Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
  • C07K16/08—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from viruses
  • C07K16/10—RNA viruses
  • C07K16/108—Orthomyxoviridae (F), e.g. influenza virus
• • 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
  • A61K39/12—Viral antigens
  • A61K39/145—Orthomyxoviridae, e.g. influenza virus
• • 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
  • A61K39/42—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum viral
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
  • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/12—Antivirals
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/12—Antivirals
  • A61P31/14—Antivirals for RNA viruses
  • A61P31/16—Antivirals for RNA viruses for influenza or rhinoviruses
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P37/00—Drugs for immunological or allergic disorders
  • A61P37/02—Immunomodulators
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P37/00—Drugs for immunological or allergic disorders
  • A61P37/02—Immunomodulators
  • A61P37/04—Immunostimulants
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
  • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
  • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
  • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
  • G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
  • G01N33/56983—Viruses
• • 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
  • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K2121/00—Preparations for use in therapy
• • 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
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K2317/00—Immunoglobulins specific features
  • C07K2317/30—Immunoglobulins specific features characterized by aspects of specificity or valency
  • C07K2317/33—Crossreactivity, e.g. for species or epitope, or lack of said crossreactivity
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K2317/00—Immunoglobulins specific features
  • C07K2317/30—Immunoglobulins specific features characterized by aspects of specificity or valency
  • C07K2317/34—Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K2317/00—Immunoglobulins specific features
  • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
  • C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K2317/00—Immunoglobulins specific features
  • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
  • C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
  • C07K2317/565—Complementarity determining region [CDR]
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K2317/00—Immunoglobulins specific features
  • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
  • C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
  • C07K2317/567—Framework region [FR]
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K2317/00—Immunoglobulins specific features
  • C07K2317/70—Immunoglobulins specific features characterized by effect upon binding to a cell or to an antigen
  • C07K2317/76—Antagonist effect on antigen, e.g. neutralization or inhibition of binding
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K2317/00—Immunoglobulins specific features
  • C07K2317/90—Immunoglobulins specific features characterized by (pharmaco)kinetic aspects or by stability of the immunoglobulin
  • C07K2317/92—Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
  • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10S435/00—Chemistry: molecular biology and microbiology
  • Y10S435/975—Kit

Definitions

• This disclosure relates to novel binding agents, e.g., peptide agents, e.g., antibody molecules, e.g., antibodies, and antigen-binding fragments thereof, that bind
• novel binding agents e.g., peptide agents, e.g., antibody molecules, e.g., antibodies, and antigen-binding fragments thereof, that bind
• hemagglutinin protein of influenza viruses and in embodiments neutralize the virus, and methods of their use.
• Influenza is an infectious disease caused by RNA viruses of the family
• Influenza viruses are classified based on core protein into three genera A, B and C that are further divided into subtypes determined by the viral envelope glycoproteins haemagglutinin (HA) and neuraminidase (NA). Influenza A viruses infect a range of mammalian and avian species, whereas type B and C infections are largely restricted to humans. Only types A and B cause human disease of any concern.
• Antigenic drift enables the virus to evade immune recognition, resulting in repeated influenza outbreaks during interpandemic years.
• Major changes in the HA antigen (“antigenic shift") are caused by reassortment of genetic material from different influenza A subtypes. Antigenic shifts resulting in new pandemic strains are rare events, occurring through reassortment between animal and human subtypes, for example in co-infected pigs.
• the disclosure is based, at least in part, on the discovery of human anti-HA antibodies comprising functional and structural properties disclosed herein, e.g., antibodies that bind a conserved region or epitope on influenza virus and uses thereof.
• binding agents e.g., antibody molecules, or preparations, or isolated preparations thereof, that bind hemagglutinin (HA) from influenza viruses.
• a binding agent e.g., an antibody molecule
• a binding agent is broad spectrum, and binds more than one HA, e.g., an HA from one or both of Group 1 or Group 2 strains of influenza A viruses and/or one or more strains of influenza B viruses. Therefore, in some embodiments, a binding agent, e.g., an antibody molecule, featured in the disclosure can treat or prevent infection by a Group 1 influenza virus and a Group 2 influenza virus.
• a binding agent e.g., an antibody molecule
• a binding agent can treat or prevent infection by an influenza A virus and an influenza B virus.
• the binding agents e.g, antibody molecules, share sufficient structural similarity with antibodies or variable regions disclosed herein such that they possess functional attributes of the antibodies disclosed herein.
• the structural similarity can be in terms of three dimensional structure or linear amino acid sequence or both.
• an anti-hemagglutinin (anti-HA) binding agent e.g., a specific binding agent, e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising one or more or all of the following properties:
• a Group 1 virus such as where the virus is an HI, H5, or H9 virus
• a Group 2 virus such as where the virus is an H3 or H7 virus
• the concentration of antibody molecule required for 50% neutralization of influenza A virus is less than 10 ⁇ g/mL;
• (k) it treats or prevents infection by an influenza B virus, e.g.,
• influenza B virus e.g., B/Wisconsin/1/2010
• q it binds an epitope other than that bound by a reference anti-HA antibody molecule, e.g., Ab 67-11, FI6, FI28, C179, F10, CR9114, or CR6261, e.g., as determined by structural analysis, e.g., by X-ray crystallography or NMR
• (r) in an embodiment it binds to an epitope, e.g., it has an epitope that overlaps with or is the same as, of an antibody disclosed herein, e.g., as determined by mutational analysis or crystal structure analysis
• the binding agent e.g., an anti-HA antibody molecule
• the anti-HA antibody molecule prevents infection by at least 1, 2, 3, 4 or 5 influenza subtypes of Group 1, and by at least 1, 2, 3, 4 or 5 influenza subtypes of Group 2; the concentration of the anti-HA antibody molecule required for 50% neutralization of influenza A virus is less than 10 ⁇ g/mL; or the anti-HA antibody molecule binds an epitope that comprises or consists of the hemagglutinin trimer interface.
• the binding agent e.g., an anti-HA antibody molecule
• treats or prevents infection by a Group 1 virus such as where the virus is an HI, H2, H5, H6, H8, H9, H12, Hll, H13, H16, or H17 virus; and treats or prevents infection by a Group 2 virus, such as where the virus is an H3, H4, H7, H10 or H15 virus.
• the binding agent e.g., an anti-HA antibody molecule, featured in the disclosure prevents infection by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11 influenza subtypes of Group 1, and by at least 1, 2, 3, 4, 5 or 6 influenza subtypes of Group 2.
• the binding agent e.g., an anti-HA antibody molecule
• the binding agent treats or prevents infection by one or more of HlNl, H2N2, H5N1, and H9N2, and also treats or prevents infection by one or more of H3N2 and H7N7.
• a binding agent e.g., antibody molecule, binds, and in embodiments, neutralizes:
• At least one strain from the Group 1 HI e.g., HI a or Hlb, cluster and at least one strain from the Group 2 H3 or H7 cluster.
• a binding agent e.g., antibody molecule, binds, and in embodiments, neutralizes:
• At least one strain from the Group 1 HI e.g., HI a or Hlb
• at least one influenza B strain e.g., B/Wisconsin/1/2010.
• a binding agent e.g., antibody molecule, binds, and in embodiments, neutralizes:
• At least one strain from the Group 2 H3 or H7 cluster and at least one influenza B strain e.g., B/Wisconsin/1/2010.
• a binding agent e.g., antibody molecule, binds, and in embodiments, neutralizes:
• At least one strain from the Group 1 HI e.g., HI a or Hlb, cluster
• at least one strain from the Group 2 H3 or H7 cluster at least one influenza B strain, e.g., B/Wisconsin/1/2010.
• the binding agent e.g., an anti-HA antibody molecule
• the binding agent treats or prevents infection by one or more of influenza B viruses, e.g., B/Wisconsin/1/2010.
• the anti-HA antibody molecule is not an anti-HA antibody molecule previously described in the art.
• the anti-HA antibody molecule is other than one or more or all of Ab 67-11 (U.S. Provisional Application No. 61/645,453), FI6 (FI6, as used herein, refers to any specifically disclosed FI6 sequence in U.S. Published Application No. 2010/0080813, US published application No. 2011/0274702, WO2013/011347 or Corti et al., Science 333:850-856, 2011, published online July 28, 2011; HGs. 12A to 12C), FI28 (U.S. Published Application No. 2010/0080813), C179 (Okuno et al., J.
• the binding agent e.g., an anti-HA antibody molecule
• binding agent e.g., an anti-HA antibody molecule
• the binding agent e.g., an anti-HA antibody molecule
• binding agent e.g., an anti-HA antibody molecule neutralizes infection with H5N1 in vivo.
• the binding agent e.g., an anti-HA antibody molecule
• neutralizes infection with an influenza B virus e.g., B/Wisconsin/1/2010, in vitro.
• the binding agent e.g., an anti-HA antibody molecule neutralizes infection with an influenza B virus, e.g., B/Wisconsin/1/2010, in vivo.
• the concentration of the binding agent, e.g., an anti- HA antibody molecule, required for 50% neutralization of influenza A virus is 10 ⁇ g/mL or less, such as 9 ⁇ g/mL or less, 8 ⁇ g/mL or less, 7 ⁇ g/mL or less, 6 ⁇ g/mL or less, or 5 ⁇ g/mL or less.
• the concentration of the binding agent, e.g., an anti- HA antibody molecule, required for 60% neutralization of influenza A virus, 50% neutralization of influenza A virus, or 40% neutralization of influenza A virus is 10 ⁇ g/mL or less, such as 9 ⁇ g/mL or less, 8 ⁇ g/mL or less, 7 ⁇ g/mL or less, 6 ⁇ g/mL or less, or 5 ⁇ g/mL or less.
• the binding agent e.g., an anti-HA antibody molecule
• the binding agent is effective for prevention or treatment of infection, e.g., in humans or mice, with H1N1 and H3N2, such as when administered at 50 mg/kg, 25 mg/kg, 10 mg/kg, 6.0 mg/kg, 5.0 mg/kg, 4.0 mg/kg, 3.0 mg/kg, 2.0 mg/kg, 1.0 mg/kg or less.
