US2019367554A1PendingUtilityA1

Preparation of oligo conjugates

Assignee: EMERALD THERAPEUTICS INCPriority: Mar 8, 2013Filed: Apr 11, 2019Published: Dec 5, 2019
Est. expiryMar 8, 2033(~6.6 yrs left)· nominal 20-yr term from priority
A61K 47/549C07H 21/00C07H 21/04Y10S977/932C07K 14/003
71
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Claims

Abstract

Conjugated molecules are prepared that comprise a predetermined number of oligo conjugation components. The conjugated molecules also may comprise one or more detectable labels. Preparation of these molecules can be implemented according to an asymmetric or a symmetric conjugation strategy.

Claims

exact text as granted — not AI-modified
1 . A method of preparing a compound of the formula 
       
         
           
           
               
               
           
         
       
       comprising 
       (i) attaching a conjugation component of the formula 
       
         
           
           
               
               
           
         
       
       wherein R 2  is R 2a  or R 2ap , to a solid support 
       
         
           
           
               
               
           
         
       
       to form a compound of the formula 
       
         
           
           
               
               
           
         
       
       (ii) when R 2  is R 2ap , converting 
       
         
           
           
               
               
           
         
       
       to 
       
         
           
           
               
               
           
         
       
       and 
       (iii) contacting 
       
         
           
           
               
               
           
         
       
       with a conjugation component of the formula 
       
         
           
           
               
               
           
         
       
       wherein R 3  is R 3a , R 3a p, hydrogen, a detectable label or an oligo; 
       to form 
       
         
           
           
               
               
           
         
       
       wherein: 
       
         
           
           
               
               
           
         
       
       is a solid support material; 
       
         
           
           
               
               
           
         
       
       and are independently an oligo;
 R 1a  and R 1b  are complementary conjugation functionalities and L 1  is conjugate linker, and R 1a , R 1b , and L 1  are 
 (a) R 1a  is azido, R 1b  is —C≡C—CR 23 , and L 1  is 
 
       
         
           
           
               
               
           
         
       
       or
 (b) R 1a  is —NHR 23 , R 1b  is carboxy, and L 1  is —NR 23 C(═O)—, or 
 (c) R 1a  is carboxy, R 1b  is —NHR 23 , and L 1  is —C(═O)NR 23 —, or 
 (d) R 1a  is —NHR 23 , R 1b  is halo, and L 1  is —NR 23 —, or 
 (e) R 1a  is —O—P(═O)(OH)(X), R 1b  is hydroxy, and L 1  is —O—P(═O)(OH)—O—, or 
 (f) R 1a  is —O—P(═O)(OH)(X), R 1b  is —NHR 23 , and L 1  is —O—P(═O)(OH)— NR 23 —, or 
 (g) R 1a  is —O—P(═O)(OH)(X), R 1b  is thio, and L 1  is —O—P(═O)(OH)—S—, or 
 (h) R 1a  is halo, R 1b  is thio, and L 1  is —S—, or 
 (i) R 1a  is 
 
       
         
           
           
               
               
           
         
       
       R 1b  is thio, and L 1  is 
       
         
           
           
               
               
           
         
       
       or
 (j) R 1a  is —C≡C—R 23 , R 1b  is azido, and L 1  is 
 
       
         
           
           
               
               
           
         
