Monoclonal antibodies against influenza virus generated by cyclical administration and uses thereof
Provided herein are methods of producing neutralizing monoclonal antibodies, by cyclical immunization, that cross-react with strains of Influenza virus of the same subtype or different subtypes. Also provided herein are compositions comprising such antibodies and methods of using such antibodies to diagnose, prevent or treat Influenza virus disease.
1. A hybridoma
(a) designated 7A7 deposited under provisions of the Budapest Treaty with the American Type Culture Collection (ATCC, 10801 University Blvd., Manassas, Va. 20110-2209) on May 22, 2009 (ATCC Accession No. PTA-10058),
(b) designated 12D1 deposited under provisions of the Budapest Treaty with the American Type Culture Collection (ATCC, 10801 University Blvd., Manassas, Va. 20110-2209) on May 22, 2009 (ATCC Accession No. PTA-10059),
(c) designated 39A4 deposited under provisions of the Budapest Treaty with the American Type Culture Collection (ATCC, 10801 University Blvd., Manassas, Va. 20110-2209) on May 22, 2009 (ATCC Accession No. PTA-10060), or
(d) designated 66A6 deposited under provisions of the Budapest Treaty with the American Type Culture Collection (ATCC, 10801 University Blvd., Manassas, Va. 20110-2209) on May 26, 2010 (ATCC Accession No. PTA-11046).
2. An isolated monoclonal antibody produced by the hybridoma designated 7A7, 12D1, 39A4, or 66A6 of claim 1 .
3. A humanized antibody generated from the monoclonal antibody of claim 2 .
4. An isolated antibody that binds to SEQ ID NO:1, SEQ ID NO:124, or SEQ ID NO:125, and neutralizes two or more strains of an Influenza A virus HA subtype.
5. An isolated antibody that binds to Influenza A virus of the H3 subtype, the antibody comprising (a) the variable heavy (VH) domain of the antibody 7A7, 12D1, 39A4, or 66A6; (b) the variable light (VL) domain of the antibody 7A7, 12D1, 39A4, or 66A6; (c) the VH domain of the antibody 7A7, 12D1, 39A4, or 66A6and the VL domain of the antibody 7A7, 12D1, 39A4, or 66A6; (d) the VH complementarity determining regions (CDRs) of the antibody 7A7, 12D1, 39A4, or 66A6; (e) the VL CDRs of the antibody 7A7, 12D1, 39A4, or 66A6; or (f) the VH CDRs of the antibody 7A7, 12D1, 39A4, or 66A6and the VL CDRs of the antibody 7A7, 12D1, 39A4, or 66A6.
6. A composition comprising the antibody of claim 3 .
7. A method of preventing an Influenza virus disease comprising administering to a subject the antibody of claim 3 .
8. A method of detecting a strain of Influenza A virus H3 subtype comprising: (a) assaying for the level of an Influenza virus HA in cells or a tissue sample of a subject using the antibody of claim 2 or 3 ; and (b) comparing the level of the Influenza virus HA assayed in (a) with the level of the Influenza virus HA in normal tissue samples not infected with Influenza virus, wherein an increase in the assayed level of Influenza virus HA compared to the control level of the Influenza virus antigen is indicative of the presence of a strain of Influenza A virus H3 subtype.
9. A kit comprising, in a container, the antibody of claim 3 .
10. An isolated nucleic acid encoding an antibody that binds to Influenza A virus of the H3 subtype, wherein the antibody comprises (a) the amino acid sequence of the VH domain of the antibody 7A7, 12D1, 39A4, or 66A6; (b) the amino acid sequence of the VL domain of the antibody 7A7, 12D1, 39A4, or 66A6; (c) the amino acid sequence of the VH domain of the antibody 7A7, 12D1, 39A4, or 66A6and the VL domain of the antibody 7A7, 12D1, 39A4, or 66A6; (d) the amino acid sequence of the VH CDRs of the antibody 7A7, 12D1, 39A4, or 66A6; (e) the amino acid sequence of the VL CDRs of the antibody 7A7, 12D1, 39A4, or 66A6; or (f) the VH CDRs of the antibody 7A7, 12D1, 39A4, or 66A6and the VL CDRs of the antibody 7A7, 12D1, 39A4, or 66A6.
11. A host cell genetically engineered to contain or express the nucleic acid of claim 10 .
12. A method of producing an antibody, comprising culturing the host cell of claim 11 under conditions in which the nucleic acid is expressed and recovering the antibody from the host cell culture.
13. The method of claim 7 , wherein the subject is a human.
14. An isolated antibody that binds to Influenza A virus of the H3 subtype, the antibody comprising
(a) a heavy chain variable region comprising a complementarity determining region (CDR)1 having the amino acid sequence of SEQ ID NO:38, a CDR2 having the amino acid sequence of SEQ ID NO:39, and a CDR3 having the amino acid sequence of SEQ ID NO:40;
and a light chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:46, a CDR2 having the amino acid sequence of SEQ ID NO:47, and a CDR3 having the amino acid sequence of SEQ ID NO:48;
(b) a heavy chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:70, a CDR2 having the amino acid sequence of SEQ ID NO:71, and a CDR3 having the amino acid sequence of SEQ ID NO:72; and a light chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:78, a CDR2 having the amino acid sequence of SEQ ID NO:79, and a CDR3 having the amino acid sequence of SEQ ID NO:80;
(c) a heavy chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:92, a CDR2 having the amino acid sequence of SEQ ID NO:93, and a CDR3 having the amino acid sequence of SEQ ID NO:94; and a light chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:100, a CDR2 having the amino acid sequence of SEQ ID NO: 101, and a CDR3 having the amino acid sequence of SEQ ID NO:102; or
(d) a heavy chain variable region comprising a CDR1 of the antibody 39A4, a CDR2 of the antibody 39A4, and a CDR3 of the antibody 39A4; and a light chain variable region comprising a CDR1 of the antibody 39A4, a CDR2 of the antibody 39A4, and a CDR3 of the antibody 39A4.
