19.10.2020

Adenine base editing

Retinal pigment epithelium (RPE) located at the back of the eye is essential for vision. It supports the photoreceptors providing molecules required for their function. One of the main proteins produced by RPE and indispensable for vision is the RPE65 enzyme, which is responsible for chemical signaling at the initial step of visual processing. Mutation in the Rpe65 gene underlies an inherited genetic disorder of the eyes resulting in blindness. Here, we harness the power of adenine base editors (ABEs) with Cas9 – single-guide RNA machinery to target de novo nonsense mutations in the Rpe65 gene for their repair. We delivered genes coding for ABEs and Cas9 system by lentiviral vector subretinally. Our therapeutic manipulation corrected the pathogenic mutation with up to 29% efficiency and with minimal formation of indel and off-target mutations. The ABE-treated mice displayed restored RPE65 expression and its activity in the visual cycle. Moreover, we have observed near-normal levels of retinal and visual functions. Our findings motivate the further testing of ABEs for the treatment of inherited retinal diseases and for the correction of pathological mutations with non-canonical protospacer-adjacent motifs.

Authors:

dr Andrzej Foik, e-mail: afoik@ichf.edu.pl & dr Anna Posłuszny, e-mail: aposluszny@ichf.edu.pl

Related paper:

Restoration of visual function in adult mice with an inherited retinal disease via adenine base editing

Susie Suh, Elliot H. Choi, Henri Leinonen, Andrzej T. Foik, Gregory A. Newby, Wei-Hsi Yeh, Zhiqian Dong, Philip D. Kiser, David C. Lyon, David R. Liu & Krzysztof Palczewski

Abstract

Cytosine base editors and adenine base editors (ABEs) can correct point mutations predictably and independent of Cas9-induced double-stranded DNA breaks (which causes substantial indel formation) and homology-directed repair (which typically leads to low editing efficiency). Here, we show, in adult mice, that a subretinal injection of a lentivirus expressing an ABE and a single-guide RNA targeting a de novo nonsense mutation in the Rpe65 gene corrects the pathogenic mutation with up to 29% efficiency and with minimal formation of indel and off-target mutations, despite the absence of the canonical NGG sequence as a protospacer-adjacent motif. The ABE-treated mice displayed restored RPE65 expression and retinoid isomerase activity, and near-normal levels of retinal and visual functions. Our findings motivate the further testing of ABEs for the treatment of inherited retinal diseases and for the correction of pathological mutations with non-canonical protospacer-adjacent motifs.

Link to publication

https://doi.org/10.1038/s41551-020-00632-6