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Amelioration of Retroviral Vector Silencing in Locus Control Region β-Globin-Transgenic Mice and Transduced F9 Embryonic Cells

By Cameron S. Osborne, Peter Pasceri, Rakesh Singal, Tanya Sukonnik, Gordon D. Ginder and James Ellis

Abstract

Retroviral vectors are transcriptionally silenced in hematopoietic stem cells, and this phenomenon must be overcome for effective gene therapy of blood diseases. The murine stem cell virus (MSCV) vector completely silences β-globin reporter genes regulated by locus control region (LCR) elements 5′HS2 to 5′HS4 in seven of eight transgenic mice. Here, we show that no single known MSCV silencer element is sufficient for complete LCR β-globin transgene silencing. However, partial silencing of high-copy transgenes is conveyed by the MSCV direct repeat and promoter elements. The CpG methylation pattern of silenced and expressed MSCV promoter transgenes is virtually identical, demonstrating that silencing does not absolutely correlate with methylation status. Combined mutations in all four MSCV silencer elements leads to expression of β-globin in 6 of 10 transgenic mice. The same mutations incorporated into the HSC1 retrovirus vector direct neo gene expression in 71% of transduced F9 embryonic carcinoma cells. These studies demonstrate that combined mutation of four retroviral silencer elements relieves complete silencing in most transgenic mice and transduced F9 cells and suggests that novel silencer elements remain. Enhanced expression of the HSC1 vector in primitive stem cells is well suited for blood gene therapy applications

Topics: Gene Therapy
Publisher: American Society for Microbiology
Year: 1999
OAI identifier: oai:pubmedcentral.nih.gov:112606
Provided by: PubMed Central
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