• the binding agent e.g., an the anti-HA antibody molecule
• is effective for prevention or treatment of infection e.g., in humans or mice, with H5N1, such as when administered at 50 mg/kg, 25 mg/kg, 10 mg/kg, 6.0 mg/kg, 5.0 mg/kg, 4.0 mg/kg, 3.0 mg/kg, 2.0 mg/kg, 1.0 mg/kg or less.
• a binding agent e.g., an anti-HA antibody molecule
• a binding agent is effective for the treatment or prevention of a Group 1 virus, where the Group 1 virus is HI, H5, or H9
• the binding agent e.g., an anti- HA antibody molecule
• the binding agent is effective for the treatment or prevention of a Group 2 virus, where the Group 2 virus is H3 or H7.
• the concentration of the binding agent, e.g., an anti- HA antibody molecule, required for 50% neutralization of influenza B virus, e.g., B/Wisconsin/1/2010 is 10 ⁇ g/mL or less, such as 9 ⁇ g/mL or less, 8 ⁇ g/mL or less, 7 ⁇ g/mL or less, 6 ⁇ g/mL or less, or 5 ⁇ g/mL or less.
• the concentration of the binding agent e.g., an anti- HA antibody molecule, required for 60% neutralization of influenza B virus, e.g., B/Wisconsin/1/2010, 50% neutralization of influenza B virus, e.g.,
• B/Wisconsin/1/2010 is 10 ⁇ g/mL or less, such as 9 ⁇ g/mL or less, 8 ⁇ g/mL or less, 7 ⁇ g/mL or less, 6 ⁇ g/mL or less, or 5 ⁇ g/mL or less.
• the binding agent e.g., an anti-HA antibody molecule
• the binding agent is a full length tetrameric antibody, a single chain antibody (scFv), a F(ab') 2 fragment, a Fab fragment, or an Fd fragment.
• the heavy chain of the antibody molecule is a ⁇ heavy chain
• the light chain of the antibody molecule is a ⁇ light chain or a ⁇ light chain.
• the anti-HA antibody molecule featured in the disclosure is an IgGl antibody.
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f:
• H3 HA2 residues A7, Ell, 118, D19, G20, W21, L38, K39, T41, Q42, A43, 145, 148, N49, L52,
• the antibody molecule has properties: a; and b.
• the antibody molecule has properties: c; and d.
• the antibody molecule has properties: a; and c or d.
• the antibody molecule has properties: b; and c or d.
• the antibody molecule has properties: c; and a or b.
• the antibody molecule has properties: d; and a or b.
• the antibody molecule has properties: a, b, c and d. In an embodiment the antibody molecule has properties: a, b, c, d, e, and f. In an embodiment, the antibody molecule has a K D for H3 of equal to or less than 10 ⁇ 6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a KD for H3 of equal to or less than 10 "6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f:
• aa it includes one, two, or all of, HI HA1 residues H31, N279, and S292;
• dd it does not include one, two, three, four, or all of, HI HA2 residues Gl, L2, F3, G4, and D46;
• R322, and 1324 are bound by both Ab 044 and FI6; or ff) it includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or all of, HI HA2 residues A7, El l, 118, D19, G20, W21, Q38, K39, T41, Q42, N43, 145, 148, T49, V52, N53, 156, and E57
• the antibody molecule has properties: aa; and bb.
• the antibody molecule has properties: cc; and dd.
• the antibody molecule has properties: aa; and cc or dd.
• the antibody molecule has properties: bb; and cc or dd. In an embodiment the antibody molecule has properties: cc; and aa or bb.
• the antibody molecule has properties: dd; and aa or bb.
• the antibody molecule has properties: aa, bb, cc and dd.
• the antibody molecule has properties: aa, bb, cc, dd, ee, and ff.
• the antibody molecule has a KD for HI of equal to or less than 10 "6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a K D for HI of equal to or less than 10 ⁇ 6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule has one, two, three or all of the following properties:
• the molecule has properties c, cc, d, and dd.
• the binding agent e.g., a specific binding agent, e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising at least, or more than, 60, 65, 70, 75, 80, 85, 87, 90, 95, 98 or 99 percent homology with a heavy chain variable region from Table 3, Table 4A, Table 4B, Fig.2, Fig 13 or Fig.17; and
• the antibody molecule comprises a heavy chain variable region
• the antibody molecule comprises a light chain variable region 52 (SEQ ID NO:52), 155 (SEQ ID NO: 155), or 45 (SEQ ID NO:45), or a structurally or functionally related variable light chain region as described herein.
• the antibody molecule comprises:
• a heavy chain variable region 25 (SEQ ID NO:25), or a structurally or functionally related variable heavy chain region as described herein;
• a light chain variable region 52 (SEQ ID NO:52), 155 (SEQ ID NO: 155), or 45 (SEQ ID NO: 45), or a structurally or functionally related variable light chain region as described herein.
• the antibody molecule comprises a heavy chain variable region comprising one, two, or all of CDRl, CDR2, and CDR3, from heavy chain variable region 25 (SEQ ID NO:25), or a structurally or functionally related variable heavy chain region as described herein.
• the antibody molecule comprises a light chain variable region comprising one, two, or all of CDRl, CDR2, and CDR3, from light chain variable region 52 (SEQ ID NO:52), 155 (SEQ ID NO: 155), or 45 (SEQ ID NO:45), or a structurally or functionally related sequence as described herein.
• the antibody molecule comprises:
• heavy chain variable region comprising one, two, or all of CDRl, CDR2, and CDR3, from heavy chain variable region 25 (SEQ ID NO:25), or a structurally or functionally related variable heavy chain region as described herein;
• a light chain variable region comprising one, two, or all of CDRl, CDR2, and CDR3, from light chain variable region 52 (SEQ ID NO:52), 155 (SEQ ID NO: 155), or 45 (SEQ ID NO:45), or a structurally or functionally related variable light chain region as described herein.
• the antibody molecule comprises a heavy chain variable region from Fig 2 or Fig 13 or a structurally or functionally related variable heavy chain region as described herein. In an embodiment the antibody molecule comprises a light chain variable region from Fig 3 or Fig 14 or a structurally or functionally related variable light chain region as described herein. In an embodiment the antibody molecule comprises one, two, or all of, a CDRl,
• CDR2, and CDR3 from a heavy chain variable region from Fig 2 or Fig 13, or a structurally or functionally related sequences as described herein.
• the antibody molecule comprises one, two, or all of, a CDRl, CDR2, and CDR3 from a light chain variable region from Fig 3 or Fig 14, or a structurally or functionally related sequences as described herein.
• the antibody molecule comprises one, two or all of, HC CDRl, HC CDR2, and HC CDR3 and one, two or all of, LC CDRl, LC CDR2, and LC CDR3 from an antibody disclosed in Table 3, or a structurally or functionally related sequences as described herein.
• the antibody molecule comprises the light chain LC45 (SEQ ID NO: 45). In yet another embodiment, the antibody comprises the light chain LC45, and the heavy chain HC25 (SEQ ID NO: 25) or 24 (SEQ ID NO: 24). In one embodiment, the antibody molecule comprises the light chain Ab032(SEQ ID NO: 45) and the heavy chain 25 (SEQ ID NO: 25). In yet another embodiment, the antibody molecule comprises light chain LC52(SEQ ID NO: 52) and heavy chain HC25 (SEQ ID NO: 25).
• the antibody molecule comprises one or more or all of FR1, FR2, FR3, or FR4, or FR sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from a heavy chain disclosed herein; and
• the antibody molecule comprises one or more or all of FR1, FR2, FR3, or FR4, or FR sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from light chain disclosed herein.
• an anti-HA antibody molecule featured in the disclosure, or preparation, or isolated preparation thereof comprises
• a heavy chain immunoglobulin variable domain comprising a sequence at least 60, 70, 80, 85, 87, 90, 95, 97, 98, or 99, e.g., 90%, homologous, to a heavy chain consensus sequence provided herein, e.g., the heavy chain consensus sequence provided in Fig. 2 or Fig 13, e.g., the heavy chain consensus sequence provided in Fig. 2, SEQ ID NO: 161; and
• a light chain immunoglobulin variable domain comprising a sequence at least 60, 70, 80, 85, 87, 90, 95, 97, 98, or 99, e.g., 95%, homologous, to a light chain consensus sequence provided herein, e.g., the light chain consensus sequence provided in Fig. 3 or Fig 14, e.g., the light chain consensus sequence provided in Fig. 3, SEQ ID NO:62.
• the anti-HA antibody molecule featured in the disclosure comprises one or both of:
• a light chain immunoglobulin variable domain comprising the sequence SEQ ID NO:62, or a sequence at least 95% identical to SEQ ID NO:62.
• influenza virus e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an H1N1 strain, e.g., A/South
• (ii) produces fewer escape mutants than does a reference anti-HA antibody molecule, e.g., Ab 67-11, FI6, FI28, C179, F10, CR9114, or CR6261, such as when tested by the method described in (i).
• a reference anti-HA antibody molecule e.g., Ab 67-11, FI6, FI28, C179, F10, CR9114, or CR6261
• the disclosure features an antibody molecule comprising one or both of:
• a heavy chain immunoglobulin variable region comprising the sequence of SEQ ID NO: 161, or a sequence that differs from SEQ ID NO: 161 by not more than 1, 2, 3, 4, 5, 6, 8, 10, 11, 12, 13, 14, 15 or 16 , e.g., by no more than 2, 3, 4, or 5 amino acids, e.g., conservative amino acids;
• a light chain immunoglobulin variable domain comprising the sequence SEQ ID NO: 62, or a sequence that differs from SEQ ID NO: 62 that differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., conservative amino acids.
• the 1, 2, 3, 4, 5, 6, 8, 10, 11, 12, 13, 14, 15 or 16 amino acid differences, e.g., conservative amino acid differences, in the heavy chain immunoglobulin variable region are in the FR regions of the heavy chain
• the 1, 2, 3, 4 or 5 amino acid differences, e.g., conservative amino acid differences, in the light chain immunoglobulin variable domain are in the FR regions of the light chain
• immunoglobulin variable region are conservative amino acid changes.
