Iron oxide nanocubes in nanomedicine: theranostic approach for cancer treatment and diagnosis
Creators: name: CASSANI, MARCO
Publisher: name: Universit� degli studi di Genova
Year: 2018

description: The present dissertation discloses the doctoral work carried out during the past three years at the Italian Institute of Technology (IIT) and the University of Genoa. The work was focused on the functionalization of iron oxide nanocubes (NCs) for different biological applications. Tuning the surface features of these magnetic nanocubes, as well as their assembly and intrinsic chemical and physical properties, resulted in the development of suitable nanotools for cancer theranostic, i.e. the combination of diagnosis and cancer therapy. The first chapter deals with the synthesis of magnetic nanoclusters, referred as magnetic nanobeads (MNBs). Here, maghemite (?-Fe2O3) nanocubes were tightly enwrapped into an amphiphilic polymer able to solubilize and stabilize them in water-based solutions. The synthetic route, reported in literature, was improved in order to obtain more stable polymeric shells that can be further functionalized with PEG-derivatized molecules. Due to the higher magnetic moment found for the MNBs, compared to that of single nanocubes, they were investigated for magnetic cell sorting. Therefore, a targeting feature was added to their surface by attaching a PEG molecule derivatized with folic acid (PEG-FA). This approach provides: 1) the binding of the MNBs to folate receptors overexpressed on the cell membrane of some cancer cells; 2) stability in complex biological media; 3) distance of the FA from the polymeric surface; 4) degree of freedom to the bioactive folic acid. A cancer cell line having high folate receptor expression profile (KB cell line) was chosen as a model for testing the sorting ability of the MNBs. The results obtained showed a significantly higher sorting efficiency for the MNBs functionalized with PEG-FA in comparison to the one observed for the MNBs functionalized with a non-biologically active PEG. This outcome reveals the potential of PEG-FA functionalized MNBs for the isolation of relevant folate receptor positive cancer cells, e.g. ovarian cancer cells, from biological tissues. In the second chapter, the structural transformation of core-shell wuestite/maghemite (FeO/?-Fe2O3) nanocubes into maghemite is reported. Here, an approach that is not often applied by material scientists was followed for the transformation under aqueous conditions. The non-interacting nature of core-shell nanocubes, due to their low magnetization, allowed for an easy and quantitative transfer in water, using a standard protocol for the coating with an amphiphilic polymer. Then, it was found that a mild oxidation process, carried out in water at 80 �C, promoted the conversion of the core-shell structure into fully maghemite nanocubes, enhancing their magnetic features, especially the specific absorption rate (SAR). Thus, the nanocubes were functionalized with PEG-FA and the annealing treatment was repeated.
Noteworthy, the oxidation strategy developed did not compromise the bio-functionality of the PEG-FA molecule. Unfortunately, the SAR values of the obtained one-phase nanocubes were found to be viscosity dependent, discouraging their use for magnetic hyperthermia in cellular environment. Instead, because annealing increased the magnetic moment of the nanocubes, they were efficiently used for the magnetic sorting of KB cells. The sorting efficiency found for these nanocubes was comparable to that of the MNBs reported in chapter 1, suggesting that a high amount of single nanocubes bound to the cell membrane increases the magnetic moment of the whole nanocubes-cell system. Thus, the methodology adopted for tuning the magnetic properties of core-shell iron oxide-based materials into one single phase NPs was proven appropriate for the preparation of nanocubes for magnetic driven cell sorting.
The third chapter discusses the use of maghemite (Fe2O3) nanocubes for developing multimodal nanotherapeutics to treat ovarian cancer. The intriguing high heat performance of the nanocubes was exploited to perform magnetic hyperthermia. At the same time, the high surface area available on the nanocubes has been used for drug delivery and specific antibody-mediated tumor targeting towards
ovarian cancer cells. The NCs were functionalized with both an oxaliplatin-derivatized PEG (PEG-Pt) and a PEG-Bis(carboxymethyl)-lysine (a nitrilotriacetic derived molecule) complex for the binding of an his-tag antibody fragment (scFv) specific for folate receptor ? (?FR). The functionalized nanocubes were able to recognize their target and to be efficiently internalized by the desired cells via endocytosis
pathway. Once inside the cells, the nanocubes delivered the Pt compound, which induced toxicity by intercalating the DNA. Thanks to their crystal structure and size, these nanocubes exhibited a viscosityindependent behavior, keeping high SAR values even in highly viscous media. Indeed, once incubated with the cells, the nanocubes were able to heat the tumor mass up to 42 �C, promoting cell death. The contribution of the cytotoxicity from both Pt delivery and hyperthermia highlighted the use of these nanocubes for cancer multitherapy. Thus, combining targeting, drug delivery and magnetic hyperthermia, a suitable platform for a synergistic treatment of cancer has been developed.