Burgeoning fields of nanomedicine and theranostics are propelled forward by the creative and systematic design of synthetic polymers. Cationic polyelectrolytes, comprising covalently-linked cations within each repeat unit, have drawn particular interest for their ability to bind nucleic acids and permeate cell membranes. Expanding the design space of these systems, we introduced a new family of polyelectrolytes based on the carbon-centered cyclopropenium cation. Cyclopropenium is a modular, aromatic building block with unique structural and electronic properties and, when coupled with modern living polymerization techniques, can be incorporated into macromolecules with precise size, shape, and composition. This thesis describes the translation of cationic polyelectrolytes based on cyclopropenium to biomedical applications and is structured into three parts. The first part evaluates cyclopropenium polymers as candidate non- viral vectors for gene therapy and demonstrates that some derivatives are both biocompatible and efficacious transfection agents. In the second part, nanoparticles comprising cyclopropenium are exploited as live-cell image contrast agents and evolved into potentially theranostic tools. The final part describes a facile route to a novel class of cyclopropenium-based polymers. Together, this thesis illustrates that cyclopropenium is a versatile component of polyelectrolytes, poised to address leading biological challenges.

No description available.

Nanomedicine, the medical application of nanotechnology, uses nanoscale objects that exist at the interface between small molecule and the macroscopic world for medical diagnosis and treatment. One of the healthcare applications of nanomedicine is drug delivery: the development of nanoscale objects to improve therapeutics' bioavailability and pharmacokinetics. Shell crosslinked knedel-like nanoparticles (SCKs), that are self assembled from amphiphilic block copolymers into polymeric micelles and then further stabilized with crosslinkers isolated throughout the peripheral shell layer, have been investigated for drug delivery applications that take advantage of their core-shell morphology and tunable surface chemistry. SCKs are attractive nanocarriers because the cores of the SCKs are used for sequestering and protecting guests. The readily adjustable shell crosslinking density allows for gating of the guest transport into and out of the core domain, while retaining the structural integrity of the SCKs. Moreover, the highly functionalizable shell surface provides opportunity for incorporation of targeting ligands for enhanced therapeutic delivery. The optimization of nanoparticle size, surface chemistry, composition, structure, and morphology has been pursued towards maximization of the SCKs' therapeutic efficacy. With distinctively different dimensions, compositions and structures of the core and shell domains of SCKs, and an ability to modify each independently, probing the effects of each is one of the major foci of this dissertation. Utilization of a living radical polymerization technique, reversible addition-fragmentation chain transfer (RAFT) polymerization, has allowed for facile manipulation of the block lengths of the polymer precursors and thus resulted in various dimensions of the nanoparticles. SCKs constructed from poly(acrylic acid)-b-polystyrene (PAA-b-PS) with various chain lengths, have been investigated on the loading and release of doxorubicin (DOX). The effect of PEGylation on paclitaxel (PTX) loaded SCKs on the cell internalization and killing was investigated. Apart from chemotherapies, the SCKs were explored as antimicrobial agents by incorporating silver species. Conjugation of the SCK surface with a protein adhesin through amidation chemistry to promote epithelial cell targeting and internalization was developed. Nanoscale assemblies with complex morphologies constructed from a linear triblock copolymer was investigated. Furthermore, a highly multifunctional nanodevice for imaging and drug delivery functionalized with a chelator for radio-labeling, polyethylene glycol (PEG) for improved biodistribution, targeting ligands, a chromophore and a therapeutic agent was evaluated in vivo as active-targeted delivery of therapeutics.

Nanomedicine is a promising and revolutionary field to improve medical diagnoses and therapies leading to a higher quality of life for everybody. Huge benefits are expected from nanomedicine applications such as in diagnostic and therapeutic field. However, nanomedicine poses several issues on risks to the human health. This thesis aims to defense a perspective of risk governance that sustains scientific knowledge process by developing guidelines and providing the minimum safety standards acceptable to protect the human health. Although nanomedicine is in an early stage of its discovery, some cautious measures are required to provide regulatory mechanisms able to response to the unique set of challenges associated to nanomedicine. Nanotechnology offers an unique opportunity to intensify a major interplay between different disciplines such as science and law. This multidisciplinary approach can positively contributes to find reliable regulatory choices and responsive normative tools in dealing with challenges of novel technologies.

