Desenvolvimento de um nanobot com propulsão biocatalítica e neutralizante das espécies reativas de oxigénio co cólon inflamado

Abstract

The development of micro and nanobots (NB) for biomedical applications has shown great potential as controlled drug release systems, presenting promising alternatives for minimally invasive and effective treatments. This thesis focuses on the development of a NB with bioactive properties, designed not only to act as a controlled drug release system, but also to neutralise reactive oxygen species (ROS), particularly in the context of intestinal inflammation, namely in the colon. Gastrointestinal diseases cover a wide range of clinical conditions, including inflammatory bowel diseases (IBD) and cancer. IBD affects around 3 million people in Europe, seriously jeopardising quality of life and generating high costs for health systems. Despite advances in medical therapy, the prevalence of IBD continues to rise, highlighting the need for new, more effective treatments. In this context, NBs appear to be a promising solution, since they can be programmed to administer drugs directly into the inflamed areas of the intestine and simultaneously reduce oxidative stress in situ, minimising the adverse effects of systemic treatments. This work describes the development of a NB based on the functionalisation of chitosan nanoparticles (NP) through biotin-avidin interaction, allowing the catalase enzyme (CAT) to be conjugated to the NP surface. CAT is responsible for the biocatalytic propulsion of NB. The chitosan NPs were characterised by scanning electron microscopy, revealing a regular spherical morphology with sizes approximately between 100 and 130 nm; these measurements were made on dehydrated particles. After functionalisation (conversion to NB), the NB movements were analysed in water (control) and in a hydrogen peroxide solution (as an ROS model) using fluorescence microscopy. The results demonstrated the promising potential of CAT-functionalized NB to respond efficiently to inflammatory environments, particularly in the presence of ROS such as H₂O₂. The significantly increased mobility of NB in H₂O₂-rich environments suggests that they can be programmed to move efficiently in areas of inflammation, using oxidative stress as fuel for biocatalytic propulsion. This behaviour highlights their potential as targeted drug delivery systems for chronic inflammatory gastrointestinal diseases such as IBD. In addition, the colloidal stability maintained, even after structural modification with biotin, avidin and CAT, ensures that NBs remain functionally viable for biological applications. Their dual ability to transport drugs and neutralise ROS reinforces their role as an innovative therapeutic tool, capable of reducing oxidative stress in areas of inflammation and improving treatment efficacy while minimising systemic side effects. These results are promising for the development of a drug delivery system within situ catalytic action, applicable in cases of inflammation in the colon. It is important to note that although this work is at an early stage, it has allowed us to explore the development of an innovative solution for the controlled release of drugs and the neutralisation of ROS in inflammatory environments, demonstrating both its sustainability and its innovative potential. However, further studies are needed to fully design a functional NB.

Date of Award	17 Dec 2024
Original language	Portuguese
Awarding Institution	Universidade Católica Portuguesa
Supervisor	Ezequiel Coscueta (Supervisor) & Maria Manuela Pintado (Co-Supervisor)