pEPito: a significantly improved non-viral episomal expression vector for mammalian cells.
BACKGROUND
The episomal replication of the prototype vector pEPI-1 depends on a transcription unit starting from the constitutively expressed Cytomegalovirus immediate early promoter (CMV-IEP) and directed into a 2000 bp long matrix attachment region sequence (MARS) derived from the human beta-interferon gene. The original pEPI-1 vector contains two mammalian transcription units and a total of 305 CpG islands, which are located predominantly within the vector elements necessary for bacterial propagation and known to be counterproductive for persistent long-term transgene expression.


RESULTS
Here, we report the development of a novel vector pEPito, which is derived from the pEPI-1 plasmid replicon but has considerably improved efficacy both in vitro and in vivo. The pEPito vector is significantly reduced in size, contains only one transcription unit and 60% less CpG motives in comparison to pEPI-1. It exhibits major advantages compared to the original pEPI-1 plasmid, including higher transgene expression levels and increased colony-forming efficiencies in vitro, as well as more persistent transgene expression profiles in vivo. The performance of pEPito-based vectors was further improved by replacing the CMV-IEP with the human CMV enhancer/human elongation factor 1 alpha promoter (hCMV/EF1P) element that is known to be less affected by epigenetic silencing events.


CONCLUSIONS
The novel vector pEPito can be considered suitable as an improved vector for biotechnological applications in vitro and for non-viral gene delivery in vivo.
Max von Pettenkofer-Institute, University of Munich, Munich, Germany.
Rudolf
R
Haase
Orestis
O
Argyros
Suet-Ping
SP
Wong
Richard P
RP
Harbottle
Hans J
HJ
Lipps
Manfred
M
Ogris
Terese
T
Magnusson
Maria G
MG
Vizoso Pinto
Jürgen
J
Haas
Armin
A
Baiker
D000818:Animals; D018899:CpG Islands; D003587:Cytomegalovirus / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors / Q000096:biosynthesis*; D006801:Humans; D008099:Liver / Q000378:metabolism; D008297:Male; D045170:Matrix Attachment Regions; D051379:Mice; D041681:NIH 3T3 Cells; D010957:Plasmids / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012093:Replicon; D014162:Transfection; D019076:Transgenes