  • the antigen-binding module that specifically binds IL-12/23 p40 The heavy chain variable region is denoted as P-VH and the light chain variable region as P-VL.
  • CDR complementarity determining region
  • frame or “FR” refers to the variable domain residues other than the CDR residues.
  • VH contains 3 CDR regions: HCDR1, HCDR2 and HCDR3;
  • VL contains 3 CDR regions: LCDR1, LCDR2 and LCDR3.
  • Each VH and VL consists of three CDRs and four FRs arranged in the following order from the amino terminus (also known as the N terminus) to the carboxyl terminus (also known as the C terminus): FR1, CDR1, FR2, CDR2, FR3, CDR3 , FR4.
  • Each VH and VL consists of three CDRs and four FRs arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4.
  • a single VH or VL may be sufficient to confer antigen binding specificity.
  • the three CDR regions in B-VH are respectively marked as B-HCDR1, B-HCDR2 and B-HCDR3, or Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3; the three CDR regions in B-VL Labeled as B-LCDR1, B-LCDR2, and B-LCDR3, or Bv-LCDR1, Bv-LCDR2, and Bv-LCDR3, respectively.
  • the three CDR regions in P-VH are respectively marked as P-HCDR1, P-HCDR2 and P-HCDR3, or Pv-HCDR1, Pv-HCDR2 and Pv-HCDR3; the three CDR regions in P-VL are respectively marked as P-LCDR1, P-LCDR2, and P-LCDR3, or Pv-LCDR1, Pv-LCDR2, and Pv-LCDR3.
  • amino acid sequence boundaries of CDRs can be determined by various known schemes, for example: “Kabat” numbering convention (see Kabat et al. (1991), “Sequences of Proteins of Immunological Interest", 5th Edition, Public Health Service, National Institutes of Health , Bethesda, MD), “Chothia” numbering sequence, “ABM” numbering sequence, "contact” numbering sequence (see Martin, ACR. Protein Sequence and Structure Analysis of Antibody Variable Domains [J].
  • the Kabat numbering convention is used for variable region and CDR sequences in this disclosure. Although in specific embodiments, the Kabat numbering rules are used to define amino acid residues, the corresponding technical solutions in other numbering systems will be regarded as equivalent technical solutions.
  • antibody fragment refers to a molecule other than an intact antibody that comprises a portion of an intact antibody that retains the antigen-binding ability of the intact antibody.
  • antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab') 2 , single domain antibody, single chain Fab (scFab), diabody, linear antibody, single chain antibody (scFv) , and multispecific antibodies formed from antibody fragments.
  • Fc region or “fragment crystallizable region” is used to define the C-terminal region of an antibody heavy chain, including native and engineered Fc regions.
  • the Fc region comprises the same or different two subunits.
  • the Fc region of a human IgG heavy chain is defined as extending from the amino acid residue at position Cys226 or from Pro230 to its carboxyl terminus.
  • Suitable Fc regions for use in the antibodies described herein include the Fc regions of human IgGl, IgG2 (IgG2A, IgG2B), IgG3 and IgG4.
  • the boundaries of the Fc region can also be varied, such as deletion of the C-terminal lysine of the Fc region (residue 447 according to the EU numbering system) or deletion of the C-terminal glycine and lysine of the Fc region (residue 447 according to the EU numbering system). system residues 446 and 447).
  • the numbering convention for the Fc region is the EU numbering system, also known as the EU index.
  • chimeric antibody refers to an antibody in which a portion of the heavy and/or light chains is from a particular source or species and the remaining portion of the heavy and/or light chains is from another, different source or species.
  • humanized antibody is an antibody that retains the reactivity of a non-human antibody while being less immunogenic in humans. Humanization can be achieved, for example, by retaining the non-human CDR regions and replacing the remainder of the antibody with their human counterparts (ie, the constant regions and the framework portion of the variable regions).
  • human antibody “human antibody”, “fully human antibody”, and “fully human antibody” are used interchangeably to refer to antibodies whose variable and constant regions are human sequences.
  • the term encompasses antibodies that are derived from human genes but have, for example, altered sequences that reduce potential immunogenicity, increase affinity, eliminate cysteines or glycosylation sites that might cause undesired folding.
  • the term encompasses such antibodies produced recombinantly in non-human cells which may confer glycosylation not characteristic of human cells.
  • the term also encompasses antibodies that have been raised in transgenic mice containing some or all of the immunoglobulin heavy and light chain loci.
  • the meaning of human antibody expressly excludes humanized antibodies comprising non-human antigen-binding residues.
  • affinity refers to the overall strength of the non-covalent interaction between a single binding site of a molecule (eg, an antigen-binding molecule of the disclosure) and its binding partner (eg, an antigen).
  • binding affinity refers to internal binding affinity, which reflects a 1:1 interaction between members of a binding pair (eg, antibody and antigen).
  • KD dissociation constant
  • the term “kassoc” or “ka” refers to the on-rate of a particular antibody-antigen interaction and the term “kdis” or “kd” refers to the dissociation rate of a particular antibody-antigen interaction.
  • KD refers to the dissociation constant, which is obtained from the ratio of kd to ka (ie, kd/ka) and is expressed as molarity (M).
  • M molarity
  • the KD value of an antibody can be determined using methods well known in the art. For example, surface plasmon resonance is measured using biosensing systems such as the system, or affinity in solution is measured by solution equilibrium titration (SET). In some embodiments, the KD value is detected by Biacore.
  • effector function refers to those biological activities attributable to an antibody Fc region (either native sequence Fc region or amino acid sequence mutated Fc region) and which vary with the antibody isotype.
  • antibody effector functions include, but are not limited to: C1q binding and complement-dependent cytotoxicity, Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, cell surface receptors (e.g., B cell receptors, body) downregulation; and B cell activation.
  • the term "monoclonal antibody” refers to a population of antibodies or members thereof that are substantially homogeneous, ie, the amino acid sequences of the antibody molecules comprised in the population are identical except for natural mutations that may be present in minor amounts.
  • polyclonal antibody preparations typically comprise multiple different antibodies with different amino acid sequences in their variable domains, often specific for different epitopes.
  • “Monoclonal” denotes the characteristics of an antibody obtained from a substantially homogeneous population of antibodies and should not be construed as requiring that the antibody be produced by any particular method.
  • the antibodies provided by the present disclosure are monoclonal antibodies.
  • antigen refers to a molecule or portion of a molecule capable of being selectively bound by an antigen-binding molecule (eg, an antibody).
  • An antigen may have one or more epitopes capable of interacting with different antigen-binding molecules (eg, antibodies).
  • epitope refers to an area (area or region) on an antigen capable of specifically binding to an antibody or antigen-binding fragment thereof.
  • An epitope may be formed from contiguous amino acids (linear epitope) or comprise non-contiguous amino acids (conformational epitope), such that non-contiguous amino acids are brought into spatial proximity by folding of the antigen, ie by tertiary folding of the antigen.
  • the difference between a conformational epitope and a linear epitope is that antibody binding to a conformational epitope is lost in the presence of denaturing solvents.
  • An epitope comprises at least 3, at least 4, at least 5, at least 6, at least 7, or 8-10 amino acids in a unique spatial conformation.
  • Antibodies that bind a particular epitope can be screened using methods routine in the art, such as, but not limited to, alanine scanning, peptide blotting, peptide cleavage analysis, epitope excision, epitope extraction, antigen Chemical modification of (see Prot. Sci.9 (2000) 487-496), and cross-blocking.
  • an antigen-binding molecule eg, an antibody
  • an antibody binds an antigen or an epitope thereof with an equilibrium dissociation constant (KD) of about 1 ⁇ 10 ⁇ 7 M or less (eg, about 1 ⁇ 10 ⁇ 8 M or less).
  • KD equilibrium dissociation constant
  • the antibody binds an antigen with a KD that is 10% or less (eg, 1%) of the antibody's KD for binding to a non-specific antigen (eg, BSA, casein).
  • KD can be measured using known methods, for example by measured by surface plasmon resonance.
  • an antibody that specifically binds to an antigen or an epitope thereof does not exclude cross-reactivity to other related antigens, e.g.
  • the corresponding antigens of cynomolgus, cyno), chimpanzee (Pan troglodytes) (chimpanzee, chimp)) or marmoset (Callithrix jacchus) (commonmarmoset, marmoset) are cross-reactive.
  • antigen binding moiety refers to a polypeptide molecule that specifically binds an antigen of interest.
  • Antigen binding moieties include antibodies and fragments thereof as otherwise defined herein.
  • the antigen binding moiety comprises an antigen binding domain of an antibody comprising an antibody heavy chain variable region and an antibody light chain variable region.
  • antigen-binding moiety that specifically binds BAFF refers to an antigen-binding moiety that is capable of binding BAFF with sufficient affinity.
  • the antigen binding moiety that specifically binds to human BAFF has an equilibrium dissociation constant (KD) of less than 10.00E-10M (e.g., less than 9.99E-10M, less than or equal to 9.00E-10M, less than or equal to 8.20E-10M, less than or equal to 7.40E-10M, less than or equal to 6.60E-10M, less than or equal to 5.20E-10M, less than or equal to 4.00E-10M, less than or equal to 3.00E-10M or less), which is measured by the Biacore method.
  • KD equilibrium dissociation constant
  • an antigen binding moiety that specifically binds BAFF binds a conserved epitope in BAFF from a different species.
  • antigen-binding moiety that specifically binds IL-12/23 refers to an antigen-binding moiety capable of binding IL-12/23 with sufficient affinity.
  • the antigen binding moiety that specifically binds IL-12/23 has an equilibrium dissociation constant (KD) of less than 6.4E-11M (e.g., less than or equal to 6.3E-11M, less than or equal to 6.2E -11M or less) for binding to human IL-23, which was detected by the Biacore method.
  • the antigen binding moiety that specifically binds IL-12/23 binds a conserved epitope in IL-12/23 from a different species.
  • Antigen binding moieties include antibody fragments as defined herein, eg Fab or scFv.
  • linker refers to a linking unit that joins two polypeptide fragments.
  • linkers appearing in the same structure may be the same or different.
  • the linker may be a peptide linker comprising one or more amino acids, typically about 1-30, 2-24 or 3-15 amino acids.
  • the linkers used herein may be the same or different.
  • “-" appears in the structural formula it means that the units on both sides are directly connected by covalent bonds.
  • bond appears in a structural unit, it means that the unit has no amino acids, and the units on either side of the unit are directly connected.
  • antibody-dependent cellular cytotoxicity is mechanisms for inducing cell death that rely on the interaction of antibody-coated target cells with lytically active effector cells (such as natural killer cells (NK), monocytes, macrophages, and neutrophils) via Fc ⁇ receptors (Fc ⁇ Rs) expressed on effector cells.
  • lytically active effector cells such as natural killer cells (NK), monocytes, macrophages, and neutrophils
  • Fc ⁇ receptors Fc ⁇ receptors
  • NK cells express FcyRIIIa
  • monocytes express FcyRI, FcyRII, and FcyRIIIa.
  • the ADCC activity of the antibodies provided herein can be assessed using an in vitro assay using antigen-expressing cells as target cells and NK cells as effector cells. Cell lysis is detected based on labels released from lysed cells, such as radioactive substrates, fluorescent dyes, or native intracellular proteins.
  • ADCP antibody-dependent cellular phagocytosis
  • complement-dependent cytotoxicity refers to a mechanism that induces cell death in which the Fc effector domains of bound antibodies on target cells bind and activate complement component C1q, which in turn activates the complement cascade, resulting in target cell die. Activation of complement can also result in the deposition of complement components on the surface of target cells that promote CDC by binding to complement receptors (eg, CR3) on leukocytes.
  • complement receptors eg, CR3
  • nucleic acid is used herein interchangeably with the term “polynucleotide” and refers to deoxyribonucleotides or ribonucleotides and polymers thereof in single- or double-stranded form.
  • the term encompasses nucleic acids containing known nucleotide analogs or modified backbone residues or linkages, synthetic, naturally occurring and non-naturally occurring, having similar binding properties to the reference nucleic acid, and defined in Metabolized in a manner similar to the reference nucleotide.
  • Examples of such analogs include, but are not limited to, phosphorothioate, phosphoramidate, methylphosphonate, chiral-methylphosphonate, 2-O-methyl ribonucleotides, peptide-nucleic acid (PNA ).
  • An "isolated" nucleic acid refers to a nucleic acid molecule that has been separated from components of its natural environment.
  • An isolated nucleic acid includes a nucleic acid molecule contained in a cell that normally contains the nucleic acid molecule, but which is present extrachromosomally or at a chromosomal location other than its natural chromosomal location.
  • An isolated nucleic acid encoding an antigen-binding molecule refers to one or more nucleic acid molecules encoding an antigen-binding molecule, including such one or more nucleic acid molecules in a single vector or in separate vectors, and one or more nucleic acid molecules present in a host cell. Such one or more nucleic acid molecules at more positions.
  • nucleic acid sequence also implicitly encompasses conservatively modified variants thereof (eg, degenerate codon substitutions) and complementary sequences as well as the explicitly indicated sequence.
  • degenerate codon substitutions can be obtained by generating sequences in which the third position of one or more selected (or all) codons is replaced by a degenerate base and/or Deoxyinosine residue substitution.
  • polypeptide and "protein” are used interchangeably herein to refer to a polymer of amino acid residues.
  • the term applies to amino acid polymers in which one or more amino acid residues are the corresponding artificial chemical mimetic of a naturally occurring amino acid, and to both naturally occurring amino acid polymers and non-naturally occurring amino acid polymers. Unless otherwise stated, a particular polypeptide sequence also implicitly encompasses conservatively modified variants thereof.
  • sequence identity refers to the degree (percentage) to which the amino acids/nucleic acids of two sequences are identical at equivalent positions when the two sequences are optimally aligned; Gaps are used to obtain the maximum percent sequence identity, and any conservative substitutions are not considered part of the sequence identity.
  • alignment can be achieved by techniques known in the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, ALIGN-2 or Megalign (DNASTAR) software. Those skilled in the art can determine suitable parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared.
  • fused or “linked” refer to the joining of components (eg, B-VH and B-VL) by a covalent bond, either directly or via one or more linkers.
  • vector means a polynucleotide molecule capable of transporting another polynucleotide to which it has been linked.
  • plasmid refers to a circular double-stranded DNA loop into which additional DNA segments can be ligated.
  • viral vector such as an adeno-associated viral vector (AAV or AAV2), in which additional DNA segments can be ligated into the viral genome.
  • AAV adeno-associated viral vector
  • Certain vectors are capable of autonomous replication in the host cells into which they are introduced (eg, bacterial vectors and episomal mammalian vectors with a bacterial origin of replication).
  • vectors can integrate into the genome of the host cell after introduction into the host cell, thereby replicating along with the host genome.
  • expression vector or "expression construct” refers to a vector that can transform a host cell and contains a vector that directs and/or controls (along with the host cell) the expression of one or more heterologous coding regions operably linked thereto.
  • Expression constructs may include, but are not limited to, sequences that affect or control transcription, translation, and, when an intron is present, RNA splicing of the coding region to which it is operably linked.
  • host cell refers to a cell into which exogenous nucleic acid has been introduced, including the progeny of such cells.
  • Host cells include “transformants” and “transformed cells,” which include the primary transformed cell and progeny derived therefrom, regardless of the number of passages.
  • Progeny may not be identical to the parental cell in nucleic acid content, but may contain mutations.
  • mutant progeny including cells having the same function or biological activity as those screened or selected in the primary transformed cells.
  • Host cells include prokaryotic and eukaryotic host cells, where eukaryotic host cells include, but are not limited to, mammalian cells, insect cell lines, plant cells, and fungal cells.
  • Mammalian host cells include human, mouse, rat, dog, monkey, pig, goat, cow, horse, and hamster cells, including but not limited to Chinese hamster ovary (CHO) cells, NSO, SP2 cells, HeLa cells, baby hamster cells Kidney (BHK) cells, monkey kidney cells (COS), human hepatocellular carcinoma cells (eg, Hep G2), A549 cells, 3T3 cells, and HEK-293 cells.
  • Fungal cells include yeast and filamentous fungal cells including, for example, Pichia pastoris, Pichia finlandica, Pichia trehalophila, Pichia koclamae, Pichia membranaefaciens, Pichia minuta (Ogataea minuta, Pichia lindneri), Pichia puntiae, Pichia thermotolerans, Pichia willow salictaria), Pichia guercuum, Pichia pijperi, Pichia stiptis, Pichia methanolica, Pichia, Saccharomycescerevisiae, Saccharomyces cerevisiae , Hansenula polymorpha, Kluyveromyces, Kluyveromyces lactis, Candida albicans, Aspergillus nidulans, Aspergillus niger, Aspergillus oryzae, Trichoderma reesei, Chrysosporium lucknowense, Fus
  • Pichia any Saccharomyces, Hansenula polymorpha, any Kluyveromyces, Candida albicans, any Aspergillus, Trichoderma reesei, Luke Mold (Chrysosporium lucknowense), any Fusarium species, Yarrowia lipolytica, and Neurospora crassa.
  • the host cell cannot develop into a plant or individual animal.
  • composition means a mixture comprising one or more antigen binding molecules described herein together with other chemical components such as physiologically/pharmaceutically acceptable carriers and excipients.
  • pharmaceutically acceptable carrier refers to an ingredient in a pharmaceutical formulation (formulation) that is different from the active ingredient and is non-toxic to the subject.
  • subject or “individual” includes humans and non-human animals.
  • Non-human animals include all vertebrates (eg, mammals and non-mammals) such as non-human primates (eg, cynomolgus monkeys), sheep, dogs, cows, chickens, amphibians, and reptiles.
  • patient or “subject” are used interchangeably herein unless otherwise indicated.
  • cyno or “cynomolgus” refers to Macaca fascicularis.
  • the individual or subject is a human.
  • administering when applied to an animal, human, experimental subject, cell, tissue, organ or biological fluid, refers to the interaction of an exogenous drug, therapeutic agent, diagnostic agent or composition with an animal, human , subjects, cells, tissues, organs or biological fluids.
  • sample refers to a collection (such as a fluid, cell, or tissue) isolated from a subject, as well as a fluid, cell, or tissue present in a subject.
  • samples are biological fluids such as blood, serum and serosal fluids, plasma, lymph, urine, saliva, cyst fluid, tears, faeces, sputum, mucous membrane secretions of secretory tissues or organs, vaginal secretions, Ascites, pleura, pericardium, peritoneum, fluids of the peritoneal cavity and other body cavities, fluid collected from bronchial lavage, synovial fluid, liquid solutions in contact with the subject or biological source, such as culture medium (including conditioned medium), perfusion Washes, etc., tissue biopsy samples, fine needle aspirations, surgically removed tissue, organ cultures, or cell cultures.
  • biological fluids such as blood, serum and serosal fluids, plasma, lymph, urine, saliva, cyst fluid, tears, faeces, sputum, mucous membrane secretions of secret
  • Treatment refers to clinical intervention applied to an individual being treated, and may be performed for prophylactic purposes, or during the course of clinical pathology. Desired effects of treatment include, but are not limited to, prevention of occurrence or recurrence of disease, alleviation of symptoms, alleviation/reduction of any direct or indirect pathological consequences of disease, prevention of metastasis, reduction of rate of disease progression, amelioration or palliation of disease state, and regression or improved prognosis .
  • the molecules of the present disclosure are used to delay the development of a disease or slow the progression of a disease.
  • an "effective amount” is generally sufficient to reduce the severity and/or frequency of symptoms, eliminate those symptoms and/or their underlying causes, prevent the occurrence of symptoms and/or their underlying causes, and/or ameliorate or ameliorate the impairment caused by or associated with the disease state amount.
  • the effective amount is a therapeutically or prophylactically effective amount.
  • a “therapeutically effective amount” is sufficient to treat a disease state or symptom, especially a state or symptom associated with the disease state, or otherwise retard, delay or reverse the progression of the disease state or any other undesirable symptom associated with the disease quantity.
  • a “prophylactically effective amount” is an amount that, when administered to a subject, will have a predetermined prophylactic effect, such as preventing or delaying the onset (or recurrence) of the disease state, or reducing the likelihood of the onset (or recurrence) of the disease state or associated symptoms .
  • Complete therapeutic or prophylactic effect does not necessarily occur after administration of one dose, but may occur after administration of a series of doses.
  • a therapeutically or prophylactically effective amount may be administered in one or more administrations.
  • “Therapeutically effective amount” and “prophylactically effective amount” can vary depending on factors such as the disease state, age, sex and weight of the individual, and the ability of the therapeutic agent or combination of therapeutic agents to elicit a desired response in the individual.
  • Exemplary indicators of an effective therapeutic agent or combination of therapeutic agents include, for example, improved health status of a patient.
  • BAFF should be broadly understood and is intended to cover various forms of molecules of BAFF in various stages of mammalian body, such as but not limited to the process of amplification, replication, transcription, splicing, processing, translation and modification of BAFF gene Produced molecules (eg, precursor BCMA, mature BAFF, membrane-expressed BAFF, BAFF splice variants, modified BAFF, or fragments thereof); the term also encompasses artificially produced or in vitro expressed BAFF.
  • IL-12 should be broadly understood and is intended to cover various forms of IL-12 molecules in various stages of mammalian body, such as but not limited to IL-12 gene in amplification, replication, transcription, splicing, processing Molecules produced during , translation, modification (e.g. precursor IL-12, mature IL-12, membrane expressed IL-12, IL-12 splice variants, modified IL-12, or fragments, subunits thereof) ; the term also encompasses artificially produced or in vitro expressed IL-12.
  • modification e.g. precursor IL-12, mature IL-12, membrane expressed IL-12, IL-12 splice variants, modified IL-12, or fragments, subunits thereof
  • IL-23 should be broadly understood and is intended to cover various forms of IL-23 molecules in various stages of mammalian body, such as but not limited to IL-23 gene in amplification, replication, transcription, splicing, processing Molecules produced during , translation, modification (e.g. precursor IL-23, mature IL-23, membrane expressed IL-23, IL-23 splice variants, modified IL-23, or fragments, subunits thereof) ; the term also encompasses artificially prepared or in vitro expressed IL-23.
  • modification e.g. precursor IL-23, mature IL-23, membrane expressed IL-23, IL-23 splice variants, modified IL-23, or fragments, subunits thereof
  • Antigen binding molecules of the present disclosure are provided.
  • the present disclosure obtains an antigen-binding molecule that specifically binds BAFF by modifying Belimumab.
  • antigen-binding molecules that specifically bind BAFF and IL-12/23 were also constructed.
  • the antigen-binding molecules of the present disclosure have many favorable properties, such as binding to antigen with high affinity, blocking the binding of BAFF to its receptors (such as BAFF-R, BCMA and/or TACI), inhibiting the induction of BAFF B cell proliferation, blocking IL-12/23 binding to its receptor, inhibiting IL-12-induced IFN ⁇ secretion, inhibiting IL-23-induced IL-17 secretion, pharmacokinetic properties and/or druggability, etc.
  • the present disclosure provides an antigen binding molecule that specifically binds BAFF, which is an anti-BAFF antibody.
  • the antibody is a full-length antibody or an antigen-binding fragment thereof (e.g., Fv, Fab, Fab', Fab'-SH, F(ab')2, single domain antibody, single chain Fab (scFab), Diabodies, linear antibodies, single chain antibodies (scFv)) having one or more of the following functional activities:
  • the antigen-binding molecule can block the binding of human BAFF to human BAFF-R; preferably, the IC50 value for blocking the binding of human BAFF to human BAFF-R is less than 11 nM (for example, less than 11 nM, less than or equal to 10 nM, less than or equal to Equal to 9nM, less than or equal to 8nM, less than or equal to 7nM, less than or equal to 6nM, less than or equal to 5nM or less), the IC50 value is detected by Elisa method; in some embodiments, the IC50 value is tested according to the present disclosure Example 2 method detection;
  • the antigen-binding molecule can inhibit BAFF-induced B cell proliferation; preferably, the antigen-binding molecule can inhibit BAFF-induced B cell proliferation; , less than or equal to 0.10 nM, less than or equal to 0.09 nM, less than or equal to 0.08 nM, less than or equal to 0.07 nM, less than or equal to 0.06 nM or less) inhibits BAFF-induced B cell proliferation; in some embodiments , the IC50 value is detected according to the method of Test Example 3 of the present disclosure;
  • the antigen binding molecule can be less than 9.99E-10M (such as less than 9.99E-10M, less than or equal to 9.00E-10M, less than or equal to 8.20E-10M, less than or equal to 7.40E-10M, less than or equal to 6.60E -10M, less than or equal to 5.20E-10M, less than or equal to 4.00E-10M, less than or equal to 3.00E-10M or less) combined with human BAFF, the KD value is detected by the Biacore method; in some implementations In the method, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can be combined with cynomolgus BAFF with a KD value of less than 5.00E-10M (for example, less than or equal to 4.00E-10M, less than or equal to 3.00E-10M or less), and the KD value is passed Biacore method detection; In some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can bind to mouse BAFF with a KD value of less than 2.20E-10M (for example, less than or equal to 2.00E-10M, less than or equal to 1.00E-10M or less), and the KD value is determined by the Biacore method Detection; In some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can block the binding of human BAFF to human BCMA; preferably, the IC50 value of blocking the binding of human BAFF to human BCMA is less than 0.9nM (for example, less than or equal to 0.4nM, less than or equal to 0.35nM, less than or equal to or equal to 0.31nM or less), the IC50 value is detected by Elisa method; in some embodiments, the IC50 value is detected according to the method of Test Example 2 of the present disclosure; or
  • the antigen-binding molecule can block the binding of human BAFF to human TACI; preferably, the IC50 value of blocking the binding of human BAFF to human TACI is less than 0.6nM (for example, less than or equal to 0.55nM, less than or equal to 0.4nM, less than or equal to or equal to 0.33nM or less), the IC50 value is detected by the Elisa method; in some embodiments, the IC50 value is detected according to the method of Test Example 2 of the present disclosure.
  • the present disclosure provides an antigen binding molecule that specifically binds BAFF and IL-12/23.
  • the antigen binding molecule is a bispecific antibody that specifically binds BAFF and IL-12/23 P40 subunits.
