Anti-SARS-CoV-2 antigen-binding polypeptide, polypeptide complex, and method of using the same

Abstract

Antigen-binding polypeptides and antigen-binding polypeptide complexes (e.g., antibodies and antigen-binding fragments thereof) having certain structural and / or functional features are disclosed. Also disclosed are polynucleotides and vectors encoding such polypeptides and polypeptide complexes; host cells, pharmaceutical compositions and kits containing such polypeptides and polypeptide complexes; and methods of using such polypeptides and polypeptide complexes.

Claims

1. An antigen-binding polypeptide complex comprising a first polypeptide and a second polypeptide; The first polypeptide is, VL1-L1-VL2-L2-VH2-L3-VH1-L4-Fc; VL1-L1-VH2-L2-VL2-L3-VH1-L4-Fc; VH1-L5-VH2-L6-VL2-L7-VL1-L8-Fc; or VH1-L5-VL2-L6-VH2-L7-VL1-L8-Fc It has a structure represented by; The second polypeptide described above is Fc; VL3-L9-VL4-L10-VH4-L11-VH3-L12-Fc; VL3-L9-VH4-L10-VL4-L11-VH3-L12-Fc; VH3-L13-VH4-L14-VL4-L15-VL3-L16-Fc; or VH3-L13-VL4-L14-VH4-L15-VL3-L16-Fc It has a structure represented by; Here, VL1 is the first immunoglobulin light chain variable region that specifically binds to the SARS-CoV-2 protein; VL2 is a second immunoglobulin light chain variable region that specifically binds to the SARS-CoV-2 protein; VL3 is a third immunoglobulin light chain variable region that specifically binds to the SARS-CoV-2 protein; VL4 is a fourth immunoglobulin light chain variable region that specifically binds to the SARS-CoV-2 protein; VH1 is the first immunoglobulin heavy chain variable region that specifically binds to the SARS-CoV-2 protein; VH2 is a second immunoglobulin heavy chain variable region that specifically binds to the SARS-CoV-2 protein; VH3 is a third immunoglobulin heavy chain variable region that specifically binds to the SARS-CoV-2 protein; VH4 is a fourth immunoglobulin heavy chain variable region that specifically binds to the SARS-CoV-2 protein; Fc is a region comprising immunoglobulin heavy chain constant region 2 (CH2), immunoglobulin heavy chain constant region 3 (CH3), and optionally an immunoglobulin hinge; and L1 to L16 are amino acid linkers. The antigen-binding polypeptide complex.

2. It comprises a first polypeptide and a second polypeptide; The first polypeptide is, VL1-L1-VL2-L2-VH2-L3-VH1-L4-Fc; or VH1-L5-VH2-L6-VL2-L7-VL1-L8-Fc It has a structure represented by; The second polypeptide described above is Fc; VL3-L9-VL4-L10-VH4-L11-VH3-L12-Fc; or VH3-L13-VH4-L14-VL4-L15-VL3-L16-Fc It has a structure represented by; Here, VL1 is the first immunoglobulin light chain variable region that specifically binds to the SARS-CoV-2 protein; VL2 is a second immunoglobulin light chain variable region that specifically binds to the SARS-CoV-2 protein; VL3 is a third immunoglobulin light chain variable region that specifically binds to the SARS-CoV-2 protein; VL4 is a fourth immunoglobulin light chain variable region that specifically binds to the SARS-CoV-2 protein; VH1 is the first immunoglobulin heavy chain variable region that specifically binds to the SARS-CoV-2 protein; VH2 is a second immunoglobulin heavy chain variable region that specifically binds to the SARS-CoV-2 protein; VH3 is a third immunoglobulin heavy chain variable region that specifically binds to the SARS-CoV-2 protein; VH4 is a fourth immunoglobulin heavy chain variable region that specifically binds to the SARS-CoV-2 protein; Fc is a region comprising the immunoglobulin heavy chain constant region 2 (CH2), the immunoglobulin heavy chain constant region 3 (CH3), and optionally the immunoglobulin hinge; L1 to L16 are amino acid linkers. The antigen-binding polypeptide complex according to claim 1.