       
       or
 (k) R 1a  is halo, R 1b  is —NHR 23 , and L 1  is —NR 23 —, or 
 (l) R 1a  is hydroxy, R 1b  is —O—P(═O)(OH)(X), and L 1  is —O—P(═O)(OH)—O—, or 
 (m) R 1a  is —NHR 23 , R 1b  is —O—P(═O)(OH)(X), and L 1  is —NR 23 —P(═O)(OH)—O—, or 
 (n) R 1a  is thio, R 1b  is —O—P(═O)(OH)(X), and L 1  is —S—P(═O)(OH)—O—, or 
 (o) R 1a  is —O—P(═O)(OH)SH, R 1b  is —X, and L 1  is —O—P(═O)(OH)—S—, or 
 (p) R 1a  is —X, R 1b  is —O—P(═O)(OH)SH, and L 1  is —S—P(═O)(OH)—O—, or 
 (q) R 1a  is —(CR 25 R 25 ) s C(═O)OR 23 , R 1b  is —NHR 23 , L 1  is —(CR 25 R 25 ) s C(═O)NR 23 —, or 
 (r) R 1a  is —NHR 23 , R 1b  is —(CR 25 R 25 ) s C(═O)OR 23 , L 1  is —NR 23 C(═O)—(CR 25 R 25 ) s —, or 
 (s) R 1a  is thio, R 1b  is halo, and L 1  is —S—, or 
 (t) R 1a  is thio, R 1b  is 
 
       
         
           
           
               
               
           
         
       
       and L 1  is 
       
         
           
           
               
               
           
         
         (u) R 1a  is —SH, R 1b  is —O—P(═O)(OH)(O—(CH 2 ) n —SH) and L 1  is —S—S—(CH 2 ) n —O—P(OH)(═O)—O—; 
         R 2a  and R 2b  are complementary conjugation functionalities and L 2  is conjugate linker, and R 2ap , R 2a , R 2b , and L 2  are 
         (a′) R 2ap  is halo, R 2a  is azido, R 2b  is —C≡C—R 23 , and L 2  is 
       
       
         
           
           
               
               
           
         
       
       or
 (b′) R 2ap  is —NR 23 Pr, R 2a  is —NHR 23 , R 2b  is carboxy, and L 2  is —NR 23 C(═O)—, or 
 (c′) R 2ap  is carboxy ester, R 2a  is carboxy, R 2b  is —NHR 23 , and L 2  is —C(═O)NR 23 —, or 
 (d′) R 2ap  is —NR 23 Pr, R 2a  is —NHR 23 , R 2b  is halo, and L 2  is —NR 23 —, or 
 (e′) R 2ap  is —OH, phosphate or phosphate ester, R 2a  is —O—P(═O)(OH)(X), R 2b  is hydroxy, and L 2  is —O—P(═O)(OH)—O—, or 
 (f′) R 2ap  is —OH, phosphate or phosphate ester, R 2a  is —O—P(═O)(OH)(X), R 2b  is —NHR 23 , and L 2  is —O—P(═O)(OH)—NR 23 —, or 
 (g′) R 2ap  is —OH, phosphate or phosphate ester, R 2a  is —O—P(═O)(OH)(X), R 2b  is thio, and L 2  is —O—P(═O)(OH)—S—, or 
 (h′) R 2a  is —X, R 2b  is thio, and L 2  is —S—, or 
 (i′) R 2a  is 
 
       
         
           
           
               
               
           
         
       
       R 2b  is thio, and L 2  is 
       
         
           
           
               
               
           
         
       
       or
 (j′) R 2a  is —C≡C—R 23 , R 2b  is azido, and L 2  is 
 
       
         
           
           
               
               
           
         
       
       or
 (k′) R 2a  is halo, R 2b  is —NHR 23 , and L 2  is —NR 23 —, or 
 (l′) R 2a  is hydroxy, R 2b  is —O—P(═O)(OH)(X), and L 2  is —O—P(═O)(OH)—O—, or 
 (m′) R 2a  is —NHR 23 , R 2b  is —O—P(═O)(OH)(X), and L 2  is —NR 23 —P(═O)(OH)—O—, or 
 (n′) R 2a  is thio, R 2b  is —O—P(═O)(OH)(X), and L 2  is —S—P(═O)(OH)—O—, or 
 (o′) R 2ap  is —O—P(═O)(OH)SR 24 , R 2a  is —O—P(═O)(OH)SH, R 2b  is —X, and L 2  is —O—P(═O)(OH)—S—, or 
 (p′) R 2a  is —X, R 2b  is —O—P(═O)(OH)SH, and L 2  is —S—P(═O)(OH)—O—, or 
 (q′) R 2a  is —(CR 25 R 25 ) s C(═O)OR 23 , R 2b  is —NHR 23 , L 2  is —(CR 25 R 25 ) s C(═O)NR 23 —, or 
 (r′) R 2a  is —NHR 23 , R 2b  is —(CR 25 R 25 ) s C(═O)OR 23 , L 2  is —NR 23 C(═O)—(CR 25 R 25 ) s —, or 
 (s′) R 2a  is thio, R 2b  is halo, and L 2  is —S—, or 
 (t′) R 2a  is thio, R 2b  is 
 