  • the antigen binding molecule has one or more of the following functional activities:
  • the antigen-binding molecule can block the binding of human BAFF to human BAFF-R; preferably, the IC50 value for blocking the binding of human BAFF to human BAFF-R is less than 11 nM (for example, less than 11 nM, less than or equal to 10 nM, less than or equal to Equal to 9nM, less than or equal to 8nM, less than or equal to 7nM, less than or equal to 6nM, less than or equal to 5nM or less), the IC50 value is detected by Elisa method; in some embodiments, the IC50 value is tested according to the present disclosure Example 2 method detection;
  • the antigen-binding molecule can inhibit BAFF-induced B cell proliferation; preferably, the antigen-binding molecule can inhibit BAFF-induced B cell proliferation; , less than or equal to 0.10 nM, less than or equal to 0.09 nM, less than or equal to 0.08 nM, less than or equal to 0.07 nM, less than or equal to 0.06 nM or less) inhibits BAFF-induced B cell proliferation; in some embodiments , the IC50 value is detected according to the method of Test Example 3 of the present disclosure;
  • the antigen binding molecule can be less than 9.99E-10M (such as less than 9.99E-10M, less than or equal to 9.00E-10M, less than or equal to 8.20E-10M, less than or equal to 7.40E-10M, less than or equal to 6.60E -10M, less than or equal to 5.20E-10M, less than or equal to 4.00E-10M, less than or equal to 3.00E-10M or less) combined with human BAFF, the KD value is detected by the Biacore method; in some implementations In the method, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can be combined with cynomolgus BAFF with a KD value of less than 5.00E-10M (for example, less than or equal to 4.00E-10M, less than or equal to 3.00E-10M or less), and the KD value is passed Biacore method detection; In some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can bind to mouse BAFF with a KD value of less than 2.20E-10M (for example, less than or equal to 2.00E-10M, less than or equal to 1.00E-10M or less), and the KD value is determined by the Biacore method Detection; In some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure;
  • the antigen-binding molecule can block the binding of human BAFF to human BCMA; preferably, the IC50 value of blocking the binding of human BAFF to human BCMA is less than 0.9nM (for example, less than or equal to 0.4nM, less than or equal to 0.35nM, less than or equal to or equal to 0.31nM or less), the IC50 value is detected by Elisa method; in some embodiments, the IC50 value is detected according to the method of Test Example 2 of the present disclosure;
  • the antigen-binding molecule can block the binding of human BAFF to human TACI; preferably, the IC50 value of blocking the binding of human BAFF to human TACI is less than 0.6nM (for example, less than or equal to 0.55nM, less than or equal to 0.4nM, less than or equal to or equal to 0.33nM or less), the IC50 value is detected by Elisa method; in some embodiments, the IC50 value is detected according to the method of Test Example 2 of the present disclosure;
  • the antigen-binding molecule can block the binding of human IL-12/23 p40 to human IL-12R ⁇ 1; preferably, the IC50 value for blocking the binding of human IL-12/23 p40 to human IL-12R ⁇ 1 is less than 3.6nM ( For example, less than or equal to 3.0nM, less than or equal to 2.0nM, less than or equal to 1.0nM, less than or equal to 0.5nM, less than or equal to 0.23nM or less), the IC50 value is detected by the Elisa method; in some embodiments, The IC50 value is detected according to the method of Test Example 2 of the present disclosure;
  • Antigen-binding molecules can inhibit IL-12-induced IFN ⁇ secretion; preferably, the IC50 value of antigen-binding molecules inhibiting IL-12-induced IFN ⁇ secretion is less than 3.0nM (for example, less than or equal to 2.6nM, less than or equal to 1.6nM or less) ; In some embodiments, the IC50 value is detected according to the method of Test Example 4 of the present disclosure;
  • J. Antigen-binding molecules can inhibit IL-23-induced IL-17 secretion; preferably, the antigen-binding molecules inhibit IL-23-induced IL-17 secretion with an IC50 value of less than 0.034nM (for example, less than or equal to 0.031nM, less than or equal to 0.030nM or less); In some embodiments, the IC50 value is detected according to the method of Test Example 4 of the present disclosure;
  • the antigen-binding molecule can bind to cynomolgus monkey IL-12/23 p40 with a KD value less than 2.7E-10M (for example, less than or equal to 2.0E-10M, less than or equal to 1.0E-10M or less), said The KD value is detected by the Biacore method; in some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure; or
  • the antigen-binding molecule can bind to human IL-23 with a KD value of less than 6.4E-11M (e.g., less than or equal to 6.3E-11M, less than or equal to 6.2E-11M or less), the KD value is determined by the Biacore method Detection; In some embodiments, the KD value is detected according to the method of Test Example 5 of the present disclosure.
  • the antigen-binding molecule according to any one of the above, which comprises a heavy chain variable region B-VH and a light chain variable region B-VL, said B-VH comprising B-HCDR1, B-HCDR2 and B-HCDR3, the B-VL comprises B-LCDR1, B-LCDR2 and B-LCDR3, wherein the amino acid sequences of the B-HCDR1, B-HCDR2 and B-HCDR3 are respectively the same as those of SEQ ID NO: 63
  • the amino acid sequences of Bv-HCDR1, Bv-HCDR2 and Bv-HCDR3 are identical, and the amino acid sequences of B-LCDR1, B-LCDR2 and B-LCDR3 are respectively the same as those of Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 in SEQ ID NO: 64.
  • Bv-LCDR3 The amino acid sequences of Bv-LCDR3 are identical.
  • the B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3, Bv-HCDR1, Bv-HCDR2, Bv-HCDR3, Bv-LCDR1, Bv-LCDR2 and Bv-LCDR3 are defined according to the same numbering convention selected from Kabat, IMGT, Chothia, AbM and Contact.
  • the antigen-binding molecules according to any one of the above, said B-HCDR1, B-HCDR2, B-HCDR3, B-LCDR1, B-LCDR2, B-LCDR3 are defined according to the Kabat numbering rules, wherein, the amino acid sequence of B-HCDR1 is NNAIN (SEQ ID NO: 18), the amino acid sequence of B-HCDR2 is X 1 IX 2 PMFGX 3 AKYSX 4 X 5 FQG (SEQ ID NO: 65), the amino acid sequence of B-HCDR3 SRDX 6 LLFPX 7 X 8 X 9 LX 10 X 11 (SEQ ID NO: 66), the amino acid sequence of B-LCDR1 is X 12 GX 13 X 14 LX 15 X 16 X 17 X 18 AS (SEQ ID NO: 67) , the amino acid sequence of B-LCDR2 is GKNNRPS (SEQ ID NO: 32) and the amino acid sequence of B-LCDR3 is X 19 SRX 20
  • X 1 to X 25 are not selected from the following combinations: X 1 is G, X 2 is I, X 3 is T, X 4 is Q, X 5 is N, X 6 is L, X 7 is H, X 8 is H, X 9 is A, X 10 is S, X 11 is P, X 12 is Q, X 13 is D, X 14 is S, X 15 is R, X 16 is S, X 17 is Y, X 18 is Y, X 19 is S, X 20 is D, X 21 is S, X 22 is S, X 23 is N, X 24 is H, and X 25 is V.
  • said B-HCDR1 is shown in SEQ ID NO: 18, said B-HCDR2 is shown in SEQ ID NO: 28, and said B-HCDR3 is shown in SEQ ID NO: 29 , and said B-LCDR1 is shown in SEQ ID NO: 43, said B-LCDR2 is shown in SEQ ID NO: 32, and said B-LCDR3 is shown in SEQ ID NO: 44; or (ii) The B-HCDR1 is shown in SEQ ID NO: 18, the B-HCDR2 is shown in SEQ ID NO: 21, the B-HCDR3 is shown in SEQ ID NO: 20, and the B-LCDR1 is shown in SEQ ID NO: 34, said B-LCDR2 as shown in SEQ ID NO: 32, and said B-LCDR3 as shown in SEQ ID NO: 35; or (iii) said B-HCDR1 as shown in SEQ ID NO: 18 As shown, the B-HCDR2 is shown in SEQ ID NO: 22, the B-HCDR3 is shown in SEQ ID NO: 29 ,
  • the antigen-binding molecule as described in any one of the above it comprises B-VH and B-VL, described B-VH is as shown in SEQ ID NO: 47, and described B-VL is as shown in SEQ ID NO: 48 shown.
  • the B-VH is as shown in SEQ ID NO: 5, and the B-VL is as shown in SEQ ID NO: 12; or the B-VH is as shown in SEQ ID NO: 6, and said B-VL as shown in SEQ ID NO: 13; or said B-VH as shown in SEQ ID NO: 7, and said B-VL as shown in SEQ ID NO: 14; or said B- VH as shown in SEQ ID NO: 8, and said B-VL as shown in SEQ ID NO: 15; or said B-VH as shown in SEQ ID NO: 9, and said B-VL as shown in SEQ ID NO : shown in 16; or the B-VH as shown in SEQ ID NO: 10, and the B-VL as shown in SEQ ID NO: 17; or the B-VH as shown in SEQ ID NO: 11, and said B-VL as shown in SEQ ID NO:2.
  • the antigen-binding molecule that specifically binds BAFF and IL-12/23 comprises an antigen-binding moiety 1 that specifically binds BAFF and an antigen-binding moiety 2 that specifically binds IL-12 and/or IL-23 ; wherein said antigen binding module 2 comprises heavy chain variable region P-VH and light chain variable region P-VL, said P-VH comprising P-HCDR1, P-HCDR2 and P-HCDR3, said P-VL Comprising P-LCDR1, P-LCDR2 and P-LCDR3; the amino acid sequences of P-HCDR1, P-HCDR2 and P-HCDR3 are respectively the same as those of Pv-HCDR1, Pv-HCDR2 and Pv-HCDR3 in SEQ ID NO: 51 The amino acid sequence is the same, and the amino acid sequence of the P-LCDR1, P-LCDR2 and P-LCDR3 is respectively identical to the amino acid sequence of Pv-LCDR1, Pv-LCDR2 and Pv
  • the antigen-binding molecule according to any one of the above, said P-HCDR1, P-HCDR2, P-HCDR3, P-LCDR1, P-LCDR2, P-LCDR3 are defined according to the Kabat numbering rules, wherein, the P-HCDR1 is shown in SEQ ID NO: 53, the P-HCDR2 is shown in SEQ ID NO: 54, the P-HCDR3 is shown in SEQ ID NO: 55, and the P-LCDR1 As shown in SEQ ID NO: 56, the P-LCDR2 is shown in SEQ ID NO: 57, and the P-LCDR3 is shown in SEQ ID NO: 58.
  • the P-VH of the heavy chain variable region is shown in SEQ ID NO:51
  • the P-VL of the light chain variable region is shown in SEQ ID NO:52.
  • amino acid sequence variants of the antigen binding molecules provided herein are contemplated.
  • Amino acid sequence variants of antibodies can be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody, or by peptide synthesis. Such modifications include, for example, deletions, and/or insertions, and/or substitutions of residues within the amino acid sequence of the antigen-binding molecule. Any combination of deletions, insertions, and substitutions can be made to arrive at the final construct, so long as the final construct possesses the desired characteristics, such as antigen-binding properties.
  • antigen binding molecule variants having one or more amino acid substitutions are provided.
  • Sites of interest for substitution mutagenesis include CDRs and FRs.
  • Conservative substitutions are shown in Table 2 under the heading "Preferred Substitutions”. More substantial changes are provided in Table 2 under the heading "Exemplary Substitutions" and are described further below with reference to amino acid side chain classes.
  • Amino acid substitutions can be introduced into an antibody of interest, and the products screened for desired activity, such as retained/improved antigen binding, reduced immunogenicity, or improved ADCC or CDC.
  • amino acids can be grouped as follows:
  • Non-conservative substitutions refer to the substitution of a member of one class for a member of another class.
  • substitutional variant involves substituting one or more CDR residues of a parent antibody (eg, a humanized or human antibody).
  • a parent antibody eg. a humanized or human antibody
  • the resulting variant selected for further study will have an altered (e.g. improved) certain biological property (e.g. increased affinity, reduced immunogenicity) relative to the parent antibody, and/or will be substantially Some of the biological properties of the parental antibody are retained.
  • An exemplary substitution variant is an affinity matured antibody, which can be conveniently produced, for example, using phage display-based affinity maturation techniques such as those described herein. Briefly, one or more CDR residues are mutated, and the variant antibodies are displayed on phage and screened for specific biological activity (eg, binding affinity).
  • Alterations can be made to the CDRs, e.g., to improve antibody affinity. Such changes can be made to CDR "hotspots" (i.e., residues encoded by codons that undergo mutation at high frequency during the somatic maturation process, and/or residues that contact antigen) while making changes to the resulting variant VH or VL test for binding affinity.
  • affinity maturation diversity is introduced into the variable genes selected for maturation by any of a variety of methods, such as error-prone PCR, strand shuffling, or oligonucleotide-directed mutagenesis middle. Then, create secondary libraries. The library is then screened to identify any antibody variants with the desired affinity.
  • CDR residues involved in antigen binding can be specifically identified, for example, using alanine scanning mutagenesis or modeling.
  • HCDR3 and LCDR3 are frequently targeted.
  • substitutions, insertions or deletions may occur within one or more CDRs, so long as such changes do not substantially reduce the ability of the antibody to bind antigen.
  • conservative changes eg, conservative substitutions, as provided herein
  • Such changes do not occur at antigen contact residues.
  • alanine scanning mutagenesis One method that can be used to identify residues or regions of an antibody that can be targeted for mutagenesis is called "alanine scanning mutagenesis".
  • residues e.g. charged residues such as Arg, Asp, His, Lys and Glu
  • neutral or negatively charged amino acids e.g. Ala or Polypropylene
  • Amino acid amino acids
  • substitutions may be introduced at amino acid positions showing functional sensitivity to the initial substitution.
  • contact points between antibody and antigen can be identified by studying the crystal structure of the antigen-antibody complex. These contact residues and neighboring residues can be targeted or eliminated as candidates for substitution. Variants can be screened to determine whether they contain desired properties.
  • Amino acid sequence insertions include: amino- and/or carboxy-terminal fusions of one residue or polypeptides with a length of 100 or more residues, and intrasequence insertions of single or multiple amino acid residues.
  • terminal insertions include antibodies with an N-terminal methionyl residue.
  • Other insertional variants of antibody molecules include fusions with enzymes (or polypeptides that extend the serum half-life of antibodies) at the N- or C-terminus of the antibody.
  • the Fc region of an antigen binding molecule of the disclosure comprises one or more amino acid substitutions that reduce its binding to an Fc receptor, e.g., its binding to an Fc ⁇ receptor, and reduce or Eliminate effector functions.
  • a native IgG Fc region specifically an IgG 1 Fc region or an IgG 4 Fc region, may result in the targeting of an antigen binding molecule of the present disclosure to cells expressing Fc receptors, rather than cells expressing antigen.
  • the engineered Fc regions of the present disclosure exhibit reduced binding affinity to Fc receptors and/or reduced effector functions.
  • the engineered Fc region has a binding affinity for Fc receptors that is reduced by more than 50%, 80%, 90%, or 95% compared to a native Fc region.
  • the Fc receptor is an Fc gamma receptor.
  • the Fc receptor is a human Fc ⁇ receptor, eg, Fc ⁇ RI, Fc ⁇ RIIa, Fc ⁇ RIIB, Fc ⁇ RIIIa.
  • the engineered Fc region has reduced binding affinity for complement (eg, C1q) compared to a native Fc region.
  • the engineered Fc region has no reduced binding affinity for neonatal Fc receptor (FcRn) compared to a native Fc region.
  • the engineered Fc region has reduced effector function, which may include, but is not limited to, one or more of the following: reduced complement-dependent cytotoxicity (CDC), reduced Antibody-dependent cell-mediated cytotoxicity (ADCC), decreased antibody-dependent cellular phagocytosis (ADCP), decreased cytokine secretion, decreased immune complex-mediated antigen uptake by antigen-presenting cells, decreased interaction with NK cells decreased binding to macrophages, decreased binding to monocytes, decreased binding to polymorphonuclear cells, decreased direct signaling-induced apoptosis, decreased dendritic cell maturation, or decreased T cells primed.
  • CDC complement-dependent cytotoxicity
  • ADCC Antibody-dependent cell-mediated cytotoxicity
  • ADCP antibody-dependent cellular phagocytosis
  • cytokine secretion decreased immune complex-mediated antigen uptake by antigen-presenting cells
  • decreased interaction with NK cells decreased binding to macrophages
  • monocytes decreased binding to monocytes
  • polymorphonuclear cells
  • amino acid residue substitutions at positions 238, 265, 269, 270, 297, 327, and 329 may reduce effector function.
  • the Fc region is a human IgG 1 Fc region, and the amino acid residues at positions 234 and 235 are A, and the numbering is based on the EU index.
  • amino acid residue substitutions at positions such as 228 may reduce effector function.
  • Antigen binding molecules may also comprise disulfide bond engineering, eg, 354C of the first subunit and 349C of the second subunit.
  • the Fc region of the present disclosure comprises modifications according to the knob-into-hole (KIH) technique, which involves the introduction of a knob at the interface of one subunit and A hole structure (hole) is introduced at the interface. This enables the protrusion structure to be positioned in the hole structure, promotes the formation of heterodimers and inhibits the generation of homodimers.
  • KH knob-into-hole
  • the bulge structure is constructed by replacing small amino acid side chains from the interface of one subunit with larger side chains (such as tyrosine or tryptophan). Instead, the pore structure is created in the interface of another subunit by replacing large amino acid side chains with smaller ones, such as alanine or threonine.
  • the protrusion structure and hole structure are prepared by changing the nucleic acid encoding the polypeptide, and the optional amino acid substitutions are shown in Table 3 below:
  • knob-and-hole technique In addition to the knob-and-hole technique, other techniques for modifying the CH3 domain of a heavy chain to achieve heterodimerization are known in the art, for example WO96/27011, WO98/050431, EP1870459, WO2007/110205, WO 007/ 147901, WO2009/089004, WO2010/129304, WO2011/90754, WO2011/143545, WO2012/058768, WO2013/157954 and WO013/096291.
  • the C-terminus of the Fc region may be a complete C-terminus ending with the amino acid residue PGK; it may also be a truncated C-terminus, for example, one or two C-terminal amino acid residues have been removed from the truncated C-terminus.
  • the C-terminus of the Fc region is a shortened C-terminus ending with PG.
  • a composition of intact antibodies can include a population of antibodies from which all K447 residues and/or G446+K447 residues have been removed.
  • a composition of intact antibodies can include a population of antibodies in which the K447 residue and/or the G446+K447 residues have not been removed.
  • the composition of whole antibodies has a population of antibodies with and without a K447 residue and/or a mixture of antibodies with G446+K447 residues.
  • Antigen binding molecules eg, antibodies
  • Antigen binding molecules can be produced using recombinant methods. For these methods, one or more isolated nucleic acids encoding the antigen binding molecule are provided.
  • the present disclosure provides an isolated nucleic acid encoding an antigen binding molecule as previously described. Such nucleic acids may independently encode any of the aforementioned polypeptide chains.
  • the present disclosure provides one or more vectors (eg, expression vectors) comprising such nucleic acids.
  • the disclosure provides host cells comprising such nucleic acids.
  • a method of making an antigen binding molecule comprisin said method comprises, under conditions suitable for expression, culturing a host cell comprising a nucleic acid encoding said antigen binding molecule, as provided above, and optionally The antigen-binding molecule is efficiently recovered from the host cell (or host cell culture medium).
  • nucleic acid encoding the protein is isolated and inserted into one or more vectors for further cloning and/or expression in host cells.
  • nucleic acids can be readily isolated and sequenced using conventional procedures, or produced by recombinant methods or obtained by chemical synthesis.
  • Suitable host cells for cloning or expressing vectors encoding antigen-binding molecules include prokaryotic or eukaryotic cells as described herein. For example, it can be produced in bacteria, especially when glycosylation and Fc effector functions are not required. After expression, it can be isolated from the bacterial cell paste in a soluble fraction and can be further purified.
  • eukaryotic microbes such as filamentous fungi or yeast are suitable cloning or expression hosts for vectors encoding antigen-binding molecules, including fungal and yeast strains.
  • Suitable host cells suitable for expression of antigen binding molecules may also be derived from multicellular organisms (invertebrates and vertebrates); examples of invertebrate cells include plant and insect cells.
  • a number of baculovirus strains have been identified for use in combination with insect cells, particularly for the transfection of Spodoptera frugiperda cells; plant cell cultures can also be used as hosts, e.g.
  • vertebrate cells can also be used as hosts, such as mammalian cell lines adapted for growth in suspension.
  • suitable mammalian host cell lines are the SV40-transformed monkey kidney CV1 line (COS-7); the human embryonic kidney line (293 or 293T cells); baby hamster kidney cells (BHK); Sertoli) cells (TM4 cells); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical cancer cells (HELA); canine kidney cells (MDCK); buffalo rat liver cells ( BRL3A); human lung cells (W138); human hepatocytes (Hep G2); mouse mammary tumor (MMT 060562); TRI cells; MRC 5 cells; and FS4 cells.
  • Suitable mammalian host cell lines include Chinese Hamster Ovary (CHO) cells, including DHFR-CHO cells; and myeloma cell lines, such as YO, NSO and Sp2/0.
  • CHO Chinese Hamster Ovary
  • myeloma cell lines such as YO, NSO and Sp2/0.
  • Antigen binding molecules provided herein can be identified, screened or characterized for their physical/chemical characteristics and/or biological activity by a variety of assays known in the art. In one aspect, antigen binding molecules of the present disclosure are tested for activity, eg, by known methods such as ELISA, Western blot, and the like.
  • the antigen binding molecules provided by the present disclosure can be used to detect the presence or level of BAFF or IL-12/23 in a biological sample.
  • the term “detection” encompasses quantitative or qualitative detection.
  • the biological sample comprises cells or tissue, such as tumor tissue.
  • an antigen binding molecule for use in a diagnostic or detection method is provided.
  • methods of detecting the presence of BAFF or IL-12/23 in a biological sample are provided.
  • the method comprises contacting a biological sample with an antigen-binding molecule under suitable conditions, and detecting whether a complex is formed between the detection reagent and the antigen.
  • antigen binding molecules are used to select subjects suitable for treatment, for example BAFF or IL-12/23 are biomarkers for patient selection.
  • Exemplary disorders that can be diagnosed using the antigen binding molecules of the disclosure such as autoimmune diseases, for example: systemic lupus erythematosus, myasthenia gravis, multiple sclerosis, insulin-dependent diabetes, Crohn's disease, rheumatoid arthritis arthritis, polyarticular juvenile rheumatoid arthritis, or psoriatic arthritis; or B-cell disorders such as neoplasms, chronic leukocytic leukemia, multiple myeloma, non-Hodgkin's lymphoma, post-transplant lymphoproliferative disease or light chain gammopathies.
  • autoimmune diseases for example: systemic lupus erythematosus, myasthenia gravis, multiple sclerosis, insulin-dependent diabetes, Crohn's disease, rheumatoid arthritis arthritis, polyarticular juvenile rheumatoid arthritis, or psoriatic arthritis
  • B-cell disorders such as neoplasms
  • Labeled antigen binding molecules include, but are not limited to, labels or moieties for direct detection (such as fluorescent, chromogenic, electron-dense, chemiluminescent, and radioactive labels), and moieties for indirect detection (e.g., indirect detection via enzymatic reactions or molecular interactions).
  • modules such as enzymes or ligands).
  • the present disclosure provides the use of an antigen binding molecule in the manufacture or preparation of a medicament.
  • the B cell disorder or autoimmune disease is a disease or condition associated with BAFF or IL-12/23.
  • the autoimmune disease is selected from the group consisting of: systemic lupus erythematosus, myasthenia gravis, multiple sclerosis, insulin-dependent diabetes mellitus, Crohn's disease, rheumatoid arthritis, polyarticular juvenile rheumatoid Arthritis and psoriatic arthritis; said B-cell disorder is selected from the group consisting of neoplasms, chronic leukaemia, multiple myeloma, non-Hodgkin's lymphoma, post-transplantation lymphoproliferative disease, and light chain gammopathies .
  • the autoimmune disease is systemic lupus erythematosus.
  • the use further comprises administering to the subject an effective amount of at least one additional therapeutic agent (e.g., one, two, three, four, five, or six additional therapeutic agents agent).
  • a “subject” according to any of the above embodiments may be a human.
  • a subject When used for therapeutic purposes, a subject is an individual who has, or is suspected of having, the disease of interest. When used for prophylactic purposes, the subject is an individual susceptible to the disease of interest.
  • a pharmaceutical composition comprising said antigen binding molecule, eg, for any of the above pharmaceutical uses or methods of treatment.
  • a pharmaceutical composition comprises any of the antigen binding molecules provided herein and a pharmaceutically acceptable carrier.
  • the pharmaceutical composition further comprises at least one additional therapeutic agent.
  • the antigen binding molecules of the present disclosure can be used alone or in combination with other agents for therapy.
  • an antibody of the present disclosure can be administered in combination (simultaneously, or sequentially) with at least one additional therapeutic agent.
  • the antigen binding molecules of the present disclosure can be administered by any suitable means, including parenteral, intrapulmonary, intranasal, and, if local treatment is desired, intralesional.
  • Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal or subcutaneous administration. Administration may be by any suitable route, eg, by injection, such as intravenous or subcutaneous injection, depending in part on whether the administration is short-term or chronic.
  • a variety of dosing schedules are contemplated herein, including, but not limited to, single or multiple administrations at multiple time points, bolus administration, and pulse infusion.
  • Antigen binding molecules of the present disclosure will be formulated, dosed and administered in a manner consistent with good medical practice (eg, GOOD MEDICAL PRACTICE Guideline, GMP). Factors considered in this context include the particular condition being treated, the particular mammal being treated, the clinical condition of the individual patient, the cause of the condition, the site of delivery of the agent, the method of administration, the timing of administration, and others known to the medical practitioner. factor. Antigen binding molecules may or may not be formulated with one or more agents currently used to prevent or treat the disorder. The effective amount of such other agents depends on the amount present in the pharmaceutical composition, the type of disorder or treatment, and other factors. These are generally used at the same dosages and routes of administration as described herein, or at about 1 to 99% of the dosages described herein, or at other dosages, and any route empirically/clinically determined to be appropriate.
  • GMP Good MEDICAL PRACTICE Guideline
  • appropriate dosages of the antigen-binding molecules of the present disclosure will depend on the type of disease to be treated, the amount of the therapeutic molecule Type, severity and course of disease, whether administered for prophylactic or therapeutic purposes, previous therapy, subject's clinical history and response to the therapeutic molecule, and the judgment of the attending physician.
  • the therapeutic molecule is suitably administered to a subject at one time or over a series of treatments.
  • a daily dosage might range from about 1 ⁇ g/kg to 100 mg/kg, depending on the factors mentioned above.
  • an article of manufacture comprising materials useful for the treatment, prevention and/or diagnosis of the disorders described above.
  • the article comprises a container and a label or package insert on or associated with the container.
