CAGW Peptide Modified Biodegradable Cationic Copolymer for Effective Gene Delivery

Abstract

In recent years, gene therapy has become a promising technology to enhance endothelialization of artificial vascular grafts. The ideal gene therapy requires a gene carrier with low cytotoxicity and high transfection efficiency. In this paper, we prepared a biodegradable cationic copolymer poly(d,l-lactide-co-glycolide)-graft-PEI (PLGA-g-PEI), grafted Cys-Ala-Gly-Trp (CAGW) peptide onto this copolymer via the thiol-ene Click-reaction, and then prepared micelles by a self-assembly method. pEGFP-ZNF580 plasmids (pDNA) were condensed by these micelles via electrostatic interaction to form gene complexes. The CAGW peptide enables these gene complexes with special recognition for endothelial cells, which could enhance their transfection. As a gene carrier system, the PLGA-g-PEI-g-CAGW/pDNA gene complexes were evaluated and the results showed that they had suitable diameter and zeta potential for cellular uptake, and exhibited low cytotoxicity and high transfection efficiency for EA.hy926 cells.

CAGW and TAT‐NLS peptides functionalized multitargeting gene delivery system with high transfection efficiency

“…The representative structures and 1 H NMR spectra of mP, PP, and PC copolymers were shown in Figure S1. The results demonstrated that these amphiphilic polymers were successfully synthesized …”

“…N‐Hydroxy succinimide (NHS), 1‐ethyl‐3‐(3‐(dimethylamino)propyl)‐carbodiimide hydrochloride (EDC), succinic anhydride, diallylcarbamyl chloride, and 2,2‐dimethoxy‐2‐phenylacetophenone (DMPA) were obtained from Tianjin Heowns Biochemical Technology Co., Ltd. CAGW (Cys‐Ala‐Gly‐Trp) and TAT‐NLS (Gly‐Arg‐Lys‐Lys‐Arg‐Arg‐Gln‐Arg‐Arg‐Arg‐Pro‐Lys‐Lys‐Lys‐Arg‐Lys‐Val) peptides were purchased from GL Biochem (Shanghai) Ltd. EA.hy926 cells (human umbilical vein endothelial cells) were purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA), and cultured as described before . Other chemicals and biological reagents were supplied by the companies referred in our previous study …”

“…To improve the transfection efficiency, CAG and REDV peptides were conjugated to gene carriers to selectively adhere ECs and promote the cellular uptake . Recently, PLGA‐g‐PEI‐g‐CAGW/pDNA targeting gene complexes were used to transfect ECs and showed relatively high transfection efficiency in our previous study …”

“…In present study, the multitargeting gene complexes based on mPEG‐b‐PLGA, PLGA‐g‐PEI‐g‐CAGW, TAT‐NLS peptide, and pEGFP‐ZNF580 (pDNA) were developed to improve the transfection of ECs. pEGFP‐ZNF580 (pDNA) was used as target gene as described before . CAGW peptide could endow the multitargeting gene complexes with selective ECs‐targeting ability, and TAT‐NLS peptide enables them with cell‐penetrating ability and nuclear accumulation capacity.…”

“…28,29 Recently, PLGA-g-PEI-g-CAGW/ pDNA targeting gene complexes were used to transfect ECs and showed relatively high transfection efficiency in our previous study. 30 To improve the cell-penetrating ability, endo/lysosomal escape ability, and nuclear-targeting ability of gene delivery systems, cellpenetrating peptides (CPPs) and nuclear localization signal (NLS) have been introduced to gene carriers. [31][32][33][34] TAT and NLS peptides possess cell-penetrating ability and nuclear localization capacity, respectively.…”