Stability of therapeutic proteins in hydrogel drug delivery systems

Protein pharmaceuticals have gained prominence as vital drug products, but their development is complex due to stability issues during the manufacturing process, potentially compromising product efficacy and safety. This thesis aimed to develop a protein delivery system and optimize it to withstand protein instability during the manufacturing process and provide storage stability for the encapsulated protein. Hydrogel microparticles prepared through an aqueous two-phase system (ATPS) were chosen as a promising protein delivery platform due to their hydrophilic nature and ability to protect proteins. Visible light photocrosslinking was used to fabricate these hydrogel microparticles, preventing potential photodegradation that could compromise protein safety and efficacy. The test protein 3β/17β-Hydroxysteroid dehydrogenase (17β-HSD) was used to study and optimize stability during encapsulation. The first chapter focuses on developing and optimizing bulk hydrogels as the basis for hydrogel microparticles. Hydroxyethyl starch (HES) was used as a biocompatible polymer, and a visible light photoinitiator system was employed for crosslinking. The optimized hydrogel formulation demonstrated excellent mechanical properties, biocompatibility, and biodegradability. However, further optimization could enhance protein release for sustained delivery. In the second chapter, hydrogel microparticles were synthesized through an ATPS based on the optimized hydrogel formulation. The particle size was adjusted to facilitate uptake by synovial macrophages for intra-articular injection in treating rheumatoid arthritis. The formulation with the optimal particle size showed favorable physicochemical properties, biocompatibility, and controlled protein release. Adjusting crosslinking density could further control the release kinetics. The third chapter investigated and optimized each formulation step's influence on the stability of the test protein 17β-HSD. The appropriate buffer and visible light photocrosslinking were identified to preserve protein activity. Incorporating HES-MA and trehalose into the final formulation significantly enhanced protein stability which was reflected in the activity of the released protein. In conclusion, hydrogel microparticles prepared via visible light photocrosslinking in ATPS showed potential as a protein delivery platform with preserved stability. Further research can focus on optimizing crosslinking density and exploring other applications for this platform. This work contributes valuable insights for developing commercially viable protein products with enhanced stability and safety.