Repository logo

Browsing by Issue Date, starting with "2022-11-11"

Filter results by year or month
Now showing 1 - 2 of 2
  • Preparation and characterization of anionic PPI dendrimers for delivery of the hydroxychloroquine into tumor cells
    Publication . Rodrigues, Ana Filipa Gonçalves; Rodrigues, João Manuel Cunha; Maciel, Dina Maria Sousa
    Cancer is one of the most lethal diseases worldwide. In 2020, 19.3 million new cancer cases were reported, with approximately 10 million deaths. Female breast cancer was the most diagnosed, with 2.3 million new cases. Hydroxychloroquine (HCQ) is an immunomodulatory drug used to treat malaria and has been shown to have therapeutic potential in oncology. It has been proven that HCQ increases tumor cell death alone or combined with targeted agents or cytotoxic chemotherapy. By combining HCQ and a nanocarrier, it is possible to direct the drug to the tumor tissues, reducing the side effects and enhancing the bioavailability. Based on the previous experience of our group, the main goal of this master thesis was to prepare anionic carboxylate and sulfonate PPI dendrimers to transport and deliver the HCQ into tumor cells. First, amine-terminated PPI dendrimers, generation 0 to 3 (G0-G3), were synthesized, with a yield in the range of 57% -99%, and characterized by 1H- and 13C- NMR. After the G0 to G3 amine-terminated PPI dendrimers were prepared and obtained, the functionalization of the terminal groups with carboxylate and sulfonate dendrimers was done. Additionally, for the first time, the carboxylate- and sulfonate-terminated dendrimers G4 was synthesized. The anionic dendrimers have the advantage of being less cytotoxic, hemotoxic, and immunogenicity than cationic dendrimers. After G1 to G3 characterization by 1H 13C NMR, ATR-FTIR, DLS, zeta potential, and MS, HCQ was encapsulated. Following this, the in vitro drug release was performed in PBS with a pH 5 and 7.4, since the cumulative release is slightly higher in PBS with a pH 5. In general, all dendrimers showed a sustained release of HCQ in both pHs. The hemotoxicity of the dendrimers and the HCQ were also evaluated and showed no hemotoxicity. Finally, the cytotoxicity of the synthesized dendrimers, non-loaded and loaded with HCQ, was evaluated in a cancer cell line (MCF-7) and a non-cancer cell line (BJ cells). The HCQ in a range of concentration between 0.5 and 50 µM are highly cytotoxic for BJ cells and less cytotoxic for MCF-7 cells. The HCQ was further tested with the CACO-2 cells and showed to be less sensitive to the drug than with the MCF-7 cells. In order to improve the cytotoxicity of the prepared dendrimers, an assay with doxorubicin (DOX) was performed, but no synergetic effect was observed.
    2022-11-11Master thesis Open access Show more
  • MALDI-TOF MS-based urinary proteomic/peptidomic signature of breast cancer as innovative diagnostic approach
    Publication . Sousa, Patrícia Maria Gonçalves; Gouveia, Rosa Maria de Sá Perestrelo; Câmara, José de Sousa
    The development of a rapid and high-sensitive breast cancer (BC) diagnostic method has been increasingly investigated by many researchers, to have a more effective therapy, through the early detection of this disease that affects millions of women worldwide. Therefore, the discovery of specific biomarkers is one of the topics in clinical medicine, however several challenges remain in the development of optimal sample preparation for proteomic/peptidomic analysis of urine, due to its highly variable content, as well as the presence of various proteins in low abundance or modified forms. Thus, this investigation aims to establish potential BC urinary protein biomarkers using one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (1D SDS-PAGE) coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometric (MALDI-TOF MS) for BC diagnosis and monitoring the efficiency of therapy. The results of the Lowry´s assay demonstrated that total protein concentration increased after precipitation and that healthy control group (HCs) have higher total protein concentrations than BC patients. Related to MALDI-TOF MS analysis, results revealed that four protein/peptide ion signatures (m/z 1046.5, 1062.4, 1237.7 and 1727.9), with variable importance in projection (VIP) > 1, allowed the discrimination between BC patients and HCs. The discrimination efficiency and accuracy of BC urine proteins/peptides were ascertained by receiver operating characteristic (ROC) curve analysis that allowed the identification of some features with high sensitivity and specificity (99.6%). Therefore, the obtained data revealed MALDI-TOF MS as a powerful tool to explore proteomic/peptidomic biosignatures, due to its speed, sensitivity, and mass accuracy, which allow establishing novel disease biomarkers. Nevertheless, a deep study using a higher number of samples must be carried out to confirm and consolidate the data obtained.
    2022-11-11Master thesis Open access Show more