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<p>Cancer is currently the second leading cause of death in the United States. Although many treatment options exist, some of the most common, including radiotherapy and chemotherapy, are restricted by dose-limiting toxicities. In addition, the largest hurdle for translating novel biological therapies such as siRNA into the clinic is lack of an efficient delivery mechanism to get the therapeutic into malignant cells. This work aims to improve this situation by engineering a minimally invasive controlled release system that specifically delivers therapeutics to the site of malignant tissue. This platform consists of two novel material components: a thermally responsive poly[N-isopropylacrylamide-co-acrylamide] (NIPAAm-co-AAm) hydrogel and gold-silica nanoshells. Therapeutic molecules are encapsulated within a poly(NIPAAm-co-AAm) hydrogel carrier, leading to increased serum stability, circulation time, and decreased exposure to off-site tissues. Additionally, gold-silica nanoshells embedded within this hydrogel will be used to optically trigger therapeutic release from the carrier. This hydrogel-nanoshell composite material was designed to be swollen under physiologic conditions (37 oC), and expel large amounts of water and absorbed molecules at higher temperatures (40-45 oC). This phase transition can be optically triggered by embedded gold-silica nanoshells, which rapidly transfer near-infrared (NIR) light energy into heat due to the surface plasmon resonance phenomena. NIR light can deeply penetrate biological tissue with little attenuation or damage to tissue, and upon exposure to such light a rapid temperature increase, hydrogel collapse, and drug expulsion will occur. Ultimately, these drug-loaded hydrogel-nanoshell composite particles would be injected intravenously, passively accumulate in tumor tissue due to the enhanced permeability and retention (EPR) effect, and then can be externally triggered to release their therapeutic payload by exposure to an external NIR laser. This dissertation describes the synthesis, characterization, and validation of such a controlled therapeutic delivery platform.</p><p>Initial validation of poly(NIPAAm-co-AAm)-gold nanoshell composites to act as a material in site-specific cancer therapeutic delivery was accomplished using bulk hydrogel-nanoparticle composite disks. The composite material underwent a phase transition from a hydrated to a collapsed state following exposure to NIR light, indicating the ability of the NIR absorption by the nanoshells to sufficiently drive this transition. The composite material was loaded with either doxorubicin or a DNA duplex (a model nucleic acid therapeutic), two cancer therapeutics with differing physical and chemical properties. Release of both therapeutics was dramatically enhanced by NIR light exposure, causing 2-5 fold increase in drug release. Drug delivery profiles were influenced by both the molecular size of the drug as well as its chemical properties. </p><p>Towards translation of this material into in vivo applications, the hydrogel-nanoshell composite material was synthesized as injectable-sized particles. Such particles retained the same thermal properties as the bulk material, collapsing in size from ~330 nm to ~270 nm upon NIR exposure. Furthermore, these particles were loaded with the chemotherapeutic doxorubicin and NIR exposure triggered a burst release of the drug payload over only 3 min. In vitro, this platform provided increased delivery of doxorubicin to colon carcinoma cells compared to free-drug controls, indicating the irradiated nanoshells may increase cell membrane permeability and increase cellular uptake of the drug. This phenomena was further explored to enhance cellular uptake of siRNA, a large anionic therapeutic which cannot diffuse into cells easily. </p><p>This work advances the development of an injectable, optically-triggered delivery platform. With continued optimization and in vivo validation, this approach may offer an novel treatment option for cancer management.</p> / Dissertation