  • Suitable containers include, for example, bottles, vials, syringes, IV solution bags, and the like.
  • Containers can be formed from various materials such as glass or plastic.
  • the container contains a composition effective, alone or in combination with another composition, for the treatment, prophylaxis and/or diagnosis of a condition, and may have a sterile access opening (e.g., the container may have a stopper pierceable by a hypodermic needle). IV solution bag or vial).
  • At least one active agent in the composition is an antigen binding molecule of the present disclosure.
  • the label or package insert indicates that the composition is used to treat the condition of choice.
  • the article of manufacture may comprise: (a) a first container having a composition therein, wherein the composition comprises an antigen binding molecule of the present disclosure; and (b) a second container having a composition therein, wherein the combination
  • the drug contains an additional cytotoxic or other therapeutic agent.
  • the article of manufacture of this embodiment of the present disclosure may further comprise a package insert indicating that the composition may be used to treat a particular condition.
  • the article of manufacture may further comprise a second (or third) container comprising a pharmaceutically acceptable buffer. It may further comprise other materials as desired from a commercial and user standpoint, including other buffers, diluents, filters, needles and syringes.
  • the article of manufacture is prepared in the form of a kit.
  • Example 1 Belimumab mutant antibody and its activity detection
  • Belimumab was modified by phage display technology. By designing mutation primers, the CDRs of the light and heavy chains of Belimumab were mutated separately to construct a mutation library. Use biotinylated BAFF protein (Sino biological, 10056-HNCH) to enrich and screen the mutant library to obtain candidate clones, determine the amino acid sequence of the cloned light and heavy chain variable regions by sequencing, and separate the light and heavy chain variable regions of the candidate clones Fused with antibody light/heavy chain constant regions respectively to construct full-length antibodies.
  • biotinylated BAFF protein Seo biological, 10056-HNCH
  • the sequence information of the Belimumab antibody is as follows:
  • the sequence is FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4
  • the underlined part is the CDR region (determined according to the Kabat numbering system)
  • the italic part is the mutated amino acid residue
  • the underlined part is not for the frame area.
  • the CDRs in the table are the CDRs determined according to the Kabat numbering system.
  • the CDR in the table is the CDR determined according to the Kabat numbering system
  • the above heavy chain variable region and light chain variable region are fused with the antibody heavy chain constant region and light chain constant region, respectively, to construct a full-length antibody.
  • the above-mentioned heavy chain variable region is fused with the heavy chain constant region of Belimumab (sequence shown in SEQ ID NO: 45), and the light chain variable region of the antibody is fused with the light chain constant region of Belimumab (sequence shown in SEQ ID NO : shown in 46) fusion to construct anti-BAFF antibodies: B1 to B7, the variable region sequences of the obtained antibodies are shown in Table 6.
  • the heavy chain variable region is B-VH6 (SEQ ID NO: 10)
  • the light chain variable region is B-VL6 (SEQ ID NO: 17)
  • the heavy chain constant region is SEQ ID NO: 45
  • the light chain constant region is SEQ ID NO: 46.
  • the framework regions of the above-mentioned anti-BAFF antibodies were modified.
  • the amino acid residue at position 44 (according to the Kabat numbering system) of the heavy chain variable region of B6 is mutated to C (ie, 44C), and at the same time, the amino acid residue at position 100 of the light chain variable region of B6 (according to The position determined by the Kabat numbering system) amino acid residue was mutated to C (i.e. 100C) in order to increase the interchain disulfide bond between VH and VL, and the variable region sequence of antibody B61 after the B6 framework region transformation was as follows:
  • Example 2 Construction of bispecific antibodies specifically binding to BAFF and IL-12/23
  • the anti-BAFF antibody obtained in Example 1 and the known anti-IL-12/23 antibody were used to construct a bispecific antibody specifically binding to BAFF and IL-12/23.
  • the IL-12/23 antibody can be derived from any suitable antibody, for example, anti-IL-12/23 p40 subunit antibody Ustekinumab (abbreviated as Umab), the sequence of Ustekinumab is as follows:
  • the double underlined part is the variable region sequence
  • the single underlined part is the CDR region sequence
  • the variable region and CDR are confirmed according to the Kabat numbering system.
  • the human IgG1 heavy chain constant region (sequence shown in SEQ ID NO: 59), human kappa light chain constant region (sequence shown in SEQ ID NO: 60), B61 antibody heavy chain variable region and Light chain variable region (sequence shown in SEQ ID NO: 47, 48) and Ustekinumab antibody heavy chain variable region and light chain variable region (sequence shown in SEQ ID NO: 51, 52), construct specific binding Bispecific antibody to BAFF and IL-12/23.
  • the C-terminal of the variable region of the light chain of the Ustekinumab antibody is fused with the N-terminal of the constant region of the human ⁇ light chain to form the second chain;
  • the C-terminal of the variable region of the heavy chain of the Ustekinumab antibody is fused with the N-terminal of the constant region of the human IgG1 heavy chain,
  • the C-terminus of the human IgG1 heavy chain constant region is fused to the N-terminus of the scFv (constructed from the heavy and light chain variable regions of the B61 antibody) to form the first chain.
  • bispecific antibodies specifically binding to BAFF and IL-12/23 were constructed: BU-1 and BU-2.
  • Both BU-1 and BU-2 are composed of 4 chains (including two identical first chains and two identical second chains), the structural diagram of BU-1 is shown in Figure 1, and the structural diagram of BU-2 is shown in Figure 1 2.
  • the amino acid sequence of the second chain of BU-1 is the same as that of the light chain of Ustekinumab, and the sequence is shown in SEQ ID NO:50.
  • the amino acid sequence of the second chain of BU-2 is the same as that of the light chain of Ustekinumab, and the sequence is shown in SEQ ID NO:50.
  • the wavy part is the heavy chain variable region of the Ustekinumab antibody
  • the underlined part is the human IgG1 heavy chain constant region
  • the double underlined part is the heavy chain of the B61 antibody.
  • the chain variable region part, the single underline part is the light chain variable region part of B61 antibody
  • the italic part is the linker sequence part.
  • the binding activity of the antibody to BAFF and IL-12/23 p40 was detected by the Elisa method.
  • the specific method is as follows:
  • the sample to be tested was diluted to 2 ⁇ g/mL with pH 7.4 PBS (B320) buffer, added to a 96-well microplate plate (Corning, 3590) at a volume of 100 ⁇ L/well, and incubated overnight at 4°C. After the liquid was discarded, 300 ⁇ L of 5% skimmed milk (BD, 232100) diluted with PBS was added to each well for blocking, and incubated at 37° C. for 2 hours.
  • PBST buffer pH7.4PBS containing 0.1% tween-20
  • BAFF ACROBiosystems, BAF-H5248
  • IL-12/ 23 p40 Sino Biological, 10052-H08H
  • the blocking activity of the antibody against the following receptors and ligands was detected by Elisa method: human BAFF and human BAFF-R, human BAFF and human BCMA, human BAFF and human TACI, and human IL-12/23 p40 and human IL-12R ⁇ 1 .
  • the specific method is as follows:
  • the receptor protein was diluted to 2 ⁇ g/mL with pH 7.4 PBS (B320) buffer, added to a 96-well microtiter plate (Corning, 3590) at a volume of 100 ⁇ L/well, and incubated overnight at 4°C. After the liquid was discarded, 200 ⁇ L of 1% Casein blocking solution (Thermo, 37528) was added to each well for blocking, and incubated at 37° C. for 2 hours. After the blocking, the blocking solution was discarded, and the plate was washed 3 times with PBST buffer (pH7.4 PBS containing 0.1% tween-20) before use.
  • PBST buffer pH7.4 PBS containing 0.1% tween-20
  • a fixed concentration of biotin-labeled ligand protein was mixed with a gradiently diluted antibody, pre-incubated at 37°C for 30 minutes, then added to the blocked microtiter plate, and incubated at 37°C for 1.5 hours. After the incubation, the plate was washed 3 times with PBST, 100 ⁇ L streptavidin-HRP (Invitrogen, 434323, diluted 1:4000) was added to each well, and incubated at 37°C for 1 hour.
  • the sources of receptors and ligand proteins used in this test example are as follows: human BAFF (Sino biological, 10056-HNCH), human IL-12/23 p40 (Sino biological, 10052-H08H), human BAFF-R (Sino biological , 16079-H02H), human BCMA (Sino biological, 10620-H02H), human TACI (ACROBiosystems, TAI-H5256), human IL-12R ⁇ 1 (ACROBiosystems, ILB-H5255).
  • the experimental results are shown in Table 10, Table 11, and Table 12.
  • the experimental results show that the antibody specifically binding to BAFF constructed in this disclosure can effectively block the binding of human BAFF and human BAFF-R, and the IC50 value is lower than that of Belimumab.
  • the bispecific antibody specifically binding to BAFF and IL-12/23 constructed in this disclosure can effectively block human BAFF and human BAFF-R, human BAFF and human BCMA, human BAFF and human TACI, and the IC50 value is less than that of Belimumab ; It can also effectively block the binding of human IL-12/23 p40 to human IL-12R ⁇ 1.
  • the B cell proliferation assay was used to detect whether the antibody could inhibit BAFF-induced B cell proliferation.
  • the experimental method is as follows:
  • the mouse spleen was taken for grinding, centrifuged at 4°C for 5 minutes to collect the lower layer of cells, washed once with washing solution (PBS+2% FBS+2mM EDTA) and centrifuged, and added RBC lysis buffer (Invitrogen, 00-4333- 57), stand at room temperature for 5 minutes until the red blood cells are completely lysed. Centrifuge again and resuspend cells for counting.
  • the cell suspension was sorted with the B cell isolation kit (Miltenyi Biotec, 130-090-862), and the isolated B cells were sorted with RPMI 1640 medium (Gibco, 11875119)+10%FBS (Gibco, 10099-141)+ 50 ⁇ M 2-mercaptoethanol (Sigma-Aldrich, M6250) was resuspended and counted, and the cells were plated in 96-well cell plates (Costar, 3903) for later use.
  • the experimental results are shown in Table 13 and Table 14 below.
  • the experimental results show that the disclosed antibody that specifically binds to BAFF and the bispecific antibody that specifically binds to BAFF and IL-12/23 can effectively inhibit the proliferation of B cells induced by BAFF, and Belimumab inhibits
  • the IC50 value of BAFF-induced B cell proliferation is more than 2 times that of the disclosed antibody specifically binding to BAFF, while ustekinumab has no function of inhibiting BAFF-induced B cell proliferation.
  • the experimental method is as follows:
  • the mouse spleen was taken for grinding, centrifuged at 4°C for 5 minutes to collect the cells in the lower layer, washed once with washing solution (PBS+2% FBS+2mM EDTA) and centrifuged, after the supernatant was removed, RBC lysis buffer (Invitrogen, 00-4333- 57), stand at room temperature for 5 minutes until the red blood cells are completely lysed. Centrifuge again and resuspend cells for counting.
  • washing solution PBS+2% FBS+2mM EDTA
  • RBC lysis buffer Invitrogen, 00-4333- 57
  • the cell suspension was sorted with the mouse CD4 cell kit (Invitrogen, 11415D), and the isolated CD4+T cells were resuspended in the medium RPMI 1640 medium (Gibco, 11875119)+10% FBS (Gibco, 10099-141) And count to spare.
  • IL-12-induced T cell differentiation 20 ⁇ g/mL anti-mouse IL-4 (BioLegend, 504122) was added to T cells, and the cell suspension was spread in a coated 96-well plate. A fixed concentration of chimeric IL-12 (human p40 fused with mouse p35) protein was mixed with serially diluted antibodies, pre-incubated at 37°C for 1 hour, added to a 96-well plate, and incubated in a 37°C cell culture incubator for 48 hours.
  • chimeric IL-12 human p40 fused with mouse p35
  • the 96-well plate was taken out, centrifuged at 1000rpm for 3 minutes, the supernatant was collected, and the content of IFN ⁇ in the supernatant was detected with mouse IFN-gamma DuoSet ELISA Kit (R&D Systems, DY485).
  • the cell suspension was spread in a well-coated 96-well plate, and a fixed concentration of IL-23 (R&D Systems, 1290-IL-010) was mixed with a serially diluted antibody for pretreatment. After incubation for 1 hour, it was added to a 96-well plate and incubated in a 37°C cell culture incubator for 48 hours. The 96-well plate was taken out, centrifuged at 1000rpm for 3 minutes, the supernatant was collected, and the IL-17 content in the supernatant was detected with the mouse IL-17 DuoSet ELISA Kit (R&D Systems, DY421).
  • the experimental results are shown in Table 15 and Table 16 below.
  • the experimental results show that the disclosed bispecific antibody that specifically binds to BAFF and IL-12/23 can effectively inhibit the secretion of IL-17 induced by IL-23, and can effectively inhibit IL-12 Induced IFN ⁇ secretion, while Belimumab could neither inhibit IL-23-induced IL-17 secretion nor IL-12-induced IFN ⁇ secretion.
  • Biosensing chip Protein A (GE, 29127556) to affinity capture a certain amount of the sample to be tested, and then flow through a series of concentration gradient antigens on the surface of the chip, and use Biacore (GE, 8K) to detect the reaction signal in real time to obtain the binding and dissolving off the curve.
  • Biacore GE, 8K
  • the biochip was cleaned and regenerated with 10 mM glycine-hydrochloric acid solution pH 1.5 (GE, BR-1003-54).
  • the experimental data was fitted with BIA evaluation version 4.1 and GE software with a 1:1 model to obtain the affinity value.
  • the relevant antigen proteins used in this test are as follows: human IL-23 (CT048-H08H, Sino biological), human BAFF (10056-HNCH, Sino biological), cynomolgus monkey IL-12/23 P40 (10215-CL, R&D Systems), cynomolgus monkey BAFF (BAF-CM412B, Kactus), mouse BAFF (BAF-M521y, Acro Biosystems).
  • the experimental results are shown in Table 17 and Table 18 below.
  • the experimental results show that, compared with Belimumab, the anti-BAFF antibody constructed in this disclosure can bind to human BAFF with a smaller KD value; in addition, the construction of this disclosure specifically binds to BAFF and IL-12
  • the /23 bispecific antibody can bind to human IL-23, human BAFF, cynomolgus monkey IL-12/23 P40, cynomolgus monkey BAFF, and mouse BAFF with high affinity.
  • mice Simultaneously stimulate mice with chimeric IL-12 (human p40 fused with mouse p35), human IL-23 and human BAFF to induce the production of cytokines such as IFN ⁇ , TNF ⁇ , IL-22 and IgA in mice, and pass the detection
  • cytokines such as IFN ⁇ , TNF ⁇ , IL-22 and IgA
  • mice SPF female C57BL/6 mice (Beijing Weitong Lihua Experimental Animal Technology Co., Ltd., 8 weeks old) were randomly divided into groups of 5 mice, and chimeric IL-12 (2 ⁇ g/hour) was mixed intraperitoneally. Rat), human IL-23 (2 ⁇ g/mouse) and human BAFF (1 mg/kg) were injected once a day for four days. The sample to be tested (Belimumab 8mpk, BU-1 10.7mpk, or BU-1 5.35mpk) was injected intraperitoneally one hour before the protein injection on the first day and the third day respectively.
