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Molecular mechanisms of tamoxifen resistance in breast cancer
Tomková, Veronika ; Truksa, Jaroslav (advisor) ; Brábek, Jan (referee) ; Mráček, Tomáš (referee)
The resistance to tamoxifen, a drug used in the adjuvant therapy for hormone sensitive breast cancer, represents a major clinical obstacle. Although various mechanisms leading to tamoxifen resistance have been described and intensively studied, a significant number of patients still become resistant to the treatment and eventually relapse. Tamoxifen therapy has been shown to enrich tumors with cancer stem cells (CSCs), which are naturally resistant, and have self-renewal ability and the potential to form secondary tumors. Metabolic rewiring, altered iron metabolism and upregulation of ATP-binding cassette (ABC) transporters have been shown to be important in the maintenance of CSC phenotype. Therefore, we investigated these mechanisms as possible contributors to tamoxifen resistance in vitro in two tamoxifen resistant (Tam5R) cell lines that we established. We show that Tam5R cells have dramatically disassembled and less active mitochondrial supercomplexes (SCs) and higher level of mitochondrial superoxide, together with a fragmented mitochondrial network. Such dysfunction of mitochondria results in the AMP-activated protein kinase (AMPK) activation and metabolic rewiring towards glycolysis. Importantly, cells lacking functional mitochondria are significantly more resistant to tamoxifen, supporting...
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Tumor suppressor NDRG1 and its regulation by iron chelators
Vondráčková, Michaela ; Truksa, Jaroslav (advisor) ; Brábek, Jan (referee)
Iron is an essential trace element required for many processes within a cell, including DNA synthesis and cell cycle progression. Moreover, it is critical for cellular respiration in mitochondria. Due to their proliferative nature, cancer cells are dependent on iron, and depleting this element via iron chelators results in the inhibition of ribonucleotide reductase, leading to cell cycle arrest and apoptosis of cancer cells. Recently, an alternative mechanism for the effect of iron chelators have been proposed, including induction of N-myc downstream regulated gene 1 (NDRG1) expression and its inhibitory effect on c-MET, EGFR, and NF-κB pathways, which can act as oncogenes in a certain context. NDRG1 is a tumour suppressor gene, which is downregulated in many cancers and its downregulation correlates with cancer progression, poor differentiation, and higher metastatic potential. It has been shown that NDRG1 expression can be regulated by intracellular iron - a decrease in intracellular iron leads to upregulation of NDRG1 at mRNA and protein level via the HIF-1-dependent mechanism by inhibiting prolyl hydroxylases. Recently, we have conceived the concept of mitochondrially targeted chelators as an effective anti-cancer agent and in this work, we focused on the evaluation of mitochondrially targeted...
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Regulation of IL-17 receptor complex signaling.
Šemberová, Tereza ; Dráber, Peter (advisor) ; Truksa, Jaroslav (referee)
Inflammatory immune response is essential for maintaining the defense against invading pathogens, although its aberrant activation leads to impaired self tolerance and development of autoimmune pathologies. Interleukin-17A (also known as IL-17), is a major proinflammatory cytokine, which contributes to the development and maintenance of inflammation and provides protection against several bacterial and yeast infections. However, extreme activation of IL-17 signaling leads to autoimmune pathologies. Thus, a strict regulation of IL-17 signal transduction is crucial to prevent progression of autoimmunity. Non-degradative ubiquitination is one of the main mechanisms regulating IL-17 signaling. Main E3 ubiquitin ligase within this signal transduction is TRAF6, which is also participating in several signaling pathways within the immune system. Non-degradative polyubiquitin chains created within inflammatory signaling complexes recruit signaling proteins such as IKK complex and TAK1 kinase, crucial for triggering of NF-κB and MAPKs downstream pathways. However, activation of these pathways upon IL-17 is very weak in comparison with other inflammatory stimuli, indicating a presence of a strong negative feedback loop. In this thesis, we establish the role of several regulatory molecules in IL-17 signaling....
