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Studium mechanismů disociace a reasociace feritinových proteinových klecí a jejich využití v nanomedicíně / Study of disassembly/reassembly mechanisms of ferritin protein cages and their utilization in nanomedicineKrausová, Kateřina January 2019 (has links)
Diploma thesis deals with the study of dissociation and reassociation of ferritin protein cages and their use in nanomedicine. Most studies that are focused on targeted transport of pharmaceuticals using ferritin cages work with horse spleen ferritin. It is, however, its origin, which leads to increasingly frequent questions about possible immunogenicity in the patient's organism, which also provides the main motivation to test the possibility of encapsulation of low-molecular drugs into ferritins originating from alternative organisms. In the practical part the method for the study of dissociation was experimentally designed. Native polyacrylamide gel electrophoresis was used to study dissociation of equine ferritin composed of different subunit, human ferritin, and archeal Pyrococcus furiosus ferritin. The obtained subunit dissociation results were used to encapsulate the low molecular chemotherapeutic drug doxorubicin and for further characterization of the ferritin-doxorubicin complex. The efficacy of the designed nanoformulations has been verified in the treatment of malignant breast cancer. Human ferritin proves to be the optimal one. Its composition of heavy subunits corresponds to a lower protein stability, thus a more efficient opening of the structure and consequent encapsulation of the cytostatics occurs. With its 60% encapsulation efficiency of doxorubicin, low polydispersity index, effective cytotoxicity of ferritin-doxorubicin complex and minimal risk of immune response to the patient's organism, human ferritin achieves better results than commonly used horse spleen ferritin.
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Ramanova spektroskopie biologicky aktivních látek a protinádorových léčiv / Raman spectroscopy of biologically active species and antitumor drugsTřeštíková, Liběna January 2009 (has links)
SERRS spektra of biological materials are very komplex, because they consist of signals from all molecules present in cells. In this text are presented SERRS spektra of antitumor drugs and its komplex with DNA. Experimental are rated on doxorubicin and another antitumor druha and on study of theirs potential by treatment for tumors. Doxorubicin belong to clase antracycline antibiotics and is used for stop of tumor cells reproduction. Scientists found still new ways, new drugs. SERRS is one of possibilities for study of this drugs and theirs interaction with DNA.
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Studium abiotického stresu u rostlin na úrovni proteomu / The proteomic study of abiotic stress of plants.Barabášová, Kamila January 2011 (has links)
Keywords: Arabidopsis thaliana, phytoremediation, abiotic stress, ibuprofene, doxorubicin, two-dimensional electrophoresis Nowadays, develop of the pharmaceutical industry is very fast. Reason of this trend is ever-increasing number of diseases, lifestyle and still increasing demand for the drugs. With this trend growing interest in the analysis of the residues of pharmaceuticals in the environment which is result of incomplete wastewater treatment. This diploma thesis is studying effect of cytostatic drugs, specifically doxorubicin and one of the most widely used analgesics - ibuprofen, at the proteome level of the model plant Arabidopsis thaliana. Proteins isolated from plants exposed to the drugs were separated by two-dimensional electrophoresis. Comparing of protein maps by PDQest program (Bio-Rad, USA) was found several proteins whose expression was affected by the presence of drugs in the culture medium. Selected proteins were identified by LC - MS / MS.
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Studium vlastností protinádorových léčiv ellipticinu, etoposidu a doxorubicinu ve formě nanočástic / The study of properties of anticancer drugs ellipticine, etoposide and doxorubicin in the forms of nanocarriersLengálová, Alžběta January 2016 (has links)
Currently available anticancer therapies are inadequate and spur demand for improved technologies. Among others, the utilization of nanocarriers for anticancer drug delivery has shown great potential in cancer treatment. Nanocarriers can improve the therapeutic efficiency of the drugs with minimization of the undesirable side effects. To evaluate potential application of this technology, two forms of nanocarriers have been studied: multi-walled carbon nanotubes (MWCNTs) and apoferritin. The aim of this study was to determine, whether given cytostatics (ellipticine, etoposide and doxorubicin) are bound to these nanotransporters and how are they released from them, especially depending on pH. Since the pH of the tumor cells is lower than the pH of healthy cells it would be preferred that the drugs would release from nanocarriers at the lower pH while at the physiological pH the release of the drug would be eliminated. The results found show that ellipticine is actually released from its MWCNT- and apoferrtin-encapsulated form at acidic pH (5.0), while at pH 7.4 its interaction with nanocarriers is stable. Ellipticine released from MWCNT is activated by microsomal enzymes to reactive metabolites (13- hydroxyellipticine and 12-hydroxyellipticine) forming DNA adducts. The results indicate that both...