  • mice in each group were collected, and the levels of IFN ⁇ , TNF ⁇ , IL-22 and IgA were detected respectively.
  • the source information of the detection kit used in this test case is as follows: Mouse IFN-gamma Quantikine ELISA Kit (R&D Systems, MIF00), Mouse TNF-alpha Quantikine ELISA Kit (R&D Systems, MTA00B), Mouse/Rat IL-22 Quantikine ELISA Kit (R&D Systems, M2200), Mouse IgA ELISA Kit (Abcam, ab157717).
  • Belimumab was used as a positive control
  • PBS was used as a negative control.
  • BU-1 10.7mpk has the same molar drug concentration as Belimumab 8mpk.
  • the experimental results are shown in Figures 3 to 6.
  • the experimental results show that the disclosed bispecific antibody specifically binding to BAFF and IL-12/23 can significantly inhibit the secretion of TNF ⁇ , IFN ⁇ , and IL-22, while Belimumab cannot inhibit the secretion of TNF ⁇ and IFN ⁇ secretion.
  • BU-1 can significantly inhibit the secretion of IgA at two doses of 10.7mpk and 5.35mpk, and the inhibitory activity is stronger than that of Belimumab.
  • the experimental results are shown in Table 19 below.
  • the experimental results show that the half-life of the disclosed bispecific antibody specifically binding to BAFF and IL-12/23 in rats is very good, and the half-life of anti-IL-12/23 p40 reaches 14 days. The half-life is longer than that of Ustekinumab.

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Abstract

一种特异性结合BAFF和IL-12/23的抗原结合分子及用途。

Description

特异性结合BAFF和IL-12/23的抗原结合分子及用途
本申请要求2021年6月30日提交的中国专利申请(申请号202110735262.2)的优先权。
技术领域
本披露属于生物技术领域，更具体地，本披露涉及一种特异性结合BAFF和IL-12/23的抗原结合分子及其应用。
背景技术
这里的陈述仅是提供与本披露有关的背景信息，而不必然地构成现有技术。
BAFF(B细胞激活因子)是属于TNF家族的一种细胞激活因子。BAFF主要在骨髓细胞膜表面表达，以三聚体的形式存在，在细胞膜表面的BAFF会被蛋白酶水解形成可溶性的BAFF进入血液循环系统，可溶性的BAFF具有多聚化的特征，最多可以形成60聚体。另外BAFF也可以跟同家族的另外一个蛋白APRIL作用形成异源的三聚体。目前已知在B细胞表面有三个BAFF的受体，分别为BAFF-R、BCMA和TACI。BAFF与这三个受体作用，参与B细胞的分化成熟、存活和调节。APRIL与BAFF有两个共同的受体，分别为BCMA和TACI，APRIL与这两个受体作用参与B细胞的存活和调节(Samy,E.,et al.,Int Rev Immunol,2017.36:p.3-19；Kamal,A.and M.Khamashta,Autoimmun Rev,2014.13:p.1094-1101)。BAFF对维持B细胞的体内平衡非常重要，BAFF信号通路的过度激活会导致自反应B细胞的存活，并产生自身抗体促进自身免疫反应(Cancro,M.P.,D.P.D'Cruz,and M.A.Khamashta,J Clin Invest,2009.119:p.1066-73)。
IL-12和IL-23是属于同一个家族的两种细胞因子。IL-12和IL-23都是由两个亚基组成的异源二聚体蛋白(Moschen,A.R.,H.Tilg,and T.Raine,Nat Rev Gastroenterol Hepatol,2019.16:p.185-196；Frieder,J.,et al.,Clin Pharmacol Ther,2018.103:p.88-101)。IL-12是由p35和p40两个亚基组成，而IL-23是由p19和p40两个亚基组成。其中p40是IL-12和IL-23共有的一个亚基(IL-12/23 P40)，因此靶向p40的抗体能同时抑制IL-12和IL-23两条信号通路。IL-12与受体作用主要激活Th1细胞的分化，同时参与刺激多种免疫细胞分泌干扰素γ和TNF。IL-23与受体作用主要激活Th17细胞的分化，并刺激多种细胞分泌IL-17、IL-22和TNF等细胞因子(Lee,E.B.,et al.,Cutis,2018.101:p.5-9；Floss,D.M.,et al.,Cytokine Growth Factor Rev,2015.26:p.569-578)。IL-12/23信号通路激活所产生的这些细胞因子进一步通过各自的途径参与系统性红斑狼疮等疾病的发展。
发明内容
本披露构建了一种特异性结合BAFF的抗原结合分子，并构建了特异性结合 BAFF和IL-12和/或IL-23的抗原结合分子。
一方面，本披露提供一种抗原结合分子，其包含特异性结合BAFF的抗原结合模块1和特异性结合IL-12和/或IL-23的抗原结合模块2，其中，所述抗原结合模块1包含重链可变区B-VH和轻链可变区B-VL，所述B-VH包含B-HCDR1、B-HCDR2和B-HCDR3，所述B-VL包含B-LCDR1、B-LCDR2和B-LCDR3，其中：所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：63中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：64中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，其中，SEQ ID NO：63为：QVQLQQSGAEVKKPGSSVRVSCKASGGTFNNNAINWVRQAPGQX 26LEWMGX 1IX 2PMFGX 3AKYSX 4X 5FQGRVAITADESTGTASMELSSLRSEDTAVYYCARSRDX 6LLFPX 7X 8X 9LX 10X 11WGX 27GTMVTVSS，
SEQ ID NO：64为：
SSELTQDPAVSVALGQTVRVTCX 12GX 13X 14LX 15X 16X 17X 18ASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCX 19SRX 20X 21X 22GX 23X 24WX 25FGX 28GTELTVL，
其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V，X 26选自G或C，X 27选自R或G，X 28选自G或C；
并且，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3不包含如下CDR组合：B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：1中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，并且B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：2中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据选自Kabat、IMGT、Chothia、AbM和Contact的相同的编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Kabat编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2 和Bv-LCDR3是根据IMGT编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Chothia编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据AbM编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Contact编号规则定义的。
在一些实施方案中，如上任一项所述的抗原结合分子，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3是根据Kabat编号规则定义的，其中，B-HCDR1包含氨基酸序列NNAIN(SEQ ID NO：18)，B-HCDR2包含氨基酸序列X 1IX 2PMFGX 3AKYSX 4X 5FQG(SEQ ID NO：65)，B-HCDR3包含氨基酸序列SRDX 6LLFPX 7X 8X 9LX 10X 11(SEQ ID NO：66)，B-LCDR1包含氨基酸序列X 12GX 13X 14LX 15X 16X 17X 18AS(SEQ ID NO：67)，B-LCDR2包含氨基酸序列GKNNRPS(SEQ ID NO：32)和B-LCDR3包含氨基酸序列X 19SRX 20X 21X 22GX 23X 24WX 25(SEQ ID NO：68)，其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V；并且所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3不包含如下CDR组合：B-HCDR1包含SEQ ID NO：18的氨基酸序列，B-HCDR2包含SEQ ID NO：19的氨基酸序列，B-HCDR3包含SEQ ID NO：20的氨基酸序列，B-LCDR1包含SEQ ID NO：31的氨基酸序列，B-LCDR2包含SEQ ID NO：32的氨基酸序列，和B-LCDR3包含SEQ ID NO：33的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，其中：
(i)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：47中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：48中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
(ii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：5中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2 和B-LCDR3分别包含SEQ ID NO：12中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
(iii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：6中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：13中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
(iv)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：7中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：14中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
(v)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：8中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：15中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
(vi)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：9中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：16中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
(vii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：10中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：17中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
(viii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：11中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：2中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Kabat编号规则定义的，其中，
(i)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：28的氨基酸序列，所述B-HCDR3包含SEQ ID NO：29的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：43的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：44的氨基酸序列；或
(ii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包 含SEQ ID NO：21的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：34的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：35的氨基酸序列；或
(iii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：22的氨基酸序列，所述B-HCDR3包含SEQ ID NO：23的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：31的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：36的氨基酸序列；或
(iv)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：24的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：37的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：38的氨基酸序列；或
(v)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：25的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：39的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：40的氨基酸序列；或
(vi)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：26的氨基酸序列，所述B-HCDR3包含SEQ ID NO：27的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：41的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：42的氨基酸序列；或
(vii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：30的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：31的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：33的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，其中，所述B-VH包含与SEQ ID NO：63具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，所述B-VL包含与SEQ ID NO：64具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，其中SEQ ID NO：63为：
QVQLQQSGAEVKKPGSSVRVSCKASGGTFNNNAINWVRQAPGQX 26LEWMGX 1IX 2PMFGX 3AKYSX 4X 5FQGRVAITADESTGTASMELSSLRSEDTAVYYCARSRDX 6 LLFPX 7X 8X 9LX 10X 11WGX 27GTMVTVSS，
SEQ ID NO：64为：
SSELTQDPAVSVALGQTVRVTCX 12GX 13X 14LX 15X 16X 17X 18ASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCX 19SRX 20X 21X 22GX 23X 24WX 25FGX 28GTELTVL，其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V，X 26选自G或C，X 27选自R或G，X 28选自G或C；并且，所述B-VH和B-VL不包含如下可变区组合：B-VH包含SEQ ID NO：1的氨基酸序列，且B-VL包含SEQ ID NO：2的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，其中，
(i)所述B-VH包含与SEQ ID NO：47具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：48具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(ii)所述B-VH包含与SEQ ID NO：5具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：12具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(iii)所述B-VH包含与SEQ ID NO：6具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：13具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(iv)所述B-VH包含与SEQ ID NO：7具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：14具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(v)所述B-VH包含与SEQ ID NO：8具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：15具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(vi)所述B-VH包含与SEQ ID NO：9具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID  NO：16具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(vii)所述B-VH包含与SEQ ID NO：10具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：17具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(viii)所述B-VH包含与SEQ ID NO：11具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：2具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列；
在一些实施方案中，如上任一项所述的抗原结合分子，其中，所述B-VH包含SEQ ID NO：47的氨基酸序列，和所述B-VL包含SEQ ID NO：48的氨基酸序列。在一些实施方案中，所述B-VH包含SEQ ID NO：5的氨基酸序列，和所述B-VL包含SEQ ID NO：12的氨基酸序列；或所述B-VH包含SEQ ID NO：6的氨基酸序列，和所述B-VL包含SEQ ID NO：13的氨基酸序列；或所述B-VH包含SEQ ID NO：7的氨基酸序列，和所述B-VL包含SEQ ID NO：14的氨基酸序列；或所述B-VH包含SEQ ID NO：8的氨基酸序列，和所述B-VL包含SEQ ID NO：15的氨基酸序列；或所述B-VH包含SEQ ID NO：9的氨基酸序列，和所述B-VL包含SEQ ID NO：16的氨基酸序列；或所述B-VH包含SEQ ID NO：10的氨基酸序列，和所述B-VL包含SEQ ID NO：17的氨基酸序列；或所述B-VH包含SEQ ID NO：11的氨基酸序列，和所述B-VL包含SEQ ID NO：2的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，其中，所述特异性结合BAFF的抗原结合模块1为scFv。在一些实施方案中，所述scFv分子具有从N端至C端的如下结构：VL-连接子-VH或VH-连接子-VL。在一些实施方案中，所述连接子为具有“L 1-(GGGGS)n-L 2”结构的肽连接子，其中，L 1是键、A、GS、GGS或GGGS，n是0、1、2、3、4、5、6、7、8、9或10，L2是键、G、GG、GGG或GGGG，并且所述肽连接子不是键。在一些实施方案中，所述肽连接子的长度为3-15个氨基酸残基。在一些实施方案中，所述连接子结构为：(G xS) y，其中x选自1-5的整数(例如1、2、3、4或5)，y选自1-6的整数(例如，1、2、3、4、5或6)。在一些实施方案中，所述连接子为GGGGSGGGGSGGGGS(SEQ ID NO：70)。
在一些实施方案中，如上任一项所述的抗原结合分子，其中所述抗原结合模块2包含重链可变区P-VH和轻链可变区P-VL，所述P-VH包含P-HCDR1、P-HCDR2和P-HCDR3，所述P-VL包含P-LCDR1、P-LCDR2和P-LCDR3；所述P-HCDR1、P-HCDR2和P-HCDR3分别包含SEQ ID NO：51中的Pv-HCDR1、Pv-HCDR2和Pv-HCDR3的氨基酸序列，和所述P-LCDR1、P-LCDR2和P-LCDR3 分别包含SEQ ID NO：52中的Pv-LCDR1、Pv-LCDR2和Pv-LCDR3的氨基酸序列；所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3、Pv-HCDR1、Pv-HCDR2、Pv-HCDR3、Pv-LCDR1、Pv-LCDR2和Pv-LCDR3是根据选自Kabat、IMGT、Chothia、AbM和Contact的相同的编号规则定义的。
在一些实施方案中，所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3、Pv-HCDR1、Pv-HCDR2、Pv-HCDR3、Pv-LCDR1、Pv-LCDR2和Pv-LCDR3是根据Kabat编号规则定义的。
在一些实施方案中，所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3、Pv-HCDR1、Pv-HCDR2、Pv-HCDR3、Pv-LCDR1、Pv-LCDR2和Pv-LCDR3是根据IMGT编号规则定义的。
在一些实施方案中，所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3、Pv-HCDR1、Pv-HCDR2、Pv-HCDR3、Pv-LCDR1、Pv-LCDR2和Pv-LCDR3是根据Chothia编号规则定义的。
在一些实施方案中，所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3、Pv-HCDR1、Pv-HCDR2、Pv-HCDR3、Pv-LCDR1、Pv-LCDR2和Pv-LCDR3是根据AbM编号规则定义的。
在一些实施方案中，所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3、Pv-HCDR1、Pv-HCDR2、Pv-HCDR3、Pv-LCDR1、Pv-LCDR2和Pv-LCDR3是根据Contact编号规则定义的。
在一些实施方案中，如上任一项所述的抗原结合分子，所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3是根据Kabat编号规则定义的，所述P-HCDR1包含SEQ ID NO：53的氨基酸序列，所述P-HCDR2包含SEQ ID NO：54的氨基酸序列，所述P-HCDR3包含SEQ ID NO：55的氨基酸序列，所述P-LCDR1包含SEQ ID NO：56的氨基酸序列，所述P-LCDR2包含SEQ ID NO：57的氨基酸序列，和所述P-LCDR3包含SEQ ID NO：58的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，所述重链可变区P-VH包含与SEQ ID NO：51具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，所述轻链可变区P-VL包含与SEQ ID NO：52具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，所述重链可变区P-VH包含SEQ ID NO：51的氨基酸序列，所述轻链可变区P-VL包含SEQ ID NO：52的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，其包括重链恒定区CH和轻链恒定区CL。在一些实施方案中，所述重链恒定区CH为人IgG重链恒定区，所述轻链恒定区CL为人λ或κ轻链恒定区。在一些实施方案中，所述重链恒定区 CH为人IgG1重链恒定区，所述轻链恒定区CL为人λ轻链恒定区。在一些实施方案中，所述重链恒定区CH为人IgG1重链恒定区，所述轻链恒定区CL为人κ轻链恒定区。在一些实施方案中，所述重链恒定区CH包含SEQ ID NO：59的氨基酸序列，所述轻链恒定区CL包含SEQ ID NO：60的氨基酸序列。
在一些实施方案中，如上任一项所述的抗原结合分子，其包括重链恒定区CH和轻链恒定区CL，其中所述重链恒定区CH包含Fc区。在一些实施方案中，所述Fc区包含能够彼此缔合的第一亚基Fc1与第二亚基Fc2。在一些实施方案中，所述Fc区为IgG的Fc区。在一些实施方案中，所述Fc区为IgG 1的Fc区。在一些实施方案中，所述Fc区具有一个或更多个减少同源二聚化的氨基酸取代；和/或所述Fc区具有一个或更多个能够减少Fc区与Fc受体结合的氨基酸取代。在一些实施方案中，所述Fc区具有YTE突变(M252Y、S254T和T256E)，L234A、L235A突变，和/或S228P突变，所述突变编号依据为EU索引。在一些实施方案中，Fc区包含能够彼此缔合的第一亚基与第二亚基，所述第一亚基和/或第二亚基具有一个或多个减少同源二聚化的氨基酸取代。在一些实施方案中，所述第一亚基具有根据杵臼技术的凸起结构，所述第二亚基具有根据杵臼技术的孔结构，或者所述第一亚基具有根据杵臼技术的孔结构，所述第二亚基具有根据杵臼技术的凸起结构。在一些实施方案中，所述第一亚基具有选自354、356、358和366的位点的一个或多个氨基酸取代，所述第二亚基具有选自349、356、358、366、368和407的位点的一个或多个氨基酸取代。在一些实施方案中，所述第二亚基具有选自354、356、358和366的位点的一个或多个氨基酸取代，所述第一亚基具有选自349、356、358、366、368和407的位点的一个或多个氨基酸取代。在一些实施方案中，所述第一亚基具有选自354C、356E、358M和366W的一个或多个氨基酸取代，所述第二亚基具有选自349C、356E、358M、366S、368A和407V的一个或多个氨基酸取代。在一些实施方案中，所述第二亚基具有选自354C、356E、358M和366W的一个或多个氨基酸取代，所述第一亚基具有选自349C、356E、358M、366S、368A和407V的一个或多个氨基酸取代。在一些实施方案中，所述第一亚基包括354C、356E、358M和366W的氨基酸取代，所述第二亚基包括349C、356E、358M、366S、368A和407V的氨基酸取代。在一些实施方案中，所述第二亚基包括354C、356E、358M和366W的氨基酸取代，所述第一亚基包括349C、356E、358M、366S、368A和407V的氨基酸取代。
在一些实施方案中，如上任一项所述的抗原结合分子，所述抗原结合模块1为scFv，所述抗原结合模块2为包含重链可变区P-VH、轻链可变区P-VL、重链恒定区CH和轻链恒定区CL的全长抗体，所述抗原结合模块1直接或通过连接子融合至抗原结合模块2的可变区的N端或抗原结合模块2的恒定区的C端；在一些实施方案中，所述scFv的N端直接或通过连接子融合至抗原结合模块2的重链恒定区CH的C端；在一些实施方案中，所述scFv的C端直接或通过连接子融合 至抗原结合模块2的重链可变区P-VH的N端。
在一些实施方案中，如上任一项所述的抗原结合分子，所述抗原结合分子包含具有式(a)所示结构的第一链和具有式(b)所示结构的第二链；或者包含具有式(c)所示结构的第一链和具有式(b)所示结构的第二链，其中，
式(a)：[P-VH]-[CH]-[连接子1]-[B-VH]-[连接子2]-[B-VL]，
式(b)：[P-VL]-[CL]，
式(c)：[P-VH]-[CH]-[连接子1]-[B-VL]-[连接子2]-[B-VH]。
在一些实施方案中，式(a)和式(c)中，其中的连接子1与连接子2是相同或不同的肽连接子。在一些实施方案中，所述肽连接子独立的具有L 1-(GGGGS)n-L 2的结构，其中，L 1是键、A、GS、GGS或GGGS，n是0、1、2、3、4、5、6、7、8、9或10，L2是键、G、GG、GGG或GGGG，并且所述肽连接子不是键。在一些实施方案中，所述肽连接子的长度为3-15个氨基酸残基。在一些实施方案中，所述连接子1和连接子2各自独立地选自式(d)连接子，其中式(d)：(G xS) y，其中x选自1-5的整数(例如1、2、3、4或5)，y选自1-6的整数(例如，1、2、3、4、5或6)。在一些实施方案中，所述连接子1选自：GGGS(SEQ ID NO：69)，所述连接子2选自：GGGGSGGGGSGGGGS(SEQ ID NO：70)。在一些实施方案中，所述的抗原结合分子，其中，所述第一链包含与SEQ ID NO：61具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述第二链包含与SEQ ID NO：50具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或者所述第一链包含与SEQ ID NO：62具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述第二链包含与SEQ ID NO：50具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列。在一些实施方案中，所述抗原结合分子具有：包含SEQ ID NO：61的氨基酸序列的第一链和包含SEQ ID NO：50的氨基酸序列的第二链，或者包含SEQ ID NO：62的氨基酸序列的第一链和包含SEQ ID NO：50的氨基酸序列的第二链。
在一些实施方案中，所述抗原结合分子具有：
(i)两条包含SEQ ID NO：61的氨基酸序列的第一链和两条包含SEQ ID NO：50的氨基酸序列的第二链，或者
(ii)两条包含SEQ ID NO：62的氨基酸序列的第一链和两条包含SEQ ID NO：50的氨基酸序列的第二链。
另一方面，本披露提供一种抗原结合分子，其特异性结合BAFF抗原，其包含重链可变区B-VH和轻链可变区B-VL，所述B-VH包含B-HCDR1、B-HCDR2和B-HCDR3，所述B-VL包含B-LCDR1、B-LCDR2和B-LCDR3，其中：所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：63中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3 分别包含SEQ ID NO：64中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，其中，SEQ ID NO：63为：
QVQLQQSGAEVKKPGSSVRVSCKASGGTFNNNAINWVRQAPGQX 26LEWMGX 1IX 2PMFGX 3AKYSX 4X 5FQGRVAITADESTGTASMELSSLRSEDTAVYYCARSRDX 6LLFPX 7X 8X 9LX 10X 11WGX 27GTMVTVSS，
SEQ ID NO：64为：
SSELTQDPAVSVALGQTVRVTCX 12GX 13X 14LX 15X 16X 17X 18ASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCX 19SRX 20X 21X 22GX 23X 24WX 25FGX 28GTELTVL，
