Antibodies against SARS-CoV and methods of use thereof

• US 20050249739A1
• Filed: 11/24/2004
• Published: 11/10/2005
• Est. Priority Date: 11/25/2003
• Status: Active Grant

• Current Assignee: Dana-Farber Cancer Institute
• Original Assignee: Dana-Farber Cancer Institute
• Inventors: Sui, Jianhua, Marasco, Wayne
• Granted Patent: US 7,750,123 B2
• Time in Patent Office: Days
• US Class Current: 424/159.100
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61K 2039/545 characterised by the dose, timing or administration schedule
• CPC Class Codes: C07K 16/1002 Coronaviridae
• CPC Class Codes: C07K 2317/21 from primates, e.g. man
• CPC Class Codes: C07K 2317/34 Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
• CPC Class Codes: C07K 2317/56 variable (Fv) region, i.e. VH and/or VL
• CPC Class Codes: C07K 2317/622 Single chain antibody (scFv)
• CPC Class Codes: C07K 2317/76 Antagonist effect on antigen, e.g. neutralization or inhibition of binding
• CPC Class Codes: C07K 2319/30 Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
• CPC Class Codes: C12N 2770/20011 Coronaviridae

Abstract

The invention provides scFv antibodies and monoclonal antibodies that neutralize SARS-CoV. Also provided are methods of treating and/or preventing a coronavirus-related disease or disorder such as SARS. The invention also provides methods of vaccinating a patient against SARS-CoV. Also provided are methods of diagnosing coronavirus-related diseases or disorders and methods of detecting the presence of a coronavirus in a sample. The invention additionally provides methods of screening for compounds that modulate the binding of SARS-CoV and the SARS-CoV receptor ACE2 as well as for compounds useful to treat SARS-CoV-related diseases or disorders.

Claims

1. A monoclonal antibody which neutralizes SARS-CoV, wherein said monoclonal antibody binds to an epitope on a region of the spike protein of SARS-CoV and neutralizes SARS-CoV.

2. The monoclonal antibody of claim 1, wherein the region of the spike protein of SARS-CoV is the S1 region.

3. The monoclonal antibody of claim 2, wherein the region is located between amino acids 318 and 510 of the S1 region.

4. The monoclonal antibody of claim 3, wherein the region is located between amino acids 324 and 503 of the S1 region.

5. The monoclonal antibody of claim 1, wherein said monoclonal antibody is monoclonal antibody 80R.

6. The monoclonal antibody of claim 1, wherein said monoclonal antibody has the binding affinity of monoclonal antibody 80R.

7. The monoclonal antibody of claim 6, wherein said binding affinity is from about 10⁻

6 M to about 10<sup>−

12</sup> M.

8. The monoclonal antibody of claim 1, wherein said monoclonal antibody inhibits binding of the S1 region of the S protein of SARS-CoV to the ACE2 receptor.

9. The monoclonal antibody of claim 1, wherein said monoclonal antibody inhibits syncytia formation between cells expressing SARS-CoV S protein and cells expressing the SARS-CoV receptor ACE2.

10. The monoclonal antibody of claim 1, wherein said monoclonal antibody competes with the binding of monoclonal antibody 80R to the S1 region.

11. The monoclonal antibody of claim 1, wherein said monoclonal antibody comprises one or more sequences selected from the group consisting of SEQ ID NOS:

2, 3, 4, and 5.

25. A method of preventing a diseaseor disorder caused by a coronavirus, the method comprising administering to a person at risk of suffering from said disease or disorder, a therapeutically effective amount of the monoclonal antibody of claim 1.

26. The method of claim 25, wherein the method further comprises administering other anti-viral drugs or entry inhibitors in addition to administering the monoclonal antibody.

27. The method of claim 25, wherein the coronavirus is SARS-CoV and wherein the monoclonal antibody is monoclonal antibody 80R.

