CA3094859A1 — Proteolipid vesicles formulated with fusion associated small transmembrane proteins

USPTO Abstract

Lipid nanoparticle (LNP) delivery platforms have undergone incremental improvements, but still suffer from cellular toxicity and a biodistribution profile limited to hepatic cells. We developed an all-encompassing proteolipid nucleic acid delivery vehicle formulated with a chimeric FAST protein, called Fusogenix. Using this approach, we demonstrate that ionizable lipids can be utilized at a minimal molar ratio for the sole purpose of neutralizing the anionic charge of nucleic acids, rather than facilitating endosomal escape. Incorporation of this chimeric FAST protein into a PLV platform enhances intracellular delivery and expression of mRNA and pDNA both in vitro and in vivo. These PLVs also display a favorable immune profile and are significantly less toxic than conventional LNPs. Gene therapy holds great promise to treat a wide array of diseases but has struggled to gain widespread regulatory approval primarily due to the lack of a safe and effective nucleic acid delivery platform that is non-immunogenic, easy to manufacture, and able to achieve systemic biodistribution. To overcome the limitations surrounding viral and non-viral vectors we combined the positive aspects of both platforms to develop a proteolipid vehicle (PLV) that can achieve efficient nucleic acid gene expression with high tolerability. We developed a library of chimeric fusion-associated small transmembrane (FAST) proteins and identified the FAST protein with a superior ability to facilitate plasma membrane fusion. Inclusion of the chimeric FAST protein into the PLV platform enables a minimal molar ratio of ionizable lipids to be utilized for the sole purpose of neutralizing the anionic charge of nucleic acids. Expression of both messenger RNA (mRNA) and plasmid DNA (pDNA) is improved following FAST incorporation. Systemically administered FAST-PLVs displayed an extensive biodistribution to all organs tested, with pDNA and mRNA inducing robust gene expression in the lungs, liver, spleen, and kidney. FAST-PLVs display a favorable immune profile and can be repeatedly dosed without losing activity. The increased safety of FAST-PLVs, even at high doses allows for comparable nucleic acid expression to conventional lipid nanoparticles with substantially less toxicity, making them an excellent clinical candidate for gene therapy approaches.