Synthesis of 1,N6-Ethano-2‘-deoxyadenosine, a Metabolic Product of 1,3-Bis(2-chloroethyl)nitrosourea, and Its Incorporation into Oligomeric DNA

1, N 6 -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 SOCl 2, 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) 3 P + MeI - in DMF gave the cyclized 1, N 6 -ethano derivative 10 in 67% yield. Desilylation of 10 with triethylamine trihydrofluoride in THF gave 1, N 6 -ethano-dA ( 1 ) in 91% yield. Tritylation of compound 1 with DMT + BF 4 - 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.