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  3. Medizin - Open Access LMU - Teil 20/22 Podcast
  4. Generation of a tumor- and tissue-specific episomal non-viral vector system
Background: A key issue for safe and reproducible gene therapy
approaches is the autologous and tissue-specific expression of
transgenes. Tissue-specific expression in vivo is either achieved
by transfer vectors that deliver the gene of interest into a
distinct cell type or by use of tissue-specific expression
cassettes. Here we present the generation of non-viral, episomally
replicating vectors that are able to replicate in a tissue specific
manner thus allowing tissue specific transgene expression in
combination with episomal replication. The episomal replication of
the prototype vector pEPI-1 and its derivatives depends exclusively
on a transcription unit starting from a constitutively active
promoter extending into the scaffold/matrix attachment region
(S/MAR). Results: Here, we exchanged the constitutive promoter in
the pEPI derivative pEPito by the tumor specific alpha fetoprotein
(AFP) or the muscle specific smooth muscle 22 (SM22) promoter
leading to specific transgene expression in AFP positive human
hepatocellular carcinoma (HUH7) and in a SM22 positive cell line,
respectively. The incorporation of the hCMV enhancer element into
the expression cassette further boosted the expression levels with
both promoters. Tissue specific-replication could be exemplary
proven for the smooth muscle protein 22 (SM22) promoter in vitro.
With the AFP promoter-driven pEPito vector hepatocellular
carcinoma-specific expression could be achieved in vivo after
systemic vector application together with polyethylenimine as
transfection enhancer. Conclusions: In this study we present an
episomal plasmid system designed for tissue specific transgene
expression and replication. The human AFP-promoter in combination
with the hCMV enhancer element was demonstrated to be a valuable
tissue-specific promoter for targeting hepatocellular carcinomas
with non-viral gene delivery system, and tissue specific
replication could be shown in vitro with the muscle specific SM22
promoter. In combination with appropriate delivery systems, the
tissue specific pEPito vector system will allow higher
tissue-specificity with less undesired side effects and is suitable
for long term transgene expression in vivo within gene
therapeutical approaches.

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