Figure 6.

SRPIN340 as eye drops inhibited choroidal neovascularization dose dependently. Twelve C57/B6 mice were subjected to four laser-lesions (250 mW; 75 μm; 01 second; 810 nm) to the back of the eye on day 0. From day 1 to 14 mice received 20 topical treatments of 10 μL SRPIN340 in a drug delivery vehicle at varying doses (100–0.1 μg/mL) in the contralateral eye and vehicle control in the ipsilateral eye. On day 14 mice were culled, eyes enucleated, and the sclerochoroidal complexes stained for isolectin-B4. Retinal protein was taken for RNA analysis. (A) Flatmounted choroids were imaged and analyzed (ImageJ). SRPIN340 demonstrated dose-dependent inhibition of neovascularisation with an EC₅₀ of 3.187 ug/ml. Images show a vehicle-treated lesion and a 100 μg/mL SRPIN340-treated lesion. (B) VEGF₁₆₅ mRNA expression relative to mouse globin was significantly reduced in 100 μg/mL SRPIN340-treated eyes compared with saline but not at 10 μg/mL or 1 μg/mL (P < 0.05; one-way ANOVA, Bonferroni post hoc). (C) Five micrograms of SRPIN340 in 10 μL was dropped on the left and right eyes of 10 CD-1 mice. Two mice were culled at each of the following time points; 1, 4, 8, 24, and 48 hours, eyes were removed, washed, and dissected into anterior and posterior compartments. Tissue was homogenized, proteins precipitated out, and sample lysate analyzed by mass spectrometry. SRPIN340 was detected in the eye after 1 hour at 175 ng/mL, 3.5% of the total applied dose. Expression in the eye tissue decreased over time with an initial half-life of 1 hour in the whole eye (one-phase exponential decay, prism). (D) The half-life in the posterior chamber was 35 minutes, and at 12 hours 0.15% of the applied dose was detected.