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WO2025168059A1 - Protéine de liaison à l'antigène multi-spécifique ciblant bcma, cd19 et cd3 et son utilisation - Google Patents

Protéine de liaison à l'antigène multi-spécifique ciblant bcma, cd19 et cd3 et son utilisation

Info

Publication number
WO2025168059A1
WO2025168059A1 PCT/CN2025/076236 CN2025076236W WO2025168059A1 WO 2025168059 A1 WO2025168059 A1 WO 2025168059A1 CN 2025076236 W CN2025076236 W CN 2025076236W WO 2025168059 A1 WO2025168059 A1 WO 2025168059A1
Authority
WO
WIPO (PCT)
Prior art keywords
seq
antibody
bcma
variable region
antigen
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/CN2025/076236
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English (en)
Chinese (zh)
Inventor
涂桂云
沈武忠
陈汉阳
杨铮
严孝强
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Itabmed Biopharmaceutical Shanghai Co Ltd
Original Assignee
Itabmed Biopharmaceutical Shanghai Co Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Itabmed Biopharmaceutical Shanghai Co Ltd filed Critical Itabmed Biopharmaceutical Shanghai Co Ltd
Publication of WO2025168059A1 publication Critical patent/WO2025168059A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

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Classifications

    • 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 [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants

Definitions

  • the present invention relates to the field of biomedicine. Specifically, the present invention relates to a multispecific antigen-binding protein targeting BCMA, CD19 and CD3 and its application.
  • the immunotherapies for treating cancer patients mainly include: 1) Checkpoint Inhibitor (CPI), 2) Chimeric Antigen Receptor T cells (CAR-T), 3) Antibody Drug Conjugate (ADC), and 4) T Cell Engagement (TCE).
  • CPI Checkpoint Inhibitor
  • CAR-T Chimeric Antigen Receptor T cells
  • ADC Antibody Drug Conjugate
  • TCE T Cell Engagement
  • the T cell engager is an artificially designed molecule that can specifically bind to two or more different antigens, one end of which binds to the CD3 antigen and the other end binds to one or more tumor-associated antigens (TAAs) and/or tumor-specific antigens (TSAs).
  • TAAs tumor-associated antigens
  • TSAs tumor-specific antigens
  • This T cell engager can cross-link tumor cells and T cells, namely the T cell driver (T cell engager), and then form synapses at the contact site, activating cytotoxic signals and leading to tumor cell lysis.
  • T cells When T cells are activated, the CD25 and CD69 cell surface markers are upregulated and cytokines are released.
  • the secreted cytokines such as interleukin-2 (IL-2), can further activate T cells and promote the proliferation of T cells to enhance the ability to kill tumor cells.
  • IL-2 interleukin-2
  • CD19 is a transmembrane glycoprotein that is primarily expressed in the early stages of human B cell development and persists on mature B cells. As part of the B cell antigen receptor complex, it plays a key role in B cell activation, proliferation, and differentiation. High expression of CD19 is not only observed on normal B cells, but is also prevalent in a variety of B cell tumors, such as B cell acute lymphoblastic leukemia (B-ALL) and non-Hodgkin's lymphoma (NHL). Therefore, CD19 has become an ideal biomarker for the treatment of targeted B cell malignancies.
  • B-ALL B cell acute lymphoblastic leukemia
  • NHL non-Hodgkin's lymphoma
  • BCMA B-cell maturation antigen
  • TNFRSF tumor necrosis factor receptor superfamily
  • BCMA expression is significantly increased and is closely associated with tumor cell proliferation and survival.
  • Tumor cells can activate BCMA through autocrine APRIL and BAFF, thereby promoting self-proliferation and evading immune system surveillance.
  • High BCMA expression is also associated with disease progression and poor prognosis, making it a clinically significant biomarker.
  • BCMA-targeted therapeutic strategies can achieve specific attacks on diseased B cells while having relatively minimal effects on normal tissues. Furthermore, BCMA-targeted therapies can work through multiple mechanisms, including directly inducing tumor cell apoptosis, blocking tumor growth signals, and activating the immune system's attack on tumor cells.
  • CD19 or BCMA targeted therapies have achieved remarkable clinical results, such as CarT therapy, their complex manufacturing process and high cost limit the widespread application of these therapies.
  • CD19 mono-targeted therapy or BCMA mono-targeted therapy the improvement of long-term survival rate, and the optimization of efficacy in children and the elderly.
  • future treatments may include the combination of targeted therapy with other treatment methods, such as radiotherapy, chemotherapy, immunomodulators or other emerging targeted therapies.
  • simultaneous targeting of CD19 and BCMA can avoid antigen escape of a single target. This multimodal treatment strategy may improve the treatment effect and provide patients with a more comprehensive treatment plan.
  • the purpose of the present invention is to provide a T cell engager targeting CD19 and BCMA.
  • an anti-BCMA antibody or an antigen-binding fragment thereof is provided, wherein the antibody comprises a heavy chain variable region and a light chain variable region,
  • the heavy chain variable region has the following complementarity determining regions CDR:
  • VH-CDR2 shown in SEQ ID NO:15, and
  • VH-CDR3 shown in SEQ ID NO:16;
  • the light chain variable region has the following complementarity determining regions (CDRs):
  • VL-CDR1 shown in SEQ ID NO: 17,
  • VL-CDR2 shown in SEQ ID NO:18.
  • VL-CDR3 shown in SEQ ID NO:19.
  • the antigen-binding fragment is selected from the group consisting of Fab, (Fab')2, scFv, or a combination thereof.
  • the antibody is a monospecific antibody, a bispecific antibody or a multispecific antibody.
  • the antibody is a single-chain antibody having a structure shown in Formula Ia or Ib from N-terminus to C-terminus:
  • VL BCMA is the light chain variable region of the anti-BCMA antibody
  • VH BCMA is the heavy chain variable region of the anti-BCMA antibody
  • L1 is a connecting peptide
  • amino acid sequence of the antibody heavy chain variable region is as shown in SEQ ID No: 25 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto, and/or
  • the amino acid sequence of the antibody light chain variable region is as shown in SEQ ID No: 26 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • L1 is a flexible connecting peptide.
  • L1 has an amino acid sequence as shown in (GGGGS)n or GG, wherein n is an integer of 1-6, preferably 2, 3 or 4.
  • L1 has an amino acid sequence as shown in SEQ ID NO:34.
  • an anti-CD3 antibody or an antigen-binding fragment thereof is provided, wherein the antibody comprises a heavy chain variable region and a light chain variable region.
  • the heavy chain variable region has the following complementarity determining regions CDR:
  • VH-CDR1 shown in SEQ ID NO: 1,
  • VH-CDR2 shown in SEQ ID NO:7, and
  • VH-CDR3 shown in SEQ ID NO:3;
  • the light chain variable region has the following complementarity determining regions (CDRs):
  • VL-CDR1 shown in SEQ ID NO:4,
  • VL-CDR2 shown in SEQ ID NO:5, and
  • VL-CDR3 shown in SEQ ID NO:6.
  • the antigen-binding fragment is selected from the group consisting of Fab, (Fab')2, scFv, or a combination thereof.
  • the antibody is a monospecific antibody, a bispecific antibody or a multispecific antibody.
  • amino acid sequence of the antibody heavy chain variable region is as shown in SEQ ID No: 22 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto, and/or
  • the antibody further comprises a light chain constant region and/or a heavy chain constant region.
  • the antibody comprises a light chain constant region CL and a heavy chain constant region CH1.
  • the light chain constant region CL has a sequence as shown in SEQ ID NO:36.
  • the heavy chain constant region CH1 has a sequence as shown in SEQ ID NO:35.
  • a multispecific antibody comprises the anti-BCMA antibody or antigen-binding fragment thereof according to the first aspect of the present invention, and/or the anti-CD3 antibody or antigen-binding fragment thereof according to the second aspect of the present invention.
  • the multispecific antibody is a trispecific antibody
  • the trispecific antibody comprises:
  • a CD19 targeting binding domain comprising one or more antigen binding domains that specifically bind to the CD19 protein
  • BCMA targeting binding domain comprising one or more antigen binding domains that specifically bind to the BCMA protein
  • CD3 targeting binding domain which comprises one or more antigen binding domains that specifically bind to CD3 protein.
  • the antigen binding domain that specifically binds to the CD19 protein includes a heavy chain variable region and a light chain variable region.
  • the heavy chain variable region has the following complementarity determining regions CDR:
  • VH-CDR2 shown in SEQ ID NO:9
  • VH-CDR3 shown in SEQ ID NO:10;
  • VL-CDR1 shown in SEQ ID NO: 11,
  • VL-CDR2 shown in SEQ ID NO:12, and
  • VL-CDR3 shown in SEQ ID NO:13.
  • the antigen binding domain that specifically binds to CD19 protein is selected from the following group: single-chain antibody, double-chain antibody, or a combination thereof.
