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CN-122029195-A - BDCA2 monoclonal antibody, fusion protein composed of BDCA2 monoclonal antibody and TACI, and method and application thereof

CN122029195ACN 122029195 ACN122029195 ACN 122029195ACN-122029195-A

Abstract

A monoclonal antibody specifically binding to BDCA2 and a bispecific fusion protein of BDCA2 and TACI constructed based on the BDCA2 monoclonal antibody are provided. Immunoconjugates, bispecific molecules and pharmaceutical compositions comprising the antibodies, and diagnostic and therapeutic methods of using the anti-BDCA 2 antibodies and BDCA2/TACI bispecific fusion proteins are also provided.

Inventors

  • WANG BAOHUI
  • SUN JIANMING
  • LI FENGXIA
  • HUANG XIAO
  • LING HONG

Assignees

  • 南京维立志博生物科技股份有限公司

Dates

Publication Date
20260512
Application Date
20240927
Priority Date
20230928

Claims (20)

  1. An anti-BDCA 2 antibody or antigen-binding fragment thereof, wherein the anti-BDCA 2 antibody or antigen-binding fragment thereof comprises a heavy chain variable region VH and/or a light chain variable region VL, wherein the heavy chain variable region VH comprises 3 CDRs, HCDR1, HCDR2 and HCDR3, and the light chain variable region VL comprises 3 CDRs, LCDR1, LCDR2 and LCDR3, wherein (I) The HCDR1, HCDR2 and HCDR3 are selected from three complementarity determining regions HCDR1, HCDR2 and HCDR3 contained in the VH as shown in SEQ ID NO:1, and the LCDR1, LCDR2 and LCDR3 are three complementarity determining regions LCDR1, LCDR2 and LCDR3 contained in the VL as shown in SEQ ID NO: 2; (ii) The HCDR1, HCDR2 and HCDR3 are selected from three complementarity determining regions HCDR1, HCDR2 and HCDR3 contained in the VH as shown in SEQ ID NO:3, and the LCDR1, LCDR2 and LCDR3 are three complementarity determining regions LCDR1, LCDR2 and LCDR3 contained in the VL as shown in SEQ ID NO:4, or (Iii) The HCDR1, HCDR2 and HCDR3 are selected from the three complementarity determining regions HCDR1, HCDR2 and HCDR3 contained in the VH as shown in SEQ ID NO:13, and the LCDR1, LCDR2 and LCDR3 are the three complementarity determining regions LCDR1, LCDR2 and LCDR3 contained in the VL as shown in SEQ ID NO:15, or (Iv) The HCDR1, HCDR2 and HCDR3 are selected from the three complementarity determining regions HCDR1, HCDR2 and HCDR3 contained in the VH as shown in SEQ ID NO:14, and the LCDR1, LCDR2 and LCDR3 are the three complementarity determining regions LCDR1, LCDR2 and LCDR3 contained in the VL as shown in SEQ ID NO:15, or (V) A sequence comprising at least one and no more than 5, 4,3, 2 or 1 amino acid changes (preferably amino acid substitutions, preferably conservative substitutions) relative to the sequence of any one of (i) - (iv) on the three HCDR regions or the three LCDR regions, e.g., the amino acid changes are mutations that enhance antibody stability; preferably, the CDRs are determined by the Kabat numbering scheme.
  2. The anti-BDCA 2 antibody or antigen-binding fragment thereof of claim 1, comprising (I) HCDR1 as shown in SEQ ID NO. 33, HCDR2 as shown in SEQ ID NO. 34, HCDR3 as shown in SEQ ID NO. 35, LCDR1 as shown in SEQ ID NO. 36, LCDR2 as shown in SEQ ID NO. 37 and LCDR3 as shown in SEQ ID NO. 38, or (Ii) HCDR1 as shown in SEQ ID NO. 39, HCDR2 as shown in SEQ ID NO. 40 or 49, HCDR3 as shown in SEQ ID NO. 41, LCDR1 as shown in SEQ ID NO. 42 or 48, LCDR2 as shown in SEQ ID NO. 43 and LCDR3 as shown in SEQ ID NO. 44, or (Iii) A sequence comprising at least one and no more than 5, 4,3, 2 or 1 amino acid changes (preferably amino acid substitutions, preferably conservative substitutions) relative to the sequence of any one of (i) or (ii) on the three HCDR regions or the three LCDR regions, e.g., the amino acid changes are mutations that enhance antibody stability.
