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Towards therapeutic use of CRISPR/Cas9-mediated prime editing in the brain

Applicant Böck Desiree
Number 196287
Funding scheme Spark
Research institution
Institution of higher education University of Zurich - ZH
Main discipline Genetics
Start/End 01.04.2021 - 31.03.2022
Approved amount 94'500.00
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All Disciplines (2)

Neurophysiology and Brain Research

Keywords (5)

CRISPR/Cas9; prime editing; gene therapy ; neurodegeneration; Alzheimer's disease

Lay Summary (German)

Neurodegenerative Erkrankungen führen zum unvermeidlichen Absterben von Neuronen. Mutationen tragen zum Auftreten und Fortschreiten vieler Gehirnerkrankungen bei. Deren Korrektur mittels CRISPR/Cas9 stellt daher eine vielversprechende Therapie dar. Dieses Projekt beschäftigt sich mit der Etablierung von CRISPR/Cas9 als sichere und effiziente Therapie für Gehirnerkrankungen.
Lay summary

Inhalt und Ziele des Forschungsprojektes

Neurodegenerative Erkrankungen werden von Genetik sowie Umwelt eines Menschen beeinflusst.  Die Ursache vieler Gehirnerkrankungen ist unklar, was die Entwicklung einer Therapie erschwert. Aktuelle Therapien beschränken sicher auf die Behandlung von akuten Symptomen, die eine temporäre Erleichterung der Symptomatik bringt, aber keine Heilung oder langfristige Verbesserung des Krankheitszustandes.

Das Projekt versucht diese Lücke mit CRISPR/Cas9 zu füllen. In der ersten Phase wird das Potential von CRISPR/Cas9, spezifische Veränderungen im Genom einzuführen, in humanen Zelllinien getestet. In einer zweiten Phase werden die getesteten CRISPR/Cas9 Komponenten in Krankheits-Mausmodellen getestet und deren therapeutische Wirksamkeit evaluiert. Unsere Experimente könnten wichtige Bausteine für die Etablierung von CRISPR/Cas9 Therapien für Gehirnerkrankungen legen.

Wissenschaftlicher und gesellschaftlicher Kontext des Forschungsprojekts

Aufgrund des kontinuierlich steigenden Alters der Weltbevölkerung, ist ein Anstieg an Patienten mit altersbedingten neurodegenerativen Erkrankungen zu erwarten. Dieser Anstieg ist stellt eine wirtschaftliche Belastung für das Gesundheitssystem dar. Das beschriebene Projekt versucht diesem Trend mit einer wirksamen Therapie entgegenzuwirken.

Direct link to Lay Summary Last update: 20.12.2020

Responsible applicant and co-applicants


In vivo prime editing of a metabolic liver disease in mice
Böck Desirée, Rothgangl Tanja, Villiger Lukas, Schmidheini Lukas, Matsushita Mai, Mathis Nicolas, Ioannidi Eleonora, Rimann Nicole, Grisch-Chan Hiu Man, Kreutzer Susanne, Kontarakis Zacharias, Kopf Manfred, Thöny Beat, Schwank Gerald (2022), In vivo prime editing of a metabolic liver disease in mice, in Science Translational Medicine, 14(636), abl9238.


Treatment of a metabolic liver disease by in vivo prime editing in mice

Author Böck, Desiree
Publication date 16.03.2022
Persistent Identifier (PID) 10.1126/scitranslmed.abl9238
Repository Gene Expression Omnibus Database (GEO)
Prime editing is a highly versatile CRISPR-based genome editing technology that works without DNA double-strand break formation. Despite rapid technological advances, in vivo application for the treatment of genetic diseases remains challenging. Here, we developed a size-reduced SpCas9 prime editor (PE) lacking the RNaseH domain (PE2ΔRnH) and an intein-split construct (PE2 p.1153) for adeno-associated virus-mediated delivery into the liver. Editing efficiencies reached 15% at the Dnmt1 locus and were further elevated to 58% by delivering unsplit PE2ΔRnH via human adenoviral vector 5 (AdV). To provide proof of concept for correcting a genetic liver disease, we used the AdV approach for repairing the disease-causing Pahenu2 mutation in a mouse model of phenylketonuria (PKU) via prime editing. Average correction efficiencies of 11.1% (up to 17.4%) in neonates led to therapeutic reduction of blood phenylalanine, without inducing detectable off-target mutations or prolonged liver inflammation. Although the current in vivo prime editing approach for PKU has limitations for clinical application due to the requirement of high vector doses (7 × 1014 vg/kg) and the induction of immune responses to the vector and the PE, further development of the technology may lead to curative therapies for PKU and other genetic liver diseases.


Group / person Country
Types of collaboration
Patriarchi group/University of Zurich Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Research Infrastructure
- Exchange of personnel


Neurodegenerative diseases (NDs) are terminal brain disorders caused by the progressive loss of neurons. Since genetic mutations strongly contribute to the onset and progression of most NDs, CRISPR/Cas9-based genome editing represents an attractive approach to treat these diseases. The classical CRISPR/Cas9 technique allows precise editing via homology directed repair (HDR) of DNA double strand breaks (DSBs) almost exclusively in mitotic cells. In contrast, the CRISPR/Cas9-derivatives base- and prime editing enable precise editing independent of HDR. While base editing performs only C to T and A to G conversions within a specific range from the protospacer adjacent motif (PAM), prime editing can introduce virtually any modifications to the genome, also distal to the PAM. Hence, prime editing represents a promising tool for gene repair in organs mainly consisting of postmitotic cells, such as the brain.Gene editing via conventional CRISPR/Cas9 endonucleases is currently tested in a handful of clinical trials, mainly for ex vivo therapy of blood disorders. In vivo genome editing in patients still faces several challenges, including limited efficiency in delivering expression cassettes large enough to encode for genome editing components, immunogenicity of Cas proteins and generation of off-target mutations that could induce oncogenic transformation. In mouse models adeno-associated vectors (AAVs) have previously enabled in vivo genome editing in different tissues using classical Cas9 nucleases as well as adenine and cytosine base editors. Protocols for efficient in vivo delivery of prime editors are yet to be developed. The proposed project addresses several of the aforementioned challenges in order to establish prime editing approaches for the treatment of brain disorders such as Alzheimer’s disease (AD). AD is characterized by the formation of amyloid plaques in the brain, leading to a loss of neuronal populations. Plaque formation is driven by mutations in several genes, e.g. amyloid precursor protein (app), resulting in pathological breakdown of the protein and subsequent aggregation. A protective mutation in app has recently been identified in Nordic populations and seems to confer resistance to amyloid plaque formation. Hence, we will first establish prime editor systems that efficiently introduce this protective mutation in the app locus in an in vitro cell model. In a parallel approach, we will reverse the most common AD mutations in the app locus. Effects on amyloid aggregation will be compared between the two approaches in an in vitro cell model. Second, we will use a previously established fluorescent reporter mouse model to test different viral vectors for in vivo delivery of prime editors to the brain. Last, we will assess the efficacy of the most promising approach in vivo in an AD mouse model with regard to amyloid plaque levels. Since AD is a multifactorial brain disorder where several mutations contribute to disease onset and progression, conventional genome editing approaches face the obstacle of correcting multiple mutations in different genes. The introduction of a protective mutation thus represents a promising, feasible and unconventional approach for treatment of this complex genetic disorder. Especially since AD and other age-related NDs will affect more people in the coming decades, the development of new and innovative therapies constitutes an important scientific and economic priority. In conclusion, the combined approach of prime editing and targeted viral delivery holds immense potential for future treatments of genetic diseases even beyond neurodegeneration.