A novel, lipid-based platform nanotechnology has been designed to overcome limitations of in vivo fluorescent imaging, multidrug resistance (MDR) phenotypes hindering breast cancer chemotherapy, and shortcomings of magnetic resonance imaging (MRI) thermometry. Using this platform, three nanoparticle systems have been developed: QD-SLN (quantum dot-loaded solid lipid nanoparticles), DMsPLN (doxorubicin and mitomycin C co-loaded polymer-lipid hybrid nanoparticles), and HLN (hydrogel-lipid hybrid nanoparticles). Stealth, near-infrared emitting QD-SLN were developed for deep tissue fluorescence imaging, which were capable of extending the depth of penetration beyond 2 cm, with near complete probe clearance and good tolerability in vivo. The QD-SLN was used to evaluate the biodistribution of non-targeted SLN and actively targeted RGD-conjugated SLN. Non-targeted SLN accumulated in breast tumors and evaded liver uptake. The RGD-SLN showed prolonged retention in breast tumor neovasculature at the cost of lesser tumor accumulation due to enhanced liver uptake. With this information, a long circulating, non-targeted DMsPLN with a synergistic cancer chemotherapeutic combination of doxorubicin and mitomycin C was formulated to overcome MDR, enhancing breast cancer chemotherapy. Extensive tumor cell uptake and perinuclear trafficking of DMsPLN overcame the MDR phenotype of breast tumor cells in vitro. The DMsPLN provided the most efficacious chemotherapy reported in literature against aggressive mouse mammary tumors in vivo with significant reduction in whole animal and cardiotoxicity as compared to clinically applied liposomal doxorubicin. In establishing our tumor models, the impact of Matrigel™ on the tumor microenvironment was investigated, demonstrating altered tumor vascular and lymphatic anatomy and physiology, and significantly impacting nanomedicines assessment in mouse models of cancer. In all in vivo studies, tumors were established without use of Matrigel™. To guide thermotherapy of solid tumors, a novel HLN was formulated for use in MRI thermometry, presenting the first contrast agent capable of indicating a tunable, absolute two-point temperature window. In using specific limitations of therapeutic and imaging modalities to inform rational nanoparticle design, this lipid-based platform nanotechnology has extended the application of fluorescence imaging in vivo, enhanced the utility of nanoparticulate chemotherapeutics against breast cancer independent of MDR status, and provided novel functionality for MRI thermometry.

A novel, lipid-based platform nanotechnology has been designed to overcome limitations of in vivo fluorescent imaging, multidrug resistance (MDR) phenotypes hindering breast cancer chemotherapy, and shortcomings of magnetic resonance imaging (MRI) thermometry. Using this platform, three nanoparticle systems have been developed: QD-SLN (quantum dot-loaded solid lipid nanoparticles), DMsPLN (doxorubicin and mitomycin C co-loaded polymer-lipid hybrid nanoparticles), and HLN (hydrogel-lipid hybrid nanoparticles). Stealth, near-infrared emitting QD-SLN were developed for deep tissue fluorescence imaging, which were capable of extending the depth of penetration beyond 2 cm, with near complete probe clearance and good tolerability in vivo. The QD-SLN was used to evaluate the biodistribution of non-targeted SLN and actively targeted RGD-conjugated SLN. Non-targeted SLN accumulated in breast tumors and evaded liver uptake. The RGD-SLN showed prolonged retention in breast tumor neovasculature at the cost of lesser tumor accumulation due to enhanced liver uptake. With this information, a long circulating, non-targeted DMsPLN with a synergistic cancer chemotherapeutic combination of doxorubicin and mitomycin C was formulated to overcome MDR, enhancing breast cancer chemotherapy. Extensive tumor cell uptake and perinuclear trafficking of DMsPLN overcame the MDR phenotype of breast tumor cells in vitro. The DMsPLN provided the most efficacious chemotherapy reported in literature against aggressive mouse mammary tumors in vivo with significant reduction in whole animal and cardiotoxicity as compared to clinically applied liposomal doxorubicin. In establishing our tumor models, the impact of Matrigel™ on the tumor microenvironment was investigated, demonstrating altered tumor vascular and lymphatic anatomy and physiology, and significantly impacting nanomedicines assessment in mouse models of cancer. In all in vivo studies, tumors were established without use of Matrigel™. To guide thermotherapy of solid tumors, a novel HLN was formulated for use in MRI thermometry, presenting the first contrast agent capable of indicating a tunable, absolute two-point temperature window. In using specific limitations of therapeutic and imaging modalities to inform rational nanoparticle design, this lipid-based platform nanotechnology has extended the application of fluorescence imaging in vivo, enhanced the utility of nanoparticulate chemotherapeutics against breast cancer independent of MDR status, and provided novel functionality for MRI thermometry.

