%0 Generic
%A Kox, Corinne
%D 2010
%F heidok:11198
%K microRNA
%R 10.11588/heidok.00011198
%T Molecular pathways important for the pathogenesis and risk stratification of childhood T-ALL
%U https://archiv.ub.uni-heidelberg.de/volltextserver/11198/
%X Paediatric T-cell acute lymphoblastic leukaemia (T-ALL) is a rare type of cancer that represents 10-15% of ALL cases. Intensified chemotherapy led to improvements of the overall survival rates of childhood T-ALL, reaching approximately 80%. However, the prognosis of relapses remains poor, emphasising the need for new molecular markers that identify high risk patients in the earliest stages of the disease. In addition, understanding of the precise mechanisms of T-ALL leukaemogenesis allows for the development of molecular targeted therapies that could potentially improve the quality of life of children with T-ALL, in general, and the response in patients that develop recurrent disease, in particular. Frequent genetic abnormalities in T-ALL are mutations in the NOTCH pathway which are thought to drive T-ALL development. In the first part of this study, the prognostic and predictive value of mutations in the NOTCH pathway was investigated in T-ALL patients, at initial presentation and at relapse. Activating mutations in the NOTCH1 gene and inactivating mutations in the FBXW7 gene, encoding for a negative regulator of the pathway, were correlated with early treatment response and long-term outcome. At initial presentation, NOTCH1 mutated patients treated according to the ALL-BFM 2000 protocol show improved early treatment response and better long-term outcome. Interestingly, the NOTCH1 effect is restricted to the ALL-BFM 2000 good responders only. In the high risk group, patients with and without NOTCH1 mutations do not show a significant difference in long-term outcome. In contrast, inactivation of the FBXW7 protein correlates with excellent early response, but does not associate with better long-term outcome. At relapse, event-free survival rates were low in the ALL-REZ 2002 study, not exceeding 40%. In particular, FBXW7 mutated patients with recurrent disease showed a poor response to treatment. This study laid foundations for the possible application of molecular markers in T-ALL risk stratification.  In the second part of this PhD work, the effect of up-regulated NOTCH signalling on microRNA expression was analysed to gain insight into the pathogenic mechanisms of NOTCH1 in T-ALL. In a cell-based assay, NOTCH1 was knocked down in a T cell line and mature microRNA expression was monitored by microarray profiling. Two normalisation methods were employed for the microarray analysis. The first method identified 5 microRNAs down-regulated upon NOTCH1 knock-down, including members of the miR-17~92 and miR-106~363 clusters. However, no major changes in miRNA expression were observed when using the second normalisation method. The validation of the microarray experiments by microRNA-specific quantitative PCR confirmed the results from the second microarray normalisation method. In comparison, a qPCR approach detected a down-regulation of primary microRNA transcripts, such as the transcript of the miR-17~92 cluster. In a second setting, the effect of NOTCH1 mutations on microRNA expression was determined in primary T-ALL patient samples. Marginal differences in expression were detected in members of the miR-17~92 cluster and the miR-181 family. Further work will be required to elucidate the mechanism of action of miRNA regulation by NOTCH1 in T-ALL.