3. The antigen-binding polypeptide complex according to claim 1, wherein VH1, VH2, VH3, and VH4 each contain the same heavy chain variable region, and VL1, VL2, VL3, and VL4 each contain the same light chain variable region.

4. The antigen-binding polypeptide complex according to claim 1, wherein VH1 contains the same heavy chain variable region as VH3, and VL1 contains the same light chain variable region as VL3.

5. The antigen-binding polypeptide complex according to claim 1, wherein VH2 contains the same heavy chain variable region as VH4, and VL2 contains the same light chain variable region as VL4.

6. The antigen-binding polypeptide complex according to claim 1, wherein two of VH1, VH2, VH3, and VH4 include the same heavy chain variable region, and two of VL1, VL2, VL3, and VL4 include the same light chain variable region.

7. The antigen-binding polypeptide complex according to claim 1, wherein VH2 contains the same heavy chain variable region as VH4, and VL2 contains the same light chain variable region as VL4.

8. The antigen-binding polypeptide complex according to claim 1, wherein VH2 contains the same heavy chain variable region as VH3, and VL2 contains the same light chain variable region as VL3.

9. The antigen-binding polypeptide complex according to claim 1, wherein VH1 includes the same heavy chain variable region as VH4, and VL1 includes the same light chain variable region as VL4.

10. The antigen-binding polypeptide complex according to claim 1, wherein VH1 contains the same heavy chain variable region as VH3, and VL1 contains the same light chain variable region as VL3.

11. The antigen-binding polypeptide complex according to claim 1, wherein the immunoglobulin hinge comprises an upper hinge region, a central hinge region, a lower hinge region, or a combination thereof.

12. The antigen-binding polypeptide complex according to claim 1, wherein each linker L1 to L16 independently has a length of 0 to about 50 amino acids.

13. The antigen-binding polypeptide complex according to claim 1, wherein linkers L1 to L16, which do not have a length of 0 amino acids, each independently contain an amino acid sequence having at least 80%, at least 85%, at least 90%, or at least 95% identity with any one of SEQ ID NOs: 1 to 34, wherein SEQ ID NO: 1 has the amino acid sequence of G, SEQ ID NO: 2 has the amino acid sequence of A, SEQ ID NO: 3 has the amino acid sequence of GSS, and SEQ ID NO: 4 has the amino acid sequence of ASG.

14. The antigen-binding polypeptide complex according to claim 1, wherein one or more of the linkers L1 to L16 are non-immunogenic.

15. The antigen-binding polypeptide complex according to claim 1, wherein one or more of the linkers L1 to L16 do not contain a consensus T cell epitope.

16. The antigen-binding polypeptide complex according to claim 1, wherein the Fc region comprises at least one knob-into-hole modification or Fc effector function knockout mutation.

17. The antigen-binding polypeptide complex is an IgG1 or IgG4 antibody, and the knob-into-hole modification is based on the EU numbering scheme. (i) Replacement of knobs in S354C and T366W and replacement of holes in Y349C, T366S, L368A and Y407V; (ii) Hole replacement of M428L and N434S or N434A; (iii) Hole replacement for M252Y, S254T and T256E; or (iv) combinations of those The antigen-binding polypeptide complex according to claim 16, comprising:

18. The antigen-binding polypeptide complex according to claim 16, wherein the antigen-binding polypeptide complex is an IgG1 or IgG4 antibody, and the Fc effector function knockout mutation is L234A, L235A, P239A, or a combination thereof, based on the EU numbering scheme.

19. The antigen-binding polypeptide complex according to claim 1, comprising a detectable label.

20. The antigen-binding polypeptide complex according to claim 19, wherein the detectable label is a radioactive label, a chemiluminescent label, a fluorescent label, an enzyme, a peptide tag, or a combination thereof.

21. The antigen-binding polypeptide complex according to claim 20, wherein the peptide tag is a polyhistidine tag consisting of about 4 to about 10 histidine residues.

22. The antigen-binding polypeptide complex according to claim 21, wherein the polyhistidine tag consists of approximately eight histidine residues.

