DISTRIBUTION AND STABILITY OF ANTISENSE PHOSPHOROTHIOATE OLIGONUCLEOTIDES IN RODENT BRAIN FOLLOWING DIRECT INTRAPARENCHYMAL CONTROLLED-RATEINFUSION

Object. High-flow microinfusion is a novel technique for delivery of c ompounds directly into brain parenchyma, bypassing the blood-brain bar rier. The feasibility of this technique has been demonstrated with low -molecular-weight compounds, macromolecular dyes, and proteins. Delive ry of antisense oligonucleotides into brain parenchyma represents an a dditional potential application of this technique not previously descr ibed. In this report the authors sought to examine the distribution an d disposition of phosphorothioate oligodeoxynucleotide (PS-ODN) for th is reason. Methods. An 18-mer S-35-PS-ODN (M-r approximately 6000) was infused over 1 hour into the caudate putamen of Fischer 344 rats. At 1, 6, 12, 24, and 48 hours after beginning the infusion, the brains we re extracted and analyzed using quantitative autoradiographic techniqu es. Cerebrospinal fluid (CSF) was also aspirated from the cisterna mag na and was analyzed to determine the radioactivity and stability of th e S-35-PS-ODN. At 1 hour, the infused ODN was uniformly distributed in brain tissue, with a maximum average concentration of 4806.5 +/- 210. 5 nCi/g. This represents a tissue concentration of 19.2 +/- 0.84 mu M. Extensive spread into surrounding parenchyma was observed over the en suing 47 hours. The S-35-PS-ODN radioactivity peaked in the CSF at the end of the 1-hour infusion, containing 1% (50 +/- 20 nCi) of the infu sed radioactivity. Activity then decayed exponentially over 11 hours, but stabilized at a lower CSF content of 0.2% (1 +/- 0.1 nCi) thereaft er. The volume of distribution was 105 +/- 7.9 mm(3) at 1 hour, repres enting a volume of distribution/volume of infusion ratio of 5.2. The v olume of distribution increased to 443 +/- 62.3 mm(3) at the end of 48 hours, whereas the average minimum tissue concentration decreased fro m 15.2 mu M to 3.2 mu M Undegraded 18-mer was observed throughout the 48-hour period by means of 20% polyacrylamide/7 M urea gel electrophor esis. The animals tolerated the infusion without evidence of toxicity and minimal structural changes in tissue were observed on histological -investigation. Conclusions. The authors found that PS-ODNs can be saf ely delivered in high concentrations to wide areas of rat brain by usi ng high-flow microinfusion and are stable even after 48 hours in situ.