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A new chromosome 14-based human artificial chromosome (HAC) vector system for efficient transgene expression in human primary cells.

Abstract：

:The use of non-integrating human artificial chromosomes (HACs) in gene therapy possibly allows for safe and reliable genetic modification of human cells without insertional mutagenesis and/or unexpected oncogene activations. Although we previously demonstrated that the HAC provides long-term therapeutic erythropoietin (EPO) production in normal human primary fibroblasts (hPFs), the expression level of EPO was too low to provide medical benefits for human therapy. Thus, the next challenge for the application of this system in therapeutic purposes is to improve the transgene expression on HACs. Here, we newly constructed chromosome 14-based HACs and examined the effects of the telomere and promoter regions on the expression level of the tansgene in hPFs. We showed that the use of natural telomere/sub-telomere and enhancers within the 5' untranslated region of the human ubiquitin C gene greatly increased (over 1000-fold) the EPO production in hPFs. Furthermore, we demonstrated the reprogramming of mouse embryonic fibroblasts by HAC-mediated introduction of four transcription factors, and established induced pluripotent stem cells with no trace of the HACs carrying multiple expression cassettes with large genome fragments. These results indicate that this HAC system could allow us to manipulate multiple transgenes efficiently in human primary cells, providing a promising tool not only for gene therapy but also for investigating genome functions in drug discoveries.

journal_name

Biochem Biophys Res Commun

journal_title

Biochemical and biophysical research communications

authors

Kakeda M,Nagata K,Osawa K,Matsuno H,Hiratsuka M,Sano A,Okazaki A,Shitara S,Nishikawa S,Masuya A,Hata T,Wako S,Osaki M,Kazuki Y,Oshimura M,Tomizuka K

doi

10.1016/j.bbrc.2011.10.088

subject

Has Abstract

pub_date

2011-11-25 00:00:00

pages

439-44

issue

3

eissn

0006-291X

issn

1090-2104

pii

S0006-291X(11)01900-0

journal_volume

415

pub_type

杂志文章

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