USE OF CYCLODEXTRIN AND ITS DERIVATIVES AS CARRIERS FOR OLIGONUCLEOTIDE DELIVERY

The use of antisense phosphorothioate oligodeoxynucleotides as tools f or modulating gene expression represents a novel strategy for designin g drugs to treat a variety of diseases. Several factors, including cel lular uptake and internalization of the phosphorothioate oligodeoxynuc leotide, are important parameters in determining the effectiveness of antisense agents as therapies. We have used cyclodextrin and its analo gs as carriers to increase cellular uptake of phosphorothioate oligode oxynucleotides. The studies were carried out using S-35-labeled and fl uorescent-labeled phosphorothioate oligodeoxynucleotide in human T cel l leukemia H9 cell line. Cellular uptake of phosphorothioate oligodeox ynucleotide in the presence of cyclodextrin was found to be concentrat ion and time dependent. Using various cyclodextrin analogs, e.g., 2-hy droxypropyl beta-cyclodextrin (HPCD), hydroxyethyl beta-cyclodextrin ( HECD), and a mixture of various hydroxypropyl beta-cyclodextrins (Enca psin), we observed increases in phosphorothioate oligodeoxynucleotide uptake, up to twofold to threefold in 48 hours. Confocal microscopy st udies confirmed that oligonucleotide was present intracellularly. Cycl odextrin itself was not toxic at the concentration used. Cyclodextrins did not seem to affect the efflux of phosphorothioate oligodeoxynucle otide from cells. Stability of phosphorothioate oligodeoxynucleotide a gainst endogenous cellular nucleases remained unchanged in the presenc e of cyclodextrins. These studies suggest that cyclodextrin and its an alogs might be used successfully as carriers for oligonucleotide and a nalogs.