Asialofetuin-Coupled Lipid-Based nanosystems to target the Asialoglycoprotein receptor: Delivering genes to hepatocytes for the treatment of Fabry disease

Exploiting the protein production capacity of hepatocytes for de novo expression of α-Galactosidase A (α-Gal A) by gene supplementation therapy represents one of the most promising strategies for the treatment of Fabry disease (FD). The asialoglycoprotein receptor (ASGPr) has proven to be one of the target receptors of choice for hepatocyte-directed nanomedicines, and natural glycoproteins such as asialofetuin (AF) can be used as specific ligands. Herein, we have developed AF-decorated solid lipid nanoparticles (SLNs), prepared by different techniques and cationic lipid compositions, for restoring the enzyme deficiency in FD by gene supplementation targeted to hepatocytes. After the physicochemical characterization of the vectors, cell association and transfection efficacy were evaluated in vitro in human hepatocytes (Hep G2), and the capacity to increase α-Gal A activity was evaluated in vivo after intravenous administration to α-Gal A knockout mice. The efficacy and targeting effect were conditioned by the type of SLN. In general, vectors containing a mixture of the cationic lipids DOTAP and DODAP showed enhanced transfection efficacy compared to their counterparts without DODAP. The incorporation of AF in the vectors formulated with SLNs prepared with DOTAP and DODAP by hot-melt emulsification significantly improved the efficacy to induce the expression of α-Gal A in hepatocytes in vitro compared to the control without AF. However, the administration to Fabry mice did not result in a significant increase in enzyme activity. The lack of in vitro-in vivo correlation corroborates the need to understand key factors influencing the behavior of non-viral vectors in biological media for nucleic acid therapies, as well as the desirability of in vivo studies in the early stages of pharmaceutical development of nucleic acid delivery systems.