High-resolution analysis of structural genomic alterations in neuroblastoma by linked-read whole-genome sequencing

• Applicants: Dr. Christoph Bartenhagen; Professor Dr. Matthias Fischer
• Subject Area: Pediatric and Adolescent Medicine ; Bioinformatics and Theoretical Biology ; Hematology, Oncology
• Term: from 2020 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 433152224

Neuroblastoma is a malignant solid tumor of childhood with a broad range of clinical courses. In roughly half of the patients, outcome is excellent with no or limited cytotoxic treatment, due to spontaneous regression of the tumor. By contrast, fatal outcome occurs frequently in the remaining cases, despite intensive multimodal treatment. We recently demonstrated that high-risk neuroblastoma is defined by the presence of telomere maintenance, whereas such mechanisms are absent in spontaneously regressing cases (Peifer et al., Nature 2015; Ackermann et al., Science 2018). In addition, genomic alterations of RAS and p53 pathway genes contribute to malignant transformation in roughly one-third of high-risk cases. It has remained unclear, however, which genes drive tumorigenesis in the remaining two-thirds of these tumors. The paucity of single nucleotide variants in cancer-related genes identified by previous studies along with the frequent occurrence of segmental chromosomal copy number alterations suggest that structural genomic alterations may play a significant role in the pathogenesis of high-risk neuroblastoma. We hypothesize that the relevance of structural alterations for the pathogenesis of high-risk neuroblastoma has been underestimated in the past. Linked-read whole-genome sequencing (WGS) provides a novel and valuable tool to accurately determine haplotypes over megabase regions of the genome, which is a prerequisite for in-depth reconstruction of complex genomic rearrangements. In the proposed study, we are therefore aiming to discover pathogenetically relevant structural alterations in high-risk neuroblastoma by linked-read sequencing of primary tumors, which will be supplemented by information on gene expression profiles, histone marks, and clinical data of the corresponding patients. To this end, we will perform linked-read WGS of 50 primary high-risk neuroblastomas and matched normal controls, as well as relapse samples of 20 of these cases. We will develop and optimize computational analysis pipelines for linked-read WGS data to facilitate reconstruction of complex structural genomic alterations occurring in cancer cells. In addition, we will assess the effect of such alterations on gene expression profiles and histone modifications by RNA sequencing and chromatin immunoprecipitation coupled to sequencing, respectively. We will also integrate results from sequencing data with clinical information of the same patients to determine potential consequences of structural variants on the clinical phenotype of the disease. Finally, associations of structural alterations with clinical variables will be validated in an independent cohort using fluorescence in situ hybridization analysis. Together, we expect that our studies will provide profound novel insights into the genetic etiology of high-risk neuroblastoma, which forms the basis for establishing biomarker-guided personalized treatment strategies in this deadly malignancy.

• DFG Programme: Research Grants
• Co-Investigator: Professor Dr. Martin Peifer