Impact of Fusion Genes and other genetic aberrations on Pediatric Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is a morphologically, immunophenotypically and genetically heterogeneous disease of the hematological system. Despite the significant improvements in overall survival rates of childhood AML patients in recent decades, there is still room for improvement in terms of event-free survival of patients150. A comprehensive genetic characterization of specific patient groups and the exploration of suitable model systems promise the identification of appropriate molecules for development of novel targeted therapies, the influence of which can be assessed in future clinical studies. In the present thesis, various genetic and cytogenetic alterations were characterized and their prognostic impact investigated in children and adolescents with newly diagnosed AML. Firstly, patients with KMT2A rearrangement (KMT2A-r) and patients with RUNX1 mutation were studied. The overall and event-free survival of patients with KMT2A-r showed a strong dependence on the KMT2A partner gene. In contrast, no influence of the most common co-occurring mutation - the KRAS mutation – was shown. The expression of CSPG4, a surface protein that was detected in 69% of pediatric AML patients with KMT2A-r, also showed no significant influence on the prognosis of the patients. Genetic characterization of this cohort showed, in addition to KRAS mutations, a lower incidence of AML-typical mutations or the mutually exclusion of KMT2A-r with other genetic aberrations. The cohort of RUNX1-mutated patients showed no prognostic impact of RUNX1 mutations on overall or event-free survival of pediatric AML patients compared to the RUNX1 wild-type group. A strong association between RUNX1 mutation and FLT3-internal tandem duplication (ITD) was demonstrated. In both analyses, KMT2A-r and RUNX1 mutations were shown to be mutually exclusive. Secondly, WT1 mutations were studied in this thesis as the third, prognostic-relevant alteration in pediatric AML. To this end, leukemic cell lines were characterized for the expression of WT1 and its mutations. In addition, to better understand the pathogenicity of WT1 in AML, its isotype distribution pattern was investigated in primary AML blasts as well as leukemic cell lines. The characterization revealed specific genetic patterns, which, in comparison with the data of analyzed patient cohorts, can serve as a suitable model system for the in vitro investigation of functional and therapeutic questions. Finally, the suitability and impact of ribonucleic acid (RNA)-based fusion screening using second-generation sequencing was analyzed for pediatric patients diagnosed with AML. It was shown that the method can improve initial diagnostics, particularly through the identification of cryptic and rare fusion transcripts. Furthermore, polymorphic fusion transcripts were identified in validation experiments. A significant impact on response diagnostics was also demonstrated, as a quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) could be designed for rare or highly divergent fusions to monitor the course of therapy. In summary, the results showed the identification of clinical and genetic characteristics of pediatric AML patients, the genetic characterization of leukemic cell lines and the establishment of a method for improved fusion diagnostics in AML.