Molecular antigen array

Abstract

The present invention is related to the fields of molecular biology, virology, immunology and medicine. The invention provides a composition comprising an ordered and repetitive antigen or antigenic determinant array. The invention also provides a process for producing an antigen or antigenic determinant in an ordered and repetitive array. The ordered and repetitive antigen or antigenic determinant is useful in the production of vaccines for the treatment of infectious diseases, the treatment of allergies and as a pharmaccine to prevent or cure cancer and to efficiently induce self-specific immune responses, in particular antibody responses.

1 . A composition comprising: 
 (a) a non-natural molecular scaffold comprising: 
 (i) a core particle comprising a virus-like particle of an RNA bacteriophage; and  
 (ii) an organizer comprising at least one first attachment site,  
 wherein said organizer is connected to said core particle by at least one covalent bond; and wherein said first attachment site is not a sulfhydryl group; and  
 (b) an antigen or antigenic determinant with at least one second attachment site,  
   wherein said second attachment site is capable of association through at least one non-peptide bond to said first attachment site; and    wherein said antigen or antigenic determinant and said scaffold interact through said association to form an ordered and repetitive antigen array.    
     
     
         2 . The composition of  claim 1 , wherein said RNA bacteriophage is selected from the group consisting of: 
 (a) bacteriophage Qβ;    (b) bacteriophage R17;    (c) bacteriophage fr;    (d) bacteriophage GA;    (e) bacteriophage SP;    (f) bacteriophage MS2;    (g) bacteriophage M11;    (h) bacteriophage MX1;    (i) bacteriophage NL95;    (j) bacteriophage f2; and    (k) bacteriophage PP7.    
     
     
         3 . The composition of  claim 1 , wherein said bacteriophage is bacteriophage Qβ.  
     
     
         4 . The composition of  claim 1 , wherein said virus-like particle of an RNA bacteriophage comprises recombinant proteins, or fragments thereof, of an RNA bacteriophage.  
     
     
         5 . The composition of  claim 4 , wherein said bacteriophage is bacteriophage Qβ.  
     
     
         6 . The composition of  claim 1 , wherein said virus-like particle of an RNA bacteriophage comprises recombinant coat proteins comprising an amino acid sequence selected from the group consisting of: 
 (a) SEQ ID NO:159;    (b) SEQ ID NO:160;    (c) SEQ ID NO:161;    (d) SEQ ID NO:162;    (e) SEQ ID NO:163;    (f) SEQ ID NO:164;    (g) SEQ ID NO:165;    (h) SEQ ID NO:166;    (i) SEQ ID NO:167;    (j) SEQ ID NO:215;    (k) SEQ ID NO:253;    (l) SEQ ID NO:217; and    (m) SEQ ID NO:254.    
     
     
         7 . The composition of  claim 1 , wherein said virus-like particle of an RNA bacteriophage comprises recombinant coat proteins having an amino acid sequence of SEQ ID NO:159, or a mixture of coat proteins having amino acid sequences of SEQ ID NO:159 and of SEQ ID NO:217.  
     
     
         8 . The composition of  claim 1 , wherein said virus-like particle of an RNA bacteriophage comprises one or more coat proteins of said RNA bacteriophage that have been modified by deletion or substitution to remove at least one naturally occurring lysine residue, or that have been modified by insertion or substitution to add at least one lysine residue.  
     
     
         9 . The composition of  claim 8 , wherein said RNA bacteriophage is Qβ.  
     
     
         10 . The composition of  claim 1 , wherein said virus-like particle of an RNA bacteriophage comprises one or more coat proteins comprising an amino acid sequence selected from the group consisting of: 
 (a) SEQ ID NO:255;    (b) SEQ ID NO:256;    (c) SEQ ID NO:257;    (d) SEQ ID NO:258;    (e) SEQ ID NO:259; and    (f) a mixture of any one of (a)-(e) and the corresponding A1 protein.    
     
     
         11 . The composition of  claim 1 , wherein said organizer is an integral part of said RNA bacteriophage.  
     
     
         12 . The composition of  claim 1 , wherein said association is by way of at least one covalent bond.  
     
     
         13 . The composition of  claim 1  further comprising an amino acid linker.  
     
     
         14 . The composition of  claim 13 , wherein said amino acid linker is bound to said antigen or said antigenic determinant by way of at least one covalent bond.  
     
     
         15 . The composition of  claim 14 , wherein said covalent bond is a peptide bond.  
     
