Optimization of production and isolation of exosomes in controlled bioreactor used for therapeutic purposes
Project no. 22868 · Status: Kapanmış · 2021
Summary
<p>Nanoparticular carrier systems are particles prepared from substances that do not harm the body, ideally 10-100 nm in size, that protect the drug or antigenic materials they carry from enzymes in the serum. Liposomes, polymeric nanoparticles, emulsions, virus-like particles, inorganic nanoparticles, and exosomes are examples of carriers. Exosomes are called natural liposomes and they are nanovesicles 20-100 nm in size released from body cells. Since the discovery of exosomes, research continues their use both as a carrier system and as a tool for early diagnosis and treatment of various diseases. However, the problems related to the production and isolation of sufficient exosomes for clinical use, and various loading strategies have not been fully resolved. In this project, the adaptation of human monocyte cell line THP-1 to a chemically defined medium that does not contain serum and animal protein (FBCD), optimization of the production of cells in a large scale, optimization of the ultrafiltration method, which is more suitable for large scale during exosome isolation from cells, characterization and determination of toxicity were performed. While THP-1 cells were adapted to the FBCD medium with gradual adaptation, it was determined that when the serum ratio fell below 5%, the cells were forced and could adapt by passing more. Adapted cells were grown in the erlenmayers to determine the initial cell concentration and then in the spinner flasks to determine the mixing speed. By keeping the specific power input constant, the 1L working volume stirred tank bioreactor was scaled up and the production parameters were optimized. It was determined that feeding the cell supernatant to the cross-flow ultrafiltration method with a flow rate of 125 rpm was optimal for exosome isolation. It was determined that the isolated exosomes were 92.5 ± 5.51 nm in size and did not show toxicity on L929 cells.</p>