This task will test the hypothesis that signal rewiring towards pro-inflammatory pathways may modulate HSC fitness in congenital CNP. This hypothesis will be tested by comparing transcriptomes, genomes and proteomes at the level of single HSPCs and their derived colonies. Also chemically defined cytokine- free media will be used to expand bulk HSCs carrying germline mutations with and without acquired somatic variants to provide the cellular substrates for single cell proteomics and genome-wide CRISPR/Cas9 screens. This task will identify functionally important pathways/dependencies that modulate HSC fitness and cancer evolution.

Evidence for a role of (pro-)inflammatory states of HSPCs in the clonal evolution of congenital CN towards malignancy has been obtained and involves activation of different inflammatory gene regulatory networks. Possible candidates igniting these states include the increased levels of reactive oxygen species in CN-HSPCs and elevated expression of transposable elements (TE), in particular so-called LINE elements. By employing already available patient-derived iPSCs, which model the progressive stages of malignant transformation of CN, this task will identify key molecules/pathways involved in these processes. CRISPR/Cas9 screens, small molecule libraries and candidate drug panels will be among the tools used in these studies. The findings will be validated in primary cells from patients and will be assessed for their value as potential biomarkers or targets for therapy.