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Role of sulfhydryl oxidase 1 in cancerogenesis
Beranová, Lea Marie ; Truksa, Jaroslav (advisor) ; Šuťák, Róbert (referee)
Disulfide bridges play a significant role in protein-folding as well as en- zyme activity and thus regulate many intra- and extracellular processes. Sulfhydryl oxidase QSOX1 forms S-S bridges de novo, modulating the activity of its substrates and thus directly or indirectly influences vital cel- lular processes. The first part of this thesis focuses on characterization of the role of QSOX1 in cancerogenesis, using breast cancer cell lines (MCF7, MDA-MB-231) and pancreatic cancer cell line (Panc-1), while the second part emphasizes the regulation of QSOX1 expression by different oxygen concentrations. To study the effect of QSOX1 on proliferation of triple-negative cancer cells MDA-MB-231, two genetically modified cell lines - QSOX1-overexpressing and QSOX1 knockout cell lines - were constructed. While increased QSOX1 protein levels do not have a significant effect, the absence of QSOX1 leads to a decreased cellular growth. Lack of QSOX1 also results in visible change in cellular morphology. QSOX1 knockout cells can be mostly characterized as more round-shaped with less noticeable or completely missing lamellipo- dia. This finding is with agreement with to-date literature suggesting that QSOX1 is important not only for cellular proliferation but also for migration and invasiveness. While authenticating the theory of...
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The role of mitochondrial respiratory chain in invasiveness and metastasis of cancer cells and possible therapeutic interventions
Legátová, Anna ; Brábek, Jan (advisor) ; Truksa, Jaroslav (referee)
The mitochondrial respiratory chain, also called the electron transport chain (ETC), has a pivotal role in key features of cancer cells e.g., proliferation, the metabolic shift from oxidative phosphorylation (OXPHOS) to aerobic glycolysis, or the ability to form metastases. This review summarizes current knowledge about ETC and its relationship to cancer, especially to invasiveness and metastases formation. Firstly, it deals with a process called the Warburg effect and with metabolic complexity in the tumor microenvironment. Then it shows how OXPHOS activity affects invasiveness of cancer cells and metastases formation, and it points out the connection between invasiveness and increased levels of ETC-generated reactive oxygen species. At the end, the review deals with possible use of ETC inhibitors in anticancer therapy.
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Molecular mechanisms of tamoxifen resistance in breast cancer
Tomková, Veronika ; Truksa, Jaroslav (advisor) ; Brábek, Jan (referee) ; Mráček, Tomáš (referee)
The resistance to tamoxifen, a drug used in the adjuvant therapy for hormone sensitive breast cancer, represents a major clinical obstacle. Although various mechanisms leading to tamoxifen resistance have been described and intensively studied, a significant number of patients still become resistant to the treatment and eventually relapse. Tamoxifen therapy has been shown to enrich tumors with cancer stem cells (CSCs), which are naturally resistant, and have self-renewal ability and the potential to form secondary tumors. Metabolic rewiring, altered iron metabolism and upregulation of ATP-binding cassette (ABC) transporters have been shown to be important in the maintenance of CSC phenotype. Therefore, we investigated these mechanisms as possible contributors to tamoxifen resistance in vitro in two tamoxifen resistant (Tam5R) cell lines that we established. We show that Tam5R cells have dramatically disassembled and less active mitochondrial supercomplexes (SCs) and higher level of mitochondrial superoxide, together with a fragmented mitochondrial network. Such dysfunction of mitochondria results in the AMP-activated protein kinase (AMPK) activation and metabolic rewiring towards glycolysis. Importantly, cells lacking functional mitochondria are significantly more resistant to tamoxifen, supporting...
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Role of sulfhydryl oxidase 1 in cancerogenesis
Beranová, Lea Marie ; Truksa, Jaroslav (advisor) ; Šuťák, Róbert (referee)
Disulfide bridges play a significant role in protein-folding as well as en- zyme activity and thus regulate many intra- and extracellular processes. Sulfhydryl oxidase QSOX1 forms S-S bridges de novo, modulating the activity of its substrates and thus directly or indirectly influences vital cel- lular processes. The first part of this thesis focuses on characterization of the role of QSOX1 in cancerogenesis, using breast cancer cell lines (MCF7, MDA-MB-231) and pancreatic cancer cell line (Panc-1), while the second part emphasizes the regulation of QSOX1 expression by different oxygen concentrations. To study the effect of QSOX1 on proliferation of triple-negative cancer cells MDA-MB-231, two genetically modified cell lines - QSOX1-overexpressing and QSOX1 knockout cell lines - were constructed. While increased QSOX1 protein levels do not have a significant effect, the absence of QSOX1 leads to a decreased cellular growth. Lack of QSOX1 also results in visible change in cellular morphology. QSOX1 knockout cells can be mostly characterized as more round-shaped with less noticeable or completely missing lamellipo- dia. This finding is with agreement with to-date literature suggesting that QSOX1 is important not only for cellular proliferation but also for migration and invasiveness. While authenticating the theory of...
