Dissertations / Theses on the topic 'Chemotherapy role'
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Allen, W. L. "The role of Fas in response to chemotherapy." Thesis, Queen's University Belfast, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.403264.
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Tran, Cuong Duy. "Intestinal zinc and metallothionein : role in chemotherapy-induced mucositis." Title page, contents and abstract only, 2001. http://web4.library.adelaide.edu.au/theses/09PH/09phc9736.pdf.
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Bowen, Joanne Marie. "Chemotherapy-induced intestinal mucositis the role of apoptosis regulators /." Click here to access, 2006. http://thesis.library.adelaide.edu.au/public/adt-SUA20060831.142913/index.html.
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Thesis (Ph.D.) -- University of Adelaide, School of Medicine, Discipline of Medicine, 2006.Includes author's previously published papers. "March 2006" Bibliography: leaves 168-191. Also available in a print form.
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Khongkow, Pasarat. "The role of FOXM1 in breast cancer chemotherapy resistance." Thesis, Imperial College London, 2015. http://hdl.handle.net/10044/1/42884.
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Resistance to chemotherapeutic agents is the main obstacle to the effective breast cancer management. Therefore, it is important to elucidate the molecular mechanisms of chemoresistance and develop novel therapeutic strategies in order to overcome drug resistance. In this work, I found that FOXM1 is a critical mediator of epirubicin and paclitaxel resistance in MCF-7 breast cancer cell lines. FOXM1 expression was upregulated in both epirubicin resistant MCF-7 (MCF-7 EpiR) and paclitaxel resistant MCF-7 (MCF-7 TaxR) cells compared to sensitive MCF-7 cells. Interestingly, its depletion dramatically impaired the clonogenic survival and significantly induced cellular senescence in the resistant cells. In addition, I identified two novel downstream FOXM1 targets, NBS1 and KIF20A, involved in epirubicin and paclitaxel resistance, respectively. Firstly, I found that FOXM1 transcriptionally regulated NBS1 expression to modulate HR-mediated DSB repair and epirubicin resistance. Overexpression of FOXM1 and NBS1 lead to the enhancement of HR efficiency to eliminate epirubicin-induced DNA damage. Conversely, similar to FOXM1, depletion of NBS1 also sensitised both MCF-7 and MCF-7 EpiR cells to epirubicin by inducing cellular senescence. Secondly, I identified the mitotic kinesin KIF20A as a direct downstream target of FOXM1, participating in the mitotic spindle formation and paclitaxel resistance. Depletion of KIF20A caused mitotic spindle abnormalities, inhibition of cell growth as well as the induction of senescent cells in both MCF-7 and MCF-7 TaxR cells. Consistently, immunohistochemical analysis of breast cancer patient samples revealed that high expression levels of FOXM1, NBS1 and KIF20A are strongly correlated with poor prognosis in breast cancer, supporting a physiological role of FOXM1 and its novel targets in genotoxic drug resistance. Collectively, these findings suggests that FOXM1 and its targets, NBS1 and KIF20A, could be reliable prognostic markers for monitoring treatment efficiency as well as promising targets for therapeutic intervention to overcome epirubicin and paclitaxel resistance in breast cancer.
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Simpson, Julie Ann. "The role of population pharmacokinetic- pharmacodynamic modelling in antimalarial chemotherapy." Thesis, Open University, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.367218.
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Gustafson, Heather Lynn. "Role of Manganese Superoxide Dismutase in Chemotherapy-induced Oxidative Stress." Diss., The University of Arizona, 2011. http://hdl.handle.net/10150/203510.
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Existing treatments for mantle cell lymphoma (MCL) are non-curative, demonstrating a need for a refined treatment approach. Recent clinical trials have shown promising results with the use of mammalian target of rapamycin inhibitors. I hypothesize that the anti-tumor effect of mTOR inhibitors in mantle cell lymphoma is mediated by an increase in manganese superoxide dismutase (MnSOD) protein expression and accumulation of hydrogen peroxide (H₂O₂). Findings indicate that the rapamycin-induced cytostatic effect is characterized by increased levels of MnSOD and H₂O₂, and is necessary for the full growth inhibitory effect of rapamycin. Furthermore, over-expression of MnSOD elevated the level of H₂O₂ and increased sensitivity to MnSOD. Treatment with rapamycin resulted in a loss of serine 473 phosphorylation of AKT and increased levels of MnSOD were found to be due to inhibition of the mTORC2 complex. These results are the first to suggest that long term treatment of MCL cells with rapamycin inhibits the mTORC2 complex. By understanding the key signaling molecules and affected pathways in the anti-tumor effects of mTOR inhibitors, we may be able to identify additional predictive markers to improve the therapeutic value, or study drug combinations that will enhance the effect of ROSinduced cytotoxicity. A retrospective study utilizing samples from lymphoma patients receiving standard anthracycline-based therapies, identified single nucleotide polymorphisms in oxidative stressrelated genes associated with survival. Individuals carrying minor allele SNPs in myeloperoxidase (MPO) and an aldo-keto reductase (AKR1C3) were found to be associated with shorter time to disease progression and death. This data suggest that some patients may benefit from a different therapy than the current standard of care and that regulation of the redox environment plays a role in aggressive lymphoma treatment response.
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Kwan, Stanley. "The role of ASK1 in arsenic trioxide-induced cell death." Thesis, McGill University, 2013. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=114442.
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Arsenic trioxide (ATO) is an effective treatment for acute promyelocytic leukemia (APL). While it is undergoing clinical trials for numerous malignancies including multiple myeloma, myelodysplastic syndrome, lymphoma and solid tumors, it has demonstrated only limited efficacy as a single agent. However, it may hold promise as part of a combination therapy. Thus investigation to elucidate the mechanisms of action underlying these clinical responses may lead to generation of rational combination therapies to increase its therapeutic spectrum. Previous work has described a pathway required for ATO-induced apoptosis in APL cells involving the generation of reactive oxygen species (ROS), and the subsequent induction of a specific mitogen-activated protein kinase (MAPK) cascade that includes both stress-activated protein kinase (SAPK)/ERK kinase 1 (SEK1) and c-Jun N-terminal kinases (JNK) activation. However, the link between ROS production and activation of SEK1 remains to be elucidated. Apoptosis signaling kinase 1 (ASK1) is a MAP3K upstream of SEK1 that has been implicated in the induction of stress-induced signaling.Using murine embryonic fibroblasts (MEFs) derived from ASK1-/- mice; we provide evidence that ASK1 is a strong mediator for ATO-induced apoptosis and JNK activation. In an APL cell line, we show that ATO activates ASK1 in a dose- and time-dependent manner. However, knockdown of ASK1 in APL cells enhanced susceptibility to ATO, undergoing apoptosis and growth inhibition more than their wild type counterparts. The same impact was observed in the knockdown of SEK1, a direct downstream MAP2K of ASK1, and the knockdown of ASK1 in MCF-7 cells, a human breast cancer cell line. This led us to postulate that ASK1 had a pro-apoptotic function in non-transformed fibroblasts, but was pro-survival in malignant cells. Indeed, transformation of ASK1-/- MEFs restored their sensitivity to ATO-induced apoptosis and growth inhibition. Taken together, these results suggest that ASK1 can have both pro-apoptotic and anti-apoptotic roles depending on the transformation state of the cells.One model of ASK1 regulation suggests that ASK1 is kept in an inactive form by reduced thioredoxin-1 (Trx1). During oxidative stress, Trx1 is oxidized and releases ASK1 for activation. Immunoprecipitation of ASK1 followed by immuoblotting for Trx1 in APL cells shows a strong basal association that is lost with ATO treatment. Furthermore, the activity of thioredoxin reductase 1 (TrxR1), an enzyme that converts oxidized Trx1 into reduced Trx1, is significantly decreased following ATO treatment. This suggests that ATO activates ASK1 signaling by ROS-mediated oxidation of Trx1 and by maintaining Trx1 in its oxidized state by decreasing TrxR1 activity. In addition, we show that inhibition of TrxR1 with the TrxR1 inhibitor Auranofin sensitizes APL cells to ATO-induced apoptosis. Overall, our results suggest that targeting Trx1 may enhance ATO-induced apoptosis in a novel combination therapy.L'arsenic trioxyde (ATO) est un traitement efficace contre la leucémie promyélocitaire aïgue (APL). Alors qu'il est testé dans le cadre de plusieurs essais cliniques sur différents cancers comme le myélome multiple, le syndrome myélodysplasique, le lymphome et les tumeurs solides, il ne montre qu'une efficacité limitée en tant qu'agent unique. Quoiqu'il en soit, il pourrait être prometteur au sein d'une thérapie