SYNTHESIS OF SMART GLYCOSIDES TO ENHANCE GLYCO-NANOMATERIALS CIRCULATION HALF-TIME

Nanoparticle (NP)-based therapies have proven to offer potential solutions for conventional medicine while minimizing its side effects. In particular, glycosylated gold NPs (AuGNPs) represent attractive imaging and/or delivery platforms combining the biological activity of carbohydrates with the biocompatibility and physicochemical versatility of the metallic core. However and despite their rapid growth, the full clinical potential of these glyconanosystems will only be reached when their underlying safety concerns are understood and accurately addressed. When placed in biological fluids, AuGNPs, interact with a high number of biomolecules forming a so-called protein corona (PC), a coating that covers them and has severe consequences for their fate, efficacy and toxicity. As a consequence, AuGNPs are often recognized by the immune system and cleared from the blood stream before they have had enough circulation time to reach their therapeutic target. In order to address the first issue, the initial chapter of the present thesis deals with the preparation of a library of six different AuGNPs functionalized with three monosaccharides (α-mannose, α-galactose and α-fucose) and two polyethylene glycol alternatives (PEG5000 and alkyl-PEG600) as an anchor to the Au nanomaterials (NMs). In collaboration with CICBiomaGUNE (Spain), the interaction of these AuGNPs with different lectins is investigated using Fluorescence Correlation Spectroscopy (FCS) to evaluate the effect of the PEG length on the interaction with proteins and the carbohydrates binding specificity. The second part of the chapter is dedicated to the preparation of an additional set of AuGNPs functionalized with negatively charged monosaccharides such as mannose-6-phosphate, mannose-6-sulfate or N-acetyl neuraminic acid. The final goal is to compare the results with neutral mannosylated AuGNPs in terms of colloidal stability and protein binding. These measurements will be performed at RCSI (Ireland) by means of Differential Centrifugal Sedimentation (DCS) and Dynamic Light Scattering (DLS). The final aim of the work described in Chapter I is to provide a better understanding of the PC formation and a correlation to the main NMs’ surface characteristics influencing it. The immunosuppressive properties of AuGNPs functionalized with N-acetyl neuraminic acid will also be tested in vivo in collaboration with Mario Negri Institute (Italy). The second chapter focuses on the synthesis of ABO blood sugar epitopes, terminal di- (Fucα1-2Galβ1-O-R) and tri-saccharides (GalNAcα1-3(Fucα1-2)Galβ1-O-R and Galα1-3(Fucα1-2)Galβ1-O-R) covering the surface of human red blood cells and other body tissues. Our synthetic derivatives have been prepared adapting previously reported procedures and include a linear aglycone with a free amino group for NM conjugation. Functionalization of NMs with these self-antigens may improve their immunotolerance and biocompatibility in comparison to PEGylated counterparts, prolonging their blood circulation time, improving their delivery efficiency and granting them new therapeutic possibilities. These assumptions are to be tested in the near future in collaboration with RCSI (Ireland). Chapter III describes the functionalization of mannose (Manα1), dimannose (Manα1-2Manα1) and trimannose (Manα1-3(Manα1-6)Manα1) derivatives with an amine-ending linker at their reducing-end for Au nanorods functionalization at VITO (Belgium). The end goal of the project is to develop a simple and label-free biosensor to detect mannose binding lectin (MBL) from human plasma samples based on optical measurements. The near IR shift due to the effective lectin binding can be correlated to potential cardiovascular disease for prevention and early onset stroke diagnosis. In summary, this thesis aims to provide a better understanding of AuGNPs’ behavior in biological fluids, to face the main concerns impairing nanomedicine translation into clinics and to pave the way towards novel glyco-nanosystems with enhanced therapeutic performance for healthcare applications.