MAMAND, D. R. A. | Search bioRxiv

bioRxiv2026

Cell Type Dependent Uptake of Extracellular Vesicles Independent of Cellular Origin

Extracellular vesicles (EVs) are promising nanocarriers for therapeutic delivery; however, the factors governing EV uptake by recipient cells remain incompletely understood. In this study, we investigated whether EV internalization is primarily influenced by donor-cell origin or recipient-cell phenotype. Fluorescently labeled EVs derived from HEK293T, or SKBR-3 cells were incubated with a range of human epithelial, immune, and murine cancer cell lines at different doses and time points. HEK293T-derived EVs showed highly variable uptake across recipient cells, with hepatocellular carcinoma cell lines Huh7 and HepG2 exhibiting the highest internalization, while parental HEK293T cells showed the lowest. THP-1 immune cells also demonstrated strong uptake, whereas Jurkat cells showed moderate uptake. In murine melanoma models, Yummer cells internalized more EVs than B16F10 cells. Importantly, similar uptake trends were observed using SKBR-3-derived EVs, where Huh7 and HepG2 again displayed the highest uptake despite originating from a different donor cell source. EV internalization increased with dose and incubation time until saturation at higher concentrations. Together, these results demonstrate that EV uptake is predominantly determined by recipient-cell characteristics rather than EV source. These findings provide important mechanistic insight for the development of EV-based therapeutics and suggest that optimizing recipient-cell targeting is essential for efficient vesicle-mediated delivery. Graphical abstractEV uptake is determined by cell membrane properties rather than by the source of the EVs. The image was created by Biorender. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=122 SRC="FIGDIR/small/726167v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@f5c1cborg.highwire.dtl.DTLVardef@860962org.highwire.dtl.DTLVardef@1d20239org.highwire.dtl.DTLVardef@9003af_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2026

Efficient delivery of gene editors using intein-engineered virus-like particles

Virus-like particles (VLPs) represent a promising next-generation drug delivery platform. However, conventional VLPs rely on multiple viral components for effective cargo encapsulation and delivery, raising safety concerns. Here, we present a novel strategy to engineer immature VLPs using a self-cleaving intein system. We employed viral Gag proteins as sorting domains, linking cargo proteins to Gag through inteins, thereby eliminating the need for the conventional protease cleavage typically mediated by the gag-pol protein. During VLP biogenesis, intein-mediated cleavage released cargo proteins into the lumen, enabling efficient intracellular delivery when VLP surfaces are pseudotyped with VSV-G. Optimal candidates for delivering Cre recombinase and gene editing tools (Cas9, Cas12a and base editors) were identified by screening various Gag proteins. Notably, these VLPs achieved robust gene editing in primary cells, including naive and activated T cells, as well as hematopoietic stem and progenitor cells (HSPCs). A single local intracerebroventricular (ICV) infusion of optimized particles induced up to 60% tdTomato expression in the brain regions of reporter mice, while intravenous injection resulted in significant recombination (up to 70%) of a variety of cell types across organs. Collectively, we developed a simplified, efficient VLP platform for intracellular cargo delivery with broad therapeutic potential for gene editing and treatment of human diseases.

molecular biology↗