Single pot organic solvent-free thermocycling technology for siRNA-ionizable LNPs: a proof-of-concept approach for alternative to microfluidics

Abstract

Ionizable LNPs are the latest trend in nucleic acid delivery. Microfluidics technology has recently gained interest owing to its rapid mixing, production of nucleic acid-ionizable LNPs, and stability of nucleic acid inside the body. Industrial scale-up, nucleic acid-lipid long-term storage instability, and high production costs prompted scientists to seek alternate solutions to replace microfluidic technology. We proposed a single-pot, organic solvent-free thermocycling technology to efficiently and economically overcome most of the limitations of microfluidic technology. New thermocycling technology needs optimization of process parameters such as sonication duration, cooling–heating cycle, number of thermal cycles, and lipid:aqueous phase ratio to formulate precisely sized particles, effective nucleic acid encapsulation, and better shelf-life stability. Our research led to the formulation of siRNA-ionizable LNPs with particle sizes of 104.2 ± 34.7 nm and PDI 0.111 ± 0.109, with 83.3 ± 4.1% siRNA encapsulation. Thermocycling siRNA-ionizable LNPs had comparable morphological structures with commercialized microfluidics ionizable LNPs imaged by TEM and cryo-TEM. When compared to microfluidics ionizable LNPs, thermocycling siRNA-ionizable LNPs had a longer shelf life at 4°C. Our thermocycling technology showed an effective alternative to microfluidics technology in the production of nucleic acid–ionizable LNPs to meet global demand.

WZB117 Decorated Metformin-Carboxymethyl Chitosan Nanoparticles for Targeting Breast Cancer Metabolism

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Folic Acid Functionalized Diallyl Trisulfide–Solid Lipid Nanoparticles for Targeting Triple Negative Breast Cancer

“…Methanol was used to extract the ultra-filtrate, which was then filtered through a 0.45 µm filter. UFLC [ 36 ] was used to analyze the supernatant. The mobile phase was acetonitrile and water (75:25, v / v ) with a flow rate of 1.1 mL/min, while the solid phase was a C18 (250 mm 4.6 mm) column.…”