Study of innovative therapies for multiple sclerosis

In recent years, nanotechnologies have acquired an increasingly important role in the development of innovative therapeutic strategies, particularly in the field of drug delivery. The interaction between nanomaterials and the immune system is an area of great interest, as it allows not only to improve the efficacy of drugs, but also to selectively modulate immune responses. In this context, the main objective of this doctoral thesis was the development and characterisation of biocompatible gold and silver nanoparticles capable of selectively interacting with specific cells of the immune system and being exploited as delivery systems for drugs with immunomodulatory and anti-inflammatory activity. The work focused on the synthesis of gold and silver nanoparticles stabilised with different surface ligands such as citrate, L-cysteine and polyethylene glycol (PEG), chosen for their known colloidal stabilisation properties and biocompatibility. The first phase of the study was dedicated to optimising the synthesis protocols in order to obtain reproducible nanoparticle systems that are stable over time and characterised by controlled chemical-physical properties. The optimisation of the experimental conditions made it possible to modulate key parameters such as size, size distribution, aggregation state and colloidal stability, which are fundamental aspects for subsequent biological use. The chemical-physical characterisation of the nanoparticles was conducted using an integrated approach that included spectroscopic techniques, size and polydispersity analysis, colloidal stability assessments, and surface interaction studies. Systematic comparison of the different formulations showed that the nature of the stabilising ligand significantly influences the properties of the nanoparticles, determining not only their physical characteristics but also their ability to interact with functionalising molecules. The overall analysis of the data allowed the selection of the most promising formulations, in particular gold nanoparticles stabilised with citrate and L-cysteine (Au-CitCys), for subsequent biological evaluations. The selected nanoparticles underwent biocompatibility studies, which demonstrated an adequate safety profile under the experimental conditions adopted. These results allowed for further study of the interactions between nanoparticles and human immune system cells, using peripheral blood mononuclear cells (PBMCs) as an experimental model. The functionalisation of nanoparticles with fluorescent probes was a fundamental tool for monitoring their association with cells and identifying the cell subpopulations most involved in the interaction. The results obtained showed that nanoparticles have an intrinsic ability to interact selectively with specific cells of the immune system. This evidence provided the rationale for the development of a “Trojan horse” drug delivery strategy, in which nanoparticles are used as vectors to facilitate the transport and action of an immunomodulatory and anti-inflammatory drug within the immune system. The functionalisation of the drug on nanoparticles has made it possible to evaluate the efficacy of the delivery system compared to the free drug. The biological data collected show that the drug delivered by nanoparticles exhibits significantly enhanced activity in modulating the immune response. In particular, the nanoparticle system is able to steer the response towards a more tolerogenic and anti-inflammatory profile, a therapeutic goal of primary importance in the treatment of diseases characterised by chronic inflammation and immune dysregulation, such as multiple sclerosis. Overall, this work demonstrates that the nanoparticles developed are not only biocompatible and capable of selectively interacting with immune system cells, but can also be effectively exploited as drug delivery systems to improve the efficacy of immunomodulatory drugs. In particular, Au-CitCys nanoparticles emerge as promising platforms for future applications in the field of immunological nanomedicine. The results obtained provide a solid basis for further preclinical studies and open up new perspectives for the development of innovative therapeutic strategies based on nanomaterials, aimed at the targeted treatment of inflammatory and immune-mediated diseases.