Nanomedicine is a promising and revolutionary field to improve medical diagnoses and therapies leading to a higher quality of life for everybody. Huge benefits are expected from nanomedicine applications such as in diagnostic and therapeutic field. However, nanomedicine poses several issues on risks to the human health. This thesis aims to defense a perspective of risk governance that sustains scientific knowledge process by developing guidelines and providing the minimum safety standards acceptable to protect the human health. Although nanomedicine is in an early stage of its discovery, some cautious measures are required to provide regulatory mechanisms able to response to the unique set of challenges associated to nanomedicine. Nanotechnology offers an unique opportunity to intensify a major interplay between different disciplines such as science and law. This multidisciplinary approach can positively contributes to find reliable regulatory choices and responsive normative tools in dealing with challenges of novel technologies.

A growing demand for information on the human health and environmental effects of materials produced using nanotechnology has led to a new area of investigation known as nanotoxicology. Research in this field has widespread implications in facilitating the medical applications of nanomaterials but also in addressing occupational and environmental toxicity concerns. Improving our understanding of these issues also has broad appeal in the stewardship of nanotechnology and its acceptance by the public. This work represents some of the early research in burgeoning field of nanotoxicology. Using a variety of in vivo and in vitro models, as well as cellular and molecular techniques I first studied a possible role for the novel cytokine endothelial monocyte activating polypeptide-II (EMAP-II) in acute lung inflammation in rats (Chapter 2). This work demonstrated a significant increase in total EMAP-II concentration in lipopolysaccharide inflamed lungs as early as 1h post-treatment (P<0.05). Increased numbers of monocytes and granulocytes were also observed in lungs treated with mature EMAP-II relative to control rats (P<0.05), and the recruitment of cells did not occur via upregulation of either Interleukin-1β or Macrophage inflammatory protein-2. I further studied whether mature EMAP-II can be induced in pulmonary nanotoxicity studies by exposure to rosette nanotubes (RNT) (Chapters 3-5). In the first in vivo experiments in mice on the RNT(1)-G0 (Chapter 3) I showed an acute inflammatory response at the 50 µg dose by 24h, but this response was resolving by 7d post-exposure as evidenced by a decreased number of cells in the bronchoalveolar lavage fluid (P<0.05) and from histological examination. The results of this study indicated that water soluble and metal-free rosette nanotubes can demonstrate a favorable acute pulmonary toxicity profile in mice. Subsequently, I studied the responses of the pulmonary epithelium using the human Calu-3 cell line (Chapter 4). This experiment indicated that RNT(2)-K1 neither reduces cell viability at 1 or 5 µg/ml doses nor does it induce a dose-dependent inflammatory cytokine response in pulmonary epithelial cells in vitro. My final experiment (Chapter 5) studied the human U937 pulmonary macrophage cell line since the macrophage is one of the key defense mechanisms to encounter RNT in the lung environment. The data indicate that this cell line lacks a robust inflammatory response upon exposure to RNT and that when RNT length is changed by altering the conditions of nanotube self-assembly, cytokine release into the supernatant is not affected profoundly. Although, EMAP-II is upregulated in a lipopolysaccharide model of lung inflammation, it does not serve as a good marker of RNT exposure. The data indicate that RNT have a favourable toxicity profile and these experiments provide a framework upon which rosette nanotubes can be investigated for a range of biomedical applications. Furthermore, in light of media and scientific reports of nanomaterials showing signs of toxicity, this work demonstrates that a biologically inspired nanostructure such as the RNT can be introduced to physiological environments without acute toxicity.