  3. The anti-BDCA 2 antibody or antigen-binding fragment thereof of claim 1 or 2, wherein the heavy chain variable region VH (I) Comprising or consisting of an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence of SEQ ID NO. 1,3, 13 or 14, or (Ii) Comprising or consisting of the amino acid sequence of SEQ ID NO.1, 3, 13 or 14.
  4. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-3, wherein the light chain variable region VL (I) Comprising or consisting of an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence of SEQ ID NO. 2,4 or 15, or (Ii) Comprising or consisting of the amino acid sequence of SEQ ID NO. 2, 4 or 15.
  5. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-4, comprising a heavy chain variable region VH and a light chain variable region VL, wherein (I) The heavy chain variable region comprises or consists of the amino acid sequence shown in SEQ ID NO. 1 or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity thereto, and the light chain variable region comprises or consists of the amino acid sequence shown in SEQ ID NO. 2 or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity thereto, or (Ii) The heavy chain variable region comprises or consists of the amino acid sequence shown in SEQ ID NO. 3 or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity thereto, and the light chain variable region comprises or consists of the amino acid sequence shown in SEQ ID NO. 4 or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity thereto, or (Iii) The heavy chain variable region comprises or consists of the amino acid sequence shown in SEQ ID NO. 13 or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity thereto, and the light chain variable region comprises or consists of the amino acid sequence shown in SEQ ID NO. 15 or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity thereto, or (Iv) The heavy chain variable region comprises or consists of the amino acid sequence shown in SEQ ID NO. 14 or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity thereto, and the light chain variable region comprises or consists of the amino acid sequence shown in SEQ ID NO. 15 or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity thereto.
  6. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-5, wherein VH and VL comprise or consist of the amino acid sequences shown below, respectively: (i) SEQ ID NO. 1 and SEQ ID NO. 2; (ii) SEQ ID NO. 3 and SEQ ID NO. 4; (iii) SEQ ID NO. 13 and SEQ ID NO. 15; (iv) SEQ ID NO. 14 and SEQ ID NO. 15.
  7. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-6, further comprising an Fc or heavy chain constant region CH, e.g., The Fc region is that of IgG1, igG2, igG3 or IgG4, preferably that of IgG1, or The antibody heavy chain constant region CH is a heavy chain constant region of IgG1, igG2, igG3 or IgG4, preferably a heavy chain constant region of IgG1, Optionally, the sequence of the heavy chain constant region has one or more amino acid substitutions compared to the native human heavy chain constant region sequence, preferably such that the ADCC and/or CDC effect of the antibody is enhanced and/or the in vivo half-life is prolonged and/or the affinity for FcRn is increased, More preferably, the one or more amino acid substitutions is a combination of amino acid substitutions M252Y/S254T/T256E according to EU numbering, e.g.the heavy chain constant region comprises or consists of the amino acid sequence of SEQ ID NO. 54.
  8. The anti-BDCA 2 antibody or antigen-binding fragment thereof of claim 7, wherein the heavy chain constant region CH (I) Comprising or consisting of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence of SEQ ID NO. 5, or (Ii) Comprising or consisting of the amino acid sequence of SEQ ID NO. 5.
  9. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-8, further comprising a light chain constant region, e.g., the light chain constant region is a lambda or kappa light chain constant region.