combinée. Ainsi, l'étude des mécanismes d'action responsables des bénéfices cliniques apportés par l'arsenic trioxide pourrait mener à la mise au point de nouvelles thérapies combinées afin d'élargir le spectre thérapeutique d'ATO. Des travaux antérieurs ont décri une voix de signalisation, requise pour l'induction de l'apoptose par l'arsenic dans des cellules d'APL, qui implique la génération d'espèces actives de l'oxygène (ROS), ainsi que l'induction subséquente d'une cascade de protéine kinases mitogène-activées (MAPK) spécifique qui inclut à la fois la protéine kinase stress-activée (SAPK)/ERK kinase 1(SEK1) ainsi que l'activation de la kinase c-jun N-terminal (JNK). Néanmoins, le lien entre la production de ROS et l'activation de SEK1 reste à être élucidé. La kinase de signalisation de l'apoptose (ASK1) est une MAP3K en amont de SEK1 qui a été impliquée dans l'induction de la signalisation induite par le stress. En utilisant des fibroblastes d'embryions de souris (MEFs) dérivées de souris ASK-/-, nous montrons qu'ASK1 est un médiateur fort de l'apoptose et de l'activation de JNK par ATO. Dans une lignée cellulaire APL, nous montrons qu'ATO active ASK1 en fonction du temps et de la dose. Par contre, une sous-régulation d'ASK1 dans des cellules APL augmente la susceptibilité envers ATO, ce qui se traduit par une diminution de la croissance et une augmentation de l'apoptose par rapport aux cellules APL sauvages. Un impact similaire est observé lors d'une sous régulation de SEK1, une MAP2K directement en aval d'ASK1, ainsi que lors d'une sous régulation d'ASK1 dans des cellules MCF-7, des cellules de cancer du sein humain. Cela nous a conduit à postuler qu'ASK1 a une fonction pro-apoptotique dans les fibroblastes non-transformés, mais anti-apoptotique dans des cellules malignes. En effet, la transformation des cellules MEFs ASK-/- a restauré leur sensibilité envers l'arrêt de la croissance et l'apoptose induits par ATO, ce qui souligne les différences entre les cellules normales et les cellules transformées. Dans l'ensemble, ces résultats suggèrent qu'ASK1 a un rôle important dans la sensibilité à l'ATO qui dépend du contexte. Un des schémas de régulation d'ASK1 suggère qu'ASK1 est séquestré sous une forme inactive par la thioredoxine-1 réduite (Trx1). Durant le stress oxydatif, Trx1 est oxydée et libère ASK1 afin qu'il puisse être activé. L'immuno-précipitation d'ASK1 suivie d'un immuno-marquage for Trx1 dans des cellules APL montre une association basale forte qui est perdue lors du traitement avec ATO. De plus, l'activité de la réductase de la thioredoxine (TrxR1), une enzyme qui convertit Trx1 oxydée en Trx1 réduite, est diminuée de façon significative après traitement avec ATO. Cela suggère qu'ATO active la signalisation d'ASK1 en oxydant Trx1, via les ROS, ainsi qu'en inhibant la réduction de Trx1 en diminuant l'activité de TrxR1. De plus, nous montrons que l'inhibiteur de TrxR1, Auranofin, sensibilise les cellules APL à l'apoptose induite par ATO. Cela suggère que la régulation d'ASK1 dépend du statut redox de Trx1. Dans l'ensemble, nos résultats suggèrent que le fait de cibler Trx1 pourrait augmenter la signalisation d'ASK1 et l'apoptose induite par ATO au sein d'une nouvelle thérapie combinée.
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Wong, Chung-lim, and 黃仲廉. "The role of GEP on chemotherapy induced alterations in hepatocellular carcinoma." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2013. http://hdl.handle.net/10722/197132.
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Hepatocellular carcinoma (HCC) is the fifth most common malignancy and the third leading cause of cancer related death worldwide. Chemo-therapy has been commonly used to treat unresectable HCC but with limited efficacy. Therefore, there is an urgent demand for the development of better therapeutic approaches. Granulin-epithelin precursor (GEP) is a novel growth factor with over-expression in more than 70% of HCCs and has been demonstrated as potential therapeutic target. The aims of this study are to examine the role of GEP in chemo-resistance and the therapeutic potential of GEP antibody therapy in combination with chemo-therapy in HCC.
The role of GEP in HCC chemo-resistance has been examined by HCC in vitro models in the first part of the study and by in vivo human HCC xenograft models in immunocompromised mice in the second part of the study. It was shown that the chemo-therapeutic agents selected HCC cells in vitro and in vivo resulted in increased cellular expression of GEP, ABCB5, hepatic cancer stem cell (CSC) marker CD133/EpCAM positive populations and demonstrated enhanced CSCs properties including colony formation ability and chemo-resistance. Over-expression and knockdown of GEP expressions respectively demonstrated that GEP levels were important in conferring resistance to the chemo-therapeutic agents and the drug-induced apoptosis.
GEP antibody therapy not only sensitized the parental HCC populations but also the chemo-resistant subpopulations to chemo-therapy induced apoptosis. Importantly, combination of GEP antibody therapy with chemo-therapy inhibited the chemo-therapy induced GEP, ABCB5 and heaptic CSCs marker over-expression through neutralization of the secretary GEP levels in the culture supernatant, and the serum GEP levels in the HCC orthotopic mice model. In human HCC xenograft models, GEP antibody treatment alone is consistently able to inhibit the tumor growth, but is unable to eliminate the established intrahepatic tumor. Cisplatin treatment, low and high dose respectively, was only able to eradicate a fraction of the intrahepatic tumor and the residual tumors grew aggressively after chemo-drug withdrawal. Combination of GEP antibody with low dose of cisplatin resulted in significant proliferation inhibition and apoptosis induction respectively. Importantly, combination of GEP antibody with high dose of cisplatin resulted in eradication of all established intrahepatic tumor.
In addition, chemo-therapy induced the Akt/PKB and MEK/ERK prosurvival pathways, disturbed the balanced between the ratio of pro-apoptotic (Bax) to anti-apoptotic (Bcl-2) member through the induction of Bcl-2. Nonetheless, combination GEP antibody therapy suppressed the chemo-therapy induced phosphorylation of PDK1, Akt, MEK, ERK, and Bcl-2 levels. It was shown that Wortmannin, the PI3K/Akt inhibitor, suppressed the expression of ABCB5 and Bcl-2 induced by chemo-therapy but showed no effect on GEP expression levels.
In summary, the study demonstrated the chemo-therapy treatment alone induced the expression of growth factor GEP, drug transporter ABCB5, hepatic cancer stem cell markers expressions, and the residual cancer cells showed enhanced CSCs properties. Combination treatment with GEP antibody reversed the signaling and cancer stem cell properties induced by chemo-therapy alone. Therefore, further investigations of this combination treatment approach may lead to the development of novel therapeutic approach for the clinical treatment of chemo-resistant HCC.published_or_final_version
Surgery
Doctoral
Doctor of Philosophy
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McDermott, Ultan. "The role of Fas in chemotherapy-induced colorectal cancer cell death." Thesis, Queen's University Belfast, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.426703.
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Gomes, Ana Rita. "The role and regulation of FOXO3a in cancer and chemotherapy resistance." Thesis, Imperial College London, 2012. http://hdl.handle.net/10044/1/14582.
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Drug resistance is the major impediment to the success of cancer therapy. The PI3K/AKT pathway mediates a plethora of cellular functions, including cell survival, proliferation and differentiation. However, increased activation of this pathway has been correlated with drug resistance mechanisms. This pathway regulates the activity of FOXO transcription factors in a negative manner through AKT-dependent phosphorylations. The modulation of FOXO activity leads to a variety of cellular outputs, including cell cycle arrest and apoptosis that define this transcription factor as a tumour suppressor. Importantly, FOXO has also been shown to mediate the effect of many anti-cancer drugs, suggesting that it has an additional role in drug sensitivity and resistance. With this work, by studying the PI3K/AKT/FOXO axis in breast cancer, I have characterised its impact in drug sensitivity and resistance. I found that this axis is deregulated in breast cancer resistant cells. By extending my in vitro findings to clinical samples, I further elucidated the potential role of AKT and FOXO3a as indicators and predictors of treatment response in breast cancer. In addition, I have also characterised three novel downstream targets of FOXO3a - FOXP1, FOXM1 and VEGF – with important roles towards breast cancer progression and in the development of drug-resistance. By characterising these FOXO3a effectors, I unravelled a potential general mechanism by which FOXO3a represses gene target expression.
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Al, Alami Usama Akram. "The role of nitric oxide in tumour biology." Thesis, University of Wolverhampton, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.327151.
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Govender, Yogeshni. "The role of short-term starvation in sensitizing breast cancer to chemotherapy." Thesis, Stellenbosch : Stellenbosch University, 2013. http://hdl.handle.net/10019.1/80007.
