Glaucoma is a multi-factorial blinding disease in which genetic factors play an important role. Elevated intraocular pressure is a highly heritable risk factor for primary open angle glaucoma and currently the only target for glaucoma therapy. Our study helps to better understand underlying genetic and molecular mechanisms that regulate intraocular pressure, and identifies a new candidate gene, Cacna2d1, that modulates intraocular pressure and a promising therapeutic, pregabalin, which binds to CACNA2D1 protein and lowers intraocular pressure significantly. Because our study utilizes a genetically diverse population of mice with known sequence variants, we are able to determine that the intraocular pressure-lowering effect of pregabalin is dependent on the Cacna2d1 haplotype. Using human genome-wide association study (GWAS) data, evidence for association of a CACNA2D1 single-nucleotide polymorphism and primary open angle glaucoma is found. Importantly, these results demonstrate that our systems genetics approach represents an efficient method to identify genetic variation that can guide the selection of therapeutic targets.

Delivery of therapeutic molecules into the deeper ocular compartments is mainly hampered by short precorneal residence and limited transmembrane permeability of topically administered drugs. Hence, the current study was undertaken to fabricate the ion-sensitive in situ gels of natamycin (NT) bilosomes (NB) for efficient ocular delivery. The effect of cholesterol and sodium taurocholate proportion on the properties of the bilosomes were studied and the formulation with better physicochemical properties was optimized and utilized to derive the in situ gelling system (IG). The impact of type/composition of gelling agent on the formation and characteristics of the hydrogel was investigated. The hydrogel formed from IG with 0.3% w/v gellan gum showed optimal viscoelastic and adhesive characteristics. The ocular safety and cytocompatibility of NB and its IG was confirmed by corneal histology and in vitro cytotoxicity evaluation. A 6- to 9-fold enhancement in the transcorneal flux of NB demonstrated efficient ocular penetration of bilosomes. Moreover, the superior mean dose normalized NT levels in the ocular tissues of rabbits treated with optimized NB and IG illustrated the effectiveness of bilosomes loaded ion-sensitive in situ hydrogels as a potential platform for the improved and prolonged ocular pharmacotherapy.

Elevated intraocular pressure (IOP) is the most significant risk factor contributing to visual field loss in glaucoma. Unfortunately, the deficiencies associated with current therapies have resulted in reduced efficacy, several daily dosings, and poor patient compliance. Previously, we identified the calcium voltage-gated channel auxiliary subunit alpha2delta 1 gene (Cacna2d1) as a modulator of IOP and demonstrated that pregabalin, a drug with high affinity and selectivity for CACNA2D1, lowered IOP in a dose-dependent manner. Unfortunately, IOP returned to baseline at 6 h after dosing. In the current study, we develop a once daily topical pregabalin-loaded multiple water-inoil-in-water microemulsion formulation to improve drug efficacy. We characterize our formulations using multiple in vitro and in vivo evaluations. Our lead formulation provides continuous release of pregabalin for up to 24 h. Because of its miniscule droplet size (<20 nm), our microemulsion has a transparent appearance and should not blur vision. It is also stable at one month of storage at temperatures ranging from 5 to 40 °C. Our formulation is nontoxic, as illustrated by a cell toxicity study and slit-lamp biomicroscopic exams. CACNA2D1 is highly expressed in both the ciliary body and the trabecular meshwork, where it functions to modulate IOP. A single drop of our lead pregabalin formulation reduces IOP by greater than 40%, which does not return to baseline until >30 h post-application. Although there were no significant differences in the amplitude of IOP reduction between the formulations we tested, a significant difference was clearly observed in their duration of action. Our multilayered microemulsion is a promising carrier that sustains the release and prolongs the duration of action of pregabalin, a proposed glaucoma therapeutic.