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Whitney Shatz, Nikhil Peer, Philip Hass, Daniela Bumbaca, Kelly Loyet, Craig Blanchette, Robert Kelley; Identification and characterization of a novel protein-PEG conjugate system for long-acting delivery to the back of the eye. Invest. Ophthalmol. Vis. Sci. 2018;59(9):243. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To identify a Fab-PEG conjugate system with biophysical and pharmacokinetic attributes desirable for long-acting delivery to the back of the eye
Polyethylene glycol (PEG) structures with varied chain length, geometry and degree of branching, were selected and coupled to fragment antibody (Fab) using maleimide chemistry. Previously, it has been demonstrated that hydrodynamic radius (RH) is a key contributor of vitreal half-life (Gadkar et al, 2015, Shatz et al., 2016) and thus RH was used to rank the conjugates in vitro. Specifically, RH of each Fab-PEG conjugate was measured using size exclusion chromatography in-line with a quasi-elastic light scattering detector (SEC-QELS). The RH screen identified two potential Fab-PEG candidates, which were further differentiated using additional biophysical analyses. Once a primary candidate was selected, a large-scale Fab-PEG production scheme was developed and half-lives of the naked Fab and Fab-PEG conjugate were compared using a cynomolgous monkey pharmacokinetic (PK) ocular in vivo model.
A suitable candidate for LAD back of the eye delivery was identified, demonstrating purity and process scalability, stability, and half-life extension. Indeed, intravitreal (IVT) injection of both molecules at similar Fab doses demonstrated an approximately 2-fold increase in half-life of Fab-PEG as compared to Fab alone in the cyno PK model.
While RH proved suitable for in vitro screening of molecules, additional biophysical analysis was necessary to differentiate the top candidates. Scale-up production of the lead candidate successfully demonstrated achievable molecular definition, even with a polydisperse molecule such as PEG. In vivo cyno PK analysis corroborated the prediction of half-life extension, further solidifying the hypothesis that a major driver of half-life extension in the vitreous is solution size.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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