| Abstract [eng] |
This thesis focuses on the molecular signaling pathways that regulate cell functions. Mitogen-activated protein kinase (MAPK) and PI3K/AKT signaling molecules are responsible for cell proliferation, regulation of cell death, and participate in signal transduction from extracellular contacts. MAPK and AKT are also involved in drug-induced signaling, and disruption of these signaling pathways is common in cancer cells and is associated with resistance to chemotherapy. Unfortunately, the knowledge about regulation and interactions of the signaling pathways is incomplete yet, and there are no anticancer drugs fully successful in restoring the balance of vital signaling. A promising strategy for cancer treatment is the combination of conventional chemotherapy with targeted drugs, such as protein kinase inhibitors, to manipulate the signals induced by conventional drugs. Therefore, another topic addressed in this work is the resistance of cancer cells to treatment. We have chosen several primary non-small cell lung cancer cell lines, a commercial A549 lung adenocarcinoma cell line, and healthy, stem-like Myo cell line as the subjects of our study, as well as models of inhibition of the extracellular contacts, in combination with the use of various protein kinase inhibitors (PI3K – LY294002, MEK – PD98059, p38 – SB203580, JNK –SP600125, etc.) and approved drugs (MEK inhibitor selumetinib, AKT inhibitor capivasertib, etc.). We investigated whether/how MAPK and AKT participate in the death of adherent cells in "weightless condition", i.e., in the anoikis process, as well as the interdependence of MAPK and AKT activity when the cells are treated with kinase inhibitors and the conventional drug cisplatin. Our studies suggest that, irrespective of cell origin/cell type, the basal phosphorylation level of the protein kinases studied is dependent on extracellular contacts. Among other results, we also showed that despite differences in cell phenotype or genotype, inhibition of the PI3K/AKT pathway promoted activation of the MAP kinase ERK and, vice versa, inhibition of the MEK/ERK pathway increased the level of phosphorylated AKT. This phenomenon was confirmed in both control and cisplatin-treated cells, also using different inhibitors of these kinases. Importantly, the ERK-AKT interaction is dependent on cell-substratum contacts and/or the kinase FAK. Since the activity of signaling molecules regulating cell fate is a principal factor in predicting cell behavior as well as response to therapy, this work highlights the dependence of the self-regulation and the response to the targeted drugs on the cellular state, proposing different molecular strategies for compact (tumor) and circulating cancer cells. |
