(452g) Invited Speaker: Lipid Nanoparticle Formulations for the Synergistic Co-Delivery of siRNA and mRNA

Despite the promise of RNA therapeutics, progress towards the clinic has been slowed by the difficulty of delivering RNA drugs, such as short interfering RNA (siRNA) and messenger RNA (mRNA), into cellular targets within the body. RNA therapeutics are large (10⁴ – 10⁶ g/mol) and negatively charged; they do not have favorable biodistribution properties in vivo nor an ability to cross the cellular membrane of target cells. In response to these challenges, our lab has created biodegradable, ionizable lipid-like materials called ‘lipidoids’ that readily formulate into nanoparticles containing RNA. Lipidoids efficiently manipulate gene expression in a variety of biological systems, including hepatocytes, white blood cells and tumors.

This talk will focus on the simultaneous delivery of siRNA and mRNA. To facilitate the treatment of diseases associated with aberrant gene upregulation and downregulation, we sought to co-formulate siRNA and mRNA in a single lipidoid nanoparticle (LNP) formulation. We accommodated the distinct molecular characteristics of mRNA and siRNA in a formulation consisting of an ionizable biodegradable lipidoid, cholesterol, DSPC, DOPE, and PEG-lipid. Surprisingly, the co-formulation of siRNA and mRNA in the same LNP enhanced the efficacy of both drugs in vitro and in vivo. Compared to LNPs encapsulating siRNA only, co-formulated LNPs improved Factor VII gene silencing in mice from 44 to 87% at an siRNA dose of 0.03 mg/kg. Co-formulation also improved mRNA delivery, as a 0.5 mg/kg dose of mRNA co-formulated with siRNA induced three times the luciferase protein expression compared to when siRNA was not included.

As not all gene therapy applications require both RNA drugs, we sought to extend the synergy of co-formulated LNPs to formulations encapsulating only a single type of RNA. We accomplished this by substituting the “helper” RNA with a negatively charged polymer. LNPs containing negatively charged polymer mediated the same level of protein silencing or expression as standard LNPs using 2-3 fold less RNA. For example, LNPs formulated with and without polymer induced 50% Factor VII silencing at siRNA doses of 0.01 and 0.03 mg/kg, respectively. Together, these studies demonstrate potent co-delivery of siRNA and mRNA and show that inclusion of a negatively charged “helper polymer” enhances the efficacy of LNP delivery systems.