Design of multimodal microgel populations for controlled protein delivery: Investigations of light and redox responsive formulations
Date
2020
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Publisher
University of Delaware
Abstract
Biologics are playing an increasingly important role in the treatment of a variety of diseases ranging from autoimmune diseases to cancers due to their high specificity and potency compared to their small molecule counterparts. These therapies represent the most advanced treatments available and as such the number of FDA approved biologics continues to grow. Despite their growth in popularity within the medical field, there are still large-scale challenges associated with protein-based therapies including stringent storage conditions, limited delivery options, and costly therapeutic regimens. On top of this, proteins face therapeutic barriers \textit{in vivo} that are inherit to the protein structure. For example, large proteins have difficulty penetrating tissues limiting delivery options and altering biodistribution, whereas small proteins suffer from short half-lives limiting therapeutic time scales. These limitations have led to the need for large doses to be administered for therapeutic efficacy to be realized, which increases the probability of adverse side effects and costs experienced by the patient. In an attempt to address these challenges, protein delivery vehicles have been engineered to provide controlled, tunable, and localized delivery motifs. ☐ Hydrogels have emerged as promising platforms for protein delivery as they can encapsulate large amounts of hydrophilic proteins and exhibit versatility in structure, size, and chemical nature. Further, hydrogel microparticles, or microgels, offer exciting opportunities for creating modular protein delivery platforms toward improved combination therapy regimens and personalized medicine. The overarching goal of this thesis was to engineer multimodal redox and light responsive microgel populations to provide controlled and on-demand protein release profiles through tunable matrix degradation properties. This goal was achieved by addressing limitations in literature associated with \textit{i)} the use of photolabile chemistries within hydrogel networks developed for controlled protein delivery, \textit{ii)} the formation of microgels using spontaneous click chemistries in combination with microfluidic flow focusing techniques, and \textit{iii)} the design of cyclic peptides with functional handles for bioconjugation. ☐ First, the light responsive behavior of poly(ethylene glycol) (PEG) based hydrogels containing different photolabile moieties within the network crosslinks were investigated. Nitrobenzyl (NB) moieties, which degrade in response to long wavelength UV, with different labile bonds were synthesized and their photolytic and hydrolytic behavior as crosslinks within hydrogels was characterized. Four different cleavable bonds, NB-ester, NB-amide, NB-carbonate, and NB-carbamate were investigated, in which the latter two had yet to be incorporated as crosslinks within hydrogels. The four different NB linkers were observed to exhibit a range of photolytic and hydrolytic properties which were utilized to control the release of model proteins in a sequential or combined manner from concentric circle hydrogels. Second, the NB-carbamate was compared to a coumarin photolabile moiety, which degrades in response to blue light, in both degradation and protein release properties. It was observed that the hydrogels containing either NB or CMR moieties could degrade rapidly in response to long wavelength UV or blue light, respectively. Further, both formulations were successfully degraded through a layer of porcine skin, a mimic for human skin, and able to effectively encapsulation and release a model bioactive antibody. These demonstrations highlight the importance in labile bond chemistry of photocleavable moieties for controlling degradation behavior and the potential utility of photolabile hydrogels as protein delivery vehicles. ☐ Next, homogenous microgels that responded to either reducing conditions or light were formed using microfluidic flow focusing. Redox responsive microgels were successfully formed using a thiol-Michael addition between thiols and maleimides by controlling the rate of hydrogel formation. Further, NB-carbamate containing microgels of the same size were formed through a strain promoted azide alkyne cycloaddition reaction indicating microgel size is controlled by the microfluidic device design and not the polymerization chemistry. The redox responsive microgels were successfully degraded on clinically relevant time scales by incubation in high reducing, low reducing, or aqueous conditions. Light responsive microgels were degraded through irradiation with long wavelength UV light. Together, these microgels provide both sustained and on-demand degradation profiles and future work will focus on combining these systems to create complex release profiles of multiple proteins. ☐ Lastly, functional cyclic RGD peptides were synthesized to contain a functional handle for subsequent conjugations using orthogonal click reactions. The cyclic RGD was found to localize to breast cancer cell membranes indicating its potential use as a targeting peptide by conjugation to protein therapeutics or delivery vehicles. Collectively, this thesis provides insights into the rational design of photolabile hydrogels for the on-demand delivery of protein therapeutics, offers design parameters for forming redox and light responsive microgels through spontaneous click chemistries, and presents a synthetic strategy for synthesizing cyclic peptides containing functional handles. These strategies can be combined to created targeted multimodal microgel populations for the sustained and on-demand release of protein therapeutics toward personalized medicine or improved therapeutic or vaccination regimens.
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Keywords
Biomaterials, Photodegradation, Protein Delivery