23. Equilibrium dissociation constant (K) of approximately 10 μM to approximately 1 pM Ｄ The antigen-binding polypeptide complex according to claim 1, which specifically binds to the SARS-CoV-2 protein.

24. The antigen-binding polypeptide complex is It is bispecific, triplicate, or quadruple specific, and Having a higher neutralizing efficacy against the SARS-CoV-2 virus than a single-specific antigen-binding polypeptide complex that specifically binds to one of the same antigens as the aforementioned bispecific, triplicate, or quadruplicate antigen-binding polypeptide complex, The antigen-binding polypeptide complex according to claim 1.

25. The antigen-binding polypeptide complex is It is bispecific, triplicate, or quadruple specific, and Having a higher neutralizing efficacy against the SARS-CoV-2 virus than a mixture of monospecific antigen-binding polypeptide complexes that specifically bind to the same antigen as the aforementioned bispecific, triplicate, or quadruplicate antigen-binding polypeptide complexes, The antigen-binding polypeptide complex according to claim 1.

26. The antigen-binding polypeptide complex according to claim 24 or 25, wherein the SARS-CoV-2 virus is the original Wuhan strain (WA1), the D614G spike protein variant (D614G), the alpha variant (B.1.1.7), the beta variant (B.1.351), the gamma variant (P.1), the delta variant, the epsilon variant (B.1.427), or the omicron variant (B.1.1.529), or a combination thereof.

27. A pharmaceutical composition comprising an antigen-binding polypeptide complex according to any one of claims 1 to 25 and a pharmaceutically acceptable carrier.

28. A pharmaceutical composition comprising an antigen-binding polypeptide complex according to any one of claims 1 to 25, an additional pharmaceutical agent, and a pharmaceutically acceptable carrier.

29. The pharmaceutical composition according to claim 28, wherein the additional pharmaceutical agent is 25-hydroxyvitamin D, a vitamin D enhancer, an antiviral agent, an antimalarial agent, an antibiotic, or a combination thereof.

30. The pharmaceutical composition according to claim 29, wherein the additional pharmaceutical agent is 25-hydroxyvitamin D.

31. The pharmaceutical composition according to claim 29, wherein the agent that enhances the action of vitamin D is a CYP24 inhibitor, a 1,25-dihydroxyvitamin D compound, or a combination thereof.

32. The pharmaceutical composition according to claim 29, wherein the antiviral agent is an antiretroviral agent, an antibody against the SARS-CoV-2 virus, a reverse transcriptase inhibitor, or a combination thereof.

33. The aforementioned antiviral agents include maraviroc, enfuvirtide, amantadine, lamivudine, nevirapine, efavirenz, dolutegravir, elvitegravir, raltegravir, acyclovir and any nucleoside analog of acyclovir, ganciclovir, cidofovir, forcalnet, ribavirin, interferon alfa, PEG-modified interferon alfa, boceprevir, atazanavir, darunavir, indinavir, oseltamivir, zanamivir, rimantadine, peremivir, and valacyclovir. The pharmaceutical composition according to claim 29, which is penciclovir, valganciclovir, foscarnet, tenofovir, adefovir, entecavir, lamivudine, terbivudine, ribavirin, glecaprevir, grazoprevir, paritaprevir, simeprevir, boxylaprevir, daclatasvir, elbasvir, ledipasvir, ombitasvir, pibrentasvir, velpatasvir, dasabuvir, famciclovir, remdesivir, trifluridine, sofobuvir, bebuterobimab, or a combination thereof.