Nanomaterials have gained great attention for biomedical applications due to their extraordinary physico-chemical and biological properties. The current dissertation presents the design and development of multifunctional nanoparticles for molecular imaging and controlled drug delivery applications which include biodegradable polymeric nanoparticles, superparamagnetic iron oxide nanoparticles (SPION)/polymeric nanocomposite for magnetic resonance imaging (MRI) and drug delivery, manganese-doped zinc sulfide (Mn:ZnS) quantum dots (QDs)/ SPION/ polymeric nanocomposites for fluorescence imaging, MRI and drug delivery.Bioimaging is an important function of multifunctional nanoparticles in this thesis. Imaging probes were made of SPION and Mn:ZnS QDs for in vitro and in vivo imaging. The SPION have been prepared through a high temperature decomposition method to be used as MRI contrast agent. SPION and Mn:ZnS were encapsulated into poly (lactic-co-glycolic) acid (PLGA) nanoparticles during the particles formation. The hydrophobic model drug, busulphan, was loaded in the PLGA vesicles in the composite particles. T2*-weighted MRI of SPION-Mn:ZnS-PLGA phantoms exhibited enhanced negative contrast with r2* relaxivity of 523 mM-1 s-1. SPION-Mn:ZnS-PLGA-NPs have been successfully applied to enhance the contrast of liver in rat model.The biodegradable and biocompatible poly (ethylene glycol)-co-poly (caprolactone) (PEG-PCL) was used as matrix materials for polymeric nanoparticles -based drug delivery system. The PEG-PCL nanoparticles have been constructed to encapsulate SPION and therapeutic agent. The encapsulation efficiency of busulphan was found to be ~ 83 %. PEG-PCL nanoparticles showed a sustained release of the loaded busulphan over a period of 10 h. The SPION-PEG-PCL phantoms showed contrast enhancement in T2*-weighted MRI. Fluorescein-labeled PEG-PCL nanoparticles have been observed in the cytoplasm of the murine macrophage cells (J774A) by fluorescence microscopy. Around 100 % cell viability were noticed for PEG-PCL nanoparticles when incubated with HL60 cell line. The in vivo biodistribution of fluorescent tagged PEG-PCL nanoparticles demonstrated accumulation of PEG-PCL nanoparticles in different tissues including lungs, spleen, liver and kidneys after intravenous administration. / <p>QC 20160516</p>

Existing methods of treating glioma are not effective for eradicating the disease. Therefore, new and innovative methods of treatment alone or in combination with existing therapies are necessary. Delivery of therapeutic agents through delivery carriers such as liposomes diminishes the harmful effects of the agent in healthy tissues and allows increased accumulation in the tumor. In addition, targeted chemotherapy using liposomes provides the opportunity for further increase in drug accumulation in tumor. However, the current targeting strategies suffer accelerated plasma clearance and are not advantageous in improving efficacy. The search for new tumor targets, novel ligands, new strategies for targeting, and particle stabilization will advance our ability to improve delivery at the tumor level while decreasing toxicity to normal tissues. The global objective of this thesis was to improve the status of current liposomal therapy to achieve higher efficacy in tumors. Here, we show a novel mechanism to increase targeting to tumor while uncompromising on the long circulation of stealth liposomes. Long circulation is essential for passive accumulation of the nanocarriers due to EPR effect, in order to see benefits of targeting. Using phage display technique, a variety of tumor specific peptides were identified for use as targeting moieties. One potential advantage of the approach proposed here is the rapid identification of patient tumor specific peptide that evades the RES. This could lead to the development of a nanocarrier system with high avidity and selectivity for tumors. Therefore, tumor accumulation of the targeted formulations will be higher than that of non‐targeted liposomes due to increased drug retention at the tumor site and uncompromised blood residence time.In addition, it has been shown that the distribution of nanocarriers, spatially within the tumor, is limited that might further hinder the distribution of the encapsulated drug, thereby limiting efficacy. It is necessary to release the drug from within the nanocarrier to promote increased efficacy. Here, we were able to address the problem of drug diffusion within the tumor interstitium using a combination therapy employing a remotely triggered thermosensitive liposomal chemotherapeutic. We fabricated a thermosensitive liposomal nanocarrier that maintained its stability at physiological temperature to minimize toxicity to healthy cells. We, then, showed a remote triggering mechanism mediated by gold nanorods heated via NIR can help in achieving precise control over the desired site for drug release. These strategies enabled increased drug availability at the tumor site and contributed to tumor retardation. Additionally, we show that the synergistic therapy employing gold nanorods and thermosensitive liposomes may have great potential to be translated to the clinic.