其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V，X 26选自G或C，X 27选自R或G，X 28选自G或C；并且，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3不包含如下CDR组合：B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：1中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，并且B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：2中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据选自Kabat、IMGT、Chothia、AbM和Contact的相同的编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Kabat编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据IMGT编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Chothia编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据AbM编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Contact编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3是根据Kabat编号规则定义的，其中所述B-HCDR1包含氨基酸序列NNAIN(SEQ ID NO：18)，B-HCDR2包含氨基酸序列X 1IX 2PMFGX 3AKYSX 4X 5FQG(SEQ ID NO：65)，B-HCDR3包含氨基酸序列SRDX 6LLFPX 7X 8X 9LX 10X 11(SEQ ID NO：66)，B-LCDR1包含氨基酸序列X 12GX 13X 14LX 15X 16X 17X 18AS(SEQ ID NO：67)，B-LCDR2包含氨基酸序列GKNNRPS(SEQ ID NO：32)和B-LCDR3包含氨基酸序列X 19SRX 20X 21X 22GX 23X 24WX 25(SEQ ID NO：68)，其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V；并且所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3不包含如下CDR组合：B-HCDR1包含SEQ ID NO：18的氨基酸序列，B-HCDR2包含SEQ ID NO：19的氨基酸序列，B-HCDR3包含SEQ ID NO：20的氨基酸序列，B-LCDR1包含SEQ ID NO：31的氨基酸序列，B-LCDR2包含SEQ ID NO：32的氨基酸序列，和B-LCDR3包含SEQ ID NO：33的氨基酸序列。
在一些实施方案中，如上任一项所述特异性结合BAFF抗原的抗原结合分子，其中：(i)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：47中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：48中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或(ii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：5中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：12中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或(iii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：6中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：13中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或(iv)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：7中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：14中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或(v)所 述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：8中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：15中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或(vi)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：9中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：16中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或(vii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：10中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：17中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或(viii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：11中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：2中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列；并且所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据选自Kabat、IMGT、Chothia、AbM和Contact的相同的编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Kabat编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据IMGT编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Chothia编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据AbM编号规则定义的。
在一些实施方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据Contact编号规则定义的。
在一些实施方案中，其中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3是根据Kabat编号规则定义的，其中(i)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：28的氨基酸序列，所述B-HCDR3包含SEQ ID NO：29的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：43的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序 列，和所述B-LCDR3包含SEQ ID NO：44的氨基酸序列；或(ii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：21的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：34的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：35的氨基酸序列；或(iii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：22的氨基酸序列，所述B-HCDR3包含SEQ ID NO：23的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：31的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：36的氨基酸序列；或(iv)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：24的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：37的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：38的氨基酸序列；或(v)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：25的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：39的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：40的氨基酸序列；或(vi)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：26的氨基酸序列，所述B-HCDR3包含SEQ ID NO：27的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：41的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：42的氨基酸序列；或(vii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：30的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：31的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：33的氨基酸序列。
在一些实施方案中，如上任一项所述特异性结合BAFF抗原的抗原结合分子，其中，所述B-VH包含与SEQ ID NO：63具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，所述B-VL包含与SEQ ID NO：64具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，其中SEQ ID NO：63为：
QVQLQQSGAEVKKPGSSVRVSCKASGGTFNNNAINWVRQAPGQX 26LEWMGX 1IX 2PMFGX 3AKYSX 4X 5FQGRVAITADESTGTASMELSSLRSEDTAVYYCARSRDX 6LLFPX 7X 8X 9LX 10X 11WGX 27GTMVTVSS，
SEQ ID NO：64为：
SSELTQDPAVSVALGQTVRVTCX 12GX 13X 14LX 15X 16X 17X 18ASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCX 19SRX 20X 21X 22GX 23X 24WX 25FGX 28GTELTVL，其中，X 1选自S、A、V或G，X 2选自A、I、M或 S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V，X 26选自G或C，X 27选自R或G，X 28选自G或C；并且所述B-VH和B-VL不包含如下可变区组合：B-VH包含SEQ ID NO：1的氨基酸序列，且B-VL包含SEQ ID NO：2的氨基酸序列。
在一些实施方案中，如上任一项所述特异性结合BAFF抗原的抗原结合分子，
(i)所述B-VH包含与SEQ ID NO：47具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：48具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(ii)所述B-VH包含与SEQ ID NO：5具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：12具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(iii)所述B-VH包含与SEQ ID NO：6具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：13具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(iv)所述B-VH包含与SEQ ID NO：7具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：14具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(v)所述B-VH包含与SEQ ID NO：8具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：15具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(vi)所述B-VH包含与SEQ ID NO：9具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：16具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，或
(vii)所述B-VH包含与SEQ ID NO：10具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：17具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同 一性的氨基酸序列，或
(viii)所述B-VH包含与SEQ ID NO：11具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：2具有至少90％(例如至少90％、95％、96％、97％、98％或99％)序列同一性的氨基酸序列；
在一些实施方案中，如上任一项所述特异性结合BAFF抗原的抗原结合分子，所述B-VH包含SEQ ID NO：47的氨基酸序列和所述B-VL包含SEQ ID NO：48的氨基酸序列，或所述B-VH包含SEQ ID NO：5的氨基酸序列，和所述B-VL包含SEQ ID NO：12的氨基酸序列；或所述B-VH包含SEQ ID NO：6的氨基酸序列，和所述B-VL包含SEQ ID NO：13的氨基酸序列；或所述B-VH包含SEQ ID NO：7的氨基酸序列，和所述B-VL包含SEQ ID NO：14的氨基酸序列；或所述B-VH包含SEQ ID NO：8的氨基酸序列，和所述B-VL包含SEQ ID NO：15的氨基酸序列；或所述B-VH包含SEQ ID NO：9的氨基酸序列，和所述B-VL包含SEQ ID NO：16的氨基酸序列；或所述B-VH包含SEQ ID NO：10的氨基酸序列，和所述B-VL包含SEQ ID NO：17的氨基酸序列；或所述B-VH包含SEQ ID NO：11的氨基酸序列，和所述B-VL包含SEQ ID NO：2的氨基酸序列。
在一些实施方案中，如上任一项所述特异性结合BAFF抗原的抗原结合分子，其包括重链恒定区CH和轻链恒定区CL。在一些实施方案中，所述重链恒定区CH为人IgG重链恒定区，所述轻链恒定区CL为人λ或κ轻链恒定区。在一些实施方案中，所述重链恒定区CH为人IgG1重链恒定区，所述轻链恒定区CL为人λ轻链恒定区。在一些实施方案中，所述重链恒定区CH为人IgG1重链恒定区，所述轻链恒定区CL为人κ轻链恒定区。在一些实施方案中，所述重链恒定区CH包含SEQ ID NO：45的氨基酸序列，所述轻链恒定区CL包含SEQ ID NO：46的氨基酸序列。在一些实施方案中，所述特异性结合BAFF抗原的抗原结合分子为全长抗体。在一些实施方案中，所述特异性结合BAFF抗原的抗原结合分子的重链恒定区具有一个或更多个能够减少其Fc区与Fc受体结合的氨基酸取代。在一些实施方案中，所述Fc区具有YTE突变(M252Y、S254T和T256E)，L234A、L235A突变，和/或S228P突变，所述突变编号依据为EU索引。
在一些实施方案中，如上任一项所述抗原结合分子(例如包含特异性结合BAFF的抗原结合模块1和特异性结合IL-12和/或IL-23的抗原结合模块2的抗原结合分子，或者特异性结合BAFF抗原的抗原结合分子)，所述抗原结合分子具有以下特征中的至少一种(例如包含以下A-G中的任意1、2、3、4、5、6或7种特征)：
A.所述抗原结合分子能阻断人BAFF与人BAFF-R的结合；优选地，阻断人BAFF与人BAFF-R结合的IC50值小于11nM(例如小于11nM、小于或等于10nM、小于或等于9nM、小于或等于8nM、小于或等于7nM、小于或等于6nM、 小于或等于5nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；
B.所述抗原结合分子能抑制BAFF诱导的B细胞增殖；优选地，抗原结合分子能以小于0.20nM(例如小于0.2nM、小于或等于0.14nM、小于或等于0.13nM、小于或等于0.12nM、小于或等于0.10nM、小于或等于0.09nM、小于或等于0.08nM、小于或等于0.07nM、小于或等于0.06nM或更小)的IC50值抑制BAFF诱导的B细胞增殖；在一些实施方式中，所述IC50值根据本披露测试例3方法检测；
C.所述抗原结合分子能以小于9.99E-10M(例如小于9.99E-10M、小于等于9.00E-10M、小于或等于8.20E-10M、小于或等于7.40E-10M、小于或等于6.60E-10M、小于或等于5.20E-10M、小于或等于4.00E-10M、小于或等于3.00E-10M或更小)的KD值与人BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
D.所述抗原结合分子能以小于5.00E-10M(例如小于或等于4.00E-10M、小于或等于3.00E-10M或更小)的KD值与食蟹猴BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
E.所述抗原结合分子能以小于2.20E-10M(例如小于或等于2.00E-10M、小于或等于1.00E-10M或更小)的KD值与鼠BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
F.所述抗原结合分子能阻断人BAFF与人BCMA的结合；优选地，阻断人BAFF与人BCMA结合的IC50值小于0.9nM(例如小于或等于0.4nM、小于或等于0.35nM、小于或等于0.31nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；或
G.所述抗原结合分子能阻断人BAFF与人TACI的结合；优选地，阻断人BAFF与人TACI结合的IC50值小于0.6nM(例如小于或等于0.55nM、小于或等于0.4nM、小于或等于0.33nM、小于或等于0.23nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测。
在一些实施方案中，如上任一项所述抗原结合分子，所述包含特异性结合BAFF的抗原结合模块1和特异性结合IL-12和/或IL-23的抗原结合模块2的抗原结合分子，其具有以下特征中的至少一种(例如包含以下H-L中的任意1、2、3、4或5种特征)：
H.所述抗原结合分子能阻断人IL-12/23 p40与人IL-12Rβ1的结合；优选地，阻断人IL-12/23 p40与人IL-12Rβ1结合的IC50值小于3.6nM(例如小于或等于3.0nM、小于或等于2.0nM、小于或等于1.0nM、小于或等于0.5nM、小于或等于0.23nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；
I.抗原结合分子能抑制IL-12诱导IFNγ分泌；优选地，抗原结合分子抑制IL-12 诱导IFNγ分泌的IC50值小于3.0nM(例如小于或等于2.6nM、小于或等于1.6nM或更小)；在一些实施方式中，所述IC50值根据本披露测试例4方法检测；
J.抗原结合分子能抑制IL-23诱导IL-17分泌；优选地，抗原结合分子抑制IL-23诱导IL-17分泌的IC50值小于0.034nM(例如小于或等于0.031nM、小于或等于0.030nM或更小)；在一些实施方式中，所述IC50值根据本披露测试例4方法检测；
K.抗原结合分子能以小于2.7E-10M(例如小于或等于2.0E-10M、小于或等于1.0E-10M或更小)的KD值与食蟹猴IL-12/23 p40结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；或
L.抗原结合分子能以小于6.4E-11M(例如小于或等于6.3E-11M、小于或等于6.2E-11M或更小)的KD值与人IL-23结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测。
另一方面，本披露提供一种药物组合物，其含有：治疗有效量的如上任一项所述抗原结合分子，以及一种或更多种药学上可接受的载体、稀释剂、缓冲剂或赋形剂。
另一方面，本披露提供分离的核酸，其编码如上任一项所述抗原结合分子。
另一方面，本披露提供宿主细胞，其包含如上任一项所述的核酸。
另一方面，本披露提供一种治疗疾病的方法，所述方法包括向受试者施用如上任一项所述抗原结合分子或药物组合物的步骤。
在一些实施方案中，所述疾病为B细胞障碍或自身免疫性疾病。在一些实施方案中，所述B细胞障碍或自身免疫性疾病是与BAFF表达相关的疾病或病症。在一些实施方案中，所述自身免疫性疾病选自：系统性红斑狼疮、重症肌无力、多发性硬化、胰岛素依赖性糖尿病、克罗恩氏病、类风湿关节炎、多关节型青少年类风湿关节炎和银屑病性关节炎；所述B细胞障碍选自：肿瘤、慢性白细胞性白血病、多发性骨髓瘤、非霍奇金淋巴瘤、移植后淋巴组织增生病以及轻链丙球蛋白病。在一些实施方案中，所述自身免疫性疾病为系统性红斑狼疮。
另一方面，本披露还提供如上任一项所述抗原结合分子或药物组合物在制备治疗前述疾病的药物中的用途。
另一方面，本披露还提供用作药物的如上任一项所述抗原结合分子或药物组合物。
附图说明
图1：BU-1的结构示意图。
图2：BU-2的结构示意图。
图3：特异性结合BAFF和IL-12/23双特异性抗体抑制TNFα分泌实验结果图。
图4：特异性结合BAFF和IL-12/23双特异性抗体抑制IgA分泌实验结果图。
图5：特异性结合BAFF和IL-12/23双特异性抗体抑制IL-22分泌实验结果图。
图6：特异性结合BAFF和IL-12/23双特异性抗体抑制IFNγ分泌实验结果图。
具体实施方式
术语
为了更容易理解本披露，以下对某些技术和科学术语进行了描述。除非在本文中另有明确定义，本文使用的全部技术和科学术语具有与本领域的普通技术人员通常所理解的相同含义。
说明书和权利要求书中所用的单数形式“一个”、“一种”和“所述”包括复数指代，除非上下文清楚表明并非如此。
除非上下文另外清楚要求，否则在专利说明书和权利要求书中，应将词语“包含”、“具有”、“包括”等理解为“包括但不仅限于”的意义，而不是排他性或穷举性意义。
术语“细胞因子”是由一个细胞群体释放的、作为细胞间介质作用于其它细胞的蛋白质/多肽的一般术语。这样的细胞因子的例子包括淋巴因子、单核因子、趋化因子和传统的多肽激素。示例性的细胞因子包括：IL-2、IFN-γ、IL-6、TNFα、IL-17和IL-5。
术语“和/或”，意指包含“和”与“或”两种含义。例如短语“A、B和/或C”旨在涵盖以下方面中的每一个：A、B和C；A、B或C；A或C；A或B；B或C；A和C；A和B；B和C；A(单独)；B(单独)；和C(单独)。本披露中，除非特别说明，“IL-12和/或IL-23”或“IL-12/23”均意指涵盖：“IL-12”、“IL-23”和“IL-12和IL-23”中的每一个。
本披露所用氨基酸三字母代码和单字母代码如J.biol.chem，243，p3558(1968)中所述。
术语“氨基酸”是指天然存在的和合成的氨基酸，以及以与天然存在的氨基酸类似的方式起作用的氨基酸类似物和氨基酸模拟物。天然存在的氨基酸是由遗传密码编码的那些氨基酸，以及后来修饰的那些氨基酸，例如羟脯氨酸、γ-羧基谷氨酸和O-磷酸丝氨酸。氨基酸类似物是指与天然存在的氨基酸具有相同基本化学结构(即与氢、羧基、氨基和R基团结合的α碳)的化合物，例如高丝氨酸、正亮氨酸、甲硫氨酸亚砜、甲硫氨酸甲基锍。此类类似物具有修饰的R基团(例如，正亮氨酸)或修饰的肽骨架，但保留与天然存在的氨基酸相同的基本化学结构。氨基酸模拟物是指具有与氨基酸的一般化学结构不同的结构，但是以与天然存在的氨基酸类似的方式起作用的化学化合物。
术语“氨基酸突变”包括氨基酸取代(也称氨基酸替换)、缺失、插入和修饰。可以进行取代、缺失、插入和修饰的任意组合来实现最终构建体，只要最终构建体拥有期望的特性，例如降低或对Fc受体的结合。氨基酸序列缺失和插入包括在 多肽链的氨基端和/或羧基端的缺失和插入。具体的氨基酸突变可以是氨基酸取代。在一个实施方式中，氨基酸突变是非保守性的氨基酸取代，即将一个氨基酸用具有不同结构和/或化学特性的另一种氨基酸替换。氨基酸取代包括由非天然存在的氨基酸或由20种天然氨基酸的衍生物(例如4-羟脯氨酸、3-甲基组氨酸、鸟氨酸、高丝氨酸、5-羟赖氨酸)替换。可以使用本领域中公知的遗传或化学方法生成氨基酸突变。遗传方法可以包括定点诱变、PCR，基因合成等。基因工程以外的改变氨基酸侧链基团的方法，如化学修饰也可能是可用的。本文中可使用各种名称来指示同一氨基酸突变。本文中，可采用位置+氨基酸残基的方式表示特定位点的氨基酸残基，例如366W，表示在366位点上的氨基酸残基为W。T366W则表示第366位点上的氨基酸残基由原来的T突变为了W。
术语“抗原结合分子”以最广义使用，涵盖各种特异性结合抗原的分子，包括但不限于抗体、其他具有抗原结合活性的多肽以及两者融合而成的抗体融合蛋白。示例性的，本文中的抗原结合分子是双特异性抗原结合分子(例如：双特异性抗体)。术语“双特异性抗原结合分子”指能够对两个不同抗原或同一抗原的至少两个不同抗原表位特异性结合的抗原结合分子。
术语“抗体”以最广义使用，并且涵盖各种抗体结构，包括但不限于单克隆抗体，多克隆抗体；单特异性抗体，多特异性抗体(例如双特异性抗体)；全长抗体和抗体片段(或抗原结合片段，或抗原结合部分)，只要它们展现出期望的抗原结合活性。“天然抗体”指天然存在的免疫球蛋白分子。例如，天然IgG抗体是约150,000道尔顿的异四聚蛋白，由二硫键结合的两条轻链和两条重链构成。从N至C端，每条重链具有一个可变区(VH)，又称作可变重域、重链可变区，接着是重链恒定区(CH)，天然IgG重链恒定区通常含三个恒定域(CH1、CH2和CH3)。类似地，从N至C端，每条轻链具有一个可变区(VL，又称作可变轻域，或轻链可变域)，接着是一个恒定轻域(轻链恒定区、CL)。术语“全长抗体”、“完整抗体”和“全抗体”在本文可互换使用，指具有与天然抗体结构基本类似的结构或具有如本文所限定的Fc区的重链的抗体。天然完整抗体轻链包括轻链可变区VL及恒定区CL，VL处于轻链的氨基末端，轻链恒定区包括κ链及λ链；重链包括可变区VH及恒定区(CH1、CH2及CH3)，VH处于重链的氨基末端(也称N端)，恒定区处于羧基末端(也称C端)，其中CH3最接近多肽的羧基末端，重链可属于任何同种型，包括IgG(包括IgG1、IgG2、IgG3及IgG4亚型)、IgA(包括IgA1及IgA2亚型)、IgM及IgE。
术语“双特异性抗体”指能够对两个不同抗原或同一抗原的两个不同抗原表位特异性结合的抗体(包括抗体或其抗原结合片段，如单链抗体)。现有技术已公开了各种结构的双特异性抗体。根据IgG分子的完整性可分为IgG样双特异性抗体和抗体片段型双特异性抗体。根据抗原结合区域的数量可分为二价、三价、四价或更多价的双特异性抗体。根据结构是否对称可分为对称结构双特异性抗体和 不对称结构双特异性抗体。其中，片段型双特异性抗体，例如缺乏Fc片段的Fab片段，其通过将2个或多个Fab片段结合在一个分子中形成双特异性抗体，其具有较低的免疫原性，且分子量小，具有较高的肿瘤组织渗透性，该类型的典型的抗体结构如F(ab) 2、scFv-Fab、(scFv) 2-Fab等。IgG样双特异性抗体(例如具有Fc片段)，这类抗体相对分子量较大，Fc片段有助于抗体的纯化，并提高其溶解性、稳定性，Fc部分还可能会与受体FcRn结合，增加抗体血清半衰期。典型的双特异性抗体结构模型如KiH、CrossMAb、Triomab quadroma、FcΔAdp、ART-Ig、BiMAb、Biclonics、BEAT、DuoBody、Azymetric、XmAb、2:1 TCBs、1Fab-IgG TDB、FynomAb、two-in-one/DAF、scFv-Fab-IgG、DART-Fc、LP-DART、CODV-Fab-TL、HLE-BiTE、F(ab)2-CrossMAb、IgG-(scFv)2、Bs4Ab、DVD-Ig、Tetravalent-DART-Fc、(scFv)4-Fc、CODV-Ig、mAb2、F(ab)4-CrossMAb等双特异性抗体(参见Aran F.Labrijn等，Nature Reviews Drug Discovery volume 18，pages585–608(2019)；Chen S1等，J Immunol Res.2019 Feb 11；2019:4516041)。
术语“可变区”或“可变域”指抗原结合分子(例如抗体)中结合抗原的域。本文中，重链可变区VH和轻链可变区VL各包含四个保守的框架区(FR)和三个互补决定区(CDR)。本文中，特异性结合BAFF的抗原结合模块中的重链可变区标示为B-VH，轻链可变区标示为B-VL；特异性结合IL-12/23 p40的抗原结合模块中的重链可变区标示为P-VH，轻链可变区标示为P-VL。术语“互补决定区”或“CDR”指可变结构域内主要促成与抗原结合的区域；“框架”或“FR”是指除CDR残基之外的可变结构域残基。VH包含3个CDR区：HCDR1、HCDR2和HCDR3；VL包含3个CDR区：LCDR1、LCDR2和LCDR3。每个VH和VL由从氨基末端(也称N末端)排到羧基末端(也称C末端)按以下顺序排列的三个CDR和四个FR构成：FR1、CDR1、FR2、CDR2、FR3、CDR3、FR4。每个VH和VL由从氨基末端排到羧基末端按以下顺序排列的三个CDR和四个FR构成：FR1、CDR1、FR2、CDR2、FR3、CDR3、FR4。单个VH或VL可能足以赋予抗原结合特异性。本披露中，B-VH中的3个CDR区分别标示为B-HCDR1、B-HCDR2和B-HCDR3，或者Bv-HCDR1、Bv-HCDR2和Bv-HCDR3；B-VL中的3个CDR区分别标示为B-LCDR1、B-LCDR2和B-LCDR3，或者Bv-LCDR1、Bv-LCDR2和Bv-LCDR3。另外，P-VH中的3个CDR区分别标示为P-HCDR1、P-HCDR2和P-HCDR3，或者Pv-HCDR1、Pv-HCDR2和Pv-HCDR3；P-VL中的3个CDR区分别标示为P-LCDR1、P-LCDR2和P-LCDR3，或者Pv-LCDR1、Pv-LCDR2和Pv-LCDR3。
可以通过各种公知方案来确定CDR的氨基酸序列边界，例如：“Kabat”编号规则(参见Kabat等(1991)，“Sequences of Proteins of Immunological Interest”，第5版，Public Health Service，National Institutes of Health，Bethesda，MD)、“Chothia”编号规则、“ABM”编号规则、“contact”编号规则(参见Martin,ACR.Protein Sequence  and Structure Analysis of Antibody Variable Domains[J].2001)和ImMunoGenTics(IMGT)编号规则(Lefranc，M.P.等，Dev.Comp.Immunol.，27，55-77(2003)；Front Immunol.2018 Oct 16；9:2278)等；各种编号系统之间的对应关系是本领域技术人员熟知的，示例性的，如下表1中所示。
表1.CDR编号系统之间的关系
CDR IMGT Kabat AbM Chothia Contact
HCDR1 27-38 31-35 26-35 26-32 30-35
HCDR2 56-65 50-65 50-58 52-56 47-58
HCDR3 105-117 95-102 95-102 95-102 93-101
LCDR1 27-38 24-34 24-34 24-34 30-36
LCDR2 56-65 50-56 50-56 50-56 46-55
LCDR3 105-117 89-97 89-97 89-97 89-96
除非另有说明，本披露中的可变区和CDR序列均适用“Kabat”编号规则。尽管在具体的实施方案中，采用了Kabat编号规则来限定氨基酸残基，但是其他编号系统所的对应技术方案将视为等同技术方案。
术语“抗体片段”指不同于完整抗体的分子，其包含完整抗体的部分，所述部分保留了完整抗体的抗原结合能力。抗体片段的实例包括但不限于Fv、Fab、Fab’、Fab’-SH、F(ab′) 2、单域抗体、单链Fab(scFab)、双抗体、线性抗体、单链抗体(scFv)，以及由抗体片段形成的多特异性抗体。
术语“Fc区”或“片段可结晶区”用于定义抗体重链的C末端区域，包括天然Fc区和改造的Fc区。在一些实施方式中，Fc区包含了相同或不同的两个亚基。在一些实施方式中，人IgG重链的Fc区定义为从Cys226位置处的氨基酸残基或从Pro230延伸至其羧基末端。用于本文所述抗体的合适Fc区包括人IgG1、IgG2(IgG2A、IgG2B)、IgG3和IgG4的Fc区。在一些实施方式中，Fc区的边界还可以变化，例如缺失Fc区的C末端赖氨酸(根据EU编号系统的残基447)或缺失Fc区的C末端甘氨酸和赖氨酸(根据EU编号系统的残基446和447)。除非另有说明，Fc区的编号规则为EU编号系统，又称作EU索引。
术语“嵌合”抗体指抗体中的重和/或轻链的一部分来自特定的来源或物种，而重和/或轻链的剩余部分来自另外的不同来源或物种的抗体。
术语“人源化”抗体是保留非人抗体的反应性同时在人中具有较低免疫原性的抗体。例如，可以通过保留非人CDR区，并用其人对应物(即，恒定区以及可变区的框架区部分)替换抗体的其余部分来实现人源化。
术语“人抗体”、“人源抗体”、“全人抗体”、“完全人抗体”可以互换使用，意指可变区及恒定区是人序列的抗体。该术语涵盖源自人基因但具有，例如，降低可能的免疫原性、增加亲和力、消除可能会引起不期望的折叠的半胱氨酸或糖基化位点等序列已发生改变的抗体。该术语涵盖这些在非人细胞(其可能会赋予不具人细胞特征的糖基化)中重组产生的抗体。该术语亦涵盖已在含有一些或所有人 免疫球蛋白重链及轻链基因座的转基因小鼠中饲养的抗体。人抗体的含义明确排除包含非人抗原结合残基的人源化抗体。
术语“亲和力”是指分子(例如，本公开的抗原结合分子)的单个结合部位与其结合配体(例如，抗原)之间非共价相互作用的总体的强度。除非另外指明，如本文所用，结合“亲和力”是指内部结合亲和力，其反映出结合对(例如，抗体与抗原)的成员之间1:1相互作用。分子X对其配体Y的亲和力通常可以由解离常数(KD)表示。亲和力可以通过本领域已知的常规方法(包括本文所述的那些方法)测量。
如本文所使用的，术语“kassoc”或“ka”指特定抗体-抗原相互作用的缔合速率，术语“kdis”或“kd”指特定抗体-抗原相互作用的解离速率。术语“KD”指解离常数，其获得自kd与ka的比率(即kd/ka)并且表示为摩尔浓度(M)。可以使用本领域公知的方法测定抗体的KD值。例如，使用生物传感系统例如系统测量表面等离子体共振，或通过溶液平衡滴定法(SET)测量溶液中的亲和力。在一些实施方式中，KD值通过Biacore检测。
术语“效应子功能”指那些可归于抗体Fc区(天然序列Fc区或氨基酸序列突变的Fc区)且随抗体同种型而变化的生物学活性。抗体效应子功能的例子包括但不限于：C1q结合和补体依赖性细胞毒性、Fc受体结合、抗体依赖性细胞介导的细胞毒性(ADCC)、吞噬作用、细胞表面受体(例如B细胞受体)下调；和B细胞活化。
术语“单克隆抗体”指基本上均质的抗体的群或其成员，即在该群中包含的抗体分子的氨基酸序列是相同的，除了可能少量存在的天然突变以外。相比之下，多克隆抗体制剂通常包含在其可变结构域具有不同氨基酸序列的多种不同抗体，其通常特异性针对不同表位。“单克隆”表示从基本上均质的抗体群体获得的抗体的特征，并且不应解释为要求通过任何特定方法来生产抗体。在一些实施方式中，本披露提供的抗体是单克隆抗体。
术语“抗原”是指能够由抗原结合分子(例如抗体)选择性结合的分子或分子部分。抗原可具有一个或多个能够与不同的抗原结合分子(例如抗体)相互作用的表位。
