Doctoral thesis

Rare diseases (RD), predominantly caused by genetic mutations, affect millions of people worldwide, although each individual disease impacts only a small population. Addressing such a challenge requires a standardized yet customizable strategy, and nucleic acid-based therapies offer a transformative approach. These therapies target specific DNA or RNA sequences and have demonstrated great potential as treatment for orphan diseases - those lacking therapeutic options, as is the case with many RD. Recent clinical advances with remarkable results validate this approach. However, significant challenges remain, such as efficient delivery, stability, and minimization of adverse effects. Overcoming these obstacles is crucial for developing pharmaceutical technologies accessible to a broad spectrum of rare diseases.

Natural oligonucleotides face significant limitations; they are prone to degradation and exhibit low cellular uptake. To overcome these issues, various chemical modifications have been implemented with a wide range of functional benefits. These modifications have already been used in a number of pharmaceuticals approved by the regulatory agencies, due to their ability to increase nuclease resistance, enhance binding affinity, and improve thermal stability. Understanding the structure of these modifications is fundamental to determining their impact on the therapeutic properties of oligonucleotides and, consequently, to designing better therapeutic agents.

The main objective of this project is to characterize the structural determinants that influence the efficiency of therapeutic oligonucleotides in gene silencing and gene expression recovery therapies. Specifically, this project focuses on a group of rare diseases whose pathophysiology is rooted in the formation of a noncanonical structure in the gene responsible for the condition. This will be achieved through a combination of structural studies using liquid and solid-state nuclear magnetic resonance (NMR) spectroscopy, followed by an in-depth analysis of the effects of chemical modifications on oligonucleotides. In the same manner, interactions between modified oligonucleotides and their noncanonical RNA or DNA targets will be evaluated. The knowledge generated will be used to design and synthesize new oligonucleotides with improved pharmaceutical properties. The generated NMR data will be applied to model and predict the behavior of modified oligonucleotides in complex biological environments, contributing to the development of the next generation of nucleic acid-based therapies for rare diseases, tailored and optimized for each patient.