Novel gene editing technology offers efficient route to future gene therapies

A new way of precisely inserting DNA sequences which is designed to overcome some key drawbacks of current gene editing techniques is reported this week in Nature Communications. The research is aimed at expanding future opportunities to repair and restore genes for a wide range of serious genetic diseases.

Since first reports that the bacterial CRISPR-Cas9 anti-viral defence system could be programmed for gene editing in living organisms in 2013, scientists have been refining the technology with the goal of making cures for genetic diseases a reality. There has been progress in gene disruption and correcting point mutations affecting single DNA nucleotides. However, precisely targeted insertions of DNA sequences to correct gene deletions in diseases such as cystic fibrosis have proved particularly challenging, because the current techniques mostly rely on DNA repair pathways that are poorly understood or not widely available across human tissues.  

Here in the Genome Engineering department at AstraZeneca R&D, our aim is to develop novel methods of promoting gene repair, with the ambition of curing genetic diseases. In our latest research paper, in Nature Communications, we report advances in DNA insertion techniques including the use of non-homologous end joining (NHEJ) – the primary pathway that human cells use for daily DNA repair.1

Introducing PEn – a new prime editor

Our new approach to targeted DNA insertions builds on a recently developed prime editing technology and uses prime editor nuclease (PEn). This combines the DNA cutting SpCas9 nuclease enzyme with a reverse transcriptase – an enzyme that catalyses transcription of RNA into DNA. We show that this approach can utilise a spectrum of efficient DNA repair mechanisms including NHEJ which robustly introduces programmed small insertions at DNA double strand breaks.

Importantly, we showed that, in contrast to conventional CRISPR-Cas9, PEn does not cause large unintended deletions at the target site which can arise when Cas9 keeps cutting DNA in a poorly controlled way. This is because PEn efficiently disrupts Cas9’s cutting cycle by destroying its binding site after a successful DNA insertion.

After two years of challenging research – much of it carried out under the demanding working conditions of the COVID-19 pandemic – it is hugely satisfying to be reporting robust strategies for promoting robust DNA insertions based on prime editing, which address the drawbacks of previous gene editing techniques. For example, a commonly used approach to replace faulty DNA, involving homology-directed repair, had the drawback that the mechanism was not active throughout the cell cycle, and previous attempts using the universally active NHEJ were typically prone to errors.

Our research, led by postdoctoral scientist, Martin Peterka, through the highly successful Postdoc Programme in our laboratories, supports PEn’s potential for precise, efficient DNA insertions.

Moving forward

The next step is to move our PEn research from the in vitro to the in vivo setting. For this, we will be able to draw on our well-established expertise in novel delivery systems, such as lipid nanoparticle technology. We are also building on the concept of localised NHEJ to develop novel methods to promote gene insertions in our therapeutic genome editing projects.

We are delighted to be collaborating with academic partners to use the PEn technology in their research. Thanks to the expert support and state of the art technologies available through the Postdoc Programme, we have been able to contribute not only to progress with AstraZeneca’s own ambitions for curing genetic diseases, but to the efforts of the wider research community.

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1. Peterka M, Akrap N, Li S,, Wimberger S, et al. Harnessing DSB repair to promote efficient homology-dependent and -independent prime editing. Nature Communications 2022; March 24th

Veeva ID: Z4-43292
Date of preparation: March 2022