       
         
           
           
               
               
           
         
       
       and L 2  is 
       
         
           
           
               
               
           
         
       
       or
 (u′) R 2a  is —SH, R 2b  is —O—P(═O)(OH)(O—(CH 2 ) n —SH) and L 2  is —S—S—(CH 2 ) n —O—P(OH)(═O)—O—; 
 X is selected from chlorine, bromine, fluorine, tosylate, mesylate, triflate, or dimethoxy triflate, 
 n is 1, 2, 3, 4, 5, or 6; 
 R 23  is selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, cycloalkynyl, substituted cycloalkynyl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic; 
 Pr is an amino protecting group; 
 R 24  is trityl or benzyl; 
 R 25  is hydrogen or C 1-6  alkyl; 
 n is 1, 2, 3, 4, 5, or 6; 
 s is an integer of greater than 1; 
 R 3ap  is selected from the group consisting of halo, —OH, —NR 23 Pr, —O—P(═O)(OH)SR 24 , carboxy ester, phosphate and phosphate ester; and 
 R 3a  is selected from the group consisting of azido, —C≡C—R 23 , —NHR 23 , carboxy, halo, hydroxy, —C(═O)OR 23 , —O—P(═O)(OH)SH and —O—P(═O)(OH)Br; 
 provided that R 2  does not react with R 1a  or R 1b , and R 3  does not react with R 2a  or R 2b ; 
 --- represents the point of connection to the part of the solid support-bound conjugated molecule that is closer to 
 
       
         
           
           
               
               
           
         
       
       and   represents the point of connection to the part of the solid support-bound conjugated molecule that is further away from 
       
         
           
           
               
               
           
         
       
     
     
         2 . The method of  claim 1  for preparing a compound of the formula 
       
         
           
           
               
               
           
         
       
       comprising 
       (i) attaching a compound of the formula 
       
         
           
           
               
               
           
         
       
       to a solid support 
       
         
           
           
               
               
           
         
       
       to form a compound of the formula 
       
         
           
           
               
               
           
         
       
       and 
       (iii) reacting 
       
         
           
           
               
               
           
         
       
       with a compound of the formula 
       
         
           
           
               
               
           
         
       
       to form 
       
         
           
           
               
               
           
         
       
       wherein: 
       
         
           
           
               
               
           
         
       
       is a solid support material; 
       
         
           
           
               
               
           
         
       
       and are independently an oligo;
 R 1a  and R 1b  are complementary conjugation functionalities and L 1  is conjugate linker, R 2a  and R 2b  are complementary conjugation functionalities and L 2  is conjugate linker, and R 1a , R 1b L 1 , R 2a , R 2b , L 2 , and R 3a  are selected from 
 
       
         
           
                 
                 
                 
               
                     
                 
                   R 1a , R 2a , or R 3a   
                   R 1b  or R 2b   
                   L 1  or L 2   
                 
                     
                 
                   —C≡C—R 23   
                   azido 
                   
                     
                       
                       
                           
                           
                       
                     
                   
                 
                     
                 
                   azido 
                   —C≡C—R 23   
                   
                     
                       
                       
                           
                           
                       
                     
                   
                 
                     
