Polymeric nanoparticles for local delivery of multicolor genetic circuits in breast cancer

• Autor(a) principal: Amorim, Joana Ramos Nunes de
• Data de Publicação: 2022
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10362/146084
• topic: breast cancer; polymeric nanoparticles; multicolor genetic circuits; oncobiology; Ciências Médicas

Breast cancer is a very heterogenous disease that affects millions each year. Although it is the most prevalent cancer type, an optimal treatment option is still far from conceived with the current options leading to a wide range of side effects. The development of molecular biology in the last years, has made obvious that new therapeutic options will rely on the modulation of nucleic acids. At the forefront of the current options are micro-RNAs (miRNAs) which are small non-coding molecules responsible for post-translational regulation of gene expression. However, the delivery of nucleic acids poses a big challenge currently as these molecules are particularly sensitive, losing their characteristics quite easily when free. This project aimed to create a system based on polymeric nanoparticles to deliver plasmid constructs to the tumour microenvironment (TME). These constructs were designed with resource to the analysis of patient samples through miRNA arrays, to discover down-regulated miRNAs specific for each tumour type. The designed plasmid constructs included a cell specific promoter as well as the sequence encoding for the micro-RNA of interest (miR125b) and a fluorescent tag. The fluorescent tag would allow for the mapping of the TME and act to monitor the therapeutic progress. To achieve the goal several steps had to be taken starting with the attempt to encapsulate the plasmid, which proved to be a difficult process and, in conjunction with poor outcomes in cellular viability, lead to a change in the path. To tackle the challenges faced, in this work, two approaches were tested: 1) the molecular modification of the polymer by inserting amines and 2) the molecular modification of the surfactant D-alpha-Tocopherol polyethylene glycol 1000 succinate (TPGS) by binding it to spermidine. The later ultimately lead to the particles having a positive surface charge that allowed for the complexation with the plasmid DNA and a decrease in cell toxicity. Although several critical aspects were achieved, further development is still needed to tackle new challenges that arose and that this project didn’t manage to solve. Of these the most important is to understand whether the particles are internalised by the cells and if they can release the plasmid DNA for translation.