术语“表位”指能够与抗体或其抗原结合片段特异性结合的抗原上的区域(area或region)。表位可以由连续氨基酸(线性表位)形成或包含非连续氨基酸(构象表位)，例如因抗原的折叠(即通过抗原的三级折叠)而使得非连续的氨基酸在空间上得以接近。构象表位和线性表位的差别在于：在变性溶剂的存在下，抗体对构象表位的结合丧失。表位包含处于独特空间构象的至少3，至少4，至少5，至少6，至少7，或8-10个氨基酸。可以使用本领域例行方法来筛选结合特定表位的抗体(即那些结合相同表位的)，例如但不限于丙氨酸扫描，肽印迹，肽切割分析，表位切除，表位提取，抗原的化学修饰(见Prot.Sci.9(2000)487-496)， 和交叉阻断。
术语“能够特异性结合”、“特异性结合”或“结合”是指相比其他抗原或表位，抗原结合分子(例如抗体)能够以更高的亲和力结合至某个抗原或其表位。通常地，抗体以约1×10 -7M或更小(例如约1×10 -8M或更小)的平衡解离常数(KD)结合抗原或其表位。在一些实施方式中，抗体与抗原结合的KD为该抗体结合至非特异性抗原(例如BSA、酪蛋白)的KD的10％或更低(例如1％)。可使用已知的方法来测量KD，例如通过
Figure PCTCN2022102621-appb-000001
表面等离子体共振测定法所测量的。然而，特异性结合至抗原或其表位的抗体不排除对其它相关的抗原具有交叉反应性，例如，对来自其它物种(同源)(诸如人或猴，例如食蟹猕猴(Macaca fascicularis)(cynomolgus，cyno)、黑猩猩(Pan troglodytes)(chimpanzee，chimp))或狨猴(Callithrix jacchus)(commonmarmoset，marmoset)的相应抗原具有交叉反应性。
术语“抗原结合模块”指特异性结合目标抗原的多肽分子。抗原结合模块包括如本文中另外定义的抗体及其片段。在一些实施方案中，抗原结合模块包括抗体的抗原结合域，其包含抗体重链可变区和抗体轻链可变区。术语“特异性结合BAFF的抗原结合模块”是指能够以足够的亲和力结合BAFF的抗原结合模块。在某些实施方案中，特异性结合人BAFF的抗原结合模块具有以下的平衡解离常数(KD)：小于10.00E-10M(例如小于9.99E-10M、小于等于9.00E-10M、小于或等于8.20E-10M、小于或等于7.40E-10M、小于或等于6.60E-10M、小于或等于5.20E-10M、小于或等于4.00E-10M、小于或等于3.00E-10M或更小)，其是通过Biacore方法测量的。在某些实施例中，特异性结合BAFF的抗原结合模块结合来自不同物种的BAFF中的保守表位。术语“特异性结合IL-12/23的抗原结合模块”是指能够以足够的亲和力结合IL-12/23的抗原结合模块。在某些实施例中，特异性结合IL-12/23的抗原结合模块具有以下的平衡解离常数(KD)：小于6.4E-11M(例如小于或等于6.3E-11M、小于或等于6.2E-11M或更小)的KD值与人IL-23结合，其是通过Biacore方法检测的。在某些实施例中，特异性结合IL-12/23的抗原结合模块结合来自不同物种的IL-12/23中的保守表位。抗原结合模块包括如本文定义的抗体片段，例如Fab或scFv。
在上下文中，“抗原结合模块1”或“抗原结合模块2”中的序数词，其目的仅在于区分不同的技术特征、化学实体；而不限定任何顺序、水平、数量。
术语“连接子”、“Linker”或“接头”指连接两个多肽片段的连接单元。在本文中，同一结构中出现的连接子可以是相同或不同的。连接子可以是肽连接子，其包含一个或多个氨基酸，典型的约1-30个、2-24个或3-15个氨基酸。应用于本文的连接子可以是相同或不同的。当“-”出现在结构式中，其表示两侧的单元直接通过共价键连接。当术语“键”出现在结构单元，其表示该单元没有氨基酸，单元两侧的单元直接连接。
术语“抗体依赖性细胞的细胞毒性”、“抗体依赖性细胞介导的细胞毒性”或“ADCC”是诱导细胞死亡的机制，该机制依赖于抗体包被的靶细胞与具有裂解活性的效应细胞(诸如自然杀伤细胞(NK)、单核细胞、巨噬细胞和中性粒细胞)经由效应细胞上表达的Fcγ受体(FcγR)发生的相互作用。例如，NK细胞表达FcγRIIIa，而单核细胞表达FcγRI、FcγRII和FcγRIIIa。本文提供的抗体的ADCC活性可使用体外测定，使用表达抗原的细胞作为靶细胞和NK细胞作为效应细胞进行评定。根据从裂解的细胞中释放的标记物(例如放射性底物、荧光染料或天然胞内蛋白)来检测细胞裂解。
术语“抗体依赖性细胞吞噬作用”(“ADCP”)是指通过吞噬细胞(诸如巨噬细胞或树突状细胞)的内化作用消除抗体包被的靶细胞的机制。
术语“补体依赖性细胞毒性”或“CDC”是指诱导细胞死亡的机制，其中靶细胞上所结合的抗体的Fc效应域结合并激活补体成分C1q，C1q继而激活补体级联，从而导致靶细胞死亡。补体的激活也可导致补体成分沉积在靶细胞表面上，这些补体成分通过结合白细胞上的补体受体(例如，CR3)来促进CDC。
术语“核酸”在本文中可与术语“多核苷酸”互换使用，并且是指呈单链或双链形式的脱氧核糖核苷酸或核糖核苷酸及其聚合物。所述术语涵盖含有已知核苷酸类似物或修饰的骨架残基或连接的核酸，所述核酸是合成的、天然存在的和非天然存在的，具有与参考核酸相似的结合特性，并且以类似于参考核苷酸的方式代谢。此类类似物的实例包括但不限于硫代磷酸酯、氨基磷酸酯、甲基膦酸酯、手性-甲基膦酸酯、2-O-甲基核糖核苷酸、肽-核酸(PNA)。
“分离的”核酸指已经与其天然环境的组分分开的核酸分子。分离的核酸包括在下述细胞中含有的核酸分子，所述细胞通常含有该核酸分子，但该核酸分子存在于染色体外或存在于不同于其天然染色体位置的染色体位置处。编码抗原结合分子的分离的核酸指编码抗原结合分子的一个或更多个核酸分子，包括在单一载体或分开的载体中的这样的一个或更多个核酸分子，和存在于宿主细胞中一个或更多个位置的这样的一个或更多个核酸分子。除非另有说明，否则特定的核酸序列还隐含地涵盖其保守修饰的变体(例如，简并密码子取代)和互补序列以及明确指明的序列。具体地，如下详述，简并密码子取代可以通过产生如下序列而获得，在这些序列中，一个或多个所选的(或全部)密码子的第三位被简并碱基和/或脱氧肌苷残基取代。
术语“多肽”和“蛋白质”在本文中可互换使用，指氨基酸残基的聚合物。该术语适用于氨基酸聚合物，其中一个或多个氨基酸残基是天然存在的氨基酸的相应人工化学模拟物，以及适用于天然存在的氨基酸聚合物和非天然存在的氨基酸聚合物。除非另外说明，否则特定的多肽序列还隐含地涵盖其保守修饰的变体。
术语序列“同一性”指，当对两条序列进行最佳比对时，两条序列的氨基酸/核酸在等价位置相同的程度(百分比)；在进行最佳比对时，必要时允许引入间隙 以获取最大序列同一性百分比，且不将任何保守性取代视为序列同一性的一部分。为测定序列同一性百分比，比对可以通过本领域技术已知的技术来实现，例如使用公开可得到的计算机软件，诸如BLAST、BLAST-2、ALIGN、ALIGN-2或Megalign(DNASTAR)软件。本领域技术人员可确定适用于测量比对的参数，包括在所比较的序列全长上达成最大比对所需的任何算法。
术语“融合”或“连接”是指部件(例如B-VH与B-VL)直接地或经由一个或多个连接子而通过共价键连接。
术语“载体(vector)”意指能够转运与其连接的另一多核苷酸的多核苷酸分子。一种类型的载体是“质粒”，其是指环状双链DNA环，其中可以连接附加的DNA区段。另一种类型的载体是病毒载体，例如腺相关病毒载体(AAV或AAV2)，其中另外的DNA区段可以连接到病毒基因组中。某些载体能够在引入它们的宿主细胞中自主复制(例如，具有细菌复制起点的细菌载体和附加型哺乳动物载体)。其他载体(例如，非附加型哺乳动物载体)可以在引入宿主细胞中后整合到宿主细胞的基因组中，从而与宿主基因组一起复制。术语“表达载体”或“表达构建体”是指可对宿主细胞进行转化，且含有指导和/或控制(连同宿主细胞一起)与其可操作地连接的一个或多个异源编码区的表达的核酸序列的载体。表达构建体可以包括但不限于影响或控制转录、翻译且在存在内含子时影响与其可操作地连接的编码区的RNA剪接的序列。
术语“宿主细胞”、“宿主细胞系”和“宿主细胞培养物”可互换使用，并且指已经导入外源核酸的细胞，包括此类细胞的后代。宿主细胞包括“转化体”和“经转化的细胞”，其包括原代的经转化的细胞及自其衍生的后代，而不考虑传代的次数。后代在核酸内容物上可以与亲本细胞不完全相同，而是可以含有突变。本文中，该术语包括突变体后代，其包括具有与在原代转化细胞中筛选或选择的细胞具有相同功能或生物学活性。宿主细胞包括原核和真核宿主细胞，其中真核宿主细胞包括但不限于哺乳动物细胞、昆虫细胞系植物细胞和真菌细胞。哺乳动物宿主细胞包括人、小鼠、大鼠、犬、猴、猪、山羊、牛、马和仓鼠细胞，包括但不限于中国仓鼠卵巢(CHO)细胞、NSO、SP2细胞、HeLa细胞、幼仓鼠肾(BHK)细胞、猴肾细胞(COS)、人肝细胞癌细胞(例如，Hep G2)、A549细胞、3T3细胞和HEK-293细胞。真菌细胞包括酵母和丝状真菌细胞，包括例如巴氏毕赤酵母(Pichiapastoris)、芬兰毕赤酵母(Pichia finlandica)、海藻毕赤酵母(Pichia trehalophila)、科克拉马毕赤酵母(Pichia koclamae)、膜状毕赤酵母(Pichia membranaefaciens)、小毕赤酵母(Pichia minuta)(Ogataea minuta、Pichia lindneri)、仙人掌毕赤酵母(Pichiaopuntiae)、耐热毕赤酵母(Pichia thermotolerans)、柳毕赤酵母(Pichia salictaria)、Pichia guercuum、皮杰普毕赤酵母(Pichia pijperi)、具柄毕赤酵母(Pichia stiptis)、甲醇毕赤酵母(Pichia methanolica)、毕赤酵母属、酿酒酵母(Saccharomycescerevisiae)、酿酒酵母属、多形汉逊酵母(Hansenula  polymorpha)、克鲁维酵母属、乳酸克鲁维酵母(Kluyveromyces lactis)、白色念珠菌(Candida albicans)、构巢曲霉(Aspergillus nidulans)、黑曲霉(Aspergillus niger)、米曲霉(Aspergillus oryzae)、里氏木霉(Trichoderma reesei)、勒克氏菌(Chrysosporium lucknowense)、镰刀菌属(Fusarium sp.)、禾谷镰刀菌(Fusarium gramineum)、菜镰刀菌(Fusarium venenatum)、小立碗藓(Physcomitrella patens)和粗糙脉孢菌(Neurospora crassa)。毕赤酵母属、任何酿酒酵母属、多形汉逊酵母(Hansenula polymorpha)、任何克鲁维酵母属、白色念珠菌(Candida albicans)、任何曲霉属、里氏木霉(Trichoderma reesei)、勒克霉菌(Chrysosporium lucknowense)、任何镰刀菌属、解脂耶氏酵母(Yarrowia lipolytica)和粗糙脉孢菌(Neurospora crassa)。
所述宿主细胞不能发育成植株或动物个体。
“任选”或“任选地”意味着随后所描述地特征可以但不必发生，将指明包括该特征发生或不发生的场合。
术语“药物组合物”表示含有一种或多种本文所述的抗原结合分子与其他化学组分的混合物，所述其他组分例如生理学/可药用的载体和赋形剂。
术语“药学上可接受的载体(vehicle)、稀释剂、缓冲剂或赋形剂”指药学制剂(formulation)中与活性成分不同的，且对受试者无毒的成分。
术语“受试者”或“个体”包括人类和非人类动物。非人动物包括所有脊椎动物(例如哺乳动物和非哺乳动物)例如非人灵长类(例如，食蟹猴)、绵羊、狗、牛、鸡、两栖动物和爬行动物。除非指出时，否则所述术语“患者”或“受试者”在本文中可互换地使用。如本文所使用的，术语“食蟹猴(cyno)”或“食蟹猴(cynomolgus)”是指食蟹猴(Macaca fascicularis)。在某些实施方案中，个体或受试者是人。
“施用”或“给予”，当其应用于动物、人、实验受试者、细胞、组织、器官或生物流体时，是指外源性药物、治疗剂、诊断剂或组合物与动物、人、受试者、细胞、组织、器官或生物流体的接触。
术语“样品”是指从受试者分离的采集物(如流体、细胞、或组织)，以及存在于受试者体内的流体、细胞或组织。示例性样品为生物流体，诸如血液、血清和浆膜液、血浆、淋巴液、尿液、唾液、囊液、泪液、排泄物、痰、分泌组织或分泌器官的粘膜分泌物、阴道分泌物、腹水、胸膜、心包、腹膜、腹腔和其它体腔的流体、由支气管灌洗液收集的流体、滑液、与受试者或生物来源接触的液体溶液，例培养基(包括条件培养基)、灌洗液等，组织活检样品、细针穿刺、手术切除的组织、器官培养物或细胞培养物。
“治疗(treatment或treat)”和“处理”(及其语法变型)指施加至所治疗个体的临床干预，并且可以为了预防目的、或者在临床病理学的过程期间进行实施。治疗的期望效果包括但不限于预防疾病的发生或复发，减轻症状，减轻/减少疾病的 任何直接或间接病理后果，预防转移，降低疾病进展速率，改善或减轻疾病状态，和消退或改善的预后。在一些实施方案中，使用本披露的分子来延迟疾病的形成或减缓疾病的进展。
“有效量”一般是足以降低症状的严重程度和/或频率、消除这些症状和/或潜在病因、预防症状和/或其潜在病因出现和/或改良或改善由疾病状态引起或与其相关的损伤的量。在一些实施例中，有效量是治疗有效量或预防有效量。
“治疗有效量”是足以治疗疾病状态或症状、尤其与该疾病状态相关的状态或症状，或者以其他方式阻碍、延迟或逆转该疾病状态或与该疾病相关的任何其他不理想症状的进展的量。
“预防有效量”是当给予受试者时将具有预定预防效应，例如预防或延迟该疾病状态的发作(或复发)，或者降低该疾病状态或相关症状的发作(或复发)可能性的量。完全治疗或预防效未必在给予一个剂量之后便发生，可能在给予一系列剂量之后发生。因而，治疗或预防有效量可以一次或多次给予的方式给予。“治疗有效量”和“预防有效量”可取决于多种因素变化：诸如个体的疾病状态、年龄、性别和体重，以及治疗剂或治疗剂组合在个体中引发期望的应答的能力。有效治疗剂或治疗剂组合的示例性指标包括例如患者改善的健康状况。
靶分子
“BAFF”应作广泛的理解，旨在涵盖BAFF在哺乳动物体内各阶段中的各种形式的分子，例如但不限于BAFF基因在扩增、复制、转录、剪接、加工、翻译、修饰过程中所产生的分子(例如前体BCMA、成熟BAFF、膜表达的BAFF、BAFF剪接变体、修饰的BAFF、或其片段)；该术语也涵盖人工制备的或体外表达的BAFF。
“IL-12”应作广泛的理解，旨在涵盖IL-12在哺乳动物体内各阶段中的各种形式的分子，例如但不限于IL-12基因在扩增、复制、转录、剪接、加工、翻译、修饰过程中所产生的分子(例如前体IL-12、成熟IL-12、膜表达的IL-12、IL-12剪接变体、修饰的IL-12、或其片段、亚基)；该术语也涵盖人工制备的或体外表达的IL-12。
“IL-23”应作广泛的理解，旨在涵盖IL-23在哺乳动物体内各阶段中的各种形式的分子，例如但不限于IL-23基因在扩增、复制、转录、剪接、加工、翻译、修饰过程中所产生的分子(例如前体IL-23、成熟IL-23、膜表达的IL-23、IL-23剪接变体、修饰的IL-23、或其片段、亚基)；该术语也涵盖人工制备的或体外表达的IL-23。
本披露的抗原结合分子
在一个方面中，本披露通过对Belimumab进行改造，获得了一种特异性结合BAFF的抗原结合分子。另外，还构建了特异性结合BAFF和IL-12/23的抗原结合分子。在一些实施方案中，本披露的抗原结合分子其具有诸多有利的特性，例如 高亲和力与抗原结合、阻断BAFF与其受体(例如BAFF-R、BCMA和/或TACI)的结合、抑制BAFF诱导的B细胞增殖、阻断IL-12/23与其受体结合、抑制IL-12诱导IFNγ分泌、抑制IL-23诱导IL-17分泌、药物代谢动力学特性和/或成药性等。
在一些实施方案中，本披露提供一种特异性结合BAFF的抗原结合分子，其为抗BAFF抗体。在一些实施方案中，所述抗体为全长抗体或其抗原结合片段(例如Fv、Fab、Fab’、Fab’-SH、F(ab′)2、单域抗体、单链Fab(scFab)、双抗体、线性抗体、单链抗体(scFv))，其具有以下一个或更多个功能活性：
A.所述抗原结合分子能阻断人BAFF与人BAFF-R的结合；优选地，阻断人BAFF与人BAFF-R结合的IC50值小于11nM(例如小于11nM、小于或等于10nM、小于或等于9nM、小于或等于8nM、小于或等于7nM、小于或等于6nM、小于或等于5nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；
B.所述抗原结合分子能抑制BAFF诱导的B细胞增殖；优选地，抗原结合分子能以小于0.20nM(例如小于0.2nM、小于或等于0.14nM、小于或等于0.13nM、小于或等于0.12nM、小于或等于0.10nM、小于或等于0.09nM、小于或等于0.08nM、小于或等于0.07nM、小于或等于0.06nM或更小)的IC50值抑制BAFF诱导的B细胞增殖；在一些实施方式中，所述IC50值根据本披露测试例3方法检测；
C.所述抗原结合分子能以小于9.99E-10M(例如小于9.99E-10M、小于等于9.00E-10M、小于或等于8.20E-10M、小于或等于7.40E-10M、小于或等于6.60E-10M、小于或等于5.20E-10M、小于或等于4.00E-10M、小于或等于3.00E-10M或更小)的KD值与人BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
D.所述抗原结合分子能以小于5.00E-10M(例如小于或等于4.00E-10M、小于或等于3.00E-10M或更小)的KD值与食蟹猴BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
E.所述抗原结合分子能以小于2.20E-10M(例如小于或等于2.00E-10M、小于或等于1.00E-10M或更小)的KD值与鼠BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
F.所述抗原结合分子能阻断人BAFF与人BCMA的结合；优选地，阻断人BAFF与人BCMA结合的IC50值小于0.9nM(例如小于或等于0.4nM、小于或等于0.35nM、小于或等于0.31nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；或
G.所述抗原结合分子能阻断人BAFF与人TACI的结合；优选地，阻断人BAFF与人TACI结合的IC50值小于0.6nM(例如小于或等于0.55nM、小于或等于0.4nM、小于或等于0.33nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测。
在一些实施方案中，本披露提供一种特异性结合BAFF和IL-12/23的抗原结合分子。在一些实施方案中，所述抗原结合分子为特异性结合BAFF和IL-12/23 P40亚基的双特异性抗体。在一些实施方案中，所述抗原结合分子具有以下一个或更多个功能活性：
A.所述抗原结合分子能阻断人BAFF与人BAFF-R的结合；优选地，阻断人BAFF与人BAFF-R结合的IC50值小于11nM(例如小于11nM、小于或等于10nM、小于或等于9nM、小于或等于8nM、小于或等于7nM、小于或等于6nM、小于或等于5nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；
B.所述抗原结合分子能抑制BAFF诱导的B细胞增殖；优选地，抗原结合分子能以小于0.20nM(例如小于0.2nM、小于或等于0.14nM、小于或等于0.13nM、小于或等于0.12nM、小于或等于0.10nM、小于或等于0.09nM、小于或等于0.08nM、小于或等于0.07nM、小于或等于0.06nM或更小)的IC50值抑制BAFF诱导的B细胞增殖；在一些实施方式中，所述IC50值根据本披露测试例3方法检测；
C.所述抗原结合分子能以小于9.99E-10M(例如小于9.99E-10M、小于等于9.00E-10M、小于或等于8.20E-10M、小于或等于7.40E-10M、小于或等于6.60E-10M、小于或等于5.20E-10M、小于或等于4.00E-10M、小于或等于3.00E-10M或更小)的KD值与人BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
D.所述抗原结合分子能以小于5.00E-10M(例如小于或等于4.00E-10M、小于或等于3.00E-10M或更小)的KD值与食蟹猴BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
E.所述抗原结合分子能以小于2.20E-10M(例如小于或等于2.00E-10M、小于或等于1.00E-10M或更小)的KD值与鼠BAFF结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；
F.所述抗原结合分子能阻断人BAFF与人BCMA的结合；优选地，阻断人BAFF与人BCMA结合的IC50值小于0.9nM(例如小于或等于0.4nM、小于或等于0.35nM、小于或等于0.31nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；
G.所述抗原结合分子能阻断人BAFF与人TACI的结合；优选地，阻断人BAFF与人TACI结合的IC50值小于0.6nM(例如小于或等于0.55nM、小于或等于0.4nM、小于或等于0.33nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；
H.所述抗原结合分子能阻断人IL-12/23 p40与人IL-12Rβ1的结合；优选地，阻断人IL-12/23 p40与人IL-12Rβ1结合的IC50值小于3.6nM(例如小于或等于3.0nM、小于或等于2.0nM、小于或等于1.0nM、小于或等于0.5nM、小于或等于 0.23nM或更小)，所述IC50值通过Elisa方法检测；在一些实施方式中，所述IC50值根据本披露测试例2方法检测；
I.抗原结合分子能抑制IL-12诱导IFNγ分泌；优选地，抗原结合分子抑制IL-12诱导IFNγ分泌的IC50值小于3.0nM(例如小于或等于2.6nM、小于或等于1.6nM或更小)；在一些实施方式中，所述IC50值根据本披露测试例4方法检测；
J.抗原结合分子能抑制IL-23诱导IL-17分泌；优选地，抗原结合分子抑制IL-23诱导IL-17分泌的IC50值小于0.034nM(例如小于或等于0.031nM、小于或等于0.030nM或更小)；在一些实施方式中，所述IC50值根据本披露测试例4方法检测；
K.抗原结合分子能以小于2.7E-10M(例如小于或等于2.0E-10M、小于或等于1.0E-10M或更小)的KD值与食蟹猴IL-12/23 p40结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测；或
L.抗原结合分子能以小于6.4E-11M(例如小于或等于6.3E-11M、小于或等于6.2E-11M或更小)的KD值与人IL-23结合，所述KD值通过Biacore方法检测；在一些实施方式中，所述KD值根据本披露测试例5方法检测。
在一些实施方案中，如上任一项所述的抗原结合分子，其包含重链可变区B-VH和轻链可变区B-VL，所述B-VH包含B-HCDR1、B-HCDR2和B-HCDR3，所述B-VL包含B-LCDR1、B-LCDR2和B-LCDR3，其中，所述B-HCDR1、B-HCDR2和B-HCDR3的氨基酸序列分别与SEQ ID NO：63中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列相同，和所述B-LCDR1、B-LCDR2和B-LCDR3的氨基酸序列分别与SEQ ID NO：64中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列相同。在一些方案中，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3、Bv-HCDR1、Bv-HCDR2、Bv-HCDR3、Bv-LCDR1、Bv-LCDR2和Bv-LCDR3是根据相同的编号规则定义的，所述编号规则选自Kabat、IMGT、Chothia、AbM和Contact。
在一些实施方案中，如上任一项所述的抗原结合分子，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3是根据Kabat编号规则定义的，其中，B-HCDR1的氨基酸序列为NNAIN(SEQ ID NO：18)，B-HCDR2的氨基酸序列为X 1IX 2PMFGX 3AKYSX 4X 5FQG(SEQ ID NO：65)，B-HCDR3的氨基酸序列为SRDX 6LLFPX 7X 8X 9LX 10X 11(SEQ ID NO：66)，B-LCDR1的氨基酸序列为X 12GX 13X 14LX 15X 16X 17X 18AS(SEQ ID NO：67)，B-LCDR2的氨基酸序列为GKNNRPS(SEQ ID NO：32)和B-LCDR3包含的氨基酸序列为X 19SRX 20X 21X 22GX 23X 24WX 25(SEQ ID NO：68)，其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I， X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V。优选地，X 1至X 25不选自以下组合：X 1为G、X 2为I、X 3为T、X 4为Q、X 5为N、X 6为L、X 7为H、X 8为H、X 9为A、X 10为S、X 11为P、X 12为Q、X 13为D、X 14为S、X 15为R、X 16为S、X 17为Y、X 18为Y、X 19为S、X 20为D、X 21为S、X 22为S、X 23为N、X 24为H和X 25为V。
在一些实施方案中，(i)所述B-HCDR1如SEQ ID NO：18所示，所述B-HCDR2如SEQ ID NO：28所示，所述B-HCDR3如SEQ ID NO：29所示，和所述B-LCDR1如SEQ ID NO：43所示，所述B-LCDR2如SEQ ID NO：32所示，和所述B-LCDR3如SEQ ID NO：44所示；或(ii)所述B-HCDR1如SEQ ID NO：18所示，所述B-HCDR2如SEQ ID NO：21所示，所述B-HCDR3如SEQ ID NO：20所示，和所述B-LCDR1如SEQ ID NO：34所示，所述B-LCDR2如SEQ ID NO：32所示，和所述B-LCDR3如SEQ ID NO：35所示；或(iii)所述B-HCDR1如SEQ ID NO：18所示，所述B-HCDR2如SEQ ID NO：22所示，所述B-HCDR3如SEQ ID NO：23所示，和所述B-LCDR1如SEQ ID NO：31所示，所述B-LCDR2如SEQ ID NO：32所示，和所述B-LCDR3如SEQ ID NO：36所示；或(iv)所述B-HCDR1如SEQ ID NO：18所示，所述B-HCDR2如SEQ ID NO：24所示，所述B-HCDR3如SEQ ID NO：20所示，和所述B-LCDR1如SEQ ID NO：37所示，所述B-LCDR2如SEQ ID NO：32所示，和所述B-LCDR3如SEQ ID NO：38所示；或(v)所述B-HCDR1如SEQ ID NO：18所示，所述B-HCDR2如SEQ ID NO：25所示，所述B-HCDR3如SEQ ID NO：20所示，和所述B-LCDR1如SEQ ID NO：39所示，所述B-LCDR2如SEQ ID NO：32所示，和所述B-LCDR3如SEQ ID NO：40所示；或(vi)所述B-HCDR1如SEQ ID NO：18所示，所述B-HCDR2如SEQ ID NO：26所示，所述B-HCDR3如SEQ ID NO：27所示，和所述B-LCDR1如SEQ ID NO：41所示，所述B-LCDR2如SEQ ID NO：32所示，和所述B-LCDR3如SEQ ID NO：42所示；或(vii)所述B-HCDR1如SEQ ID NO：18所示，所述B-HCDR2如SEQ ID NO：30所示，所述B-HCDR3如SEQ ID NO：20所示，和所述B-LCDR1如SEQ ID NO：31所示，所述B-LCDR2如SEQ ID NO：32所示，和所述B-LCDR3如SEQ ID NO：33。
在一些实施方案中，如上任一项所述的抗原结合分子，其包含B-VH和B-VL，所述B-VH如SEQ ID NO：47所示，和所述B-VL如SEQ ID NO：48所示。在一些实施方案中，所述B-VH如SEQ ID NO：5所示，和所述B-VL如SEQ ID NO：12所示；或所述B-VH如SEQ ID NO：6所示，和所述B-VL如SEQ ID NO：13所示；或所述B-VH如SEQ ID NO：7所示，和所述B-VL如SEQ ID NO：14所示；或所述B-VH如SEQ ID NO：8所示，和所述B-VL如SEQ ID NO：15所示； 或所述B-VH如SEQ ID NO：9所示，和所述B-VL如SEQ ID NO：16所示；或所述B-VH如SEQ ID NO：10所示，和所述B-VL如SEQ ID NO：17所示；或所述B-VH如SEQ ID NO：11所示，和所述B-VL如SEQ ID NO：2所示。
在一些实施方案中，特异性结合BAFF和IL-12/23的抗原结合分子，其包含特异性结合BAFF的抗原结合模块1和特异性结合IL-12和/或IL-23的抗原结合模块2；其中所述抗原结合模块2包含重链可变区P-VH和轻链可变区P-VL，所述P-VH包含P-HCDR1、P-HCDR2和P-HCDR3，所述P-VL包含P-LCDR1、P-LCDR2和P-LCDR3；所述P-HCDR1、P-HCDR2和P-HCDR3的氨基酸序列分别与SEQ ID NO：51中的Pv-HCDR1、Pv-HCDR2和Pv-HCDR3的氨基酸序列相同，和所述P-LCDR1、P-LCDR2和P-LCDR3的氨基酸序列分别与SEQ ID NO：52中的Pv-LCDR1、Pv-LCDR2和Pv-LCDR3的氨基酸序列相同；所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3、Pv-HCDR1、Pv-HCDR2、Pv-HCDR3、Pv-LCDR1、Pv-LCDR2和Pv-LCDR3是根据选自Kabat、IMGT、Chothia、AbM和Contact的相同的编号规则定义的。在一些实施方案中，如上任一项所述的抗原结合分子，所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2、P-LCDR3是根据Kabat编号规则定义的，其中，所述P-HCDR1如SEQ ID NO：53所示，所述P-HCDR2如SEQ ID NO：54所示，所述P-HCDR3如SEQ ID NO：55所示，和所述P-LCDR1如SEQ ID NO：56所示，所述P-LCDR2如SEQ ID NO：57所示，所述P-LCDR3如SEQ ID NO：58所示。在一些实施方案中，所述重链可变区P-VH如SEQ ID NO：51所示，所述轻链可变区P-VL如SEQ ID NO：52所示。
抗原结合分子的变体
在某些实施方案中，涵盖本文中提供的抗原结合分子的氨基酸序列变体。例如，可以期望改善抗体的结合亲和力和/或其它生物学特性。可以通过将合适的修饰引入编码抗体的核苷酸序列中，或者通过肽合成来制备抗体的氨基酸序列变体。此类修饰包括例如对抗原结合分子的氨基酸序列内的残基的删除、和/或插入、和/或取代。可以进行删除、插入、和取代的任何组合以得到最终的构建体，只要最终的构建体拥有期望的特征，例如抗原结合特性。
包含取代、插入、和删除的变体
在某些实施方案中，提供了具有一处或多处氨基酸取代的抗原结合分子变体。取代诱变感兴趣的位点包括CDR和FR。保守取代在表2中在“优选的取代”的标题下显示。更实质的变化在表2中在“示例性取代”的标题下提供，并且如下文参照氨基酸侧链类别进一步描述的。可以将氨基酸取代引入感兴趣的抗体中，并且对产物筛选期望的活性，例如保留/改善的抗原结合，降低的免疫原性，或改善的ADCC或CDC。
表2.氨基酸的取代
原始残基 示例性取代 优选的取代
Ala(A) Val；Leu；Ile Val
Arg(R) Lys；Gln；Asn Lys
Asn(N) Gln；His；Asp,Lys；Arg Gln
Asp(D) Glu；Asn Glu
Cys(C) Ser；Ala Ser
Gln(Q) Asn；Glu Asn
Glu(E) Asp；Gln Asp
Gly(G) Ala Ala
His(H) Asn；Gln；Lys；Arg Arg
Ile(I) Leu；Val；Met；Ala；Phe；正亮氨酸 Leu
Leu(L) 正亮氨酸；Ile；Val；Met；Ala；Phe Ile
Lys(K) Arg；Gln；Asn Arg
Met(M) Leu；Phe；Ile Leu
Phe(F) Trp；Leu；Val；Ile；Ala；Tyr Tyr
Pro(P) Ala Ala
Ser(S) Thr Thr
Thr(T) Ser Ser
Trp(W) Tyr；Phe Tyr
Tyr(Y) Trp；Phe；Thr；Ser Phe
Val(V) Ile；Leu；Met；Phe；Ala；正亮氨酸 Leu
依照常见的侧链特性，氨基酸可以如下分组:
(1)疏水性的：Nle，Met，Ala，Val，Leu，Ile；
(2)中性，亲水性的：Cys，Ser，Thr，Asn，Gln；
(3)酸性的：Asp，Glu；
(4)碱性的：His，Lys，Arg；
(5)影响链取向(orientation)的残基：Gly，Pro；
(6)芳香族的：Trp，Tyr，Phe。
非保守取代是指，用一个类别的成员替换另一个类别的成员。
一类取代变体涉及取代亲本抗体(例如人源化或人抗体)的一个或多个CDR残基。一般地，经选择用于进一步研究的所得变体相对于亲本抗体会具有某些生物学特性(例如升高的亲和力，降低的免疫原性)的改变(例如改善)，和/或会基本上保留亲本抗体的某些生物学特性。一种示例性的取代变体是亲和力成熟的抗体，可以例如使用基于噬菌体展示的亲和力成熟技术(如本文所述的那些技术)，便利地产生所述抗体。简言之，将一个或多个CDR残基突变，并将变体抗体在噬菌体上展示，并对其筛选特定的生物学活性(例如结合亲和力)。可以对CDR做 出改变(例如取代)，例如以改善抗体亲和力。可以对CDR“热点”(即在体细胞成熟过程期间以高频率经历突变的密码子所编码的残基，和/或接触抗原的残基)做出此类改变，同时对所得的变体VH或VL测试结合亲和力。在亲和力成熟的一些实施方案中，通过多种方法(例如易错PCR、链改组、或寡核苷酸指导的诱变)的任一种，将多样性引入所选择用于成熟的可变基因中。然后，创建次级文库。然后，筛选文库以鉴定具有期望的亲和力的任何抗体变体。另一种引入多样性的方法涉及CDR定向的方法，其中将几个CDR残基(例如一次4-6个残基)随机化。可以例如使用丙氨酸扫描诱变或建模来特异性鉴定涉及抗原结合的CDR残基。特别地，HCDR3和LCDR3经常被作为靶点。
在某些实施方案中，取代、插入或缺失可以在一个或多个CDR内发生，只要此类变化不实质性降低抗体结合抗原的能力。例如，可以对CDR做出保守变化(例如保守取代，如本文中提供的)，其不实质性降低结合亲和力。此类变化不发生在抗原接触残基。
一种可用于鉴定抗体中可以作为诱变靶位的残基或区域的方法称作“丙氨酸扫描诱变”。在这种方法中，鉴定一个残基或残基组(例如带电荷的残基，诸如Arg、Asp、His、Lys和Glu)，并且用中性或带负电荷的氨基酸(例如，Ala或聚丙氨酸)替换以确定该抗体与抗原的相互作用是否受影响。可以在对初始取代显示功能敏感性的氨基酸位置引入进一步的取代。此外，可通过研究抗原-抗体复合物的晶体结构来鉴定抗体与抗原间的接触点。这些接触残基及邻近残基可以作为取代候选物被打靶或消除。可以筛选变体以确定它们是否含有期望的特性。
氨基酸序列插入包括：在氨基和/或羧基端融合1个残基或长度为100或更多个残基的多肽，和单个或多个氨基酸残基的序列内插入。在末端插入的例子包括具有N端甲硫氨酰基残基的抗体。抗体分子的其它插入变体包括，在抗体的N或C端融合有酶(或延长抗体的血清半衰期的多肽)的融合物。
Fc区的改造