  • the antigen binding domain that specifically binds to CD19 protein is an anti-CD19 single-chain antibody or Fab.
  • the anti-CD19 single-chain antibody has a structure shown in the following formula IIa or IIb from N-terminus to C-terminus:
  • VL CD19 is the light chain variable region of the anti-CD19 antibody
  • VH CD19 is the heavy chain variable region of the anti-CD19 antibody
  • L2 is a connecting peptide
  • amino acid sequence of the heavy chain variable region of the anti-CD19 antibody is as shown in SEQ ID No: 23 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • amino acid sequence of the light chain variable region of the anti-CD19 antibody is as shown in SEQ ID No: 24 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • L2 is a flexible connecting peptide.
  • L2 has an amino acid sequence as shown in (GGGGS)n or GG, wherein n is an integer of 1-6, preferably 2, 3 or 4.
  • L2 has an amino acid sequence as shown in SEQ ID NO:34.
  • the antigen-binding domain that specifically binds to the BCMA protein is the anti-BCMA antibody or antigen-binding fragment thereof as described in the first aspect of the present invention.
  • the antigen-binding domain that specifically binds to the BCMA protein is a single-chain antibody or Fab, preferably a single-chain antibody represented by Formula Ia or Ib.
  • the CD3 targeting binding domain includes a heavy chain variable region and a light chain variable region.
  • the heavy chain variable region has the following complementarity determining regions CDR:
  • VH-CDR1 shown in SEQ ID NO: 1,
  • VH-CDR2 shown in SEQ ID NO: 2 or 7, and
  • VH-CDR3 shown in SEQ ID NO:3;
  • the light chain variable region has the following complementarity determining regions (CDRs):
  • VL-CDR1 shown in SEQ ID NO:4,
  • VL-CDR2 shown in SEQ ID NO:5, and
  • VL-CDR3 shown in SEQ ID NO:6.
  • the CD3 targeting binding domain has a heavy chain variable region with an amino acid sequence as shown in SEQ ID No: 20 or 22, and/or a light chain variable region with an amino acid sequence as shown in SEQ ID No: 21.
  • the CD3 targeting binding domain further includes a light chain constant region and/or a heavy chain constant region.
  • the CD3 targeting binding domain is an anti-CD3 single-chain antibody or Fab.
  • the CD3 targeting binding domain is Fab, which includes a light chain constant region CL and a heavy chain constant region CH1.
  • the constant region is a human constant region or a mouse constant region.
  • the light chain constant region CL has a sequence as shown in SEQ ID NO:36.
  • the heavy chain constant region CH1 has a sequence as shown in SEQ ID NO:35.
  • the CD3 targeting binding domain is the anti-CD3 antibody or antigen-binding fragment thereof as described in the second aspect of the present invention.
  • the CD19 targeting binding domain is connected to the N-terminus of the heavy chain variable region of the CD3 targeting binding domain; and the BCMA targeting binding domain is connected to the N-terminus of the light chain variable region of the CD3 targeting binding domain.
  • the CD19 targeting binding domain is connected to the N-terminus of the light chain variable region of the CD3 targeting binding domain; and the BCMA targeting binding domain is connected to the N-terminus of the heavy chain variable region of the CD3 targeting binding domain.
  • the light chain and/or heavy chain of the trispecific antibody further comprises an Fc domain.
  • the Fc domain is the Fc domain from IgG protein.
  • the trispecific antibody has a structure as shown in Formula IIIa, IIIb, IIIc, IIId or IIIe:
  • “-” is each independently a bond or a peptide linker
  • ScFv CD19 is the CD19 targeting binding domain
  • ScFv BCMA is the BCMA targeting binding domain
  • ScFv CD3 is the CD3 targeting binding domain
  • VH CD3 is the heavy chain variable region of the CD3 targeting binding domain
  • VL CD3 is the light chain variable region of the CD3 targeting binding domain
  • VH BCMA is the heavy chain variable region of the BCMA targeting binding domain
  • VL BCMA is the light chain variable region of the BCMA targeting binding domain
  • VH CD19 is the heavy chain variable region of the CD19 targeting binding domain
  • VL CD19 is the light chain variable region of the CD19 targeting binding domain
  • CH1 is the heavy chain constant region 1;
  • CL is the light chain constant region
  • Fc1 and Fc2 are each independently free or Fc domain
  • L3 and L4 are each independently a connecting peptide
  • the trispecific antibody has the structure shown in IIIa or IIIb, and Fc1 and Fc2 are absent.
  • L3 and L4 are flexible connecting peptides.
  • L3 and L4 each independently have an amino acid sequence as shown in (GGGGS)n or GG, wherein n is an integer of 1-6, preferably 1, 2, or 3.
  • L3 has an amino acid sequence as shown in SEQ ID NO:33.
  • L4 has an amino acid sequence as shown in SEQ ID NO:33.
  • Fc1 has an amino acid sequence as shown in SEQ ID NO:46.
  • Fc2 has an amino acid sequence as shown in SEQ ID NO:47.
  • the CD19 targeting binding domain includes a heavy chain variable region as shown in SEQ ID NO: 23 and a light chain variable region as shown in SEQ ID NO: 24.
  • the BCMA targeting binding domain includes a heavy chain variable region as shown in SEQ ID NO: 25 and a light chain variable region as shown in SEQ ID NO: 26.
  • the CD3 targeting binding domain includes a heavy chain variable region as shown in SEQ ID NO: 20 and a light chain variable region as shown in SEQ ID NO: 21.
  • the CD3 targeting binding domain includes a heavy chain variable region as shown in SEQ ID NO: 22 and a light chain variable region as shown in SEQ ID NO: 21.
  • the heavy chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO: 27 or 31, or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto;
  • the light chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO:28 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • the heavy chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO: 29 or 32, or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto;
  • the light chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO:30 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • the heavy chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO: 39 or 45, or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto;
  • the light chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO:40 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • the heavy chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO:41 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto;
  • the light chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO:42 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • the heavy chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO: 43 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto;
  • the light chain amino acid sequence of the trispecific antibody is as shown in SEQ ID NO:44 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • the trispecific antibody has a heavy chain and a light chain selected from the group consisting of:
  • the heavy chain amino acid sequence of the trispecific antibody is shown as SEQ ID NO:31; and the light chain amino acid sequence of the trispecific antibody is shown as SEQ ID NO:28.
  • the heavy chain amino acid sequence of the trispecific antibody is shown as SEQ ID NO:45; and the light chain amino acid sequence of the trispecific antibody is shown as SEQ ID NO:40.
  • the multispecific antibody is a bispecific antibody.
  • the bispecific antibody comprises:
  • a BCMA targeting binding domain comprising one or more antigen binding domains that specifically bind to a BCMA protein
  • CD3 targeting binding domain which comprises one or more antigen binding domains that specifically bind to CD3 protein.
  • the N-terminus of the heavy chain variable region and/or light chain variable region of the CD3 targeting binding domain is connected to the BCMA targeting binding domain, preferably through a flexible connecting peptide.
  • the connecting peptide has an amino acid sequence as shown in (GGGGS)n or GG, wherein n is an integer of 1-6, preferably 1, 2, or 3.
  • the bispecific antibody comprises:
  • a CD19 targeting binding domain comprising one or more antigen binding domains that specifically bind to the CD19 protein
  • the bispecific antibody has a structure as shown in Formula IIIf, IIIg, or IIIh:
  • “-” is each independently a bond or a peptide linker
  • ScFv CD19 is the CD19 targeting binding domain
  • ScFv BCMA is the BCMA targeting binding domain
  • VH CD3 is the heavy chain variable region of the CD3 targeting binding domain
  • VL CD3 is the light chain variable region of the CD3 targeting binding domain
  • CH1 is the heavy chain constant region 1;
  • CL is the light chain constant region
  • the heavy chain amino acid sequence of the bispecific antibody is as shown in SEQ ID NO: 29 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto;
  • the light chain amino acid sequence of the bispecific antibody is as shown in SEQ ID NO: 28 or has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence homology thereto.
  • the heavy chain amino acid sequence of the bispecific antibody is shown in SEQ ID NO: 29;
  • the light chain amino acid sequence of the bispecific antibody is shown in SEQ ID NO:28.
  • the heavy chain amino acid sequence of the bispecific antibody is shown in SEQ ID NO: 31;
  • the light chain amino acid sequence of the bispecific antibody is shown in SEQ ID NO:37.
  • the heavy chain amino acid sequence of the bispecific antibody is shown in SEQ ID NO: 38;
  • the light chain amino acid sequence of the bispecific antibody is shown in SEQ ID NO:28.
  • the EC50 of the antibody binding to target cells expressing CD19 and BCMA is ⁇ 1 ⁇ 10 ⁇ 10 M, preferably ⁇ 6 ⁇ 10 ⁇ 11 M, and more preferably ⁇ 2 ⁇ 10 ⁇ 11 M.
  • a recombinant protein comprising:
  • the tag sequence includes a 6His tag, a GGGS sequence, and a FLAG tag.
