Comprehensive analysis of structural variants in breast cancer genomes using single-molecule sequencing

Abstract

Improved identification of structural variants (SVs) in cancer can lead to more targeted and effective treatment options as well as advance our basic understanding of the disease and its progression. We performed whole-genome sequencing of the SKBR3 breast cancer cell line and patient-derived tumor and normal organoids from two breast cancer patients using Illumina/10x Genomics, Pacific Biosciences (PacBio), and Oxford Nanopore Technologies (ONT) sequencing. We then inferred SVs and large-scale allele-specific copy number variants (CNVs) using an ensemble of methods. Our findings show that long-read sequencing allows for substantially more accurate and sensitive SV detection, with between 90% and 95% of variants supported by each long-read technology also supported by the other. We also report high accuracy for long reads even at relatively low coverage (25×–30×). Furthermore, we integrated SV and CNV data into a unifying karyotype-graph structure to present a more accurate representation of the mutated cancer genomes. We find hundreds of variants within known cancer-related genes detectable only through long-read sequencing. These findings highlight the need for long-read sequencing of cancer genomes for the precise analysis of their genetic instability.

The application of Nanopore sequencing for variant calling on the human mitochondrial DNA

“…The main advantages of MinION discussed in the literature are easiness in handling, rapid development, expected cost reduction of medical examination in the coming years, and the ability to generate reads up to megabases long, which makes them of great potential to be included in clinical practice (Edge and Bansal, 2019). Nanopore reads find extensive implementation in forensic sequencing (Cornelis et al, 2019), characterizing structural variations and somatic mutations in cancer samples (Orsini et al, 2018;Cumbo et al, 2019;Aganezov et al, 2020), typing STEC O157:H7 Shiga toxin-producing Escherichia coli isolates for health monitoring (Greig et al, 2019). The usage of long reads for mitochondrial genome sequencing was reported for vertebrate species identification (Franco-Sierra and Díaz-Nieto, 2020), equine genetics (Dhorne-Pollet et al, 2020), the structural-wise grouping of plan mitochondrial genomes (Masutani et al, 2021), and clinical diagnostics (Wood et al, 2019).…”

“…At the same time, long reads have found application in revealing structural variants (Aganezov et al, 2020), phasing analysis with haplotype reconstruction (Maestri et al, 2020;Popitsch et al, 2020), and the nanoporebased assay for analyzing leukemic samples (Orsini et al, 2018;Cumbo et al, 2019). Nevertheless, reports on long reads' usage with regard to mitochondrial genetics are still scarce.…”

PRINCESS: comprehensive detection of haplotype resolved SNVs, SVs, and methylation

“…Current instruments allow the assessment of human genomes at unprecedented accuracy [ 4 , 5 ] and scale [ 1 , 6 , 7 ]. LRS enables further investigation into many biological questions such as assessment of highly repetitive regions (e.g., SMN1,2 ) [ 8 ], resolving complex regions (e.g., MHC, KIRR ) [ 9 , 10 ], and improving our understanding of structural variants (SVs) [ 1 , 7 , 11 ]. More recent papers show that LRS enable a more detailed characterization of SVs especially over insertions [ 12 , 13 ].…”

“…Thus, LRS established its utility as one of the main components for genomic sequencing [ 15 , 16 ]. Given these advancements, we see novel insights in human diseases [ 7 , 17 , 18 ], evolution [ 6 , 14 , 19 ], and other areas of biology and medical research [ 11 ].…”

Evaluation of Single-Molecule Sequencing Technologies for Structural Variant Detection in Two Swedish Human Genomes

“…All human genomes are different, and to a large extent the genetic variation between individuals can be explained as insertions, deletions, duplications, or translocations [1,2]. Taken together, such structural variation (SV) events affect a large portion of the genomic sequence when comparing any given individual to the existing reference genome, and although our knowledge about SVs is still incomplete, they have been shown to contribute to a number of human diseases and conditions [3][4][5][6][7]. In recent years, SV analysis has become a key endeavour for genomics research [8][9][10][11][12] and these efforts will likely lead to an increased understanding of disease etiology and progression and possibly also improved clinical therapy.…”