Supramolecular nanosubstrate-mediated delivery system enables CRISPR-Cas9 knockin of hemoglobin beta gene for hemoglobinopathies.
basic_science · Level V
Where this comes from
- Record sourced from PubMed, PMID 33097539.
- Also identified by DOI 10.1126/sciadv.abb7107 and PMC identifier 7608838.
- Licence recorded as CC BY-NC.
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Abstract
Leveraging the endogenous homology-directed repair (HDR) pathway, the CRISPR-Cas9 gene-editing system can be applied to knock in a therapeutic gene at a designated site in the genome, offering a general therapeutic solution for treating genetic diseases such as hemoglobinopathies. Here, a combined supramolecular nanoparticle (SMNP)/supramolecular nanosubstrate-mediated delivery (SNSMD) strategy is used to facilitate CRISPR-Cas9 knockin of the hemoglobin beta (HBB) gene into the adeno-associated virus integration site 1 (AAVS1) safe-harbor site of an engineered K562 3.21 cell line harboring the sickle cell disease mutation. Through stepwise treatments of the two SMNP vectors encapsulating a Cas9•single-guide RNA (sgRNA) complex and an HBB/green fluorescent protein (GFP)-encoding plasmid, CRISPR-Cas9 knockin was successfully achieved via HDR. Last, the HBB/GFP-knockin K562 3.21 cells were introduced into mice via intraperitoneal injection to show their in vivo proliferative potential. This proof-of-concept demonstration paves the way for general gene therapeutic solutions for treating hemoglobinopathies.
Medical subject headings
- CRISPR-Cas Systems
- Hemoglobinopathies