  • the recombinant protein is a fusion protein.
  • the recombinant protein is a monomer, a dimer, or a multimer.
  • a polynucleotide which encodes the anti-BCMA antibody or antigen-binding fragment thereof as described in the first aspect of the present invention, the anti-CD3 antibody or antigen-binding fragment thereof as described in the second aspect of the present invention, or the multispecific antibody as described in the third aspect of the present invention.
  • a vector is provided, wherein the vector contains the polynucleotide according to the fifth aspect of the present invention.
  • the expression vector is a viral vector, such as a lentivirus, adenovirus, AAV virus, retrovirus, or a combination thereof.
  • a genetically engineered host cell wherein the host cell contains the vector as described in the sixth aspect of the present invention, or an exogenous polynucleotide as described in the fifth aspect of the present invention integrated into its genome.
  • the host cell includes a prokaryotic cell or a eukaryotic cell.
  • the host cell is selected from the group consisting of Escherichia coli, yeast cells, and mammalian cells.
  • an antibody conjugate comprising:
  • conjugated moiety conjugated to the antibody portion, wherein the conjugated moiety is selected from the group consisting of a detectable label, a drug, or a combination thereof.
  • the detectable marker comprises a radionuclide.
  • the drugs include toxins, cytokines, and enzymes.
  • the conjugate is selected from: fluorescent or luminescent markers, radioactive markers, MRI (magnetic resonance imaging) or CT (computer tomography) contrast agents, or enzymes capable of producing detectable products, radionuclides, biotoxins, cytokines (such as IL-2, etc.), antibodies, antibody Fc fragments, antibody scFv fragments, gold nanoparticles/nanorods, viral particles, liposomes, nanomagnetic particles, prodrug-activating enzymes (for example, DT-diaphorase (DTD) or biphenyl hydrolase-like protein (BPHL)), chemotherapeutic agents (for example, cisplatin) or any form of nanoparticles, etc.
  • fluorescent or luminescent markers for example, radioactive markers, MRI (magnetic resonance imaging) or CT (computer tomography) contrast agents, or enzymes capable of producing detectable products, radionuclides, biotoxins, cytokines (such as IL-2, etc.), antibodies, antibody Fc fragment
  • the antibody portion and the coupling portion are coupled via a chemical bond or a linker.
  • the disease is cancer or tumor.
  • the cancer or tumor is a solid tumor or a hematological tumor.
  • the cancer or tumor is selected from the group consisting of multiple myeloma, refractory or relapsed B-cell acute lymphoblastic leukemia (B-ALL), non-Hodgkin's lymphoma (NHL), or a combination thereof.
  • B-ALL B-cell acute lymphoblastic leukemia
  • NHL non-Hodgkin's lymphoma
  • a pharmaceutical composition comprising:
  • an active ingredient selected from the group consisting of the anti-BCMA antibody or antigen-binding fragment thereof according to the first aspect of the present invention, the anti-CD3 antibody or antigen-binding fragment thereof according to the second aspect of the present invention, the multispecific antibody according to the third aspect of the present invention, the recombinant protein according to the fourth aspect of the present invention, the antibody conjugate according to the eighth aspect of the present invention, or a combination thereof; and
  • the pharmaceutical composition is a liquid preparation.
  • the pharmaceutical composition is an injection.
  • the pharmaceutical composition is used to prevent and/or treat diseases associated with high expression of CD19 and/or BCMA.
  • Figure 1 shows the structures of trispecific antibody molecules (IMP-2023-031, IMP-2023-032, IMP-2023-033, IMP-2023-034), bispecific molecules (IMP-2023-035), control molecules (IMC-035 and IMC-036), and long-acting molecules (IMP-2023-036, IMP-2023-037, IMP-2023-038, IMP-2023-039).
  • FIG2 shows a heavy chain transient expression vector and a light chain transient expression vector.
  • Figure 3A-D shows cell binding curves.
  • A is a T cell binding curve;
  • B-D are NCI-H929, K562-CD19, and Daudi cell binding curves, respectively.
  • Figure 4 shows CD19 and BCMA expression.
  • Figures 5A-J show target cell killing curves of IMP-2023-033.
  • the effector cells were hPBMCs, the target cells were Raji, and the E:T ratio was 10:1; in Figure F, the effector cells were hPBMCs T cells, the target cells were hPBMCs B cells, and the incubation time was 24 hours. In the remaining experiments, the effector cells were Pri-T cells, the E:T ratio was 4:1, and the incubation time was 24 hours.
  • Figures 6A-B show target cell killing curves for IMP-2023-033 and control molecules.
  • A) Effector cells are T cells, target cells are K562-BCMA, E:T ratio is 4:1, and incubation time is 24 hours.
  • B) Effector cells are T cells, target cells are K562-CD19, E:T ratio is 4:1, and incubation time is 24 hours.
  • FIG8 shows the results of IMP-2023-033 stimulating T cell proliferation.
  • Figure 12 shows the results of IMP-2023-induced target cell killing.
  • the effector cells are T cells.
  • the target cells in Figures A, B, and C are NCI-H929 expressing BCMA, K562-CD19 expressing CD19, and Daudi expressing both BCMA and CD19, respectively.
  • the E:T ratio is 4:1, and the incubation time is 24 hours.
  • FIG13 shows the T cell proliferation results.
  • FIG14 shows the results of dexamethasone inhibition of cytokine release.
  • This multispecific antigen-binding protein is composed of an anti-BCMA scFv, an anti-CD19 scFv, and an anti-CD3 Fab linked together.
  • the multispecific antigen-binding protein binds BCMA, CD19, and CD3 simultaneously with high affinity, and can function as a T cell adaptor, connecting target cells and T cells, inducing T cell activation and proliferation, and promoting T cell cytotoxicity. This is the basis for the present invention.
  • treatment refers to administering an internal or external therapeutic agent, including antibodies against respiratory syncytial virus fusion protein (preferably pre-fusion F protein) and compositions thereof, to a patient experiencing one or more symptoms of a disease for which the therapeutic agent is known to have a therapeutic effect.
  • the therapeutic agent is administered to the patient in an amount effective to alleviate one or more symptoms of the disease (a therapeutically effective amount).
  • sequence identity refers to the degree of identity between two nucleic acid or amino acid sequences when optimally aligned and compared with appropriate mutations such as substitutions, insertions, or deletions.
  • sequence identity between a sequence described herein and a sequence to which it is identical may be at least 85%, 90%, or 95%, preferably at least 95%. Non-limiting examples include 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, and 100%.
  • antibody or "immunoglobulin” are heterotetrameric glycoproteins of approximately 150,000 daltons with identical structural features, consisting of two identical light chains (L) and two identical heavy chains (H). Each light chain is linked to a heavy chain by one covalent disulfide bond, while the number of disulfide bonds varies between heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bonds. Each heavy chain has a variable region (VH) at one end, followed by multiple constant regions.
  • VH variable region
  • Each light chain has a variable region (VL) at one end and a constant region at the other end; the constant region of the light chain is opposite the first constant region of the heavy chain, and the variable region of the light chain is opposite the variable region of the heavy chain.
  • VL variable region
  • Specific amino acid residues form an interface between the variable regions of the light and heavy chains.
  • variable refers to certain parts of the variable region of an antibody that differ in sequence, which contributes to the binding and specificity of each specific antibody for its specific antigen. However, variability is not evenly distributed throughout the variable region of an antibody. It is concentrated in three segments called complementarity determining regions (CDRs) or hypervariable regions in the variable regions of the light and heavy chains. The more conserved parts of the variable region are called framework regions (FRs).
  • CDRs complementarity determining regions
  • FRs framework regions
  • the variable regions of natural heavy and light chains each contain four FR regions, which are generally in a ⁇ -sheet configuration and are connected by three CDRs that form a connecting loop, which in some cases can form a partial ⁇ -sheet structure.
  • the CDRs in each chain are closely together through the FR region and together with the CDRs of the other chain form the antigen-binding site of the antibody (see Kabat et al., NIH Publ. No. 91-3242, Vol. I, pp. 647-669 (1991)).
  • the constant regions are not directly involved in the binding of the antibody to the antigen, but they exhibit different effector functions, such as participation in the antibody's antibody-dependent cellular toxicity.
  • variable regions Generally, an antibody's antigen-binding properties are described by three specific regions located in the variable regions of the heavy and light chains, known as the variable regions (CDRs). These regions are divided into four framework regions (FRs). The amino acid sequences of the four FRs are relatively conserved and do not directly participate in the binding reaction. These CDRs form a ring structure, spatially close to each other through the ⁇ -sheet formed by the FRs between them. The CDRs on the heavy chain and the corresponding CDRs on the light chain constitute the antibody's antigen-binding site. The amino acid sequences of antibodies of the same type can be compared to determine which amino acids constitute the FR or CDR regions.
  • antibody fragment refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen. It has been shown that fragments of full-length antibodies can be used to perform the antigen-binding function of an antibody.
