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Bacteria-based cancer therapy (BBCT) holds immense promise in addressing the limitations in treatment of solid tumors. Bacterial strains used for BBCT are engineered to express therapeutics, facilitate precise navigation within the tumor microenvironment by enhancing bacteria's motility, chemotaxis (movement toward or away from specific chemicals), or other mechanisms that aid in reaching and infiltrating the tumor tissue effectively, and complementing traditional chemotherapy and immunotherapies while minimizing side effects. Bacterial motility not only influences the ability of bacteria to navigate within the tumor but also plays a pivotal role in optimizing drug delivery, treatment efficacy, and minimizing potential obstacles associated with the complex microenvironment of human tissues. However, the current understanding of bacterial motility remains limited. In this thesis, we use a reductionist approach and study bacteria motile behavior within human tissue phantoms (collagen and agar) and the bacteria-hydrogel interactions. Apart from motility, it is important to analyze the mechanical properties of the hydrogels the bacteria interact with as they play a vital role in overall behavior and physics of bacteria movement. To that extent, there exists a gap in our understanding of the viscoelastic properties of hydrogels. Lastly, systematic and comprehensive investigation of bacteria behavior in hydrogels requires tracking of thousands of individual cells. Thus, there is an unmet need to develop new automated techniques to reduce the labor-intensive manual tracking of bacteria in low-contrast hydrogel environments, with feature sizes comparable to that of bacteria. To address these gaps, this thesis proposes a trident approach towards mechanistic understanding of bacteria motility in time-invariant agar and temporally evolving collagen hydrogels to bridge critical gaps in understanding bacterial motile behavior in these media, non-destructive microelastography-based mechanical characterization of hydrogels with less than 4.7% error compared with rheology, and the development of deep learning-enabled automated bacteria tracking tools with 77% precision. / Master of Science / There exists a huge scope for improvement in cancer therapies. The gold standard chemotherapy and immunotherapies are responsible for a lot of side effects. Over a century ago, bacteria-based cancer therapy started to develop and over the period, it was discovered that they can be effective when used with traditional therapies improving precision and reducing side effects. The motility of bacteria is shown to improve bacterial distribution in solid tumors. However, the physical underpinnings of bacteria motility in the tumor environment remains understudied. This thesis proposes a trident approach, investigating bacteria motility in tissue-like environments (hydrogels), characterizing the mechanical properties of hydrogels using acoustic waves to capture bacteria-induced alterations, and developing deep-learning-enabled automated bacterial tracking approach for high throughput analysis of experimental data. We report bacteria behavior and motility patterns in hydrogels, the mechanics of these hydrogels with less than 4.7% error compared with standard characterization methods, and automated bacteria tracking with 77% precision to inform the development and advancement of bacteria-based drug delivery systems. In summary, these tools can help improve our understanding of bacteria-hydrogel interactions, allowing us to develop innovative bacteria-based cancer therapies in the long term.

<p>The mechanical and chemical properties of hydrogels make them excellent vehicles to deliver drugs. However, current systems encounter difficulties with loading hydrophobic molecules into the aqueous gel network and the subsequent release of the drug from the gel matrix. Cyclodextrins (CDs) offer a potential solution to this drug delivery challenge. CDs have the unique property of possessing a hydrophilic exterior and a hydrophobic interior pocket which is capable of hydrophobic drug binding. CD molecules complexed with hydrophobic drugs have been demonstrated to significantly increase the bioavailability of those drugs in free solution. Thus, if these nanodomains are introduced into microgels or hydrogels, we anticipate that significantly higher hydrophobic drug loadings may be achieved together with improved controlled release of these drugs based on the properties of the hydrogel or microgel phase. We have fabricated <em>in situ</em> gellable and degradable hydrogels and microgels based on combinations of CDs and either functionalized carbohydrates (dextran) or thermosensitive synthetic polymers (poly(N-isopropylacrylamide), PNIPAM). To achieve this goal, we designed a series of microgels with grafted or immobilized CD groups and used multi-functional CD as a reactive crosslinker for making injectable bulk hydrogels.</p> / Master of Applied Science (MASc)