在一个方面，本披露的抗原结合分子的Fc区包含一个或多个氨基酸取代，所述一个或多个氨基酸取代减少其与Fc受体的结合，例如其与Fcγ受体的结合，并且降低或消除效应子功能。天然IgG Fc区，具体地是IgG 1Fc区或IgG 4Fc区，可能导致本披露的抗原结合分子靶向表达Fc受体的细胞，而不是表达抗原的细胞。本披露改造的Fc区表现出降低的对Fc受体的结合亲和力和/或降低的效应子功能。在一些实施方案中，改造的Fc区与天然Fc区相比，对Fc受体的结合亲和力下降50％、80％、90％或95％以上。在一些实施方案中，所述的Fc受体是Fcγ受体。在一些实施方案中，所述Fc受体是人Fcγ受体，例如FcγRI、FcγRIIa、FcγRIIB、FcγRIIIa。在一些实施方案中，改造的Fc区与天然Fc区相比，对补体(如C1q)的结合亲和力降低。在一些实施方案中，改造的Fc区与天然Fc区相比，对新生儿Fc受体(FcRn)的结合亲和力不降低。在一些实施例中，改造的Fc区具有降 低的效应子功能，所述降低的效应子功能可以包括但不限于以下中的一个或多个：降低的补体依赖性细胞毒性(CDC)、降低的抗体依赖性细胞介导的细胞毒性(ADCC)、降低的抗体依赖性细胞吞噬(ADCP)、减少的细胞因子分泌、减少的免疫复合物介导的抗原呈递细胞的抗原摄取、减少的与NK细胞的结合、减少的与巨噬细胞的结合、减少的与单核细胞的结合、减少的与多形核细胞的结合、减少的直接信号传导诱导性细胞凋亡、降低的树突细胞成熟或减少的T细胞引发。
对于IgG 1Fc区，在238、265、269、270、297、327和329等位置的氨基酸残基取代可降低的效应子功能。在一些实施方案中，所述Fc区是人IgG 1Fc区，并且在234和235位置的氨基酸残基为A，编号依据为EU索引。对于IgG 4Fc区，在228等位置的氨基酸残基取代可降低的效应子功能。
抗原结合分子还可包含二硫键改造，例如第一亚基的354C和第二亚基的349C。
抗原结合分子包含与Fc区的两个亚基融合的结合模块时，可能导致不期望的同源二聚化。为了提高产率和纯度，因此在本披露的抗原结合分子的Fc区中引入促进异源二聚化的修饰将是有利的。在一些实施方式中，本披露的Fc区包含根据杵臼(knob-into-hole，KIH)技术的改造，该方法涉及在一个亚基的界面处引入凸起结构(knob)以及在另一个亚基的界面处引入孔结构(hole)。使得所述凸起结构可以定位在孔结构中，促进异源二聚体的形成并抑制同源二聚体的产生。凸起结构是通过用较大侧链(例如酪氨酸或色氨酸)取代来自一个亚基的界面的小氨基酸侧链而构建的。而孔结构是通过用较小的氨基酸侧链(例如丙氨酸或苏氨酸)取代大氨基酸侧链而在另一个亚基的界面中创建的。凸起结构和孔结构通过改变编码多肽的核酸来制备，可选的氨基酸取代如下表3所示：
表3.KIH突变组合
Figure PCTCN2022102621-appb-000002
除了杵臼技术外，用于修饰重链的CH3结构域以实现异源二聚化的其他技术也是本领域中已知的，例如WO96/27011、WO98/050431、EP1870459、WO2007/110205、WO 007/147901、WO2009/089004、WO2010/129304、WO2011/90754、WO2011/143545、WO2012/058768、WO2013/157954和WO013/096291。
Fc区的C末端可以是以氨基酸残基PGK结束的完整C末端；也可以是截短的C末端，例如在所述截短的C末端中已经去除了一个或两个C末端氨基酸残基。在一个优选的方面中，Fc区的C末端是以PG结束的缩短的C末端。因此，在一些实施方式中，完整抗体的组合物可以包括去除了所有K447残基和/或G446+K447 残基的抗体群体。在一些实施方式中，完整抗体的组合物可以包括没有去除K447残基和/或G446+K447残基的抗体群体。在一些实施方式中，完整抗体的组合物具有带有和不带有K447残基和/或G446+K447残基的抗体混合物的抗体群体。
重组方法
抗原结合分子(例如抗体)可以使用重组方法来产生。对于这些方法，提供编码抗原结合分子的一个或更多个分离的核酸。
在一个实施方案中，本披露提供了编码如前所述的抗原结合分子的分离的核酸。此类核酸可以独立编码前述的任一多肽链。在另一方面中，本披露提供了包含此类核酸的一种或多种载体(例如表达载体)。在另一方面中，本披露提供了包含此类核酸的宿主细胞。在一个实施方案中，提供制备抗原结合分子的方法，其中所述方法包括，在适合表达的条件下，培养包含编码所述抗原结合分子的核酸的宿主细胞，如上文所提供的，和任选地从宿主细胞(或宿主细胞培养基)回收所述抗原结合分子。
为了重组产生抗原结合分子，将编码蛋白的核酸分离并插入一个或更多个载体中，用于在宿主细胞中进一步克隆和/或表达。此类核酸可以使用常规程序容易地分离和测序，或者通过重组方法产生或通过化学合成获得。
用于克隆或表达编码抗原结合分子的载体的适当宿主细胞包括本文描述的原核或真核细胞。例如，可在细菌中产生，特别是当不需要糖基化和Fc效应子功能时。在表达后，可以在可溶级分中从细菌细胞糊状物分离，并且可进一步纯化。
除了原核生物以外，真核微生物诸如丝状真菌或酵母也是用于编码抗原结合分子的载体的合适的克隆或表达宿主，包括真菌和酵母菌株。适于表达抗原结合分子的合适的宿主细胞也可源自多细胞生物体(无脊椎动物和脊椎动物)；无脊椎动物细胞的例子包括植物和昆虫细胞。已经鉴定了许多杆状病毒株，其可与昆虫细胞联合使用，特别是用于草地贪夜蛾(Spodoptera frugiperda)细胞的转染；还可利用植物细胞培养物作为宿主，例如US5959177、US 6040498、US6420548、US 7125978和US6417429；也可将脊椎动物细胞用作宿主，例如适应于在悬浮液中生长的哺乳动物细胞系。适宜的哺乳动物宿主细胞系的其它例子是经SV40转化的猴肾CVl系(COS-7)；人胚肾系(293或293T细胞)；幼仓鼠肾细胞(BHK)；小鼠塞托利(sertoli)细胞(TM4细胞)；猴肾细胞(CV1)；非洲绿猴肾细胞(VERO-76)；人宫颈癌细胞(HELA)；犬肾细胞(MDCK)；水牛鼠(buffalo rat)肝细胞(BRL3A)；人肺细胞(W138)；人肝细胞(Hep G2)；小鼠乳房肿瘤(MMT 060562)；TRI细胞；MRC 5细胞；和FS4细胞。其它适宜的哺乳动物宿主细胞系包括中国仓鼠卵巢(CHO)细胞，包括DHFR-CHO细胞；以及骨髓瘤细胞系，如Y0、NS0和Sp2/0。关于适合产生抗体的某些哺乳动物宿主细胞系的综述参见例如Yazaki，P.和Wu，A.M.，Methods in Molecular Biology，Vol.248，Lo，B.K.C.(编)，Humana Press，Totowa，NJ(2004)，第255-268页。
检测与诊断
本文提供的抗原结合分子可以通过本领域已知的多种测定法对其物理/化学特征和/或生物学活性进行鉴定、筛选或表征。在一个方面中，例如通过已知方法如ELISA、蛋白印迹法等，测试本披露的抗原结合分子活性。
在某些实施方案中，本披露提供的抗原结合分子可用于检测生物学样品中BAFF或IL-12/23的存在或水平。在用于本文时，术语“检测”涵盖定量或定性检测。在某些实施方案中，生物学样品包含细胞或组织，诸如肿瘤组织。
在一个实施方案中，提供了在诊断或检测方法中使用的抗原结合分子。一方面，提供了检测生物学样品中BAFF或IL-12/23的存在的方法。在某些实施方案中，该方法包括在适宜条件下使生物学样品与抗原结合分子接触，并检测是否在检测试剂与抗原之间形成复合物。此类方法可以是体外或体内方法。在一个实施方案中，使用抗原结合分子来选择适合治疗的受试者，例如BAFF或IL-12/23是用于选择患者的生物标志物。
可使用本披露的抗原结合分子来诊断的示例性病症，例如身免疫性疾病，例如：系统性红斑狼疮、重症肌无力、多发性硬化、胰岛素依赖性糖尿病、克罗恩氏病、类风湿关节炎、多关节型青少年类风湿关节炎或银屑病性关节炎；或者B细胞障碍，例如：肿瘤、慢性白细胞性白血病、多发性骨髓瘤、非霍奇金淋巴瘤、移植后淋巴组织增生病或轻链丙球蛋白病。
在某些实施方案中，提供了经标记的抗原结合分子。标记物包括但不限于直接检测的标记物或模块(诸如荧光、发色、电子致密、化学发光、和放射性标记物)，和间接检测的模块(例如，经由酶反应或分子相互作用间接检测的模块，诸如酶或配体)。
治疗方法与施用途径
本文提供的任何抗原结合分子(例如抗体)可用于治疗方法。在又一个方面，本披露提供抗原结合分子在药物的制造或制备中的用途。在一些实施方案中，所述B细胞障碍或自身免疫性疾病是与BAFF或IL-12/23相关的疾病或病症。在一些实施方案中，所述自身免疫性疾病选自：系统性红斑狼疮、重症肌无力、多发性硬化、胰岛素依赖性糖尿病、克罗恩氏病、类风湿关节炎、多关节型青少年类风湿关节炎和银屑病性关节炎；所述B细胞障碍选自：肿瘤、慢性白细胞性白血病、多发性骨髓瘤、非霍奇金淋巴瘤、移植后淋巴组织增生病以及轻链丙球蛋白病。在一些实施方案中，所述自身免疫性疾病为系统性红斑狼疮。在一个此类实施方案中，所述用途进一步包括向受试者施用有效量的至少一种另外的治疗剂(例如一种、两种、三种、四种、五种或六种另外的治疗剂)。
根据任意以上实施方案的“受试者”可以是人。当用于治疗目的时，受试者是已患有、疑似患有目标疾病的个体。当用于预防目的时，受试者是易感于目标疾病的个体。
在又一个的方面，提供包含所述抗原结合分子的药物组合物，例如，其用于以上任何制药用途或治疗方法。在一个实施方案中，药物组合物包含本文提供的任何抗原结合分子和药学上可接受的载体。在另一个实施方案中，药物组合物还包含至少一种另外的治疗剂。
本披露的抗原结合分子可单独使用或与其他试剂联合用于治疗。例如，本披露的抗体可与至少一种另外的治疗剂联合施用(同时、或先后施用)。
本披露的抗原结合分子(和任何另外的治疗剂)可通过任何合适的手段施用，包括肠胃外、肺内和鼻内，并且如果需要局部治疗，则病灶内施用。肠胃外输注包括肌肉内、静脉内、动脉内、腹膜内或皮下施用。给药可以通过任何适当的途径，例如，通过注射，诸如静脉内或皮下注射，这部分取决于施用是短期的还是长期的。本文考虑多种给药时间方案，包括但不限于，单次或在多个时间点多次施用，推注施用和脉冲输注。
本披露的抗原结合分子将以符合良好医疗实践(如GOOD MEDICAL PRACTICE Guideline，GMP)的方式配制、给药和施用。在此背景下考虑的因素包括所治疗的具体病症、所治疗的具体哺乳动物、个体患者的临床状况、病症的起因、试剂的递送部位、施用方法、施用时间安排以及医学从业者已知的其他因素。抗原结合分子可以与或不与目前用于预防或治疗所述病症的一种或更多种试剂一起配制。此类其它试剂的有效量取决于药物组合物中存在的量、病症或治疗的类型以及其它因素。这些通常以与本文所述相同的剂量和施用路径使用，或以本文所述剂量的约1至99％使用，或以其它剂量使用，并通过经验/临床确定为合适的任何途径使用。
为了预防或治疗疾病，本披露的抗原结合分子(当单独使用或与一种或更多种其他另外的治疗剂组合使用时)的适当的剂量将取决于待治疗的疾病的类型，治疗分子的类型，疾病的严重性和病程，是为预防还是治疗目的施用，之前的治疗，受试者的临床病史和对治疗分子的响应，和主治医师的判断。治疗分子恰当地以一次或经过一系列治疗施用于受试者。例如，每日剂量可能在约1μg/kg至100mg/kg，具体取决于上文提及的因素。
制品
在本披露的另一方面中，提供一种制品，所述制品包含可用于治疗、预防和/或诊断上述病症的材料。该制品包含容器和在容器上或与容器联合的标签或包装插页(package insert)。合适的容器包括，例如，瓶子、管形瓶、注射器、IV溶液袋等。容器可以自各种材料诸如玻璃或塑料形成。容器装有单独或与另一种组合物组合有效治疗，预防和/或诊断疾患的组合物，并且可具有无菌的存取口(例如，容器可以是具有由皮下注射针可刺穿的塞子的静脉内溶液袋或管形瓶)。组合物中的至少一种活性试剂是本披露的抗原结合分子。标签或包装插页指示使用该组合物是来治疗选择的病况。此外，制品可以包含：(a)其中装有组合物的第 一容器，其中所述组合物包含本披露的抗原结合分子；和(b)其中装有组合物的第二容器，其中所述组合物包含另外的细胞毒性剂或其他方面的治疗剂。本披露的该实施方案中的制品可进一步包含包装插页，所述包装插页指示所述组合物可以用于治疗特定病况。备选地，或另外地，制品可进一步包含第二(或第三)容器，所述第二(或第三)容器包含药学上可接受的缓冲液。从商业和用户立场，它可进一步包括所需的其他材料，包括其他缓冲剂、稀释剂、滤器、针头和注射器。作为一个示例，制品制备成药盒(kit)的形式。
实施例与测试例
以下结合实施例和测试例进一步描述本披露，但这些实施例和测试例并非限制着本披露的范围。本披露实施例和测试例中未注明具体条件的实验方法，通常按照常规条件，如冷泉港的抗体技术实验手册，分子克隆手册；或按照原料或商品制造厂商所建议的条件。未注明具体来源的试剂，为市场购买的常规试剂。
实施例1：Belimumab突变抗体及其活性检测
采用噬菌体展示技术对Belimumab进行改造。通过设计突变引物，对Belimumab轻重链的CDR分别进行突变，构建成突变文库。利用生物素化的BAFF蛋白(Sino biological，10056-HNCH)对突变文库进行富集和筛选，获得候选克隆，通过测序确定克隆轻重链可变区氨基酸序列，将候选克隆轻/重链可变区分别与抗体轻/重链恒定区融合，进而构建全长抗体。其中，
Belimumab抗体的序列信息如下：
>B-VH0(Belimumab重链可变区)的氨基酸序列：
Figure PCTCN2022102621-appb-000003
>B-VL0(Belimumab轻链可变区)的氨基酸序列：
Figure PCTCN2022102621-appb-000004
>Belimumab重链的氨基酸序列：
Figure PCTCN2022102621-appb-000005
Figure PCTCN2022102621-appb-000006
>Belimumab轻链的氨基酸序列：
Figure PCTCN2022102621-appb-000007
Belimumab改造后重链可变区和轻链可变区：
>B-VH1的氨基酸序列：
Figure PCTCN2022102621-appb-000008
>B-VH2的氨基酸序列
Figure PCTCN2022102621-appb-000009
>B-VH3的氨基酸序列：
Figure PCTCN2022102621-appb-000010
>B-VH4的氨基酸序列：
Figure PCTCN2022102621-appb-000011
>B-VH5的氨基酸序列：
Figure PCTCN2022102621-appb-000012
>B-VH6的氨基酸序列：
Figure PCTCN2022102621-appb-000013
>B-VH7的氨基酸序列：
Figure PCTCN2022102621-appb-000014
>B-VL1的氨基酸序列：
Figure PCTCN2022102621-appb-000015
>B-VL2的氨基酸序列：
Figure PCTCN2022102621-appb-000016
>B-VL3的氨基酸序列：
Figure PCTCN2022102621-appb-000017
>B-VL4的氨基酸序列：
Figure PCTCN2022102621-appb-000018
>B-VL5的氨基酸序列：
Figure PCTCN2022102621-appb-000019
>B-VL6的氨基酸序列：
Figure PCTCN2022102621-appb-000020
备注：上述可变区序列中，依次为FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4，下划线部分为CDR区(依照Kabat编号系统确定)，斜体部分为突变的氨基酸残基，无下划线部分为框架区。
表4.重链CDR序列表
Figure PCTCN2022102621-appb-000021
Figure PCTCN2022102621-appb-000022
备注：表中CDR为依照Kabat编号系统确定的CDR。
表5.轻链CDR序列表
Figure PCTCN2022102621-appb-000023
Figure PCTCN2022102621-appb-000024
备注：表中CDR为依照Kabat编号系统确定的CDR
将上述重链可变区和轻链可变区分别与抗体重链恒定区和轻链恒定区融合，构建全长抗体。示例性地，将上述重链可变区与Belimumab的重链恒定区(序列如SEQ ID NO：45所示)融合，抗体轻链可变区与Belimumab的轻链恒定区(序列如SEQ ID NO：46所示)融合，构建抗BAFF抗体：B1至B7，获得的抗体的可变区序列见表6。
>Belimumab抗体重链恒定区的氨基酸序列
Figure PCTCN2022102621-appb-000025
>Belimumab抗体轻链恒定区的氨基酸序列
Figure PCTCN2022102621-appb-000026
表6.抗BAFF抗体的可变区序列
抗体编号 重链可变区 轻链可变区
B1 B-VH1(SEQ ID NO：5) B-VL1(SEQ ID NO：12)
B2 B-VH2(SEQ ID NO：6) B-VL2(SEQ ID NO：13)
B3 B-VH3(SEQ ID NO：7) B-VL3(SEQ ID NO：14)
B4 B-VH4(SEQ ID NO：8) B-VL4(SEQ ID NO：15)
B5 B-VH5(SEQ ID NO：9) B-VL5(SEQ ID NO：16)
B6 B-VH6(SEQ ID NO：10) B-VL6(SEQ ID NO：17)
B7 B-VH7(SEQ ID NO：11) B-VL0(SEQ ID NO：2)
备注：表中，例如B6抗体，其重链可变区为B-VH6(SEQ ID NO：10)，轻链可变区为B-VL6(SEQ ID NO：17)，重链恒定区为SEQ ID NO：45，轻链恒定区为SEQ ID NO：46。
通过体外活性实验，检测构建的B1至B7抗体阻断人BAFF与BAFF-R结合的活性(参见本披露的测试例2)、抑制BAFF诱导的B细胞增殖的活性(参见本披露的测试例3)以及与人BAFF结合的亲和力(参见本披露的测试例5)。实验结果显示，本披露的B1至B7抗体能以高亲和力与人BAFF结合，与母抗体Belimumab相比，其阻断BAFF与BAFF-R结合能力以及抑制BAFF诱导的B细 胞增殖的活性更好。
另外，对上述抗BAFF抗体的框架区进行改造。示范性地，将B6的重链可变区第44位(依照Kabat编号系统确定的位点)氨基酸残基突变为C(即44C)，同时将B6的轻链可变区第100位(依照Kabat编号系统确定的位点)氨基酸残基突变为C(即100C)，以便增加VH与VL间的链间二硫键，B6框架区改造后的抗体B61的可变区序列如下：
>B61的重链可变区的氨基酸序列
Figure PCTCN2022102621-appb-000027
>B61的轻链可变区的氨基酸序列
Figure PCTCN2022102621-appb-000028
实施例2：特异性结合BAFF和IL-12/23的双特异性抗体的构建
利用实施例1获得的抗BAFF抗体以及已知的抗IL-12/23抗体构建特异性结合BAFF和IL-12/23的双特异性抗体。IL-12/23抗体可以是来源于任意适宜的抗体，示例性地，例如抗IL-12/23 p40亚基抗体Ustekinumab(简称Umab)，Ustekinumab的序列如下：
>Ustekinumab重链的氨基酸序列
Figure PCTCN2022102621-appb-000029
>Ustekinumab轻链的氨基酸序列
Figure PCTCN2022102621-appb-000030
Figure PCTCN2022102621-appb-000031
>Ustekinumab重链可变区的氨基酸序列
Figure PCTCN2022102621-appb-000032
>Ustekinumab轻链可变区的氨基酸序列
Figure PCTCN2022102621-appb-000033
备注：上述序列中，其中双下划线部分为可变区序列，单下划线部分为CDR区序列，所述可变区和CDR根据Kabat编号系统确认。
Ustekinumab的CDR序列见下表7：
表7.Ustekinumab的CDR序列
Figure PCTCN2022102621-appb-000034
备注：上述CDR根据Kabat编号系统确认。
示例性地，以下利用人IgG1重链恒定区(序列如SEQ ID NO：59所示)、人κ轻链恒定区(序列如SEQ ID NO：60所示)、B61抗体重链可变区和轻链可变区(序列如SEQ ID NO：47、48所示)和Ustekinumab抗体重链可变区和轻链可变区(序列如SEQ ID NO：51、52所示)，构建特异性结合BAFF和IL-12/23的双特异性抗体。其中，Ustekinumab抗体轻链可变区的C端与人κ轻链恒定区的N端融合形成第二链；Ustekinumab抗体重链可变区的C端与人IgG1重链恒定区的N端融合，人IgG1重链恒定区的C端与scFv(由B61抗体的重链可变区和轻链可变区构建而成)的N端融合，从而形成第一链。最终构建特异性结合BAFF和IL-12/23的双特异性抗体：BU-1和BU-2。BU-1和BU-2均由4条链组成(包含两条相同的第一链和两条相同的第二链)，BU-1的结构示意图如图1，BU-2的结构示意图如图2。
>人IgG1重链恒定区的氨基酸序列
Figure PCTCN2022102621-appb-000035
Figure PCTCN2022102621-appb-000036
>人κ轻链恒定区的氨基酸序列
Figure PCTCN2022102621-appb-000037
>BU-1的第一链的氨基酸序列
Figure PCTCN2022102621-appb-000038
>BU-1的第二链的氨基酸序列与Ustekinumab轻链相同，序列如SEQ ID NO：50所示。
>BU-2的第一链的氨基酸序列
Figure PCTCN2022102621-appb-000039
Figure PCTCN2022102621-appb-000040
>BU-2的第二链的氨基酸序列与Ustekinumab轻链相同，序列如SEQ ID NO：50所示。
备注：上述BU-1和BU-2的第一链中，其中波浪线部分为Ustekinumab抗体的重链可变区部分，点下划线部分为人IgG1重链恒定区部分，双下划线部分为B61抗体的重链可变区部分，单下划线部分为B61抗体的轻链可变区部分，斜体部分为连接子序列部分。
测试例1：Elisa结合实验
以下通过Elisa的方法检测抗体对BAFF和IL-12/23 p40的结合活性。具体方法如下：
用pH7.4的PBS(源培生物，B320)缓冲液将待测样品稀释至2μg/mL，以100μL/孔的体积加入96孔酶标板(Corning，3590)中，4℃过夜孵育。弃去液体后，每孔加入300μL用PBS稀释的5％脱脂牛奶(BD，232100)进行封闭，37℃孵育2小时。封闭结束后，弃去封闭液，并用PBST缓冲液(pH7.4PBS含0.1％tween-20)洗板3次后，每孔加入100μL梯度稀释的BAFF(ACROBiosystems，BAF-H5248)或IL-12/23 p40(Sino Biological，10052-H08H)溶液，于37℃孵育1小时。孵育结束后用PBST洗板3次。每孔加入100μL Anti-His-HRP(Sino biological，105327-MM02T-H，1:2000稀释)。用PBST洗板3次后，每孔加入100μL TMB显色底物(KPL，5120-0077)，室温孵育10至15分钟，每孔加入50μL 1M H 2SO 4终止反应，用酶标仪读取在450nm处的吸收值，用软件拟合出抗体与抗原的结合曲线，计算出EC50值。实验结果见下表8和表9，实验结果表明，本披露构建的特异性结合BAFF和IL-12/23双特异性抗体均能有效结合BAFF和IL-12/23 p40。
表8.抗体与IL-12/23 p40结合实验结果
样品 与IL-12/23 p40结合的EC50(nM)
BU-1 2.537
BU-2 3.432
表9.抗体与BAFF结合实验结果
样品 与BAFF结合的EC50(nM)
BU-1 3.709
BU-2 6.638
测试例2：配体和受体的阻断实验
通过Elisa方法检测抗体对以下受体与配体的阻断活性：人BAFF与人 BAFF-R、人BAFF与人BCMA、人BAFF与人TACI、以及人IL-12/23 p40与人IL-12Rβ1。具体方法如下：
用pH7.4的PBS(源培生物，B320)缓冲液将受体蛋白稀释至2μg/mL，以100μL/孔的体积加入96孔酶标板(Corning，3590)中，4℃过夜孵育。弃去液体后，每孔加入200μL 1％Casein封闭液(Thermo，37528)进行封闭，37℃孵育2小时。封闭结束后，弃去封闭液，并用PBST缓冲液(pH7.4 PBS含0.1％tween-20)洗板3次后备用。将固定浓度的生物素(Biotin)标记的配体蛋白与梯度稀释的抗体混合后37℃预孵育30分钟后加入封闭好的酶标板中，37℃孵育1.5小时。孵育结束后用PBST洗板3次，每孔加入100μL链和亲霉素-HRP(Invitrogen，434323，1:4000稀释)，37℃孵育1小时。去上清，用PBST洗板3次后每孔加入100μL TMB显色底物(KPL，5120-0077)，室温孵育10至15分钟，每孔加入50μL 1M H 2SO 4终止反应，用酶标仪读取在450nm处的吸收值，用软件拟合出抑制配体和受体结合的曲线，计算出IC50值。本测试例中所用受体和配体蛋白的来源信息如下：人BAFF(Sino biological，10056-HNCH)，人IL-12/23 p40(Sino biological，10052-H08H)，人BAFF-R(Sino biological，16079-H02H)，人BCMA(Sino biological，10620-H02H)，人TACI(ACROBiosystems，TAI-H5256)，人IL-12Rβ1(ACROBiosystems，ILB-H5255)。
实验结果见表10、表11、表12，实验结果表明，本披露构建的特异性结合BAFF抗体能有效阻断人BAFF与人BAFF-R结合，且IC50值小于Belimumab。另外，本披露构建的特异性结合BAFF和IL-12/23双特异性抗体既能有效阻断人BAFF与人BAFF-R、人BAFF与人BCMA、人BAFF与人TACI，且IC50值小于Belimumab；也能有效阻断人IL-12/23 p40与人IL-12Rβ1的结合。
表10.抗体阻断人BAFF与人BAFF-R结合实验结果
抗体编号 阻断人BAFF与人BAFF-R结合的IC50(nM)
Belimumab 19.03
B1 10.93
B2 8.647
B3 9.424
B4 6.556
B5 10.38
B6 7.694
B7 10.66
表11.抗体阻断人BAFF与其受体、阻断IL-12/23 p40与其受体结合实验结果
Figure PCTCN2022102621-appb-000041
Figure PCTCN2022102621-appb-000042
备注：“-”表示未检测
表12.双特异性抗体阻断人BAFF与人BAFF-R结合实验结果
样品 阻断人BAFF与人BAFF-R结合的IC50(nM)
BU-1 4.44
BU-2 6.208
测试例3：B细胞增殖实验
通过B细胞增殖实验检测抗体是否能抑制BAFF诱导的B细胞增殖。实验方法如下：
取小鼠脾脏进行研磨，4℃离心5分钟收集下层细胞，用洗涤溶液(PBS+2％FBS+2mM EDTA)清洗一次并离心，去上清后加入RBC裂解缓冲液(Invitrogen，00-4333-57)，室温静置5分钟至红细胞完全裂解。再次离心并重悬细胞进行计数。细胞悬液用B细胞分离试剂盒(Miltenyi Biotec，130-090-862)进行分选，将分离的B细胞用RPMI 1640培养基(Gibco，11875119)+10％FBS(Gibco，10099-141)+50μM 2-巯基乙醇(Sigma-Aldrich，M6250)重悬并计数，细胞铺在96孔细胞板(Costar，3903)中备用。将BAFF(R&D Systems，7537-BF)蛋白稀释至固定浓度，并加入梯度稀释的抗体混匀，37℃预孵育30分钟后加入96孔细胞板中，37℃细胞培养箱中培养48小时。取出细胞培养板，每孔加入50μL Celltiter Glo检测液(Promega，G7573)，室温孵育10分钟，用酶标仪检测生物发光信号，将检测结果用软件拟合出抑制曲线，计算出IC50值。
实验结果见下表13、表14，实验结果表明，本披露的特异性结合BAFF抗体以及特异性结合BAFF和IL-12/23双特异性抗体能有效抑制BAFF诱导的B细胞增殖，且Belimumab抑制BAFF诱导B细胞增殖的IC50值是本披露的特异性结合BAFF抗体的2倍以上，而ustekinumab无抑制BAFF诱导B细胞增殖功能。
表13.抗BAFF抗体抑制BAFF诱导的B细胞增殖实验结果
样品 抑制BAFF诱导的B细胞增殖的IC50(nM)
Belimumab 0.249
B1 0.1162
B2 0.09694
B3 0.08038
B4 0.06931
B5 0.05736
B6 0.05031
B7 0.106
表14.双特异性抗体抑制BAFF诱导B细胞增殖实验结果
样品 抑制BAFF诱导的B细胞增殖的IC50(nM)
BU-1 0.05704
BU-2 0.1396
Ustekinumab 无活性
测试例4：IFNγ和IL-17分泌实验检测
通过IFNγ和IL-17分泌实验检测抗体是否能抑制IL-12和IL-23诱导的T细胞分化。实验方法如下：
在96孔板(Corning，3599)中每孔加入100μL 2μg/mL抗鼠CD3抗体(BioLegend，100238)和2μg/mL抗鼠CD28抗体(BioLegend，102116)，37℃孵育1小时，用PBS洗涤2次后备用。取小鼠脾脏进行研磨，4℃离心5分钟收集下层细胞，用洗涤溶液(PBS+2％FBS+2mM EDTA)清洗一次并离心，去上清后加入RBC裂解缓冲液(Invitrogen，00-4333-57)，室温静置5分钟至红细胞完全裂解。再次离心并重悬细胞进行计数。细胞悬液用鼠CD4细胞试剂盒(Invitrogen，11415D)进行分选，将分离的CD4+T细胞用培养基RPMI 1640培养基(Gibco，11875119)+10％FBS(Gibco，10099-141)重悬并计数备用。
检测IL-12诱导的T细胞分化时，在T细胞中加入20μg/mL抗鼠IL-4(BioLegend，504122)后将细胞悬液铺在包被好的96孔板中。将固定浓度的嵌合IL-12(人p40与鼠p35融合)蛋白与梯度稀释的抗体混匀，37℃预孵育1小时后加入96孔板中，37℃细胞培养箱中培养48小时。取出96孔板，1000rpm离心3分钟，收集上清液，用小鼠IFN-gamma DuoSet ELISA Kit(R&D Systems，DY485)检测上清中IFNγ的含量。
检测IL-23诱导的T细胞分化时，将细胞悬液铺在包被好的96孔板中，将固定浓度的IL-23(R&D Systems，1290-IL-010)与梯度稀释的抗体混合预孵育1小时后加入96孔板中，37℃细胞培养箱中培养48小时。取出96孔板，1000rpm离心3分钟，收集上清液，用小鼠IL-17 DuoSet ELISA Kit(R&D Systems，DY421)检测上清中IL-17的含量。
实验结果见下表15和表16，实验结果表明，本披露的特异性结合BAFF和IL-12/23双特异性抗体能有效抑制IL-23诱导IL-17分泌，并能有效抑制IL-12诱导IFNγ分泌，而Belimumab既不能抑制IL-23诱导IL-17分泌，也不能抑制IL-12诱导IFNγ分泌。
表15.抗体抑制IL-12诱导IFNγ分泌实验结果
样品 抑制IL-12诱导IFNγ分泌的IC50(nM)
BU-1 1.595
Ustekinumab 3.075
Belimumab 无活性
表16.抗体抑制IL-23诱导IL-17分泌实验结果
样品 抑制IL-23诱导IL-17分泌的IC50(nM)
BU-1 0.02983
BU-2 0.03052
Belimumab 无活性
测试例5：亲和力测试
用生物传感芯片Protein A(GE，29127556)亲和捕获一定量的待测样品，然后于芯片表面流经一系列浓度梯度的抗原，利用Biacore(GE，8K)实时检测反应信号从而获得结合和解离曲线。在每个循环解离完成后，用10mM甘氨酸-盐酸溶液pH 1.5(GE，BR-1003-54)将生物芯片洗净再生。实验数据用BIA evaluation version 4.1，GE软件以1:1模型进行拟合，从而得出亲和力数值。本测试中用到的相关抗原蛋白如下：人IL-23(CT048-H08H，Sino biological)，人BAFF(10056-HNCH，Sino biological)，食蟹猴IL-12/23 P40(10215-CL，R&D Systems)，食蟹猴BAFF(BAF-CM412B，Kactus)，鼠BAFF(BAF-M521y，Acro Biosystems)。
实验结果见下表17、表18，实验结果表明，相比Belimumab，本披露构建的抗BAFF抗体能以更小的KD值与人BAFF结合；另外，本披露构建特异性结合BAFF和IL-12/23双特异性抗体能以高亲和力与人IL-23、人BAFF、食蟹猴IL-12/23 P40、食蟹猴BAFF、鼠BAFF结合。
表17.抗体与人BAFF亲和力检测实验结果
Figure PCTCN2022102621-appb-000043
表18.抗体与相关抗原亲和力检测实验结果
Figure PCTCN2022102621-appb-000044
测试例6：体内活性评价
用嵌合IL-12(人p40与鼠p35融合)，人IL-23和人BAFF三种蛋白同时刺激小鼠，诱导小鼠体内产生IFNγ、TNFα、IL-22和IgA等细胞因子，通过检测这些细胞因子的水平来评价抗体的体内活性。
SPF级雌性C57BL/6小鼠(北京维通利华实验动物技术有限公司，8周龄)，将小鼠进行随机分组，每组5只，通过腹腔混合注射嵌合IL-12(2μg/小鼠)、人IL-23(2μg/小鼠)和人BAFF(1mg/kg)三种蛋白，每天注射一次，持续四天。分别在第一天和第三天注射蛋白前一小时腹腔注射待测样品(Belimumab 8mpk，BU-1 10.7mpk，或BU-1 5.35mpk)。第五天收集各组小鼠血浆样品，分别检测IFNγ、TNFα、IL-22和IgA的水平。本测试例中所用的检测试剂盒来源信息如下：Mouse IFN-gamma Quantikine ELISA Kit(R&D Systems，MIF00)，Mouse TNF-alpha Quantikine ELISA Kit(R&D Systems，MTA00B)，Mouse/Rat IL-22 Quantikine ELISA Kit(R&D Systems，M2200)，Mouse IgA ELISA Kit(Abcam，ab157717)。Belimumab作为阳性对照，PBS作为阴性对照。BU-1 10.7mpk与Belimumab 8mpk的药物摩尔浓度相同。
实验结果见图3至图6，实验结果表明，本披露的特异性结合BAFF和IL-12/23双特异性抗体能显著抑制TNFα、IFNγ、IL-22的分泌，而Belimumab无法抑制TNFα、IFNγ的分泌。另外，BU-1在10.7mpk、5.35mpk两个剂量下均能显著的抑制IgA的分泌，并且抑制活性强于Belimumab。
测试例7.大鼠体内药代动力学实验
用SD大鼠进行体内药代动力学测试。雄性SD大鼠(浙江维通利华实验动物技术有限公司)随机分组，每组4只，静脉注射给药，给药组于给药前及给药后5分钟、8小时、24小时、48小时、84小时、9天、10天、14天、21天、28天采集全血0.2mL，不加抗凝，取血后在4℃放置30分钟，1000g离心15分钟，取上层血清置于EP管中，于-80℃保存。用ELISA法检测血清中的血药浓度，用Winnolin软件计算受试药物的药代动力学参数和体内半衰期。
实验结果见下表19，实验结果表明，本披露的特异性结合BAFF和IL-12/23双特异性抗体在大鼠中的半衰期很好，抗IL-12/23 p40端的半衰期达14天，半衰期比Ustekinumab更长。
表19.抗体大鼠体内药代动力学实验结果
名称 BU-2 Ustekinumab
给药剂量 4mg/kg 3mg/kg
抗IL-12/23 p40端半衰期 14.44±1.05天 12.02±0.87天

Claims (16)

  1. 一种抗原结合分子，其包含特异性结合BAFF的抗原结合模块1和特异性结合IL-12和/或IL-23的抗原结合模块2，其中，所述抗原结合模块1包含重链可变区B-VH和轻链可变区B-VL，所述B-VH包含B-HCDR1、B-HCDR2和B-HCDR3，所述B-VL包含B-LCDR1、B-LCDR2和B-LCDR3，其中：
