Yanni Lin, TJ Cradick, Gang Bao and colleagues from Georgia Tech and Emory reported recently in Nucleic Acids Research on how the CRISPR/Cas9 gene editing system can sometimes miss its mark.
CRISPR/Cas9 has received abundant coverageÂ fromÂ science-focused mediaÂ outlets.Â Basically, it is a convenient system for cutting DNA in cells in a precise way. This paper shows that the CRISPR/Cas9 system can sometimes cut DNA in places that donâ€™t exactly match the designed target.
Here we show that CRISPR/Cas9 systems can have off-target cleavage when DNA sequences have an extra base or a missing base at various locations compared with the corresponding RNA guide strandâ€¦Our results suggest the need to perform comprehensive off-target analysis by considering cleavage due to DNA and sgRNA bulges in addition to base mismatches.
CRISPR/Cas9 could be used to develop therapies for humans for genetic blood diseases such as sickle cell or thalassemia, and this paper does not change that potential. But the authors are cautioning fellow scientists that they need to design their tools carefully and perform quality control. Other investigators have made similarÂ findings.
An excellent example of the use of CRISPR gene editing technology came up at the Emory-Children’s Pediatric Research Center’s Innovation Conference this week.
Marcela Preininger, who is working with cardiomyocyte stem cell specialist Chunhui Xu, described her work (poster abstract 108) on cells derived from a 12 year old patient with an inherited cardiac arrhythmia syndrome: catecholaminergic polymorphic ventricular tachycardia or CPVT. Her team has obtained skin fibroblasts from the patient, and converted those cells into induced pluripotent stem cells, which can then be differentiated into cardiac muscle cells or cardiomyocytes.
Working with TJ Cradick, director of the Protein Engineering Facility at Georgia Tech, Preininger is testing out CRISPR gene editing as a means of correcting the defect in this patient’s cells, outside the body. Cradick says that while easy and efficient, RNA-directed CRISPR can be lower in specificity compared to the protein-directed TALEN technology.
From Preininger’s abstract:
Once the mutation has been corrected at the stem cell level, we will investigate whether the repaired (mutation-free) iPS cells can be differentiated into functional cardiomyocytes with normal Ca2+ handling properties, while closely monitoring the cells for mutagenic events. Pharmacological restoration of the normal myocardial phenotype will also be optimized and explored in our model.