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Poly(ethylene imine)-poly(ethylene glycol) copolymers facilitate efficient delivery of antisense oligonucleotides to nuclei of mature muscle cells of mdx mice
Journal article   Peer reviewed

Poly(ethylene imine)-poly(ethylene glycol) copolymers facilitate efficient delivery of antisense oligonucleotides to nuclei of mature muscle cells of mdx mice

Shashank R Sirsi, Jason H Williams and Gordon J Lutz
Human gene therapy, v 16(11), pp 1307-1317
Nov 2005
DOI: https://doi.org/10.1089/hum.2005.16.1307
PMID: 16259564
Featured in Collection :   UN Sustainable Development Goals @ Drexel

Abstract

Animals Base Sequence Cell Nucleus - metabolism Mice Mice, Inbred mdx Microscopy, Confocal Microscopy, Fluorescence Muscle Fibers, Skeletal - cytology Muscle Fibers, Skeletal - metabolism Muscle Fibers, Skeletal - ultrastructure Muscles - cytology Muscles - metabolism Muscles - ultrastructure Muscular Dystrophy, Duchenne - therapy Oligonucleotides, Antisense - administration & dosage Polyethylene Glycols - administration & dosage Polyethyleneimine - administration & dosage
Antisense oligonucleotides (AO) can facilitate dystrophin expression via targeted exon skipping in cultured cells of Duchenne muscular dystrophy (DMD) patients and in the mouse model of DMD (mdx mice). However, the lack of effective means to deliver AO to myonuclei remains the foremost limitation to their usefulness in DMD gene therapy. In this study we show that copolymers of cationic poly(ethylene imine) (PEI) and poly(ethylene glycol) (PEG) facilitated efficient cellular uptake and nuclear delivery of AO in mature skeletal muscle fibers isolated from mdx mice. Confocal analysis of dual fluorescently tagged PEG-PEI-AO polyplexes, 24 hr after transfection, showed that the copolymer and AO were colocalized within punctate membrane- associated structures. Importantly, AO was efficiently translocated into myonuclei, whereas the copolymer was mostly excluded. The morphology of all transfected myofibers was perfectly maintained with no indication of damage or cytotoxicity. Quantitative fluorescence analysis showed that transfection with PEG-PEI-AO resulted in a 6-fold higher uptake of AO into myonuclei compared with transfections of AO alone. Interestingly, transfections with rhodamine-labeled PEG-PEI copolymers yielded an approximately 2- fold higher uptake of AO into myonuclei compared with transfections of unlabeled copolymers. Attempts to further increase AO delivery by addition of insulin-transferrin-selenium (ITS) to the medium showed no further improvement in AO delivery. Dose-response analysis indicated saturation of endocytotic uptake of the polyplex. Overall, we conclude that PEG-PEI copolymers represent high-capacity, nontoxic carriers for efficient delivery of AO to nuclei of mature myofibers.

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Web of Science research areas
Biotechnology & Applied Microbiology
Genetics & Heredity
Medicine, Research & Experimental
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