PEGylation Improves Nanoparticle Formation and Transfection Efficiency of Messenger RNA
Cationic polymers have been intensively investigated for plasmid-DNA (pDNA), but few studies addressed their use for messenger-RNA (mRNA) delivery. We analyzed two types of polymers, linear polyethylenimine (l-PEI) and poly-N,N-dimethylaminoethylmethacrylate P(DMAEMA), to highlight specific requirements for the design of mRNA delivery reagents. The effect of PEGylation was investigated using P(DMAEMA-co-OEGMA) copolymer.
The influence of polymer structure on mRNA binding and particle formation was assessed in a side-by-side comparison with pDNA by methods such as agarose-retardation assay and scanning probe microscopy. Transfection studies were performed on bronchial epithelial cells.
Binding of cationic polymers inversely correlated with type of nucleic acid. Whereas P(DMAEMA) bound strongly to pDNA, only weak mRNA binding was observed, which was vice versa for l-PEI. Both polymers resulted in self-assembled nanoparticles forming pDNA complexes of irregular round shape; mRNA particles were significantly smaller and more distinct. Surprisingly, PEGylation improved mRNA binding and transfection efficiency contrary to observations made with pDNA. Co-transfections with free polymer improved mRNA transfection.
Gene delivery requires tailor-made design for each type of nucleic acid. PEGylation influenced mRNA-polymer binding efficiency and transfection and may provide a method of further improving mRNA delivery.
KEY WORDScationic polymers gene delivery mRNA
- 15.Van Driessche A, Van de Velde AL, Nijs G, Braeckman T, Stein B, De Vries JM, et al. Clinical-grade manufacturing of autologous mature mRNA-electroporated dendritic cells and safety testing in acute myeloid leukemia patients in a phase I dose-escalation clinical trial. Cytotherapy. 2009;11:653–68.PubMedCrossRefGoogle Scholar
- 19.Kormann MS, Hasenpusch G, Aneja MK, Nica G, Flemmer AW, Herber-Jonat S, Huppmann M, Mays LE, Illenyi M, Schams A, Griese M, Bittmann I, Handgretinger R, Hartl D, Rosenecker J, Rudolph C. Expression of therapeutic proteins after delivery of chemically modified mRNA in mice. Nat Biotechnol. 29:154–7Google Scholar
- 20.Van Tendeloo VF, Ponsaerts P, Lardon F, Nijs G, Lenjou M, Van Broeckhoven C, et al. Highly efficient gene delivery by mRNA electroporation in human hematopoietic cells: superiority to lipofection and passive pulsing of mRNA and to electroporation of plasmid cDNA for tumor antigen loading of dendritic cells. Blood. 2001;98:49–56.PubMedCrossRefGoogle Scholar
- 24.Orgis M, Wagner E. Linear polyethylenimine: synthesis and transfection procedures for in vitro and in vivo. In: Friedman T, Rossi J, editors. Gene transfer: delivery and expression of cDNA and RNA, a laboratory manual. New York: Cold Spring Habor Laboratory Press; 2007. p. 521–8.Google Scholar
- 26.Üzgün S, Akdemir Ö, Hsenpusch G, Maucksch C, Golas MM, Sander B, Stark H, Imker R, Lutz JF, Rudolph C. Characterization of tailor-made copolymers of oligo(ethylene glycol) methyl ether methacrylate (OEGMA) and N,N-dimethylaminoethyl methacrylate (DMAEMA) as nonviral gene transfer agents—influence of macromolecular structure on gene vector particle properties and transfection efficiency. Biomacromolecules. in pressGoogle Scholar
- 27.Wagner E, Plank C, Zatloukal K, Cotten M, Birnstiel ML. Influenza virus hemagglutinin HA-2 N-terminal fusogenic peptides augment gene transfer by transferrin-polylysine-DNA complexes: toward a synthetic virus-like gene-transfer vehicle. Proc Natl Acad Sci USA. 1992;89:7934–8.PubMedCrossRefGoogle Scholar