A growing demand for information on the human health and environmental effects of materials produced using nanotechnology has led to a new area of investigation known as nanotoxicology. Research in this field has widespread implications in facilitating the medical applications of nanomaterials but also in addressing occupational and environmental toxicity concerns. Improving our understanding of these issues also has broad appeal in the stewardship of nanotechnology and its acceptance by the public. This work represents some of the early research in burgeoning field of nanotoxicology. Using a variety of in vivo and in vitro models, as well as cellular and molecular techniques I first studied a possible role for the novel cytokine endothelial monocyte activating polypeptide-II (EMAP-II) in acute lung inflammation in rats (Chapter 2). This work demonstrated a significant increase in total EMAP-II concentration in lipopolysaccharide inflamed lungs as early as 1h post-treatment (P<0.05). Increased numbers of monocytes and granulocytes were also observed in lungs treated with mature EMAP-II relative to control rats (P<0.05), and the recruitment of cells did not occur via upregulation of either Interleukin-1β or Macrophage inflammatory protein-2. I further studied whether mature EMAP-II can be induced in pulmonary nanotoxicity studies by exposure to rosette nanotubes (RNT) (Chapters 3-5). In the first in vivo experiments in mice on the RNT(1)-G0 (Chapter 3) I showed an acute inflammatory response at the 50 µg dose by 24h, but this response was resolving by 7d post-exposure as evidenced by a decreased number of cells in the bronchoalveolar lavage fluid (P<0.05) and from histological examination. The results of this study indicated that water soluble and metal-free rosette nanotubes can demonstrate a favorable acute pulmonary toxicity profile in mice. Subsequently, I studied the responses of the pulmonary epithelium using the human Calu-3 cell line (Chapter 4). This experiment indicated that RNT(2)-K1 neither reduces cell viability at 1 or 5 µg/ml doses nor does it induce a dose-dependent inflammatory cytokine response in pulmonary epithelial cells in vitro. My final experiment (Chapter 5) studied the human U937 pulmonary macrophage cell line since the macrophage is one of the key defense mechanisms to encounter RNT in the lung environment. The data indicate that this cell line lacks a robust inflammatory response upon exposure to RNT and that when RNT length is changed by altering the conditions of nanotube self-assembly, cytokine release into the supernatant is not affected profoundly. Although, EMAP-II is upregulated in a lipopolysaccharide model of lung inflammation, it does not serve as a good marker of RNT exposure. The data indicate that RNT have a favourable toxicity profile and these experiments provide a framework upon which rosette nanotubes can be investigated for a range of biomedical applications. Furthermore, in light of media and scientific reports of nanomaterials showing signs of toxicity, this work demonstrates that a biologically inspired nanostructure such as the RNT can be introduced to physiological environments without acute toxicity.

Nanoscience and nanotechnology have advanced rapidly in recent years and have made a profound impact in the medical field. Nanoparticles have attracted great attention for their potential as diagnostic and/or therapeutic tools in oncology owing to their unique properties. Theranostics are nanodevices with diagnostic, therapeutic and possibly treatment-monitoring functions for treating cancers. Different noble metal nanoparticles can provide the basic unit for theranostics. Suitably designed and developed noble metal nanoparticle-based theranostics will have multiple functions. In this project, the design, fabrication and performance of novel multifunctional nanodevices for cancer detection and treatment were investigated. The foundation of this project was laid by investigating different types of hybrid nanoparticles for novel theranostics. Different approaches were developed for fabricating core-shell structured hybrid nanoparticles. Highly branched gold and gold-silver bimetallic nanoparticles were firstly made. pH-sensitive folic acid-chitosan (CS-FA) conjugate was then introduced on these nanoparticles to form hybrid nanoparticles with a metal core (Au@CS-FA and Au-Ag@CS-FA). Poly(lactide-co-glycolide) (PLGA) and chitosan (CS) micro- or nanoparticles were also produced to serve as the polymer core for forming hybrid particles with a gold or gold-silver nanoshell (PLGA@Au, CS@Au and PLGA@Ag-Au). Furthermore, Fe3O4@Au nanoparticles having both magnetic and plasmonic properties were investigated. Thermo-sensitive poly(N-isopropylacrylamide) (pNIPAm) polymer or pH-sensitive CS-FA was then coated on Fe3O4@Au nanoparticles, forming new hybrid nanoparticles. The formation mechanisms of nanoparticles and hybrid nanoparticles were studied. Raman reporters (Rhodamine B or 4-mercaptobenzoic acid) and anti-cancer drugs (paclitaxel or 5-fluorouracil) were loaded into the polymer core or shell of hybrid nanoparticles to form multifunctional nanodevices. While the noble metal unit in the nanodevices provided high light-scattering enhancement for achieving photothermal effect, the polymer component encapsulated Raman reporter molecules and put them close to the metal nanoparticles for generating high surface enhanced Raman scattering (SERS) signals. These nanodevices could also serve as excellent drug carriers, and the stimulus-triggered release of incorporated drug was studied. It was shown in this project that the conjugation of targeting ligand (e.g. folic acid) or antibody (e.g. anti-HER2 monoclonal antibody) on hybrid nanoparticles had formed novel theranostics which allowed selective detection, continuous imaging of intracellular behavior and killing of targeted cancer cells. These theranostics could be taken up by specific cancer cells through receptor-mediated endocytosis and internalized into cytoplasma of the cell. These theranostics as stable SERS-active tags and imaging agents for HeLa cells, SK-BR-3 cancer cells or MCF-7 cancer cells were demonstrated. The targeting ability and intracellular uptake of these theranostics were studied. The photothermal effect of the theranostics was investigated using different laser irradiation powers. The anti-cancer treatment could be significantly improved by the synergistic effects of chemo- and photothermal therapy when these theranostics were also tasked as the carrier of anti-cancer drugs. Therefore, combining plasmonic metal nanoparticles with targeting ligand or antibody, magnetic nanoparticles, polymer shell or core, and anti-cancer drug has created advanced theranostics for the early detection and effective treatment of cancers. These novel theranostics have greatly improved capability for cancer detection and can provide multifunctions for cancer cell targeting, sensing/imaging and combined therapy. / published_or_final_version / Mechanical Engineering / Doctoral / Doctor of Philosophy

