Abstract
The advent of next-generation genome engineering tools like CRISPR-Cas9 has transformed the field of gene therapy, rendering targeted treatment for several incurable diseases. Hematopoietic stem and progenitor cells (HSPCs) continue to be the ideal target cells for gene manipulation due to their long-term repopulation potential. Among the gene manipulation strategies such as lentiviral gene augmentation, non-homologous end joining (NHEJ)-mediated gene editing, base editing and prime editing, only the homology-directed repair (HDR)-mediated gene editing provides the option of inserting a large transgene under its endogenous promoter or any desired locus. In addition, HDR-mediated gene editing can be applied for the gene knock-out, correction of point mutations and introduction of beneficial mutations. HSPC gene therapy studies involving lentiviral vectors and NHEJ-based gene-editing studies have exhibited substantial clinical progress. However, studies involving HDR-mediated HSPC gene editing have not yet progressed to the clinical testing. This suggests the existence of unique challenges in exploiting HDR pathway for HSPC gene therapy. Our review summarizes the mechanism, recent progresses, challenges, and the scope of HDR-based gene editing for the HSPC gene therapy.
Highlights
Recent estimates suggest that over 8000 diseases are of monogenic in origin, often manifesting during childhood and causing premature deaths in severe cases
The results showed marginal levels of homology-directed repair (HDR) (2–18%) with post-transplant reduction in HDR frequency (0.27–2%) [10,11,12]
With CRISPR-Cas9 and AAV6 donors, the insertion frequency was achieved up to 45% in patient Hematopoietic stem and progenitor cells (HSPCs) in vitro and functional correction was achieved in 10–20% of LT-HSCs in vivo [30]
Summary
Recent estimates suggest that over 8000 diseases are of monogenic in origin, often manifesting during childhood and causing premature deaths in severe cases. The burden of genetic disorders remains alarmingly high as the clinical management of many such diseases is largely inefficient. For monogenic disorders, such as β-hemoglobinopathies (β-thalassemia and sickle cell disease (SCD)), cystic fibrosis, hemophilia, Huntington’s disease and Duchenne muscular dystrophy, targeted therapeutic strategies are in high demand. Gene therapy aims to correct the root cause of monogeneic disorders by directly acting at DNA level and by employing a wide array of viral or nuclease-based strategies such as gene supplementation, silencing, correction or disruption. The development of customizable DNA cleaving endonucleases such as meganucleases, zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats-associated RNA-guided Cas (CRISPRCas9) revolutionized the field of gene editing and allowed facile gene manipulation
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