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Mechanisms of resistance and iron metabolism in cancer stem cells
Lettlová, Sandra ; Truksa, Jaroslav (advisor) ; Kovář, Jan (referee) ; Brábek, Jan (referee)
(EN) Analogously to normal stem cells within the tissues, cancer stem cells (CSCs) have been proposed to be responsible for maintenance and growth of tumours. CSCs represent a small fraction of cells within the tumour, which is characterised by self-renewal capacity and ability to give rise to a tumour when grafted into immunocompromised mice. Cells with increased stemness properties are believed to be responsible for tumour resistance, metastases formation and relapse after tumour treatment. The first part of this work concentrates on resistance of the tumours, which is often associated with increased expression of ATP-binding cassete (ABC) transporters pumping chemotherapeutics out of the cells. For the purposes of this study, we utilized an in vitro model of CSCs, based on cultivation of cells as 3D "spheres". Expression profiling demonstrates that our model of CSCs derived from breast and prostate cancer cell lines express higher mRNA level of ABC transporters, particularly ABCA1, ABCA3, ABCA5, ABCA12, ABCA13, ABCB7, ABCB9, ABCB10, ABCC1, ABCC2, ABCC3, ABCC5, ABCC8, ABCC10, ABCC11 and ABCG2 among the cell lines tested. The protein level of ABC transporters tested in breast CSCs showed higher expression of ABCB8, ABCC1, ABCC2, ABCC10 and ABCG2 but downregulation of ABCB10 and ABCF2 proteins....
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Analyzing the role of the p130Cas SH3 domain in p130Cas-mediated signaling
Gemperle, Jakub ; Rösel, Daniel (advisor) ; Vomastek, Tomáš (referee) ; Truksa, Jaroslav (referee)
The adaptor protein p130Cas (CAS, BCAR1) represents a nodal signaling platform for integrin and growth factor receptor signaling, and influences normal development and tissue homeostasis. Its altered expression drives many pathological conditions including tumor growth, metastasis and drug resistance in many cancer types. How p130Cas contributes to many of these pathologies is still poorly understood. Therefore, the overall aim of my PhD work was to provide new insights to p130Cas signaling and its regulation. The SH3 domain is indispensable for p130Cas signaling, but the ligand binding characteristics of the p130Cas SH3 domain, and the structural determinants of its regulation were not well understood. To be able to study various aspects of p130Cas signaling we identified an atypical binding motif in p130Cas SH3 domain by establishing collaborations with Dr Veverka (Structural biology) and Dr Lepšík (Computational biochemistry; Academy of Sciences, CZ) which gave new insight into this binding interface. Through these collaborations I generated chimeras of p130Cas SH3 domain with its ligands for structural NMR analysis and learned how to visualize and analyze structures. Furthermore, my work expanded our knowledge of p130Cas SH3 ligand binding regulation and led to a novel model of Src-p130Cas- FAK...
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Male infertility in context of testicular cancer
Cimlerová, Markéta ; Komrsková, Kateřina (advisor) ; Truksa, Jaroslav (referee)
This bachelor thesis focuses on male infertility in a connection to testicular cancer. Testicular cancer is the most common malignancy among young men in a reproductive age and the worldwide incidence of testicular cancer is on the rise. A lot of attention is also given to an increasing rate of infertility in a context of testicular cancer. For these reasons, the aim of this thesis is to clarify several non-physiological changes, such as hormone levels, spermatogenesis and sperm parameters, which take place in a male body and influence the chance to become a biological father. These pathological changes can be due to the disease itself but also due to the treatment. A cryopreservation of semen is also going to be discussed as the way to preserve male fertility and as an option for couples with the infertility problems due to male factor which rely on help of the assisted reprodiction. Kye words: testicular cancer, cancer treatment, male infertility, sperm parameters, mitochondria, assisted reproduction
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