Nanomedicine utilises biocompatible nanomaterials for therapeutic as well as imaging purposes, for the treatment of various diseases including cancer, neurological disorders and wound infections. Graphene, a material composed of a single layer of carbon atoms, has recently shown great potential to improve diagnostics and therapeutics, owing to its small size, large surface-area-to-volume ratio and unique physicochemical properties. However, the limited fabrication, in vitro and in vivo functionalities published in the literature indicate inconsistencies regarding the factors affecting metabolic fate, biodistribution as well as toxicity patterns of graphene. This thesis focuses on the biological effects of graphene-based materials, including graphene oxide (GO), reduced graphene oxide (rGO), graphene nanopores (GNPs), graphene quantum dots (GQDs) and three-dimensional graphene foam (GF). These can be used to closely mimic therapeutic functions and thereby open up new pathways to anticancer nanomedicine. In this work, a biocompatible GO-based anti-metastatic enzyme cancer therapy approach has been introduced for the first time to target the extracellular pro-metastatic and pro- tumourigenic enzymes of cathepsin D and cathepsin L, which are typically overexpressed in ovarian and breast cancers. Definitive binding and modulation of cathepsin- D and -L with GO has revealed that both of the enzymes were adsorbed onto the surface of GO through its cationic and hydrophilic residues under the biologically relevant condition of acidic pH. It has been demonstrated that low concentrations of rGO were shown to significantly produce late apoptosis and necrosis rather than early apoptotic events in lung cancer cells (A549 and SKMES-1), suggesting that it was able to disintegrate the cellular membranes in a dose-dependent manner. GNPs at lower concentrations (250μg/ml) induce upregulation of phosphatidylserine on cell surface membrane (i.e. early apoptotic event), which does not significantly disintegrate the cell membrane in the aforementioned lung cancer cells, while higher concentrations of GNPs (5 and 15 mg/kg) in rats (when intraperitoneally injected) exhibited sub-chronic toxicity in a period of 27 days. The interaction of GQDs and trypsin has revealed the strong bonding capacity of GQDs with trypsin, owing to their surface charge and surface functionalities evidencing the high bioavailability of GQDs in enzyme engineering. Finally, 3D GF was developed to probe the role of graphene-based scaffold cues in the field of regenerative medicine revealing their cell attachment to in vitro cell cultures. Furthermore, GF was shown to maintain remarkable biocompatibility with in vitro and in vivo toxicity screening models when exposed for 7 days at doses of 5, 10 and 15 mg/l. Taken together, graphene and its modified structures developed in this thesis promise to revolutionise clinical settings across the board in nanomedicine which include, but are not limited to, ultra-high sensitive enzyme adsorbents, high throughput biosensors, enzyme modulators and smart scaffolds for tissue regeneration.

