| This dissertation focused on development and in vitro/in vivo evaluation of a novel PLGA and starch-based hydrogel composite microsphere delivery system for sustained release of proteins. PLGA acts as an effective barrier and controls biodegradation, thereby providing sustained release characteristics. The hydrogel's propensity for water uptake and its hydrophilic nature offered a friendly microenvironment for proteins.; The composite microspheres were prepared by soaking protein into blank acryloyl hydroxyethyl starch (AcHES) hydrogel microparticles followed by encapsulation into the PLGA matrix. Bovine serum albumin, a common model protein, was selected to develop the fabrication process and scanning electron microscopy showed a PLGA matrix containing many AcHES particles. AcHES 5–10% and 5–10% BSA target load provided high drug encapsulation efficiency. The composite exhibited a favorable profile with a reduced initial burst, higher incremental and more complete release than that from PLGA alone. Enzymatically active proteins, maltase and horseradish peroxidase retained over 70% specific activity after encapsulation, suggesting that sensitive proteins could survive the composite manufacturing process.; To further characterize the composite microsphere system, lysozyme, similar in size to many commercial biopharmaceuticals, was selected to study the in vitro-in vivo correlation (IVIVC). Lysozyme displayed high conformational stability and lack of adsorption to PLGA in acetate and glycine buffers, whereas in PBS, the protein conformational stability was low with a trend toward aggregation and significant protein adsorption was evident. The adsorption resulted in slow and incomplete release in PBS while the release in acetate and glycine buffers was complete within 40 and 70 days, respectively. The in vivo profile qualitatively correlated with in vitro release in glycine buffer, suggesting that protein stability and adsorption are critical factors controlling protein release kinetics and optimization of IVIVC.; Finally, the composite system was adapted to a therapeutic protein, insulin. Insulin integrity was confirmed by SDS-PAGE and MALDI-TOF Mass Spectrometry. An extraction and HPLC analytical method was developed to determine insulin loading. Microspheres with low initial burst batches were achieved by sonication and they exhibited a sustained in vitro release pattern over 28 days. Glucose suppression correlated well with serum insulin levels and animal growth for ten days after single dose microsphere treatment. Multiple dosing based on glucose titration showed sustained suppression of glucose as well as repetitive efficacy and blood insulin levels from each dose. These results indicate that the composite microsphere system can be costumed for a target therapeutic protein and can be engineered to meet desired therapeutic needs. |
