Scientists discovered that prime editing in human pluripotent stem cells (hPSCs) can greatly complement existing clustered regularly interspaced short palindromic repeats (CRISPR) genome editing tools to potentially treat alpha-1 antitrypsin deficiency (AATD), as published in Nucleic Acids Research.

hPSCs have the potential to differentiate into cells that come from the 3 embryonic germ layers. As such, their use in genome editing has resulted in vast progress in disease modeling, developmental biology, and the development of new stem cell-based therapeutics.

The CRISPR-Cas9 system is a major existing genome editing tool. It works by generating a double strand break (DSB) in DNA sequences, triggering a response from the cell’s repair systems and leading to targeted genome editing. The main sticking point is that even a single DSB can result in chromosomal rearrangements, depletions, or deletion, especially in hPSCs, leading to apoptosis.


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DNA base editors are able to generate single-nucleotide substitutions without inducing a DSB. Prime editors, on the other hand, are genome editing tools that can introduce all kinds of base-to-base conversions, as well as small insertions and deletions without introducing a DSB.

“We comprehensively tested prime editing in hPSCs by generating a doxycycline-inducible prime editing platform,” Habib and colleagues wrote. “Moreover, we compared prime editing and base editing for correcting a disease-related mutation in induced pluripotent stem cells derived from a patient with AATD.”

The researchers also conducted whole-genome sequencing, which demonstrated that the reverse transcriptase domain of a prime editor does not cause unintended mutations in the genome.

“In summary, prime editing adds a new dimension to CRISPR-based approaches and broadens the scope of genome editing in hPSCs by complementing other methods,” the authors concluded.

“It is likely that an understanding of the repair mechanisms involved in prime editing and further improvement of prime editors will accelerate translation of this technology into therapeutic applications.”

Reference

Habib O, Habib G, Hwang GH, Bae S. Comprehensive analysis of prime editing outcomes in human embryonic stem cellsNucleic Acids Res. 2022;50(2):1187-1197. doi:10.1093/nar/gkab1295