Our laboratory has used the 1,4,5,8 Naphthalenetetracarboxylic diimine (NDI) unit to develop threading polyintercalators that bind DNA with the NDI units intercalated in between GpG steps and two different peptide linkers, which connect the NDI units, situated in either the major or minor grooves. The first generation bisintercalators, G₃K and [beta]Ala₃K, were shown to bind two different sequences of DNA, where the peptide linkers reside in the major and minor grooves respectively. These binding modules were then combined to generate threading polyintercalators that bound different DNA sequences with simultaneous occupation of both grooves. In particular, a cyclic bisintercalator was designed and DNAse I footprinting revealed a strong preference for the sequence 5'-GGTACC-3'. NMR structural studies of the complex with d(CGGTACCG)₂ verified a pseudocatenane structure in which the NDI units reside four base pairs apart, with one linker located in the minor groove and the other in the major groove. This was the first structurally well-characterized pseudocatenane complex between a sequence-specific cyclic bisintercalator and its preferred binding sequence. The ability to simultaneously occupy both groves of the same sequence is interesting for several reasons. Most significantly, it raises questions about a complex DNA intercalator's ability to locate its preferred sequence within a long strand of DNA. In order to directly assess this, the intercalator was modified (CBI-Cys) to incorporate a gold nanoparticle probe to allow for the direct visualization of the intercalator locating its preferred sequence within a long DNA strand. The appropriate protocols to visualize DNA using electron and atomic force microscopy were unsuccessful; however, the foundation has been set for future work to develop the appropriate method to determine the mechanism by which the cyclic bisintercalator locates its preferred sequence. Additionally, the bisintercalators developed in our laboratory offered a unique opportunity to exploit their sequence specificity for controlled nanoparticle assembly. Over the past decade, nanoparticles and DNA have been used to develop novel nanoparticle assembly systems with the goal of developing electronic devices and nanomaterials. The G₃K bisintercalator was synthetically modified to incorporate a gold nanoparticle probe. This intercalator-nanoparticle conjugate, BisKC·Au, maintained its binding specificity (5'-GGTACC-3') to a modified DNA fragment containing multiple G₃K binding sites. The atomic force microscope has become the most promising tool in visualizing individual DNA molecules. A modified procedure utilized APS to allow for the direct visualizing of plasmid DNA. The framework is now in place to confirm the controlled assembly of the gold nanoparticles. This protocol can then be used for the [beta]Ala₃K bisintercalator to lead to the development of a nanoparticle assembly system that can precisely control the organization of multiple types of nanoparticles. / text

A significant challenge in the surgical resection of tumors is accurate identification of tumor margins. Current methods for margin detection are time-intensive and often result in incomplete tumor excision and recurrence of disease. The objective of this project was to develop a near-infrared narrowband imaging (NIR NBI) system to image tumor and its margins in real-time during surgery utilizing the contrast provided by gold nanoparticles (GNPs). NIR NBI images narrow wavelength bands to enhance contrast from plasmonic particles in a widefield, portable and non-contact device that is clinically compatible for real-time tumor margin demarcation. GNPs have recently gained significant traction as nanovectors for combined imaging and photothermal therapy of tumors. Delivered systemically, GNPs preferentially accumulate at the tumor site via the enhanced permeability and retention effect, and when irradiated with NIR light, produce sufficient heat to treat tumor tissue. The NIR NBI system consists of 1) two LED's: green (530 nm) and NIR (780 nm) LED for illuminating the blood vessels and GNP, respectively, 2) a filter wheel for wavelength selection, and 3) a CCD to collect reflected light from the sample. The NIR NBI system acquires and processes images at a rate of at least 6 frames per second. We have developed custom control software with a graphical user interface that handles both image acquisition and processing/display in real-time. We used mice with a subcutaneous tumor xenograft model that received intravenous administration and topical administration of gold nanoshells and gold nanorods. We determined the GNP's distribution and accumulation pattern within tumors using NIR NBI. Ex vivo NIR NBI of tumor xenografts accumulated with GNPs delivered systemically, demonstrated a highly heterogeneous distribution of GNP within the tumor with higher accumulation at the cortex. GNPs were observed in unique patterns surrounding the perivascular region. The GNPs clearly defined the tumor while surrounding normal tissue did not indicate the presence of particles. In addition, we present results from NBI of tumors that received topical delivery of conjugated GNPs. We determined that tumor labeling using topical delivery approach resulted in a more homogenous distribution of GNPs compared to the systemic delivery approach. Finally, we present results from the on-going in vivo tumor margin imaging studies using NIR NBI. Our results demonstrate the feasibility of NIR NBI in demarcating tumor margins during surgical resection and potentially guiding photo-thermal ablation of tumors. / text

