Functional characterization of potentially oncogenic somatic mutations of cardiac sarcomas

• Applicant: Professor Dr. Andreas Bräuninger
• Subject Area: Cardiac and Vascular Surgery
• Term: from 2009 to 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 159503538

Cardiac sarcomas, mainly angiosarcomas and sarcomas NOS, have an unfavorable prognosis and the pathogenic mechanisms are largely unknown. Using targeted next generation sequencing and genome-wide SNP array analysis we identified recurrent genomic aberrations in cardiac angiosarcomas. In most cases in addition to receptor tyrosine kinases like KDR and signalling molecules activated by receptor tyrosine kinases also chromatin modifiers were affected by genomic aberrations. For a recurrent PLCG1 mutation we could show that the mutation causes permanent PLCG1 activation with subsequent activation of the transcription factor NFAT and increased apoptosuis resistance. Among the chromatin modifiers MLL2 was most frequently affected by inactivating mutations.Therefore we aim to analyse how the recurrent PLCG1 mutation and MLL2 inactivation contribute to the pathogenesis of cardiac angiosarcomas. Chromatin modifiers like MLL2 as well as PLCG1 via activation of the transcription factor NFAT influence gene expression. Therfore we will perform genome-wide gene expression analysis using RNA next generation sequencing to analyse the effects of PLCG1 activation by ectopic expression of the mutant and MLL2 inactivation by genome editing using the CRISPR-Cas9 system, individually and in combination, in primary endothelial cells. Complementary we will identifiy differentially expressed genes in cardiac angiosarcomas by comparison with primary cardiac endothelial cells. By these approaches we should identify genes aberrantly expressed due to PLCG1 activation and MLL2 inactivation and thereby also identify potential therapeutic targets downstream of PLCG1 and MLL2.In the gene expression analysis also miRNAs will be included. miRNA expression profiles of cardiac angiosarcomas and sarcomas NOS have already been generated. For the identification of differentially expressed miRNAs expression profiles of primary cardiac endothelial cells as reference have to be generated.A further goal is the establishment of a model for cardiac angiosarcomas. We shall establish cell lines with stable expression of the PLCG1 mutation and MLL2 inactivation using genome editing of primary human endothelial cells. With this model inhibitors of the KDR/PLCG1 pathway as well as targetd interventions downstream of PLCG1 and MLL2 like inactivation or inhibition of aberrantly expressed proteins could be evaluated.Finally, we will continue our analysis of genomic aberrations in cardiac sarcomas NOS to identify and functionally analyse, like in cardiac angiosarcomas, potentially pathogenic mutations.

• DFG Programme: Research Grants