  10. The anti-BDCA 2 antibody or antigen-binding fragment thereof of claim 9, wherein the light chain constant region (I) Comprising or consisting of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence of SEQ ID NO. 6, or (Ii) Comprising or consisting of the amino acid sequence of SEQ ID NO. 6.
  11. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-10, comprising a heavy chain, wherein the heavy chain comprises or consists of the amino acid sequence of SEQ ID NO 9, 11, 16, or 17, or an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to the amino acid sequence.
  12. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-11, comprising a light chain, wherein the light chain comprises the amino acid sequence of SEQ ID NO:10, 12 or 18, or an amino acid sequence comprising or consisting of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to said amino acid sequence.
  13. The anti-BDCA 2 antibody or antigen-binding fragment thereof of claim 11 or 12, comprising a heavy chain and a light chain, wherein (I) The heavy chain comprises or consists of the amino acid sequence of SEQ ID NO. 9, or comprises or consists of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence, and the light chain comprises or consists of the amino acid sequence of SEQ ID NO. 10, or comprises or consists of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence; (ii) The heavy chain comprises or consists of the amino acid sequence of SEQ ID NO. 11, or comprises or consists of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence, and the light chain comprises or consists of the amino acid sequence of SEQ ID NO. 12, or comprises or consists of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence; (iii) The heavy chain comprises or consists of the amino acid sequence of SEQ ID NO. 16, or comprises or consists of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence, and the light chain comprises or consists of the amino acid sequence of SEQ ID NO. 18, or comprises or consists of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence, or consists of the amino acid sequence (Iv) The heavy chain comprises or consists of the amino acid sequence of SEQ ID NO. 17, or comprises or consists of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence, and the light chain comprises or consists of the amino acid sequence of SEQ ID NO. 18, or comprises or consists of an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence.
  14. The anti-BDCA 2 antibody or antigen-binding fragment thereof of claim 13, wherein the heavy and light chains comprise or consist of the amino acid sequence set forth in SEQ ID NO: (i) SEQ ID NO. 9 and SEQ ID NO. 10; (ii) SEQ ID NO. 11 and SEQ ID NO. 12; (iii) SEQ ID NO. 16 and SEQ ID NO. 18; (iv) SEQ ID NO. 17 and SEQ ID NO. 18.
  15. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-14, wherein the antibody or antigen-binding fragment thereof is ADCC-enhancing and/or half-life-extending, e.g., it has an altered glycosylation pattern, e.g., is hypofucosylation or defucosylation or comprises a combination of amino acid substitutions with increased bisected GlcNac structure, and/or carries, e.g., M252Y/S254T/T256E.
  16. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-14, which is a humanized or chimeric antibody.
  17. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-16, which is a monoclonal antibody.
  18. The anti-BDCA 2 antibody or antigen-binding fragment thereof of any one of claims 1-17, wherein the antigen-binding fragment is selected from the group consisting of Fab, fab '-SH, fv, single chain antibody (e.g., scFv), (Fab') 2 , single domain antibody such as VHH, dAb (domain anti) or linear antibody.
  19. A fusion protein molecule that specifically binds to human BDCA2, human BAFF and/or APRIL.
  20. The fusion protein molecule of claim 19, comprising a first target binding region and a second target binding region, wherein the first target binding region specifically binds BDCA2 and the second target binding region specifically binds BAFF and/or APRIL.