• the binding agent e.g., an antibody molecule
• binds to an epitope e.g., it has an epitope that overlaps with or is the same as, of an antibody disclosed herein, e.g., as determined by mutational analysis or crystal structure analysis.
• the antibody molecule comprises one or both of:
• FRs framework regions from heavy chain consensus sequence disclosed herein, e.g., the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from heavy chain consensus sequence disclosed herein; and
• FRs framework regions from light chain consensus sequence disclosed herein, e.g., the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from light chain consensus disclosed herein.
• the binding agent e.g., an antibody molecule, specifically binds the HA antigen.
• the disclosure features, a binding agent, e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of Ab 044.
• a binding agent e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of Ab 044.
• the antibody molecule competes with a reference antibody molecule, e.g., an antibody molecule described herein, for binding to a substrate, e.g., an HA.
• a reference antibody molecule e.g., an antibody molecule described herein
• the reference antibody molecule can be:
• an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO: 1
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y- A-D-S-V-Q-G
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W- L-S-Q-G-Y-F-N-P (SEQ ID NO:70); and ii) a light chain variable region segment comprising: a CDR1 comprising the sequence Q-S-I-T-F-D-Y-K-N-Y-L-A (SEQ ID NO: 145);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• an antibody molecule comprises one or both of: (i) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO: 25; and (ii) a light chain variable region segment comprising SEQ ID NO:52; or
• the HA can be from a Group 1 strain, e.g., an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/California/04/2009, or an H5N1 strain, e.g., A/Indonesia/5/2005 or A/Vietnam/1203/2004.
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecules or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, e.g.
• H1N1 strain e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009
• H5N1 strain e.g., A/Indonesia/5/2005 or
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more.
• the antibody molecule binds to the same epitope, or a portion thereof, which the reference antibody molecule binds. In an embodiment the antibody molecule does not bind to the same epitope, or a portion thereof, which the reference antibody molecule binds. In an embodiment the antibody molecule binds to the same epitope, or a portion thereof, on HA, as does a reference antibody molecule, e.g. an antibody molecule disclosed herein.
• the reference antibody molecule can be:
• an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-
• L-S-Q-G-Y-F-N-P (SEQ ID NO:70); and ii) a light chain variable region segment comprising:
• a CDR1 comprising the sequence Q-S-I-T-F-D-Y-K-N-Y-L-A (SEQ ID NO: 145);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• an antibody molecule comprises one or both of: (i) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO: 25; and (ii) a light chain variable region segment comprising SEQ ID NO:52; or
• the HA can be HA1 or HA5, e.g., from an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/California/04/2009, or an H5N1 strain, e.g., A/Indonesia/5/2005 or A/Vietnam/1203/2004Binding to the same epitope, or a portion thereof, can be shown by one or more of:
• a) mutational analysis e.g., binding to HA, or binding affinity for HA, is decreased or abolished if a residue is mutated;
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, from, e.g., an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009, or an H5N1 strain, e.g.,
• Reduction of the ability to bind can be evaluated by methods in the art.
• Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• a light chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 52.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• each HC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding CDR of SEQ ID NO: 25 and each LC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding CDR of SEQ ID NO: 52.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• antibody molecule comprises 1, 2, 3, 4, 5, or all of:
• a HC CDR1 comprising: S at the 1st position and A at the 3rd position in HC CDR1;
• a HC CDR2 comprising one or both, e.g., one of: V at the 2 nd position; or N at the 7 th position and Q at the 16 th position in HC CDR2;
• a HC CDR3 comprising: R at the 3rd position (and optionally, L at the 3 rd position);
• a LC CDR1 comprising one or both of, e.g., one of: I at the 3rd position; or D at the 6th position in LC CDR1;
• a LC CDR2 comprising one, two, or three of, e.g., one of: G at the 2 nd position; Y at the 4* position; or L at the 5 th position in LC CDR2;
• a LC CDR3 comprising: S at the 9 th position in LC CDR3;
• the binding agent e.g., an antibody molecule, comprises:
• a heavy chain immunoglobulin variable region segment comprising SEQ ID NO:25 (or a sequence that differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., conservative amino acids, therefrom);
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q-
• G-Y-F-N-P (SEQ ID NO:70) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom); and
• a CDR1 comprising the sequence:
• Q-S-I-T-F-D-Y-K-N- Y-L-A (SEQ ID NO: 145) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom).
• the binding agent e.g., an antibody molecule, comprises one or both of:
• LC CDRl-3 that collectively, differ from the AB 044 LC CDRl-3 by no more than, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, e.g., 1, 2, 3, or 4, amino acids, e.g., conservative amino acids;
• the antibody molecule comprises one or both of:
• the binding agent is an antibody molecule comprising one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68) (or a sequence that differs by no more than, 1, 2, or 3, e.g., 1 or 2, amino acids, e.g., conservative amino acids, there from, optionally provided that at least 1 or 2 of the highlighted residue are not changed, e.g., both S and A are not changed);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least 1, 2, or 3 of the highlighted residues are not changed, e.g., V or both N and Q or all three of V, N, and Q are not changed);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that R is not changed); and
• a CDR1 comprising the sequence:
• Q-S-I-T-F-D-Y-K-N-Y-L-A (SEQ ID NO: 145) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least lor 2 of the highlighted residues are not changed, e.g., I or D is not changed);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided thatat least 1, 2 or 3 of the highlighted residues are not changed, e.g., 1, 2 or all of G, Y, and L are not changed); a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least 1 or both of the highlighted residues are not changed, e.g., S is not changed).
• a CDR of the light or heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR, (i.e., while other residues in that CDR might be changed, the highlighted residue or combination of residues, are not changed).
• V or both N and Q, for heavy chain CDR2 are not changed.
• a CDR of the light and a CDR of the heavy chain each includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of two CDRs in the antibody molecule includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR. In embodiments both are in the light chain. In embodiments both are in the heavy chain.
• each of the three CDRs in the heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of the three CDRs in the light chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of the six CDRs in the heavy and light chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• the binding agent is an antibody molecule that comprises one or more or all of the following properties:
• the antibody molecule comprises 1, 2, 3, 4, 5, or all 6 properties selected from (a) to (f).
• the antibody molecule comprises a heavy chain having a one or more properties selected from (a), (b), and (c) and a light chain having one or more properties selected from (d), (e), and (f).
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68); a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70);
• a CDR1 comprising the sequence Q-S-I-T-F-D-Y-K-N-Y-L-A (SEQ ID NO: 1
• CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72); and a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• the antibody molecule comprises one or more or all of the following properties: (i) it fails to produce any escape mutants as determined by the failure of a viral titer to recover following at least 10, 9, 8, 7, 6, or 5 rounds of serial infections in cell culture with a mixture of the antibody molecule and an influenza virus (e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an HlNl strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/California/04/2009, or an H5N1 strain, e.g., A/Indonesia/5/2005 or A/Vietnam/1203/2004, or an influenza B virus, e.g., B/Wisconsin/1/2010); and (ii) it produces fewer escape mutants than does a reference anti-HA antibody molecule, such as Ab 67-11, FI6, FI28, C179, F10,
• an influenza virus
• FRs framework regions from SEQ ID NO: 25
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from SEQ ID NO: 25; and
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from SEQ ID NO: 52.
• the antibody molecule comprises:
• an FRl comprising the sequence Q-V-Q-L-L-E-T-G-G-G-L-V-K-P-G-Q-S-L- K-L-S-C-A-A-S-G-F-T-F- ⁇ (SEQ ID NO:74) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that T is not changed);
• an FR2 comprising the sequence W-V-R-Q-P-P-G-K-G-L-E- W-V-A (SEQ ID NO:75) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that W is not changed, or that if changed, is other than R);
• an FR3 comprising the sequence R-F-T-J-S-fl-D-N-S-K-N-T-L-Y-L-Q-M-N- S-L-R-A-E-D-T-A-V-Y-Y-C-A-K (SEQ ID NO:76) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that one, two or three of I, R, or L is not changed, or that if I is changed it is other than G, if R is changed it is other than P.
• an FR4 comprising the sequence W-G-Q-G-T-T-L-T-V-S-S (SEQ ID NO:77) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom) or W-G-Q-G-T-T-V-T-V-S-S(SEQ ID NO: 171) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom); and
• an FR1 comprising the sequence D-I-Q-M-T-Q-S-P-S-S-L-S-A-S-V-G-D-R- V-T-I-T-C-fl-S-S (SEQ ID NO:78) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that R is not changed);
• an FR2 comprising the sequence W-Y-Q-Q-K-P-G-K-A-P-K-L-L-I-Y (SEQ ID NO:79) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• an FR3 comprising the sequence G-V-P-S-R-F-S-G-S-G-S-G-T-D-F-T-L-T-I-
• S-S-L-Q-P-E-D-F-A-T-Y-Y- (SEQ ID NO:80) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that C is not changed, or if changed, is other than P); and
• an FR4 comprising the sequence F-G-Q-G-T-K-V-E-I-K (SEQ ID NO:81) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom).
• a FR of the light or heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR, (i.e., while other residues in that FR might be changed, the highlighted residue or combination of residues, are not changed).
• one, two or three of /, R, or L for heavy chain FR3 is not changed.
• a FR of the light and a FR of the heavy chain each includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• each of two FRs in the antibody molecule includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• both are in the light chain.
• both are in the heavy chain.
• each of FR2 and FR3 in the heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• each of FR1 and FR2 in the heavy and light chain includes one of the h ighlighted residues for that FR.
• sequence of FR1 of the heavy chain variable region segment is Q-V-Q-L-L-E-T-G-G-G-L-V-K-P-G-Q-S-L-K-L-S-C-A-A-S-G-F-T-F- ⁇ (SEQ ID NO:74).