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Theranostic Nanoparticles Folic acid-Carbon Dots-Drug(s) for CancerBABANYINAH, GODWIN KWEKU 18 March 2021 (has links)
The main aim of this study is to synthesize theranostic nanoparticles (NPs) that will drastically increase the diagnostics and therapeutic efficacy for cancer. In this research, we had prepared the NPs which constitute carbon dots (CDs), the imaging agent, Folic acid, the targeting agent, and Doxorubicin (DOX) or Gemcitabine (GEM) as the chemotherapy agents. The prepared NPs include noncovalent FA-CDs-DOX, covalent CDs-FA-DOX, and covalent FA-CDs-GEM. The spectroscopy, ultraviolet-visible spectroscopy (UV-vis), fluorescence spectroscopy, and Fourier transform-infrared spectroscopy (FT-IR), were used to confirm the successful fabrication of these complexes. Through UV-vis analysis, the drug loading capacity (DLC) and drug loading efficiency (DLE) of the complexes were determined. The noncovalent series had a higher DLE of about 83% while the covalent series showed higher DLC, 70% on average indicating high drug content. The in-vitro pH-dependent drug release shows that the noncovalent FA-CDs-DOX and the covalent FA-CDs-GEM series release more drugs into the cancer cells (pH of 5.0) than into healthy normal (pH of 7.4). The sizes of NPs were measure around 2-5 nm with Dynamic light Scattering (DLS). The toxicity of CDs, CDs-drug, and FA-CDs-drug on MDA-MB468 breast cancer cell was tested through the methylthiazolytetrazolium (MTT) assay and found that the FA bonded NPs exhibited strong therapeutic efficacy. More pharmaceutical data towards the cancer cells are investigated by our research collaborators – the pharmaceutical department at ETSU and Xavier University at Louisiana.
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The Role of Activating Transcription Factor 3 as a Regulator of DNA-Damaging Chemotherapy CytotoxicityHasim, Mohamed Shaad 29 November 2019 (has links)
DNA-damaging chemotherapeutics are a consistently employed for the treatment of cancer, and are regularly part of first-line combination therapeutic regimens. However, these regimens often display limited activity in cancers including of the lung and breast. Novel therapeutic strategies are urgently required. Understanding the molecular mechanisms regulating DNA-damaging chemotherapeutics activity will lead to such novel strategies.
Activating transcription factor 3 (ATF3) is a stress inducible gene that plays a significant role in regulating cellular stress including DNA damage. This thesis investigates the role of ATF3 in mediating the cytotoxic effects of two commonly employed DNA-damaging chemotherapeutics, cisplatin and doxorubicin. This study also identifies other independent ATF3 inducing agents in potential novel combination therapeutic strategies.
In this study, cell line models of cisplatin-resistance were generated independently from two non-small cell lung cancer (NSCLC) cell lines, Calu6 and H23. Full transcriptome RNA-sequencing analysis identified ATF3 as the most highly dysregulated apoptosis regulatory gene in the cisplatin-resistant compared to their respective parental cell lines following treatment with cisplatin. Further characterization identified cisplatin induced activation of JNK as the key regulator of ATF3 induction in these cell lines. Restoring JNK activity resulted in induced ATF3 expression and re-sensitization of the resistant cell lines to cisplatin treatment. FDA-approved 1200 compound library screens were employed to identify agents that can enhance cisplatin cytotoxicity as well as whose cytotoxicity was dependent on ATF3 expression. Vorinostat, an HDAC inhibitor, was identified in both screens and importantly displayed synergistic cytotoxicity in combination with cisplatin.
In addition, ATF3 was also induced with treatments of the DNA damaging agent doxorubicin in these NSCLC cell lines including in the cisplatin resistant models. This work suggested a different mechanism of ATF3 induction by doxorubicin and the potential role of ATF3 in mediating doxorubicin cytotoxicity was further investigated in breast cancer cells. Doxorubicin robustly increased ATF3 expression in human breast cancer cell lines and tumor tissue ex-vivo. Loss of ATF3 in MEFs resulted in reduced sensitivity to doxorubicin treatment compared to wild-type MEFs. Employing the same library screen as above, compounds with ATF3 dependent mechanisms that could enhance doxorubicin cytotoxicity were identified. Vorinostat, as well as, the nucleoside analogues trifluridine and 6-mercaptopurine induced ATF3 expression and enhanced doxorubicin cytotoxicity.
