Background: Lung cancer is a leading cause of cancer-related death worldwide. Somatic DNA alterations, including structural variants (SVs) and DNA methylation, drive tumour development and inform therapy selection. However, detecting SVs is complicated due to intra-tumour heterogeneity which can obscure subclonal mutations with low allele frequencies, and the presence of complex SV events comprised of multiple breakpoints. Complex SVs may arise from genomic instability, DNA damage, and aberrant DNA repair processes and have been linked to events such as localized hypermutation (kataegis) or methylation changes. Hypothesis: We hypothesise that aberrant methylation patterns and kataegis are associated with SV formation and drive genomic instability. This instability may disrupt gene loci, and regulatory elements or induce DNA damage at mutation-prone sites, further contributing to cancer progression.
Methods: To address these complexities, we performed direct DNA long-read whole genome sequencing of lung cancer samples comprising 31 paired tumour and non-tumour samples of different subtypes (including small cell carcinoma, non-small cell carcinoma, squamous cell carcinoma and adenocarcinoma). Sequencing was performed using the latest ONT PromethiON chemistry using direct DNA sequencing allowing simultaneous genome-wide methylation profiling without bisulfite conversion.
Results: Through a comprehensive benchmark of 7 approaches to call somatic SV events we found that long-read sequencing detects more events than short-read sequencing and allows phasing of reads across genomic regions. Using the long-read whole genome data we identified somatic SV events and characterised the methylation and kataegis patterns around SV breakpoints. We found distinct classes of SV events that are associated with methylation patterns. Notably, we detected methylation changes around the SV breakpoints, which may influence gene expression and contribute to lung cancer progression. By integrating kataegis, methylation and SV events, we provide new insights into the development of SVs in lung cancer and their impact on driver genes in tumour development.