  • binding fragments encompassed by the term "antigen-binding fragment of an antibody” include (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL, and CH1 domains; (ii) a F(ab')2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge on the hinge region; (iii) an Fd fragment consisting of the VH and CH1 domains; and (iv) an Fv fragment consisting of the VH and VL domains of a single arm of an antibody.
  • Fv antibodies are the smallest antibody fragments that contain the variable regions of the heavy and light chains of an antibody, but lack the constant region, and possess all the antigen-binding sites.
  • Fv antibodies also contain a polypeptide linker between the VH and VL domains, and are capable of forming the structure required for antigen binding.
  • the present invention includes not only complete monoclonal antibodies, but also antibody fragments with immunological activity, such as Fab or (Fab') 2 fragments; antibody heavy chains; and antibody light chains.
  • epitope refers to a site on an antigen to which an immunoglobulin or antibody specifically binds.
  • An epitope typically comprises at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 contiguous or non-contiguous amino acids in a unique spatial conformation.
  • An epitope can be a discontinuous three-dimensional site on an antigen that is recognized by the antibodies or antigen-binding fragments of the present invention.
  • binding refers to the binding of an antibody to a predetermined epitope on an antigen.
  • the antibody binds with an affinity (KD) of less than about 10-7 M, such as less than about 10-8 M, 10-9 M, or 10-10 M or less.
  • the present invention includes not only complete antibodies, but also fragments of antibodies with immunological activity or fusion proteins formed by antibodies and other sequences. Therefore, the present invention also includes fragments, derivatives and analogs of the antibodies.
  • antibodies include murine, chimeric, humanized or fully human antibodies prepared using techniques well known to those skilled in the art.
  • Recombinant antibodies such as chimeric and humanized monoclonal antibodies, including human and non-human parts, can be obtained by standard DNA recombinant techniques, and they are all useful antibodies.
  • a chimeric antibody is a molecule in which different parts are derived from different animal species, such as a chimeric antibody having a variable region from a mouse monoclonal antibody and a constant region from a human immunoglobulin (see, for example, U.S. Patent No. 4,816,567 and U.S. Patent No. 4,816,397, which are hereby incorporated by reference in their entirety).
  • a humanized antibody refers to an antibody molecule derived from a non-human species, having one or more complementary determining regions (CDRs) derived from a non-human species and a framework region derived from a human immunoglobulin molecule (see U.S. Patent No. 5,585,089, which is hereby incorporated by reference in its entirety).
  • CDRs complementary determining regions
  • These chimeric and humanized monoclonal antibodies can be prepared using DNA recombinant techniques well known in the art.
  • the antibodies can be monospecific, bispecific, trispecific, or more multispecific.
  • CDR refers to one of the six hypervariable regions within the variable domain of an antibody that primarily contributes to antigen binding.
  • One of the most commonly used definitions of the six CDRs is provided by Kabat E.A. et al. (1991) Sequences of proteins of immunological interest. NIH Publication 91-3242.
  • heavy chain constant region includes an amino acid sequence derived from an immunoglobulin heavy chain.
  • a polypeptide comprising a heavy chain constant region includes at least one of the following: a CH1 domain, a hinge region (e.g., an upper, middle, and/or lower hinge region) domain, a CH2 domain, a CH3 domain, or a variant or fragment thereof.
  • the term "light chain constant region” includes an amino acid sequence derived from an antibody light chain.
  • the light chain constant region includes at least one of a constant kappa domain or a constant lambda domain.
  • Fab refers to a monovalent fragment consisting of the VL, VH, CL and CH1 domains, which form a dimer via a disulfide bond between the CL domain of the light chain and the CH1 domain of the heavy chain.
  • multispecific antibody and “multispecific antigen-binding protein” are used interchangeably to refer to molecules that are able to simultaneously bind to multiple different types of epitopes, which can be located on the same target cell or on different target cells.
  • the present invention provides a bispecific antibody (also known as a bispecific T cell engager) that simultaneously targets BCMA and CD3.
  • the present invention provides a trispecific antibody (also known as a trispecific T cell engager) that simultaneously targets CD19, BCMA, and CD3. Its advantage is that it can redirect specific polyclonal immune cells (such as T cells and NK cells) to tumor cells to enhance tumor killing.
  • the antibodies of the present invention comprise antigen-binding fragments from two or more (e.g., three) monoclonal antibodies and rely on their antigen-binding ability to exert their effects.
  • the antibodies of the present invention comprise Fab fragments from anti-CD3 antibodies.
  • the antibodies of the present invention comprise scFv from anti-BCMA antibodies.
  • the antibodies of the present invention comprise scFv from anti-CD19 antibodies.
  • Bispecific or trispecific antibodies can be produced by chemical crosslinking or hybridoma technology.
  • bispecific or trispecific antibody molecules can be produced by recombinant technology, for example, by linking one or more scFv molecules to Fab molecules via a connecting peptide.
  • the length of the connecting peptide is between 2 and 15 amino acids.
  • the connecting peptide can be composed of multiple amino acids, such as repeating units of GGGGS.
  • the trispecific antibodies of the present invention may further comprise an Fc domain, thereby extending the antibody half-life and forming a long-acting molecule.
  • the Fc domain is derived from an IgG protein.
  • the Fc domain is linked to the C-terminus of the CH1 domain and the CL domain of the Fab fragment.
  • the multispecific antibody has a structure as shown in any one of the following formulae IIIa-IIIh:
  • “-” is each independently a bond or a peptide linker
  • ScFv CD19 is the CD19 targeting binding domain
  • ScFv BCMA is the BCMA targeting binding domain
  • ScFv CD3 is the CD3 targeting binding domain
  • VH CD3 is the heavy chain variable region of the CD3 targeting binding domain
  • VL CD3 is the light chain variable region of the CD3 targeting binding domain
  • VH BCMA is the heavy chain variable region of the BCMA targeting binding domain
  • VL BCMA is the light chain variable region of the BCMA targeting binding domain
  • VH CD19 is the heavy chain variable region of the CD19 targeting binding domain
  • Fc1 and Fc2 are each independently free or Fc domain
  • the present invention also provides a nucleic acid encoding the above-mentioned antibody or recombinant protein of the present invention or a chimeric antigen receptor (CAR) construct of the antibody of the present invention.
  • CAR chimeric antigen receptor
  • the base sequence encoding the amino acid sequence of the aforementioned protein can be appropriately substituted, deleted, altered, inserted, or added to provide a polynucleotide homolog.
  • the polynucleotide homologs of the present invention can be prepared by substituting, deleting, or adding one or more bases in the gene encoding the protein sequence while maintaining antibody activity.
  • the present invention also provides polynucleotide molecules encoding the above-mentioned antibodies or fragments thereof.
  • the polynucleotides of the present invention may be in the form of DNA or RNA.
  • DNA forms include cDNA, genomic DNA, or artificially synthesized DNA.
  • DNA may be single-stranded or double-stranded.
  • DNA may be a coding strand or a non-coding strand.
  • the coding region sequence encoding the mature polypeptide may be identical to the coding region sequence of the antibody of the present invention or a degenerate variant.
  • degenerate variant in the present invention refers to a nucleic acid sequence encoding an amino acid sequence identical to that of the polypeptide of the present invention, but having a different coding region sequence.
  • Polynucleotides encoding mature polypeptides of the present invention include: coding sequences encoding only the mature polypeptide; coding sequences for the mature polypeptide and various additional coding sequences; coding sequences for the mature polypeptide (and optional additional coding sequences) and non-coding sequences.
  • the term "polynucleotide encoding a polypeptide" may include a polynucleotide encoding the polypeptide or a polynucleotide further including additional coding and/or non-coding sequences.
  • the present invention also relates to polynucleotides that hybridize with the above-mentioned sequences and have at least 50%, preferably at least 70%, and more preferably at least 80% identity between the two sequences.
  • the present invention particularly relates to polynucleotides that can hybridize with the polynucleotides of the present invention under stringent conditions.
  • stringent conditions refer to: (1) hybridization and elution at relatively low ionic strength and relatively high temperature, such as 0.2 ⁇ SSC, 0.1% SDS, 60°C; or (2) the addition of a denaturing agent during hybridization, such as 50% (v/v) formamide, 0.1% calf serum/0.1% Ficoll, 42°C, etc.; or (3) hybridization only occurs when the identity between the two sequences is at least 90%, and more preferably at least 95%.
  • the polypeptide encoded by the hybridizable polynucleotide has the same biological function and activity as the mature polypeptide constituting the antibody of the present invention.
  • the full-length nucleotide sequence of the antibody of the present invention or its fragments can generally be obtained by PCR amplification, recombinant methods, or artificial synthesis methods.
  • One feasible method is to synthesize the relevant sequence by artificial synthesis, especially when the fragment length is relatively short. Generally, by first synthesizing multiple small fragments and then ligating them, very long fragments of sequence can be obtained.
