In vivo gene transfer using sulfhydryl cross-linked PEG-peptide/glycopeptide DNA co-condensates.
Recent interest in sulfhydryl cross-linked nonviral gene delivery systems, designed to trigger the intracellular release of DNA, has inspired studies to establish their utility in vitro. To determine if this concept can be extrapolated to in vivo gene delivery, sulfhydryl cross-linking peptides (dp 20), derivatized with either an N-glycan or polyethylene glycol (PEG), were used to generate sulfhydryl cross-linked gene formulations. The biodistribution, metabolism, cell-type targeting, and gene expression of sulfhydryl cross-linked PEG-peptide/glycopeptide DNA co-condensates were examined following i.v. dosing in mice. Optimal targeting to hepatocytes was achieved by condensing (125)I-DNA with an add-mixture of 10 mol % triantennary glycopeptide, 5 mol % PEG-peptide, and 85 mol % backbone peptide. Four backbone peptides were substituted into the formulation to examine the influence of peptide metabolism and disulfide bond strength on the rate of DNA metabolism and the level of gene expression in vivo. The half-life of DNA in liver was extended from 1 to 3 h using a backbone peptide composed of d-amino acids, whereas substituting penicillamine for cysteine failed to further increase the metabolic stability of DNA. Optimized gene delivery formulations transiently expressed secreted alkaline phosphatase in mouse serum for 12 days. The results suggest that disulfide bond reduction in liver hepatocytes proceeds rapidly, followed by peptide metabolism, ultimately limiting the metabolic half-life of sulfhydryl cross-linked DNA condensates in vivo.
Kwok
Kai Y
KY
Department of Pharmaceutics and Medicinal Chemistry, College of Pharmacy, University of Michigan, Ann Arbor, Michigan 48109-1065, USA.
Park
Youmie
Y
Yang
Yongsheng
Y
McKenzie
Donald L
DL
Liu
Yahong
Y
Rice
Kevin G
KG
D000469:Alkaline Phosphatase / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D001682:Biological Availability; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D003432:Cross-Linking Reagents / Q000737:chemistry; D004247:DNA / Q000737:chemistry / Q000493:pharmacokinetics*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D006020:Glycopeptides / Q000737:chemistry; D006801:Humans; D007728:Kupffer Cells / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry; D010920:Placenta / Q000201:enzymology; D010957:Plasmids; D011092:Polyethylene Glycols / Q000737:chemistry; D013438:Sulfhydryl Compounds / Q000737:chemistry
D003432:Cross-Linking Reagents; D006020:Glycopeptides; D009842:Oligopeptides; D013438:Sulfhydryl Compounds; D011092:Polyethylene Glycols; D004247:DNA; D000469:Alkaline Phosphatase