Yeast extracellular vesicles and their application as a new-generation drug delivery platform

The extracellular vesicles could be considered as cell-derived double-layer phospholipid membrane particles, excreted by cells of all organisms. They are able to secrete different macromolecules, including proteins, nucleic acids, lipids, polysaccharides, etc. This natural feature enables them to participate in plenty of physiological and pathological events, including intercellular communication and disease development in humans and animals. Extracellular vesicles, derived from pathogenic yeast species, could be a transmitter of various virulence factors, which may be harmful to the host organism. Nevertheless, slightly known about the vesicles secreted by probiotic yeast. A few studies have shown that they do not negatively influence human cells, therefore it is possible that they could be effectively used as a diagnostic, therapeutic tool, and even drug delivery platform, including being carrier for various vaccines. The overall topic of my Master's thesis project was an investigation of the biotechnological properties of yeast extracellular vesicles. Especially, the studies were done on the two world widely known probiotic yeast species, such as: Kluyveromyces marxianus (WUT 240 strain, Warsaw University of Technology Yeast Collection) and Saccharomyces boulardii (CNCM I-745 strain). Clearly known from the literature, that described yeast species are not harmful to humans, and have probiotic features. Should be mentioned, that initially we seeded cultures on two types of media: Sabouraud Dextrose Agar (SAB), and Yeast Extract Peptone Dextrose (YPD), and perform subsequent isolation of EVs from the culture liquid. The isolated EVs have been further subjected to various studies to estimate their size and loading efficacy. As well as was conducted series of trials, in order to develop an effective method of insertion of the selected biologically active substance (doxorubicin) inside the vesicles, with subsequent contemplation of releasement kinetics of doxorubicin substance out of extracellular vesicles under the application of different physicochemical conditions. The parameters were fairly similar to the human physiological values of the gastrointestinal tract, such as pH, the temperature of incubation, etc. The final investigation step was done to understand the activity of EVs, prefilled with doxorubicin, on the human colorectal adenocarcinoma cell lines (HT-29 and HCT 116). Generally, the conducted research showed the valuable properties of extracellular vesicles derived from Kluyveromyces marxianus and Saccharomyces boulardii, in the role of the carrier for selected biologically active substances. Undoubtedly, should be performed additional rounds of trials to deeply understand the properties of vesicles, as a drug delivery platform.