CHARACTERIZATION OF CANCER CELL BEHAVIORS AFTER WASHOUT FROM DIFFERENT MICROTUBULE-TARGETING AGENTS

Abstract

Paclitaxel (Taxol), Epothilone B, Nocodazole, and Vinorelbine are the four microtubule-targeting agents widely used in clinics and research as anticancer chemotherapeutic drugs. These drugs have microtubules stabilizing and destabilizing effects, inducing mitotic arrest. As a result of prolonged mitotic arrest, some cells undergo apoptosis in their mitotic stage, whereas surviving cells undergo mitotic slippage or abnormal division and become multinucleated. It was generally thought that multinucleated cells are short-lived and eventually die. However, some studies reported that multinucleated cells produce mononucleated aneuploid progeny, which might be linked to a cancer relapse and tumorigenesis. Also, there is a scientific gap associated of how the drug clearance, a physiological washout of drug from the body, changes the cell behavior and affects clonogenic properties of cancer cells. This thesis aimed to investigate the differences in the behavior of cancer cells as a result of mitotic arrest between the washout and non-washout cell groups. Also, the difference in the behavior seems to be related to the type of drug applied. Finally, the clonogenic potential of the cells after drug washout is studied. Overall, there is a lack of data on the effect drug clearance has on cancer cell behavior and clonogenic capacities. Thus, the frequency of cell division, the time of first colony proliferation, and the ploidy of cancer cells treated with microtubule-targeting agents were studied in-depth. The methods included time-lapse microscopy observations, long-term microscopy observations and flow cytometry. After the drug washout the abnormal division event becomes predominant while non-washout cell groups experience more mitotic slippage. The four microtubule-targeting drugs (Nocodazole, Vinorelbine, Paclitaxel, and Epothilone B) induced different cell behavior after drug washout, various clonogenicity results due to their various modes of action on microtubules. While Nocodazole and Vinorelbine was not efficient to block normal mitosis, the Paclitaxel and Epothilone B drugs were capable to a more extent to shift cells to the abnormal division and mitotic slippage. As a result, the latter two drugs were efficient to shift cancer cell population to the multinucleated state which is featured with low clonogenic properties and rise of mononuclear colonies after relatively long period of time. The obtained results are beneficial in understanding the cell fate of cancer cells after drug washout and obtaining new insights about cancer relapse and treatment strategies.