<p>Coated calcium-alginate beads are the basis of many encapsulation methods used in pursuit of cell-based enzyme and hormone replacement therapies. The standard alginate - poly-L-lysine - alginate (APA) capsules consist of a calcium-alginate hydrogel core containing cells designed to express a therapeutic product, coated with permeability controlling poly-L-lysine (PLL, a polycation) followed by an exterior layer of polyanionic alginate. Although this approach is promising, the required long-term survival of the implanted cells has remained largely elusive as the current APA capsules suffer from several biocompatibility and mechanical strength issues, one of which is the weakening of ionic crosslinks over time, exposing the encapsulated cells to the host.</p> <p>This thesis aims to replace the exterior layer of alginate with a Temporarily Reactive Polyelectrolyte (TPR) to reinforce AP capsules by forming covalently crosslinked shells. TRPs are polyanions that possess reactive electrophilic groups capable of forming permanent covalent crosslinks with the underlying polyamine (such as PLL), and subsequently hydrolyze, increasing the net negative charge of the polyanion. TRPs are thought to improve the biocompatibility and strength of the microcapsules by forming stable inert amide bonds, as well as increasing the net negative charge of the capsule through the liberation of carboxylates. This thesis will focus primarily on two TRPs: 50% hydrolyzed poly(methyl vinyl ether-<em>alt</em>-maleic anhydride), PMM₅₀ , and poly(methacrylic acid-co-2-vinyl-4,4-dimethylazlactone) with a 50:50 co-monomer ratio, PMV₅₀ . Their synthesis, rates of hydrolysis and capsule formation around encapsulated C2C12 cells for <em>in-vitro</em> and<em> in-vivo</em> studies will be described. Additionally the synthesis and rates of hydrolysis of other 2-vinyl-4,4-dimethylazlactone (VDMA)-copolymers are presented as potential candidates for future TRPs.</p> / Doctor of Philosophy (PhD)

<p>Endocrine disrupting substances have been frequently reported to exist in potent concentrations in wastewater treatment plant effluent and other surface waters. Common techniques of wastewater treatment have varied effectiveness to remove estrogens from wastewater. A thermo-responsive smart membrane technology is investigated for its use in adsorptive removal of 17β-estradiol from a background electrolyte solution. A simplified fabrication method is adapted for hydrogel-substrate composite thermo-responsive membranes. Deposition of hydrogel occurs through aqueous polymerization in a coating process dissimilar to common grafting techniques. Acrylamide and acrylic acid monomers are polymerized in two different structures, a random copolymer as well as an interpenetrating network, to form a positive volume-phase transition hydrogel coating. Subsequent membranes experience high permeability at low temperatures with a gating mechanism reducing permeability upon heating. The effects of crosslinker content, monomer ratio, mass loading and butylmethacrylate content are investigate. Only mass loading was found to have significant influence on the behaviour of the membranes in all cases. The variations of the other factors were too little to have great influence. The membranes with the most stable permeability response function were then used in 17β-estradiol adsorption tests, investigating the binding capacity at both colder water temperatures (10oC) and warmer water temperatures (40oC). In the collapse and swelling of the volume-phase transitions, the membranes changed their solution properties which were hypothesized to also alter surface functionality. After introducing the estradiol sample, the membranes were subjected to temperature change with the expectation that any bound material would elute once the surface functionality of the membranes became adequately altered. Only some membranes produced an elution fraction while others appeared to undergo irreversible binding with a possible delayed elution. Removal of dosed 17β-estradiol is reported as adsorbed mass per area of membrane.</p> / Master of Applied Science (MASc)

