US2012128684A1PendingUtilityA1

Conserved Hemagglutinin Epitope, Antibodies to the Epitope and Methods of Use

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Assignee: MARASCO WAYNEPriority: Aug 25, 2008Filed: Aug 25, 2009Published: May 24, 2012
Est. expiryAug 25, 2028(~2.1 yrs left)· nominal 20-yr term from priority
C12N 2760/16034C07K 2317/92C07K 2299/00C07K 2317/76A61K 2039/505C12N 2760/16022C07K 2319/40A61P 31/16C07K 14/005A61P 37/04C07K 16/108A61K 39/00
59
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Claims

Abstract

Disclosed are antibodies that bind to the stem region of influenza hemagglutinin in the neutral pH conformation, hemagglutinin epitopes in the stem region, and methods of making and using both.

Claims

exact text as granted — not AI-modified
1 . An immunogen comprising an epitope or epitope unit recognized by a protective monoclonal antibody having the specificity for the stem region of hemagglutinn protein of an influenza virus. 
     
     
         2 . The immunogen of  claim 1 , wherein said antibody binds both the HA1 and HA2 peptide. 
     
     
         3 . The immunogen of  claim 1 , wherein said epitope is the F10 epitope. 
     
     
         4 . The immunogen of  claim 1 , wherein the antibody is monoclonal antibody D7, D8, F10, G17, H40, A66, D80, E88, E90, or H98 or a monoclonal antibody that competes with the binding of monoclonal antibody D7, D8, F10, G17, H40, A66, D80, E88, E90, or H98 to the HA protein. 
     
     
         5 . The immunogen of  claim 1 , wherein said hemagglutin protein is in the neutral pH conformation. 
     
     
         6 . The immunogen of  claim 1 , wherein said immunogen is a peptide or a synthetic peptide. 
     
     
         7 . The immunogen of  claim 1 , wherein the conformation of said epitope is defined by amino acid residues 18, 38, 39, 40 and 291 of HA1 and 18, 19, 20, 21, 38, 41, 42, 45, 49, 52, 53, and 56 of HA2 when said hemagglutinin in the neutral pH conformation. 
     
     
         8 . The immunogen of  claim 1 , wherein said immunogen comprises a peptide comprising one or more of the following amino acid sequences 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 125) 
                 
                     
                   a) [Xaa 0 ] m -Xaa 1 -Xaa 2 - [Xaa 0 ] p   
                 
             
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, 
         Xaa 0 , is independently any amino acid,
 Xaa 1  is S, T, F H or Y, and 
 Xaa 2  is H, Y, M, L or Q; 
 
       
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 126) 
                 
                     
                   b) [Xaa 0 ] m -Xaa 1 -Xaa 2 - [Xaa 0 ] p   
                 
             
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, 
         Xaa 0 , is independently any amino acid, and
 Xaa 1  is H, Q, Y, S, D, N or T, 
 Xaa 2  is Q, E, K, I, V, M, E, R or T; 
 
       
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 127) 
                 
                     
                   c) [Xaa 0 ] m -Xaa 1 -Xaa 2 -Xaa 3 -Xaa 4 - [Xaa 0 ] p   
                 
             
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, 
         Xaa 0 , is independently any amino acid, and
 Xaa 1  is I, V, M, or L; 
 Xaa 2  is D, N, H, Y, D, A, S or E, 
 Xaa 3  is G or A, and 
 Xaa 4  is W, R, or G; or 
 
       
       
         
           
                 
                 
               
                   d) 
                     
                 
                   (SEQ ID NO: 128) 
                     
                 
                   [Xaa 0 ] m -Xaa 1 -[Xaa 0 ] q  Xaa 2 -Xaa 3 -[Xaa 0 ] q  Xaa 4 -[Xaa 0 ] r  Xaa 5 -[Xaa 0 ] q -Xaa 6  Xaa 7  - 
                     
                 
                     
                 
                   [Xaa 0 ] q  -Xaa 8  -[Xaa 0 ] p   
                 
             
                
                
                
