Light Enables Nonviral Delivery of Prime Editing Complexes to Cardiomyocytes
Background
A cell study reported nonviral delivery of prime editing (PE) ribonucleoprotein (RNP) complexes into human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). The editing frequency measured under optimized conditions reached a maximum of 8.46%.
PE performs precise genomic modifications without inducing DNA double-strand breaks. Applying it to cardiomyocytes has posed the challenge of delivering editing tools efficiently while avoiding genomic integration of the genetic material used for delivery.
Key findings
The researchers used LumiSense nanosensitizers in a light-based delivery method called photoporation. After first confirming cytosolic delivery and the feasibility of editing in HEK293T cells, they applied the method to iPSC-CMs, which are difficult to deliver material into. The delivered material was a PE RNP complex, not PE protein alone.
Fluorescence imaging confirmed intracellular entry of PE RNPs, and prime editing frequency was measured by droplet digital polymerase chain reaction (ddPCR). The maximum under optimized conditions was 8.46%; this does not represent the average across all experimental conditions or the cellular uptake rate. The available abstract does not report the target gene, the mutation being corrected, the number of replicates, or results for each comparison group.
Significance and outlook
The findings demonstrate the potential to deliver PE RNPs into human iPSC-derived cardiomyocytes and perform editing using a physical, virus-free method. The researchers presented it as a nonintegrating, controllable delivery strategy.
However, delivery and editing results in cells do not establish therapeutic efficacy or safety for heart disease. The available abstract alone does not allow assessment of cell viability and function, off-target editing, or effects in animals or patients. It is also difficult to determine whether the method outperforms existing delivery approaches.
Source: Biomedicine & Pharmacotherapy article abstract
Prime editing (PE) offers precise genome modification without inducing double-strand breaks; however, its application in cardiomyocytes remains constrained by the lack of efficient and non-integrative delivery strategies. Here, we report a tunable photoporation-based approach for the non-viral delivery of PE ribonucleoprotein (RNP) complexes into human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), a clinically relevant model for cardiac disease. Using LumiSense nanosensitizers, we first validated cytosolic delivery and editing feasibility in HEK293T cells, and subsequently achieved efficient photoporation in hard-to-transfect iPSC-CMs. Fluorescence imaging confirmed intracellular uptake of PE RNPs, and droplet digital PCR revealed prime-editing frequencies of up to 8.46% under optimized conditions. This study demonstrates a non-integrative and controllable physical strategy for PE RNP delivery into human cardiomyocytes, providing a promising foundation for the development of genome editing-based therapeutic interventions for cardiac disorders.
Research on gene editing in cardiomyocytes must consider not only the editing technology but also how to get the tools into cells. This study presents the potential of a nonviral physical approach for delivering PE RNPs into human iPSC-derived cardiomyocytes. Follow-up studies could examine whether the approach can be applied to cardiomyocyte models carrying disease-associated variants. They would need to assess editing reproducibility at each target, cell survival and cardiomyocyte function after treatment, and off-target editing. The maximum editing frequency of 8.46% alone cannot establish whether the level is sufficient for treatment; delivery to the heart in vivo and therapeutic effects in patients require separate validation.