Actuellement, la chirurgie représente la première indication pour la thérapie du cancer. Néanmoins, la résection complète du tissu tumoral, la détection des micrométastases et la préservation des tissus sains pendant l'intervention représentent un enjeu majeur et influencent fortement le pronostic du patient. Les récents développements technologiques en imagerie pour la chirurgie guidée des cancers ont conduit à des résultats précliniques prometteurs et les premiers essais cliniques utilisant des traceurs non-spécifiques confirment déjà le potentiel de ces systèmes pour l'amélioration de la chirurgie. De plus, le diagnostic précoce des tumeurs, ainsi que le développement de thérapies ciblées sont également des axes majeurs de recherche en cancérologie. Dans ce contexte notre équipe a précédemment développé un vecteur synthétique ciblant un récepteur cellulaire l'intégrine αVβ3. Ce vecteur est constitué d'un châssis décapeptidique cyclique RAFT (Regioselectively Addressable Functionalized Template) et présentant deux domaines indépendants permettant de séparer les deux fonctions du vecteur. Sur un domaine, la fonction de ciblage est assurée par la présentation multivalente de ligands -RGD- spécifiques du récepteur. L'autre domaine du vecteur porte les molécules d'intérêt à vectoriser, agents thérapeutiques ou de détection pour l'imagerie médicale. Dans la première partie de ces travaux, nous avons évalué la combinaison de ce vecteur couplé à un fluorophore avec une sonde portative pour imager et guider le chirurgien pendant la chirurgie des fibrosarcomes spontanés chez le chat. Cette étude représente une preuve de concept pour la translation clinique chez l'homme. Les résultats ont montré que l'injection du traceur ne provoquait pas d'effets toxiques chez le chat et permettait un marquage spécifique de la tumeur avec un bon ratio tumeur/tissu sain, qui devrait améliorer la qualité de la résection tumorale en aidant le chirurgien à mieux délimiter les marges du tissu tumoral. Dans la seconde partie de ces travaux nous avons développé un nouveau vecteur bi-fonctionnel dérivé du RAFT-RGD. Au composé d'origine a été ajoutée une séquence peptidique clivable par la matrixmetalloprotease-9, une enzyme surexprimée dans la tumorigénèse. Cette molécule à fluorescence activable a montré une amélioration du ciblage tumoral in vitro et in vivo comparée au RAFT-RGD suggérant un effet additionnel lié au double ciblage. Ces résultats préliminaires encouragent la poursuite de sa caractérisation pour son potentiel de « pro-drug » mais également pour l'étude des interactions entre l'intégrine et l'environment tumoraux. / Cancer surgery is still the gold standard therapy in most cancers. Nevertheless, total tumor resection and metastasis detection while preserving healthy tissues represent a crucial point for further prognosis. Development of imaging technologies for intra-operative guided surgery provided promising results and efficient application in preclinical studies and first clinical trials using non-specific tracers already confirmed the improved out-come in surgery. Moreover early and precise diagnosis and targeted therapies are major domains of cancer research. In this context our team previously developed a synthetic vector based on a cyclic decapeptide scaffold RAFT (Regioselectively Addressable Functionalized Template) which allows the independent functionalizing of two domains: a targeting domain with multivalent RGD-ligand targeting the cell receptor integrin αVβ3, and a vehicle domain grafted with a pro-drug or an imaging agent. One part of this work consisted in the evaluation of the combination of this molecule carrying a fluorophore with a portable fluorescent imaging device for image-guided surgery of natural occurring feline fibrosarcomas. This study represents a proof of concept for further translation into human clinics. No toxic effects in cats after administration of the tracer could be reported. Furthermore the tumors were specifically labeled showing a good tumor-to-healthy tissue ratio. This should improve tumor resection by helping the surgeon to delineate tumor margins. In parallel we developed a bi-functinal derivative of the RAFT-RGD. Therefore we engrafted a peptide sequence flanked by two fluorophores, which is activatable by matrixmetalloprotease-9, an enzyme overexpressed in tumors. This molecule showed an improved tumor labeling in vitro and in vivo compared to the conventional RAFT-RGD, suggesting an additional effect of the double targeting. These preliminary results encourage further caracterisation for its potential as pro-drug vehicle, as well as for studying interactions between the integrin and the tumor environment.