  • the coding sequence of the heavy chain can be fused with an expression tag (such as 6His) to form a fusion protein.
  • sequence of the DNA molecule of the antibody or fragment thereof of the present invention can be obtained using conventional techniques, such as PCR amplification or genomic library screening.
  • the relevant sequence can be obtained in large quantities by recombinant methods. This is usually done by cloning it into a vector, then transferring it into cells, and then isolating the relevant sequence from the propagated host cells by conventional methods.
  • sequences can also be synthesized by artificial synthesis, especially when the fragment length is shorter.
  • a long fragment can be obtained by synthesizing multiple small fragments and then connecting them.
  • DNA sequences encoding the antibodies (or fragments thereof, or derivatives thereof) of the present invention can be obtained entirely through chemical synthesis. This DNA sequence can then be introduced into various existing DNA molecules (or vectors) and cells known in the art. In addition, mutations can also be introduced into the protein sequences of the present invention through chemical synthesis.
  • the present invention also relates to vectors comprising the above-mentioned appropriate DNA sequence and appropriate promoter or control sequence. These vectors can be used to transform appropriate host cells to enable them to express proteins.
  • the host cell can be a prokaryotic cell, such as a bacterial cell, a lower eukaryotic cell, such as a yeast cell, or a higher eukaryotic cell, such as a mammalian cell.
  • Preferred animal cells include (but are not limited to): CHO-S and HEK-293 cells.
  • the transformed host cells are cultured under conditions suitable for expression of the antibodies of the present invention.
  • the antibodies of the present invention are then purified using conventional immunoglobulin purification procedures, such as protein A-Sepharose, hydroxyapatite chromatography, gel electrophoresis, dialysis, ion exchange chromatography, hydrophobic chromatography, molecular sieve chromatography, or affinity chromatography, among other conventional separation and purification methods well known to those skilled in the art.
  • the antibodies of the present invention can be expressed intracellularly, on the cell membrane, or secreted extracellularly. If necessary, the recombinant protein can be separated and purified by various separation methods utilizing its physical, chemical, and other properties. These methods are well known to those skilled in the art. Examples of these methods include, but are not limited to, conventional renaturation treatment, treatment with a protein precipitant (salting out method), centrifugation, osmotic shock, ultrasonic treatment, ultracentrifugation, molecular sieve chromatography (gel filtration), adsorption chromatography, ion exchange chromatography, high performance liquid chromatography (HPLC), and various other liquid chromatography techniques and combinations of these methods.
  • conventional renaturation treatment treatment with a protein precipitant (salting out method), centrifugation, osmotic shock, ultrasonic treatment, ultracentrifugation, molecular sieve chromatography (gel filtration), adsorption chromatography, ion exchange chromatography,
  • the present invention also provides an antibody-drug conjugate (ADC) based on the antibody of the present invention.
  • ADC antibody-drug conjugate
  • the antibody-drug conjugate comprises the antibody and an effector molecule, wherein the antibody is conjugated to the effector molecule, preferably chemically conjugated.
  • the effector molecule is preferably a therapeutically active drug.
  • the effector molecule may be one or more of a toxic protein, a chemotherapeutic drug, a small molecule drug, or a radionuclide.
  • the antibody of the present invention and the effector molecule can be coupled via a coupling agent.
  • the coupling agent may include any one or more of a non-selective coupling agent, a coupling agent utilizing a carboxyl group, a peptide chain, and a coupling agent utilizing a disulfide bond.
  • the non-selective coupling agent refers to a compound that forms a covalent bond between the effector molecule and the antibody, such as glutaraldehyde.
  • the coupling agent utilizing a carboxyl group may include any one or more of a cis-aconitic anhydride coupling agent (such as cis-aconitic anhydride) and an acylhydrazone coupling agent (where the coupling site is an acylhydrazone).
  • a cis-aconitic anhydride coupling agent such as cis-aconitic anhydride
  • an acylhydrazone coupling agent where the coupling site is an acylhydrazone
  • antibodies are used to connect to a variety of functional groups, including imaging agents (such as chromophores and fluorescent groups), diagnostic agents (such as MRI contrast agents and radioisotopes), stabilizers (such as ethylene glycol polymers) and therapeutic agents.
  • imaging agents such as chromophores and fluorescent groups
  • diagnostic agents such as MRI contrast agents and radioisotopes
  • stabilizers such as ethylene glycol polymers
  • therapeutic agents such as ethylene glycol polymers
  • Antibodies can be coupled to functional agents to form antibody-functional agent conjugates.
  • Functional agents such as drugs, detection reagents, stabilizers
  • Functional agents can be directly or indirectly connected to antibodies through linkers.
  • Antibodies can be conjugated to drugs to form antibody-drug conjugates (ADCs).
  • ADCs contain a linker positioned between the drug and the antibody.
  • the linker can be degradable or non-degradable.
  • Degradable linkers typically readily degrade in the intracellular environment, for example, at the target site, thereby releasing the drug from the antibody.
  • Suitable degradable linkers include, for example, enzymatically degradable linkers, including linkers containing peptidyl groups that can be degraded by intracellular proteases (e.g., lysosomal proteases or endosomal proteases), or sugar linkers, such as glucuronide-containing linkers that can be degraded by glucuronidases.
  • Peptide linkers can include, for example, dipeptides such as valine-citrulline, phenylalanine-lysine, or valine-alanine.
  • Other suitable degradable linkers include, for example, pH-sensitive linkers (e.g., linkers that hydrolyze at a pH below 5.5, such as hydrazone linkers) and linkers that degrade under reducing conditions (e.g., disulfide linkers).
  • Non-degradable linkers typically release the drug when the antibody is hydrolyzed by proteases.
  • the linker Prior to attachment to the antibody, the linker has an active reactive group capable of reacting with certain amino acid residues, and attachment is achieved via the active reactive group.
  • Thiol-specific active reactive groups are preferred and include, for example, maleimides, haloamides (e.g., iodinated, brominated, or chlorinated); haloesters (e.g., iodinated, brominated, or chlorinated); halomethylketones (e.g., iodinated, brominated, or chlorinated); benzyl halides (e.g., iodinated, brominated, or chlorinated); vinyl sulfones, pyridyl disulfides; mercury derivatives such as 3,6-di-(mercurymethyl)dioxane, where the counter ion is acetate, chloride, or nitrate; and polymethylene dimethyl sulfide thiosulfonate.
  • Useful drug classes include, for example, anti-tubulin drugs, DNA minor groove binding agents, DNA replication inhibitors, alkylating agents, antibiotics, folate antagonists, antimetabolites, chemosensitizers, topoisomerase inhibitors, vinca alkaloids, and the like.
  • particularly useful cytotoxic drugs include, for example, DNA minor groove binding agents, DNA alkylating agents, and tubulin inhibitors.
  • Typical cytotoxic drugs include, for example, auristatins, camptothecins, duocarmycins, etoposides, maytansines and maytansinoids (e.g., DM1 and DM4), taxanes, benzodiazepines or benzodiazepine-containing drugs (e.g., pyrrolo[1,4]benzodiazepines (PBDs), indolinobenzodiazepines, and oxazolidinobenzodiazepines), and vinca alkaloids.
  • auristatins camptothecins, duocarmycins, etoposides
  • maytansines and maytansinoids e.g., DM1 and DM4
  • taxanes e.g., benzodiazepines or benzodiazepine-containing drugs (e.g., pyrrolo[1,4]benzodiazepines (PBDs), indolinobenzodiazepines, and oxazolid
  • drug-linkers can be used to form ADCs in a single step.
  • bifunctional linker compounds can be used to form ADCs in a two-step or multi-step process. For example, a cysteine residue is reacted with a reactive moiety of a linker in a first step, and in a subsequent step, the functional group on the linker reacts with the drug to form an ADC.
  • Other useful functional groups include, for example, ketones and aldehydes (suitable for reaction with hydrazides and alkoxyamines), phosphines (suitable for reaction with azides); isocyanates and isothiocyanates (suitable for reaction with amines and alcohols); and activated esters, such as N-hydroxysuccinimide esters (suitable for reaction with amines and alcohols).
  • ketones and aldehydes suitable for reaction with hydrazides and alkoxyamines
  • phosphines suitable for reaction with azides
  • isocyanates and isothiocyanates suitable for reaction with amines and alcohols
  • activated esters such as N-hydroxysuccinimide esters (suitable for reaction with amines and alcohols).
  • the methods of the present invention comprise conjugating an antibody to a bifunctional linker compound under conditions sufficient to form an antibody-linker conjugate. In these embodiments, the methods of the present invention further comprise conjugating the antibody-linker conjugate to a drug moiety under conditions sufficient to covalently attach the drug moiety to the antibody via the linker.
  • the present invention also provides a composition.
  • the composition is a pharmaceutical composition comprising the above-mentioned antibody, active fragment thereof, fusion protein thereof, or ADC, and a pharmaceutically acceptable carrier.
