Research Progress Report by Morgan Fedorchak, PhD, and Ken Nischal, MD, FRCO, University of Pittsburgh

Morgan Fedorchak, PhD, Assistant Professor, and Ken Nischal, MD, FRCO, Professor, Chair of Pediatric Ophthalmology at the University of Pittsburgh, received their most recent grant award in spring 2020 for their work to develop a controlled release cysteamine eye drop. CRF has supported and funded their important research for corneal cystinosis since 2016 with $880,258 in research grants. This report provides the most recent research progress as of Fall 2021.

Controlled Release Cysteamine Eye Drop Development

Corneal cystine crystal accumulation in patients with cystinosis is treated by hourly administration of topical cysteamine eye drops. Topical ophthalmic cysteamine has proven to be effective in dissolving corneal cystine crystals but requires extremely frequent administration, up to once per waking hour, to achieve clinically relevant results. Cysteamine is also highly susceptible to oxidative degradation, thus requiring that the eye drops be frozen until opening, stored in the refrigerator, and disposed of within one week. The strict dosing regimen and high concentration of drug per drop make this treatment inconvenient and painful for patients. The purpose of this study is to develop and test a controlled release formulation that provides one full day of cysteamine therapy in a single eye drop.

To this end, our group has developed a thermoresponsive gel drop that contains spray-dried, cysteamine-loaded microspheres. We recently published a peer-reviewed article in the journal Drug Delivery and Translational Research describing this system, click here to download the published article. This combined system is designed to reside beneath the lower eyelid and release drug in a sustained fashion. We have also validated a robust and novel spectroscopy method for assessing drug stability over time, with secondary validation using chromatography. This allows us to closely monitor shelf-life and activity of the stored drug formulation. The current formulation is stable for over 7 weeks at 4C.

In one recent study, we quantified drug levels over time eluting from the gel drop in our healthy rabbit biodistribution model. All samples were analyzed using a LC/MS method. To summarize our findings, drug concentration in cornea and aqueous humor were comparable between the cysteamine gel eye drop and standard eye drops after 24 hours (1 drop versus 12). While standard eye drops showed higher drug levels at earlier time points, it is important to note that those are discrete measurements after which drug concentration would decrease rapidly before the next hourly drop. In the case of the gel eye drop, the drug level can be assumed to decrease in a much more gradual, linear fashion. Thus the total amount of drug presented is the area under the curve for the full 24 hours. Vitreous humor concentration for the gel and aqueous drops were comparable at all time points and suggests high levels of intraocular absorption. As expected, plasma levels were lower than those in the eye at all time points. IOP was within normal range at all time points and didn’t not show any evidence of trending upward or downward. Rabbit grimace scores suggest a mild, transient sensitivity from all treatments resolving within one hour of treatment. Again it is important to note that for the gel drop this sensitivity would be once per day, while for traditional drops this effect would occur repeatedly throughout the day.

We have confirmed that drug release from the scalable, GMP-grade spray-dried formulation, as with our previous double emulsion microspheres, is sustained longer in the presence of gel. We would currently recommend a dosing frequency of once per day based on our rabbit data. We sought to test the efficacy of that dosing frequency in the CTNS-/- mouse. These studies were completed using a small Tegaderm film to hold the blank or cysteamine-loaded gel in place, as the inferior fornix volume on a mouse is of insufficient volume to properly test the system. Our initial testing showed that mice were able to eat and drink normally fitted with the Tegaderm covering and an Elizabethan collar for several weeks. We used longitudinal OCT imaging to semi-quantitatively evaluate crystal density and location using pixel intensity for comparison.

Pixel intensity can be interpreted to suggest that cysteamine treatment is decreasing crystal density compared to the control for a ratio of OD/OS <1. OD/OS=1 would suggest no net positive or negative effect from treatment, and OD/OS>1 would suggest an increase in crystal density (or some other change to the cornea).  As expected, the cysteamine eye drops showed OD/OS<1. Both the blank and cysteamine loaded gel drops showed OD/OS>1 and a marked increase in central corneal thickness. We interpret these results to indicate that the application method (Tegaderm plus Elizabethan collar) interfered with our ability to determine the efficacy of the gel drop. Corneal health deteriorated over time for several mice in those groups, mostly in the form of corneal edema and slight opacification, but none of our results to date have suggested any toxicity of the test materials. Similarly, the fact that this occurred in both the blank and cysteamine groups (and that the blank materials have undergone substantial preclinical testing in other projects) further suggests it is an issue with the application and not the materials. For this reason, we are planning to repeat the mouse study without Tegaderm on the eye.