     
         16 . The composition of  claim 13 , wherein said amino acid linker comprises said second attachment site.  
     
     
         17 . The composition of  claim 13 , wherein said amino acid linker comprises a sulfhydryl group.  
     
     
         18 . The composition of  claim 13 , wherein said amino acid linker comprises a cysteine residue.  
     
     
         19 . The composition of  claim 1 , wherein said first attachment site comprises: 
 (a) an amino group;    (b) a chemical group reactive to an amino group;    (c) a carboxyl group;    (d) a chemical group reactive to a carboxyl group;    (e) biotin;    (f) avidin;    (g) strepavidin; or    (h) interacting leucine zipper polypeptide.    
     
     
         20 . The composition of  claim 1 , wherein said second attachment site comprises: 
 (a) an amino group;    (b) a chemical group reactive to an amino group;    (c) a carboxyl group;    (d) a chemical group reactive to a carboxyl group;    (e) a sulfhydryl group;    (f) a chemical group reactive to a sulfhydryl group.    (g) biotin;    (h) avidin;    (i) strepavidin; or    (j) interacting leucine zipper polypeptide.    
     
     
         21 . The composition of  claim 1 , wherein said first attachment site and said second attachment site are associated through a heterobifunctional linker.  
     
     
         22 . The composition of  claim 21 , wherein said heterobifunctional linker is selected from the group consisting of: 
 (a) a maleimidocaproic acid N-hydroxysuccinimide ester;    (b) N-Succinimidyl 3-(2-pyridyldithio)propionate (SPDP); and    (c) Sulfo-MBS.    
     
     
         23 . The composition of  claim 1 , wherein said first attachment site comprises an amino group.  
     
     
         24 . The composition of  claim 1 , wherein said first attachment site is an amino group or is a lysine residue.  
     
     
         25 . The composition of  claim 1 , wherein said second attachment site comprises a sulfhydryl group.  
     
     
         26 . The composition of  claim 1 , wherein said second attachment site is a sulfhydryl group or is a cysteine residue.  
     
     
         27 . The composition of  claim 1 , wherein said second attachment site does not naturally occur within said antigen or antigenic determinant.  
     
     
         28 . The composition of  claim 1 , wherein said first attachment site comprises an amino group and said second attachment site comprises a sulfhydryl group.  
     
     
         29 . The composition of  claim 1 , wherein said first attachment site is an amino group and said second attachment site is a sulfhydryl group.  
     
     
         30 . The composition of  claim 1 , wherein said first attachment site is a lysine residue and said second attachment site is a cysteine residue.  
     
     
         31 . The composition of  claim 1 , wherein only one of said second attachment sites associates with said first attachment site through at least one non-peptide covalent bond leading to a single and uniform type of binding of said antigen to said core particle, wherein said only one second attachment site that associates with said first attachment site is a sulfhydryl group.  
     
     
         32 . The composition of  claim 1 , wherein said antigen is a protein or a fragment thereof, being selected from the group consisting of: 
 (a) proteins suitable to induce an immune response against allergens,    (b) proteins suitable to induce an immune response against cancer cells,    (c) proteins suitable to induce an immune response against infectious diseases, and    (d) proteins suitable to induce an immune response in farm animals or pets.    
     
     
         33 . The composition of  claim 1 , wherein said antigen is: 
 a recombinant protein of bee sting allergy,    a recombinant protein of nut allergy,    a recombinant protein of food allergies,    a recombinant protein of asthma,    a recombinant protein of breast cancer cells,    a recombinant protein of kidney cancer cells,    a recombinant protein of prostate cancer cells,    a recombinant protein of skin cancer cells,    a recombinant protein of brain cancer cells,    a recombinant protein of leukemia cells,    a recombinant protein of Influenza virus,    a recombinant protein of HIV,    a recombinant protein of Hepatitis C virus,    a recombinant protein of  Toxoplasma,      a recombinant protein of  Plasmodium falciparum,      a recombinant protein of  Plasmodium vivax,      a recombinant protein of  Plasmodium ovale,      a recombinant protein of  Plasmodium malariae,      a recombinant protein of  Chlamydia.      
     
     
         34 . The composition of  claim 1 , wherein said antigen is suitable to prevent or treat an infectious disease of viral etiology.  
     
     
         35 . The composition of  claim 34 , wherein said infectious disease of viral etiology is selected from the group consisting of: 
 HIV,    influenza,    herpes,    viral hepatitis,    Epstein Bar, or    viral encephalitis.    
     
