Persistence of plasmid‐mediated expression of transgenes in human mesenchymal stem cells depends primarily on CpG levels of both vector and transgene

Abstract

The persistence of plasmid vector expression in human MSCs is determined primarily by CpG content of both vector and transgene. The data obtained in the present study indicate that the pMBR2 vector with a minimized number of CpG motifs is appropriate for extended plasmid-mediated expression of transgenes in MSCs and possibly other types of stem cells.

Advances in the Development and the Applications of Nonviral, Episomal Vectors for Gene Therapy

“…Nevertheless, “harmful” DNA sequences exist in a replicating episome, and can reside in the promoter, 50 transgene, 51 or backbone. 23 These sequences are often targets of epigenetic gene silencing, 23 , 52 as well as the genes within the vector that are necessary for plasmid propagation in bacteria. The deletion of such sequences generally produces smaller constructs, which may improve the delivery into (and movement within) the recipient cell.…”

“…These observations by Bruter et al 65 suggested that the requirements of vector CpG-free sequences to optimize gene expression can vary depending on the tissue to be transfected. Interestingly, the same group reported that for mouse liver 65 or human mesenchymal stem cells, 52 both the vector and the transgene should be CpG free, whereas in mouse skeletal muscle, 65 removing the CpG motifs from the vector alone was sufficient to maintaining expression for about 100 days relative to control vector. Noticeably, other vectors with similar dual S/MAR regions (IFNβ S/MAR and an immunoglobulin S/MAR 66 or short chimeric MARs 67 ) promote higher colony formation, transgene expression levels, and maintenance in CHO cells relative to one S/MAR, suggesting that more than one S/MAR may augment the vector's activity and function further.…”