published_or_final_version / Chemistry / Master / Master of Philosophy

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The Mt. Milligan deposit is a tilted (~45°) Cu-Au alkalic porphyry located 155 km northwest of Prince George, B.C., Canada. It is the youngest of the BC alkalic porphyry deposits, all of which formed between 210 to 180 Ma in an extensive belt of K-enriched rocks related to the accretion of the Quesnellia-Stikinia superterrane to ancestral North America. Mt. Milligan has a measured and indicated resource of 205.9 million tonnes at 0.60 g/t Au and 0.25% Cu containing 3.7 million oz. gold, and 1.12 billion lb. copper. Shoshonitic volcanic and volcaniclastic andesites host mineralization. These have been intruded by a composite monzonitic stock (MBX stock), and associated sill (Rainbow Dike). Early disseminated chalcopyrite-magnetite and accessory quartz veins are associated with K-feldspar alteration in the MBX stock. A halo of biotite alteration with less extensive magnetite replaces host rocks within a ~150 m zone surrounding the stock, while K-feldpsar alteration extends along the Rainbow Dike and permeable epiclastic horizons. Peripheral albite-actinolite-epidote assemblages surround the K-silicate zone. Albite-actinolite occurs at depth, and epidote dominates laterally. Copper and Au grade are maximal where the albite-actinolite assemblage overprints biotite alteration. Gold grade is moderate in association with epidote, whereas Cu is depleted. The post-mineral Rainbow Fault separates the core Cu-rich zone from a downthrown Au-rich zone. A similar zonation of metals occurs in the hanging-wall (66 zone), where a Cu-bearing, potassically-altered trachytic horizon transitions to a funnel-shaped zone of pyrite-dolomite-sericite-chlorite alteration with elevated gold. Sulfide S-isotope compositions range from -4.79 δ34S in the central Cu-Au orebody to near-zero values at the system periphery, typical of alkalic porphyries. Sulfur isotope contours reflect the magmatic-hydrothermal fluid evolution, and indicate late-stage ingress of peripheral fluids into the Cu-Au zone. Carbonate C- and O-isotope compositions corroborate the magmatic fluid path from the Cu-Au rich zone to Au-rich zone with decreasing depth. Strontium isotopic compositions of peripheral alteration minerals indicate a laterally increasing meteoric fluid component. Changes in major- and trace element composition of epidote and pyrite across the deposit are also systematic. These provide additional vectors to ore, and confirm the kinematics of the Rainbow Fault.

Part of the Slave Structural Province, the Hope Bay Greenstone Belt is a 82 km long north-striking sequence of supracrustal rocks dominated by mafic volcanic rocks with lesser felsic volcanic and sedimentary rocks. Mapping of two transects in the southern section and two transects in the northern section have contributed to a robust stratigraphic framework the belt. Three recently discovered Archean lode gold deposits in the Hope Bay Greenstone belt have associations with major structures and specific lithologies (Fe-Ti enriched basalts). The Flake Lake and the Clover Transects are in the southern part of the belt and the Wolverine and Doris-Discovery Transects are in the northern part of the belt. This work subdivides the volcanic rocks into distinct suites based upon field, petrologic, geochemical, and geochronologic criteria. Some of the suites are stratigraphically continuous and can be correlated tens of kilometres along strike thereby linking the two parts of the Hope Bay Greenstone Belt. U-Pb geochronology supports work by Hebel (1999) concluded that virtually all the supracrustal rocks in the Hope Bay Greenstone Belt were deposited over at least 53 m.y. (2716-2663 Ma), with the majority of the volcanism occurring after 2700 Ma. A number of basalt groups are identified and include the normal basalt, the LREE-enriched basalt, the Ti-enriched basalt and the Ti-enriched Al-depleted basalt groups. They have chemical signatures that vary in trace elements particularly HFSE and REE’s, and can be easily be distinguished by geochemical screening. The felsic volcanic suites are also divided into three main groups, tholeiitic rhyolite, calc-alkaline dacite and calc-alkaline rhyolite groups. Nd and Hf isotope signatures are consistent with trace element signatures in identifying mafic and felsic volcanic groups, with the tholeiitic rhyolite showing highly variable signature. The Hope Bay Greenstone Belt has been show to have a number of felsic and volcanic cycles. An early construction phase of the belt is made up of primarily mafic volcanics which is followed by felsic volcanism equalled mafic volcanism which lacks basalts enriched in Ti. The geodynamic environment that created the Hope Bay Greenstone Belt can be explained by plume influenced subduction zone.