     
         36 . The composition of  claim 1 , wherein said antigen is suitable to prevent or treat an infectious disease of bacterial etiology.  
     
     
         37 . The composition of  claim 36 , wherein said infectious disease of bacterial etiology is a pneumonia, syphillis or tuberculosis.  
     
     
         38 . The composition of  claim 1 , wherein said antigen is suitable to prevent or treat an infectious disease of parasitic etiology.  
     
     
         39 . The composition of  claim 38 , wherein said infectious disease of parasitic etiology is malaria, trypanosomiasis, leishmaniasis, trichomoniasis or amoebiasis.  
     
     
         40 . The composition of  claim 1 , wherein said antigen or antigenic determinant is a peptide, a protein, or a fragment of a protein, selected from the group consisting of: 
 (a) an Influenza M2 protein;    (b) a phospholipase A2 protein; and    (c) a Der p I peptide.    
     
     
         41 . The composition of  claim 1 , wherein said antigen or said antigenic determinant is an influenza M2 protein, or a fragment thereof.  
     
     
         42 . The composition of  claim 1 , wherein said antigen or antigenic determinant is the M2 protein of SEQ ID NO: 213 or a fragment thereof being at least 6 amino acids in length.  
     
     
         43 . The composition of  claim 1 , wherein said antigen or antigenic determinant is a peptide comprising the amino acids 2 to 24 of SEQ ID NO: 213 or a fragment thereof being at least 6 amino acids in length.  
     
     
         44 . The composition of  claim 1 , wherein said antigen or antigenic determinant is a peptide consisting of the amino acids 2 to 24 of SEQ ID NO: 171 or a fragment thereof being at least 6 amino acids in length.  
     
     
         45 . The composition of  claim 1 , wherein said antigen or antigenic determinant with said second attachment site is the M2 peptide of SEQ ID NO: 170.  
     
     
         46 . The composition of  claim 1 , wherein said antigen or antigenic determinant is Der p I peptide, or a fragment thereof comprising an amino acid sequence selected from the group consisting of:  
       
         
           
                 
                 
                 
                 
               
                     
                     
                 
                     
                   (a) 
                   CGNQSLDLAEQELVDCASQHGCH; 
                     
                 
                     
                   and 
                 
                     
                     
                 
                     
                   (b) 
                   CQIYPPNANKIREALAQTHSA. 
                 
                     
                     
                 
             
                
                
                
                
                
                
               
            
           
         
       
     
     
         47 . The composition of  claim 1 , wherein said antigen or said antigenic determinant is a phospholipase A2 protein, or a fragment thereof.  
     
     
         48 . The composition of  claim 1 , wherein said antigen or antigenic determinant is a phospholipase A2 protein or fragment thereof comprising an amino acid sequence selected from the group consisting of: 
 (a) the amino acid sequence of SEQ ID NO:168;    (b) the amino acid sequence of SEQ ID NO:169;    (c) the amino acid sequence of SEQ ID NO:170;    (d) the amino acid sequence of SEQ ID NO:171;    (e) the amino acid sequence of SEQ ID NO:172;    (f) the amino acid sequence of SEQ ID NO:173;    (g) the amino acid sequence of SEQ ID NO:174; and    (h) the amino acid sequence of SEQ ID NO:175.    
     
     
         49 . A pharmaceutical composition comprising: 
 (a) the composition of  claim 1;  and    (b) an acceptable pharmaceutical carrier.    
     
     
         50 . A method of immunization of an animal comprising administering to said animal the composition of  claim 1 , wherein an immune response against said antigen or antigenic determinant is produced in said animal.  
     
     
         51 . An immunogenic composition comprising the composition of  claim 1  and an adjuvant.  
     
     
         52 . A process for producing a non-naturally occurring, ordered and repetitive antigen array comprising: 
 (a) providing a non-natural molecular scaffold comprising: 
 (i) a core particle comprising a virus-like particle of an RNA bacteriophage; and  
 (ii) an organizer comprising at least one first attachment site,  
 wherein said organizer is connected to said core particle by at least one covalent bond; and wherein said first attachment site is not a sulfhydryl group; and  
   (b) providing an antigen or antigenic determinant with at least one second attachment site, wherein said second attachment site is capable of association through at least one non-peptide bond to said first attachment site; and    (c) combining said non-natural molecular scaffold and said antigen to form an ordered and repetitive antigen array.