Modified amino acids comprising tetrazine functional groups, methods of preparation, and methods of their use

Abstract

Provided herein are modified amino acids comprising a tetrazine groups according to Formula I: polypeptides, antibodies, payloads and conjugates comprising these modified amino acid residues derived from the modified amino acids, and methods of producing the polypeptides, antibodies, payloads and conjugates comprising the modified amino acid residues. The polypeptides, antibodies, payloads and conjugates are useful in methods of treatment and prevention, methods of detection and methods of diagnosis.

What is claimed is: 
     
         1 . A compound according to formula I: 
       
         
           
           
               
               
           
         
         or a salt thereof, wherein:
 Ar is 
 
       
       
         
           
           
               
               
           
         
         
           V is a single bond, lower alkylene, or —W 1 -W 2 —; 
           one of W, and W 2  is absent or lower alkylene, and the other is —NH—, —O—, or —S—; 
           each X 1  is independently —NH—, —O—, or —S—; 
           one of Z 1 , Z 2 , and Z 3  is —CH— or —N— and the others of Z 1 , Z 2 , and Z 3  are each independently —CH—; and 
           R is lower alkyl; 
           wherein when Ar is 
         
       
       
         
           
           
               
               
           
         
       
       and V is —NH—, then one of Z 1 , Z 2 , and Z 3  is —N—. 
     
     
         2 . The compound of  claim 1 , wherein V is a single bond, —NH—, or —CH 2 NH—. 
     
     
         3 . The compound of  claim 1 , wherein:
 V is a single bond, —NH—, or —CH 2 NH—; and   Ar is   
       
         
           
           
               
               
           
         
       
     
     
         4 . The compound of  claim 1 , wherein:
 V is a single bond, —NH—, or —CH 2 NH—; and   Ar is   
       
         
           
           
               
               
           
         
       
     
     
         5 . The compound of  claim 1 , wherein:
 V is a single bond, —NH—, or —CH 2 NH—; and   Ar is   
       
         
           
           
               
               
           
         
       
     
     
         6 . The compound of  claim 1  according to formula Ia: 
       
         
           
           
               
               
           
         
       
     
     
         7 . The compound of  claim 1  according to formula II: 
       
         
           
           
               
               
           
         
         or a salt thereof, wherein V is a single bond or —CH 2 NH—. 
       
     
     
         8 . The compound of  claim 1  according to formula III: 
       
         
           
           
               
               
           
         
         or a salt thereof, wherein V is a single bond or —CH 2 NH—. 
       
     
     
         9 . The compound of  claim 1  according to formula IV, V, VI or VII: 
       
         
           
           
               
               
           
         
         or a salt thereof, wherein V is a single bond, —NH—, or —CH 2 NH—. 
       
     
     
         10 . The compound of  claim 1  according to formula VIII or IX: 
       
         
           
           
               
               
           
         
         or a salt thereof, wherein V is a single bond, —NH—, or —CH 2 NH—. 
       
     
     
         11 . The compound of  claim 1  according to any of formulas 1-10: 
       
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         or a salt thereof. 
       
     
     
         12 . A polypeptide comprising an amino acid residue corresponding to the compound of any of  claims 1 - 11 . 
     
     
         13 . A polypeptide comprising at least two amino acid residues corresponding to the compound of any of  claims 1 - 11 . 
     
     
         14 . The polypeptide of any of  claims 12 - 13 , further comprising at least one amino acid residue comprising an azide functional group. 
     
     
         15 . The polypeptide of  claim 14 , wherein the amino acid residue comprising an azide functional group is selected from the group consisting of a compound according to any of formulas A1-A30, A40, and combinations thereof: 
       
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
       
     
     
         16 . A conjugate comprising the polypeptide of any of  claims 12 - 15  linked to a payload and optionally comprising a linking moiety between the polypeptide and the payload. 
     
     
         17 . An antibody comprising an amino acid residue corresponding to the compound of any of  claims 1 - 11 . 
     
     
         18 . An antibody comprising at least two amino acid residues corresponding to the compound of any of  claims 1 - 11 . 
     
     
         19 . The antibody of any of  claims 17 - 18 , further comprising at least one amino acid residue comprising an azide functional group. 
     
     
         20 . The antibody of  claim 19 , wherein the amino acid residue comprising an azide functional group is selected from the group consisting of a compound according to any of formulas A1-A30, A40, and combinations thereof. 
     
     
         21 . A conjugate comprising the antibody of  claim 17 - 20  linked to a payload and optionally comprising a linking moiety between the antibody and the payload. 
     
     
         22 . An orthogonal tRNA aminoacylated with an amino acid residue corresponding to the compound of any of  claims 1 - 11 . 
     
     
         23 . A method of producing a polypeptide, comprising contacting a polypeptide with the orthogonal tRNA of  claim 22  under conditions suitable for incorporating the amino acid residue into the polypeptide. 
     
     
         24 . The method of  claim 23 , wherein the orthogonal tRNA base pairs with a codon that is not normally associated with an amino acid. 
     
     
         25 . The method of  claim 23 , wherein the contacting occurs in a reaction mixture which comprises a tRNA synthetase capable of aminoacylating the orthogonal tRNA with the compound of any of  claims 1 - 11 . 
     
     
         26 . The method of  claim 25 , wherein the tRNA synthetase is selected from the group consisting of SEQ ID NOs: 4-5.