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The role of iron in the regulation of proteins connected with tamoxifen-resistance
Potomová, Petra ; Truksa, Jaroslav (advisor) ; Balušíková, Kamila (referee)
Cancer cells are highly dependent on nutrient uptake to sustain their increased proliferation, one of these nutrients being iron. In recent years, a heightened dependency on iron was observed in cancer cells, allowing for the proper function of numerous enzymes, DNA synthesis and mitochondrial respiration. Here, we further delve into the iron metabolism of malignant cells, attempting to understand the differences between tamoxifen-sensitive and resistant (Tam5R) ones using two breast cancer cell lines of luminal A origin, MCF7 and T47D. These cells show numerous changes in iron homeostasis and iron-dependent mechanisms. Based on alterations in proteomes of Tam5R cell lines, we focused on iron regulation of proteins that are deregulated in tamoxifen resistance - assessing their regulation on transcriptional (mRNA) and post-transcriptional level (protein) as well as comparing their responsiveness to their sensitive parental cell line. We assessed two main types of regulation - iron-responsive element interaction with iron-regulatory proteins (IRE-IRP pathway) and tristetraprolin (TTP) driven mRNA degradation via AU-rich elements (ARE). Using iron loading and chelation, we challenged the cells - confirming the known IRE-IRP regulation of ferritin heavy chain (FTH), transferrin receptor 1 (TfR1),...
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Proteome analysis of anti-cancer drug effects and characterisation of drug resistance
Hrabáková, Rita
Despite significant progress in the development of anti-cancer drugs, there is still a need for novel therapeutic strategies that would improve the outcome of cancer patients. Using proteomic technologies and cell lines with different phenotype of p53 tumour suppressor, we monitored cancer cell response to anti-cancer treatment with focus on the development of drug resistance. The different levels of metabolic proteins were identified in our study which may help to explain different anti-cancer activity of drugs with only a subtle difference in structure. More importantly, proteins associated with the development of drug resistance were identified and such expression changes have become a focus of interest. Our findings demonstrate a higher protein level of serine hydroxymethyltransferase, serpin B5 and calretinin in cancer cells resistant to Aurora kinase inhibitors. Such proteins promote the tumour growth with no apparent impact of p53 phenotype whilst voltage-dependent anion-selective channel protein 2 contributes to the development of resistance only in cells with functional p53 which is accompanied by the decreased level of elongation factor 2. On the other hand, cancer cells with loss of p53 appear to amplify alternative mechanisms such as protection against oxidative stress. The results...
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The role of autophagy and selected beta-tubulin isotypes in taxane resistance in breast cancer cells
Kábelová, Adéla ; Jelínek, Michael (advisor) ; Truksa, Jaroslav (referee)
Drug resistance in cancer cells is a frequent cause of breast cancer therapy failure. The aim of this thesis was to elucidate mechanisms of resistance to taxanes, that are used in therapy of various types of cancer, including breast cancer. We particularly assessed the role of autophagy and changes in βII- and βIII isotype gene expression in development of taxane resistance. As model of breast cancer we used human sensitive cell lines SK-BR-3, MCF-7 a T47-D and resistant sublines SK-BR-3-PAC/REZ a MCF-7- PAC/REZ which grow in paclitaxel concentration lethal for sensitive sublines. In cell lines SK-BR-3 and MCF-7, taxane application decreased the level of autophagy, however in cell line T47-D led to its activation. We detected no difference between basal levels of autophagy in sensitive subline SK-BR-3 compared to resistant subline SK-BR-3-PAC/REZ, but we observed increased basal level of autophagy in sensitive subline MCF-7 compared to the resistant subline. Increase or decrease level of autophagy did not affect taxane resistance, except activation of autophagy in resistant subline SK-BR-3-PAC/REZ, that further increased the resistance to paclitaxel. Taxane application in cell line T47-D increased the levels of βII- and βIII-tubuline expression, however we did not find any similar effect in other tested...
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Molecular mechanisms of the resistence of breast cancer cells to taxanes: the role of ABC transporters
Kopperová, Dana ; Kovář, Jan (advisor) ; Hrdý, Jiří (referee)
Resistance to chemotherapeutics is a widespread phenomenon in cancer cells that may counteract the successful therapy of many patients. In resistant cells, higher level of ABC transporters, among others, often can be detected. This high level of ABC transporters represents a suspected mechanism of acquired cancer resistance. We studied the molecular mechanism of resistance to taxanes in cancer cells using SK-BR-3 and MCF-7 breast cancer cell lines. We analyzed the effect of paclitaxel on apoptosis induction in the originally sensitive cells of these lines as compared to their counterpart resistant cells, developed by gradual adaptation to paclitaxel. In resistant cells of the SK-BR-3 and MCF-7 lines, we did not detected ongoing induction of apoptosis but we did detect significantly increased expression of ABCB1 transporter after paclitaxel application. By silencing the expression of the transport via employment of small interfering RNA (siRNA), we tested the role of the ABCB1 transporter in cells resistant to paclitaxel. We found that resistant cells with silenced expression of the ABCB1 transporter had a statistically significant increase of sensitivity to paclitaxel as compared to control resistant cells with high expression of this transporter. Along with increased sensitivity, we demonstrated...
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Proteome analysis of anti-cancer drug effects and characterisation of drug resistance
Hrabáková, Rita ; Kovářová, Hana (advisor) ; Hernychová, Lenka (referee) ; Šulc, Miroslav (referee)
Despite significant progress in the development of anti-cancer drugs, there is still a need for novel therapeutic strategies that would improve the outcome of cancer patients. Using proteomic technologies and cell lines with different phenotype of p53 tumour suppressor, we monitored cancer cell response to anti-cancer treatment with focus on the development of drug resistance. The different levels of metabolic proteins were identified in our study which may help to explain different anti-cancer activity of drugs with only a subtle difference in structure. More importantly, proteins associated with the development of drug resistance were identified and such expression changes have become a focus of interest. Our findings demonstrate a higher protein level of serine hydroxymethyltransferase, serpin B5 and calretinin in cancer cells resistant to Aurora kinase inhibitors. Such proteins promote the tumour growth with no apparent impact of p53 phenotype whilst voltage-dependent anion-selective channel protein 2 contributes to the development of resistance only in cells with functional p53 which is accompanied by the decreased level of elongation factor 2. On the other hand, cancer cells with loss of p53 appear to amplify alternative mechanisms such as protection against oxidative stress. The results...
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Proteome analysis of anti-cancer drug effects and characterisation of drug resistance
Hrabáková, Rita
Despite significant progress in the development of anti-cancer drugs, there is still a need for novel therapeutic strategies that would improve the outcome of cancer patients. Using proteomic technologies and cell lines with different phenotype of p53 tumour suppressor, we monitored cancer cell response to anti-cancer treatment with focus on the development of drug resistance. The different levels of metabolic proteins were identified in our study which may help to explain different anti-cancer activity of drugs with only a subtle difference in structure. More importantly, proteins associated with the development of drug resistance were identified and such expression changes have become a focus of interest. Our findings demonstrate a higher protein level of serine hydroxymethyltransferase, serpin B5 and calretinin in cancer cells resistant to Aurora kinase inhibitors. Such proteins promote the tumour growth with no apparent impact of p53 phenotype whilst voltage-dependent anion-selective channel protein 2 contributes to the development of resistance only in cells with functional p53 which is accompanied by the decreased level of elongation factor 2. On the other hand, cancer cells with loss of p53 appear to amplify alternative mechanisms such as protection against oxidative stress. The results...
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