Maniar, LEG, Maniar, JM, Chen, Z-Y, Lu, J, Fire, AZ and Kay, MA. Minicircle DNA vector achieve sustained expression reflected by active chromatin and transcriptional level. Mol Ther 21: 131-138

Department of Genetics, Stanford University School of Medicine, Stanford, California, USA.
Molecular Therapy (Impact Factor: 6.23). 11/2012; 21(1). DOI: 10.1038/mt.2012.244
Source: PubMed


Current efforts in nonviral gene therapy are plagued by a pervasive difficulty in sustaining therapeutic levels of delivered transgenes. Minicircles (plasmid derivatives with the same expression cassette but lacking a bacterial backbone) show sustained expression and hold promise for therapeutic use where persistent transgene expression is required. To characterize the widely-observed silencing process affecting expression of foreign DNA in mammals, we used a system in which mouse liver presented with either plasmid or minicircle consistently silences plasmid but not minicircle expression. We found that preferential silencing of plasmid DNA occurs at a nuclear stage that precedes transport of mRNA to the cytoplasm, evident from a consistent >25-fold minicircle/plasmid transcript difference observed in both nuclear and total RNA. Among possible mechanisms of nuclear silencing, our data favor chromatin-linked transcriptional blockage rather than targeted degradation, aberrant processing, or compromised mRNA transport. In particular, we observe dramatic enrichment of H3K27 trimethylation on plasmid sequences. Also, it appears that Pol II can engage the modified plasmid chromatin, potentially in a manner that is not productive in the synthesis of high levels of new transcript. We outline a scenario in which sustained differences at the chromatin level cooperate to determine the activity of foreign DNA.Molecular Therapy (2012); doi:10.1038/mt.2012.244.

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Available from: Mark A Kay, Jun 28, 2014
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    • "The size of minicircles, which is smaller than that of conventional plasmids, increases the possibility of delivering these vectors into cells. Moreover, it has been reported that minicircles escape gene silencing and show sustained expression because of the unique methylation patterns found in these vectors8. "
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    • "However, significant enhancement of transgene expression was also observed in the FLPo-based system lacking oriP/EBNA1 (unpublished data); thus, the enhancer activity of oriP is not the key determinant for the enhanced expression. Conversely, it was shown that minicircles confer stronger and longer transgene expression than their plasmid counterpart (52), which was associated with chromatin-linked transcription blockade (53) but was independent of CpG methylation (52). As BV genome (134 kb) is significantly larger than the minicircle (<10 kb), one may envisage that BV genome, when compared with the smaller minicircle, forms a chromatin structure that less favors persistent transgene expression. "
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