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Updated: Aug 14, 2026

CRISPR/Cas9 Gene Editing of Hematopoietic Stem and Progenitor Cells for Gene Therapy Applications
Published on: August 9, 2022
A genetic strategy to treat sickle cell anemia by coregulating globin transgene expression and RNA interference
Selda Samakoglu1, Leszek Lisowski, Tulin Budak-Alpdogan
1Laboratory of Gene Transfer and Gene Expression, Sloan-Kettering Institute, New York, New York 10021, USA.
Abstract:
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 gamma-globin gene. Expression of both gamma-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 gamma-globin transgene and concomitant reduction of endogenous beta(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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