mRNA delivery and safety evaluation of arginine-rich amphipathic cationic lipopeptides in vivo and in vitro
10.16438/j.0513-4870.2023-1443
- VernacularTitle:富含支化精氨酸的两亲性阳离子脂质肽体内外mRNA基因递送能力及安全性评价
- Author:
Yi-chun WANG
1
;
Yi-yao PU
1
;
Qun-jie BI
1
;
Xiang-rong SONG
2
;
Rong-rong JIN
1
;
Yu NIE
1
Author Information
1. National Engineering Research Centre for Biomaterials/College of Biomedical Engineering, Sichuan University, Chengdu 610064, China
2. State Key Laboratory of Biotherapy, Sichuan University, Chengdu 610041, China
- Publication Type:Research Article
- Keywords:
branched arginine;
amphiphilic cationic lipid;
gene vector;
mRNA delivery;
zebrafish
- From:
Acta Pharmaceutica Sinica
2024;59(4):1079-1086
- CountryChina
- Language:Chinese
-
Abstract:
mRNA gene therapy has attracted much attention due to its advantages such as scalability, modification, no need to enter the nucleus and no integration of host genes. In gene therapy, safe and effective delivery of mRNA into cells is critical for the success of gene therapy. In this study, we designed and synthesized an amphiphilic cationic lipopeptide gene vector (dendritic arginine & disulfide bond-containing cationic lipopeptide, RLS) enriched with branched arginine. We achieved a 1.5-fold higher mRNA transfection efficiency in zebrafish compared to the commercial reagent Lipofectamine 2000, and confirmed its good biosafety by in vitro cytotoxicity and in vivo biosafety. First, we characterized the chemical composition of the cationic lipid peptides by nuclear magnetic resonance hydrogen spectroscopy (1H NMR) and time-of-flight mass spectrometry (MS). The results of particle size and potential tested by dynamic light scattering particle size analysis showed that at a nitrogen/phosphorus (N/P) ratio of 20, the RLS/mRNA composite assemblies formed homogeneous nanoparticles with an average particle size of about 220 nm and a surface ζ potential of about +21 mV. In vitro gene transfection, the transfection experiments demonstrated that RLS exhibited 1.2-fold higher transfection efficiency in human embryonic kidney 293 cells (HEK293) and 3-fold higher transfection efficiency in rat mesenchymal stem cells (MSC) compared to Lipofectamine 2000. In addition, after microinjection of RLS into zebrafish embryos, we evaluated the survival, hatching, and teratogenicity rates, all of which confirmed its favorable in vivo safety profile. Thus, this amphiphilic cationic lipid peptide RLS, enriched with branched arginine, exhibits excellent mRNA delivery properties and safety. These findings highlight its potential as a promising gene therapy tool.