Description

BDCA2 monoclonal antibody, fusion protein composed of BDCA2 monoclonal antibody and TACI, and method and application thereof Technical Field The present invention is in the fields of molecular biology and biologicals. In particular, the invention relates to monoclonal antibodies or fragments thereof, and bispecific fusion proteins comprising monoclonal antibodies or fragments thereof, as well as methods of making and uses thereof. Background Systemic lupus erythematosus (SLE, systemic lupus erythematosus) is a chronic, systemic autoimmune disease affecting a number of organs throughout the body through immune complexes, autoantibodies, cellular immunity and inflammatory responses, including the skin, joints, central nervous system and kidneys (Kaul A,Gordon C,Crow MK,Touma Z,Urowitz MB,van Vollenhoven R,Ruiz-Irastorza G,Hughes G.Systemic lupus erythematosus.Nat Rev Dis Primers.2016Jun 16;2:16039.).pDCs(Plasmacytoid dendritic cells) are a special class of dendritic cells (DCs, DENDRITIC CELLS), which can sense nucleic acids through TLR7 or TLR9, produce large amounts of type I Interferons (IFNs), and cause the body's innate immune response .(Saadeh D,Kurban M,Abbas O.Update on the role of plasmacytoid dendritic cells in inflammatory/autoimmune skin diseases.Exp Dermatol.2016Jun;25(6):415-21). to play an important role in disease progression in SLE in addition to pDCs. B cells can secrete not only autoantibodies, but also as antigen presenting cells, affecting T cell function, and thus secreting a range of pro-inflammatory factors (Chan,VF.,Tsang,HL.,Tam,RY.et al.B-cell-targeted therapies in systemic lupus erythematosus.Cell Mol Immunol 10,133–142(2013))., thus targeting pDCs and/or B cells is an effective approach to treat SLE. BDCA2 (Blood DC ANTIGEN 2) is a type C lectin receptor specifically expressed on the surface of human plasma cell-like dendritic cells (plasma cell DENDRITIC CELL, pDC), and through binding with the pDCs membrane surface receptor fcεRIgamma, the BDCA2 activates the signal path regulated by ITAM to cause downstream protein phosphorylation, and finally can inhibit CpG-induced pDCs from generating type I interferon (Gilliet M,Cao W,Liu YJ.Plasmacytoid dendritic cells:sensing nucleic acids in viral infection and autoimmune diseases.Nat Rev Immunol.2008Aug;8(8):594-606)., thus, BDCA2 is targeted, the BDCA2 downstream signal path is further activated, and the generation of type I interferon is inhibited, so that the BDCA2 is an effective path for treating SLE. TACI (Transmembrane activator and calcium-modulating cyclophilin ligand interactor) is a B cell surface receptor whose extracellular domain contains 2 cysteine-rich regions, capable of binding to BAFF (B CELL ACTIVATING factor of the tumor necrosis factor family) and APRIL (proliferation-inducing ligand). BAFF is a type II transmembrane protein, belonging to the TNF ligand superfamily, and is produced mainly by myeloid cells, such as monocytes, macrophages, neutrophils and DCs. BAFF is mainly composed of two forms-membrane-bound and soluble, its receptors including TACI, BCMA (B cell maturation antigen) and BAFF-R (BAFF receptor). APRIL is also a type II transmembrane protein, belonging to the TNF ligand superfamily, produced mainly by myeloid cells. Unlike BAFF, APRIL is secreted only. APRIL shares two receptors, BCMA and TACI, with BAFF. After TACI is combined with BAFF and APRIL, class conversion of IgG and IgA of B cells can be induced, and differentiation and survival of plasma cells can be promoted .(Zhang Y,Li J,Zhang YM,Zhang XM,Tao J.Effect of TACI signaling on humoral immunity and autoimmune diseases.J Immunol Res.2015;2015:247426.) The invention aims to construct a bifunctional molecule for simultaneously constructing a targeting BDCA2 and BAFF/APRIL, and on one hand, the invention eliminates pDCs, activates a BDCA2 downstream signal path and inhibits the expression of type I interferon. On the other hand, blocking the binding of BAFF/APRIL to its receptor, inhibiting B cell over-activation, may show better therapeutic effect on SLE. Disclosure of Invention The present invention provides novel anti-BDCA 2 antibodies or antigen binding fragments thereof that have the advantages of high affinity and high specificity for human and cynomolgus monkey BDCA 2. The invention also provides a novel bispecific fusion protein which targets BDCA2 and BAFF/APRIL simultaneously. Polynucleotides encoding said antibodies, bispecific fusion proteins or fragments thereof, vectors comprising said polynucleotides, host cells comprising said polynucleotides or vectors, methods of treating diseases associated with BDCA2 and/or BAFF/APRIL using said antibodies, bispecific fusion proteins, and the use of said antibodies, bispecific fusion proteins in the treatment, prevention and/or diagnosis of diseases associated with BDCA2 and/or BAFF/APRIL in an individual. The anti-BDCA 2 antibodies, BDCA2 xbaff/APRIL bispecific fusion proteins, or antigen/tar