15. The antibody of claim 14 , wherein the antibody comprises a heavy chain variable region comprising a complementarity determining region (CDR)1 having the amino acid sequence of SEQ ID NO:38, a CDR2 having the amino acid sequence of SEQ ID NO:39, and a CDR3 having the amino acid sequence of SEQ ID NO:40; and a light chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:46, a CDR2 having the amino acid sequence of SEQ ID NO:47, and a CDR3 having the amino acid sequence of SEQ ID NO:48.
16. The antibody of claim 14 , wherein the antibody comprises a heavy chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:70, a CDR2 having the amino acid sequence of SEQ ID NO:71, and a CDR3 having the amino acid sequence of SEQ ID NO:72; and a light chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:78, a CDR2 having the amino acid sequence of SEQ ID NO:79, and a CDR3 having the amino acid sequence of SEQ ID NO:80.
17. The antibody of claim 14 , wherein the antibody comprises a heavy chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:92, a CDR2 having the amino acid sequence of SEQ ID NO:93, and a CDR3 having the amino acid sequence of SEQ ID NO:94; and a light chain variable region comprising a CDR1 having the amino acid sequence of SEQ ID NO:100, a CDR2 having the amino acid sequence of SEQ ID NO: 101, and a CDR3 having the amino acid sequence of SEQ ID NO:102.
18. The antibody of claim 14 , wherein the antibody comprises a heavy chain variable region comprising a CDR1 of the antibody 39A4, a CDR2 of the antibody 39A4, and a CDR3 of the antibody 39A4; and a light chain variable region comprising a CDR1 of the antibody 39A4, a CDR2 of the antibody 39A4, and a CDR3 of the antibody 39A4.
19. The antibody of claim 15 , wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:6, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:7.
20. The antibody of claim 16 , wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:12, and the light chain variable region comprising the amino acid sequence of SEQ ID NO:13.
21. The antibody of claim 17 , wherein the heavy chain variable region comprising the amino acid sequence of SEQ ID NO:16, and the light chain variable region comprising the amino acid sequence of SEQ ID NO:17.
22. The antibody of claim 18 , wherein the heavy chain variable region comprising the amino acid sequence of the heavy chain variable region of antibody 39A4, and the light chain variable region comprising the amino acid sequence of the heavy chain variable region of antibody 39A4.
23. The isolated antibody of claim 14 , wherein the antibody is a humanized or chimeric antibody.
24. A method of preventing an Influenza virus disease comprising administering to a subject the antibody of claim 14 .
25. The method of claim 24 , wherein the subject is a human.
26. A method of detecting a strain of Influenza A virus H3 subtype comprising: (a) assaying for the level of an Influenza virus HA in cells or a tissue sample of a subject using the antibody of claim 14 ; and (b) comparing the level of the Influenza virus HA assayed in (a) with the level of the Influenza virus HA in normal tissue samples not infected with Influenza virus, wherein an increase in the assayed level of Influenza virus HA compared to the control level of the Influenza virus antigen is indicative of the presence of a strain of Influenza A virus H3 subtype.
27. An isolated nucleic acid encoding the antibody of claim 14 .
28. A host cell genetically engineered to contain or express the nucleic acid of claim 26 .
29. A method of producing an antibody, comprising culturing the host cell of claim 28 under conditions in which the nucleic acid is expressed and recovering the antibody from the host cell culture.
30. A method of preventing an Influenza virus disease comprising administering to a subject the antibody of claim 4 .
31. The method of claim 28 , wherein the subject is a human.
32. A method of detecting a strain of Influenza A virus H3 subtype comprising: (a) assaying for the level of an Influenza virus HA in cells or a tissue sample of a subject using the antibody of claim 4 ; and (b) comparing the level of the Influenza virus HA assayed in (a) with the level of the Influenza virus HA in normal tissue samples not infected with Influenza virus, wherein an increase in the assayed level of Influenza virus HA compared to the control level of the Influenza virus antigen is indicative of the presence of a strain of Influenza A virus H3 subtype.
33. The hybridoma of claim 1 , which is designated 7A7 deposited under provisions of the Budapest Treaty with the American Type Culture Collection (ATCC, 10801 University Blvd., Manassas, Va. 20110-2209) on May 22, 2009 (ATCC Accession No. PTA-10058).
34. The hybridoma of claim 1 , which is designated 12D1deposited under provisions of the Budapest Treaty with the American Type Culture Collection (ATCC, 10801 University Blvd., Manassas, Va. 20110-2209) on May 22, 2009 (ATCC Accession No. PTA-10059).
35. The hybridoma of claim 1 , which is designated 39A4deposited under provisions of the Budapest Treaty with the American Type Culture Collection (ATCC, 10801 University Blvd., Manassas, Va. 20110-2209) on May 22, 2009 (ATCC Accession No. PTA-10060).
36. The hybridoma of claim 1 , which is designated 66A6deposited under provisions of the Budapest Treaty with the American Type Culture Collection (ATCC, 10801 University Blvd., Manassas, Va. 20110-2209) on May 26, 2010 (ATCC Accession No. PTA-11046).