Fullerenes and metallofullerenes have been discovered to have a lot of applications in the biomedical area, for instance, they have been shown to have antioxidant, anti-virus, anti-cancer, immunological properties, etc. However, the hydrophobicity nature of fullerenes and metallofullerenes raises the need for functionalized hydrophilic fullerenes and metallofullerenes. Also, the advancement of the purification techniques of fullerenes and metallofullerenes makes the isolation of new fullerenes and metallofullerenes possible. Therefore, discovering the biomedical applications of these newly found fullerenes and metallofullerenes is also of vital importance. In Chapter 1, we provided a comprehensive background on the history of fullerenes and metallofullerenes, synthesis and purification methods of fullerenes and metallofullerenes, and some of their biological applications, including antioxidant applications and fluorescence applications. Some important fullerene and metallofullerenes and milestones in this area were also discussed. In Chapter 2, we demonstrated the antioxidant and anti-inflammation ability of a conjugate, FIFIFK(Cy5)PEG24K(NH2)CONH2-C60, that consisted of a peptide that binds specifically to a formyl peptide receptor-1 (FPR-1), which expresses on activated macrophages, and a carboxyl-group-functionalized C60, which is the first discovered and most used fullerene is history. We showed that the fullerene-peptide conjugate had great ability as a radical scavenger and to reduce the volume of inflammatory tissue. In Chapter 3, we demonstrated the antioxidant and anti-inflammation ability of several metallofullerene derivatives, including amino-Gd3N@C80, amino-Sc3N@C80, carboxy-Gd3N@C80, and carboxyl-Sc3N@C80. Amino-group-functionalized metallofullerenes were found to have higher radical scavenging ability and anti-inflammation ability. In Chapter 4, we developed fluorescent metallofullerene derivatives, including Tb3N@C80(NH2)9(OH)4NO2 and Tb3N@C80(CH2CH2COOH)21(OH)18. The fluorescence properties of the derivatives of fullerenes or metallofullerenes were normally realized by the addition of fluorescent moieties on the carbon cage of fullerenes or metallofullerenes. However, the fluorescence of our newly developed metallofullerene derivative was realized by the tri-terbium nitride cluster inside the carbon cage. This saved the multi-step synthesis of the fluorescence probe and maintained the simplicity of the structure of the metallofullerene derivative. These derivatives of terbium-containing metallofullerene also showed radical scavenging ability towards hydroxyl radicals. In Chapter 5, we developed another fluorescent Tb3N@C80 derivative, Tb3N@C80[DiPEG2000]. The advantage of this fluorescent metallofullerene derivative was that it was easier to purify compared to small-functional-groups-functionalized Tb3N@C80 derivatives. This newly developed fluorescent Tb3N@C80 derivative also showed great fluorescent ability and radical scavenging ability. In Chapter 6, we provided a summary of the studies on the antioxidant and fluorescent properties of fullerenes and metallofullerenes' derivatives that were discussed in this dissertation. / Doctor of Philosophy / Fullerenes and metallofullerenes' have large carbonaceous outer structures that give them possibilities for a lot of chemical reactions. The conjugated carbonaceous outer structures also endow them with exceptional antioxidant ability by reacting with the oxidant species. Therefore, fullerenes and metallofullerenes can be considered as great antioxidants, which are substances that can prevent or slow damage to cells caused by oxidants, in most cases free radicals or other unstable oxidant molecules that the body produces as reactions to environmental and other pressures. Fullerenes and metallofullerenes are sometimes called "free-radical scavengers". However, fullerenes and metallofullerenes are not soluble in biological systems, therefore, it's necessary to attach functional groups onto the outer cage-like structures of fullerenes and metallofullerenes to make them water-soluble, while maintaining their great antioxidant properties. Fullerenes and metallofullerenes are not limited to antioxidant applications, and with the attachment of specialized moieties, for instance, short peptides, they can be made of additional applications. Different metals in these metallofullerenes also provide them with specialized applications. Herein, we have developed a series of water-soluble fullerenes and metallofullerenes and compared their antioxidant properties. We have also developed a water-soluble C60-peptide conjugate that can specifically bind to inflammatory cells and ease the inflammatory condition. We also synthesized a series of metallofullerene derivatives that had dual modalities fluorescent properties and antioxidant properties.