31. A method of treating a coronavirus-related disease or disorder, the method comprising administering a therapeutically effective amount of the monoclonal antibody of claim 1 to a patient suffering from a coronavirus-related disease or disorder.

32. The method of claim 31, wherein the coronavirus related disease or disorder is SARS and wherein the monoclonal antibody is monoclonal antibody 80R.

35. A method for vaccinating a patient against SARS-CoV, the method comprising administering an immunogenically effective amount of a vaccine candidate peptide to a patient in need of vaccination, wherein said vaccine candidate peptide binds to the monoclonal antibody of claim 1.

36. The method of claim 35, wherein the monoclonal antibody is monoclonal antibody 80R.

37. The method of claim 35, wherein the vaccine candidate peptide comprises the monoclonal antibody 80R epitope.

38. A method of diagnosing a coronavirus-related disease or disorder, the method comprising:

a) contacting a test sample obtained from a patient suspected of having the coronavirus-related disease or disorder with the monoclonal antibody of claim 1;

and

b) detecting the presence or absence of an antibody-antigen complex,

wherein the presence of the antibody-antigen complex indicates that the patient has the coronavirus-related disease or disorder.

39. The method of diagnosing of claim 38, wherein the coronavirus-related disease or disorder is SARS.

40. The method of claim 38, wherein the monoclonal antibody is monoclonal antibody 80R.

41. The method of claim 38, wherein the sample is obtained from blood, hair, cheek scraping, saliva, biopsy, or semen.

42. The method of claim 38, wherein the detecting occurs in vivo.

43. A method of detecting the presence of a coronavirus in a sample, the method comprising:

a) contacting the sample with the monoclonal antibody of claim 1;

and

b) detecting the presence or absence of an antibody-antigen complex,

thereby detecting the presence of a coronavirus in a sample.

44. The method of claim 43, wherein the coronavirus is SARS-CoV.

45. The method of claim 43, wherein the monoclonal antibody is monoclonal antibody 80R.

46. The method of claim 43, wherein the detecting occurs in vivo.

47. The method of claim 43, wherein the sample is obtained from blood, hair, cheek scraping, saliva, biopsy, or semen.

48. A composition comprising the monoclonal antibody of claim 1 and a carrier.

49. A kit comprising, in one or more containers, the composition of claim 48.

50. A passive vaccine against SARS-CoV comprising the composition of claim 48.

52. A method for identifying compounds useful to treat a SARS-Cov-related disease or disorder, the method comprising

a) providing at least one SARS-CoV protein;

b) exposing said protein to the monoclonal antibody of claim 1,

c) detecting the formation of an antibody-antigen complex;

d) introducing one or more candidate compounds; and

e) determining whether said one or more candidate compounds disrupt the antibody-antigen complex,

wherein the disruption of the antibody-antigen complex indicates that the one or more candidate compounds is useful to treat a SARS-CoV-related disease or disorder.

53. The method of claim 52, wherein the SARS-CoV-related disease or disorder is SARS.

54. The method of claim 53, wherein the monoclonal antibody is monoclonal antibody 80R.

55. The method of claim 53, wherein the at least one SARS-CoV protein comprises the S1 region of the spike protein of SARS-CoV.

58. The method of claim 53, wherein the at least one SARS-CoV protein is provided as a SARS-CoV molecule.

59. The method of claim 53, wherein the at least one SARS-CoV protein is provided in a cell infected with the SARS-CoV.

12. A monoclonal antibody capable of neutralizing SARS-CoV, wherein said monoclonal antibody binds to the same epitope as the 80R monoclonal antibody.

13. An scFv antibody which neutralizes SARS-CoV, wherein said scFv antibody binds to an epitope on a region of the spike protein of SARS-CoV and neutralizes SARS-CoV.

