Barriers to horizontal cell transformation by extracellular vesicles containing oncogenic H-ras

Abstract

Extracellular vesicles (EVs) enable the exit of regulatory, mutant and oncogenic macromolecules (proteins, RNA and DNA) from their parental tumor cells and uptake of this material by unrelated cellular populations. Among the resulting biological effects of interest is the notion that cancer-derived EVs may mediate horizontal transformation of normal cells through transfer of mutant genes, including mutant ras. Here, we report that H-ras-mediated transformation of intestinal epithelial cells (IEC-18) results in the emission of exosome-like EVs containing genomic DNA, HRAS oncoprotein and transcript. However, EV-mediated horizontal transformation of non-transformed cells (epithelial, astrocytic, fibroblastic and endothelial) is transient, limited or absent due to barrier mechanisms that curtail the uptake, retention and function of oncogenic H-ras in recipient cells. Thus, epithelial cells and astrocytes are resistant to EV uptake, unless they undergo malignant transformation. In contrast, primary and immortalized fibroblasts are susceptible to the EV uptake, retention of H-ras DNA and phenotypic transformation, but these effects are transient and fail to produce a permanent tumorigenic conversion of these cells in vitro and in vivo, even after several months of observation. Increased exposure to EVs isolated from H-ras-transformed cancer cells, but not to those from their indolent counterparts, triggers demise of recipient fibroblasts. Uptake of H-ras-containing EVs stimulates but fails to transform primary endothelial cells. Thus, we suggest that intercellular transfer of oncogenes exerts regulatory rather than transforming influence on recipient cells, while cancer cells may often act as preferential EV recipients.

The Impact of Oncogenic EGFRvIII on the Proteome of Extracellular Vesicles Released from Glioblastoma Cells

“…S6). Nonetheless, the cumulative ability to uptake EVs was greater in the case of EGFRvIII-transformed glioma cells, which is consistent with the role of oncogeneses in regulating EV uptake (58,59). Thus, EGFRvIIImediated oncogenic transformation alters the magnitude and specificity of the EV internalization by changing the surface properties of both EVs and cellular recipients.…”

“…Genetic and epigenetic driver events have already been implicated by us and others as regulators of the EV release and intercellular communication in cancer, including intercellular trafficking of transforming mutant macromolecules, such as HRAS, EGFR and EGFRvIII (12,17,18,58,62).…”

Molecular subtypes and differentiation programmes of glioma stem cells as determinants of extracellular vesicle profiles and endothelial cell‐stimulating activities

“…EVs isolated from cells were labelled with PKH26 (#MINI26, Sigma-Aldrich) red fluorescent dye as described earlier [33,34]. Briefly, EVs were re-suspended in 250 μL of 2× Diluent C. The 2 × Dye Solution (4 × 10 –6  M) in Diluent C is prepared by adding 1 μL of the PKH26 ethanolic dye solution (# P9691) to 250 mL of Diluent C in a polypropylene centrifuge tube and mixed well to disperse.…”

Extracellular vesicle docking at the cellular port: Extracellular vesicle binding and uptake

“…Interestingly, the interaction and uptake of EVs by recipient cells may be dependent on the specific properties of the recipient cells. For example, epithelial cells and astrocytes were unable to internalize EVs from transformed cells [43]. However, when the same cell types were transformed through ectopic expression of oncogenic forms of Ras or c-Src, they efficiently internalized the EVs [43].…”