ACS biomaterials science & engineeringJun 24, 2026

Creating Engineered Cell Factories for Large-Scale Production of Modified Tiny Cell Particles Using Gene Editing

CRISPR Gene Editing Weekly Brief ↗PubMed ↗DOI ↗OA ↗

Updated Jun 25, 2026

Abstract

Engineered cells produced extracellular vesicles (EVs) with a uniform size of 120-130 nm.

A synthetic-biology-based manufacturing platform was developed using CRISPR-Cas9 to create stable HEK293T cell lines for continuous EV production.
The system integrates a fusion construct into the AAVS1 safe-harbor locus, allowing for consistent expression of EV membrane proteins.
EVs produced from these engineered cells preserved canonical markers (CD63 and ALIX) and displayed enhanced surface features compared to transiently transfected controls.
These vesicles exhibited effective cellular uptake and maintained their structure and functionality over more than 25 passages (approximately 3 months).
The platform addresses issues of batch-to-batch variability associated with transient production methods, providing a reliable approach for engineered EV biomanufacturing.

Simplified

Extracellular vesicles (EVs) are versatile biological nanoparticles with applications in therapeutics, diagnostics, and biotechnology. Current production methods relying on transient transfection or chemical conjugation suffer from high variability, limited scalability, and heterogeneous EV populations. Here, we present a synthetic-biology-based biomaterial manufacturing platform that uses CRISPR-Cas9 genome editing to generate stable HEK293T cell lines for continuous production of surface-functionalized EVs. A fusion construct encoding mCherry-C1C2 was site-specifically integrated into the AAVS1 safe-harbor locus, enabling consistent and heritable expression of EV membrane proteins without repeated transfection. Engineered cells produced EVs with uniform size (120-130 nm), preserved canonical markers (CD63 and ALIX), and enhanced surface-display efficiency compared with transiently transfected controls. These vesicles exhibited robust cellular uptake and maintained structural and functional stability for over 25 passages (∼3 months), confirming durable genome-encoded production. Overall, this platform eliminates batch-to-batch variability inherent to transient systems and provides a genetically defined route to biofunctional nanomaterial fabrication. This approach links genetic design to nanoscale surface functionality, establishing a versatile foundation for reproducible biomanufacturing of engineered EVs for biomaterial, therapeutic, and diagnostic applications.

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Creating Engineered Cell Factories for Large-Scale Production of Modified Tiny Cell Particles Using Gene Editing

Abstract

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