• sequence of FR1 of the heavy chain variable region segment is E-V-Q-L-L-E-S-G-G-G-L-V-K-P-G-Q-S-L-K-L-S-C-A-A-S-G-F-T-F-J (SEQ ID NO: 173).
• the binding agent e.g., an antibody molecule
• an influenza virus e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an HlNl strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif orn
• hemagglutinin (HA) of an influenza B virus e.g., B/Wisconsin/1/2010; (1) it treats or prevents infection by an influenza B virus, e.g., B/Wisconsin/1/2010; (m) it is effective for prevention or treatment of infection, e.g., in humans or mice, with an influenza B virus, e.g., B/Wisconsin/1/2010 when administered at 50 mg/kg, 25 mg/kg, 10 mg/kg, 6 mg/kg, 5 mg/kg, 4 mg/kg, 3 mg/kg, 2 mg/kg or 1 mg/kg; (n) the concentration of antibody molecule required for 50% neutralization of influenza A virus is less than 10 ⁇ g/mL; (o) the concentration of antibody molecule required for 50% neutralization of influenza B virus, e.g., B/Wisconsin/1/2010, is less than 10 ⁇ g/mL; (p) it prevents or minimizes secondary infection (e.g., secondary bacterial infection) or effects thereof on
• the binding agent e.g., an antibody molecule, specifically binds the HA antigen.
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f:
• H3 HA2 residues A7, Ell, 118, D19, G20, W21, L38, K39, T41, Q42, A43, 145, 148, N49, L52,
• the antibody molecule has properties: a; and b.
• the antibody molecule has properties: c; and d.
• the antibody molecule has properties: a; and c or d.
• the antibody molecule has properties: b; and c or d.
• the antibody molecule has properties: c; and a or b.
• the antibody molecule has properties: d; and a or b. In an embodiment the antibody molecule has properties: a, b, c and d.
• the antibody molecule has properties: a, b, c, d, e, and f.
• the antibody molecule has a KD for H3 of equal to or less than 10 ⁇ 6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a KD for H3 of equal to or less than 10 "6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f:
• aa it includes one, two, or all of, HI HA1 residues H31, N279, and S292;
• dd it does not include one, two, three, four, or all of, HI HA2 residues Gl, L2, F3, G4, and D46;
• ee it includes one, two, or all of, HI HA1 residues T319, R322, and 1324 are bound by both Ab 044 and FI6; or ff) it includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or all of, HI HA2 residues A7, El l, 118, D19, G20, W21, Q38, K39, T41, Q42, N43, 145, 148, T49, V52, N53, 156, and E57.
• the antibody molecule has properties: aa; and bb.
• the antibody molecule has properties: cc; and dd.
• the antibody molecule has properties: aa; and cc or dd.
• the antibody molecule has properties: bb; and cc or dd. In an embodiment the antibody molecule has properties: cc; and aa or bb.
• the antibody molecule has properties: dd; and aa or bb.
• the antibody molecule has properties: aa, bb, cc and dd.
• the antibody molecule has properties: aa, bb, cc, dd, ee, and ff.
• the antibody molecule has a KD for HI of equal to or less than 10 "6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a K D for HI of equal to or less than 10 ⁇ 6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule has one, two, three or all of the following properties:
• the molecule has properties c, cc, d, and dd.
• the disclosure features, a binding agent, e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of Ab 069.
• a binding agent e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of Ab 069.
• the antibody molecule competes with a reference antibody molecule, e.g., an antibody molecule described herein, for binding to a substrate, e.g., an HA.
• a reference antibody molecule e.g., an antibody molecule described herein
• the reference antibody molecule can be:
• an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68); a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W- L-S-Q-G-Y-F-N-P (SEQ ID NO:70); and ii) a light chain variable region segment comprising:
• a CDR1 comprising the sequence Q-S-I-T-F-E-Y-K-N-Y-L-A (SEQ ID NO : 172);;
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• an antibody molecule comprises one or both of: (i) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO: 25; and (ii) a light chain variable region segment comprising SEQ ID NO: 155; or
• the HA can be HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more. In an embodiment the antibody molecule binds to the same epitope, or a portion thereof, which the reference antibody molecule binds. In an embodiment the antibody molecule does not bind to the same epitope, or a portion thereof, which the reference antibody molecule binds.
• the antibody molecule binds to the same epitope, or a portion thereof, on HA, as does a reference antibody molecule, e.g. an antibody molecule disclosed herein.
• the reference antibody molecule can be:
• an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W- L-S-Q-G-Y-F-N-P (SEQ ID NO:70); and ii) a light chain variable region segment comprising:
• a CDR1 comprising the sequence Q-S-I-T-F-E-Y-K-N-Y-L-A (SEQ ID NO : 172);;
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• an antibody molecule comprises one or both of: (i) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO: 25; and (ii) a light chain variable region segment comprising SEQ ID NO: 155; or
• the HA can be HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South
• Binding to the same epitope, or a portion thereof, can be shown by one or more of: a) mutational analysis, e.g., binding or lack thereof to mutant HA, e.g., if a residue is mutated;
• HA e.g., HA1 or HA5
• H1N1 strain e.g., A/South Carolina/1/1918, A/Puerto
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009, or an H5N1 strain, e.g.,
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• a light chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 155,
• each HC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding CDR of SEQ ID NO: 25 and each LC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding CDR of SEQ ID NO: 155.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• a light chain variable region comprising at least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 155,
• antibody molecule comprises 1, 2, 3, 4, 5, or all of:
• a HC CDR1 comprising: S at the 1st position and A at the 3rd position in HC CDR1;
• a HC CDR2 comprising one or both, e.g., one of: V at the 2nd position; or N at the 7 th position and Q at the 16 th position in HC CDR2;
• a HC CDR3 comprising: R at the 3rd position (and optionally, L at the 3 rd position);
• a LC CDR1 comprising one or both of, e.g., one of: ; I at the 3rd position; or E at the 6th position in LC CDR1;
• a LC CDR2 comprising one, two or three of, e.g., one of: G at the 2nd position; Y at the 4 th position; or L at the 5 th position in LC CDR2;
• a LC CDR3 comprising: S at the 9 th position in LC CDR3;
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain immunoglobulin variable region segment comprising SEQ ID NO:25 (or a sequence that differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., conservative amino acids, therefrom);
• the antibody molecule comprises one or both of:
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom); and
• a CDR1 comprising the sequence:
• Q-S-I-T-F-E-Y-K-N-Y-L-A (SEQ ID NO: 172) or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom); a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom).
• the binding agent e.g., an antibody molecule, comprises one or both of:
• LC CDRl-3 that collectively, differ from the AB 069 LC CDRl-3 by no more than, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, e.g., 1, 2, 3, or 4, amino acids, e.g., conservative amino acids;
• HC CDRl-3 that collectively, differ from the AB 069 HC CDRl-3 by no more than, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, e.g., 1, 2, 3, or 4, amino acids, e.g., conservative amino acids.
• the binding agent is an antibody molecule comprising one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68) (or a sequence that differs by no more than, 1, 2, or 3, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least 1 or 2 of the highlighted residues are not changed, e.g., both S and A are not changed);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom , optionally provided that at least 1, 2, or 3 of the highlighted residues are not changed, e.g., V or both N and Q or all three of V, N, and Q are not changed);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom optionally provided that, R is not changed); and (b) a light chain variable region segment comprising
• a CDR1 comprising the sequence:
• Q-S-I-T-F-E-Y-K-N-Y-L-A (SEQ ID NO: 172) or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least 1 or 2 of the highlighted residues are not changed, e.g., I_or E is not changed);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided thatat least 1, 2, or 3 of the highlighted residues are not changed, e.g., 1, 2 or all of G, Y, and L are not changed);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P ⁇ (SEQ ID NO:73) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that, at least one or both of the highlighted residues are not changed, e.g., S is not changed).
• a CDR of the light or heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR, (i.e., while other residues in that CDR might be changed, the highlighted residue or combination of residues, are not changed).
• a CDR of the light and a CDR of the heavy chain each includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of two CDRs in the antibody molecule includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR. In embodiments both are in the light chain. In embodiments both are in the heavy chain.
• each of the three CDRs in the heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of the three CDRs in the light chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of the six CDRs in the heavy and light chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• the binding agent is an antibody molecule that comprises one or more or all of the following properties:
• the antibody molecule comprises 1, 2, 3, 4, 5, or all 6 properties selected from (a) to (f).
• the antibody molecule comprises a heavy chain having a one or more properties selected from (a), (b), and (c) and a light chain having one or more properties selected from (d), (e), and (f).
• the antibody molecule comprises one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68); a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• the antibody molecule comprises one or more or all of the following properties: (i) it fails to produce any escape mutants as determined by the failure of a viral titer to recover following at least 10, 9, 8, 7, 6, or 5 rounds of serial infections in cell culture with a mixture of the antibody molecule and an influenza virus (e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an HlNl strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/California/04/2009, or an H5N1 strain, e.g., A/Indonesia/5/2005 or A/Vietnam/1203/2004, or an influenza B virus, e.g., B/Wisconsin/1/2010); and (ii) it produces fewer escape mutants than does a reference anti-HA antibody molecule, such as Ab 67-11, FI6, FI28, C179, F10,
• an influenza virus
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from SEQ ID NO: 25; and
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from SEQ ID NO: 155.
