(W1130-01-01) Development of an Intraocular In Situ-Forming Implant of a Novel Compound to Target the Retina in a Rabbit Model

Purpose: Vision loss and blindness affect more than 1 billion people worldwide. The leading causes of irreversible blindness are posterior segment-related diseases including age-related macular degeneration, retinitis pigmentosa, and diabetic retinopathy. The posterior segment of the eye poses many challenges for effective drug delivery that may be bypassed with an extended-release dosage form. Here, we present our efforts to develop a long-acting intraocular injection for a novel compound with a target in the retina. The formulation reported is the initial formulation for use during preclinical testing. The ultimate goal is to develop an extruded implant as a model for the final dosage form. This work is part of a collaborative project and represents one step in an ongoing investigation of a series of compounds. The data presented is for the representative compound in the series.

Methods: In vivo pharmacokinetic(PK) testing was conducted in New Zealand white rabbits. In the first PK study (PK1), the compound was dissolved in dimethyl sulfoxide(DMSO) to a concentration of 12 ug/uL. A volume of 50 uL (600 ug) was injected into the vitreous space of the animal model. At set timepoints the animals were sacrificed, and eyes harvested, to provide samples at 45 minutes, 9 days, 12 days, and 16 days. Retina and vitreous humor samples were isolated and stored at -80°C. For PK2, n-methyl pyrrolidone(NMP) was used to dissolve the compound, both with and without a polymer, to create immediate and long-acting formulations. Poly lactic-co-glycolic acid 50-50 (PLGA) was chosen for the polymer. The formulations were made so a 50 uL injection would deliver a 600 ug dose. Each formulation was injected into the eye of the animal model. At set timepoints, 1 day and 16 days, animals were sacrificed, eyes harvested, and retina and vitreous humor isolated and stored at -80°C. PK3 was designed based on results from previous studies. DMSO was used as the solvent, only the implant formulation was used, and the duration of the study extended, to timepoints 1 day, 15 days, and 30 days. All tissue samples were homogenized and underwent a purification process before being analyzed via LCMS for drug concentration. Finally, an in vitro release study of the in situ-forming implant was started. A 50 uL dose of the DMSO formulation was injected into a vitreous humor substitute. The implants were placed in an incubator at 37°C under agitation and allowed to harden before being transferred to a dialysis apparatus in a vial containing phosphate-buffered saline(PBS). At set timepoints, the implants are transferred to a vial of fresh solution. Samples of starting vitreous humor substitute and PBS are stored at -80°C until LCMS analysis.

Results: PK1 measured concentration of the compound in the retina and vitreous humor over a 16-day period. After 45 minutes, retina and vitreous humor contained 16985.76±4442.93 ng and 13881.64±927.49 ng respectively. On day 9, 1680.09±722.64 ng and 30369.07±37346.69 ng of compound was measured in the retina and vitreous humor. On day 12, retina and vitreous humor contained 230.58±284.41 ng and 2273.05±3205.31 ng respectively. Drug in retina and vitreous humor samples was still detectable on day 16, measuring 472.01±680.09 ng and 3513.52±4957.06 ng respectively. PK2 compared the immediate and long-acting formulations. At day 1, retina and vitreous humor treated with the immediate formulation contained 11640.89±9484.68 ng and 120895.3±49698.29 ng of DKR-1677; the retina and vitreous humor treated with the long-acting formulation contained 711.29±359.14 ng and 6119.8±346.95 ng of compound. At day 16, the retina and vitreous humor samples treated with the immediate and long-acting formulations contained 94.79±227.61 ng, 189.52±431.6 ng, and 1355.26±902.82 ng, 2165.99±369.37 ng of drug respectively. Unfortunately, the solvent, NMP, was found to irritate the eye of the animal model and was not used for PK3. PK3 tested the long-acting formulation with DMSO as the solvent and lengthened the study duration. At day 1, retina and vitreous humor contained 21971.88±6724.19 ng and 69648.86±39272.94 ng of drug respectively. At day 15, retina and vitreous humor contained 62417.16±30602.49 ng and 83299.62±27341.24 ng. At day 30, retina and vitreous humor contained 634.31±181.21 ng and 1634.23±744.43 ng of drug respectively. Results from the on-going in vitro release study will be compared to data shown here.

Conclusion: Our goal was to develop an in-situ-forming implant for intravitreal injection. The PK studies reported here demonstrated the compound is a good candidate for this drug target and it is retained in the retina for a sufficient duration when formulated with a polymer. The long-acting formulation slowed the release of the drug resulting in an increased drug concentration in both the retina and vitreous humor at day 16, with levels detectable at day 30. The PK data shown here indicates the in-situ-forming implant is a viable formulation approach to extend drug presence at the target site during preclinical testing. Future work includes developing an extruded implant and improving biorelevance of in vitro release methods.

References: 1. Awwad, S., et al., Principles of pharmacology in the eye. Br J Pharmacol, 2017. 174(23): p. 4205-4223.
2. Kim, H.M. and S.J. Woo, Ocular Drug Delivery to the Retina: Current Innovations and Future Perspectives. Pharmaceutics, 2021. 13(1).

Acknowledgements: This work was supported by the Gund Harrington Foundation and the Foundation for Fighting Blindness.