Lipid-like materials for low-dose, in vivo gene silencing

Abstract

Significant effort has been applied to discover and develop vehicles which can guide small interfering RNAs (siRNA) through the many barriers guarding the interior of target cells. While studies have demonstrated the potential of gene silencing in vivo, improvements in delivery efficacy are required to fulfill the broadest potential of RNA interference therapeutics. Through the combinatorial synthesis and screening of a different class of materials, a formulation has been identified that enables siRNA-directed liver gene silencing in mice at doses below 0.01 mg/kg. This formulation was also shown to specifically inhibit expression of five hepatic genes simultaneously, after a single injection. The potential of this formulation was further validated in nonhuman primates, where high levels of knockdown of the clinically relevant gene transthyretin was observed at doses as low as 0.03 mg/kg. To our knowledge, this formulation facilitates gene silencing at orders-of-magnitude lower doses than required by any previously described siRNA liver delivery system.

Multiparametric approach for the evaluation of lipid nanoparticles for siRNA delivery

“…To generate meaningful relationships between structure, biological function, and biological activity, we created a diverse set of lipids for formulation with siRNAs into LNPs. Recent high-throughput screens of lipidlike materials indicate that structures with multiple short alkyl tails stemming from a polyamine core are able to facilitate efficient gene silencing (22,23). To synthesize pure multitailed structures with varying polyamine head-group architectures, we designed a rapid and efficient two-step solvent-free synthetic route that requires neither protection/deprotection steps nor the use of costly and time-consuming chromatographic purification.…”

“…To address some of these challenges, single-parameter studies that evaluate the effect of chemical structure on a single biological property or on delivery performance have been carried out (16)(17)(18)(19)(20)(21). Furthermore, high-throughput synthetic methods have been exploited for the accelerated discovery of potent lipid nanoparticles (LNP) and evaluation of structure activity relationships (SAR) (22,23). Despite these efforts, the relationships between physicochemical properties of nanoparticles and biological barriers, and that between biological barriers and genesilencing activity remain unclear.…”

“…LNPs were used in this study because they have been shown to facilitate efficient delivery to hepatic and immune targets (23,(25)(26)(27). In the past few years, LNPs formulated with siRNAs against transthyretin (TTR) and proprotein convertase subtilisin/kexin type 9 (PCSK9) in rodents and nonhuman primates have produced promising preclinical results and several clinical trials involving LNPs are currently underway (28)(29)(30).…”

Human macrophage and dendritic cell-specific silencing of high-mobility group protein B1 ameliorates sepsis in a humanized mouse model

“…An ionizable cationic lipid called DLinKC2DMA, which was originally shown to deliver siRNA to liver cells (26), was also reported to deliver siRNA to mouse macrophages and DCs (27). Similarly, a lipidoid nanoparticle called C12-200, also developed originally for liver-specific siRNA delivery (28), was recently shown to target monocytes, and silencing CCR2 with this reagent was shown to reduce a number of inflammatory conditions in mice (18). siRNAs synthetically linked to C-phosphate-G (CpG) oligonucleotide has also been used to deliver siRNA to toll-like receptor (TLR)-9 + murine macrophages and B cells (29).…”

“Programmed packaging” for gene delivery

“…Because of these virtues associated with pH-sensitive carriers, several groups have reported on some original pH sensitive systems [29][30][31][32]. To date, one of the most effective systems is lipid nano particles (LNPs), which consist of a hydrophilic moiety containing a tertiary amino group, a linker and two hydrophobic dialkene groups [33].…”