  • these substances are formulated in a non-toxic, inert, and pharmaceutically acceptable aqueous carrier medium, wherein the pH is generally about 5-8, preferably about 6-8, although the pH value may vary depending on the nature of the substance being formulated and the condition to be treated.
  • the pharmaceutical composition of the present invention is used to prevent and/or treat diseases associated with high expression of CD19 and/or BCMA.
  • the pharmaceutical composition of the present invention is used to prevent and/or treat cancers or tumors with high expression of CD19 and/or BCMA.
  • the pharmaceutical composition of the present invention contains a safe and effective amount (e.g., 0.001-99 wt%, preferably 0.01-90 wt%, more preferably 0.1-80 wt%) of the above-mentioned monoclonal antibody of the present invention (or its conjugate) and a pharmaceutically acceptable carrier or excipient.
  • a pharmaceutically acceptable carrier or excipient include (but are not limited to): saline, buffer, glucose, water, glycerol, ethanol, and combinations thereof.
  • the pharmaceutical preparation should match the mode of administration.
  • the pharmaceutical composition of the present invention can be prepared in the form of an injection, for example, using physiological saline or an aqueous solution containing glucose and other adjuvants by conventional methods.
  • compositions such as injections and solutions are preferably manufactured under sterile conditions.
  • the dosage of the active ingredient is a therapeutically effective amount, for example, about 1 ⁇ g/kg body weight to about 5 mg/kg body weight per day.
  • the polypeptide of the present invention can also be used in conjunction with other therapeutic agents.
  • the multispecific antibodies of the present invention can be used together with a glucocorticoid to inhibit cytokine release without affecting target cell killing activity. Therefore, the pharmaceutical composition of the present invention can comprise the multispecific antibodies of the present invention and a glucocorticoid (e.g., dexamethasone).
  • a glucocorticoid e.g., dexamethasone
  • the pharmaceutical composition of the present invention further comprises one or more pharmaceutical carriers.
  • the pharmaceutical carrier is a conventional pharmaceutical carrier in the art, and the pharmaceutical carrier can be any suitable physiologically or pharmaceutically acceptable pharmaceutical excipient.
  • the pharmaceutical excipient is a conventional pharmaceutical excipient in the art, and preferably includes a pharmaceutically acceptable excipient, filler or diluent, etc. More preferably, the pharmaceutical composition comprises 0.01 to 99.99% of the above-mentioned protein and 0.01 to 99.99% of a pharmaceutical carrier, and the percentages are the mass percentages of the pharmaceutical composition.
  • a safe and effective amount of the immunoconjugate is administered to a mammal, wherein the safe and effective amount is generally at least about 10 ⁇ g/kg body weight, and in most cases does not exceed about 50 mg/kg body weight.
  • the dose is about 10 ⁇ g/kg body weight to about 20 mg/kg body weight.
  • the specific dose should also take into account factors such as the route of administration and the patient's health status, which are all within the skill of a skilled physician.
  • the multispecific T cell engager of the present invention can simultaneously target BCMA, CD19, and CD3 with high binding force, thereby connecting target cells and T cells and promoting T cell killing.
  • the multispecific T cell engager of the present invention can induce T cell activation and proliferation in human PBMCs.
  • the multispecific T cell engager of the present invention when used in combination with dexamethasone, can inhibit cytokine release without affecting the killing activity induced by the T cell engager, and has good safety.
  • the trispecific antibody molecule contains three antibody sequences that bind to different antigen targets (CD19, BCMA and CD3).
  • the three different target antibody sequences were amplified by overlap extension PCR (overlap PCR) to obtain the first peptide chain gene (heavy chain) and the second peptide chain gene (light chain), and both peptide chains carried their own Kozak sequence and signal peptide sequence.
  • the DNA fragments encoding the heavy and light chains of the trispecific antigen-binding protein were double-digested with restriction endonucleases (EcoR I and Not I) and cloned into a transient expression vector to obtain a transient expression vector expressing the heavy and light chains ( Figure 2).
  • the transient expression vector contains an ampicillin resistance gene and a strong promoter CMV gene, which can promote high-level expression of the target gene in eukaryotic cells.
  • the colonies were picked and sent for sequencing, and after sequence alignment, they were consistent with the theoretical sequence.
  • the positive clone plasmid was transformed into competent cells DH5a, and the selected clone was inoculated into 50 mL of E. coli culture medium and cultured overnight. The next day, the cells were collected and the transfection-grade plasmid was extracted. The transfection-grade plasmid was extracted according to the instructions of Plamid Plus Midi Kit (100) (QIAGEN, Cat No. 12945).
  • CHO-S cells were cultured and expanded using ExpiCHO Expression Medium (Gibco, Cat No. A29100). CHO-S cells were transiently transfected according to the instructions of the ExpiFectamine CHO Transfection Kit (Gibco, Cat No. A29129). The specific steps are as follows:
  • the supernatant was collected by centrifugation and filtered through a 0.22 ⁇ M filter.
  • the target protein was purified using an IgG-CH1 affinity chromatography column (Thermo Fisher Scientific).
  • the IgG-CH1 affinity column was equilibrated with phosphate buffer (10 mM sodium phosphate buffer + 50 mM sodium chloride, pH 7.2).
  • the cell culture supernatant was loaded onto the column, which was then rinsed with phosphate buffer (10 mM sodium phosphate buffer + 50 mM sodium chloride, pH 7.2).
  • the target protein was eluted with 50 mM NaAc buffer (pH 3.5). The pH of the eluate was adjusted to 4.5-5.5 using Tris buffer.
  • the three-target antigen affinity of IMP-2023-031 and IMP-2023-033 was detected using a Gator instrument using an anti-human IgG Fc GenII (HFCII) probe (Gator; Cat: 160024). All three antigen proteins are fusion proteins of the extracellular region of the protein and the human Fc (CD19/Fc was purchased from Acro Biosystems, and BCMA/Fc and CD3/Fc were purchased from Sino Biological). The antigen concentration was diluted to 2.5-5.0ug/mL with PBST buffer and immobilized on the anti-human IgG Fc probe.
  • HFCII anti-human IgG Fc GenII
  • the drug molecules to be tested were gradiently diluted into the wells of a 96-well black microplate and reacted with the antigen immobilized on the probe.
  • the test results were analyzed using Gator data analysis software.
  • the KD value for IMP-2023-031 binding to the CD3 antigen protein was 1.83 ⁇ 10 ⁇ 9 M, while the KD value for IMP-2023-033 binding to the CD3 antigen protein was 1.04 ⁇ 10 ⁇ 8 M, a difference of approximately 10-fold.
  • the KD value for IMP-2023-031 binding to the CD19 antigen protein was 5.49 ⁇ 10 ⁇ 9 M, while the KD value for IMP-2023-033 binding to the CD19 antigen protein was 5.36 ⁇ 10 ⁇ 9 M, indicating that both have similar binding abilities to the CD19 antigen protein.
  • the KD values for binding to the BCMA antigen protein were both within the order of 10 ⁇ 9 M.
  • T cell preparation Human peripheral blood mononuclear cells (hPBMCs) (purchased from OriBiotech) were activated for 3 days in the presence of 5 ⁇ g/mL CD3 antibody (eBioscience, 16-0037-85) and 1 ⁇ g/mL CD28 antibody (eBioscience, 16-0289-85), then incubated with 10 ng/mL IL-2 (PeproTech, 200-02) for 3 days, and the cells were collected and frozen. When necessary, the cells were revived and cultured in the presence of 10 ng/mL IL-2 for 1-3 days, and the cells were collected for later use.
  • the culture medium used in the experiment was RPMI-1640 (Life technologies) containing 10% fetal bovine serum (Life technologies), hereinafter referred to as complete culture medium.
  • test antibody contains a human lambda light chain
  • lambda(+)-positive cells represent cells that bind to the test antibody.
  • Data were analyzed using GraphPad Prism software, plotting T cell binding rate against drug concentration using a four-parameter curve fit. EC50 values were calculated. The results are shown in Table 2 and Figure 3.
  • the EC50 values of IMP-2023-031 or IMP-2023-033 binding to T cells (CD3+), NCI-H929 cells (BCMA+), K562-CD19 cells (CD19+), and Daudi cells (BCMA+CD19+) were in the range of 10-9 M, 10-10 M, and 10-11 M, respectively, indicating that the binding abilities of the two molecules were similar.
  • the cells used in this experiment included Raji, Su-DHL-4, Daudi, K562, K562-CD19, K562-CD20, K562-BCMA, hPBMCs, NCI-H929, and RPMI-8226. All cells were suspension culture.
  • the culture medium used was RPMI-1640 (Life Technologies) supplemented with 10% fetal bovine serum (Life Technologies), a complete medium.
  • the three cell lines K562-CD19, K562-CD20 and K562-BCMA are artificially constructed cells obtained by stably transfecting CD19, CD20 or BCMA into K562 cells, respectively.
  • the cells tested for protein expression were used as target cells, and the specific preparation was as described above.
