|dc.description.abstract||Delivery of therapeutics to the brain through non-invasive administration is a difficult task due to the blood-brain barrier (BBB), which prevents the transport of 98% of therapeutics. In GM1 gangliosidosis, patients are missing β-galactosidase (βgal), an enzyme necessary for cellular digestion, with major central nervous system (CNS) manifestation. GM1 gangliosidosis is fatal in infancy with no clinically available treatment. We have designed and characterized the first nanoparticle-mediated treatment of GM1 gangliosidosis using self-assembled polymersomes for IV enzyme delivery. When coupled with apolipoprotein, delivery through the BBB and to the lysosome of neural cells will occur, treating patients without invasive surgery.
Poly(ethylene glycol)-b-Poly(lactic acid) (PEG-b-PLA) polymersome formed via passive mixing in water at average polymersome diameters of 237.2 ± 66.5 nm over 150 minutes. Empty polymersomes increased to 4.63 ± 0.01 times their size after lyophilization, showing lack of long-term stability. The use of lyoprotectants, mannitol and inulin, to maintain particle size distribution (PSD) was studied. The incorporation of both molecules was confirmed. Differences in moisture content were found after lyophilization between samples incorporating inulin and mannitol. It was hypothesized that lyoprotectants replaced water, maintaining polymersome structure under stressful processing conditions. The ability to reconstitute polymersome drug delivery carriers without altering size distribution is paramount to the creation of effective and efficient drug delivery systems.
PEG-b-PLA polymersomes also formed via solvent injection with 2 wt%/v mannitol at an average diameter of 145 ± 21 nm. PEG-b-PLA polymersomes encapsulate βgal at 72.0 ± 12.2% efficiency and demonstrate simultaneous encapsulation and ligand attachment at 86.7 ± 11.6% efficiency. Amine-reactive PEG facilitated the attachment of CF 350 Amine, a blue fluorescent ligand, for fluorescent imaging, and apolipoprotein E (ApoE), a target to the low density lipoprotein family of receptors, for BBB delivery, to the polymersome surface. In vitro, PEG-b-PLA polymersomes demonstrate limited release in physiologic environment, pH 7.4, with a burst release upon membrane poration in lysosomal environment, pH 4.8. Cellular studies, using GM1 gangliosidosis-diseased fibroblasts, confirm that βgal-loaded polymersomes increase enzyme activity to normal levels with doses as low as 0.7 mg/cm2 and the use of an ApoE-tag increases enzyme activity to normal levels with doses as low as 0.175 mg/cm2. Results are promising towards the goal of creating the first clinical treatment for GM1 gangliosidosis, using a combination of enzyme replacement therapy and nanotechnology methods to cross the BBB.||en_US