Modulators of coagulation factors with enhanced stability

Abstract

The invention provides improved nucleic acid ligands with enhanced stability that inhibit coagulation and improved modulators of the nucleic acids to provide ideal modulators of coagulation. These improved nucleic acids and modulators are particularly useful for inhibiting coagulation in a host undergoing a therapeutic regime such as surgery or coronary artery bypass.

1 . A composition comprising a nucleic acid sequence of one of SEQ ID NOs:9-59 linked to a polyethylene glycol molecule at a 5′ terminus via a linker moiety, or a pharmaceutically acceptable salt thereof.  
     
     
         2 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
       
       wherein ligand is selected from one of SEQ ID NO:9-59 or a pharmaceutically acceptable salt thereof.  
     
     
         3 . The composition of  claim 1  wherein the composition is produced by conjugating a linker precursor to a nucleic acid molecule to form a nucleic acid linker-composition and further, conjugating a polyethylene glycol precursor moiety to the linker of the nucleic acid-linker composition.  
     
     
         4 . The composition of  claim 3  in which the linker precursor is selected from the group consisting of: 
 6-(trifluoroacetamido)hexanol (2-cyanoethyl-N,N-diisopropyl)phosphoramidite of the structure:                          TFA-amino C4 CED phosphoramidite of the structure:                          5′-amino modifier C3 TFA of the structure:                          5′-Amino-Modifier C3-TFA    5′-amino modifier 5 of the structure:                          5′-Amino Modifier 5,    5′-amino modifier C12 of the structure:                          , and 5′thiol-modifier C6 of the structure:                          
     
     
         5 . The composition of  claim 3  wherein the polyethylene glycol precursor is selected from:  
       
         
           
           
               
               
           
         
         
           
           
               
               
           
         
       
     
     
         6 . The composition of  claim 1  wherein the polyethylene glycol has a total mass of about 40 kD.  
     
     
         7 . The composition of  claim 1  wherein the composition comprises two polyethylene glycol moieties.  
     
     
         8 . The composition of  claim 7  wherein each moiety has a total mass of about 20 kD.  
     
     
         9 . The composition of  claim 1  wherein the nucleic acid comprises SEQ ID NO: 19.  
     
     
         10 . The composition of  claim 1  wherein the nucleic acid comprises SEQ ID NO: 39.  
     
     
         11 . The composition of  claim 1  comprising the structure: 
 5′-O-[6-[N 2 -(monomethoxy 20K polyethylene glycol carbamoyl)-N 6 -(monomethoxy 20K polyethylene glycol carbamoyl)]-lysylamido]hexyl]-2′-methoxy-2′-deoxguanylyl-(3′-5′)-2′-methoxy-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyguanyiyl-(3-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-methoxy-2′-deoxyuridylyl-(3′-5′)-riboguanylyl-(3′-5′)-2′-methoxy-2′-deoxycytidylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxcytidylyl-(3′-5′)-2′-methoxy-2′-deoxytidylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-methoxy-2′-deoxcytidylyl-(3′-3)-thymidine.    
     
     
         12 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         13 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         14 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         15 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         16 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         17 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         18 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         19 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         20 . The composition of claim I of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         21 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         22 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         23 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         24 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         25 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         26 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         27 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         28 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         29 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         30 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         31 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         32 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         33 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         34 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         35 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         36 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt thereof.  
       
     
     
         37 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt therof.  
       
     
     
         38 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt therof.  
       
     
     
         39 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt therof.  
       
     
     
         40 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt therof.  
       
     
     
         41 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt therof  
       
     
     
         42 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt therof.  
       
     
     
         43 . The composition of  claim 1  of the formula:  
       
         
           
           
               
               
           
         
         n=approximately 450  
         , or a pharmaceutically acceptable salt therof.  
       
     
     
         44 . A method of regulating blood coagulation comprising contacting a FIXa molecule with a composition comprising a nucleic acid sequence of one of SEQ ID NOs:9-59 linked to a polyethylene glycol molecule at a 5′ terminus via a linker moiety, or a pharmaceutically acceptable salt thereof.  
     
     
         45 . The method of  claim 44  wherein the aptamer comprises structure: 
 5′-O-[6-[N 2 -(monomethoxy 20K polyethylene glycol carbamoyl)-N 6 -(monomethoxy 20K polyethylene glycol carbamoyl)]-lysylamido]hexyl]-2′-methoxy-2′-deoxguanylyl-(3′-5′)-2′-methoxy-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxyguanylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-methoxy-2′-deoxyuridylyl-(3′-5′)-riboguanylyl-(3′-5′)-2′-methoxy-2′-deoxycytidylyl-(3′-5′)-2′-fluoro-2′-deoxycytidylyl-(3′-5′)-2′-fluoro-2′-deoxyuridylyl-(3′-5′)-2′-methoxy-2′-deoxcytidylyl-(3′-5′)-2′-methoxy-2′-deoxytidylyl-(3′-5′)-2′-methoxy-2′-deoxyadenylyl-(3′-5′)-2′-methoxy-2′-deoxcytidylyl-(3′-3)-thymidine, or a pharmaceutically acceptable salt thereof.    
     
     
         46 . The method of  claim 44  further comprising contacting the composition with an antidote oligonucleotide.