Nanoparticelle per la terapia dei tumori: preparazione, caratterizzazione e tossicità

The work of this doctoral thesis can be mainly divided in three parts. In the first part, the synthesis and characterization of different types of inorganic nanoparticles (NP) is described. The full protocol to prepare NP-GS, NP-Si)GS, and NP-Si)Au was found in literature. Stöber method-synthesized NP-Si were used as a platform to create NP-Si)GS, and NP-Si)Au. A novel method to conjugate antitumor drugs (s-Azl 6 and s-Azl 38) to Au-NP was also developed. Studies on Azl 6 and Azl 38 showed that they are highly hydrophobic, but when conjugated with Au-NPs, in the presence of an amphiphilic molecule (C), the complex (DNPC) passed through water. NP-Si, NP-GS, NP-Si)GS, and NP-Si)Au were characterized with electron microscopy, x-ray diffraction (EDS), and DLS. Although synthesis appeared to be easy to perform, results showed that NP-Si)Au did not form, while NP-GS and NP-Si)GS were obtained. NP-GS have a small diameter (2-3 nm on average), while for NP-Si and NP-Si)GS a diameter of 178-205,52 nm, and 164,80-178,66 nm respectively was calculated. TEM and SEM microscopy also indicated that NP-Si)Au corroded, and the gold envelope, that was supposed to coat the whole particule, was observed detached from the silica NP and in the form of small spheres. The reason why the synthesis of NP-Si)Au did not succeed, maybe relies on the particular type of Si-NP used. Further studies will deepen this aspect. The culture of three different types of human cell phenotypes is accurately described in the second part. For our experiments, we decided to cultivate human melanocytes, a human melanoma cell line, and human keratinocytes. Melanocytes appeared characterized by a dendritic-like morphology, and they showed a very slow growth rate. On the contrary, we observed that their corresponding tumor cells (melanoma) grew quickly and, although they were sold as adherent cells, floating living cells were also found in cultures. Finally, in human keratinocytes two morphotypes were observed. The first (the most diffused) had the typical “cobblestone shape” of keratinocytes, while the second appeared bigger than the first, flattened, and with a jagged contour. Finally, in the third part of this work, the toxicity of inorganic NP (i.e., NP-Si, NP-GS, and NP-Si)GS), DNPC, inorganic rhodaminated NP conjugated with heparin, and lipid NP containing paclitaxel (NC-Pacli), was tested on the cell phenotypes mentioned above. CellTiter Blue reagent was selected to quantify cell viability, and it was chosen among other dyes and assays for its non-toxicity and easiness of use. It was shown that among inorganic NP, only Si-NP have toxic effect, especially on healthy cells. NC-Pacli, on the contrary, caused the decrease of cell viability on healthy and tumor cells.