Nature Biotechnology 24, 89 - 94 (2005)
Published online: 25 December 2005; | doi:10.1038/nbt1176
A genetic strategy to treat sickle cell anemia by coregulating globin transgene expression and RNA interferenceSelda Samakoglu1, Leszek Lisowski1, Tulin Budak-Alpdogan2, 3, Yelena Usachenko1, Santina Acuto4, Rosalba Di Marzo4, Aurelio Maggio4, Ping Zhu1, John F. Tisdale5, Isabelle Rivière2, 6
& Michel Sadelain1, 61
Laboratory of Gene Transfer and Gene Expression, Sloan-Kettering Institute, New York, New York 10021 USA. 2
Gene Transfer and Somatic Cell Engineering Facility, Sloan-Kettering Institute, New York, New York 10021 USA. 3
Department of Medicine, University of Medicine and Dentistry of New Jersey, New Brunswick, New Jersey 08901 USA. 4
Div. Ematologia II con Talassemia, Ospedale V.Cervello, Palermo 90146 Italy. 5
MCHB/NIDDK National Institutes of Health/DHHS, Bethesda, Maryland 20892 USA. 6
Department of Medicine and Immunology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021 USA.
Correspondence should be addressed to Michel Sadelain m-sadelain@ski.mskcc.org The application of RNA interference (RNAi) to stem cell–based therapies will require highly specific and lineage-restricted gene silencing. Here we show the feasibility and therapeutic potential of coregulating transgene expression and RNAi in hematopoietic stem cells. We encoded promoterless small-hairpin RNA (shRNA) within the intron of a recombinant  -globin gene. Expression of both -globin and the lariat-embedded small interfering RNA (siRNA) was induced upon erythroid differentiation, specifically downregulating the targeted gene in tissue- and differentiation stage–specific fashion. The position of the shRNA within the intron was critical to concurrently achieve high-level transgene expression, effective siRNA generation and minimal interferon induction. Lentiviral transduction of CD34+ cells from patients with sickle cell anemia led to erythroid-specific expression of the -globin transgene and concomitant reduction of endogenous  S transcripts, thus providing proof of principle for therapeutic strategies that require synergistic gene addition and gene silencing in stem cell progeny.
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