• the antibody molecule comprises:
• a heavy chain immunoglobulin variable region segment that further comprises one or more or all of: an FR1 comprising the sequence Q-V-Q-L-L-E-T-G-G-G-L-V-K-P-G-Q-S-L- K-L-S-C-A-A-S-G-F-T-F-r (SEQ ID NO:74) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that T is not changed);
• an FR2 comprising the sequence W-V-R-Q-P-P-G-K-G-L-E- W-V-A (SEQ ID NO:
• NO:75 (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that W is not changed, or that if changed, is other than R);
• an FR3 comprising the sequence R-F-T-J-S-fl-D-N-S-K-N-T-L-Y-L-Q-M-N- S-L-R-A-E-D-T-A-V-Y-Y-C-A-K (SEQ ID NO:76) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that one, two or three of I, R, or L is not changed, or that if I is changed it is other than G, if R is changed it is other than P. or if L is changed it is other than A); and
• the light chain immunoglobulin variable region segment comprises one or more or all of
• an FR1 comprising the sequence D-I-Q-M-T-Q-S-P-S-S-L-S-A-S-V-G-D-R- V-T-I-T-C-fl-S-S (SEQ ID NO:78) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that R is not changed);
• an FR2 comprising the sequence W-Y-Q-Q-K-P-G-K-A-P-K-L-L-I-Y (SEQ ID NO:79) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• an FR3 comprising the sequence G-V-P-S-R-F-S-G-S-G-S-G-T-D-F-T-L-T-I- S-S-L-Q-P-E-D-F-A-T-Y-Y- (SEQ ID NO:80) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that C is not changed, or if changed, is other than P); and
• an FR4 comprising the sequence F-G-Q-G-T-K-V-E-I-K (SEQ ID NO:81) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom).
• a FR of the light or heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR, (i.e., while other residues in that FR might be changed, the highlighted residue or combination of residues, are not changed).
• one, two or three of I, R, or L for heavy chain FR3 is not changed.
• a FR of the light and a FR of the heavy chain each includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• each of two FRs in the antibody molecule includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR. In embodiments both are in the light chain. In embodiments both are in the heavy chain.
• each of FR2 and FR3 in the heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• each of FR1 and FR2 in the heavy and light chain includes one of the h ighlighted residues for that FR.
• the binding agent e.g., an antibody molecule
• an influenza virus e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an HlNl strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif orn
• A/Vietnam/1203/2004, or an influenza B virus e.g., B/Wisconsin/l/2010
• it produces fewer escape mutants than does a reference anti-HA antibody molecule e.g., Ab 67-11, FI6, FI28, C179, or CR6261, e.g., when tested by the method described in (a);
• HA hemagglutinin
• the binding agent e.g., an antibody molecule, specifically binds the HA antigen.
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f:
• H3 HA2 residues A7, Ell, 118, D19, G20, W21, L38, K39, T41, Q42, A43, 145, 148, N49, L52,
• the antibody molecule has properties: a; and b.
• the antibody molecule has properties: c; and d.
• the antibody molecule has properties: a; and c or d.
• the antibody molecule has properties: b; and c or d.
• the antibody molecule has properties: c; and a or b.
• the antibody molecule has properties: d; and a or b.
• the antibody molecule has properties: a, b, c and d. In an embodiment the antibody molecule has properties: a, b, c, d, e, and f.
• the antibody molecule has a K D for H3 of equal to or less than 10 ⁇ 6 , wherein said K D is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a KD for H3 of equal to or less than 10 "6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f: aa) it includes one, two, or all of, HI HA1 residues H31, N279, and S292;
• dd it does not include one, two, three, four, or all of, HI HA2 residues Gl, L2, F3, G4, and D46; ee) it includes one, two, or all of, HI HA1 residues T319, R322, and 1324 are bound by both Ab 044 and FI6; or ff) it includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or all of, HI HA2 residues A7, El l, 118, D19, G20, W21, Q38, K39, T41, Q42, N43, 145, 148, T49, V52, N53, 156, and E57.
• the antibody molecule has properties: aa; and bb.
• the antibody molecule has properties: cc; and dd.
• the antibody molecule has properties: aa; and cc or dd.
• the antibody molecule has properties: bb; and cc or dd. In an embodiment the antibody molecule has properties: cc; and aa or bb.
• the antibody molecule has properties: dd; and aa or bb.
• the antibody molecule has properties: aa, bb, cc and dd.
• the antibody molecule has properties: aa, bb, cc, dd, ee, and ff.
• the antibody molecule has a KD for HI of equal to or less than 10 "6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a K D for HI of equal to or less than 10 ⁇ 6 , wherein said K D is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule has one, two, three or all of the following properties:
• the molecule has properties c, cc, d, and dd.
• the molecule has properties c, cc, d, and dd.
• the disclosure features, a binding agent, e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of Ab 032.
• a binding agent e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of Ab 032.
• the antibody molecule competes with a reference antibody molecule, e.g., an antibody molecule described herein, for binding to a substrate, e.g., an HA.
• a reference antibody molecule e.g., an antibody molecule described herein
• the reference antibody molecule can be:
• an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y- A-D-S-V-Q-G (SEQ ID NO: 69);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W- L-S-Q-G-Y-F-N-P (SEQ ID NO:70); and ii) a light chain variable region segment comprising:
• a CDR1 comprising the sequence Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO: 71);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• an antibody molecule comprises one or both of: (i) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO: 25; and (ii) a light chain variable region segment comprising SEQ ID NO:45; or
• the HA can be HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, e.g.
• H1N1 strain e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009
• H5N1 strain e.g., A/Indonesia/5/2005 or
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more.
• the antibody molecule binds to the same epitope, or a portion thereof, which the reference antibody molecule binds. In an embodiment the antibody molecule does not bind to the same epitope, or a portion thereof, which the reference antibody molecule binds.
• the antibody molecule binds to the same epitope, or a portion thereof, on HA, as does a reference antibody molecule, e.g. an antibody molecule disclosed herein.
• the reference antibody molecule can be:
• an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W- L-S-Q-G-Y-F-N-P (SEQ ID NO:70); and ii) a light chain variable region segment comprising:
• a CDR1 comprising the sequence Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO: 71);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• an antibody molecule comprises one or both of: (i) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO: 25; and (ii) a light chain variable region segment comprising SEQ ID NO:45; or
• the HA can be HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South
• Binding to the same epitope, or a portion thereof, can be shown by one or more of:
• a) mutational analysis e.g., binding to HA, or binding affinity for HA, is decreased or abolished if a residue is mutated
• analysis e.g., comparison, of the crystal structure of the antibody molecule and HA and the crystal structure of a reference antibody and HA, e.g., to determine the touch points of each;
• HA e.g., HA1 or HA5
• H1N1 strain e.g., A/South Carolina/1/1918, A/Puerto
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, from, e.g., an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009, or an H5N1 strain, e.g.,
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• a light chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 45.
• the binding agent e.g., an antibody molecule, comprises one or both of: a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• each HC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding CDR of SEQ ID NO: 25 and each LC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding CDR of SEQ ID NO: 45.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• antibody molecule comprises 1, 2, 3, 4, 5, or all of:
• a HC CDR1 comprising: S at the 1st position and A at the 3rd position in HC CDR1;
• a HC CDR2 comprising one or both, e.g., one of: V at the 2 nd position; or N at the 7 th position and Q at the 16 th position in HC CDR2;
• a HC CDR3 comprising: R at the 3rd position (and optionally, L at the 3 rd position);
• a LC CDR2 comprising one, two, or three of, e.g., one of: G at the 2 nd position; Y at the 4* position; or L at the 5 th position in LC CDR2;
• a LC CDR3 comprising: S at the 9 th position in LC CDR3;
• the binding agent e.g., an antibody molecule, comprises one or both of: (a) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO:25 (or a sequence that differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., conservative amino acids, therefrom); and
• a light chain variable region segment comprising SEQ ID NO: 155 (or a sequence that differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., conservative amino acids, therefrom).
• the antibody molecule comprises one or both of:
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom); and
• a CDR1 comprising the sequence:
• Q-S-I-T-F N-Y-K-N-Y-L-A (SEQ ID NO:71) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72)
• the binding agent e.g., an antibody molecule, comprises one or both of:
• LC CDRl-3 that collectively, differ from the AB 032 LC CDRl-3 by no more than, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, e.g., 1, 2, 3, or 4, amino acids, e.g., conservative amino acids;
• HC CDRl-3 that collectively, differ from the AB 032 HC CDRl-3 by no more than, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, e.g., 1, 2, 3, or 4, amino acids, e.g., conservative amino acids.
• the binding agent is an antibody molecule comprising one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68) (or a sequence that differs by no more than, 1, 2, or 3, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least 1 or 2 of the highlighted residues are not changed, e.g., both S and A are not changed);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, provided that, e,g., at least 1, 2, or 3 of the highlighted residues are not changed, e.g., V or both N and Q or all three of V, N, and Q are not changed);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that R is not changed); and
• a CDR1 comprising the sequence: Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO: 71) or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided thatat least 1 or 2 of the highlighted residues are not changed, e.g., I is not changed);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least 1, 2, or 3 of the highlighted residues are not changed, e.g., 1, 2 or all of G, Y, and L are not changed);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P ⁇ (SEQ ID NO:73) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least one or both of the highlighted residues are not changed, e.g., S is not changed).
• a CDR of the light or heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR, (i.e., while other residues in that CDR might be changed, the highlighted residue or combination of residues, are not changed).
• a CDR of the light and a CDR of the heavy chain each includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of two CDRs in the antibody molecule includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR. In embodiments both are in the light chain. In embodiments both are in the heavy chain.
• each of the three CDRs in the heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of the three CDRs in the light chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of the six CDRs in the heavy and light chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• the binding agent is an antibody molecule that comprises one or more or all of the following properties:
• the antibody molecule comprises 1, 2, 3, 4, 5, or all 6 properties selected from (a) to (f).
• the antibody molecule comprises a heavy chain having a one or more properties selected from (a), (b), and (c) and a light chain having one or more properties selected from (d), (e), and (f).
• the antibody molecule comprises one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68); a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70);
• a CDR1 comprising the sequence Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO: 1
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• the antibody molecule comprises one or more or all of the following properties: (i) it fails to produce any escape mutants as determined by the failure of a viral titer to recover following at least 10, 9, 8, 7, 6, or 5 rounds of serial infections in cell culture with a mixture of the antibody molecule and an influenza virus (e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an HlNl strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/California/04/2009, or an H5N1 strain, e.g., A/Indonesia/5/2005 or A/Vietnam/1203/2004, or an influenza B virus, e.g., B/Wisconsin/1/2010); and (ii) it produces fewer escape mutants than does a reference anti-HA antibody molecule, such as Ab 67-11, FI6, FI28, C179,
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from SEQ ID NO: 25; and
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from SEQ ID NO: 45.