This study demonstrated that ATF3 plays an important role in mediating the cytotoxic effect of the DNA-damaging chemotherapeutics cisplatin and doxorubicin. It also provides rationale for ATF3 as a potential therapeutic target, and for the incorporation of ATF3 inducing agents in novel combination therapeutic strategies.
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Overexpression of Angiopoietin-1 Reduces Doxorubicin-Induced Apoptosis in CardiomyocytesRen, Danyang, Zhu, Quan, Li, Jiantao, Ha, Tuanzhu, Wang, Xiaohui, Li, Yuehua 01 November 2012 (has links)
Doxorubicin (Dox) is a major anticancer chemotherapeutic agent. However, it causes cardiomyopathy due to the side effect of cardiomyocyte apoptosis. We have previously reported that angiopoietin-1 significantly reduced myocardial infarction after ischemic injury and protected cardiomyocytes from oxidative stress-induced apoptosis. It is hypothesized that angiopoietin-1 may protect cardiomyocytes from Dox-induced apoptosis. Cardiomyocytes H9C2 were transfected with adenovirus expressing angiopoietin-1 (Ad5-Ang-1) 24 h before the cells were challenged with Dox at a concentration of 2 μmol/L. Ad5-GFP served as the vector control. Cardiomyocyte apoptosis was evaluated using Annexin V-FITC staining and caspase-3 and caspase-8 activity was determined by Western blotting. The results showed that Dox treatment significantly induced cardiomyocyte apoptosis as evidenced by the greater number of Annexin V-FITC stained cells and increases in caspase-3 and caspase-8 activity. In contrast, overexpression of angiopoietin-1 significantly prevented Dox-induced cardiomyocyte apoptosis. To elucidate the mechanisms by which angiopoietin-1 protected cells from Dox-induced apoptosis, we analyzed both extrinsic and intrinsic apoptotic signaling pathways. We observed that angiopoietin-1 prevented Dox-induced activation of both extrinsic and intrinsic apoptotic signaling pathways. Specifically, angiopoietin-1 prevented DOX-induced in-creases in FasL and Bax levels and cleaved caspase-3 and caspase-8 levels in H9C2 cells. In addition, overexpression of angiopoietin-1 also activated the pro-survival phosphoinositide-3 kinase (PI3K)/Akt signaling pathway and decreased Dox-induced nuclear factor-kappaB (NF-κB) activation. Our data suggest that promoting the expression of angiopoietin-1 could be a potential approach for reducing Dox-induced cardiomyocyte cytoxicity.
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Over-Expression of a Modified Bifunctional Apoptosis Regulator Protects Against Cardiac Injury and Doxorubicin-Induced Cardiotoxicity in Transgenic MiceChua, Chu C., Gao, Jinping, Ho, Ye S., Xu, Xingshun, Kuo, I. C., Chua, Kaw Y., Wang, Hong, Hamdy, Ronald C., Reed, John C., Chua, Balvin H. 01 January 2009 (has links)
Aims: Bifunctional apoptosis regulator (BAR) is an endoplasmic reticulum protein that interacts with both the extrinsic and intrinsic apoptosis pathways. We hypothesize that over-expression of BARΔRING prevents apoptosis and injury following ischaemia/reperfusion (I/R) and attenuates doxorubicin (DOX)-induced cardiotoxicity. Methods and results: We generated a line of transgenic mice that carried a human BARΔRING transgene under the control of the mouse α-myosin heavy chain promoter. The RING domain, which binds ubiquitin conjugating enzymes, was deleted to prevent auto-ubiquitination of BAR and allow accumulation of the BAR protein, which binds apoptosis-regulating proteins. High levels of human BARΔRING transcripts and 42 KDa BARΔRING protein were expressed in the hearts of transgenic mice. When excised hearts were reperfused ex vivo for 45 min as Langendorff preparations after 45 min of global ischaemia, the functional recovery of the hearts, expressed as left ventricular developed pressure x heart rate, was 23 ± 1.7% in the non-transgenic hearts compared with 51.5 ± 4.3% in the transgenic hearts (P < 0.05). For in vivo studies, mice were subjected to 50 min of ligation of the left descending anterior coronary artery followed by 4 h of reperfusion. The infarct sizes following I/R injury, expressed as the percentage of the area at risk, were significantly smaller in the transgenic mice than in the non-transgenic mice (29 ± 4 vs. 55 ± 4%, P < 0.05). In hearts of mice subjected to cardiac I/R injury, BAR transgenic hearts had significantly fewer in situ oligo-ligation-positive cardiac cells (5.0 ± 0.4 vs. 13.4 ± 0.5%, P < 0.05). Over-expression of BARΔRING also significantly attenuated DOX-induced cardiac dysfunction and apoptosis. Conclusion: Our results demonstrate that over-expression of BARΔRING renders the heart more resistant to I/R injury and DOX-induced cardiotoxicity, and this protection correlates with reduced cardiomyocyte apoptosis.
