Gene Therapy Progress and Prospects: Episomally maintained self-replicating systems

Abstract

The use of nonviral gene therapy vectors has been hampered by low level of transfection efficiency and lack of sustained gene expression. Episomal self-replicating systems may overcome these hurdles through their large packaging capacity, stability and reduced toxicity. This article reviews three classes of episomal molecules that have been tested with possible therapeutic genes: (1) self-replicating circular vectors, containing the Epstein-Barr virus (EBV) elements oriP and EBNA1; (2) small circular vectors containing scaffold/matrix attachment regions (S/MARs) as cis-acting elements to maintain the episomal status of the vector; (3) chromosomal vectors, based on the functional elements of the natural chromosomes. The studies reported validate the use of episomal vectors to obtain stable and prolonged gene expression, although reveal some limitations that necessitate additional work.

Persistent episomal transgene expression in liver following delivery of a scaffold/matrix attachment region containing non-viral vector

“…40,41 Additionally, no prolonged luciferase expression or DNA persistence was observed after intravenous lung administration of an S/MAR-containing pEPI-1 vector with a CMV-driven expression cassette. 42 Together with our data, these results suggest that S/MAR-containing vectors are prone to promoter attenuation in vivo if not combined with constitutive or tissue-specific promoters. The exact mechanism of how an S/MAR element may protect a particular promoter in vivo from methylation now requires further characterization but it can be assumed that cell-or tissue-specific factors play a decisive role in this process.…”

Gene transfer into human hematopoietic progenitor cells with an episomal vector carrying an S/MAR element

“…Alternatively or in addition to, silencing of the CMV promoter could also account for this, especially since pCMV has been reported to be prone to silencing in primary human cells. 43,44 Given that the presence of an active transcription unit upstream of the S/MAR is believed to play a very important role in vector's maintenance, 20,27 the expression cassette of this plasmid is likely to have an impact on properties of the vector other than transgene expression rate and it is possible that the use of alternative constitutive viral promoters or tissue-specific promoters might also enhance vector stability. We are currently examining this hypothesis by testing the effect that different transcriptional cisregulatory elements have on overall vector performance in CD34 + cells.…”

Intracellular partitioning of cell organelles and extraneous nanoparticles during mitosis

“…Moreover, Tessadori et al [178] concluded that the regulatory mechanisms that act on episomal genes seem to be similar to those that control host genes. The episomal chromatin undergoes the same modifications as the host chromatin e.g.…”