ATR-FTIR spectroscopy as a reliable analytical tool in cancer research: Tracking glycosylation-induced protein and lipid alteration in extracellular vesicles.

The rising incidence of cancer creates an urgent need to develop faster methods for early detection. Fourier Transform Infrared Spectroscopy (FTIR) spectroscopy is a promising technique to analyse molecular content in biological samples. FTIR has also been used to study structural changes in extracellular vesicles (EVs), revealing alterations in protein secondary structure, protein-to-lipid ratio, and protein phosphorylation levels. We investigated how changes in glycosylation, which naturally occur during tumor progression, affect the previously mentioned parameters and whether these cellular changes are reflected in EV composition. The WM266-4 melanoma cell line, treated with two glycosylation inhibitors, was used as a model. After treatment, EVs were isolated and ATR-FTIR measurements were performed, followed by a comprehensive analysis of the data. Significant differences in protein and lipid content were observed between EV populations treated with glycosylation inhibitors, with greater changes in EVs compared to cells. Glycosylation inhibitors also increased the level of phosphorylated proteins in EVs. Our findings showed that changes in EVs were more dynamic than at the cellular level cells. The presented analysis shows that subtle molecular changes can significantly impact EV cargo and properties. FTIR is a powerful tool for detecting these changes and may aid future cancer diagnostics.