Clinical trials reveal that plasmid DNA (pDNA)-based gene delivery must be improved to realize its potential to treat human disease. Current pDNA platforms suffer from brief transgene expression, primarily due to the spread of transcriptionally repressive chromatin initially deposited on plasmid bacterial backbone sequences. Minicircle (MC) DNA lacks plasmid backbone sequences and correspondingly confers higher levels of sustained transgene expression upon delivery, accounting for its success in preclinical gene therapy models. In this study, we show for the first time that MC DNA also functions as a vaccine platform. We used a luciferase reporter transgene to demonstrate that intradermal delivery of MC DNA, relative to pDNA, resulted in significantly higher and persistent levels of luciferase expression in mouse skin. Next, we immunized mice intradermally with DNA encoding a peptide that, when presented by the appropriate major histocompatibility complex class I molecule, was recognized by endogenous CD8(+) T cells. Finally, immunization with peptide-encoding MC DNA, but not the corresponding full-length (FL) pDNA, conferred significant protection in mice challenged with Listeria monocytogenes expressing the model peptide. Together, our results suggest intradermal delivery of MC DNA may prove more efficacious for prophylaxis than traditional pDNA vaccines.
Plasmid DNA (pDNA)-based gene delivery has the potential to advance vaccine development due to advantages such as low cost, ease of production, safety (e.g. pDNA does not cause insertional mutagenesis), and the flexibility to introduce several genes at once. To date, there are more than 300 clinical trials involving pDNA-based gene transfer. Although promising, these trials have revealed that poor transfection efficiency, transient protein production, and low immunogenicity limit therapy efficacy. We are investigating the use of minicircle (MC) DNA as a means of circumventing these limitations. MC vectors contain the expression cassette of full-length plasmids, but lack the bacterial backbone sequences that are targeted for transcriptional repression in mammalian cells. Removal of these plasmid backbone sequences is achieved via site-specific recombination in Escherichia coli prior to episomal (i.e. MC) DNA isolation. The absence of these prokaryotic elements also reduces MC size relative to its parental pDNA, leading to enhanced transfection efficiencies. Data will be presented showing that vaccination with MC DNA results in significantly higher and more prolonged reporter transgene expression. Furthermore, we compare specific responses of adoptively transferred and endogenous CD8+ T cells following vaccination of mice with either parental pDNA or MC-encoded antigen.
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