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Glutathione Peroxidase 1-Deficient Mice Are More Susceptible to Doxorubicin-Induced CardiotoxicityGao, Jinping, Xiong, Ye, Ho, Ye Shih, Liu, Xuwan, Chua, Chu Chang, Xu, Xingshun, Wang, Hong, Hamdy, Ronald, Chua, Balvin H.L. 01 October 2008 (has links)
Doxorubicin (DOX)-induced cardiotoxicity is thought to be mediated by the generation of superoxide anion radicals (superoxide) from redox cycling of DOX in cardiomyocyte mitochondria. Reduction of superoxide generates H2O2, which diffuses throughout the cell and potentially contributes to oxidant-mediated cardiac injury. The mitochondrial and cytosolic glutathione peroxidase 1 (Gpx1) primarily functions to eradicate H2O2. In this study, we hypothesize that Gpx1 plays a pivotal role in the clearance of H2O2 generated by DOX. To test this hypothesis, we compared DOX-induced cardiac dysfunction, mitochondrial injury, protein nitration, and apoptosis in Gpx1-deficient and wild type mouse hearts. The Gpx1-deficient hearts showed increased susceptibility to DOX-induced acute functional derangements than wild type hearts, including impaired contractility and diastolic properties, decreased coronary flow rate, and reduced heart rate. In addition, DOX treatment impaired the mitochondrial function of Gpx1-deficient hearts. Specifically, Gpx1-deficient hearts treated with DOX demonstrated an increased rate of NAD-linked state 4 respiration and a decline in the P/O ratio relative to wild type hearts, suggesting that DOX uncouples the electron transfer chain and oxidative phosphorylation in Gpx1-deficient hearts. Finally, apoptosis and protein nitration were significantly increased in Gpx1-deficient mouse hearts compared to wild type hearts. These studies suggest that Gpx1 plays significant roles in protecting DOX-induced mitochondrial impairment and cardiac dysfunction in the acute phase.
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Over-Expression of Heat Shock Protein 27 Attenuates Doxorubicin-Induced Cardiac Dysfunction in MiceLiu, Li, Zhang, Xiaojin, Qian, Bo, Min, Xiaoyan, Gao, Xiang, Li, Chuanfu, Cheng, Yunlin, Huang, Jun 01 August 2007 (has links)
Background: Oxidative stress and myocyte apoptosis are thought to play an important role in the pathogenesis, progression and prognosis of heart failure (HF). Heat shock protein 27 (Hsp27) has been found to confer resistance to oxidative stress in cultured cells; however, the role of Hsp27 in in-vivo hearts remains to be determined. Aim: To investigate the effects of Hsp27 over-expression on doxorubicin-induced HF. Methods and Results: Transgenic mice (TG) with cardiac specific over-expression of Hsp27 and their wild type littermates (WT) were challenged with doxorubicin (25 mg/kg, IP) to induce HF. At day 5, TG mice had significantly improved cardiac function and viability and decreased loss of heart weight following doxorubicin exposure compared with WT. In another parallel experiment, doxorubicin-induced increased levels of reactive oxygen species, protein carbonylation, apoptosis and morphologic changes were detected in the mitochondria in WT hearts, whereas these effects were markedly attenuated in TG hearts. In addition, upregulation of heat shock protein 70 and heme oxygenase-1 was present in the TG hearts after doxorubicin stimulation in comparison to WT hearts. Conclusion: These findings indicate that Hsp27 may play a key role in resistance to doxorubicin-induced cardiac dysfunction.
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