Preparation and in vitro evaluation of redox-sensitive polypeptide vector for co-delivery of gene and chemotherapeutic drugs
10.16781/j.0258-879x.2018.02.0144
- Author:
Chun-Ai GONG
1
Author Information
1. Department of Pharmacy, Changhai Hospital, Second Military Medical University
- Publication Type:Journal Article
- Keywords:
Chemotherapy;
Co-delivery;
Cytosol-localizing internalization peptide 6;
Docetaxel;
Gene therapy;
Green fluorescent protein
- From:
Academic Journal of Second Military Medical University
2018;39(2):144-151
- CountryChina
- Language:Chinese
-
Abstract:
Objective To prepare a lipoic acid (LA) modified intrinsically disordered protein-cytosol-localizing internalization peptide 6 (CL) nanocomplex (LA-CL) entering cells by non-endocytosis mechanism for co-delivery of gene and chemotherapeutic drugs, and to investigate its transfection efficiency and cellular uptake on human embryonic kidney cell line HEK293 cells and its release behavior in vitro. Methods We synthesized four disulfide cross-linked lipoic acid modified LA-CLss(1-4) at different cross-linked degrees using different mass fractions (2.5%, 5%, 10% and 20%) of cysteine as cross-linking agent. The construction of LA-CLss was characterized by1H nuclear magnetic resonance (1HNMR) and gel permeation chromatography. The LA-CLss/plasmid enhanced green fluorescent protein (pEGFP) nanocomplexes were self-assembled with LA-CLss and pEGFP at different nitrogen/phosphorus (N/P) ratios (2.5, 5, 10, 20, 40 and 80). The size and zeta potential of LA-CLss/pEGFP nanocomplexes were determined by particle size analyzer, and the pEGFP enrichment capacity of LA-CLss was determined by agarose gel electrophoresis. The docetaxel (DTX)-loaded micelles were prepared by ultrasonic emulsification, and the critical micelle concentration of LA-CLss3 was determined by pyrene fluorescence probe spectroscopy. The LA-CLss/pEGFP nanocomplexes were co-cultured with HEK293 cells, and the transfection efficiencies of LA-CLss/pEGFP nanocomplexes at different cross-linked degrees were investigated. Results1 HNMR results showed the LA-CLss was successfully synthesized. When N/P ratio was 40, the transfection efficiency of LA-CLss3/pEGFP nanocomplex by HEK293 cells was significantly higher than that of LA-CL/pEGFP, LA-CLss1/pEGFP, LA-CLss2/pEGFP and LA-CLss4 nanocomplexes. The encapsulation efficiency and drug loading of docetaxel-loaded micelles prepared by ultrasonic emulsification were (85.25±0.04)% and (8.81±0.02)%, respectively. Cellular uptake test showed that the gene could be effectively delivered into the HEK293 cells by the LA-CLss micelles. In vitro release experiments showed that the LA-CLss micelles had redox-responsive drug release behavior. Conclusion The prepared LA-CLss/DTX/pEGFP nanocomplex is expected to become an efficient vector for co-delivery of gene and chemotherapeutic drugs.
