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• W2014328248 startingPage "3632" @default.
• W2014328248 abstract "Antisense oligonucleotides (ODN) targeted to specific genes have shown considerable potential as therapeutic agents. The polyanionic charges carried by these molecules, however, present a barrier to efficient cellular uptake and consequently their biological effects on gene regulation are compromised. To overcome this obstacle, a rationally designed carrier system is desirable for antisense delivery. This carrier should assist antisense ODN penetrate the cell membrane and, once inside the cell, then release the ODN and make them available for target binding. We have developed a carrier formulation employing programmable fusogenic vesicles (PFV) as the antisense delivery mediator. This study investigates the intracellular fate of PFV–ODN and bioavailability of antisense ODN to cells. The subcellular distribution of PFV and ODN was examined by monitoring the trafficking of FITC‐labeled ODN and rhodamine/phosphatidylethanolamine (Rh‐PE)‐labeled PFV using confocal microscopy. Fluorescently tagged ODN were first co‐localized with the liposomal carrier in the cytoplasm, presumably in endosome/lysosome compartments, shortly after incubation of PFV–ODN with HEK 293 and 518A2 cells. Between 24 and 48 h incubation, however, separation of FITC–ODN from the carrier and subsequent accumulation in the nucleus was observed. In contrast, the Rh‐PE label was localized to the cell cytoplasm. The enhanced cellular uptake achieved using the PFV carrier, compared to incubation of free ODN with cells, and subsequent release of ODN from the carrier resulted in significant down‐regulation of mRNA expression. Specifically, G3139, an antisense construct targeting the apoptotic antagonist gene bcl‐2 , was examined in the human melanoma cell line 518A2. Upon exposure to PFV‐encapsulated G3139, cells displayed a time‐dependent reduction in bcl‐2 message levels. The bcl‐2 mRNA level was reduced by 50% after 24 h treatment and by ∼80% after 72 h when compared to cells treated with free G3139, empty PFV or PFV–G3622, a control ODN sequence. Our results establish that ODN can be released from PFV after intracellular uptake and can then migrate to the nucleus and selectively down‐regulate target mRNA." @default.
• W2014328248 created "2016-06-24" @default.
• W2014328248 creator A5056585446 @default.
• W2014328248 date "2002-08-01" @default.
• W2014328248 modified "2023-09-25" @default.
• W2014328248 title "Subcellular trafficking of antisense oligonucleotides and down-regulation of bcl-2 gene expression in human melanoma cells using a fusogenic liposome delivery system" @default.
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• W2014328248 doi "https://doi.org/10.1093/nar/gkf448" @default.
• W2014328248 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/137064" @default.
• W2014328248 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/12177306" @default.
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