Abstract 261: Long-read sequencing characterization of a patient with bilateral Wilms tumor of unknown etiology

Abstract Wilms tumor is the most common childhood kidney cancer. 15% of patients with Wilms tumor have germline pathogenic variants in genes or regions such as WT1 or the 11p15 region. Variants in these regions can include structural or copy number alterations or alterations in methylation. In the majority of cases of Wilms tumor no known pathogenic variant could be found using the state-of-the-art technologies, including comprehensive approaches such as Illumina whole exome sequencing. One explanation for this is that such technologies have difficulties in detecting structural variants (SVs) in areas associated with repeat or low complexity sequence. In addition, Illumina technology does not immediately support direct methylation detection. Therefore, we hypothesized that analysis of bilateral Wilms tumor of unknown etiology using long-read sequencing could reveal molecular events of potential clinical interest. We performed in-depth genomic analysis on a whole blood DNA sample from a patient with a bilateral Wilms tumor. This patient had no significant family history of cancer, and previously tested negative for Beckwith-Wiedemann syndrome by methylation testing of the 11p15 region; clinical exome sequencing of the patient's germline detected no variants associated with Wilms tumors. We sequenced the genome at 40x depth using PromethION Nanopore sequencing. 29180 SVs (deletions or insertions larger than 30 base pairs) were detected using Sniffles and 26480 were detected with SVIM. Only SVs that were detected by both methods were considered for downstream analysis. Variants were annotated and filtered using a short-read catalog of SVs (gnomAD-SV), a long-read catalog, the Database of Genomic Variants catalog, and by comparison to 11 in-house genomes. We focused on SVs, copy number variants, and methylation events affecting genes previously associated with Wilms tumor. Our long-read sequencing approach detected compound heterozygotes using phased variant calls. A heterozygous missense mutation was identified in haplotype one, while a 300 base pair insertion in an ALU element was present in haplotype two. These two compound heterozygous variants overlap an exon of the OVCH2 gene, and the ALU element was not detected by the prior Illumina analysis. Additionally, we determined the frequency of methylation in CpG sites genomewide using nanopolish. Using a normal blood sample from an unrelated individual as a control, we searched for extreme differences across large and gene promoter regions. Hypermethylation in the promoter regions of genes in the 11p15.5 locus was observed in the patient as compared to the control. Hypomethylation in this region is associated with Beckwith-Wiedemann syndrome. In conclusion, nanopore technology is able to detect variants missed by Illumina sequencing, and has the potential to yield new findings of interest in a case of a child with suspected cancer predisposition syndrome. Citation Format: Allison R. Cheney, Jean Monlong, Holly C. Beale, Hugh Olsen, Ellen Towle Kephart, Katrina Learned, Shanna White, Julian A. Martinez-Agosto, Noah Federman, Mark Akeson, Miten Jain, Vivian Y. Chang, Olena M. Vaske. Long-read sequencing characterization of a patient with bilateral Wilms tumor of unknown etiology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 261.