Novel antisense oligonucleotides targeting TGF-beta inhibit in vivo scarring and improve surgical outcome.

The scarring response is an important factor in many diseases throughout the body. In addition, it is a major problem in influencing results of surgery. In the eye, for example, post-operative scarring can determine the outcome of surgery. This is particularly the case in the blinding disease glaucoma, where several anti-scarring regimens are currently used to improve glaucoma surgery results, but are of limited use clinically because of severe complications. We have recently identified transforming growth factor-beta (TGF-beta) as a target for post-operative anti-scarring therapy in glaucoma, and now report the first study of novel second-generation antisense phosphorothioate oligonucleotides against TGF-beta in vivo. Single applications of a TGF-beta OGN at the time of surgery in two different animal models closely related to the surgical procedure performed in glaucoma patients, significantly reduced post-operative scarring (P<0.05) and improved surgical outcome. Our findings suggest that TGF-beta antisense oligonucleotides have potential as a new therapy for reducing post-surgical scarring. Its long-lasting effects after only a single administration at the time of surgery make it particularly attractive clinically. Furthermore, although we have shown this agent to be useful in the eye, it could have widespread applications anywhere in the body where the wound-healing response requires modulation.

Antiviral activity and ocular kinetics of antisense oligonucleotides designed to inhibit CMV replication.

PURPOSE: To compare the antiviral activity and ocular distribution of first- and second-generation antisense oligonucleotides intended for the treatment of cytomegalovirus (CMV) retinitis. METHODS: The antiviral activity of ISIS 13312 and ISIS 2922 (Isis Pharmaceuticals, Inc., Carlsbad, CA) against 10 clinical CMV isolates was compared with a plaque-reduction assay. The ocular pharmacokinetics were compared after intravitreal injection in rabbits (36-90 microg) and monkeys (125-500 microg). Vitreous and/or retina were collected after single and multiple injections to characterize ocular distribution, clearance, and accumulation. Oligonucleotide concentrations were measured by capillary gel electrophoresis and immunohistochemical techniques. RESULTS: ISIS 13312 and ISIS 2922 demonstrated comparable antiviral activity that was consistent among the 10 clinical isolates examined (50% inhibitory concentration [IC(50)], <1 microM). Activity was independent of the resistance of CMV isolates to DNA polymerase inhibitors. After intravitreal injection, the kinetics of ISIS 2922 and ISIS 13312 were characterized by clearance from vitreous and distribution to the retina; however, ISIS 2922 was cleared more quickly from the retina than ISIS 13312. The half-life of ISIS 13312 in the monkey retina was approximately 2 months. Retinal concentrations of ISIS 13312 were dose dependent, with approximately a twofold increase in concentration after once-monthly doses compared with single-dose concentrations. Immunohistochemical analysis indicated that both oligonucleotides were efficiently distributed to numerous ocular tissues, including retina, ciliary body, and optic nerve. CONCLUSIONS: ISIS 13312 possesses antiviral activity and pharmacokinetic properties that favor its use as a therapeutic agent in treatment of CMV retinitis. The half-life of ISIS 13312 in retina is longer than that of ISIS 2922, potentially allowing for less frequent administration.

Absorption of antisense oligonucleotides in rat intestine: effect of chemistry and length.

An in situ single-pass perfusion model was used to assess the effect of chemical modification and length on permeability and absorption of various oligonucleotides in rat intestine. Phosphorothioate oligodeoxynucleotides (PS-ODN) were compared with oligoribonucleotides with 2'-methoxyethyl (MOE) or 2'-O-methyl (OMe) modifications. A 25-mer PS-OMe-modified oligonucleotide showed relatively poor permeability in this model, as did unmodified 20-mer PS-ODN (permeability coefficient [P(eff)] = 2-8 X 10(-6)cm/sec). Modifying some or all of the oligonucleotides with 2'-MOE groups on deoxyribose and 5'-methylation of the cytosines substantially increased intestinal permeability of oligonucleotides. Both partially and fully modified PS-MOE oligonucleotides showed a (2-4)-fold increase in permeability as compared with unmodified PS-ODN. The presence of a phosphodiester backbone in MOE-modified compounds led to further increases in intestinal permeability. PS-MOE composed of 6, 8, 10, 12, 14, 16, 18, 20, and 22 nucleotides were also examined. It was found that the permeability of these oligonucleotides increased linearly with decreasing length.