• the antibody molecule comprises:
• an FRl comprising the sequence Q-V-Q-L-L-E-T-G-G-G-L-V-K-P-G-Q-S-L- K-L-S-C-A-A-S-G-F-T-F-r (SEQ ID NO:74) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that T is not changed);
• an FR2 comprising the sequence W-V-R-Q-P-P-G-K-G-L-E- W-V-A (SEQ ID NO:
• NO:75 (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that W is not changed, or that if changed, is other than R);
• an FR3 comprising the sequence R-F-T-J-S-fl-D-N-S-K-N-T-L-Y-L-Q-M-N- S-L-R-A-E-D-T-A-V-Y-Y-C-A-K (SEQ ID NO:76) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2, amino acids, e.g., conservative amino acids, therefrom, optionally provided that one, two or three of I, R, or L is not changed, or that if I is changed it is other than G, if R is changed it is other than P. or if L is changed it is other than A); and
• an FR4 comprising the sequence W-G-Q-G-T-T-L-T-V-S-S (SEQ ID NO:77) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom)or W-G-Q-G-T-T-V-T-V-S-S (SEQ ID NO: 171) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom); and
• the light chain immunoglobulin variable region segment comprises one or more or all of
• an FR1 comprising the sequence D-I-Q-M-T-Q-S-P-S-S-L-S-A-S-V-G-D-R- V-T-I-T-C-fl-S-S (SEQ ID NO:78) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that R is not changed);
• an FR2 comprising the sequence W-Y-Q-Q-K-P-G-K-A-P-K-L-L-I-Y (SEQ ID NO:
• ID NO:79 (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• an FR3 comprising the sequence G-V-P-S-R-F-S-G-S-G-S-G-T-D-F-T-L-T-I- S-S-L-Q-P-E-D-F-A-T-Y-Y- (SEQ ID NO:80) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that C is not changed, or if changed, is other than P); and
• an FR4 comprising the sequence F-G-Q-G-T-K-V-E-I-K (SEQ ID NO:81) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom).
• a FR of the light or heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR, (i.e., while other residues in that FR might be changed, the highlighted residue or combination of residues, are not changed).
• one, two or three of I, R, or L for heavy chain FR3 is not changed.
• a FR of the light and a FR of the heavy chain each includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• each of two FRs in the antibody molecule includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR. In embodiments both are in the light chain. In embodiments both are in the heavy chain.
• each of FR2 and FR3 in the heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• each of FR1 and FR2 in the heavy and light chain includes one of the h ighlighted residues for that FR.
• the binding agent e.g., an antibody molecule
• an influenza virus e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an HlNl strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif orn
• A/Vietnam/1203/2004, or an influenza B virus e.g., B/Wisconsin/l/2010
• it produces fewer escape mutants than does a reference anti-HA antibody molecule e.g., Ab 67-11, FI6, FI28, C179, or CR6261, e.g., when tested by the method described in (a);
• HA hemagglutinin
• the binding agent e.g., an antibody molecule, specifically binds the HA antigen.
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f:
• H3 HA2 residues A7, Ell, 118, D19, G20, W21, L38, K39, T41, Q42, A43, 145, 148, N49, L52,
• the antibody molecule has properties: a; and b.
• the antibody molecule has properties: c; and d.
• the antibody molecule has properties: a; and c or d.
• the antibody molecule has properties: b; and c or d.
• the antibody molecule has properties: c; and a or b.
• the antibody molecule has properties: d; and a or b.
• the antibody molecule has properties: a, b, c and d. In an embodiment the antibody molecule has properties: a, b, c, d, e, and f.
• the antibody molecule has a K D for H3 of equal to or less than 10 ⁇ 6 , wherein said K D is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a KD for H3 of equal to or less than 10 "6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f: aa) it includes one, two, or all of, HI HA1 residues H31, N279, and S292;
• dd it does not include one, two, three, four, or all of, HI HA2 residues Gl, L2, F3, G4, and D46; ee) it includes one, two, or all of, HI HA1 residues T319, R322, and 1324 are bound by both Ab 044 and FI6; or ff) it includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or all of, HI HA2 residues A7, El l, 118, D19, G20, W21, Q38, K39, T41, Q42, N43, 145, 148, T49, V52, N53, 156, and E57.
• the antibody molecule has properties: aa; and bb.
• the antibody molecule has properties: cc; and dd.
• the antibody molecule has properties: aa; and cc or dd.
• the antibody molecule has properties: bb; and cc or dd. In an embodiment the antibody molecule has properties: cc; and aa or bb.
• the antibody molecule has properties: dd; and aa or bb.
• the antibody molecule has properties: aa, bb, cc and dd.
• the antibody molecule has properties: aa, bb, cc, dd, ee, and ff.
• the antibody molecule has a KD for HI of equal to or less than 10 "6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a K D for HI of equal to or less than 10 ⁇ 6 , wherein said K D is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule has one, two, three or all of the following properties:
• the molecule has properties c, cc, d, and dd.
• the disclosure features, a binding agent, e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of Ab 031.
• a binding agent e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of Ab 031.
• the antibody molecule competes with a reference antibody molecule, e.g., an antibody molecule described herein, for binding to a substrate, e.g., an HA.
• a reference antibody molecule e.g., an antibody molecule described herein
• the reference antibody molecule can be:
• an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y- A-D-S-V-Q-G
• a CDR1 comprising the sequence Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO:71);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• an antibody molecule comprises one or both of: (i) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO: 24; and (ii) a light chain variable region segment comprising SEQ ID NO:45; or
• the HA can be HAl or HA5, e.g. from an H1N1 strain, e.g., A/South
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HAl or HA5, e.g.
• H1N1 strain e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009
• H5N1 strain e.g., A/Indonesia/5/2005 or
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more.
• the antibody molecule binds to the same epitope, or a portion thereof, which the reference antibody molecule binds.
• the antibody molecule does not bind to the same epitope, a portion thereof, which the reference antibody molecule binds. In an embodiment the antibody molecule binds to the same epitope, or a portion thereof, on HA, as does a reference antibody molecule, e.g. an antibody molecule disclosed herein.
• the reference antibody molecule can be:
• an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W- L-S-Q-G-Y-F-N-P (SEQ ID NO:70); and ii) a light chain variable region segment comprising:
• a CDR1 comprising the sequence Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO:71);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73).
• an antibody molecule comprises one or both of: (i) a heavy chain immunoglobulin variable region segment comprising SEQ ID NO: 24; and (ii) a light chain variable region segment comprising SEQ ID NO:45; or
• the HA can be HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South
• Binding to the same epitope, or a portion thereof, can be shown by one or more of:
• a) mutational analysis e.g., binding to HA, or binding affinity for HA, is decreased or abolished if a residue is mutated;
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, from, e.g., an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009, or an H5N1 strain, e.g.,
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 24;
• a light chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 45.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 24;
• each HC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding CDR of SEQ ID NO: 24 and each LC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding CDR of SEQ ID NO: 45.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with SEQ ID NO: 25;
• antibody molecule comprises 1, 2, 3, 4, 5, or all of:
• a HC CDR1 comprising: S at the 1st position and A at the 3rd position in HC CDR1;
• a HC CDR2 comprising one or both, e.g., one of: V at the 2nd position; or N at the 7 th position and Q at the 16 th position in HC CDR2;
• a HC CDR3 comprising: R at the 3rd position (and optionally, L at the 3 rd position);
• a LC CDR2 comprising one, two, or three of, e.g., one of: G at the 2nd position; Y at the 4 th position; or L at the 5 th position in LC CDR2;
• a LC CDR3 comprising: S at the 9 th position in LC CDR3;
• the binding agent comprises an antibody molecule comprising:
• a heavy chain immunoglobulin variable region segment comprising SEQ ID NO:24(or a sequence that differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., conservative amino acids, therefrom);
• the antibody molecule comprises one or both of:
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom); and
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom); and
• a CDR1 comprising the sequence Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO:71) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO: 72) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom); and
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P-S (SEQ ID NO:73) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom).
• the binding agent e.g., an antibody molecule, comprises one or both of: a) LC CDRl-3, that collectively, differ from the AB 031 LC CDRl-3 by no more than, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, e.g., 1, 2, 3, or 4, amino acids, e.g., conservative amino acids; and
• HC CDRl-3 that collectively, differ from the AB 031 HC CDRl-3 by no more than, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, e.g., 1, 2, 3, or 4, amino acids, e.g., conservative amino acids.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain immunoglobulin variable region segment comprising a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68) (or a sequence that differs by no more than, 1, 2, or 3, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided thatat least 1 or 2 of the highlighted residues are not changed, e.g., both S and A are not changed);
• a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom , provided that, e,g., at least 1, 2, or 3 of the highlighted residues are not changed, e.g., V or both N and Q or all three of V, N, and Q are not changed);
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom optionally provided that, e.g., R is not changed); and
• a CDR1 comprising the sequence Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO: 71) or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided thatat least 1 or 2 of the highlighted residues are not changed, e.g., I is not changed);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least 1, 2, or 3 of the highlighted residues are not changed, e.g., 1, 2 or all of G, Y, and L are not changed);
• a CDR3 comprising the sequence Q-Q-H-Y-R-T-P-P ⁇ (SEQ ID NO:73) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that at least one or both of the highlighted residues are not changed, e.g., S is not changed).
• a CDR of the light or heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR, (i.e., while other residues in that CDR might be changed, the highlighted residue or combination of residues, are not changed).
• a CDR of the light and a CDR of the heavy chain each includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of two CDRs in the antibody molecule includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR. In embodiments both are in the light chain. In embodiments both are in the heavy chain.
• each of the three CDRs in the heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of the three CDRs in the light chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• each of the six CDRs in the heavy and light chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that CDR.
• the binding agent is an antibody molecule that comprises one or more or all of the following properties:
• both S and A in HC CDR1 are unchanged.
• V or both N and Q or all three of V, N, and Q in HC CDR2 are unchanged.
