Nucleosides and Oligonucleotides for Studies on Reversal of Cytotoxic and Mutagenic Damage of DNA and as Diagnostics Tools

Abstract

The present invention is directed to n-alkylated synthetic nucleosides of high regiospecific purity and oligonucleotides that can be utilized for studies on reversal of cytotoxic and mutagenic DNA damage, and as diagnostic tools.

What is claimed is: 
     
         1 . A nucleoside of the formula (II), 
       
         
           
           
               
               
           
         
       
       wherein,
 R 1  is methyl; 
 X is hydrogen, a lower alkyl containing 1 to 6 carbon atoms, or a branched chain lower alkyl; 
 Y is monomethoxytrityl, dimethoxytrityl, trimethoxytrityl, or tetrahydropyranyl; 
 R′ is a phosphoramidite, said phosphoramidite is
   P—N(R 2 ,R 3 )(OCH 2 CH 2 CN) or P—N(R 2 ,R 3 )(OR 4 ),
 
 
 
       wherein R 2  and R 3  are selected from the group consisting of diisopropyl, diethyl, dimethyl, morpholino, and pyrrolidino; and R 4  is a lower alkyl containing 1-6 carbon atoms; and
 Z is hydrogen, fluoro, or protected amino. 
 
     
     
         2 . A nucleoside of the formula (VIII), 
       
         
           
           
               
               
           
         
       
       wherein,
 R 1  is methyl; 
 Y is monomethoxytrityl, dimethoxytrityl, trimethoxytrityl, or tetrahydropyranyl; 
 R′ is a phosphoramidite, said phosphoramidite is
   P—N(R 2 ,R 3 )(OCH 2 CH 2 CN) or P—N(R 2 ,R 3 )(OR 4 ),
 
 
 
       wherein R 2  and R 3  are selected from the group consisting of diisopropyl, diethyl, dimethyl, morpholino, and pyrrolidino; and R 4  is a lower alkyl containing 1-6 carbon atoms;
 Z is hydrogen, fluoro, or protected amino; and 
 R 5  is phenoxy acetyl, para or ortho chloro phenoxy acetyl, para or ortho nitro phenoxy acetyl, para or ortho bromo phenoxy acetyl, pivaloyl, 9-fluorenylmethoxycarbonyl (9-FMOC), or a dialkylformamidine. 
 
     
     
         3 . A method of synthesizing a defined sequence of an oligo-2′-deoxyribonucleotide, comprising the step of:
 reacting the nucleoside phosphoramidite of one of  claim 1  or  2  with an oligo-2′-deoxynucleotide to produce said defined sequence of oligo-2′-deoxyribonucleotide. 
 
     
     
         4 . The defined sequence of oligo-2′-deoxyribonucleotide of  claim 3 , wherein the nucleoside exhibits at least 95% regiospecific purity. 
     
     
         5 . An in vitro diagnostic agent for reversal of cytotoxic and mutagenic damage to DNA, comprising:
 said defined sequence of  claim 3 , wherein the nucleoside phosphoramidite exhibits at least 95% regiospecific purity.   
     
     
         6 . An in vivo diagnostic agent for reversal of cytotoxic and mutagenic damage to DNA, comprising:
 said defined sequence of  claim 3 , wherein the nucleoside phosphoramidite exhibits at least 95% regiospecific purity.   
     
     
         7 . A process for preparing N3 alkyl-deoxycytidine, said process comprising the steps of:
 reacting N-4-benzoyl-5′-dimethoxytrityl-2′-deoxycytidine with an alkyl iodide to produce N-3 alkyl N-4-benzoyl-5′-dimethoxytrityl-2′-deoxycytidine; and   converting N-3 alkyl N-4-benzoyl-5′-dimethoxytrityl-2′-deoxycytidine into the corresponding phosphoramidite.   
     
     
         8 . The process of  claim 7 , further comprising the step of:
 removing protecting groups N-4-benzoyl and 5′-dimethoxytrityl from N-3 alkyl N-4-benzoyl-5′-dimethoxytrityl-2′-deoxycytidine to produce N-3-alkyl-2′-deoxycytidine.   
     
     
         9 . An oligo-2′-deoxyribonucleotide, comprising at least one of the nucleoside of  claims 1  and  2 .