Ripasudil liposomes with transferrin targeting for Fuchs endothelial corneal dystrophy

Abstract

Purpose : Currently, surgery is the only definitive treatment for Fuchs endothelial corneal dystrophy (FECD). We aimed to develop Ripasudil loaded transferrin (TF) conjugated corneal endothelial cell (CEC) targeting liposomal eye drops to prevent FECD progression with reduced side-effects. TF was used as it targets transferrin receptors, which we have shown previously to have increased expression in FECD.

Methods : Liposomes were synthesized using lipids and DSPE-PEG(2000)COOH with microfluidics. DSPE-PEG(2000)COOH was conjugated to TF by EDC-NHS chemistry. Ripasudil was loaded in liposomes. Liposomes were characterized by measuring size and charge using Zetasizer. Drug loading and encapsulation efficiency were quantified using HPLC-DAD. Formulation stability was examined for 3 months at 4°C and 25°C. TF liposome ability to penetrate mouse corneas and produce controlled drug release was evaluated by LC-MS/MS. TF ligand effect on liposome uptake was studied. Safety and efficacy of Ripasudil TF liposomes, sham TF liposomes, and commercial Ripasudil solution were compared by clinical examination in mice with a well-characterized FECD mutation (129S6/SvEvTac Col8a2^Q455K), followed by postmortem RNA-seq analysis using ΔΔCt method for ZO-1, N-Cadherin and Na⁺/K⁺ ATPase.

Results : Liposomal particle size was determined to be 96.69±1.69 nm with a narrow polydispersity index (PDI) of 0.14±0.01 (n=4). Ripasudil TF liposomes were stable for 3 months at 4°C and at 25°C. At 5-, 10- and 30-min time points after drug administration, Ripasudil TF liposomes achieved a longer drug residence compared to commercial solution, which rapidly declined over time (each data point represents, drug pooled from 4 CEC peels). Liposomal therapy had a good safety profile and low off-target side-effects in this mouse model. RNA-seq data showed elevated levels of ZO-1, N-Cadherin and Na⁺/K⁺ ATPase in mouse groups receiving Ripasudil TF liposomes.

Conclusions : Findings indicate the successful development of Ripasudil loaded TF conjugated liposomes as a targeted drug delivery strategy for CECs. This formulation increases residence time in CECs and may reduce conventional dosing frequency without off-target effects compared to commercial drug solution.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.