• the antibody molecule comprises 1, 2, 3, 4, 5, or all 6 properties selected from (a) to (f).
• the antibody molecule comprises a heavy chain having a one or more properties selected from (a), (b), and (c) and a light chain having one or more properties selected from (d), (e), and (f).
• the antibody molecule comprises one or both of:
• a CDR1 comprising the sequence S-Y-A-M-H (SEQ ID NO:68); a CDR2 comprising the sequence V-V-S-Y-D-G-N-Y-K-Y-Y-A-D-S- V-Q-G (SEQ ID NO:69); and
• a CDR3 comprising the sequence D-S-R-L-R-S-L-L-Y-F-E-W-L-S-Q- G-Y-F-N-P (SEQ ID NO:70);
• a CDR1 comprising the sequence Q-S-I-T-F-N-Y-K-N-Y-L-A (SEQ ID NO:71);
• a CDR2 comprising the sequence W-G-S-Y-L-E-S (SEQ ID NO:72);
• the antibody molecule comprises one or more or all of the following properties: (i) it fails to produce any escape mutants as determined by the failure of a viral titer to recover following at least 10, 9, 8, 7, 6, or 5 rounds of serial infections in cell culture with a mixture of the antibody molecule and an influenza virus (e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an HlNl strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/California/04/2009, or an H5N1 strain, e.g., A/Indonesia/5/2005 or A/Vietnam/1203/2004, or an influenza B virus, e.g., B/Wisconsin/1/2010); and (ii) it produces fewer escape mutants than does a reference anti-HA antibody molecule, e.g., Ab 67-11, FI6, FI28, C179,
• the antibody molecule comprises one or both of:
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from SEQ ID NO: 24; and
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from SEQ ID NO: 45.
• the antibody molecule comprises:
• an FRl comprising the sequence E-V-Q-L-L-E-S-G-G-G-L-V-K-P-G-Q-S-L- K-L-S-C-A-A-S-G-F-T-F-r (SEQ ID NO:82) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids,
• an FR2 comprising the sequence W-V-R-Q-P-P-G-K-G-L-E- W-V-A (SEQ ID NO:75) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that W is not changed, or that if changed, is other than R);
• an FR3 comprising the sequence R-F-T-J-S-fl-D-N-S-K-N-T-L-Y-L-Q-M-N- S-L-R-A-E-D-T-A-V-Y-Y-C-A-K (SEQ ID NO:76) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that one, two or three of I, R, or L is not changed, or that if I is changed it is other than G, if R is changed it is other than P. or if L is changed it is other than A); and
• an FR4 comprising the sequence W-G-Q-G-T-T-L-T-V-S-S (SEQ ID NO:77)
• W-G-Q-G-T-T-V-T-V-S-S SEQ ID NO: 171
• SEQ ID NO: 171 W-G-Q-G-T-T-V-T-V-S-S
• a light chain immunoglobulin variable region segment further comprises one or more or all of:
• an FR1 comprising the sequence D-I-Q-M-T-Q-S-P-S-S-L-S-A-S-V-G-D-R- V-T-I-T-C-fl-S-S (SEQ ID NO:78) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, , optionally provided that R is not changed);
• an FR2 comprising the sequence W-Y-Q-Q-K-P-G-K-A-P-K-L-L-I-Y (SEQ ID NO:79) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom);
• an FR3 comprising the sequence G-V-P-S-R-F-S-G-S-G-S-G-T-D-F-T-L-T-I- S-S-L-Q-P-E-D-F-A-T-Y-Y- (SEQ ID NO:80) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom, optionally provided that C is not changed, or if changed, is other than P); and
• an FR4 comprising the sequence F-G-Q-G-T-K-V-E-I-K (SEQ ID NO:81) (or a sequence that differs by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids, therefrom).
• a FR of the light or heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR, (i.e., while other residues in that FR might be changed, the highlighted residue or combination of residues, are not changed).
• one, two or three of /, R, or L for heavy chain FR3 is not changed.
• a FR of the light and a FR of the heavy chain each includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• each of two FRs in the antibody molecule includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR. In embodiments both are in the light chain. In embodiments both are in the heavy chain.
• each of FR2 and FR3 in the heavy chain includes one of the highlighted residues, or one of the highlighted combinations of residues, for that FR.
• each of FR1 and FR2 in the heavy and light chain includes one of the h ighlighted residues for that FR.
• the antibody molecule comprises:
• the heavy chain immunoglobulin variable region segment comprises one or more or all of
• an FR1 comprising the sequence E-V-Q-L-L-E-S-G-G-G-L-V-K-P-G- Q-S-L-K-L-S-C-A-A-S-G-F-T-F-KSEQ ID NO:82);
• an FR2 comprising the sequence W-V-R-Q-P-P-G-K-G-L-E-W-V-A (SEQ ID NO:75);
• an FR3 comprising the sequence R-F-T-I-S-R-D-N-S-K-N-T-L-Y-L- Q-M-N-S-L-R-A-E-D-T-A-V-Y-Y-C-A-K (SEQ ID NO:76); and
• an FR4 comprising the sequence W-G-Q-G-T-T-L-T-V-S-S (SEQ ID NO:77) or W-G-Q-G-T-T-V-T-V-S-S (SEQ ID NO: 171);
• an FR2 comprising the sequence W-Y-Q-Q-K-P-G-K-A-P-K-L-L-I-Y (SEQ ID NO: 79);
• an FR3 comprising the sequence G-V-P-S-R-F-S-G-S-G-S-G-T-D-F-
• an FR4 comprising the sequence F-G-Q-G-T-K-V-E-I-K (SEQ ID NO: 1
• the antibody molecule comprises one or more or all of the following properties: (a) it fails to produce any escape mutants as determined by the failure of a viral titer to recover following at least 10, 9, 8, 7, 6, or 5 rounds of serial infections in cell culture with a mixture of the antibody molecule and an influenza virus (e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/California/04/2009, or an H5N1 strain, e.g., A/Indonesia/5/2005 or A/Vietnam/1203/2004, or an influenza B virus, e.g., B/Wisconsin/1/2010); (b) it produces fewer escape mutants than does a reference anti-HA antibody molecule, e.g., Ab 67-11, FI6, FI28, C179
• B/Wisconsin/1/2010 when administered at 50 mg/kg, 25 mg/kg, 10 mg/kg, 6 mg/kg, 5 mg/kg, 4 mg/kg, 3 mg/kg, 2 mg/kg, or 1 mg/kg;
• the concentration of antibody molecule required for 50% neutralization of influenza A virus is less than 10 ⁇ g/mL;
• the concentration of antibody molecule required for 50% neutralization of influenza B virus, e.g., B/Wisconsin/1/2010 is less than 10 ⁇ g/mL;
• a heavy chain immunoglobulin variable region segment comprising SEQ ID NO:24 (or a sequence that differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., conservative amino acids, therefrom); and (b) a light chain variable region segment comprising SEQ ID NO:45 (or a sequence that differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., conservative amino acids, therefrom).
• the antibody molecule comprises one or more or all of the following properties: (i) it fails to produce any escape mutants as determined by the failure of a viral titer to recover following at least 10, 9, 8, 7, 6, or 5 rounds of serial infections in cell culture with a mixture of the antibody molecule and an influenza a virus, e.g., a Group 1 strain, e.g., an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or
• the binding agent e.g., an antibody molecule, specifically binds the HA antigen.
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f:
• H3 HA2 residues A7, Ell, 118, D19, G20, W21, L38, K39, T41, Q42, A43, 145, 148, N49, L52, N53, 156,and E57.
• the antibody molecule has properties: a; and b.
• the antibody molecule has properties: c; and d.
• the antibody molecule has properties: a; and c or d
• the antibody molecule has properties: b; and c or d
• the antibody molecule has properties: c; and a or b
• the antibody molecule has properties: d; and a or b In an embodiment the antibody molecule has properties: a, b, c and d.
• the antibody molecule has properties: a, b, c, d, e, and f .
• the antibody molecule has a KD for H3 of equal to or less than 10 ⁇ 6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a KD for H3 of equal to or less than 10 "6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f: aa) it includes one, two, or all of, HI HA1 residues H31, N279, and S292;
• dd it does not include one, two, three, four, or all of, HI HA2 residues Gl, L2, F3, G4, and D46; ee) it includes one, two, or all of, HI HA1 residues T319, R322, and 1324 are bound by both Ab 044 and FI6; or
• ff it includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or all of, HI HA2 residues A7, El l, 118, D19, G20, W21, Q38, K39, T41, Q42, N43, 145, 148, T49, V52, N53, 156, and E57.
• the antibody molecule has properties: aa; and bb.
• the antibody molecule has properties: cc; and dd.
• the antibody molecule has properties: aa; and cc or dd.
• the antibody molecule has properties: bb; and cc or dd.
• the antibody molecule has properties: cc; and aa or bb.
• the antibody molecule has properties: dd; and aa or bb.
• the antibody molecule has properties: aa, bb, cc and dd.
• the antibody molecule has properties: aa, bb, cc, dd, ee, and ff.
• the antibody molecule has a KD for HI of equal to or less than 10 "6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of: aa) HI HA1 residues H31, N279, and S292;
• the antibody molecule has a KD for HI of equal to or less than 10 ⁇ 6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule has one, two, three or all of the following properties:
• the molecule has properties c, cc, d, and dd.
• the disclosure features, a binding agent, e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of one or both a heavy chain variable region and a light chain variable region disclosed herein.
• a binding agent e.g., an antibody molecule, or preparation, or isolated preparation thereof, comprising a structural or functional property of one or both a heavy chain variable region and a light chain variable region disclosed herein.
• the antibody molecule competes with a reference antibody molecule, e.g., an antibody molecule described herein, for binding to a substrate, e.g., an HA.
• a reference antibody molecule e.g., an antibody molecule described herein
• the reference antibody molecule can be:
• an antibody molecule comprising the heavy and light CDRs from :
• an antibody molecule that comprises: (i) a heavy chain immunoglobulin variable region segment from Table 3, Table 4A, Table 4B, Fig.2, Fig 13, or Fig. 17; and (ii) a light chain variable region segment from Table 3, Table 4A,
• the HA can be HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, e.g.