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Thesis (MSc)--Stellenbosch University, 2013.ENGLISH ABSTRACT: Introduction: Breast cancer is a major contributor to mortality in women worldwide. Although, anthracyclines, such as doxorubicin, are among the most valuable treatments for breast cancer, their clinical use is limited due to detrimental side-effects such as cardiotoxicity. Additionally, evidence suggests that cancer cells are becoming increasingly resistant to chemotherapeutic agents. The consequence of poor vascularisation within tumours subsequently leads to a nutrient deprived microenvironment which cancer cells are known to adapt to via metabolic remodelling and increasing autophagy. Autophagy is an intracellular degradation system, which is induced as a survival mechanism in response to starvation and other environmental stressors. Recent studies have shown that starvation protects non-tumourigenic cells against chemotherapy-induced cell death. Furthermore, patients who starved prior to chemotherapy reported reduced side-effects. However, these studies investigated the effects of long-term starvation, which maybe clinically challenging. Therefore, this concept, under shorter and more tolerable periods of starvation still needs to be investigated. We hypothesis, that short-term starvation will sensitize breast cancer cells to doxorubicin-induced cell death. In order to test this hypothesis this study was approached by the following aims: (i) to establish a time point at which MCF12A breast epithelial cells are protected against starvation; (ii) to determine the effect of short-term starvation on doxorubicin induced cell death; (iii) to assess autophagy and; (iv) to assess these above mentioned aims using an in vivo model. Methods: MDAMB231 cells and MCF12A cells were starved for 0, 3, 6, 12, 24 and 48 hours using Hanks Balanced Salt Solution. Cell viability was assessed using the trypan blue, MTT and Caspase-Glo assays. MDAMB231 cells and MCF12A cells were subjected to the following conditions: (1) control; (2) 5 μM doxorubicin; (3) starvation of 3 hours and (4) a combination of starvation and doxorubicin. Following treatment an MTT assay to assess cell viability was performed. MDAMB231 cells were further examined using Live-Cell Imaging and western blot analysis. C57BL6 tumour bearing mice were treated with doxorubicin (5 mg/kg) or in combination with starvation of 24 hours. Upon termination of the protocol, tumour tissue was assessed using western blot analysis. In both in vitro and in vivo analyses cleaved-caspase 3 and cleaved-PARP were used as markers for apoptosis, LC3 and p62 as autophagic markers and p-AMPK and p-mTOR as markers of oxygen and energy sensing, respectively. Results and discussion: Three hours of starvation was chosen for in vitro experiments since no significant reduction in cell viability or increases in apoptosis occurred at this time-point in the normal MCF12A breast epithelial cells. As expected, doxorubicin induced a significant decrease in cell viability in the cancerous MDAMB231 cells. Short-term starvation in combination with doxorubicin treatment caused a further significant decrease in cell viability in MDAMB231 cells compared to the doxorubicin group alone. Interestingly, starved MCF12A cells were protected against doxorubicin-induced cytotoxicity as cell viability significantly increased. A significant decrease in autophagy was further observed with the combined treatment of doxorubicin and starvation which corresponded with a significant increase in cell death. In contrast, although the in vivo study also demonstrated a significant elevation in cell death and autophagy in response to doxorubicin treatment, the combined treatment (starvation and doxorubicin) did not have an additive effect when compared to the doxorubicin group alone. Conclusion: Our in vitro results clearly demonstrate that short-term starvation sensitizes breast cancer cells to doxorubicin-induced cell death. Additionally, decreased levels of autophagy appear to contribute to this phenomenon of sensitization. Although doxorubicin treatment resulted in increased apoptosis in vivo, 24 hours starvation in combination with doxorubicin did not sensitize the tumours to doxorubicin treatment. Thus, for future in vivo studies more time points should be considered in order to translate the beneficial effects of short-term starvation observed in our in vitro study to an animal model.
AFRIKAANSE OPSOMMING: Inleiding: Borskanker is ‘n belangrike faktor wat bydrae tot sterftes in vrouens wêreldwyd. Alhoewel antrasikliene soos doxorubicin, waardevol is vir die behandeling van borskanker, word die kliniese gebruik daarvan beperk deur newe-effekte soos kardiotoksisiteit. Verder, word daar al hoe meer bewys dat kankerselle toenemend weeerstandbiedend word teen chemoterapeutiese middels. Swak vaskularisasie van tumore lei tot ‘n mikro-omgewing met beperkte voedingstowwe waaby kankerselle kan aanpas deur middel van metaboliese hermodelering en ‘n toename in autofagie. Autofagie is ‘n intrasellulêre degraderingsisteem wat as ‘n oorlewingsmeganisme aangewend word tydens verhongering en ander omgewingstressors. Onlangse studies het getoon dat verhongering nie-tumourigeniese (normale) selle teen chemoterapie-geïnduseerde seldood beskerm. Verder is daar ook geraporteer dat pasiënte wat gevas het voor chemoterapie, verminderde newe-effekte getoon het. Hierdie studies het egter gefokus op ‘n relatief lang-termyn vas, wat klinies nogal uitdagend kan wees. Daarom moet hierdie konsep nog op korter, meer hanteerbare tye getoets word. Ons hipotese is dus dat kort-termyn vas borskankerselle kan sensitiseer tot doxorubicin-geïnduseerde seldood. Om hierdie hipotese te toets, is die volgende doelwitte gestel: (i) om ‘n tydspunt te bepaal waar MCF12A borsepiteelselle beskerm is teen verhongering; (ii) om die effek van kort-termyn verhongering op doxorubicin-geïnduseerde seldood te toets; (iii) om autofagie te karakteriseer in ons model en; (iv) om hierdie doelwitte ook in ‘n in vivo model te toets. Metodes: MDAMB231 en MCF12A selle is verhonger vir 0, 3, 6, 12, 24 and 48 ure deur van Hanks se gebalanseerde soutoplossing gebruik te maak. Sellewensvatbaarheid is bepaal deur middel van trypan blou, MTT en die Caspase-Glo tegnieke. MDAMB231 en MCF12A selle is onderwerp aan die volgende omstandighede: (1) kontrole; (2) 5 μM doxorubicin; (3) verhongering van 3 ure en (4) ‘n kombinasie van verhongering en doxorubicin. Na behandeling is die sellewensvatbaarheid deur middel van die MTT tegniek bepaal. MDAMB231 selle is verder ondersoek deur middel van “Live-Cell Imaging” en die westelike klad tegniek. C57BL6 tumor-draende muise is behandel met doxorubicin (5 mg/kg) of met ‘n kombinasie van verhongering van 24 ure en doxorubicin. Aan die einde van die protokol, is die kankerweefsel geanaliseer deur die westelike klad tegniek. In beide in vitro en in vivo analises, is gekliefde- caspase 3 en -PARP as merkers vir apoptose, LC3 and p62 as merkers vir autofagie en p-AMPK en p-mTOR as suurstof- en energie sensors respektiewelik gemeet. Resultate en bespreking: Vir die in vitro eksperimente, is ‘n tydspunt van 3 ure gekies as gevolg van die feit dat geen afname in sellewensvatbaarheid en ‘n toename in apoptose in hierdie tydsgleuf tydens verhongering in die normale MCF12A borsepiteelselle plaasgevind het nie. Soos verwag, het doxorubicin behandeling ‘n insiggewende afname in sellewensvatbaarheid in die kankeragtige MDAMB231 selle veroorsaak. Die kombinasie-terapie van verhongering en doxorubicin het ‘n verdere verhoging in seldood teweeg gebring in die MDAMB231 selle, maar het die normale MCF12A borsepiteelselle teen doxorubicin-geïnduseerde toksisiteit beskerm. Die kombinasie-behandeling is ook geassosieer met ‘n afname in autofagie. Alhoewel, die in vivo studie ook getoon het dat doxorubicin alleen insiggewende hoeveelheid seldood teweeggebring het, het die kombinasie-behandeling nie die additiewe effek, soos in die in vitro studie, teweeg gebring nie. Gevolgtrekking: Die in vitro resultate het duidelik getoon dat kort-termyn verhongering borskankerselle kan sensitiseer vir doxorubicin terapie. Verder het dit geblyk dat ‘n afname in autofagie tot die fenomeen van sensitisering bygedrae het. Alhoewel doxorubicin behandeling in vivo tot ‘n toename in apoptose in die tumor gelei het, het die kombinasie behandeling nie die kankerweefsel ten op sigte van doxorubicin gesensitiseer nie. Daar sal dus vir toekomstige in vivo studies meer tydsgleuwe van behandeling ondersoek moet word om die optimum verhongeringsperiode te vind sodat die in vitro resultate ook in vivo van toepassing kan wees.
NRF and CANSA for financial support
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Kondratska-Klymenko, Kateryna. "Role of calcium-permeable channels in pancreatic ductal adenocarcinoma resistance to chemotherapy." Thesis, Lille 1, 2015. http://www.theses.fr/2015LIL10099/document.