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, q is 2, and r is 3 
         Xaa 0 , is independently any amino acid, and
 Xaa 1  is K, Q, R, N, L, G, F, H or Y, 
 Xaa 2  is S or T, 
 Xaa 3  is Q or P, 
 Xaa 4  is F, V, I, M, L, or T, 
 Xaa 5  is I, T, S, N, Q, D, or A, 
 Xaa 6  is I, V, M, or L, 
 Xaa 7  is N, S, T, or D 
 Xaa 8  is I, F, V, A, or T; 
 
       
       
         
           
                 
                 
               
                   e) 
                     
                 
                   (SEQ ID NO: 129) 
                     
                 
                   [Xaa 0 ] m -Xaa 1 -[Xaa 0 ] q  Xaa 2 -Xaa 3 -[Xaa 0 ] q  Xaa 4 -[Xaa 0 ] r  Xaa 5 -[Xaa 0 ] q -Xaa 6  Xaa 7  - 
                     
                 
                     
                 
                   [Xaa 0 ] s  - [Xaa 8 ] t  -[Xaa 0 ] p   
                 
             
                
                
                
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, q is 2, r is 3, s is 0 or 2, and t is 0 or 1, 
         Xaa 0 , is independently any amino acid, and
 Xaa 1  is K, Q, R, N, L, G, F, H or Y, 
 Xaa 2  is S or T, 
 Xaa 3  is Q or P, 
 Xaa 4  is F, V, I, M, L, or T, 
 Xaa 5  is I, T, S, N, Q, D, or A, 
 Xaa 6  is I, V, M, or L, 
 Xaa 7  is N, S, T, or D, 
 Xaa 8  I, F, V, A, or T; 
 
       
     
     
         9 . A nucleic acid encoding the immunogen of  claim 8 . 
     
     
         10 . The immunogen of  claim 1 , further comprising an adjuvant. 
     
     
         11 . The immunogen of  claim 1 , wherein said immunogen is conjugated to a carrier. 
     
     
         12 . A composition comprising the immunogen of  claim 1  together with one or more pharmaceutically acceptable excipients, diluents, and/or adjuvants. 
     
     
         13 . The composition of  claim 12 , wherein said composition further comprises a anti-influenza antibody of antigen binding fragment thereof. 
     
     
         14 . The composition of  claim 13 , wherein said antibody is monoclonal antibody D7, D8, F10, G17, H40, A66, D80, E88, E90, or H98 or a monoclonal antibody that competes with the binding of monoclonal antibody D7, D8, F10, G17, H40, A66, D80, E88, E90, or H98 to the HA protein. 
     
     
         15 . A conjugate comprising one or more peptides or peptide fragments optionally linked to a backbone wherein the peptides or peptide fragments are spatially positioned relative to each other so that they together form a non-linear sequence which mimics the tertiary structure of an F10 epitope, wherein said conjugate competes with the binding of monoclonal antibody F10 to the HA protein. 
     
     
         16 . The conjugate of  claim 15 , wherein said peptide or peptide fragment comprises one or more of the following amino acids sequences: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 125) 
                 
                     
                   a) [Xaa 0 ] m -Xaa 1 -Xaa 2 - [Xaa 0 ] p   
                 
             
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, 
         Xaa 0 , is independently any amino acid,
 Xaa 1  is S, T, F H or Y, and 
 Xaa 2  is H, Y, M, L or Q; 
 
       
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 126) 
                 
                     
                   b) [Xaa 0 ] m -Xaa 1 -Xaa 2 - [Xaa 0 ] p   
                 
             
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, 
         Xaa 0 , is independently any amino acid, and
 Xaa 1  is H, Q, Y, S, D, N or T, 
 Xaa 2  is Q, E, K, I, V, M, E, R or T; 
 
       
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 127) 
                 
                     
                   c) [Xaa 0 ] m -Xaa 1 -Xaa 2 -Xaa 3 -Xaa 4 - [Xaa 0 ] p   
                 
             
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, 
         Xaa 0 , is independently any amino acid, and
 Xaa 1  is I, V, M, or L; 
 Xaa 2  is D, N, H, Y, D, A, S or E, 
 Xaa 3  is G or A, and 
 Xaa 4  is W, R, or G; or 
 
