LONG-TERM GDNF DELIVERY FROM PLGA MICROSPHERES AFTER SINGLE INTRAVITREAL INJECTION IN RABBITS

Abstract

Purpose: Local long term delivery of the Glial cell line Derived Neurotrophic Factor (GDNF) from a novel formulation of VitE-PLGA microspheres (MPs) has demonstrated to protect the retinal ganglion cells (RGCs) in an animal model of glaucoma (unilateral IOP elevation after episcleral injection of a hypertonic solution in rats) for at least three months. However, the pharmacokinetic of the neurotrophic factor after intravitreal (IVT) injection of the biodegradable particles has not been studied. The aim of this work was to evaluate the GDNF levels after a single intravitreal administration of GDNF/VitE-PLGA MPs.

Methods: GDNF was purchased from R&D (Minneapolis, MN USA) and PLGA 50:50 (Resomer® 503) from EVONIK (Darmstadt, Germany). Adult female New Zealand White Rabbits (2-3Kg, SAEA, Zaragoza University, Spain) were used for the in-vivo studies (n=30). Animal experiments were performed in compliance with the ARVO Statement for the Use of Animals in Ophthalmic Vision Research. GDNF- MPs were prepared by the S/O/W emulsion-solvent evaporation technique and characterized in terms of morphology, particle size, GDNF loading and in vitro release studies. Animals received a single IVT injection (50μL) of GDNF/ VitE-PLGA -MPs (4%w/v) (24 eyes), unloaded VitE-PLGA MPs (4%w/v) (24 eyes), GDNF solution (50ng/mL) (3 eyes) and Balanced Salt solution (BSS) (7 eyes). At pre-set times (24 hours,1,4,6,12 and 24 weeks), the eyes were enucleated and the IVT levels of GDNF were quantified (ELISA).

Results: GDNF loaded microspheres (74.74ng/mg MPs) ranged from 20-40μm in size and were spherical with porous surfaces. The neurotrophic factor was released in-vitro in a controlled fashion up to 24 weeks. Due to the eye response after injection, non loaded PLGA-MPs caused an initial increase of the basal levels of GDNF (10.24 ± 4.03 pg/mL ) compared to non injected eyes (3.69 ± 1.60 pg/mL). The highest intraocular GDNF levels were observed at 24 hours (763.08 ± 113.92 pg/mL) and 4 weeks (726.87 ± 66.42 pg/mL) in eyes receiving GDNF/vitE-PLGA MPs. The concentration of the neurotrophic factor determined at six months post-injection of GDNF loaded MPs (20.36 ± 7.60 pg/mL) resulted significantly higher than the basal GDNF levels.

Conclusions: Single injection of the novel formulation of GDNF/vitE- PLGA microspheres provided a sustained controlled release of the neurotrophic factor during long term (at least 6 months).