Uptake dynamics and retinal tolerance of phosphorothioate oligonucleotide and its direct delivery into the site of choroidal neovascularization through subretinal administration in the rat

This study aimed to investigate uptake dynamics and retinal tolerance of ph osphorothioate oligonucleotides (PS-oligos) following subretinal injection. A fluorescent-labeled PS-oligo (FL-oligo) with random sequence was adminis tered into the subretinal space of rat by transsclera-choroid-retinal pigme nt epithelium (RPE) injection at doses of 0.129, 1.29, and 12.9 mug in 2.0 mul solution. The uptake dynamics were evaluated by fundus fluorescent phot ography in real time and by fluorescence microscopy using flat mounts and c ryosections. Immunophenotyping for CD4(+), CD8(+) cytotoxic lymphocytes, an d CD68(+) macrophages was performed to assess cellular infiltration in the retina. In addition, the FL-oligo was injected subretinally in a rat model of choroidal neovascularization (CNV) for direct delivery into the site of CNV. Subretinal administration of FL-oligo resulted in both dose-dependent and time-dependent distribution in the retina, where it accessed the RPE an d all layers of the neuroretina. CD4(+), CD8(+) cytotoxic lymphocytes, and CD68+ macrophages were observed at the site of needle penetration. However, in areas far from the injection site where the FL-oligo appeared strongly, cellular infiltration was absent, and the retinal morphology was preserved very well. The FL-oligo was successfully delivered into the site of intens e laser photocoagulation. It was predominantly localized to the RPE, macrop hages, and some choroid cells and remained detectable for at least 56 days after injection. Our results demonstrate for the first time that subretinal injection efficiently introduced PS-oligo into the RPE and neuroretina wit h an acceptable level of safety. Subretinal administration of antiangiogeni c oligonucleotides may hold great potential for the treatment of CNV.