14. The scFv antibody of claim 13, wherein the region of the spike protein of SARS-CoV is the S1 region.

15. The scFv antibody of claim 14, wherein the region is located between amino acids 318 and 510 of the S1 region.

16. The scFv antibody of claim 15, wherein the region is located between amino acids 324 and 503 of the S1 region.

17. The scFv antibody of claim 13, wherein said scFv antibody is scFv antibody 80R.

18. The scFv antibody of claim 13, wherein said scFv antibody has the binding affinity of scFv antibody 80R.

19. The scFv antibody of claim 18, wherein said binding affinity is from about 10⁻

6 M to about 10<sup>−

12</sup> M.

20. The scFv antibody of claim 13, wherein said scFv antibody inhibits binding of the S1 region of the S protein of SARS-CoV to the ACE2 receptor.

21. The scFv antibody of claim 13, wherein said scFv antibody inhibits syncytia formation between cells expressing SARS-CoV S protein and cells expressing the SARS-CoV receptor ACE2.

22. The scFv antibody of claim 13, wherein said scFv antibody competes with the binding of scFv antibody 80R to the S1 region.

23. The scFv antibody of claim 13, wherein said scFv antibody has the sequence of SEQ ID NO:

1.

28. A method of preventing a disease or disorder caused by a coronavirus, the method comprising administering to a person at risk of suffering from said disease or disorder, a therapeutically effective amount of the scFv antibody of claim 13.

29. The method of claim 28, wherein the method further comprises administering other anti-viral drugs or entry inhibitors in addition to administering the scFv antibody.

30. The method of claim 28, wherein the coronavirus is SARS-CoV and wherein the scFv antibody is scFv antibody 80R.

33. A method of treating a coronavirus-related disease or disorder, the method comprising administering a therapeutically effective amount of the scFv antibody of claim 13 to a patient suffering from a coronavirus-related disease or disorder.

34. The method of claim 33, wherein the coronavirus-related disease or disorder is SARS and wherein the scFv antibody is scFv antibody 80R.

24. An scFv antibody capable of neutralizing SARS-CoV, wherein said scFv antibody binds to the same epitope as the 80R scFv antibody.

51. A method of screening for compounds that modulate the interaction between SARS-CoV and the SARS-CoV receptor ACE2, the method comprising

a) introducing a candidate compound to an antibody-antigen complex, wherein said antibody is monoclonal antibody 80R and said antigen is located on the S1 region of the S protein of SARS-CoV;

and

b) determining whether said candidate compound disrupts the antibody-antigen complex,

wherein the disruption of the antibody-antigen complex indicates that the candidate compound modulates the interaction between SARS-CoV and ACE2.

56. The method of claim 51, wherein the SARS-CoV protein comprises amino acid residues 318 to 510 of the S1 region.

57. The method of claim 56, wherein the SARS-CoV protein comprises amino acid residues 324 to 503 of the S1 region.

60. A method of enhancing the neutralization activity of MAb 80R, the method comprising

a) performing complementarity determining region mutagenesis on the MAb 80R binding site;

b) determining the neutralization activity of the mutated MAbs; and

c) identifying those mutated MAbs having increased neutralization activity.

62. The method of claims 60 or 61, wherein complementarity region determining mutagenesis is accomplished by light and heavy chain swapping or by CDR doping.

61. A method of enhancing the neutralization activity of scFv 80R, the method comprising

a) performing complementarity determining region mutagenesis on the scFv 80R binding site;

b) determining the neutralization activity of the mutated scFvs; and

c) identifying those mutated scFvs having increased neutralization activity.

63. A monoclonal antibody which neutralizes SARS-CoV, wherein said antibody has a light chain with three CDRs comprising an amino acid sequence selected from the group consisting of the amino acid sequences of RASQVRSNLA (SEQ ID NO:

37);

DASTAT (SEQ ID NO;

38); and

QQRSNWPPT (SEQ ID NO;

39).

64. A monoclonal antibody which neutralizes SARS-CoV, wherein said antibody has a heavy chain with three CDRs comprising an amino acid sequence selected from the group consisting of the amino acid sequences of AH;

VYDNK (SEQ ID NO;

     35); and

RSYYL (SEQ ID NO;

36).