A genetic strategy to treat sickle cell anemia by coregulating globin transgene expression and RNA interference

• Published in: Nature biotechnology Vol. 24; no. 1; pp. 89 - 94
• Main Authors: Sadelain, Michel, Samakoglu, Selda, Lisowski, Leszek, Budak-Alpdogan, Tulin, Usachenko, Yelena, Acuto, Santina, Di Marzo, Rosalba, Maggio, Aurelio, Zhu, Ping, Tisdale, John F, Rivière, Isabelle
• Format: Journal Article
• Language: English
• Published: New York, NY Nature 01.01.2006; Nature Publishing Group
• Subjects: Anemia, Sickle Cell - genetics; Anemia, Sickle Cell - metabolism; Anemia, Sickle Cell - pathology; Anemia, Sickle Cell - therapy; Biological and medical sciences; Biotechnology; Cells, Cultured; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation - genetics; Gene Silencing; Gene Targeting - methods; Genetic Therapy - methods; Globins - genetics; Globins - metabolism; Globins - therapeutic use; Health. Pharmaceutical industry; Hematopoietic Stem Cells - metabolism; Humans; Industrial applications and implications. Economical aspects; Miscellaneous; RNA Interference; Stem cells; Treatment Outcome; Sickle cell anemia; Cell therapy; RNA interference; Targeting; Stem cell; Retroviridae; Hematopoietic cell; Gene overexpression; Gene expression; Lentivirus; Virus; Globin; Gene silencing; Messenger RNA; Gene therapy; Application
• ISSN: 1087-0156; 1546-1696
• DOI: 10.1038/nbt1176

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 γ-globin gene. Expression of both γ-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 γ-globin transgene and concomitant reduction of endogenous βS transcripts, thus providing proof of principle for therapeutic strategies that require synergistic gene addition and gene silencing in stem cell progeny.