Synthesis of 1,N⁶Ethano-2′-deoxyadenosine, a Metabolic Product of 1,3-Bis(2-chloroethyl)nitrosourea, and Its Incorporation into Oligomeric DNA

1,N⁶-Ethano-2′-deoxyadenosine (1) is one of the adducts formed during DNA reaction with the antitumor agent, 1,3-bis(2-chloroethyl)nitrosourea (BCNU), and was synthesized and incorporated into a site-specific deoxyoligonucleotide for the first time. The product 6-chloropurine-2′- deoxyriboside (11) was prepared in high yield by the reaction of 2′-deoxyinosine (6) with SOCl₂, which then was derivatized to give compound 12 using tert-butyldimethylsilyl chloride, which was then reacted with 2-hydroxyethylamine to produce compound 13 in 86% yield. Reaction of 13 with (PhO)₃P^-MeI^- in DMF gave the cyclized 1,N⁶-ethano derivative 10 in 67% yield. Desilylation of 10 with triethylamine trihydrofluoride in THF gave 1N⁶-ethano-dA (1) in 91% yield. Tritylation of compound 1 with DMT⁺BF₄^- gave the 5′-O-DMT product 14 in 62% yield, which then was phosphitylated with 2-cyanoethyl N,N-diisopropylchlorophosphoramidite, which yielded a 1:1 mixture of the diastereoisomers 15 in 71% yield. This fully protected compound 15 was incorporated site-specifically into a 25-mer oligonucleotide. The coupling efficiency of ethano-dA phosphoramidite was 93%. Enzymatic hydrolysis and analysis by HPLC confirmed the incorporation of ethano-dA and base composition of the DNA oligomer. The latter is now under investigation for its biochemical and physical properties.