Effect of pre‑freezing and saccharide types in freeze‑drying of siRNA lipoplexes on gene‑silencing effects in the cells by reverse transfection

Tang,Min; Hu,Subin; Hattori,Yoshiyuki

doi:10.3892/mmr.2020.11419

Effect of pre‑freezing and saccharide types in freeze‑drying of siRNA lipoplexes on gene‑silencing effects in the cells by reverse transfection

Authors:
- Min Tang
- Subin Hu
- Yoshiyuki Hattori
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Affiliations: Department of Molecular Pharmaceutics, Hoshi University, Shinagawa, Tokyo 142‑8501, Japan
Published online on: August 5, 2020 https://doi.org/10.3892/mmr.2020.11419
Pages: 3233-3244
Copyright: © Tang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Our previous study reported that reverse (Rev)‑transfection with small interfering RNA (siRNA)/cationic liposome complexes (siRNA lipoplexes) freeze‑dried in trehalose or sucrose solution resulted in high gene‑silencing activity in cells. The current study investigated whether pre‑freezing or saccharide types present during the freeze‑drying of siRNA lipoplexes affected gene‑silencing in cells after Rev‑transfection. Three types of cationic cholesterol derivatives and three types of dialkyl or trialkyl cationic lipids were used for the preparation of cationic liposomes. Additionally, six types of siRNA lipoplexes were vacuum‑dried in trehalose or sucrose solution without a pre-freezing process in multi‑well plates. A strong gene‑silencing activity after Rev‑transfection was observed regardless of the cationic lipid types in the cationic liposomes. It was also investigated whether saccharide types in the freeze‑drying of siRNA lipoplexes affected gene‑silencing after Rev‑transfection. siRNA lipoplexes freeze‑dried in monosaccharides (glucose, fructose, galactose or mannose), disaccharides (maltose, lactose, lactulose or cellobiose) and trisaccharide solution (raffinose or melezitose) demonstrated high gene‑silencing activity. However, following Rev‑transfection with siRNA lipoplexes freeze‑dried in monosaccharides or trisaccharides, certain saccharides induced cytotoxicity and/or off‑target effects. The results of the current study indicated that disaccharides may be suitable for the preparation of vacuum‑dried or freeze‑dried siRNA lipoplexes for Rev‑transfection.

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