The effect of JV-GL1 in ocular hypertensive mice
We then examined the effect of JV-GL1 in C57BL/6J mice with ocular hypertension. We induced ocular hypertension by periocular injection of dexamethasone-eluting nanoparticles given bilaterally. This elevated IOP by 4.2 [3.5, 4.9] mmHg relative to baseline, and this elevation was sustained over 2 weeks following a single nanoparticle injection without a significant loss of body weight (n=18; Supplemental Figure 2). Ocular hypertensive mice were treated with a single topical drop of JV-GL1 (10 µl, 0.01%) given bilaterally. As with the normotensive mice, IOP was reduced after 3 hrs relative to the pre-treatment IOP (δIOP = -2.3 [-3.3, -1.2] mmHg; P =0.01; n=9 mice, Fig. 2A). However, unlike the normotensives, the IOP reduction persisted for up to 6 days when δIOP = -2.8 [-3.7, -2.0] mmHg (P =0.001; n=9). Thus, a single treatment of JV-GL1 reduces IOP for several days in mice with steroid-induced ocular hypertension, in contrast to results from a different cohort of normotensive mice where the IOP-lowering effects of JV-GL1 lasted for less than 1 day (Fig. 1A)
After 9 days, the effects of JV-GL1 in hypertensives had diminished and IOP had returned to pre-treatment levels (δIOP = -0.5 [-1.5, 0.5] mmHg; P =1; n=9), which were still elevated with respect to baseline due to dexamethasone. We then administered a second dose of JV-GL1 (10 µl, 0.01%) to one randomly-chosen eye of each mouse, while the contralateral eye received vehicle (Fig. 2A second red arrow ). After 3 days, IOP was again reduced in the JV-GL1-treated eye by -2.3 [-3.4, -1.2] mmHg (P= 0.04; n=9) relative to the contralateral eye, demonstrating at least partial repeatability. The mice were then euthanized 3 days after JV-GL1 treatment, and eyes were enucleated to measure outflow facility. In eyes treated with JV-GL1, outflow facility was not statistically different from that in contralateral vehicle-treated eyes (18% [-7, 50], P =0.2, n=9 pairs; Fig 2B). Thus, the IOP reduction observed in response to JV-GL1 in ocular hypertensive mice does not appear to coincide with an increase in outflow facility, similar to the absence of a facility response observed in normotensive mice.
To confirm that the long-lasting IOP reduction of JV-GL1 is not attributable to cohort differences between the normotensive and hypertensive groups, we compared the effects of JV-GL1 between wildtype littermates with or without ocular hypertension. Hypertension was induced by bilateral periocular injection of dexamethasone-eluting nanoparticles, while normotensives received a bilateral injection of unloaded nanoparticles. In the group receiving dexamethasone-eluting nanoparticles, IOP increased by 4.7 [3.4, 6.0] mmHg after 1 week, while IOP remained near baseline in the group receiving unloaded nanoparticles (∆IOP = 0.0 [-1.6, 1.6] mmHg; Supplemental Figure 3). In the hypertensives, a single topical drop of JV-GL1 given unilaterally (10 µl, 0.01%) reduced IOP after 3 hrs by 3.1 [-4.4, -1.8] mmHg (P =0.01, n=8; Fig. 3). The IOP reduction in hypertensives persisted for up to 4 days (∆IOP = -2.8 [-3.8, -1.8] mmHg,P =0.04), but had returned to pre-treatment levels by 6 days (∆IOP = -1.4 [-2.9, 0.2] mmHg, P =0.5). In normotensive mice, however, the IOP reduction was short-lived and was observed only at 3 hrs (∆IOP = -1.6 [-2.7, -0.5] mmHg, P =0.08; n=9). This confirms that a single dose of JV-GL1 is capable of reducing IOP for several days in mice with steroid-induced ocular hypertension, but the effect of JV-GL1 is relatively short-lived in normotensive C57BL/6J littermates.
To examine whether the long duration of JV-GL1 action was distinct from typical EP2 agonism, we treated ocular hypertensive mice with a single bilateral dose of butaprost (10 µl, 0.01%). Butaprost reduced IOP after 3 hrs (∆IOP = -2.5 [-4.2, -1.0] mmHg;P =0.07; n=9; Fig. 2A) but, as observed in normotensive mice, IOP returned to pre-treatment values by 2 days (∆IOP = ‑0.8 [-1.5, 0.0] mmHg; P =0.6; n=9). This reveals that the long-lasting IOP reduction attributable to JV-GL1 under hypertensive conditions is distinct from that of typical EP2 agonists.