Extracellular Vesicles as a platform with potential clinical and industrial applications: from cancer diagnosis to cell-free therapy.

Extracellular Vesicles (EVs) are released by all cell types and have a role in cell-to-cell communication, in both physiological and pathological conditions. EVs are exploited in industry since they are a platform with complementary clinical applications. In fact, they carry biomolecules that can be detected in many biofluids and exploited for diagnostic purposes. Moreover, EVs by themselves (like EVs from menchymal stem cells), or engineered for drug delivery, can be used as therapeutic agents for many diseases. Hypoxia, which can be a physiological condition or a pathological stress, may affect the release of vesicles and their content. The first part of this thesis is focused on the effects of hypoxia on the release of EVs in melanoma. It has been demonstrated that the expression of a promising marker of tumor hypoxia, Carbonic Anhydrase IX (CA-IX), is associated to hypoxia-induced EVs. CA-IX protein has a role in maintaining a favorable intracellular pH for tumor cells, thus increasing their survival and growth. CA-IX-carrying EVs have been found also in plasma of metastatic melanoma patients, suggesting their potential application as biomarkers for liquid biopsy. The second part of this thesis is focused on the characterization of Mesenchymal Stem Cells (MSC)-derived EVs, purified with different methods enabling to provide different grades of purity of final EVs. MSC-EVs have been tested in vitro to investigate their biological properties related to regeneration, anti-inflammatory and/or tumor suppression. In literature, it has been reported that an in vitro hypoxic environment can ameliorate the proliferation of MSCs, and that MSC-EVs generated under hypoxia have improved angiogenic and anti-inflammatory effects. It could be useful, in the future, investigate if an hypoxic treatment can further enhance the functional properties of MSC-EVs that we observed.