Undeniably, the future of therapeutics is moving away from panacea miracle drugs and gravitating towards personalized treatments using gene therapy. Furthermore, the discovery of iPSCs verified the idea that individuals could have their own stem cell line available for therapeutic purposes—therapies that are made possible by editing the genome of iPSCs.
Transcription activator-like effector (TALEN) and clustered regulatory interspaced short palindromic repeat (CRISPR)/Cas9 have ascended to fame with the speed of a reality television star due to their genome editing prowess. These endonucleases work by producing double stranded breaks at specific DNA sequences, which initiate the natural repair mechanisms of the cell. Donor template DNA template is incorporated by taking advantage of these repair mechanisms.
The success of these editing techniques in human iPSCs is dependent upon their accuracy and precision. Previous studies sounded the alarms for off target effects when using Cas9 systems (Fu et al., 2013; Hsu et al., 2013), while others reported minimal off target effects, but looked only at highly mutable regions.
That’s why we were excited to see Cory Smith and colleagues shed some much needed light on the situation. The team recently used whole genome sequencing to determine the frequency of off target mutational effects when using CRISPR/Cas9 and TALEN to edit human iPSC cell lines.
Here’s what they found:
- By aligning whole genome sequence data to a reference genome, naturally occurring SNVs and indels can be detected.
- Sequence variation between clones and parental lines can be analyzed to distinguish between mutations arising during gene editing, and natural sequence variants.
- After accounting for the naturally occurring single nucleotide variants (SNVs) and indels, Cas9 and TALEN both showed no off target effects.
The study demonstrated high specificity of these two techniques in editing human iPSCs providing more optimism for using genome editing as a potential therapy.
Still don’t believe the hype? Check out the paper at Cell Stem Cell, July 2014 (Smith, C. et al).