Allergoids Derived from Allergens

Abstract

Modified allergens having reduced allergenicity compared to corresponding native allergenic material, wherein all or a part of the primary amine groups of the lysine and arginine residues of the allergenic molecules are functionalized and the modified allergens have the structure wherein R and R 2 are independently selected from H, C1-C5 alkyl, phenyl, phenyl substituted in ortho, meta, or para with a hydroxy, C1-C4 alkoxy, halogen, amino, alkylamino, dialkylamino, mercapto, C1-C4 alkylmercapto group; X represents O, S, or NR3, where R3 is H, alkyl with 1-6 carbon atoms, phenyl, or CN; R1 represents H, alkyl with 1-8 carbon atoms, phenyl or arylalkyl with up to 8 carbon atoms, or alkyl containing a heterocyclic ring; prot represents the protein residue of the allergen; n is the number of functionalized arginine groups and ranges between 1 and the number of arginine groups present in the allergen; m is the number of functionalized lysine groups and ranges between 1 and the number of lysine groups present in the allergen.

1 - 12 . (canceled) 
     
     
         13 . Modified allergens having reduced allergenicity compared to corresponding native allergenic material, wherein all or a part of primary amine groups of lysine and arginine residues of allergenic molecules are functionalized as shown in the structure (I), and said modified allergens have the structure: 
       
         
           
           
               
               
           
         
       
       in which
 R and R 2  are independently selected from H, C1-C5 alkyl, phenyl, phenyl substituted in ortho, meta, or para with a hydroxy, C1-C4 alkoxy, halogen, amino, alkylamino, dialkylamino, mercapto, C1-C4 alkylmercapto group, 
 X represents O, S, or NR3, where R3 is H, alkyl with 1-6 carbon atoms, phenyl, or CN, 
 R1 represents H, alkyl with 1-8 carbon atoms, phenyl or arylalkyl with up to 8 carbon atoms, or alkyl containing a heterocyclic ring, 
 prot represents the protein residue of the allergen; 
 n is the number of functionalized arginine groups, and ranges between 1 and the number of arginine groups present in the allergen, and 
 m is the number of functionalized lysine groups and ranges between 1 and the number of lysine groups present in the allergen. 
 
     
     
         14 . The modified allergens according to  claim 13 , wherein said native allergenic material subjected to modification is obtained from mites, pollens, epitelia of animals, mycophyta, proteins of food origin, selected from proteins of milk, egg, cereals, peach, apple, by extraction of the allergenic proteins with suitable solvent. 
     
     
         15 . The modified allergens according to  claim 13 , wherein said native allergenic material is composed of proteins purified from the above-listed raw materials. 
     
     
         16 . The modified allergens according to  claim 13 , wherein said native allergenic material is obtained in a recombinant form. 
     
     
         17 . The modified allergens according to  claim 13 , wherein the average percentage of modified primary amine groups of the lysine ranges between 75% and 100%, preferably about 90%; and the average percentage of the substituted arginine residues ranges between 25 and 100%, preferably about 40%. 
     
     
         18 . The modified allergens according to  claim 13 , wherein R is phenyl, phenyl substituted in ortho, meta, or para with a hydroxy, C1-C4 alkoxy, halogen, amino, alkylamino, dialkylamino, mercapto, C1-C4 alkylmercapto group, and R2 is hydrogen. 
     
     
         19 . The modified allergens according to  claim 13 , wherein X is O or S, and R1 is hydrogen. 
     
     
         20 . The modified allergens according to  claim 13 , wherein R is phenyl, R1 and R2 are hydrogen, and X is O or S. 
     
     
         21 . A method for obtaining modified allergens according to  claim 13 , comprising the following steps:
 a) carbamylation or thiocarbamylation reaction of all or a part of the lysine residues of a raw native allergenic material;   b) subsequently, reaction of all or a part of the arginine residues of said allergenic material of the step a) with a dialdehyde or diketal.   
     
     
         22 . The method according to  claim 21 , wherein said carbamylation reaction of step a) is carried out by reacting said native allergenic material with potassium cyanate to a final concentration of the salt ranging between 0.1 M and 1.5 M, and by keeping the pH between 7 and 11, at a temperature ranging between the room temperature and 50° C., for a total reaction time ranging between 12 and 36 hours. 
     
     
         23 . The method according to  claim 21 , wherein said carbamylation reaction of step a) is carried out by reacting said native allergenic material with potassium cyanate to a final concentration of the salt ranging between 0.4 M and 0.8 M, and by keeping the pH between 9 and 9.6; at a temperature ranging between 35 and 40° C., for a total reaction time ranging between 16 and 24 hours. 
     
     
         24 . The method according to  claim 21 , wherein, between said step a) and said step b), the thus-modified native allergenic material is subjected to gel-filtration to remove the excess reagent and is equilibrated with a suitable saline solution. 
     
     
         25 . The method according to  claim 21 , wherein said step b) is carried out by reaction with phenylglyoxal, in a molar excess ranging between 100 and 1600, for a period of time ranging between 30 minutes and 8 hours, at temperatures ranging between 20 and 37° C. 
     
     
         26 . The method according to  claim 21 , wherein said step b) is carried out by reaction with phenylglyoxal, in a molar excess ranging between 400 and 800, for a period of 4 hours, at a temperature of 25° C. 
     
     
         27 . A pharmaceutical composition comprising an efficient dose for immunotherapy of one or more modified allergens according to  claim 13 , in combination with pharmaceutically acceptable excipients. 
     
     
         28 . The modified allergens according to  claim 13 , for the use in specific immunotherapy, wherein said modified allergens have the ability to induce specific IgG antibodies capable of recognizing also the corresponding native allergen, but reduced capability of linkage to the specific IgE antibodies, as compared to unmodified allergen.