<p>Despite the long history of topical eye drops and their use in delivering therapeutic agents to the anterior of the eye, efficient sustained delivery continues to be an elusive goal. The robust and effective clearance mechanisms that the eye is endowed with are significant delivery challenges and result in short drug residence times and low ocular bioavailability. The work carried out in this thesis focused on developing, synthesizing and characterizing silicone hydrogels and evaluating their potential as drug eluting inserts for more effective delivery of ocular pharmaceuticals. The first strategy (Chapter 2) focused on incorporating a novel hydrogel additive, hyaluronic acid, to promote hydrogel-drug ionic interactions that can function to increase drug loading and subsequent release dosage. Hydrogels composed of a hydrophilic monomer, N,N-dimethlacrylamide (DMA) or 2-hydroxyethyl methacrylate (HEMA), and a hydrophobic monomer, methacryloxypropyltris(trimethylsiloxy)silane (TRIS), were used as model contact lenses. By combining ionic interactions with molecular imprinting techniques within a single hydrogel, it was shown that this can produce a compound effect on drug uptake and release. Although greater control over release dosage was achieved, there was limited capacity for these materials to delivery timolol for extended periods with drug release occurring rapidly over a period of 1-2 days. However, there were clear differences in the release duration from the p(DMA-<em>co</em>-TRIS) and p(HEMA-<em>co</em>-TRIS) hydrogel formulations. Therefore, the second study (Chapter 3) aimed to better understand the relationship between the hydrogel chemical composition and the resultant material properties on the drug release characteristics. A range of hydrogels were synthesized with varying hydrophilic and hydrophobic monomers, which were then characterized by their water content, transparency, optical haze and surface wettability. The previous generation materials were evolved by incorporating a modified siloxy methacrylate TRIS(OH), a methacrylated polydimethylsiloxane macromonomer (mPDMS) and a polymerizable silicone surfactant (ACR). The properties of the hydrogels were dramatically affected by the nature and relative contribution of hydrophobic and hydrophilic monomers. The release of dexamethasone (DEX), an anti-inflammatory medication, was shown to vary significantly depending on the hydrogel formulations; often displaying faster release in high water content materials and slow release in low water content hydrogels. The mechanism of diffusion for lipophilic DEX in these hydrogel systems appeared to be through the internal aqueous network channels within the bulk. Over the range of hydrogels formulations that were tested, the release from them varied from approximately seven days to greater than two weeks.</p> / Master of Applied Science (MASc)

Osteoarthritis and cartilage deterioration are favored by meniscetomy, which is the ablation of the meniscus from the knee joint. Meniscectomy can be partial or total. This procedure is performed when meniscus lesions and tears or the degeneration of the meniscus caused by its natural dehydration occur. There is a peak of meniscal lesions observed between 20 and 29 years old. Alternative methods such as sutures fail in that they present a short term solution which is ideal for a less active, older generation. A long term solution is needed for a younger population to reduce the number of surgical procedures over the lifetime of this active group. There is a crucial need for a functional implant designed in the image of the native meniscus. Blends of poly (vinyl alcohol) PVA and poly (vinyl pyrrolidone) (PVP) present a potential solution. PVA has shown similar characteristics to soft tissues. PVP further stabilizes the hydrogel network. This work is the mechanical characterization of PVA/PVP (99:1) hydrogels under physiological conditions. Equilibrium swelling in a medium replicating the ionic and the osmotic content of the synovial fluid was investigated during 35 days. The mass retention of hydrogels was characterized using data obtained from the swelling study and was examined as a function of the cross link density and the polymer content. The modulus of hydrogels was obtained in unconfined compression, first at a strain rate slow enough to ignore fluid flow in and out of the gels, and subsequently at a physiological strain rate of walking. Results indicate that PVA/PVP hydrogels volume swelling ratio and weight swelling ratio show no significant difference for most formulations by the 14th day of immersion. A few hydrogels would reach equilibrium by day 21. Additionally, percentage polymer mass retention increases with the cross link density. However, there is no consistent trend with the polymer content. All formulations with 10% wt of polymer show the highest mass retention while 15% wt show the lowest. Interestingly, the mechanical characterization of hydrogels at 100%/min strain rate shows that 15% wt is the only formulation whose compressive modulus falls within the targeted range whereas 10% wt proves to not be stiff enough. 20% wt and 25% wt are always too stiff. Results obtained from unconfined compression at the physiological strain rate, that is 1920%/min, are rather inconclusive. There is not enough consistency in the literature to narrow the results down to one successful candidate formulation. The modulus range obtained at physiological strain rate encompasses the range obtained at 100%/min strain rate. The highest modulus value obtained is 10 times higher at physiological strain rate than the modulus of a real human meniscus obtained at 100%/min strain rate. It is not reasonable at this time to make a choice of a formulation at physiological strain rate due to high variability of the modulus of a human meniscus as a result of its intrinsic anisotropy. All formulations tested would be considered successful candidates, which is irrational considering the difference in their stiffness. / Mechanical Engineering