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L'adénocarcinome pancréatique (PDAC) est la forme la plus fréquente de néoplasme de cet organe puisqu’il représente environ 90% de toutes les tumeurs pancréatiques et constitue l'une des principales causes de décès par cancer chez l’Homme. Le taux de survie à 5 ans n’est que de 6%. L'une des raisons à cela est que, au début du développement du cancer du pancréas, il n'y a pas de symptômes, et donc la majorité des cas sont diagnostiqués à des stades tardifs métastatiques ou invasifs pour lesquels une intervention chirurgicale n’est plus possible. Il a été montré que les cellules du cancer du pancréas présentent plusieurs mutations génétiques qui conduisent à la prolifération incontrôlée des cellules, ainsi qu’à l'évasion de l'apoptose. Les changements de concentration du Ca2+ cytosolique jouent un rôle central dans de nombreux processus cellulaires fondamentaux, et la perturbation des mécanismes de régulation de l'homéostasie du Ca2+ conduit à une grande variété de pathologies graves, dont le cancer. C’est notamment le cas pour les canaux calciques de type SOC, qui régulent une variété de processus cellulaires dépendants du calcium. Cependant, bien que le rôle du Ca2+ et des canaux calciques soit bien établi dans de nombreuses voies de signalisation de différents types cellulaires, les informations sur le rôle des canaux calciques dans le PDAC sont limitées. Donc, l'identification de la nature moléculaire ainsi que des fonctions des canaux calciques revêt une grande importance dans ces cellules car elle pourrait à termes fournir de nouvelles approches relatives au traitement du cancer du pancréas par le ciblage des processus dépendants du calciumPancreatic ductal adenocarcinoma (PDAC) representing the most prevalent pancreatic neoplasm accounting for about 90% of all pancreatic tumors, is one of the leading causes of cancer death in men and women. The current five-year relative survival rate is about 6% . One of the reasons of this is that early stage pancreatic cancer usually has no symptoms and thus the majority of cases are diagnosed at the late metastatic or invasive stages which are not suitable for surgery. Pancreatic cancer cells have been shown to exhibit a number of genetic mutations leading to uncontrolled cell proliferation, as well as evasion of programmed cell death (apoptosis). Changes in the cytosolic free Ca2+ concentration, play a central role in many fundamental cellular processes and disturbance of the Ca2+ homeostasis regulatory mechanisms leads to a vast variety of severe pathologies, including cancer. Among these, store-operated calcium channels (SOCs) have been shown to regulate a variety of calcium dependent cellular processes altered in different cancers. However, although the role of Ca2+ and calcium-permeable channels is well established in many signaling pathways in a variety of cell types, the information of the role of calcium-permeable channels in PDAC cells is limited. Therefore, identification of the molecular nature as well as functions of calcium-permeable channels in these cells is of great importance as it can reveal novel approaches for treating pancreatic cancer through targeting calcium-dependent processes
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Weaver, Andrew. "Dose intensive chemotherapy in ovarian cancer and the role of stem cell factor." Thesis, Imperial College London, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.287931.
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Wilson, T. R. "The role of c-FLIP in mediating resistance to chemotherapy in colorectal cancer." Thesis, Queen's University Belfast, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.426918.
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Kennedy, R. D. "Investigation of the role played by BRCA1 in modulating the response to chemotherapy." Thesis, Queen's University Belfast, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.411374.
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Lo, Maisie K. Y. "Role of transporters in pancreatic cancer drug resistance." Thesis, University of British Columbia, 2007. http://hdl.handle.net/2429/361.
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Pancreatic cancer (PC) is known to be highly resistant to chemotherapy. Transporters, which regulate the influx and efflux of substrates across the plasma membrane, may play a role in PC drug resistance. ABC transporters are a large family of transmembrane proteins with diverse physiological functions, several of which play major roles in cancer drug resistance. Given that 90% of PC express a mutant K-ras oncogene and that PC are highly hypoxic, I postulated that constitutive K-ras activation and/or hypoxia may correlate with ABC transporter expression, which in turn may promote drug resistance in PC. Using normal and PC cell lines either overexpressing mutant K-ras or subjected to hypoxic treatment, mRNA expression was profiled for 48 ABC transporters. My findings indicate that expression of mutant K-ras and hypoxic treatment, as well as long-term exposure to chemotherapy, may contribute to the development of drug resistance in PC cells in part by inducing the expression of ABC transporters.
Similar to ABC transporters, I investigated whether amino acid transporters would mediate drug resistance in PC. The xc" amino acid transporter (xc") mediates cellular uptake of cystine for the biosynthesis of glutathione, a major detoxifying agent. Because the xc" has been regulates the growth of various cancer cell types, and x," is expressed in the pancreas, I postulated that the xc" may be involved in growth and drug resistance in PC. The xc" transporter is differentially expressed in normal pancreatic tissues and is overexpressed in PC in vivo. UsingPC cell lines, I found that cystine uptake via the N.: was required for growth and survival in response to oxidative stress, and that expression of the xc" correlated with gemcitabine resistance. Accordingly, inhibition of xc" expression via siRNA reduced PC cell proliferation and restored sensitivity to gemcitabine. I also identified the anti-inflammatory drug sulfasalazine as a mixed inhibitor of the x,-, which acts to inhibit cell proliferation via reducing xc" activity and not by reducing NFKB activity. My findings thus indicate that the xc" plays a role in PC growth in part by contributing to glutathione synthesis to promote PC cell proliferation, survival, and drug resistance.
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Finn, Nnenna Adimora. "Role of redox systems in doxorubicin metabolism and doxorubicin-mediated cell signaling: a computational analysis." Diss., Georgia Institute of Technology, 2011. http://hdl.handle.net/1853/41149.
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Insensitivity to chemotherapy is an ongoing issue in cancer treatment, one that appears to be highly dependent on patient-specific variations. It has been shown clinically that while a subset of patients will successfully respond to a particular chemotherapeutic regimen, there exists another subset of patients who when exposed to the same course of therapy will remain resistant to treatment or exhibit signs of relapse after treatment has been administered. This discrepancy raises interesting questions regarding the role that patient-specific variations play in controlling the efficacy of chemotherapy treatment regimens. Doxorubicin (Dox) is a common chemotherapeutic agent used in the treatment of a variety of solid tumors and leukemias and resistance to Dox treatment is a major issue in cancer chemotherapy, oftentimes leading to patient relapse. To gain a deeper understanding of the processes that influence Dox resistance, we must first understand the mechanisms that underlie and contribute to Dox's toxicity. To this end, the metabolic reactions that activate Dox have been implicated as major determinants of Dox cytoxicity and as possible factors that control Dox resistance in cancer cells.
There are several lines of evidence that redox-dependent metabolism plays a large role in Dox toxicity. The Dox bioactivation network is comprised of a system of reduction/oxidation (redox) reactions that lead to the formation of toxic Dox metabolites and reactive oxygen species (ROS). Moreover, multi-drug resistant acute lymphoblastic leukemia cells derived from relapsed patients have elevated levels of the antioxidant glutathione and show insensitivity to Dox treatment. The redox dependence of Dox bioactivation, the understanding that Dox treatment generates ROS, and the evidence that Dox resistant cells exhibit increased antioxidant capacity, suggest the possibility that redox pathways modulate the efficacy of Dox treatment in cancer cells. The overall objectives of the proposed dissertation, therefore, were to investigate how the redox properties of the Dox bioactivation network influence Dox toxicity in acute lymphoblastic leukemia cells, and to provide evidence that cell-specific variations in the intracellular levels of these redox components influences the degree to which Dox treatment will induce cancer cell death.
The significant findings of this study are that the redox reactions involved in Dox metabolism are dual-natured, containing a toxicity-generating module characterized by nicotinamide adenine dinucleotide phosphate (NADPH)-dependent Dox reductive conversion, as well as an ROS signal-generating module characterized by NADPH- and oxygen-dependent Dox redox cycling. The balance between the coupled redox reactions that comprise the toxicity- and ROS signal-generating modules of Dox bioactivation determines the sensitivity-phenotype of leukemia cells and phenotypic changes in the Dox-sensitivity of leukemia cells can be induced by the successful modulation of the Dox bioactivation network through the pharmacological inhibition of NADPH in a concentration- and cell type-dependent manner.
This study highlights the importance of the intracellular redox network in controlling chemotherapy-induced ROS. The unequal distribution in antioxidant burden across the various intracellular antioxidant enzymes suggests a significant role for NADPH supply, as controlled by the enzyme glucose-6-phosphate dehydrogenase (G6PD), to the intracellular ROS buffering capacity of cells during instances of oxidative stress. Changes in G6PD activity were shown to promote protein-S-glutathionylation during oxidative stress conditions, thereby implicating G6PD in the modulation of redox-sensitive signal transduction pathways. The intracellular glutathione redox balance, a measure of the intracellular redox environment, can effectively regulate Dox-induced NF-κB signal transduction in leukemia cells. The systematic modulation of intracellular glutathione redox balance in leukemia cells by N-acetylcysteine (NAC) revealed an important role for protein S-glutathionylation mechanisms in the control of NF-κB signal transduction induced by Dox treatment. These findings identify the glutathione redox network as a potential therapeutic target for the systematic modulation of Dox sensitivity in cancer cells and elucidate the complex role that antioxidants such as NAC can play in modulating the effectiveness of Dox chemotherapy treatment regimens.
Lastly, this study highlights the need for and the capacity of computational models to accurately describe the complex redox-reactions that contribute to Dox metabolism in leukemia cells. This study is groundbreaking in its use of computational modeling to analyze reversible electron transfer events between proteins using mass-action kinetics. The models developed in this study can accurately explain cytosolic doxorubicin bioactivation, intracellular hydrogen peroxide clearance, and kinase-specific S-glutathionylation, thereby showing that the use of comprehensive and/or relatively simple computational models can provide semi-quantitative predictions about the behavior of redox systems in mammalian cells as they relate to Dox-induced toxicity and Dox-induced cell signaling.
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Doherty, J. E. "The role of ADAM 17 in regulating resistance to Chemotherapy Treatment in Colorectal Cancer." Thesis, Queen's University Belfast, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.527684.
