Genetic treatment of a molecular disorder: gene therapy approaches to sickle cell disease

Abstract

Effective medical management for sickle cell disease (SCD) remains elusive. As a prevalent and severe monogenic disorder, SCD has been long considered a logical candidate for gene therapy. Significant progress has been made in moving toward this goal. These efforts have provided substantial insight into the natural regulation of the globin genes and illuminated challenges for genetic manipulation of the hematopoietic system. The initial γ-retroviral vectors, next-generation lentiviral vectors, and novel genome engineering and gene regulation approaches each share the goal of preventing erythrocyte sickling. After years of preclinical studies, several clinical trials for SCD gene therapies are now open. This review focuses on progress made toward achieving gene therapy, the current state of the field, consideration of factors that may determine clinical success, and prospects for future development.

Figure 1

Vector schematic: general organization of proviral form of lentiviral vectors for gene therapy for SCD. (A) Antisickling β-globin vector containing the T87Q mutation. The SIN lentiviral vector is noninsulated and contains a modified LCR with HS2, HS3, and HS4. (B) Antisickling β-globin vector containing the G16D, E22A, and T87Q mutations (AS3). The SIN lentiviral vector has an FB element for enhancer-blocking activity and contains a modified LCR with HS2, HS3, and HS4. (C) A γ/β-globin hybrid vector containing the coding regions of γ-globin and the noncoding regions of β-globin. γ-globin (dark blue boxes), β-globin, or antisickling β-globin gene cassettes (blue boxes) with β-globin 5′ and 3′ untranslated regions (short blue boxes) under the control of a β-globin promoter (blue arrow) and modified β-globin LCR (red). Gene cassette is in reverse with respect to viral transcription to avoid aberrant splicing during packaging due to presence of globin intronic sequences (light blue boxes). All vectors are SIN (ΔU3) lentiviral vectors (green). Vectors are intended as basic schematics of gene therapy clinical vectors for SCD and are neither drawn to scale nor are all details included. ψ, packaging signal; cPPT, central polypurine tract; HS, DNase I hypersensitive site; LTR, long terminal repeats (U3, R, and U5); RRE, rev-responsive element; WPRE, woodchuck hepatitis virus posttranscriptional regulator element.

Figure 2

Strategies for gene therapy for SCD: schematic overview of various approaches for correcting the sickle phenotype via gene therapy. Gene correction: targeted genome engineering leads to correction of the sickle mutation such that β^S is repaired as β^A. HbF induction: multiple strategies for induction of γ-globin expression include shRNA-mediated knockdown of BCL11A, targeted disruption of the +58 DNase I HS site in the BCL11A erythroid-specific enhancer, and forced chromatin looping to promote association of the β-globin LCR with the γ-globin genes. Gene addition: integrating lentiviral vector carrying a β-globin, γ-globin, or antisickling β-globin cassette. Ldb1, transcription factor; ZF/SA, zinc-finger self-association domain.