34. The pharmaceutical composition according to claim 33, wherein the antiviral agent is bebuterobimab.

35. The aforementioned antiviral agents are Retrovir® (3'-azido-3'-deoxypyrimidine, zidovudine), 3'-azido-3'-deoxythymidine (AZT), HMD® (2',3'-dideoxycytidine, zalcitabine), VidexEC® (2',3'-dideoxyinosine, didanosine), Epivir® (lamivudine), and Zerit® (registered trademark). ) (Stabzine), Viread® (Tenofovir DF), Ziagen® (Abacavir), Emtriva® (Emtricitabine, FTC), Rescriber® (Delavirdin), Sustiva® (Efavirenz), Viramune® (Nevirapine, 11-Cyclopropyl-4-methyl-5,11-Dihydro-6) H-dipyride[3,2-b:2',3'-e][1,4]diazepine-6-one), trisodium phosphonoformate, ammonium-21-tungstenato-9-antimonate, 1-β-D-ribofuranoxyl-1,2,4-triazole-3-carboxamide, Aganerase® (amprenavir), Reyataz® (atazanavir), Lexiva® (Fosamprenavir), Crixivan® (Indinavir), Viracept® (Nelfinavir), Norvir® (Ritonavir), Fortovase® or Invirase® (Saquinavir), Lasinavir (5(S)-(tert-butoxycarbonylamino)-4(S)-hydroxy-6-phenyl-2(R)(2,3,4-Trimethoxyphenylmethyl)-Hexanoyl-(L)-Valyl-N-(2-Methoxyethyl)-Amid), Adriamycin, KVX-478, VX-478, 141W94, AG-1343, KNI-272, U-96988, BILA-2011BS (Palinavir), Polymannol acetate, Fuzeon® (Enfuvirtide, T-20), Epzicom® ( The pharmaceutical composition according to claim 29, which is abacavir and lamivudine), Trizivir® (abacavir, lamivudine and zidovudine), Truvada® (emtricitabine and tenofil DF), Combivir® (lamivudine and zidovudine), Kaletra® (lopinavir and ritonavir), bebuterobimab, or a combination thereof.

36. A kit comprising an antigen-binding polypeptide complex according to any one of claims 1 to 25.

37. (i) an antigen-binding polypeptide complex according to any one of claims 1 to 25, and (ii) Additional pharmaceutical agents A kit that includes this.

38. A pharmaceutical composition for preventing or treating SARS-CoV-2 virus infection in a subject in need, comprising a therapeutically effective amount of the antigen-binding polypeptide complex according to any one of claims 1 to 25.

39. A pharmaceutical composition for preventing or treating coronavirus disease 2019 (COVID-19) in a subject in need, comprising a therapeutically effective amount of the antigen-binding polypeptide complex according to any one of claims 1 to 25.

40. An in vitro or ex vivo method for assisting in the diagnosis of SARS-CoV-2 virus infection, (i) a step of contacting a sample obtained from a subject with an antigen-binding polypeptide complex according to any one of claims 1 to 25; and (ii) A step of detecting the presence or absence of a virus complex containing the polypeptide complex. The method comprising, if the virus complex is detected, the presence of the complex indicates that the subject is infected with the SARS-CoV-2 virus or is suspected to be infected with the SARS-CoV-2 virus.

41. An in vitro or ex vivo method for assisting in the diagnosis of SARS-CoV-2 virus infection, (i) a step of contacting a sample obtained from a subject with an antigen-binding polypeptide complex according to any one of claims 1 to 25; and (ii) A step of detecting the presence or absence of a virus complex containing the polypeptide complex. The method comprising, wherein if the virus complex is not detected, its absence indicates that the subject is not infected with the SARS-CoV-2 virus or is not suspected of being infected with the SARS-CoV-2 virus.

42. An in vitro or ex vivo method for assisting in the diagnosis of COVID-19, (i) a step of contacting a sample obtained from a subject with an antigen-binding polypeptide complex according to any one of claims 1 to 25; and (ii) A step of detecting the presence or absence of a virus complex containing the polypeptide complex. The method comprising, if the virus complex is detected, the presence thereof indicates that the subject has COVID-19 or is suspected of having COVID-19.

43. An in vitro or ex vivo method for assisting in the diagnosis of COVID-19, (i) a step of contacting a sample obtained from a subject with an antigen-binding polypeptide complex according to any one of claims 1 to 25; and (ii) A step of detecting the presence or absence of a virus complex containing the polypeptide complex. The method comprising, wherein if the virus complex is not detected, its absence indicates that the subject does not have COVID-19 or is not suspected of having COVID-19.

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