                 
                   carboxy 
                   —NHR 23   
                   —C(═O)NR 23 — 
                 
                   —NHR 23   
                   carboxy 
                   —NR 23 C(═O)— 
                 
                   halo 
                   —NHR 23   
                   —NR 23 — 
                 
                   —NR 23   
                   halo 
                   —NR 23 — 
                 
                   hydroxy 
                   —O—P(═O)(OH)(X) 
                   —O—P(═O)(OH)—O— 
                 
                   —O—P(═O)(OH)(X) 
                   hydroxy 
                   —O—P(═O)(OH)—O— 
                 
                   —NHR 23   
                   —O—P(═O(OH)(X) 
                   —NR 23 —P(═O)(OH)—O— 
                 
                   —O—P(═O)(OH)(X) 
                   —NHR 23   
                   —O—P(═O)(OH)—NR 23 — 
                 
                   thio 
                   —O—P(═O)(OH)(X) 
                   —S—P(═O)(OH)—O— 
                 
                   —O—P(═O)(OH)(X) 
                   thio 
                   —O—P(═O)(OH)—S— 
                 
                   thio 
                   —X 
                   —S— 
                 
                   —X 
                   thio 
                   —S— 
                 
                   —O—P(═O)(OH)SH 
                   —X 
                   —O—P(═O)(OH)—S— 
                 
                   —X 
                   —O—P(═O)(OH)SH 
                   —S—P(═O)(OH)—O— 
                 
                   —(CR 25 R 25 ) s C(═O)OR 23   
                   —NHR 23   
                   —(CR 25 R 25 ) s C(═O)NR 23 — 
                 
                   —NHR 23   
                   —(CR 25 R 25 ) s C(═O)OR 23   
                   —NR 23 C(═O)—(CR 25 R 25 ) s — 
                 
                     
                 
                   thio 
                   
                     
                       
                       
                           
                           
                       
                     
                   
                   
                     
                       
                       
                           
                           
                       
                     
                   
                 
                     
                 
                   
                     
                       
                       
                           
                           
                       
                     
                   
                   thio 
                   
                     
                       
                       
                           
                           
                       
                     
                   
                 
                     
                 
                   —SH 
                   —O—P(═O)(OH)(O—(CH 2 ) n —SH) 
                   —S—S—(CH 2 ) n —O—P(OH)(═O)—O— 
                 
                     
                 
             
                
                
                
               
               
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
               
            
           
         
         wherein the selection of R 1a , R 2a , or R 3a  is independent of one another provided that L 1  and L 2  are different, R 2a  does not react with R 1a  or R 1b , and R 3a  does not react with R 2a  or R 2b ; 
         X is selected from chlorine, bromine, fluorine, tosylate, mesylate, triflate, or dimethoxy triflate; 
         R 23  is selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, cycloalkynyl, substituted cycloalkynyl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic; and 
         R 25  is hydrogen or C 1-6  alkyl; 
         s is an integer of greater than 1; 
         --- represents the point of connection to the part of the solid support-bound conjugated molecule that is closer to 
       
       
         
           
           
               
               
           
         
       
       and   represents the point of connection to the part of the solid support-bound conjugated molecule that is further away from 
       
         
           
           
               
               
           
         
       
     
     
         3 . A method of preparing a compound of the formula 
       
         
           
           
               
               
           
         
       
       comprising 
       (i) attaching a compound of the formula 
       
         
           
           
               
               
           
         
       
       to a solid support 
       
         
           
           
               
               
           
         
       
       to form a compound of the formula 
       
         
           
           
               
               
           
         
       
       (ii) converting 
       
         
           
           
               
               
           
         
       
       to 
       
         
           
           
               
               
           
         
       
       and 
       (iii) contacting 
       
         
           
           
               
               
           
         
       
       with a compound of the formula 
       
         
           
           
               
               
           
         
       
       to form 
       
         
           
           
               
               
           
