Beyond precision: evaluation of off-target clustered regularly interspaced short palindromic repeats/Cas9-mediated genome editing.

Abstract

The gene editing field has advanced rapidly since the development of the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system because of its applicability in precisely modifying the genome. Among its multiple applications, the correction of genetic diseases has emerged as a potential curative treatment for many disorders that have eluded a cure to date. Despite its efficiency in achieving therapeutic levels of correction, the unexpected adverse effects of editing due to CRISPR/Cas9 nuclease activity are a major concern when translating these new strategies to the clinic. Multiple in silico tools and empirical methods have been developed to evaluate these off-target edits as well as other adverse alterations of the genome, including rearrangements, not only in ex vivo experiments but also in in vivo experiments. In this review, we summarize the available methods for the assessment of off-target effects of CRISPR/Cas9 systems, highlighting their advantages and limitations. Special attention will be paid to their application in pre-clinical studies and clinical trials, both in the manufacturing product and in the long-term follow-up of patients.