Addition of the Human Interferon β Scaffold Attachment Region to Retroviral Vector Backbones Increases the Level of in Vivo Transgene Expression among Progeny of Engrafted Human Hematopoietic Stem Cells

Abstract

Absence of durable high-level expression of transgenes from Moloney murine leukemia (Mo-MuLV) retroviral vectors has been a hurdle in bringing effective gene therapy to the clinic. In this study we have analyzed transgene expression among the progeny of mobilized hematopoietic stem cells (HSCs), comparing Mo-MuLV and mouse stem cell virus (MSCV) vectors, with or without addition of a scaffold attachment region (SAR) from the human interferon β gene. Retroviral (RV) vector supernatant quality was assessed by comparing NGFR transgene expression by HEL cells, and transgene delivery and expression by CD34⁺ cells 72 hr after transduction, using real-time PCR and FACS analysis. This is the first description of the effect of SAR within both Mo-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, transgene expression was observed among a mean of 10% of donor HSC progeny in the SCID-hu bone (range, 0.6-43%). The predominant effect of SAR was to increase 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).