         
       
       wherein: 
       
         
           
           
               
               
           
         
       
       is a solid support material; 
       
         
           
           
               
               
           
         
       
       and are independently an oligo;
 R ap , R a , R b , and L are selected from 
 (a) R ap  is halo, R a  is azido, R b  is —C≡C—R 23 , and L is 
 
       
         
           
           
               
               
           
         
       
       or
 (b) R ap  is —NR 23 Pr, R a  is —NHR 23 , R b  is carboxy, and L is —NR 23 C(═O)—, or 
 (c) R ap  is carboxy ester, R a  is carboxy, R b  is —NHR 23 , and L is —C(═O)NR 23 —, or 
 (d) R ap  is —NR 23 Pr, R a  is —NHR 23 , R b  is halo, and L is —NR 23 —, or 
 (e) R ap  is —OH, phosphate or phosphate ester; R a  is —O—P(═O)(OH)(X), R b  is hydroxy, and L is —O—P(═O)(OH)—O—, or 
 (f) R ap  is —OH, phosphate or phosphate ester; R a  is —O—P(═O)(OH)(X), R b  is —NHR 23 , and L is —O—P(═O)(OH)—NR 23 —, or 
 (g) R ap  is —O—P(═O)(OH)SR 24 , R a  is —O—P(═O)(OH)SH, R b  is —X, and L is —O—P(═O)(OH)—S—, or 
 (h) R ap  is —OH, phosphate or phosphate ester; R a  is —O—P(═O)(OH)Br, R b  is thio, and L is —O—P(═O)(OH)—S—, or 
 (i) (i) R ap  is —SR 24 , R a  is —SH, R b  is —O—P(═O)(OH)SH and L is —O—P(═O)(OH)S—S—; 
 wherein R 23  is selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, cycloalkynyl, substituted cycloalkynyl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic; 
 X is selected from chlorine, bromine, fluorine, tosylate, mesylate, triflate, or dimethoxy triflate; 
 R 24  is trityl or benzyl; 
 --- represents the point of connection to the part of the solid support-bound conjugated molecule that is closer to 
 
       
         
           
           
               
               
           
         
       
       and   represents the point of connection to the part of the solid support-bound conjugated molecule that is further away from 
       
         
           
           
               
               
           
         
       
     
     
         4 . A method of preparing a compound of the formula 
       
         
           
           
               
               
           
         
       
       comprising
 cleaving the bond between 
 
       
         
           
           
               
               
           
         
       
       and L 1  of the compound of the formula 
       
         
           
           
               
               
           
         
       
       thereby obtaining 
       
         
           
           
               
               
           
         
       
       wherein:
 the compound of 
 
       
         
           
           
               
               
           
         
       
       is prepared according to the method of  claim 1 ; and 
       
         
           
           
               
               
           
         
       
       L 1 , L 2 , R 3 , 
       
         
           
           
               
               
           
         
       
       are as defined in  claim 1 ; and
 Z is selected from —OH, OH—(C 1 -C 10 )alkylene-, —COOH, NH 2 C(O)—, NH 2 NH—C(O)—, COOH—(C 1 -C 10 )alkylene-, NH 2 C(O)—(C 1 -C 10 )alkylene-, NH 2 NH—C(O)—(C 1 -C 10 )alkylene-, CH 2 ═CH—(C 1 -C 10 )alkylene-, C≡C—(C 1 -C 10 )alkylene- or HS—(C 1 -C 10 )alkylene- and any alkylene is optionally substituted by one or more groups selected from —OH, halogen, —NHR″, —NHC(O)—(C 1 -C 10 )alkylene-C≡CH, or —NHC(O)—(C 1 -C 10 )alkylene-CH═CH 2 ; 
 R″ is selected from (C 1 -C 10 )alkyl, (C 3 -C 10 )cycloalkyl, or (C 3 -C 10 )aryl. 
 
     
     
         5 .- 8 . (canceled)

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