A novel non-peptide bradykinin B 2 agonist lowers intraocular pressure ( IOP ) in ocular hypertensive cynomolgus monkeys

Program Number: 2883 Poster Board Number: B0281 Presentation Time: 8:30 AM–10:15 AM BK2A78: A novel non-peptide bradykinin B2 agonist lowers intraocular pressure (IOP) in ocular hypertensive cynomolgus monkeys Ganesh Prasanna1, Naj Sharif1, Byron H. Li1, Mark Hellberg1, Terri Krause1, Shenouda Yacoub1, Daniel Scott1, Curtis R. Kelly1, Iok-Hou Pang2, Keith Combrink1. 1Bio Med (NIBR)/Ophthalmology Research, Novartis, Cambrige, MA; 2Pharmaceutical Sciences, UNT Health Science Center, Fort Wort, TX. Purpose: Bradykinin (BK), a nonapeptide has been shown to regulate IOP in different species including rabbits and monkeys. Presently we intended to characterize the IOP lowering effects of BK2A78, a novel and selective non-peptide bradykinin B2 receptor agonist in ocular hypertensive (OHT) cynomolgus monkeys. Methods: BK2A78 was evaluated in several in-vitro efficacy and receptor binding assays using human cloned B1 and B2 CHO cells and in human ciliary smooth muscle (HCM) cells. Intracellular calcium mobilization ([Ca2+]i) was assessed using FLIPR assay and prostaglandin (PG) release was measured using an EIA assay. Ocular safety assessments including slit lamp examinations were performed in monkeys following topical ocular application of BK2A78. IOP changes were measured using an Alcon computerized pneumatonometer in normal and hypertensive eyes of cynomolgus monkeys. Results: BK2A78 is a selective B2 receptor agonist with EC50 values of 13 ± 5 nM (Emax = 92 ± 1%; BK response was 100%) in [Ca2+]i assay and 12 ± 7 nM (Emax = 100 ± 14%) in the PG release assays respectively. BK2A78 exhibited comparable high affinity binding to B2 receptors (Ki = 3 – 10 nM) with no detectable affinity towards B1 receptors. Topical ocular dosing of BK2A78 (3 ug x 3 times, 1 hour apart) to sedated cynomolgus monkeys caused no flare or cells to appear in the anterior chamber as observed up to 24h post-dose. No other adverse effects were noted. A single topical ocular application of BK2A78 (0.03 – 3 ug) caused a dose-dependent IOP reduction up to 25% from baseline between 6 – 24h post-dose in the hypertensive eyes of conscious cynomolgus monkeys. Maximal percent IOP reduction of 25% was observed at 0.9 – 3 ug doses. The duration of action appeared to last >24h for BK2A78. Conclusions: Bradykinin B2 receptor agonism appears to cause IOP lowering. Unlike the issues surrounding topical ocular application of BK peptide, including non-selectivity against B1 and B2 receptors, poor ocular penetration and susceptibility to rapid degradation by angiotensin converting enzyme, BK2A78 offers several new therapeutic advantages. BK2A78 is a selective non-peptide B2 receptor agonist capable of robust and long lasting IOP reduction that appears to also be well tolerated following topical ocular dosing in OHT monkeys. Commercial Relationships: Ganesh Prasanna, Novartis Inst Biomedical Research (E); Naj Sharif, Alcon Research Ltd (E); Byron H. Li, NIBR (E); Mark Hellberg, NIBR (E); Terri Krause, NIBR (E); Shenouda Yacoub, NIBR (E); Daniel Scott, NIBR (E); Curtis R. Kelly, NIBR (E); Iok-Hou Pang, None; Keith Combrink, None Program Number: 2884 Poster Board Number: B0282 Presentation Time: 8:30 AM–10:15 AM The Use of Tissue Plasminogen Activator to Reduce Elevated Intraocular Pressure Induced by Prednisolone in Sheep Oscar A. Candia1, Rosana Gerometta2, John Danias3. 1Ophthalmology, Mount Sinai School of Medicine, New York, NY; 2Oftalmologia, UNNE, Corrientes, Argentina; 3Ophthalmology, SUNY Downstate Medical Center, Brooklyn, NY. Purpose: We have previously shown (ARVO 2013) that tissue plasminogen activator (tPA) injected into the vitreous of sheep reduced or prevented the elevation of the intraocular pressure (IOP) normally produced by the instillation of 1% prednisolone. In this presentation we report the effect of tPA when injected into the anterior chamber (AC) in amounts of 0.01, 0.001 and 0.0001 μg diluted in a volume of 50 μL. Methods: Lyophilized tPA, obtained as Actilyse ® 50 mg from Boehringer Ingelheim S.A. (Buenos Aires) containing arginine, was used. Five sheep of the Coriedale breed were selected. Initially all eyes received instillation of 1% prednisolone 3 times/day for 10 days to elevate their IOP from 10 mm Hg to about 23 mm Hg. Then, 0.0001 μg was injected into one of the eyes and its effect was followed for up to 55:00 hrs while the instillation of prednisolone continued in both eyes. The same protocol was implemented for the 0.001 and 0.01 μg amounts (after extended washout) in the contra lateral eyes. 0.423 μg of arginine, which is associated with 0.01 μg tPA, was injected alone and had no effect. Results: Injection of 0.0001 μg into the AC had no effect on IOP of 23.0 mm Hg at 6:00 and 30:00 hrs after injection. 0.001 μg reduced IOP from 23.1 to 18.6 mm Hg at 6:00 hr but IOP recovered to 22.3 mm Hg at 30:00 hr. Injection of 0.01 μg produced a marked and prolonged reduction of IOP. From a baseline of 23.4, IOP was reduced to 14.2, 19.0, 20.9, and 22.3 mm Hg at 6:00, 30:00, 48:00 and 55:00 hrs, respectively. Conclusions: Recombinant human tPA is effective in reversing steroid-induced IOP elevation in sheep. The reduction of IOP elevation may be the result of an effect on extra-cellular matrix turnover in the TM. These findings suggest that tPA may be useful as a therapeutic agent in steroid-induced glaucoma. Commercial Relationships: Oscar A. Candia, None; Rosana Gerometta, None; John Danias, None Support: NIH EY020670, RPB (Candia & Danias); UNNE CONICET (Gerometta)