       
       
         
           
                 
                 
               
                   d) 
                     
                 
                   (SEQ ID NO: 128) 
                     
                 
                   [Xaa 0 ] m -Xaa 1 -[Xaa 0 ] q  Xaa 2 -Xaa 3 -[Xaa 0 ] q  Xaa 4 -[Xaa 0 ] r  Xaa 5 -[Xaa 0 ] q -Xaa 6  Xaa 7  - 
                     
                 
                     
                 
                   [Xaa 0 ] q  -Xaa 8  -[Xaa 0 ] p   
                 
             
                
                
                
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, q is 2, and r is 3 
         Xaa 0 , is independently any amino acid, and
 Xaa 1  is K, Q, R, N, L, G, F, H or Y, 
 Xaa 2  is S or T, 
 Xaa 3  is Q or P, 
 Xaa 4  is F, V, I, M, L, or T, 
 Xaa 5  is I, T, S, N, Q, D, or A, 
 Xaa 6  is I, V, M, or L, 
 Xaa 7  is N, S, T, or D 
 Xaa 8  is I, F, V, A, or T; 
 
       
       
         
           
                 
                 
               
                   e) 
                     
                 
                   (SEQ ID NO: 129) 
                     
                 
                   [Xaa 0 ] m -Xaa 1 -[Xaa 0 ] q  Xaa 2 -Xaa 3 -[Xaa 0 ] q  Xaa 4 -[Xaa 0 ] r  Xaa 5 -[Xaa 0 ] q -Xaa 6  Xaa 7  - 
                     
                 
                     
                 
                   [Xaa 0 ] s  - [Xaa 8 ] t  -[Xaa 0 ] p   
                 
             
                
                
                
                
                
               
            
           
         
         wherein, m, and p are independently 0 or 1-10, q is 2, r is 3, s is 0 or 2, and t is 0 or 1, 
         Xaa 0 , is independently any amino acid, and
 Xaa 1  is K, Q, R, N, L, G, F, H or Y, 
 Xaa 2  is S or T, 
 Xaa 3  is Q or P, 
 Xaa 4  is F, V, I, M, L, or T, 
 Xaa 5  is I, T, S, N, Q, D, or A, 
 Xaa 6  is I, V, M, or L, 
 Xaa 7  is N, S, T, or D, 
 Xaa 8  I, F, V, A, or T; 
 
       
     
     
         17 . A nucleic acid encoding the conjugate of  claim 16 . 
     
     
         18 . The conjugate of  claim 15 , further comprising an adjuvant. 
     
     
         19 . The conjugate of  claim 15 , wherein said conjugate is linked to a carrier. 
     
     
         20 . A composition comprising the conjugate of  claim 15  together with one or more pharmaceutically acceptable excipients, diluents, and/or adjuvants. 
     
     
         21 . The composition of  claim 20 , wherein said composition further comprises an anti-influenza antibody of antigen binding fragment thereof. 
     
     
         22 . The composition of  claim 21 , wherein said antibody is monoclonal antibody D7, D8, F10, G17, H40, A66, D80, E88, E90, or H98 or a monoclonal antibody that competes with the binding of monoclonal antibody D7, D8, F10, G17, H40, A66, D80, E88, E90, or H98 to the HA protein. 
     
     
         24 . A method preventing a disease or disorder caused by an influenza virus, comprising administering to person at risk of suffering from said disease or disorder the composition of  claim 12 . 
     
     
         25 . The method of  claim 24 , wherein the method further comprises administering an anti-viral drug, a viral entry inhibitor or a viral attachment inhibitor. 
     
     
         26 . The method of  claim 25 , wherein said anti-viral drug is a neuraminidase inhibitor, a HA inhibitor, a sialic acid inhibitor or an M2 ion channel. 
     
     
         27 . The method of  claim 26 , wherein said M2 ion channel inhibitor is amantadine or, rimantadine. 
     
     
         28 . The method of  claim 26 , wherein said neuraminidase inhibitor zanamivir, or oseltamivir phosphate. 
     