  • tumor target cells were washed and centrifuged, then stained with CFSE (carboxyfluorescein succinimidyl amino ester) (eBioscience, 65-0850-85) at room temperature for 10 minutes. The cells were then washed and resuspended in complete culture medium. If the effector cells were T cells, the cells were seeded at a 4:1 effector cell:target cell ratio. If the effector cells were hPBMCs, the cells were seeded at a 10:1 effector cell:target cell ratio. For hPBMCs autologous B cell cytotoxicity assays, hPBMCs were directly plated. The molecule to be tested was added and incubated.
  • CFSE carboxyfluorescein succinimidyl amino ester
  • 7-AAD (BD, 51-68981E or Biolegend, 420404) was added to the corresponding wells of the 96-well plate. After staining for 15 minutes, the cells were analyzed by flow cytometry.
  • hPBMCs autologous B cell killing assay Before adding 7-AAD, add PE-anti-CD20 (Biolegend, 375503) and incubate at room temperature for 30-60 minutes. Finally, analyze the cells by flow cytometry.
  • CD20 + represents target cells
  • 7- AAD- represents viable cells
  • CD20 + 7- AAD- represents viable target cells.
  • Cytotoxicity can be calculated using the following formula:
  • GraphPad Prism software was used to analyze the data. The cell killing rate was used as the ordinate and the drug concentration was used as the abscissa. A four-parameter curve was fitted to plot the data and calculate the EC50 value.
  • the results are shown in Figure 5.
  • the target cells in Figures 5A-D are cells that express both CD19 and BCMA, and the EC 50 of IMP-2023-033 against them is between 0.045-0.844pM.
  • the target cells in Figures 5C-D are all Raji, and the effector cells are hPBMCs and T cells, respectively.
  • the results show that the killing effects of hPBMCs and T cells are basically the same.
  • the target cells in Figures 5E-F are cells that express CD19 but do not express BCMA, and the EC 50 of IMP-2023-033 against them is between 3.3-13pM, indicating that IMP-2023-033 can exert its effect through anti-CD19.
  • the target cells in Figures 5G-I are cells that express BCMA but do not express CD19, and the EC 50 of IMP-2023-033 against them is between 16-34pM, indicating that IMP-2023-033 can exert its effect through anti-BCMA.
  • the target cells in Figure 5J are K562-CD20, which express neither CD19 nor BCMA.
  • IMP-2023-033 has no killing activity against them, while the positive molecule IMP-2023-026 (anti-CD3 x anti-CD20) has killing activity against them. This shows that IMP-2023-033 mediates specific target cell killing, has no activity against cells that do not express the target protein, and has good safety.
  • the EC50 of IMP-2023-033 against target cells expressing only CD19 is between 3.3-13pM
  • the EC50 against target cells expressing only BCMA is between 16-34pM
  • the EC50 against target cells expressing both CD19 and BCMA is between 0.045-0.844pM, indicating that dual-expressing target cells are more easily killed by IMP-2023-033.
  • IMC-035 anti-CD3 x anti-CD19
  • IMC-036 anti-CD3 x anti-BCMA
  • IMP-2023-033 demonstrated high cytotoxicity against target cells expressing both antigens and reduced off-target activity compared to the two control molecules, IMC-035 and IMC-036.
  • hPBMCs were resuspended in complete culture medium and seeded into 96-well cell culture plates.
  • the detection antibody was diluted to different concentrations with complete culture medium, and the test antibody IMP-2023-033 or diluent was added to the corresponding wells.
  • the 96-well cell culture plate containing cells was placed in a CO2 incubator and incubated at 37°C.
  • APC-anti-CD4 antibody (BD Pharmingen, 551980), PE-anti-CD8 antibody (Biolegend, 301008) and FITC-anti-CD69 antibody (BD Pharmingen, 555530) were added and incubated at room temperature for 30-60 minutes.
  • the cells were analyzed using a flow cytometer to calculate the proportion of CD69-positive cells in the CD4 + T or CD8 + T subsets.
  • CD69 is a marker of T cell activation. This study examined CD69 expression on T cells to confirm whether IMP-2023-033 mediates T cell activation.
  • Graphpad Prism software was used to perform four-parameter equation fitting analysis on the flow cytometry data, yielding the dose-response curve and the half-maximal effective concentration (EC 50 ) , as shown in Figure 7.
  • the experimental results showed that IMP-2023-033 could induce CD69 expression in CD4 + T cells or CD8 + T cells in a concentration-dependent manner, with EC 50 values of 26 and 29 pM, respectively.
  • IMP-2023-033 can induce T cell activation in human PBMCs in a concentration-dependent manner.
  • hPBMCs Resuspend hPBMCs in complete medium and seed into 96-well cell culture plates. Dilute the test antibody to various concentrations in complete medium, and add the test antibody IMP-2023-033 or its dilution to the corresponding wells. Finally, incubate the 96-well cell culture plates containing cells in a CO2 incubator at 37°C.
  • T cells were collected and processed using the Fixation/Permeabilization Solution Kit (Invitrogen, 00-5523-00).
  • APC-anti-CD4 antibody Biolegend, 344614
  • PE-anti-CD8 antibody Biolegend, 301008
  • FITC-anti-Ki67 antibody BD Pharmingen, 556026
  • Ki67 is a marker protein for cell proliferation.
  • hPBMCs and Raji cells were resuspended in complete culture medium and seeded into 96-well plates at a 10:1 effector cell:target cell ratio.
  • Serial dilutions of IMP-2023-033 or IMP-2023-033 plus dexamethasone (Dex) (Sigma, 50-02-2) were added and incubated.
  • the LDH kit (Promega, batch number: G1782) was used to determine the killing curve of IMP-2023-033-mediated T cells against Raji cells in the presence or absence of Dex according to the method provided in the instructions.
  • the specific method is: take the cell supernatant or cell lysate, add the LDH substrate, stop the reaction after 30 minutes, and measure the absorbance value at 490nm (OD490). LDH is released by dead cells, so the LDH content in the cell supernatant is related to the degree of cell death.
  • the cell death rate was calculated according to the following formula:
  • [OD490] represents the minimum absorbance value of the supernatant of the wells containing Raji cells and PBMCs;
  • [OD490] background represents the absorbance value of the mixed liquid in the cell culture medium and the cell lysis solution added during the test.
  • IL-2 ELISA kit R&D, DY202
  • IL-6 ELISA kit R&D, DY206
  • TNF- ⁇ ELISA kit R&D, DY210
  • IFN- ⁇ ELISA kit R&D, DY285B
  • T cell activation and proliferation can cause cytokine release syndrome (CRS), and fulminant CRS can be life-threatening.
  • Glucocorticoids such as dexamethasone (Dex)
  • Dex dexamethasone
  • Human PBMCs were resuspended in PBS and the cell density was adjusted to 2.5 ⁇ 10 7 /mL. 5 ⁇ 10 6 cells were injected intraperitoneally into each mouse for immune reconstitution 5 days before tumor cell inoculation (Day-5). Each mouse was injected with 0.2 mL.
  • NCI-H929 cells in the logarithmic growth phase of in vitro culture were collected, resuspended in PBS and the cell density was adjusted to 3 ⁇ 10 7 /mL. 3 ⁇ 10 6 cells were injected subcutaneously into each mouse, and 0.1 mL was injected into each mouse.
  • mice with normally distributed tumor volumes were randomly divided into five groups: a vehicle control group (PBS), a subcutaneous low-dose IMP-2023-033 group (300 ⁇ g/kg, SC), a subcutaneous medium-dose IMP-2023-033 group (1000 ⁇ g/kg, SC), a subcutaneous high-dose IMP-2023-033 group (3000 ⁇ g/kg, SC), and an intraperitoneal administration group (1000 ⁇ g/kg, IP).
  • PBS vehicle control group
  • SC subcutaneous low-dose IMP-2023-033 group
  • SC subcutaneous medium-dose IMP-2023-033 group
  • a subcutaneous high-dose IMP-2023-033 group 3000 ⁇ g/kg, SC
  • IP intraperitoneal administration group
  • the subcutaneous high-dose group (IMP-2023-033, 3000 ⁇ g/kg, SC) and the intraperitoneal administration group (IMP-2023-033, 1000 ⁇ g/kg, IP) were administered for 2 weeks, and the other groups were administered for 3 weeks, after which the drug was discontinued for observation.
  • the long diameter (L) and short diameter (W) of tumors in tumor-bearing mice were measured twice a week using a vernier caliper, and the tumor volume (TV) and tumor growth inhibition rate (TGI%) were calculated according to the following formula: avT 0 and avT i are the average tumor volumes of mice in the treatment group at grouping (Day 7) and Day 7 + i, respectively; avC 0 and avC i are the average tumor volumes of mice in the vehicle control group at grouping (Day 7) and Day 7 + i, respectively.
  • Subcutaneous injection of IMP-2023-033 showed dose-dependent antitumor efficacy in NCI-H929 tumor-bearing mice reconstituted with human PBMC immunity.