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Tse, Tak-fong, and 謝德芳. "Role of RON activation on chemoresistance in gastric cancer." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2007. http://hub.hku.hk/bib/B38592253.
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Yang, Sitian, and 楊斯恬. "Novel role of drug-induced non homologous end joining factor 1 in the chemoresistance of hepatocellular carcinoma." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2015. http://hdl.handle.net/10722/209505.
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Hasim, Mohamed Shaad. "The Role of Activating Transcription Factor 3 as a Regulator of DNA-Damaging Chemotherapy Cytotoxicity." Thesis, Université d'Ottawa / University of Ottawa, 2019. http://hdl.handle.net/10393/39897.
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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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Errington, Fiona. "An investigation into the cytotoxic mechanisms of DNA topoisomerase II poisons and catalytic inhibitors : the role of DNA topoisomerase II alpha and beta." Thesis, University of Newcastle Upon Tyne, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.340718.
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Tai, Kin-ki Emily, and 戴健琦. "Defining the protective role of cathelicidin on ulcerative colitis in mice." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2007. http://hub.hku.hk/bib/B40203542.
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Carser, J. C. "The role of BRCA1 as a marker of clinical outcome following chemotherapy in sporadic ovarian cancer." Thesis, Queen's University Belfast, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.517246.
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Hotama, Peggy Yulia [Verfasser]. "The role of Herpes Simplex Virus in chemotherapy-induced Mucositis in pediatric patients / Peggy Yulia Hotama." Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2014. http://d-nb.info/1052530168/34.
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Hall, Andrew G. "The role of glutathione S-transferases in the development of resistance to chemotherapy in lymphoid malignancies." Thesis, University of Newcastle Upon Tyne, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.287172.
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Sobhakumari, Arya. "Dual Role of Oxidative Stress in Head and Neck Cancer Chemotherapy: Cytotoxicity and Pro-survival Autophagy." Diss., University of Iowa, 2013. https://ir.uiowa.edu/etd/4913.
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Cancer cells are believed to exist in a condition of metabolic oxidative stress compared to normal cells because of inherent mitochondrial dysfunction. Cancer cells up regulate antioxidant defense mechanisms to combat the toxic effect of reactive oxygen species (ROS). Many anticancer agents block ROS detoxification mechanisms and utilize oxidative stress to cause cytotoxicity to cancer cells. However, ROS also up-regulate many pro-survival signaling pathways that may mediate resistance to chemotherapy. I hypothesize that ROS induces both cytotoxicity and pro-survival mechanisms in cells treated with chemotherapeutic agents such as the EGFR inhibitor erlotinib. This thesis explores how oxidative stress may induce both pro-survival and pro-death mechanisms in HNSCC cells and how this can be exploited to increase the cytotoxicity of erlotinib. The combined use of buthionine-[S,R]-sulfoximine, an inhibitor of glutathione and auranofin, an inhibitor of thioredoxin metabolism enhanced human head and neck cancer cell killing by a mechanism involving oxidative stress both in vitro and in vivo and sensitized cells to erlotinib in vitro. However, in other studies erlotinib as a single agent induced oxidative stress and this was mediated by NADPH oxidase 4 (NOX4). NOX4 mediated oxidative stress activated a process called autophagy which protected cancer cells from cytotoxic effect of erlotinib and inhibition of autophagy sensitized cells to erlotinib in vitro. These studies show that oxidative stress may have a dual role in cancer chemotherapy. ROS generated from various drug treatments can cause oxidative damage of cells culminating in cell death. However, it may also activate autophagy protecting cells against the stress and leading to decreased efficacy of the treatment. Hence inhibiting autophagy and hydroperoxide metabolism can be effective treatment modalities to enhance the cytotoxicity of erlotinib and achieve maximum therapeutic efficacy.
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Yao, Jia. "Synthesis of silver nanoparticles and their role against a thiazolekinase enzyme from Plasmodium falciparum." Thesis, Rhodes University, 2014. http://hdl.handle.net/10962/d1020894.
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Malaria, a mosquito-borne infectious disease, caused by the protozoan Plasmodium genus, is the greatest health challenges worldwide. The plasmodial vitamin B1 biosynthetic enzyme PfThzK diverges significantly, both structurally and functionally from its counterpart in higher eukaryotes, thereby making it particularly attractive as a biomedical target. In the present study, PfThzK was recombinantly produced as 6×His fusion protein in E. coli BL21, purified using nickel affinity chromatography and size exclusion chromatography resulting in 1.03% yield and specific activity 0.28 U/mg. The enzyme was found to be a monomer with a molecular mass of 34 kDa. Characterization of the PfThzK showed an optimum temperature and pH of 37°C and 7.5 respectively, and it is relatively stable (t₁/₂=2.66 h). Ag nanoparticles were synthesized by NaBH₄/tannic acid, and characterized by UV-vis spectroscopy and transmission electron microscopy. The morphologies of these Ag nanoparticles (in terms of size) synthesized by tannic acid appeared to be more controlled with the size of 7.06±2.41 nm, compared with those synthesized by NaBH₄, with the sized of 12.9±4.21 nm. The purified PfThzK was challenged with Ag NPs synthesized by tannic acid, and the results suggested that they competitively inhibited PfThzK (89 %) at low concentrations (5-10 μM) with a Ki = 6.45 μM.
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Santos, Cravo Ana Maria. "The role of epithelial cell de-differentiation in the context of improved chemotherapy applied to pancreatic cancer." Thesis, University of Bath, 2015. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.642055.
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In cancer, epithelial cell de-differentiation is a feature of rapidly dividing cells under non-controlled growth and it often reflects a change in the gene expression pattern; however, the relationship between proliferation and alterations in cellular differentiation has not yet been identified. This work examined how changes in the characteristics of cells that discriminate their differentiated and proliferative states can be used to improve on current pancreatic cancer chemotherapeutic strategies. PepT1, a high substrate-capacity and low-affinity transporter system, has been suggested as an attractive drug delivery target for pancreatic cancer. Through a combination of immunological assays, PepT1 normally restricted to the apical surfaces in polarised intestinal epithelial cells, was shown to distribute at the cell membrane of non-polarised cancerous ductal cells. Anti-inflammatory or anti-cancer agents, like ibuprofen or gemcitabine, were conjugated to selected amino acids to enhance their uptake via PepT1. Studies with these conjugates demonstrated enhanced uptake into pancreatic cancer cells, AsPc-1 and HPAFII. Subsequent studies investigated how cell polarity that is typically disrupted in cancer can be modulated to affect the balance of epithelial differentiation versus proliferation. Pharmacological attenuation of YAP (c-Yes associated protein) using a β adrenergic agonist, dobutamine, increased functional tight junction (TJ) structures and diminished proliferation rates of two pancreatic cancer cells, AsPc-1 and HPAFII. Dobutamine also primed an apoptotic cell response. When given in combination with gemcitabine, dobutamine further reduced cell proliferation. Overall, these studies have provided support for using PepT1 as a method to target pancreatic cancer cells for the delivery of anti-cancer agents. Additionally, dobutamine was identified as a potential pharmacological agent to suppress the proliferation of pancreatic cancer cells by altering increasing cell programming that drives epithelial cell differentiation.
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Rodrigues, Andréia Catarina Dias. "Predictive factors for chemotherapy-induced oral mucositis in colorectal cancer patients: role of inflammatory related genes polymorphisms." Master's thesis, Instituto de Ciências Biomédicas Abel Salazar, 2008. http://hdl.handle.net/10216/9150.
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Forestrania, Roshamur Cahyan. "Secondary Metabolites from Garcinia daedalanthera Pierre Leaves and Their Role in Cancer Chemotherapy and Chemoprevention." The Ohio State University, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=osu1557191838034824.
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Middleton, Justin David. "Defining the early stages of chemotherapy-exacerbated cancer colonization of the lung: A role for host-ATF3." The Ohio State University, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=osu1594999911940819.
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Rodrigues, Andréia Catarina Dias. "Predictive factors for chemotherapy-induced oral mucositis in colorectal cancer patients: role of inflammatory related genes polymorphisms." Dissertação, Instituto de Ciências Biomédicas Abel Salazar, 2008. http://hdl.handle.net/10216/9150.
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Yarana, Chontida. "ROLE OF OXIDIZED EXTRACELLULAR VESICLES AS EARLY BIOMARKERS AND INFLAMMATORY MEDIATORS IN CHEMOTHERAPY-INDUCED NORMAL TISSUE INJURY." UKnowledge, 2018. https://uknowledge.uky.edu/toxicology_etds/23.
