Chemical and Biomolecular Engineering - Theses
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ItemCapsosomes: en route toward synthetic cellular systems( 2011)Engineering artificial cells is currently an emerging area of research that involves constructing mimics of biological cells. These biomimetic cellular structures hold tremendous promise for the creation of next-generation therapeutic tools due to their ability to restore lost cellular functions. Amongst their potential applications, replenishing absent or malfunctioning enzymatic activities to degrade waste products or to support the synthesis of medically relevant biomolecules is a chief goal, which can provide long-term therapeutic solutions for chronic diseases. Artificial cells do not require the complex multifunctionality of their biological counterparts and can be more simply designed to perform a specific activity. A key approach in designing a cell-like system is a subcompartmentalized assembly, which is one of the features of biological cells that enable the performance of multiple complex biochemical reactions within confined environments. This thesis focuses on developing a bottom-up approach to assemble micron-sized vessels with a controlled number of enzyme-loaded subcompartments toward cell mimicry. Capsosomes, polymer hydrogel capsules containing controlled amounts of intact cargo-loaded liposomal subcompartments, were developed in this thesis and they represent a novel class of carrier system toward the design of bioinspired vehicles. Polymer capsules, assembled via the sequential deposition of interacting polymers onto particle templates (layer-by-layer technique, LbL) followed by core removal, serve as structurally stable scaffolds with tunable permeability that allow exchange of reagents and nutrients between the internal and external milieu – resembling cell membranes. On the other hand, liposomes divide the interior of the capsules into subcompartments and can stably encapsulate fragile hydrophobic and hydrophilic cargo, e.g., enzymes in order to conduct encapsulated catalysis – resembling cell organelles. The creation of (bio)degradable capsosomes is based on the sequential assembly of cargo-loaded liposomes and polymers onto sacrificial particle templates, followed by the LbL deposition of poly(N-vinylpyrrolidone) (PVP) and thiol-functionalized poly(methacrylic acid) (PMASH) via hydrogen bonding. Upon crosslinking the thiols of the PMASH and dissolution of the particle templates, colloidally stable capsosomes are obtained. The coassembly of polymers and liposomes was optimized via novel noncovalent linkage concept using tailor-made cholesterol-modified polymers and this unique approach facilitates stable incorporation of intact liposomes into polymer films. Spatial position of the subcompartments can be controlled, which yields capsosomes containing membrane-associated or “free-floating” subunits. Capsosomes exhibit size-dependent retention of the encapsulated cargo within the liposomal subunits. To prolong the stability of the liposomes in the compartmentalized assembly against degradative enzymes, the outer membrane of the capsosomes was surface functionalized with poly (ethylene glycol) (PEG). The functionality of capsosomes was demonstrated by triggered encapsulated (two-step) enzymatic catalysis. Capsosomes encapsulating glutathione reductase were able to generate glutathione, a potent antioxidant, while simultaneously releasing small therapeutic molecules, which highlights the ability of this subcompartmentalized assembly in addressing the complexity in therapeutic cell mimicry. The phase transition temperature of the liposomes was used as a trigger to initiate the enzymatic reactions, allowing capsosomes to be repeatedly used for multiple subsequent catalysis. Capsosomes with tailored properties present new opportunities en route to the development of functional cell mimics and the presented studies highlight crucial aspects for the successful applications of capsosomes as therapeutic artificial cells.