CRISPR-Heart: Engineering CRISPR editors for modeling and treating heart disease

• Applicant: Professor Dr. Julian Grünewald
• Subject Area: Cardiology, Angiology
• Term: since 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 502106724

A number of heart diseases would be prime candidates for gene editing approaches, including some forms of dilated cardiomyopathy (DCM) which is considered to be the leading cause of heart transplantation. CRISPR gene editing holds enormous potential for the treatment of genetic diseases. Current phase I trials show promising first results in humans. Base editors (BEs) and prime editors (PEs) are new platforms that function via CRISPR-guided deamination or reverse transcription, respectively. Together, they enable the precise introduction of any single-base substitution, small insertion, or deletion while avoiding DNA double-strand breaks and subsequent insertion/deletion byproducts (indels). However, these new editors come with new challenges, including larger sizes and differences in editing efficiencies between different cell types which may impede their translational use. For disease modeling in cardiology, new editors are commonly applied in induced pluripotent stem cells (iPSCs) that are subsequently differentiated to cardiomyocytes (iPSC-CMs). Yet, to develop clinically relevant gene editing strategies, the direct testing and optimization of these editors in heart cells would be transformative. Therefore, I aim to expand the therapeutic potential of CRISPR nucleases, BEs, and PEs for treating heart diseases, such as DCM, by (1) profiling these editors, their delivery, and functional outcomes after gene editing directly in iPSC-CMs as well as in human ex vivo heart slices. In addition, I will (2) engineer new editors with reduced sizes for enhanced delivery and I will use both transcriptomic profiling and CRISPR interference screens to elucidate how DNA damage response and repair pathways affect gene editing outcomes specifically in cardiomyocytes. Targeted modulation of these pathways will then be used to develop CRISPR editors that are custom-engineered for highly efficient and precise gene editing in cardiomyocytes. Finally, I will (3) use the latest CRISPR base and prime editing technologies to model and optimize a genetic rescue strategy for a DCM-causing TTN mutation in iPSC-CMs and a mouse model. With this approach, I aim to investigate the relationship between gene editing frequencies and phenotypical outcomes. Moreover, I will assess the optimal timing (i.e. age) for therapeutic gene editing to rescue the DCM phenotype in this mouse model. In sum, this project - CRISPR-Heart - will unlock highly efficient and precise „DNA interventions“ for heart disease.

• DFG Programme: Independent Junior Research Groups