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Significant advances in the efficacy of cancer therapy have been accompanied by an escalation of side effects that result from therapy-induced injury to normal tissues. Patients with high grade cancer or metastasis are often treated with chemotherapy, 50% of which are associated with reactive oxygen species generation and cellular oxidative stress. Heart is the normal tissue most susceptible to chemotherapy-induced oxidative stress and heart disease is the most common leading cause of death in cancer survivors. However, early and sensitive biomarkers to identify heart disease are still lacking. Extracellular vesicles (EVs) are released from cells during oxidative stress and send oxidized proteins into the circulation as a compensatory mechanism that prevents cellular proteotoxicity. Thus, the protein contents of EVs released during the pre-degeneration stage reveal that oxidative stress is occurring early in the damaged tissue. Using a mouse model of doxorubicin (DOX)-induced cardiac injury, we demonstrated that EVs can be used as an early diagnostic tool for tissue injury as they are oxidatively modified with 4-hydroxynonenal and contain tissue specific proteins—glycogen phosphorylase brain/heart, muscle, and liver isoforms—that indicate their origins. These biomarkers increased early, before the changes of conventional biomarkers occurred.
EVs also mediate intercellular communication by transferring bioactive molecules between cells. In the cell culture system, EVs play an important role in oxidative stress response by inducing macrophage polarization. EVs from cardiomyocytes promoted both proinflammatory (M1) and anti-inflammatory (M2) macrophage polarization evidenced by higher pro- and anti-inflammatory cytokines and nitric oxide generation, as well as mitochondrial oxidative phosphorylation suppression and glycolysis enhancement. In contrast, EVs from the hepatocytes supported anti-inflammatory macrophage (M2) by enhancing oxidative phosphorylation and anti-oxidant proteins. DOX promoted the immunostimulatory effects of cardiomyocyte EVs but not hepatocyte EVs. The differential functions of EVs on macrophage phenotype switching are due to their different effects on Thioredoxin 1 redox state, which regulates activities of redox sensitive transcription factors NFκB and Nrf-2. Our findings shed light on the role of EVs as a redox active mediator of immune response during chemotherapy.
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Ingram, Judy. "An explanation of the role of family participation in a medication information program on schizophrenic clients' medication adherence behaviors." Thesis, The University of Arizona, 1987. http://hdl.handle.net/10150/276561.
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The purpose of this study was to describe family members' influence on medication adherence rates for schizophrenic clients following an educational program presented simultaneously to both client and family member. Three chronic schizophrenic clients participated in this study, two were females, and the other was male. The two family members who provided data were husbands of the two female clients. The obtained scores and responses of two questionnaires was indicative of improved medication adherence for clients and family members. The level of adherence was similarly perceived by the clients and their family members as obtained scores and responses were similar across both time periods. However, family members' attendance at the program presentation did not influence the level of reported medicated adherence behaviors of their wives as compared to the client who attended the program alone because the greatest increase in obtained scores was reported by the client who attended the program alone.
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Hassan, Mohammed Hashim Abdalraheem. "Characterization of ATP-binding cassette drug transporters and their role in breast cancer treatment using in silico approach." University of the Western Cape, 2019. http://hdl.handle.net/11394/7255.
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>Magister Scientiae - MScBreast cancer is the most common cancer in women worldwide, and is the second most common cancer in the world, responsible for more than 500 000 deaths annually. Estimates are that 1 in 8 women will develop breast cancer in their lifetime. In South Africa, breast cancer in women affects about 16.6 % of the population and could see a 78 % increase in cases by 2030. Comprehensive therapy on breast cancer including surgical operation, chemotherapy, radiotherapy, endocrinotherapy, etc. could help, but still has serious side effects. The Chemotherapy resistance against anticancer drugs is an emerging concern. Biomarkers have been identified as a viable option for early detection and progression of disease. Examples of biological indicators for disease could be the ATP-binding cassette (ABC) drug transporters that utilizes the energy derived from ATP hydrolysis to efflux many chemically diverse compounds across the plasma membrane, thereby playing a critical and important physiological role in protecting cells from xenobiotics. These transporters are also implicated in the development of multidrug resistance (MDR) in cancer cells that have been treated with chemotherapeutics. High expression of these membrane proteins as a family of ABC drug transporters are one of the main reasons for drug resistance by increasing the efflux rate of the anti-cancer drug from cancer cells. ABC drug transporters are considered to be one of the largest protein families in living organisms. There are 48 genes in the human genome that encode ABC transporters, which are divided into seven subfamilies (ABCA-ABCG). Studies revealed that ABC transporter genes has been shown to be associated with tumour development, progression and response to therapy, suggesting their possible use as diagnostic, prognostic and predictive biomarkers. The aim of this study was to investigate and identify novel ABC transporter genes that could be implicated in breast cancer and MDR and potentially would be a therapeutic target for successful chemotherapy treatment and disease progression and survival in breast cancer patients. An in silico approach was used to identify 10 ABC transporter genes (ABCB2, ABCB9, ABCB10, ABCC1, ABCC4, ABCC5, ABCC10, ABCC11, ABCC12, ABCD1) implicated in breast cancer by conferring drug resistance through over-expression in cancer cells. The in silico study investigated the tissue expression specificity, protein interaction/s, pathways, and comparative toxicogenomics of the identified ABC transporter genes using several computational software such as Tissue-specific Gene Expression and Regulation (TiGER), the Human Protein Atlas (HPA), Search Tool for the Retrieval of Interacting Genes/Proteins (STRING), and The Comparative Toxicogenomics Database (CTD). The 48 ABC transporter genes were shortlisted through very selective criteria that narrowed the genes down to 10. Differential expression analysis of the genes using TiGER and HPA compared expression in normal versus cancerous tissue of the candidate genes. The result showed that ABCC11 was preferentially expressed in breast tissue with an enrichment value higher than 10.0. The results also showed ABCC10 overexpressed in breast cancer tissue, making these two genes top candidates for further analysis. Result from STRING database showed a strong functional interaction network between the prioritized genes through protein homology, co-expression and text mining as evidence for the observed interactions. Furthermore, the prioritized list of genes was submitted to the CTD for intersectional analysis to obtain the toxicity relationship between the genes and the Tamoxifen as the first line chemotherapeutic treatment for breast cancer. Venn diagrams obtained from CTD showed intersectional relation between ABCB2, ABCC1, ABCC4, ABCC11, and ABCD1 genes and Tamoxifen. Furthermore, an in silico validation of the prognostic/predictive values of the 10 prioritized genes (list 2) was carried out using an online biomarker validation tool and database for cancer gene expression data using survival analysis (SurvExpress) and gene expression based survival analysis web application for multiple cancer (PROGGENE). Results obtained from the PROGGENE survival and predictive analysis showed good prognostic values for the genes ABCB2, ABCC1, ABCC4, ABCC10 and ABCC12 with their significance measured by the probability value (Pv) (0.053, 0.001118, 0.01286, 0.00604, 0.00157 respectively). From this study ABCC1, ABCC4, ABCC5, ABCC10, and ABCC11 genes could serve as putative therapeutic target biomarkers for breast cancer treatment following further in depth analysis. However, the variance in the effectiveness of individual genes suggests that the set of genes would perform better than individual gene in the management of breast cancer. The modulating roles of ABCC4, ABCC5 ABCC10, and ABCC11 in drug induced apoptosis, suggest they could probably play an important role in personalized medicine and could serve as biomarkers to monitor the prognosis and/or therapeutic outcome of chemotherapy drugs in breast cancer patients. The use of modern genomics, proteomics, bioinformatics, and systems biology approaches has resulted in a substantial increase in our ability to identify molecular mechanisms that are involved in MDR in cancer and to find drugs that may block or reverse the development of drug resistance. By using an in silico approach in this study, a list of five ABC transporter genes were identified, of which two (ABCC10 and ABCC11) could potentially serve as prognostic and predictive biomarkers for the management of breast cancer treatment.
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Purcell, C. "Evaluating the role of chemokine and nuclear factor kappa B signalling in mediating chemotherapy resistance in gastrointestinal cancer." Thesis, Queen's University Belfast, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.517058.
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Demgenski, Janine [Verfasser]. "Role of Bim and Kinases in the Synergistic Induction of Cell Death by TRAIL and Chemotherapy / Janine Demgenski." Konstanz : KOPS Universität Konstanz, 2019. http://d-nb.info/1204829349/34.
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Swami, Priyanka. "Understanding the Role of the Receptor for Advanced Glycation End-Products (Rage) in Pancreatic Cancer." Diss., North Dakota State University, 2019. https://hdl.handle.net/10365/29865.
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Expression of the Receptor for Advanced Glycation End Products (RAGE) and is upregulated in a several cancers. Based on published studies, we hypothesized that RAGE, when overexpressed in pancreatic cancer cells, will promote cell proliferation and migration. To study the role of RAGE in pancreatic cancer, we selected the human pancreatic cancer cell-line PANC-1, and stably transfected the cells with full length RAGE to generate model cell-lines that overexpress RAGE. We obtained two cell-lines PANC-1 FLR2 and PANC-1 FLR3 and examined the influence of RAGE on cellular properties. A significant increase in proliferation but a reduction in migratory abilities of PANC-1 FLR2 and PANC-1 FLR3 cells was observed. The increase in proliferation and reduction in migration was reverted upon knockdown of RAGE in PANC-1 FLR2 cells with siRNA specific for RAGE. The reduction in migration was supported by the reduced levels of vimentin and several integrins in RAGE transfected cells. Furthermore, we observed a downregulation in FAK, AKT, ERK1/2 and NF-κB activity.
