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Wan Wang, Pu Shi, Suhaas Aluri, Pang-Yu Hsueh, Maria Edman, Denise Ryan, Robert McKown, Sarah Hamm-Alvarez, Gordon Laurie, John MacKay, Department of Pharmacology and Pharmaceutical Sciences, University of Southern California; Bridging ocular therapeutics and contact lenses via thermo-responsive protein polymers. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2668.
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To influence the bioavailability of the prosecretory mitogen lacritin, we fused it with elastin-derived protein polymers, elastin-like polypeptides (ELPs) that have a range of assembly properties. ELPs reversibly phase separated onto ProclearTM contact lens and may provide an alternative route for delivery of protein therapeutics to the ocular surface system.
Lac-ELP genes were expressed in E. coli and purified utilizing Inverse Transition Cycling (ITC) followed by size exclusion chromatography. Phase behavior and nanoparticle formation of fusion proteins were characterized by optical density, Dynamic Light Scattering (DLS), TEM and Cryo-TEM. In vitro activities were tested using rabbit lacrimal gland acinar cells (LGACs) and SV40-immortalized human corneal epithelial cells (HCE-T). In vitro ProclearTM contact lens retention was characterized by fluorescence detection.
A novel fusion protein library based on recombinant human lacritin and elastin-like-polypeptides (ELPs) was designed as a potential therapeutic for the ocular surface. The Lacritin-ELP fusion proteins imparted thermo-sensitive assembly of micron-sized coacervates or 130-140nm diameter micelles depending on the sequence of their ELP tag. Exogenous Lac-ELPs promoted β-hexosaminidase secretion from rabbit lacrimal gland acinar cells (LGACs); evoked Ca2+ wave propagation across Human Corneal Epithelial Cells (HCE-T) and were taken up by both LGACs and HCE-T cells. ELPs reversibly attached onto ProclearTM contact lens and the total retention time was temperature and Tt dependent.
Exploration of thermo-responsive prosecretory mitogen lacritin-ELPs may provide an alternative approach to adjust their ocular retention and interaction with target cellular receptor. Moreover, biocompatible elastin-like polypeptides (ELPs) appear to provide a reversible, temperature dependent bridge between potential ocular therapeutics and other polymers used on the ocular surface.
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