Efficient, Scalable Manufacturing Of Virus-Like Particles For The Delivery Of Gene Editor Ribonucleoproteins Using A GMP-Compatible Electroporation Platform

By Isabel Daher, Tien Nguyen, Peter Gee, Sean Menarguez, Andrew Mancini, Marianna Romito, Camille Ciesliga, Ashley B. Strickland-Dietz, Lauren E. Unsworth, Jason Papademetriou, and James Brady – MaxCyte, Inc., Rockville, MD, USA

Genome editing tools like CRISPR-Cas9, base editors, and prime editors offer immense potential for treating genetic diseases. However, safe and efficient in vivo delivery remains a significant challenge. Viral vectors, while effective, have limitations in cargo size.

Virus-like particles (VLPs) provide a promising solution. Derived from retroviral proteins, VLPs can be engineered to carry specific genetic payloads. They are safer than traditional viral vectors as they lack a viral genome but can still leverage the host cell's machinery for efficient delivery.

Recent studies have shown that VLPs can effectively deliver base and prime editors to mice, opening doors for therapeutic applications. To fully realize this potential, scalable manufacturing methods are crucial.

In this study, we utilized the MaxCyte® ExPERT GTx™ electroporation system to produce VLPs loaded with CRISPR-Cas9 or base editor RNPs. This method significantly outperformed traditional transfection reagents, yielding over 10-fold higher VLP titers. We demonstrated successful genome editing at the B2M locus using both CRISPR-Cas9 and base editor VLPs.

Electroporation offers several advantages:

• Efficiency: Higher VLP yields and enhanced editing efficiency.
• Speed: Shorter production timelines.
• Scalability: Consistent performance across a wide volume range.

Our findings highlight the potential of electroporation as a robust and scalable method for VLP production. This advancement brings us closer to the clinical application of VLP-based gene therapies.