Addition of the human interferon beta scaffold attachment region to retroviral vector backbones increases the level of in vivo transgene expression among progeny of engrafted human hematopoietic stem cells

Absence of durable high-level expression of transgenes from Moloney murine leukemia (Mo-MuLV) retroviral vectors has been a hurdle in bringing effecti ve gene therapy to the clinic. In this study we have analyzed transgene exp ression among the progeny of mobilized hematopoietic stem cells (HSCs), com paring Mo-MuLV and mouse stem cell virus (MSCV) vectors, with or without ad dition of a scaffold attachment region (SAR) from the human interferon beta gene. Retroviral (RV) vector supernatant quality was assessed by comparing NGFR transgene expression by HEL cells, and transgene delivery and express ion by CD34(+) cells 72 hr after transduction, using real-time PCR and FAGS analysis. This is the first description of the effect of SAR within both M o-MuLV and MSCV vector backbones on long-term RV transgene expression among in vivo HSC progeny in HSC repopulation assays (SCID-hu bone and NOD/SCID) , After transduction of mobilized CD34(+) cells with MSCV-SAR vector, trans gene expression was observed among a mean of 10% of donor HSC progeny in th e SCID-hu bone (range, 0.6-43%). The predominant effect of SAR was to incre ase the mean fluorescence intensity (MFI) of transgene expression among HSC progeny in both in vivo bone repopulation models (three- to fourfold), and after long-term stromal cultures (twofold),.