Within the last decade, neurodegenerative diseases such as Alzheimer’s and Parkinson’s have emerged as one of the top 5 leading causes of death globally, and there is currently no cure. All neurodegenerative diseases lead to loss of the functional cells in the nervous system, the neurons. One therapeutic approach is to replace the damaged and lost neurons with new, healthy neurons. Unfortunately, this is a difficult endeavor since mature neurons are not capable of cell division. Instead, researchers are turning to neural stem cells, which are able to self-renew and be rapidly expanded before being differentiated into functional cell phenotypes, such as neurons, allowing for large numbers of cells to be generated in vitro. Controlled differentiation of human neural stem cells into new neurons has been of interest due to the immense potential for improving clinical outcomes. Adult neural stem cell behavior, however, is not well understood and the transplanted stem cells are at risk for tumorigenesis. The focus of this dissertation is the development of engineered biomaterials as tools to study human neural stem cell behavior and neurogenesis (differentiation). A novel cell penetrating peptide was developed to enhance intracellular delivery of retinoic acid, a bioactive lipid known to induce differentiation. A hydrogel platform fabricated from hyaluronic acid, a naturally-occurring polysaccharide found in brain extracellular space, was designed to serve as a biomimetic soft substrate with similar mechanical properties to the brain. The biological behavior of the stem cells was characterized in response to chemical and physical cues. / Bioengineering

When a biomaterial is inserted into the body, the interaction of the surface with the surrounding biological environment is crucial. Given the importance of the surface, the ability to alter the surface properties to support a compatible environment is therefore desirable. Silicone hydrogel contact lenses (CL) allow for improved oxygen permeability through the incorporation of siloxane functional groups. These groups however are extremely surface active and upon rotation, can impart hydrophobicity to the lens surface, decreasing lens wettability and increasing protein and lipid deposition. Lens biofouling may be problematic and therefore surface modification of these materials to increase compatibility is exceedingly recognized for importance in both industry and research. The current work focuses on the creation of a novel anti-fouling polymer surface by the incorporation of 2-methacryoyloxyethyl phosphorylcholine (MPC), well known for its biomimetic and anti-fouling properties. A controlled polymerization method was used to generate a unique double-grafted architecture to explore the effect of increasing surface density of polyMPC chains on corresponding anti-fouling properties. The novel free polymer was synthesized by a 3-step atom transfer radical polymerization (ATRP). First, poly(2-hydroxyethyl methacrylate) (polyHEMA) was polymerized by ATRP, where the hydroxyl (OH) groups of the polymer then underwent an esterification to create macroinitiating sites. From these sites, a second ATRP of poly(MPC) varying in length occurred, yielding the double-grafted polymer poly(2(2-bromoisobutyryloxy-ethyl methacrylate)-graft-poly(2-methacryloyloxyethyl phosphorylcholine (pBIBEM-g-pMPC). The polymer was designed for resistance to protein adsorption through a possible synergistic effect between the surface induced hydration layer by surrounding PC groups coupled with steric repulsion of the densely grafted chains. To test its potential as a surface modifier, the polymer was grafted from model silicone hydrogel CL through a 4-step surface initiated ATRP (SI-ATRP) in a similar manner to the free polymer. First, the ATRP initiator was immobilized from the HEMA OH groups of the unmodified CL, generating Intermedate-1. A polyHEMA brush was grafted from the initiating sites yielding pHEMA-50, followed by the generation of a second initiator layer (Intermediate- 2). A sequential ATRP of poly(MPC) then generated the target pMPC-50/pMPC-100 surfaces. For the free pBIBEM-g-pMPC polymer analysis, 1H-NMR and GPC determined polymers formed with a predictable MW and low polydispersity (PDI). For surface grafting, using a sacrificial initiator, 1H-NMR and GPC indicated that the pHEMA-50 and pMPC-50/pMPC-100 polymers were well-controlled, with a MW close to the theoretical and a low PDI. For surface chemical composition, ATR-FTIR showed the presence of the ATRP initiator (Intermediate-1 and 2) by the appearance of a C-Br peak and disappearance of the OH peak. XPS confirmed the chemical composition of the 4-step synthesis by a change in the fraction of expected surface elements. Both the surface wettability and EWC of the materials increased upon pMPC modification, further improving upon increasing pMPC chain length. The contact angle was as low as 16.04 ± 2.37º for pMPC-50 surfaces and complete wetting for pMPC-100. Finally, the single protein adsorption using lysozyme and bovine serum albumin (BSA) showed significantly decreased protein levels for pMPC-50/100 lenses, as much as 83% (p 0.00036) for lysozyme and 73% (p 0.0076) for BSA, with no significant difference upon chain length variation. The aforementioned data demonstrates that the novel polymer has potential in providing an anti-fouling and extremely wettable surface, specifically regarding silicone hydrogel CL surfaces. / Thesis / Master of Applied Science (MASc)