    所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：63中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和
    所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：64中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，其中，
    SEQ ID NO：63为：
    QVQLQQSGAEVKKPGSSVRVSCKASGGTFNNNAINWVRQAPGQX 26LEWMGX 1IX 2PMFGX 3AKYSX 4X 5FQGRVAITADESTGTASMELSSLRSEDTAVYYCARSRDX 6LLFPX 7X 8X 9LX 10X 11WGX 27GTMVTVSS，
    SEQ ID NO：64为：
    SSELTQDPAVSVALGQTVRVTCX 12GX 13X 14LX 15X 16X 17X 18ASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCX 19SRX 20X 21X 22GX 23X 24WX 25FGX 28GTELTVL，
    其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V，X 26选自G或C，X 27选自R或G，X 28选自G或C；
    并且，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3不包含如下CDR组合：B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：1中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，并且B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：2中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列；
    优选地，
    所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3是根据Kabat编号规则定义的，其中，
    B-HCDR1包含氨基酸序列NNAIN(SEQ ID NO：18)，
    B-HCDR2包含氨基酸序列X 1IX 2PMFGX 3AKYSX 4X 5FQG(SEQ ID NO：65)，
    B-HCDR3包含氨基酸序列SRDX 6LLFPX 7X 8X 9LX 10X 11(SEQ ID NO：66)，
    B-LCDR1包含氨基酸序列X 12GX 13X 14LX 15X 16X 17X 18AS(SEQ ID NO：67)，
    B-LCDR2包含氨基酸序列GKNNRPS(SEQ ID NO：32)和
    B-LCDR3包含氨基酸序列X 19SRX 20X 21X 22GX 23X 24WX 25(SEQ ID NO：68)，
    其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V；
    并且，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3不包含如下CDR组合：B-HCDR1包含SEQ ID NO：18的氨基酸序列，B-HCDR2包含SEQ ID NO：19的氨基酸序列，B-HCDR3包含SEQ ID NO：20的氨基酸序列，B-LCDR1包含SEQ ID NO：31的氨基酸序列，B-LCDR2包含SEQ ID NO：32的氨基酸序列，和B-LCDR3包含SEQ ID NO：33的氨基酸序列。
  2. 根据权利要求1所述的抗原结合分子，其中：
    (i)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：47中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：48中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (ii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：5中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：12中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (iii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：6中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：13中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (iv)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：7中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：14中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (v)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：8中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：15中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (vi)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：9中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：16中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (vii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：10中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：17中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (viii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：11中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：2中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列；
    优选地，
    所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2、B-LCDR3是根据Kabat编号规则定义的，其中，
    (i)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：28的氨基酸序列，所述B-HCDR3包含SEQ ID NO：29的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：43的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和和所述B-LCDR3包含SEQ ID NO：44的氨基酸序列；或
    (ii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：21的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：34的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：35的氨基酸序列；或
    (iii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：22的氨基酸序列，所述B-HCDR3包含SEQ ID NO：23的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：31的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：36的氨基酸序列；或
    (iv)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：24的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：37的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：38的氨基酸序列；或
    (v)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包 含SEQ ID NO：25的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：39的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：40的氨基酸序列；或
    (vi)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：26的氨基酸序列，所述B-HCDR3包含SEQ ID NO：27的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：41的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：42的氨基酸序列；或
    (vii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：30的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：31的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：33的氨基酸序列。
  3. 根据权利要求1或2所述的抗原结合分子，其中，所述B-VH包含与SEQ ID NO：63具有至少90％序列同一性的氨基酸序列，所述B-VL包含与SEQ ID NO：64具有至少90％序列同一性的氨基酸序列，其中
    SEQ ID NO：63为：
    QVQLQQSGAEVKKPGSSVRVSCKASGGTFNNNAINWVRQAPGQX 26LEWMGX 1IX 2PMFGX 3AKYSX 4X 5FQGRVAITADESTGTASMELSSLRSEDTAVYYCARSRDX 6LLFPX 7X 8X 9LX 10X 11WGX 27GTMVTVSS，
    SEQ ID NO：64为：
    SSELTQDPAVSVALGQTVRVTCX 12GX 13X 14LX 15X 16X 17X 18ASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCX 19SRX 20X 21X 22GX 23X 24WX 25FGX 28GTELTVL，
    其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V，X 26选自G或C，X 27选自R或G，X 28选自G或C；
    并且，所述B-VH和B-VL不包含如下可变区组合：B-VH包含SEQ ID NO：1的氨基酸序列，且B-VL包含SEQ ID NO：2的氨基酸序列；
    优选地，
    (i)所述B-VH包含与SEQ ID NO：47具有至少90％序列同一性的氨基酸序 列，和所述B-VL包含与SEQ ID NO：48具有至少90％序列同一性的氨基酸序列，或
    (ii)所述B-VH包含与SEQ ID NO：5具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：12具有至少90％序列同一性的氨基酸序列，或
    (iii)所述B-VH包含与SEQ ID NO：6具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：13具有至少90％序列同一性的氨基酸序列，或
    (iv)所述B-VH包含与SEQ ID NO：7具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：14具有至少90％序列同一性的氨基酸序列，或
    (v)所述B-VH包含与SEQ ID NO：8具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：15具有至少90％序列同一性的氨基酸序列，或
    (vi)所述B-VH包含与SEQ ID NO：9具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：16具有至少90％序列同一性的氨基酸序列，或
    (vii)所述B-VH包含与SEQ ID NO：10具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：17具有至少90％序列同一性的氨基酸序列，或
    (viii)所述B-VH包含与SEQ ID NO：11具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：2具有至少90％序列同一性的氨基酸序列；
    更优选地，
    所述B-VH包含SEQ ID NO：47的氨基酸序列，和所述B-VL包含SEQ ID NO：48的氨基酸序列。
  4. 根据权利要求1至3中任一项所述的抗原结合分子，其中所述抗原结合模块2包含重链可变区P-VH和轻链可变区P-VL，所述P-VH包含P-HCDR1、P-HCDR2和P-HCDR3，所述P-VL包含P-LCDR1、P-LCDR2和P-LCDR3；
    所述P-HCDR1、P-HCDR2和P-HCDR3分别包含SEQ ID NO：51中的Pv-HCDR1、Pv-HCDR2和Pv-HCDR3的氨基酸序列，和所述P-LCDR1、P-LCDR2和P-LCDR3分别包含SEQ ID NO：52中的Pv-LCDR1、Pv-LCDR2和Pv-LCDR3的氨基酸序列；
    优选地，所述P-HCDR1、P-HCDR2、P-HCDR3、P-LCDR1、P-LCDR2和P-LCDR3是根据Kabat编号规则定义的，所述P-HCDR1包含SEQ ID NO：53的 氨基酸序列，所述P-HCDR2包含SEQ ID NO：54的氨基酸序列，所述P-HCDR3包含SEQ ID NO：55的氨基酸序列，所述P-LCDR1包含SEQ ID NO：56的氨基酸序列，所述P-LCDR2包含SEQ ID NO：57的氨基酸序列，和所述P-LCDR3包含SEQ ID NO：58的氨基酸序列；
    更优选地，所述重链可变区P-VH包含与SEQ ID NO：51具有至少90％序列同一性的氨基酸序列，和所述轻链可变区P-VL包含与SEQ ID NO：52具有至少90％序列同一性的氨基酸序列；
    最优选地，所述重链可变区P-VH包含SEQ ID NO：51的氨基酸序列，和所述轻链可变区P-VL包含SEQ ID NO：52的氨基酸序列。
  5. 根据权利要求1至4中任一项所述的抗原结合分子，其包括重链恒定区CH和轻链恒定区CL；
    优选地，所述重链恒定区CH为人IgG1重链恒定区，所述轻链恒定区CL为人κ轻链恒定区；
    更优选地，所述重链恒定区CH包含SEQ ID NO：59的氨基酸序列，所述轻链恒定区CL包含SEQ ID NO：60的氨基酸序列。
  6. 根据权利要求5所述的抗原结合分子，其中，所述抗原结合模块1为scFv，所述抗原结合模块2为包含重链可变区P-VH、轻链可变区P-VL、重链恒定区CH和轻链恒定区CL的全长抗体，所述抗原结合模块1直接或通过连接子融合至抗原结合模块2的可变区的N端或恒定区的C端；
    优选地，所述抗原结合分子包含：
    具有式(a)所示结构的第一链和具有式(b)所示结构的第二链；或者
    具有式(c)所示结构的第一链和具有式(b)所示结构的第二链，
    式(a)[P-VH]-[CH]-[连接子1]-[B-VH]-[连接子2]-[B-VL]，
    式(b)[P-VL]-[CL]，
    式(c)[P-VH]-[CH]-[连接子1]-[B-VL]-[连接子2]-[B-VH]，
    更优选地，式(a)和式(c)中，其中的连接子1和连接子2是相同或不同的肽连接子；
    最优选地，式(a)和式(c)中，其中连接子1的氨基酸序列如SEQ ID NO：69所示，连接子2的氨基酸序列如SEQ ID NO：70所示。
  7. 根据权利要求6所述的抗原结合分子，其中，
    所述第一链包含与SEQ ID NO：61具有至少90％序列同一性的氨基酸序列，和所述第二链包含与SEQ ID NO：50具有至少90％序列同一性的氨基酸序列，或
    所述第一链包含与SEQ ID NO：62具有至少90％序列同一性的氨基酸序列， 和所述第二链包含与SEQ ID NO：50具有至少90％序列同一性的氨基酸序列；
    优选地，所述抗原结合分子具有：
    (i)两条包含SEQ ID NO：61的氨基酸序列的第一链和两条包含SEQ ID NO：50的氨基酸序列的第二链，或者
    (ii)两条包含SEQ ID NO：62的氨基酸序列的第一链和两条包含SEQ ID NO：50的氨基酸序列的第二链。
  8. 一种抗原结合分子，其特异性结合BAFF抗原，其包含：
    重链可变区B-VH和轻链可变区B-VL，
    所述B-VH包含B-HCDR1、B-HCDR2和B-HCDR3，
    所述B-VL包含B-LCDR1、B-LCDR2和B-LCDR3，其中：
    所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：63中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：64中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，其中，
    SEQ ID NO：63为：
    QVQLQQSGAEVKKPGSSVRVSCKASGGTFNNNAINWVRQAPGQX 26LEWMGX 1IX 2PMFGX 3AKYSX 4X 5FQGRVAITADESTGTASMELSSLRSEDTAVYYCARSRDX 6LLFPX 7X 8X 9LX 10X 11WGX 27GTMVTVSS，
    SEQ ID NO：64为：
    SSELTQDPAVSVALGQTVRVTCX 12GX 13X 14LX 15X 16X 17X 18ASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCX 19SRX 20X 21X 22GX 23X 24WX 25FGX 28GTELTVL，
    其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V，X 26选自G或C，X 27选自R或G，X 28选自G或C；
    并且，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3不包含如下CDR组合：B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：1中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，并且B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：2中的B-LCDR1、B-LCDR2和B-LCDR3；
    优选地，
    所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3 是根据Kabat编号规则定义的，其中，
    B-HCDR1包含氨基酸序列NNAIN(SEQ ID NO：18)，
    B-HCDR2包含氨基酸序列X 1IX 2PMFGX 3AKYSX 4X 5FQG(SEQ ID NO：65)，
    B-HCDR3包含氨基酸序列SRDX 6LLFPX 7X 8X 9LX 10X 11(SEQ ID NO：66)，
    B-LCDR1包含氨基酸序列X 12GX 13X 14LX 15X 16X 17X 18AS(SEQ ID NO：67)，
    B-LCDR2包含氨基酸序列GKNNRPS(SEQ ID NO：32)和
    B-LCDR3包含氨基酸序列X 19SRX 20X 21X 22GX 23X 24WX 25(SEQ ID NO：68)，
    其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、H或R，X 25选自L或V；
    并且，所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3不包含如下CDR组合：B-HCDR1包含SEQ ID NO：18的氨基酸序列，B-HCDR2包含SEQ ID NO：19的氨基酸序列，B-HCDR3包含SEQ ID NO：20的氨基酸序列，B-LCDR1包含SEQ ID NO：31的氨基酸序列，B-LCDR2包含SEQ ID NO：32的氨基酸序列，和B-LCDR3包含SEQ ID NO：33的氨基酸序列。
  9. 根据权利要求8所述的抗原结合分子，其中：
    (i)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：47中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：48中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (ii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：5中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：12中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (iii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：6中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：13中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (iv)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：7中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：14中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3 的氨基酸序列，或
    (v)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：8中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：15中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (vi)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：9中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：16中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (vii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：10中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：17中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列，或
    (viii)所述B-HCDR1、B-HCDR2和B-HCDR3分别包含SEQ ID NO：11中的Bv-HCDR1、Bv-HCDR2和Bv-HCDR3的氨基酸序列，和所述B-LCDR1、B-LCDR2和B-LCDR3分别包含SEQ ID NO：2中的Bv-LCDR1、Bv-LCDR2和Bv-LCDR3的氨基酸序列；
    优选地，
    所述B-HCDR1、B-HCDR2、B-HCDR3、B-LCDR1、B-LCDR2和B-LCDR3是根据Kabat编号规则定义的，并且，
    (i)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：28的氨基酸序列，所述B-HCDR3包含SEQ ID NO：29的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：43的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：44的氨基酸序列；或
    (ii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：21的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：34的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：35的氨基酸序列；或
    (iii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：22的氨基酸序列，所述B-HCDR3包含SEQ ID NO：23的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：31的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：36的氨基酸序列；或
    (iv)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包 含SEQ ID NO：24的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：37的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：38的氨基酸序列；或
    (v)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：25的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：39的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：40的氨基酸序列；或
    (vi)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：26的氨基酸序列，所述B-HCDR3包含SEQ ID NO：27的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：41的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：42的氨基酸序列；或
    (vii)所述B-HCDR1包含SEQ ID NO：18的氨基酸序列，所述B-HCDR2包含SEQ ID NO：30的氨基酸序列，所述B-HCDR3包含SEQ ID NO：20的氨基酸序列，和所述B-LCDR1包含SEQ ID NO：31的氨基酸序列，所述B-LCDR2包含SEQ ID NO：32的氨基酸序列，和所述B-LCDR3包含SEQ ID NO：33的氨基酸序列。
  10. 根据权利要求8或9所述的抗原结合分子，其中，所述B-VH包含与SEQ ID NO：63具有至少90％序列同一性的氨基酸序列，所述B-VL包含与SEQ ID NO：64具有至少90％序列同一性的氨基酸序列，其中
    SEQ ID NO：63为：
    QVQLQQSGAEVKKPGSSVRVSCKASGGTFNNNAINWVRQAPGQX 26LEWMGX 1IX 2PMFGX 3AKYSX 4X 5FQGRVAITADESTGTASMELSSLRSEDTAVYYCARSRDX 6LLFPX 7X 8X 9LX 10X 11WGX 27GTMVTVSS，
    SEQ ID NO：64为：
    SSELTQDPAVSVALGQTVRVTCX 12GX 13X 14LX 15X 16X 17X 18ASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCX 19SRX 20X 21X 22GX 23X 24WX 25FGX 28GTELTVL，
    其中，X 1选自S、A、V或G，X 2选自A、I、M或S，X 3选自T或G，X 4选自Q、E、G或K，X 5选自N、G或Q，X 6选自L或P，X 7选自H、Q或D，X 8选自H或D，X 9选自A或G，X 10选自S或L，X 11选自P或S，X 12选自Q或H，X 13选自D、A或N，X 14选自S或I，X 15选自K、R或T，X 16选自S、W、T或D，X 17选自H、Y或S，X 18选自Y或R，X 19选自G或S，X 20选自A或D，X 21选自E或S，X 22选自S或A，X 23选自V、E、A、N或W，X 24选自G、K、 H或R，X 25选自L或V，X 26选自G或C，X 27选自R或G，X 28选自G或C；
    并且，所述B-VH和B-VL不包含如下可变区组合：B-VH包含SEQ ID NO：1的氨基酸序列，且B-VL包含SEQ ID NO：2的氨基酸序列；
    优选地，
    (i)所述B-VH包含与SEQ ID NO：47具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：48具有至少90％序列同一性的氨基酸序列，或
    (ii)所述B-VH包含与SEQ ID NO：5具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：12具有至少90％序列同一性的氨基酸序列，或
    (iii)所述B-VH包含与SEQ ID NO：6具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：13具有至少90％序列同一性的氨基酸序列，或
    (iv)所述B-VH包含与SEQ ID NO：7具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：14具有至少90％序列同一性的氨基酸序列，或
    (v)所述B-VH包含与SEQ ID NO：8具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：15具有至少90％序列同一性的氨基酸序列，或
    (vi)所述B-VH包含与SEQ ID NO：9具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：16具有至少90％序列同一性的氨基酸序列，或
    (vii)所述B-VH包含与SEQ ID NO：10具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：17具有至少90％序列同一性的氨基酸序列，或
    (viii)所述B-VH包含与SEQ ID NO：11具有至少90％序列同一性的氨基酸序列，和所述B-VL包含与SEQ ID NO：2具有至少90％序列同一性的氨基酸序列；
    更优选地，
    所述B-VH包含SEQ ID NO：47的氨基酸序列，和所述B-VL包含SEQ ID NO：48的氨基酸序列，或
    所述B-VH包含SEQ ID NO：5的氨基酸序列，和所述B-VL包含SEQ ID NO：12的氨基酸序列，或
    所述B-VH包含SEQ ID NO：6的氨基酸序列，和所述B-VL包含SEQ ID NO：13的氨基酸序列，或
    所述B-VH包含SEQ ID NO：7的氨基酸序列，和所述B-VL包含SEQ ID NO： 14的氨基酸序列，或
    所述B-VH包含SEQ ID NO：8的氨基酸序列，和所述B-VL包含SEQ ID NO：15的氨基酸序列，或
    所述B-VH包含SEQ ID NO：9的氨基酸序列，和所述B-VL包含SEQ ID NO：16的氨基酸序列，或
    所述B-VH包含SEQ ID NO：10的氨基酸序列，和所述B-VL包含SEQ ID NO：17的氨基酸序列，或
    所述B-VH包含SEQ ID NO：11的氨基酸序列，和所述B-VL包含SEQ ID NO：2的氨基酸序列。
  11. 根据权利要求8至10中任一项所述的抗原结合分子，其包括重链恒定区CH和轻链恒定区CL；
    优选地，所述重链恒定区CH为人IgG1重链恒定区，所述轻链恒定区CL为人λ轻链恒定区；
    更优选地，所述重链恒定区CH包含SEQ ID NO：45的氨基酸序列，所述轻链恒定区CL包含SEQ ID NO：46的氨基酸序列。
  12. 根据权利要求1至11中任一项所述的抗原结合分子，所述抗原结合分子具有一种或更多种以下特征：
    A.所述抗原结合分子能阻断人BAFF与人BAFF-R的结合；优选地，阻断人BAFF与人BAFF-R结合的IC50值小于11nM，所述IC50值通过Elisa方法检测；
    B.所述抗原结合分子能抑制BAFF诱导的B细胞增殖；优选地，抗原结合分子能以小于0.2nM的IC50值抑制BAFF诱导的B细胞增殖；
    C.所述抗原结合分子能以小于9.99E-10M的KD值与人BAFF结合，所述KD值通过Biacore方法检测；
    D.所述抗原结合分子能以小于5.00E-10M的KD值与食蟹猴BAFF结合，所述KD值通过Biacore方法检测；
    E.所述抗原结合分子能以小于2.20E-10M的KD值与鼠BAFF结合，所述KD值通过Biacore方法检测；
    F.所述抗原结合分子能阻断人BAFF与人BCMA的结合；优选地，阻断人BAFF与人BCMA结合的IC50值小于0.9nM，所述IC50值通过Elisa方法检测；或
    G.所述抗原结合分子能阻断人BAFF与人TACI的结合；优选地，阻断人BAFF与人TACI结合的IC50值小于0.6nM，所述IC50值通过Elisa方法检测。
  13. 一种药物组合物，其含有：
    治疗有效量的权利要求1至12中任一项所述的抗原结合分子，以及
    一种或更多种药学上可接受的载体、稀释剂、缓冲剂或赋形剂。
  14. 分离的核酸，其编码权利要求1至12中任一项所述的抗原结合分子。
  15. 宿主细胞，其包含如权利要求14所述的分离的核酸。
  16. 一种治疗疾病的方法，所述方法包括向受试者施用权利要求1至12中任一项所述的抗原结合分子或权利要求13所述的组合物的步骤；
    优选地，所述疾病是B细胞障碍或自身免疫性疾病；
    更优选地，所述自身免疫性疾病选自：系统性红斑狼疮、重症肌无力、多发性硬化、胰岛素依赖性糖尿病、克罗恩氏病、类风湿关节炎、多关节型青少年类风湿关节炎和银屑病性关节炎；
    所述B细胞障碍选自：肿瘤、慢性白细胞性白血病、多发性骨髓瘤、非霍奇金淋巴瘤、移植后淋巴组织增生病和轻链丙球蛋白病；
    最优选地，所述疾病为系统性红斑狼疮。
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