     
         29 . The method of  claim 24 , wherein the method comprises further administering one or more antibodies specific to a Group I influenza virus and or an Group II influenza virus 
     
     
         30 . The method of  claim 24 , wherein said antibody is administered prior to or after exposure to influenza virus. 
     
     
         31 . The method of  claim 29 , wherein said antibody is administered at a dose sufficient to neutralize said influenza virus. 
     
     
         32 . A method of treating a subject, the method comprising administering to the subject the stem region of influenza hemagglutinin in the neutral pH conformation in isolation from other components of influenza virus, wherein the subject produces an immune response to the stem region. 
     
     
         33 . A method of treating a subject, the method comprising administering to the subject the stem region of influenza hemagglutinin in the neutral pH conformation in isolation from the head region of hemagglutinin, wherein the subject produces an immune response to the stem region. 
     
     
         34 . A method of treating a subject, the method comprising administering to the subject influenza hemagglutinin in the neutral pH conformation in isolation from other components of influenza virus, wherein the head region of the hemagglutinin is modified to reduce the antigenicity of the head region, wherein the subject produces an immune response to the stem region. 
     
     
         35 . The method of  claim 34 , wherein the head region of the hemagglutinin is modified by removing or replacing glycosylation sites. 
     
     
         36 . The method of  claim 34 , wherein the head region of the hemagglutinin is modified by adding glycosylation sites. 
     
     
         37 . The method of  claim 34 , wherein the head region of the hemagglutinin is modified by removing all or a portion of the head region. 
     
     
         38 . The method of  claim 34 , wherein the subject produces an immune response that prevents or reduces the severity of an influenza infection. 
     
     
         39 . The method  claim 34 , wherein the immune response is reactive to influenza viruses within a subtype. 
     
     
         41 . The method  claim 34 , wherein the immune response is reactive to influenza viruses in each subtype within a cluster. 
     
     
         42 . The method  claim 34 , wherein the immune response is reactive to influenza viruses in each cluster within a group. 
     
     
         43 . The method  claim 34 , wherein the immune response is reactive to all influenza viruses in each subtype within a group. 
     
     
         44 . The method of  claim 43 , wherein the immune response is reactive to influenza viruses within group 1. 
     
     
         45 . A method, the method comprising screening antibodies reactive to hemagglutinin for binding to hemagglutinin immobilized on a surface, thereby identifying antibodies of interest. 
     
     
         46 . A method, the method comprising screening antibodies reactive to hemagglutinin for binding to the stem region of influenza hemagglutinin in the neutral pH conformation in isolation from the head region of hemagglutinin, thereby identifying antibodies of interest. 
     
     
         47 . A method, the method comprising screening antibodies reactive to hemagglutinin for binding to influenza hemagglutinin in the neutral pH conformation in isolation from other components of influenza virus, wherein the head region of the hemagglutinin is modified to reduce the antigenicity of the head region, thereby identifying antibodies of interest. 
     
     
         48 . The method of  claim 47 , wherein the head region of the hemagglutinin is modified by removing or replacing glycosylation sites. 
     
     
         49 . The method of  claim 47 , wherein the head region of the hemagglutinin is modified by adding glycosylation sites. 
     
     
         50 . The method of  claim 47 , wherein the head region of the hemagglutinin is modified by removing all or a portion of the head region. 
     
     
         51 . The method of  claim 47 , further comprising screening the antibodies of interest for competing with antibody F10 for binding to hemagglutinin, thereby identifying F10-competing antibodies. 
     
     
         52 . The method of  claim 47 , wherein the hemagglutinin is hemagglutinin from a group 2 influenza virus. 
     
     
         53 . The method of  claim 47 , wherein the hemagglutinin is hemagglutinin from a group 1 influenza virus. 
     
     
         54 . The method of  claim 47 , further comprising producing the identified antibodies. 
     
     
         55 . An antibody produced by the method of  claim 54 . 
     
     
         56 . A method of screening a compound for binding to an F10 antibody comprising contacting said F10 antibody with a compound of interest and detecting a compound-antibody complex.

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