• H1N1 strain e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009
• H5N1 strain e.g., A/Indonesia/5/2005 or
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more. In an embodiment the antibody molecule binds to the same epitope, or a portion thereof, which the reference antibody molecule binds. In an embodiment the antibody molecule does not bind to the same epitope, or a portion thereof, which the reference antibody molecule binds.
• the antibody molecule binds to the same epitope, or a portion thereof, on HA, as does a reference antibody molecule, e.g. an antibody molecule disclosed herein.
• the reference antibody molecule can be:
• an antibody molecule comprising the heavy and light CDRs from :
• an antibody molecule that comprises: (i) a heavy chain immunoglobulin variable region segment from Table 3, Table 4A, Table 4B, Fig.2, Fig 13, or Fig. 17; and (ii) a light chain variable region segment from Table 3, Table 4A,
• the HA can be HA1 or HA5, e.g. from an H1N1 strain, e.g., A/South
• Binding to the same epitope, or a portion thereof, can be shown by one or more of:
• a) mutational analysis e.g., binding to HA, or binding affinity for HA, is decreased or abolished if a residue is mutated;
• HA e.g., HA1 or HA5
• H1N1 strain e.g., A/South Carolina/1/1918, A/Puerto
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, from, e.g., an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/California/04/2009, or an H5N1 strain, e.g.,
• Reduction of the ability to bind can be evaluated by methods in the art. Reduction of the ability to bind can be evaluated, e.g., by one or more of:
• the antibody molecule can compete with the reference antibody such that binding of the reference antibody is decreased by 50% or more.
• HA e.g., HA1 or HA5
• H1N1 strain e.g., A/South Carolina/1/1918, A/Puerto
• Competition between the antibody molecule and a reference antibody molecule can be determined by evaluating the ability of one of the antibody molecule or the reference antibody molecule to decrease binding of the other to a substrate, e.g., HA, e.g., HA1 or HA5, from, e.g., an H1N1 strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif ornia/04/2009, or an H5N1 strain, e.g.,
• the binding agent e.g., an antibody molecule, comprises one or both of:
• a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with a reference heavy chain from Table 3, Table 4A, Table 4B, Fig.2, Fig 13 or Fig.17; and
• a light chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with reference light chain from Table 3, Table 4A, Table 4B, Fig.3, Fig. 14 or Fig 17,
• each HC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the corresponding HC CDR from its reference heavy chain and each LC CDR differs by no more than 1, 2, 3, 4 or 5 amino acids, e.g., 1 or 2, e.g., conservative amino acids, from the
• the binding agent e.g., an antibody molecule, comprises: a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with a heavy chain from Table 3 and a light chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with the corresponding light chain from Table 3.
• the binding agent e.g., an antibody molecule, comprises: a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with a heavy chain from Table 4A and a light chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with the corresponding light chain from Table 4A.
• the binding agent e.g., an antibody molecule, comprises: a heavy chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with a heavy chain from Table 4B and a light chain variable region comprising least 60, 70, 80, 85, 90, 95, 98 or 99 percent homology with the corresponding light chain from Table 4B.
• the binding agent e.g., an antibody molecule, comprises one or both of:
• the binding agent e.g., an antibody molecule, comprises: a heavy chain variable region from Table 3 and the corresponding light chain from Table 3;
• the binding agent e.g., an antibody molecule
• the binding agent comprises both of: (a) a heavy chain immunoglobulin variable region segment comprising a CDRl, a CDR2 and a CDR3 from a heavy chain sequence of Table 3, Table 4A, Table 4B, FIG. 2, FIG. 13, or FIG. 17 (or CDRs that, individually or collectively, differ therefrom by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids)); and
• a light chain immunoglobulin variable region segment comprising a CDRl, a CDR2 and a CDR3 from a light chain sequence of Table 3, Table 4A, Table 4B, FIG. 3, FIG. 14, or FIG. 17 (or CDRs that, individually or collectively, differ therefrom by no more than, 1, 2, 3, 4, or 5, e.g., 1 or 2 amino acids, e.g., conservative amino acids).
• the binding agent e.g., an antibody molecule, comprises one or both of:
• the binding agent e.g., an antibody molecule, comprises one or both of:
• the binding agent e.g., an antibody molecule, comprises one or both of:
• the binding agent e.g., an antibody molecule
• an influenza virus e.g., an influenza A virus, e.g., a Group 1 strain, e.g., an HlNl strain, e.g., A/South Carolina/1/1918, A/Puerto Rico/08/1934, or A/Calif orn
• a reference anti-HA antibody molecule e.g., Ab 67-11, FI6, FI28, C179, F10, CR9114, or CR6261, e.g., when tested by the method described in (i); and (iii) it is other than Ab 67-11 and FI6.
• the antibody molecule comprises one or both of:
• the antibody molecule comprises:
• the heavy chain immunoglobulin variable region further comprises an Isoleucine- Aspartate (He- Asp) dipeptide at the N-terminus.
• the light chain immunoglobulin variable region further comprises an Ile- Asp dipeptide at the N-terminus.
• both the heavy chain immunoglobulin variable region and the light chain immunoglobulin variable region or an antibody featured in the disclosure further comprises an lie- Asp dipeptide at the N-terminus.
• the He- Asp dipeptide is absent from one or both the heavy and light chain.
• the binding agent e.g., an antibody molecule, further comprises one or more or all of the following: (a) it treats or prevents infection by at least 1, 2, 3, 4 or 5 influenza subtypes of Group 1, and by at least 1, 2, 3, 4 or 5 influenza subtypes of Group 2; (b) it inhibits fusogenic activity of the targeted HA; (c) it treats or prevents infection by a Group 1 virus, wherein the virus is an HI, H5, or H9 virus; and treats or prevents infection by a Group 2 virus, wherein the virus is an H3 or H7 virus; (d) it treats or prevents infection by influenza A strains H1N1 and H3N2; (e) it is effective for prevention or treatment of infection, e.g., in humans or mice, with H1N1 and H3N2 when administered at 50 mg/kg, 25 mg/kg, 10 mg/kg, 6 mg/kg, 5 mg/kg, 4 mg/kg, 3 mg/kg, 2 mg/kg, or 1 mg/kg; (a) it treats
• B/Wisconsin/1/2010 treats or prevents infection by an influenza B virus, e.g., B/Wisconsin/1/2010; (j) it is effective for prevention or treatment of infection, e.g., in humans or mice, with an influenza B virus, e.g., B/Wisconsin/1/2010 when
• the concentration of antibody molecule required for 50% neutralization of influenza A virus is less than 10 ⁇ g/mL; (1) the concentration of antibody molecule required for 50% neutralization of influenza B virus, e.g.,
• B/Wisconsin/1/2010 is less than 10 ⁇ g/mL; (m) it prevents or minimizes secondary infection (e.g., secondary bacterial infection) or effects thereof on a subject; (n) it is effective for preventing or minimizing secondary infection (e.g., secondary bacterial infection) or effects thereof on a subject when administered at 50 mg/kg, 25 mg/kg, 10 mg/kg, 6 mg/kg, 5 mg/kg, 4 mg/kg, 3 mg/kg, 2 mg/kg, or 1 mg/kg; (o) it binds an epitope which comprises or consists of the hemagglutinin trimer interface; and (p) it binds an epitope other than that bound by a reference anti-HA antibody molecule, e.g., Ab 67-11, FI6, FI28, C179, F10, CR9114, or CR6261, e.g., when tested by a method disclosed herein, e.g., by competition in an ELISA assay.
• the antibody molecule comprises one or more or all of FR1, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from heavy chain disclosed herein; and
• the antibody molecule comprises one or more or all of FRl, FR2, FR3, or FR4, or sequences that differ individually, or collectively, by no more than 1, 2, 3, 4, of 5 amino acid residues, e.g., conservative residues, from light chain disclosed herein.
• the binding agent e.g., an antibody molecule, specifically binds the HA antigen.
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f:
• H3 HA2 residues A7, Ell, 118, D19, G20, W21, L38, K39, T41, Q42, A43, 145, 148, N49, L52,
• the antibody molecule has properties: a; and b.
• the antibody molecule has properties: c; and d.
• the antibody molecule has properties: a; and c or d In an embodiment the antibody molecule has properties: b; and c or d In an embodiment the antibody molecule has properties: c; and a or b In an embodiment the antibody molecule has properties: d; and a or b In an embodiment the antibody molecule has properties: a, b, c and d. In an embodiment the antibody molecule has properties: a, b, c, d, e, In an embodiment, the antibody molecule has a KD for H3 of equal to or less than 10 ⁇ 6 , wherein said KD is increased by at least 2, 5, 10, or 100 fold, by a mutation or mutations in any of:
• the antibody molecule has a KD for H3 of equal to or less than 10 "6 , wherein said KD is increased by no more than 2, or 5 fold, by a mutation or mutations in any of:
• the antibody molecule binds an epitope that has one, two, three, four, five, or all of, the following properties a-f: aa) it includes one, two, or all of, HI HA1 residues H31, N279, and S292;
• dd it does not include one, two, three, four, or all of, HI HA2 residues Gl, L2, F3, G4, and D46; ee) it includes one, two, or all of, HI HA1 residues T319, R322, and 1324 are bound by both Ab 044 and FI6; or
• the antibody molecule has properties: aa; and bb.
• the antibody molecule has properties: cc; and dd.
• the antibody molecule has properties: aa; and cc or dd.
• the antibody molecule has properties: bb; and cc or dd.
• the antibody molecule has properties: cc; and aa or bb. In an embodiment the antibody molecule has properties: dd; and aa or bb.
• the antibody molecule has properties: aa, bb, cc and dd.
• the antibody molecule has properties: aa, bb, cc, dd, ee, and ff.
• the antibody molecule has a KD for HI of equal to or less than 10 ⁇ 6 , wherein said KD is increased by at least 2, 5,10, or 100 fold, by a mutation or mutations in any of:

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