Nanoparticles containing constrained phospholipids deliver mRNA to liver immune cells in vivo without targeting ligands

Abstract

Once inside the cytoplasm of a cell, mRNA can be used to treat disease by upregulating the expression of any gene. Lipid nanoparticles (LNPs) can deliver mRNA to hepatocytes in humans, yet systemic non-hepatocyte delivery at clinical doses remains difficult. We noted that LNPs have historically been formulated with phospholipids containing unconstrained alkyl tails. Based on evidence that constrained adamantyl groups have unique properties that can improve small molecule drug delivery, we hypothesized that a phospholipid containing an adamantyl group would facilitate mRNA delivery in vivo. We quantified how 109 LNPs containing "constrained phospholipids" delivered mRNA to 16 cell types in mice, then using a DNA barcoding-based analytical pipeline, related phospholipid structure to in vivo delivery. By analyzing delivery mediated by constrained phospholipids, we identified a novel LNP that delivers mRNA to immune cells at 0.5 mg/kg. Unlike many previous LNPs, these (a) did not preferentially target hepatocytes and (b) delivered mRNA to immune cells without targeting ligands. These data suggest constrained phospholipids may be useful LNP components.

Lipid nanoparticles for mRNA delivery

“…Using DNA barcode-labelled oligonucleotides, the distribution of different lipid nanoparticle formulations can be analysed in a high-throughput manner in vivo 121 , for example, to quantify targeted delivery of nucleic acids in multiple tissues 121 . Based on this method, a series of phosphatidylcholines containing constrained adamantyl groups has been explored for mRNA delivery, including analysis of distribution in different cell types 122 .…”

Non-Immunotherapy Application of LNP-mRNA: Maximizing Efficacy and Safety

“…The importance of LNP composition optimization in screening carried out by Sago et al resulted in finding 7C2 and 7C3 LNPs that efficiently target endothelial cells, as previously noted [ 147 ]. Using the same screening methodology as Sago et al, Gan et al recently tested a library of 109 LNPs composed of “constrained phospholipids” that contained an adamantylhydrocarbon chain [ 154 ]. This study identified a novel LNP that delivers mRNA to Kupffer cells instead of hepatocytes, without targeting ligands [ 154 ].…”

“…Using the same screening methodology as Sago et al, Gan et al recently tested a library of 109 LNPs composed of “constrained phospholipids” that contained an adamantylhydrocarbon chain [ 154 ]. This study identified a novel LNP that delivers mRNA to Kupffer cells instead of hepatocytes, without targeting ligands [ 154 ]. Recently, Zukancic et al used Onpattro ® LNP, where the authors replaced 1, 2-Distearoyl-sn-glycero-3-phosphoethanolamine-Poly(ethylene glycol) (PEG-DSPE) with Tween 20 containing short (C11) PEG alkyl chain [ 155 ].…”

Non-Viral Targeted Nucleic Acid Delivery: Apply Sequences for Optimization

“…By applying a siGFP/DNA-barcoded system in transgenic eGFP mice, the group found that constrained lipid nanoparticles (cLNPs) containing a conformationally constrained adamantane tail can deliver siRNA and sgRNA to splenic T cells in vivo at low doses [ 197 ], and in contrast to standard LNPs do not preferentially target hepatocytes. Based on this, the group further demonstrated cLNP containing adamantyl phospholipids can delivery Cre mRNA to liver immune cells at low doses [ 198 ]. Like reported before [ 188 ], delivery takes place in a size-independent and chemical composition-dependent manner, providing a potential alternative for further active targeting based on chemical modification and not targeting ligands.…”