Biomarker Study on Oral Cancer Exosomes

Objectives: Extracellular vesicles (EVs) are released by almost all cell types. They are involved in intracellular communications as vehicles for transfer of functional membranes, cytosolic proteins, lipids, RNAs and DNA. EVs are classified according to their mechanism of origin, composition, size and density. Exosomes are nano-size particles, measuring 40-100 nm. Depending on their origin they are capable of altering the fate of recipient cells through the transferred information. In cancer biology exosomes offer both diagnostic and therapeutic advantage. Their involvement in cell-cell communication indicates their influence in tumour development, progression, metastasis and therapeutic efficacy. Exosomes released by cancerous cells carry numerous biomarkers, which are passed on to healthy cells. In this study we characterised and investigated the functional significance of exosomes derived from SVpgC2a (oral epithelial pre-cancerous) and SVFN10 (transformed) cell lines.
Methods: The characteristics of exosomes were first confirmed by Scanning Electron Microscopy, Transmission Electron Microscopy, Zetasizer and Nanosight Tracking Analysis, along with western blot for specific exosomal membrane proteins. Further we validated the presence of RNA within exosomes that remained stable after treatment with proteinase K and RNase. The quality and size distribution of RNA extracted from exosomes were analysed on Agilent BioAnalyser.
Results: We found that, similar to parental cell line SVFN10, exosomes derived from SVFN10, but not SVpgC2a, were abundant in FOXM1 mRNA, an oncogene involved in cancer initiation and progression. Exosomes derived from SVFN10 were found to be 20-50% larger than exosomes derived from SVpgC2a cells. Exposure of SVpgC2a cells with exosomes derived from SVFN10 induced a distinct morphological change in the SVpgC2a cells within 24 hours. Exposure of SVpgC2a cells with its own exosomes or exosome-free supernatant did not induce significant morphological change.
Conclusions: This is the first evidence showing the presence of oncogene mRNA within exosomes secreted by a transformed malignant oral keratinocyte cell line. This finding has tremendous potential for clinical translation into a non-invasive oral cancer diagnostic tool.