Authors
Yuki Kawai-Harada, SunYoung You, Thomas Scarborough, Nayeema Siraj, Jayadeep Yedla, Tiffany Rennells, S Patrick Walton, Christina Chan, Masako Harada
Published in
ACS biomaterials science & engineering. Jun 24, 2026. Epub Jun 24, 2026.
Abstract
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.
PMID:
42340184
Bibliographic data and abstract were imported from PubMed on 24 Jun 2026.
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