CORRELATING STRUCTURE AND STABILITY OF DNA DUPLEXES WITH INCORPORATED2'-O-MODIFIED RNA ANALOGS

Chemically modified nucleic acids are currently being evaluated as pot ential antisense compounds for therapeutic applications. 2'-O-Ethylene glycol substituted oligoribonucleotides are second-generation antisen se inhibitors of gene expression with promising features for in vivo u se. Relative to DNA, they display improved RNA affinity and higher nuc lease resistance. Moreover, chimeric oligonucleotides with 2'-O-methox yethyl ribonucleoside wings and a central DNA phosphorothioate window have been shown to effectively reduce the growth of tumors in animal m odels at low doses. Using X-ray crystallography, we have determined th e structures of three A-form DNA duplexes containing the following 2'- O-modified ribothymidine building blocks: 2'-O-methoxyethyl ribo-T, 2' -O-methyl[tri(oxyethyl)] ribo-T, and 2'-O-ethoxymethylene ribo-T. In c ontrast to 2'-O-ethylene glycol substituents, the presence of a 2'-O-e thoxymethylene group leads to slightly reduced RNA affinity of the cor responding oligonucleotides. The three structures allow a qualitative rationalization of the differing stabilities of duplexes between oligo nucleotides comprising these types of 2'-O-modified ribonucleotides an d complementary RNAs. The stabilizing 2'-O-ethylene glycol substituent s are conformationally preorganized for the duplex state. Thus, the pr esence of one or several ethylene glycol moieties may reduce the confo rmational space of the substituents in an oligonucleotide single stran d. In addition, most of these preferred conformations appear to be com patible with the minor groove topology in an A-type duplex. Factors th at contribute to the conformational rigidity of the 2'-O-substituents are anomeric and gauche effects, electrostatic interactions between ba ckbone and substituent, and bound water molecules.