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Projeto de investigação
Biointeration studies of recombinant HPV-16 oncoproteins to discover therapeutic inhibitors
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Development and Characterization of Quercetin-Loaded Delivery Systems for Increasing Its Bioavailability in Cervical Cancer Cells
Publication . Ferreira, Miguel; Gomes, Diana; Neto, Miguel; Passarinha, Luís A.; Costa, Diana; Sousa, Ângela; UCIBIO - Applied Molecular Biosciences Unit; DQ - Departamento de Química
Quercetin is a natural flavonoid with high anticancer activity, especially for related-HPV cancers such as cervical cancer. However, quercetin exhibits a reduced aqueous solubility and stability, resulting in a low bioavailability that limits its therapeutic use. In this study, chitosan/sulfonyl-ether-β-cyclodextrin (SBE-β-CD)-conjugated delivery systems have been explored in order to increase quercetin loading capacity, carriage, solubility and consequently bioavailability in cervical cancer cells. SBE-β-CD/quercetin inclusion complexes were tested as well as chitosan/SBE-β-CD/quercetin-conjugated delivery systems, using two types of chitosan differing in molecular weight. Regarding characterization studies, HMW chitosan/SBE-β-CD/quercetin formulations have demonstrated the best results, which are obtaining nanoparticle sizes of 272.07 ± 2.87 nm, a polydispersity index (PdI) of 0.287 ± 0.011, a zeta potential of +38.0 ± 1.34 mV and an encapsulation efficiency of approximately 99.9%. In vitro release studies were also performed for 5 kDa chitosan formulations, indicating a quercetin release of 9.6% and 57.53% at pH 7.4 and 5.8, respectively. IC50 values on HeLa cells indicated an increased cytotoxic effect with HMW chitosan/SBE-β-CD/quercetin delivery systems (43.55 μM), suggesting a remarkable improvement of quercetin bioavailability.
Specific Six-Transmembrane Epithelial Antigen of the Prostate 1 Capture with Gellan Gum Microspheres
Publication . Batista-Silva, João; Gomes, Diana; Barroca-Ferreira, Jorge; Gallardo, Eugénia; Sousa, Ângela; Passarinha, Luís A.; UCIBIO - Applied Molecular Biosciences Unit; DQ - Departamento de Química; MDPI - Multidisciplinary Digital Publishing Institute
This work demonstrates the potential of calcium- and nickel-crosslinked Gellan Gum (GG) microspheres to capture the Six-Transmembrane Epithelial Antigen of the Prostate 1 (STEAP1) directly from complex Komagataella pastoris mini-bioreactor lysates in a batch method. Calcium-crosslinked microspheres were applied in an ionic exchange strategy, by manipulation of pH and ionic strength, whereas nickel-crosslinked microspheres were applied in an affinity strategy, mirroring a standard immobilized metal affinity chromatography. Both formulations presented small diameters, with appreciable crosslinker content, but calcium-crosslinked microspheres were far smoother. The most promising results were obtained for the ionic strategy, wherein calcium-crosslinked GG microspheres were able to completely bind 0.1% (v/v) DM solubilized STEAP1 in lysate samples (~7 mg/mL). The target protein was eluted in a complexed state at pH 11 with 500 mM NaCl in 10 mM Tris buffer, in a single step with minimal losses. Coupling the batch clarified sample with a co-immunoprecipitation polishing step yields a sample of monomeric STEAP1 with a high degree of purity. For the first time, we demonstrate the potential of a gellan batch method to function as a clarification and primary capture method towards STEAP1, a membrane protein, simplifying and reducing the costs of standard purification workflows.
In silico approaches
Publication . Gomes, Diana; Silvestre, Samuel; Duarte, Ana Paula; Venuti, Aldo; Soares, Christiane P.; Passarinha, Luís; Sousa, Ângela; UCIBIO - Applied Molecular Biosciences Unit; DCV - Departamento de Ciências da Vida; DQ - Departamento de Química; Molecular Diversity Preservation International (MDPI)
Cervical cancer (CC) is the fourth most common pathology in women worldwide and presents a high impact in developing countries due to limited financial resources as well as difficulties in monitoring and access to health services. Human papillomavirus (HPV) is the leading cause of CC, and despite the approval of prophylactic vaccines, there is no effective treatment for patients with pre-existing infections or HPV-induced carcinomas. High-risk (HR) HPV E6 and E7 oncoproteins are considered biomarkers in CC progression. Since the E6 structure was resolved, it has been one of the most studied targets to develop novel and specific therapeutics to treat/manage CC. Therefore, several small molecules (plant-derived or synthetic compounds) have been reported as blockers/inhibitors of E6 oncoprotein action, and computational-aided methods have been of high relevance in their discovery and development. In silico approaches have become a powerful tool for reducing the time and cost of the drug development process. Thus, this review will depict small molecules that are already being explored as HR HPV E6 protein blockers and in silico approaches to the design of novel therapeutics for managing CC. Besides, future perspectives in CC therapy will be briefly discussed.
Progress and opportunities in Gellan gum-based materials
Publication . Gomes, D.; Batista-Silva, J. P.; Sousa, A.; Passarinha, L. A.; UCIBIO - Applied Molecular Biosciences Unit; DQ - Departamento de Química; Elsevier
Gellan gum, a microbial exopolysaccharide, is biodegradable and has potential to fill several key roles in many fields from food to pharmacy, biomedicine and tissue engineering. In order to improve the physicochemical and biological properties of gellan gum, some researchers take advantage of numerous hydroxyl groups and the free carboxyl present in each repeating unit. As a result, design and development of gellan-based materials have advanced significantly. The goal of this review is to provide a summary of the most recent, high-quality research trends that have used gellan gum as a polymeric component in the design of numerous cutting-edge materials with applications in various fields.
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Fundação para a Ciência e a Tecnologia
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2020.06792.BD