Browsing by Subject "Epidermaler Wachstumsfaktor-Rezeptor"

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Der Einfluss des Stammzellmarkers ALDH und des EGFR-PI3 Kinase-Akt Signalwegs auf die Strahlenresistenz humaner Tumorzelllinien
(2014) Mihatsch, Julia; Rodemann, H.-Peter
Cancer is the second leading cause of death in industriated nations. Besides surgury and chemotherapy, radiotherapy (RT) is an important approch by which about 60% of patients are treated. The response of these patients to RT is very heterogenous. On the one hand, there are patients with tumors which are radiosensitive and can be cured, but on the other hand patients bear tumors which are quite resistant to radiotherapy. A Radioresistant phenotype of tumor cells causes treatment failure consequently leading to a limited response to radiotherapy. It is proposed, that radiotherapy outcome mainly depends on the potential of radiation on controlling growth, proliferation and survival of a specific population of tumor cells called cancer stem cells (CSCs) or tumor-initiating cells. Based on experimental studies so far reported it is assumed that the population of CSC varies in tumors from different entities and is relatively low compared to the tumor bulk cells in general. According to the CSC hypothesis, it might be concluded that the differential response of tumors to radiotherapy depends on CSC populations, since these supposedly slow replicating cells are able to initiate a tumor, to self renew indefinitely and to generate the differentiated progeny of a tumor. Besides the role of cancer stem cells in radiotherapy response, ionizing radiation (IR) activates the epidermal growth factor receptor (EGFR) and its downstream signaling pathways such as phosphoinositide 3-kinase (PI3K)/Akt, mitogen-activated protein kinase (MAPK) and Janus kinase/Signal Transducer and Activator of Transcription (JAK/STAT) pathways. Among these pathwas, PI3K/Akt is one of the most important pathways involved in post-irradiation survival: Activation of Akt results in activation of DNA-dependent protein kinase, catalytic subunit (DNA-PKcs). DNA-PKcs is a core enzyme involved in repair of IR-induced DNA-double strand breaks (DNA-DSB) through non-homologous end joining (NHEJ). The aim of the present study was to investigate the role of CSCs in resistance of radioselected subclones of non-small cell lung cancer (NSCLC) and breast cancer cells to irradiation. Additionally, the role of EGFR dependent PI3K/Akt/DNA-PKcs signaling in the context of CSC-mediated radiotherapy resistance was investigated. The following major results were obtained: 1) Radioresistant tumor cells from NSCLC-A549 cells as well as SK-BR-3 breast cancer cells could be isolated in vitro by a radioselection process. 2) In line with the proposed CSC biological behaviors radioselected cells presented extended population doubling time and decreased plating efficiency. 3) Among identified potential CSC markers such as CD133, Oct-4, Sox2 or aldehyde dehydrogenase (ALDH) expression, solely expression of the embryonic stem cell marker Oct-4 was increased in the radio-selected SK-BR-3 cells. However, increased ALDH activity but not enhanced ALDH protein expression was associated with radioresis-tance of A549 cells. 4) Respectively, ALDH activity was found to be involved in radio-resistance partially through PI3K pathway. 5) Using an siRNA approach, a differential effect of ALDH1 vs ALDH2 in terms of post-irradiation survival of tumor cells was demonstrated. In this context and in contrast to the role of ALDH2 a prosurvival effect of ALDH1 could be observed. 6) Radioresistance of IR-selected tumor cells was partially mediated through EGFR/PI3K/DNA-PKcs-dependent accelerated repair of DNA-DSBs. Thus, based on the described major findings in this study it is proposed that targeting of PI3K/Akt pathway and ALDH1 might be effective approaches towards overcoming CSC-mediated radiotherapy resistance.
Untersuchungen zum molekularen Wirkmechanismus des Radioprotektors O-Phospho L-Tyrosin : Wechselwirkungen von Phosphotyrosin mit Aktivierungsprozessen des epidermalen Wachstumsfaktorrezeptors
(2008) Wanner, Gabriele; Rodemann, H.-Peter
Summary Cancer is, after cardiovascular diseases, the main cause of death in Germany. The five-year-survival-rate averages at 35% to 45% and is supported by tumour treatment with ionizing irradiation. Radiotherapy is among surgical interventions and chemotherapeutical methods one of the most important treatment procedure for oncological diseases. In spite of technical improvements and modern therapy designs the ineluctable damage of the tumour surrounding normal tissue acts dose limiting and may lead to acute and late normal tissue reactions and associated side effects. Radioprotectors are applied to protect healthy tissue in the radiation field, but should not protect tumour tissue against radiation effects. The potential dose escalation due to enhanced radiation-tolerance would be associated with increased tumour control. The intention of this work was to shed light on the molecular mode of action of the radioprotector O-Phospho L-Tyrosine (pTyr) and the associated modulation of radiation-induced effects on signalling cascades. The data presented provide evidence that preincubation of human fibroblasts with pTyr leads to a significant increase of cell survival after irradiation. The pTyr-mediated radioprotection was found to be dependent on the availability of the tumour suppressor TP53. Only cells characterized by a wildtype TP53, but not cells with mutated TP53 cells were protected by a pTyr-treatment. Given that a large proportion of human tumours express a mutated TP53, more than 50% of tumours are considered to be treated with pTyr and irradiation in combination. Preliminary work on the field of pTyr-induced molecular mechanism showed an interaction between pTyr and the EGFR-associated signalling pathway, which positively influences radiation-induced DNA-repair (Dittmann et al., JBC 2005). Based on these data we investigate the influence of pTyr treatment on a molecular level. The following results were obtained: 1.pTyr treatment stimulates accumulation of the EGFR in the nucleus, which is involved in regulation of DNA-PK-dependent DNA-repair (NHEJ). 2.pTyr-mediated stimulation of DSB-DNA-repair processes is dependent on functional tumour suppressor TP53. 3.Ionizing radiation and pTyr are able to modulate the phosphorylation status of nuclear EGFR at the position No. 654. This phosphorylation site plays an important role in accumulation of EGFR in the nucleus. 4.The isoform ε of the protein kinase C family is responsible for the pTyr- and radiation-induced T654 phosphorylation of the EGFR in the nucleus. 5.The kinase activity of the nuclear PKCε is essential for nuclear EGFR accumulation and EGFR-associated stimulation of the DNA-PK-dependent DNA-repair. 6.The considered second messenger in the pTyr- and radiation-induced activation of PKCε is diacylglycerol, which is limited by its regulatory kinase DGK θ. 7. Diacylglycerol is generated by hydrolyzation of phosphoinositol biphosphate, which can be stimulated by pTyr treatment and ionizing irradiation. 8.pTyr enhances complex formation between nuclear EGFR, phosphorylated form of DNA-PK and DNA after irradiation. Aim of this work was to elucidate molecular vertices of a pTyr treatment in order to promote the clinical application of pTyr in the context of radiooncological therapy. From present data we suggest that combined treatment of TP53-mutated tumours with pTyr and irradiation improves survival of tumour surrounding normal tissue by influencing DNA-repair processes positively and therefore takes advantage in radiotherapy-treatment. Furthermore this work gives a better insight into radiation-mediated and radioprotective signalling pathways and helps to achieve a deeper understanding in molecular mechanisms after irradiation and radiation-associated survival signals.