  • the TGI% of the subcutaneous low-dose group (IMP-2023-033, 300 ⁇ g/kg, SC), medium-dose group (IMP-2023-033, 1000 ⁇ g/kg, SC), and high-dose group (IMP-2023-033, 3000 ⁇ g/kg, SC) on Day 27 were 43.1%, 65.9%, and 94.2%, respectively.
  • the P values were 0.0303, 0.0009, and 0.0002, respectively, and the differences were statistically significant.
  • mice After subcutaneous administration of 3000 ⁇ g/kg or intraperitoneal administration of 1000 ⁇ g/kg IMP-2023-033 to NCI-H929 tumor-bearing mice reconstituted with human PBMCs, 60% (3/5) and 80% (4/5) of the mice showed tumor regression, respectively.
  • IMP-2023-036 and IMP-2023-037 were 2.2 ⁇ 10-8 M and 1.4 ⁇ 10-8 M, respectively, while those of IMP-2023-038 and IMP-2023-039 were 1.3 ⁇ 10-9 M and 2.6 ⁇ 10-9 M, respectively.
  • IMP-2023-037 In binding to NCI-H929 cells (BCMA+), IMP-2023-037 had an EC50 value in the 10-8 M range, while the other EC50 values were all in the 10-9 M range.
  • IMP-2023-037 In binding to K562-CD19 cells (CD19+), IMP-2023-037 had the weakest binding, with an EC50 value of 1.0 ⁇ 10-8 M; the other three molecules all had EC50 values in the 10-10 M range.
  • IMP-2023-037 had the weakest binding, with an EC50 value of 1.7 ⁇ 10-9 M; IMP-2023-039 had an EC50 value of 1.6 ⁇ 10-10 M; while the EC50 values of the other two molecules were both at the 10-11 M level.
  • the cells tested for protein expression were used as target cells, and the specific preparation was as described above.
  • target cells were washed and centrifuged, then stained with CFSE (eBioscience, 65-0850-85) for 10 minutes at room temperature. The cells were then washed and resuspended in complete culture medium. The cells were then seeded at a 4:1 ratio of effector (T) cells to target cells and incubated with the test molecule. After 24 hours, 7-AAD (Biolegend, 420404) was added to the corresponding wells of the 96-well plate. After staining for 10-15 minutes, the cells were analyzed by flow cytometry.
  • CFSE eBioscience, 65-0850-85
  • CFSE + represents target cells
  • 7-AAD - represents viable cells
  • CFSE + 7-AAD - represents viable target cells
  • GraphPad Prism software was used to analyze the data. The cell killing rate was used as the ordinate and the drug concentration was used as the abscissa. A four-parameter curve was fitted to plot the data and calculate the EC50 value.
  • the results are shown in Figure 12 and Table 7.
  • the target cells in Figure 12A were NCI-H929 cells expressing BCMA but not CD19.
  • the data showed that IMP-2023-037 and IMP-2023-038 had cytotoxicity EC50 values greater than 1000 pM, while IMP-2023-036 and IMP-2023-039 had cytotoxicity EC50 values of 242 pM and 80 pM, respectively, indicating that IMP-2023-039 had better activity.
  • the target cells in Figure 12B were K562-CD19 cells expressing CD19 but not BCMA.
  • IMP-2023-038 had the best cytotoxicity, with an EC50 of 1.3 pM, while IMP-2023-037 had the weakest cytotoxicity, with an EC50 of 128 pM.
  • IMP-2023-036 and IMP-2023-039 were intermediate, with EC 50 values of 7.2 pM and 14 pM, respectively.
  • Figure 12C shows that the target cells are Daudi cells expressing both CD19 and BCMA.
  • the data show that IMP-2023-036, IMP-2023-038, and IMP-2023-039 showed similar cytotoxicity, with EC 50 values of 4.4 pM, 2.3 pM, and 1.9 pM, respectively.
  • IMP-2023-037 showed weak cytotoxicity, with an EC 50 of 204 pM.
  • IMP-2023-037 and IMP-2023-038 showed very weak cytotoxicity against NCI-H929 cells, which only express BCMA, with EC 50 values greater than 1000 pM. However, they showed cytotoxicity against K562-CD19 and Daudi cells, which express CD19. This suggests that IMP-2023-037 and IMP-2023-038 may act through CD19 rather than BCMA, making them inappropriate triple-target antibodies. IMP-2023-036 and IMP-2023-039 showed cytotoxicity against all three target cell lines, making them triple-target antibodies.
  • IMP-2023-039 should be superior to IMP-2023-036, so IMP-2023-039 was selected for further experiments.
  • hPBMCs and Daudi were seeded into 96-well cell culture plates at a ratio of 10:1, and then the test antibody IMP-2023-039 was added. Finally, the 96-well cell culture plates containing cells were placed in a CO2 incubator and incubated at 37°C.
  • Ki67 is a marker protein for cell proliferation.
  • hPBMCs and CFSE-prestained Daudi were resuspended in complete culture medium at a ratio of 10:1 and seeded into 96-well plates.
  • Serial dilutions of IMP-2023-039 or IMP-2023-039 plus dexamethasone (Dex) (Sigma, 50-02-2) were added and incubated for 24 hours for cell killing experiments. Specific methods are as described above.
  • Cytokine detection experiment hPBMCs and Daudi were resuspended in complete medium at a ratio of 10:1 and seeded into 48-well plates. Serial dilutions of IMP-2023-039 or IMP-2023-039 plus dexamethasone (Dex) (Sigma, 50-02-2) were added and incubated. After 24 hours, the 48-well plates containing cells and supernatant were frozen at -80°C to prepare cell lysates.
  • Dex dexamethasone
  • the prepared cell lysates were thawed and the relevant cytokines were detected according to the IL-2 ELISA kit (R&D, DY202), IL-6 ELISA kit (R&D, DY206), TNF- ⁇ ELISA kit (R&D, DY210), and IFN- ⁇ ELISA kit (R&D, DY285B).

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Abstract

L'invention concerne une protéine de liaison à l'antigène multi-spécifique ciblant BCMA, CD19 et CD3. Plus particulièrement, l'invention concerne un adaptateur de lymphocyte T tri-spécifique formé par liaison d'un scFv anti-BCMA, d'un scFv anti-CD19 et d'un Fab anti-CD3. L'adaptateur de lymphocyte T multi-spécifique est capable de cibler simultanément BCMA, CD19 et CD3 avec une affinité de liaison élevée, liant ainsi des cellules cibles et des lymphocytes T, et favorisant l'effet destructeur de lymphocytes T. La présente invention a la perspective d'être appliquée dans le domaine antitumoral.
PCT/CN2025/076236 2024-02-08 2025-02-07 Protéine de liaison à l'antigène multi-spécifique ciblant bcma, cd19 et cd3 et son utilisation Pending WO2025168059A1 (fr)

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Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20190263904A1 (en) * 2016-06-21 2019-08-29 Teneobio, Inc. Cd3 binding antibodies
WO2022199555A1 (fr) * 2021-03-23 2022-09-29 Guangzhou Lintonpharm Co., Ltd. Protéine de liaison à un antigène multispécifique se liant à cd3 et son utilisation
US20220340673A1 (en) * 2019-09-30 2022-10-27 Harbour Biomed (suzhou) Co., Ltd. Antibody targeting bcma, bispecific antibody, and use thereof
US20230002489A1 (en) * 2019-11-26 2023-01-05 Shanghai Epimab Biotherapeutics Co., Ltd. Antibodies to cd3 and bcma, and bispecific binding proteins made therefrom
WO2023029089A1 (fr) * 2021-09-03 2023-03-09 苏州近岸蛋白质科技股份有限公司 Anticorps humanisé anti-cd3
US20230348630A1 (en) * 2022-04-28 2023-11-02 Beijing Mabworks Biotech Co., Ltd Antibodies binding bcma and cd3 and uses thereof

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20190263904A1 (en) * 2016-06-21 2019-08-29 Teneobio, Inc. Cd3 binding antibodies
US20220340673A1 (en) * 2019-09-30 2022-10-27 Harbour Biomed (suzhou) Co., Ltd. Antibody targeting bcma, bispecific antibody, and use thereof
US20230002489A1 (en) * 2019-11-26 2023-01-05 Shanghai Epimab Biotherapeutics Co., Ltd. Antibodies to cd3 and bcma, and bispecific binding proteins made therefrom
WO2022199555A1 (fr) * 2021-03-23 2022-09-29 Guangzhou Lintonpharm Co., Ltd. Protéine de liaison à un antigène multispécifique se liant à cd3 et son utilisation
WO2023029089A1 (fr) * 2021-09-03 2023-03-09 苏州近岸蛋白质科技股份有限公司 Anticorps humanisé anti-cd3
US20230348630A1 (en) * 2022-04-28 2023-11-02 Beijing Mabworks Biotech Co., Ltd Antibodies binding bcma and cd3 and uses thereof

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