Lipid-mRNA Nanoparticle Designed to Enhance Intracellular Delivery Mediated by Shock Waves

Abstract

Cellular membranes are, in general, impermeable to macromolecules (herein referred to as macrodrugs, e.g., recombinant protein, expression plasmids, or mRNA), which is a major barrier for clinical translation of macrodrug-based therapies. Encapsulation of macromolecules in lipid nanoparticles (LNPs) can protect the therapeutic agent during transport through the body and facilitate the intracellular delivery via a fusion-based pathway. Furthermore, designing LNPs responsive to stimuli can make their delivery more localized, thus limiting the side effects. However, the principles and criteria for designing such nanoparticles remain unclear. We show that the thermodynamic state of the lipid membrane of the nanoparticle is a key design principle for acoustically responsive fusogenic nanoparticles. We have optimized a cationic LNP (designated LNPLH) with two different phase transitions near physiological conditions for delivering mRNA. A bicistronic mRNA encoding a single domain intracellular antibody fragment and green fluorescent protein (GFP) was introduced into a range of human cancer cell types using LNPLH, and the protein expression was measured via fluorescence corresponding to the GFP expression. The LNPLH/mRNA complex demonstrated low toxicity and high delivery, which was significantly enhanced when the transfection occurred in the presence of acoustic shock waves. The results suggest that the thermodynamic state of LNPs provides an important criterion for stimulus responsive fusogenic nanoparticles to deliver macrodrugs to the inside of cells.

Realizing Cancer Precision Medicine by Integrating Systems Biology and Nanomaterial Engineering

“…DNA in the form of a circular plasmid or messenger RNA (mRNA), which are longer than RNAi molecules, can also be loaded into nanocarriers to effectively express a protein in a target cell. [73][74][75][76] Moffett et al and Smith et al each developed a nanocarrier that specifically targets the T cells of patients and delivers mRNA or a DNA plasmid into the T cells to transform them into chimeric antigen receptor (CAR) T cells in situ or ex vivo. [77,78] In this nanocarrier, mRNA or a DNA plasmid encoding the CAR is condensed by incubation with poly(beta-amino ester) (PBAE) 447 polymer and a polyglutamic acid-antibody recognizing cluster of differentiation 3 (CD3), a protein found on the surface of T cells.…”

Interaction Kinetics of Peptide Lipids-Mediated Gene Delivery

“…Confocal laser scanning microscopy (CLSM) and TEM were used to investigate the intracellular transport of lipoplexes [20,21]. Hoechst 33342, NBD-PE and Cy5-DNA were used here in order to detect the intracellular distribution of liposome/DNA lipoplexes.…”

Thermodynamic state of the interface during acoustic cavitation in lipid suspensions

“…Cavitation is the formation of vapor cavities in a liquid due to a tensile stress (1) and is assumed to be one of the key mechanisms for biophysical effects of ultrasound and in particular shock waves (2,3). It has been suggested as a possible mechanism both in transmembrane and intracellular drug delivery (4) as well as acoustic excitation of the peripheral nervous system (5). The key biophysical system in both these cases is the cellular membrane, which is essentially a self-assembled macromolecular bilayer mainly composed of lipids.…”