Decoding the Germline Genome of Paediatric Cancer: Detecting Novel Variants and Determining Their Role in Relation to Cancer Predisposition

Abstract

Cancer development in children does not typically require accumulated somatic mutations introduced by ageing and environmental mutagens, as seen in adult-onset. Rather, germline variants (present in constitutional DNA) are suggested to increase risk, known as cancer predisposition. Studies estimate 10-15% of children diagnosed with cancer carry a germline pathogenic variant in a cancer predisposition gene (CPG), however those studies primarily focus on detecting small variants within protein-coding regions in CPGs, limiting detection rates and discovery of potential novel mechanisms of childhood cancer predisposition. Hence, we analysed whole genome sequencing of germline DNA from newly diagnosed patients (≤21 years old; n=230 patients) recruited in the PREDICT study (NSW; 2021-2023). Five bioinformatics pipelines were established to detect different types of germline alterations (protein-coding, non-coding, splicing, structural variants (SVs) and fusions) across 1,253 genes, curating both clinically relevant and research variants-of-interest. The pipelines detected small (≤50 bp) and large (>50 bp) variants, including in understudied genomic regions (e.g. promoter and deep intronic regions). Sixty patients (26%) were detected to carry at least 1 clinically relevant variant across protein-coding, splicing and SV pipelines. Novel variants-of-interest were detected in 81 patients (35%) across all pipelines, including multiple predicted variants located in KLF4 promoter region, warranting further investigation. Three RAD54L candidate variants identified in haematological malignancies were stably transduced in Ba/F3 cells, which demonstrated altered cell cycle profile and increased sensitivity to DNA-damaging reagents. This study highlights the significance of interrogating the whole genome to identify novel childhood cancer predisposition mechanisms. Consequently, a personalised medicine approach can be developed to ultimately improve quality of life.