Growing evidence supports that RAGE is essential for pancreatic cancer progression. It has also been shown that RAGE facilitates pancreatic tumor cell survival by enhancing autophagy and inhibiting apoptosis. The goal of our study was to determine the effect of RAGE inhibition during gemcitabine chemotherapy on the growth of pancreatic tumor. Hence, we investigated the effect of RAGE inhibitors and their combination with gemcitabine in an orthotopic mouse model of pancreatic cancer using mouse pancreatic cancer cell-line KPC 5508. We used two RAGE inhibitors, an anti-RAGE monoclonal antibody (IgG2A11) and a small molecule RAGE inhibitor (FPS-ZM1). We observed a significant reduction in tumor weights of the mice treated with the combination of IgG2A11 and gemcitabine as compared to gemcitabine alone treated mice. The reduction in tumor growth was accompanied with increase in p62 levels (marker of autophagy) and increase in levels of cleaved PARP (marker of apoptosis). We also observed reduction in HMGB1 and phosphorylation levels of ERK1/2 in tumors from the group treated with the combination as compared to the gemcitabine alone treated group.North Dakota State University. College of Health Professions
NIH Grant # P20 GM109024 from the National Institute of General Medicine
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Geng, Wei, and 耿瑋. "The role of proline rich tyrosine kinase 2 (Pyk2) on cisplatin resistance in HCC." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2009. http://hub.hku.hk/bib/B43224106.
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Söderdahl, Gunnar. "Liver transplantation and the role of adjuvant therapy for advanced primary liver tumours /." Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-579-8/.
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Eklund, Birgitta I. "Role of Multiple Glutathione Transferases in Bioactivation of Thiopurine Prodrugs : Studies of Human Soluble Glutathione Transferases from Alpha, Kappa, Mu, Omega, Pi, Theta, and Zeta Classes." Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis : Universitetsbiblioteket [distributör], 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-7102.
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Martinsson, Lisa. "Quality improvement in palliative care : the role of a national quality register and perceptions of information during palliative chemotherapy." Doctoral thesis, Umeå universitet, Onkologi, 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-102264.
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Introduction There is a need in palliative care for development of structured methods to assess quality and support improvement. This need is present both within and outside specialised palliative care. Honest information from physicians is regarded as an important part of palliative care. Information about the transition to end-of-life care (ITEOL), conveyed by physicians to patients, has also been described as a ‘breakpoint conversation’. Quality registers enable monitoring and improvement of quality of care, and clinical research. The Swedish Register of Palliative Care (SRPC) is a Swedish national quality register that collects data from hospitals, hospices, nursing homes and home care, with an end-of-life questionnaire (ELQ) about palliative care content focusing on the last week of life. Aims Study I – The aim was to examine the validity of the ELQ from the SRPC. Study II – The aim was to examine whether participation in the SRPC increases the quality of palliative care over a three-year period regarding eight items in the ELQ. Study III – The aim was to examine whether an educational intervention for physicians and nurses in nursing homes and hospitals increases the proportion of patients who receive ITEOL. Study IV – The aim was to describe how patients with cancer perceive the information they receive from physicians during palliative chemotherapy. Methods Study I – Data from 100 medical records and data from the paper versions of the ELQ filled in at a specialised palliative unit were compared with data reported to the SRPC. Study II – Data from eight items in the ELQ reported to the SRPC from all healthcare units that had reported patients continuously during a three-year period were compared year-by-year with logistic regression. Study III – Two municipalities (in charge of nursing homes) and two hospitals were randomised to receive an interactive half-day course about ITEOL for physicians and nurses. Four hospitals and four municipalities were assigned matched controls. The proportion of patients who received ITEOL before and after the intervention was measured with the ELQ. The effect of the intervention was analysed with a multivariable logistic regression model. Study IV – Fifteen semi-structured interviews with patients with incurable cancer were conducted, transcribed verbatim and analysed with qualitative content analysis. Results Study I – The questions in the ELQ showed a congruity of 22% to 100% when comparing data from medical records with data reported to the SRPC. Eight questions fell below 80%. The paper versions filled in at the unit and the data reported to the SRPC showed a congruity of between 91% and 100%. Study II – The prevalence of six symptoms decreased. The prescription of PRN (pro re nata – ‘as needed’) medications for pain, nausea, anxiety and death rattle increased. A higher proportion of patients died in their place of preference. The patient’s next of kin was more often offered a follow-up appointment after the patient’s death. There was no change in the proportion of patients or next of kin who received ITEOL. Study III – The proportion of patients in the intervention group who received ITEOL increased from 35.1% (during a six-month period before the intervention) to 42.0% (during a six-month period after the intervention). The proportion in the control group increased from 30.4% to 33.7%. The effect of the intervention was significant (p=0.005) in the multivariable model. Study IV – Three categories were disclosed during the analytical process: ‘living with a death sentence’, ‘depending on chemotherapy’, and ‘living with uncertainty’. Conclusions A national quality register has the potential of contributing to quality improvement in palliative care. Study I showed varied validity in the ELQ, implying a need to modify the questionnaire. Study II showed that participation in the SRPC was covariant with quality improvements in end-of-life care over time. Study III showed that more patients received ITEOL after an educative half-day intervention directed to physicians and nurses. Study IV showed that patients undergoing palliative chemotherapy perceived that their disease was incurable and deadly, and that conditions for future treatment and survival were uncertain.
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Barthe, Anaïs. "Role of the p53 family in chemotherapy-related side effects on the enteric nervous system and skeletal muscle homeostasis." Thesis, Strasbourg, 2019. http://www.theses.fr/2019STRAJ072.
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Une des grandes complications des traitements anticancéreux s’explique par les nombreux effets secondaires liés aux composés de chimiothérapie classique dont les sels de platine et les anthracyclines. Ils causent respectivement une altération du système nerveux entérique (SNE) et une sévère atrophie musculaire pouvant être létales ; malheureusement ; leurs origines sont encore mal connues et peu de traitements sont efficaces ou développés pour les limiter. L’objectif de ce travail a été d’identifier les mécanismes moléculaires responsables de la perte neuronale du SNE et de la dégradation abusive des protéines du muscle pour empêcher ces altérations et développer de nouvelles approches thérapeutiques. Ces travaux ont prouvé pour la première fois l’existence de la famille p53 dans le SNE via son implication dans la mort cellulaire liée au cisplatine, ainsi que son rôle majeur dans l’activation du processus catabolique dans le muscle lié à la doxorubicine. Dans l’ensemble, ces travaux montrent l’importance de la famille p53 dans les toxicités des chimiothérapies et font émerger de potentiels outils thérapeutiques basés sur le ciblage de la famille p53One of the major complications of cancer treatments is the important side effects associated with conventional chemotherapy compounds including platinum salts and anthracyclines. They cause respectively an alteration of the enteric nervous system (ENS) and severe muscular atrophy which can be lethal; unfortunately, their origins are still poorly understood, and few treatments are effective or developed to limit them. The objective of this work was to identify the molecular mechanisms responsible for the neuronal loss of the ENS and the excessive degradation of muscle proteins to prevent these alterations and develop new therapeutic approaches. This work proved for the first time the existence of the p53 family in the ENS through its involvement in cisplatin-related cell death, as well as its major role in activating the doxorubicin-related catabolic process in muscle. Overall, these results show the importance of the p53 family in the toxicities of chemotherapy and point out the emergence of potential therapeutic tools based on the targeting of the p53 family
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Fung, Tak-kwan James, and 馮德焜. "Development of anti-HBs in patients with chronic hepatitis B after liver transplantation using lamivudine prophylaxis: the possible role of adoptive immunity transfer." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2003. http://hub.hku.hk/bib/B31980934.
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Pich, Roselló Oriol 1992. "The Role of mutational processes in the evolution of somatic tissues and malignancy." Doctoral thesis, TDX (Tesis Doctorals en Xarxa), 2021. http://hdl.handle.net/10803/671532.
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Cells of our body are constantly exposed to events that can create lesions in the
DNA. If unremoved by the DNA repair mechanisms, the lesion may lead to a
mutational event. Some of these events might drive the transformation of a healthy
cell to a malignant one. This thesis focuses on understanding the generation of
mutations and their positive selection in somatic tissues. First, we study the coding
and noncoding driver landscape of more than 2500 patients. We also attempt to
distinguish driver and passenger mutations in driver genes, and link them to
mutational processes. We then study the mutations related to chemotherapies in
both tumoral and healthy cells, as well as the effects of treatment in clonal
hematopoiesis (CH). We also perform a systematic discovery of CH driver genes.
Lastly, we study the interplay of DNA damage and repair in nucleosome-bound DNA,
uncovering a 10bp periodic mutation rate which might have influenced the genome
composition in eukarya.Les cèl·lules del nostre cos estan constantment exposades a esdeveniments que poden lesionar l’ADN. Si aquestes lesions no són eliminades pels mecanismes de reparació, poden causar mutacions. Alguns d’aquests podrien ajudar a transformar una cèl·lula sana en una maligna. Aquesta tesi doctoral té com objectiu estudiar la generació de les mutacions i com són seleccionades positivament als teixits somàtics. Primer, estudiem el panorama de drivers en zones codificant i no codificant en més de 2500 pacients. També intentem distingir quines mutacions són driver de les passenger, i les relacionem amb els processos mutacionals. Després estudiem les mutacions relacionades amb la quimioteràpia en cèl·lules sanes i tumorals, i també l’efecte que tenen a la hematopoesi clonal. D’aquest últim fenomen en descobrim els gens drivers. Per últim, estudiem la relació entre el dany a l’ADN i els mecanismes de reparació quan la molècula s’empaqueta en nucleosomes. Aquesta relació deixa una periodicitat de 10 parells de bases que pot haver influenciat la composició del genoma dels eucariotes.
