It has been reported that the presence of a hydrophobic moiety at the 5-position of 2'-deoxyuridine increases duplex stability when incorporated into an oligonucleotide. A series of 2'-deoxyuridine analogues modified at the 5-position have been synthesised. The stability of a 12mer duplex containing these residues was analysed by ultraviolet melting studies. Of the groups tested 5-propynyl-2'-deoxyuridine imparted greatest stability on the duplex. Thermodynamic parameters show that the reason for increased stability arises from a decrease in enthalpy which is related to the base stacking process. Ultra high field NMR was used to elucidate the 3D structures of a deoxydodecanulceotide containing 5-propynyl-2'-deoxyuridine residues at two different positions. These structures show an increased π-π overlap with the base on the 5'-side of the modified base. This suggests the degree of stabilisation imparted by the propyne moiety may be sequence dependent. Cellular uptake of antisense oligonucleotides is particularly low. In an attempt to improve uptake a cholesterol moiety linked by two different lengths of alkyl spacer was attached to an antisense oligonucleotide. Anti-HIV activity of the cholesteryl conjugated oligonucleotide was improved relative to the unmodified oligonucleotide. Pharmacokinetic properties of the cholesteryl conjugated oligonucleotide was less than that of the unmodified oligonucleotide.