Study of the Endosomal Distribution of Polyplexes and Lipid Nanoparticles and their in-vitro Dissociation for Improved Gene Delivery

Nucleic acid-based therapies are currently booming with the advent of mRNA vaccines formulation. Hence, gene delivery systems employing polyplexes or lipid nanoparticles (LNPs) as versatile carriers have shown great promise for therapeutic applications due to the advantages in terms of tuneable properties and biocompatibility. Key to the success of these therapies is the development of efficient and controlled delivery systems capable of transporting the genetic material into target cells, releasing it intracellularly, and minimizing cytotoxicity. However, the efficient release of genetic material from its carriers within the endosomal compartments of target cells remains a critical challenge. This project will investigate the endosomal distribution and in-vitro dissociation mechanisms of polyplexes and LNPs, aiming to provide an outstanding insight on the structure/activity relation of nucleic acids careers. Employing a combination of state-of-the-art microscopy and spectroscopy techniques such as Förster Resonance Energy Transfer (FRET), live-cell imaging, and physicochemical approaches, this research will uncover vital insights into the intracellular dynamics of these gene delivery carriers for the better understanding of endosomal escape and particles dissociation. Furthermore, we will explore the relations between pH distribution of endosomes at various maturation levels and the corresponding accumulation of particles. This research contributes valuable insights into the intracellular fate of gene delivery carriers and provides strategies to overcome endosomal entrapment. Improved understanding of endosomal distribution and in-vitro dissociation mechanisms will ultimately facilitate the development of more efficient and targeted gene delivery systems for therapeutic applications. This will bridge the gap between laboratory research and clinical applications for a diverse spectrum of diseases.