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Chen, Jie, and 陈洁. "The role of FOXO3a in the development of chemoresistance in breast cancer." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2011. http://hub.hku.hk/bib/B47234519.
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Breast cancer is the most common malignancy in women and represents one of the major causes of death worldwide. The PI3K-Akt-FOXO3a signalling pathway has been shown to play a crucial role in tumorigenesis and the development of drug resistance in many cancer types. However, previous studies on FOXO3a using breast cancer tissues were controversial. So this study aims at better understanding of the role of FOXO3a in the development of drug resistance, especially endocrine resistance and anthracycline resistance in breast cancer.
Examination of FOXO3a and phosphorylated-Akt (P-Akt) expressions in breast cancer tissue microarrays revealed nuclear FOXO3a was significantly associated with poor prognosis (p=0.014) and lymph node positivity (p=0.052) in invasive ductal carcinoma. Using the tamoxifen and anthracycline-sensitive and -resistant breast cancer cell lines as models, we found that the nuclear accumulation of FOXO3a was associated with enhanced anthracycline-resistance but not tamoxifen-resistance. This was consistent with the finding that sustained nuclear FOXO3a was associated with poor prognosis, as cytotoxic chemotherapy resistance is linked to limited therapeutic options and poor prognosis. We demonstrated a possible feedback mechanism in which induction of FOXO3a activity in the anthracycline-sensitive MCF-7 cells induced Akt phosphorylation and promoted cell proliferation arrest. Using MDA-MB-231-FOXO3a(A3):ER cells in which FOXO3a activity could be induced by 4-hydroxytamoxifen, we showed that FOXO3a induction could up-regulate PI3K-Akt activity but had little effect on cell proliferation, which indicates impaired Akt-FOXO3a axis in chemoresistant cell models.
To further uncover the precise mechanism of Akt-FOXO3a deregulation in the development of chemoresistance, we have explored the post-translational regulation of FOXO3a by miRNAs. Through a series of Gain-and-Loss functional experiments and luciferase reporter assays in vitro, three miRNAs, including miR-222, miR-221 and miR-29a, were found to suppress FOXO3a protein expression through binding directly to FOXO3a 3’UTR. Moreover, the aberrant expressions of the miR-222/221 cluster and miR-29a in drug resistant cell lines could confer a proliferation advantage to cancer cells through suppressing FOXO3a expression. We further demonstrated that FOXO3a as a transcription factor could transactivate the oncogenic miR-222 and miR-221 expressions under certain chemotherapy stimulation. This suggests the existence of a feedback regulatory loop composed of the miR-222/221 cluster and FOXO3a which may not only play a self-protective role under drug treatment in chemosensitive cells, but also partially explain the tolerated nuclear FOXO3a in the breast cancer with poor prognosis.
Taken together, our study suggested that lymph node metastasis and poor survival in invasive ductal breast carcinoma are linked to an uncoupling of the Akt-FOXO3a signalling axis, as in these breast cancers the nuclear-located FOXO3a was unable to induce cell death or cell cycle arrest. We also demonstrated post-translational regulation of FOXO3a by miR-222/221 and miR-29a, while aberrant expressions of miR-222/221 and miR-29a may promote cell resistance to therapy through directly suppressing FOXO3a. FOXO3a could further contribute to the deregulation of the miR-222/221 cluster as a transcription factor in breast cancer. Studying this Akt-FOXO3a-miRNAs signalling circuit will provide us better understanding in predicting and monitoring treatment response in breast cancer and other malignancies.published_or_final_version
Pathology
Doctoral
Doctor of Philosophy
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Steer, Henry John. "Combining chemotherapy with immunotherapy to treat mesothelioma : an investigation into the role of CD4+ T cells in a murine model." Thesis, University of Nottingham, 2013. http://eprints.nottingham.ac.uk/13672/.
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Cytotoxic chemotherapy remains the mainstay of treatment for patients with cancer, however immunotherapy is starting to emerge as an additional modality of treatment. Evidence suggests that chemotherapy can synergise with immunotherapy to improve responses. Although CD8 T cells have been regarded as the main anti-tumour effector cell, the role of CD4 T cells in orchestrating CD8 and other anti-tumour responses is increasingly recognised. However, the CD4 T cell population contains effector and suppressive subsets with diverse and opposing functions. This thesis describes the establishment of a murine mesothelioma model with which to study the effects of different CD4 subsets on anti-tumour immune responses, and investigate their capacity to provide cognate help to tumour antigen specific CD8 T cells. Haemagluttin (HA) specific CD4 T cells from transgenic mice were polarised in vitro into Th1, Th2, Th17 and Treg subsets and adoptively transferred alongside HA specific CD8 T cells into mice bearing HA expressing tumours derived from a mesothelioma cell line. The effects of the different CD4 subtypes on tumour growth and their capacity to provide ‘help’ to CD8 T cells was investigated in a prophylactic treatment model and in the context of treatment with gemcitabine chemotherapy. Results showed that survival and behaviour of in vitro differentiated CD4 subtypes after adoptive transfer was highly variable and that only Th1s displayed anti-tumour activity when injected prophylactically, prior to tumour inoculation. Cytotoxic chemotherapy did not provide a favourable environment for adoptive transfer of in vitro differentiated CD4 cells. No antitumour activity was seen against established tumours, which may have been due to overriding tumour induced immunosuppressive mechanisms. Successful treatment of established tumours that had been treated with chemotherapy required both the provision of HA specific CD8 cells and the prior removal of an established, endogenous regulatory CD4 T cell population.
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Carey, Hulyer Alex Robert. "Bioenergetic coupling in P-glycoprotein: determining the relative position, topography and role of transmembrane helices six and twelve." Phd thesis, Canberra, ACT : The Australian National University, 2018. http://hdl.handle.net/1885/156454.
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Multidrug efflux pump P-glycoprotein (P-gp) has been the subject of significant research interest since the discovery of the protein in multi-drug resistant cancer cells 40 years ago. Overexpression of P-gp can confer resistance to a broad range of structurally and functionally distinct drugs. The mechanism of this polyspecificity is poorly understood, yet the mechanism is known to involve a complex series of binding, hydrolysis and dissociation events that are coupled to an array of conformational changes. A wealth of biochemical data implicates TM6 & 12 as mediators of bioenergetic coupling in P-gp. A major issue in attempts to determine the molecular mechanism is the lack of a high-resolution structure of human P-gp in the basal, inwardfacing state. Several crystal structures of murine P-gp in the basal state are available, and molecular dynamics have been used to simulate the structure of human P-gp. Both of these structural models are subject to controversy. Crystallisation requires solubilisation of P-gp into a detergent micelle, an environment in which the protein has been demonstrated to be inactive; the crystal structures published to date also differ significantly. MD simulations are an increasingly powerful tool for predicting protein structure; however, these must be treated with caution when not supported by experimental data. The present investigation determined the relative position of two transmembrane helices, six and twelve (TM6 & 12), by measuring the distances between residues. Site directed mutagenesis was used to generate nine double mutants, each containing one cysteine on TM6 and one on TM12. Double mutants were expressed using the baculovirus/insect cell expression system. Crude membranes containing P-gp were isolated, followed by purification by affinity chromatography. P-gp mutants were reconstituted into proteoliposomes; retention of function in the P-gp double mutants was confirmed using ATP hydrolysis assays. Inter-residue distance measurements were performed using intramolecular cross-linking with homobifunctional reagents and 5 double resonance EPR. Further ATP hydrolysis assays were used to determine the effect of intramolecular cross-linking between TM6 & 12 on the activity of P-gp. All the double mutants were functional, and most inter-residue distances were measured successfully by both techniques. The topographical map generated was compared to distance measurements in the crystal structures and MD simulations to judge whether either of these models are a good representation of human P-gp in a lipid bilayer. It was found that the 4M1M structure of murine P-gp is the most accurate representation among models derived by crystallography, while MD simulations based on this are closer still to experimental observations. The experimental results strongly suggest that the side chain orientations are accurate in the models in the membrane embedded portion of TM6 & 12. This investigation therefore demonstrated that 4M1Mbased structural models can be relied upon in the important membrane-integral region, which has been shown to be the site of drug binding. This can give greater weight to in silico methods for predicting molecular interactions with P-gp. Further details of the role of the two helices in the P-gp mechanism were added by analysis of functional changes in P-gp containing intramolecular cross-links between TM6 & 12. Specifically, the results presented here indicate that different parts of the helices require relative movement during drug-stimulated ATP hydrolysis in comparison to basal hydrolysis, suggesting that the conformational transitions of P-gp may be different in the two cases.