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In a recent edition of Cell Genomics, a group of European researchers investigated childhood tumors and elucidated some of the somatic genetic and epigenetic landscapes of these tumors. Their work involved using long-read sequencing to explore cancer genomes from a specific type of brain tumor called medulloblastoma.
"[N]anopore sequencing enabled us to discover a complex pattern of DNA rearrangement 'missed' by short-read sequencing and such patterns may prove to be potentially relevant for precision medicine in the future," said Jan Korbel one of the corresponding authors and researcher from The European Molecular Biology Laboratory’s European Bioinformatics Institute.
Using Oxford Nanopore Technologies’ long-read instruments, the team performed whole-genome sequencing to evaluate medulloblastoma samples from an individual with Li-Fraumeni syndrome (OMIM: 151623). This syndrome is exhibited by an increase in cancer susceptibility, most commonly by mutations in the TP53 gene. In addition, they performed genome sequencing on a sample from a post-treatment relapse tumor along with a blood sample from the same individual.
Analysis of the long-read sequences showed methylation and complex structural variant that were further evaluated in other cancer genome data produced with short-read sequencing. While reviewing the methylation patterns, the team noticed that the long-read sequence could better distinguish important features such as allele-specific methylation and methylation shifts located at cancer driver gene promoter sequences or defined complex rearrangement events.
After integrating the long-read data with short reads generated from Illumina instruments, they mapped hundreds of structural variants throughout the primary medulloblastoma tumor. A closer investigation of these structural changes using in situ hybridization allowed them to better characterize the variants, particularly a 1.55-megabase chromothripsis event that included stretches of chromosomes 11 and 17. Chromothripsis is a chromosomal rearrangement characterized by many, potentially thousands, of cluster chromosomal rearrangements in a specific region in the genome.
In addition, the analysis of the long-read sequences also led to the detection of a novel complex rearrangement pattern called templated insertion (TI) threads. The TI threads copy and link large numbers of sub-kilobase-sized TIs in both the forward and reverse orientation, which results in amplified sequences that range from a few kilobases to tens of kilobases in size. Originally the TI threads were not detected in whole genome sequencing data from short-read instruments, but after a close examination, the researchers identified common features linked with TI threads used to detect them in cancer genomes built with shorter reads.
“Our study shows the advantage of long-read sequencing in the discovery and characterization of complex somatic rearrangements,” wrote the study authors. They also added that their work shows the benefits of long reads for “refining complex and repetitive rearrangement patterns such as TI threads and telomere-associated SVs, and of integrating these with ASM and expression changes.”
While the study didn’t have a clinical focus, the researchers wanted to show the practicality of using long-read sequencing to characterize cancer genomes and epigenomes, with the potential for clinical use. "Nanopore sequencing has a very rapid turnaround, and reveals both genetic changes and epigenetic changes in tumors," said Kobel. "Therefore, it is likely to contribute to accelerated diagnosis and clinical decision-making in the future.”
"[N]anopore sequencing enabled us to discover a complex pattern of DNA rearrangement 'missed' by short-read sequencing and such patterns may prove to be potentially relevant for precision medicine in the future," said Jan Korbel one of the corresponding authors and researcher from The European Molecular Biology Laboratory’s European Bioinformatics Institute.
Using Oxford Nanopore Technologies’ long-read instruments, the team performed whole-genome sequencing to evaluate medulloblastoma samples from an individual with Li-Fraumeni syndrome (OMIM: 151623). This syndrome is exhibited by an increase in cancer susceptibility, most commonly by mutations in the TP53 gene. In addition, they performed genome sequencing on a sample from a post-treatment relapse tumor along with a blood sample from the same individual.
Analysis of the long-read sequences showed methylation and complex structural variant that were further evaluated in other cancer genome data produced with short-read sequencing. While reviewing the methylation patterns, the team noticed that the long-read sequence could better distinguish important features such as allele-specific methylation and methylation shifts located at cancer driver gene promoter sequences or defined complex rearrangement events.
After integrating the long-read data with short reads generated from Illumina instruments, they mapped hundreds of structural variants throughout the primary medulloblastoma tumor. A closer investigation of these structural changes using in situ hybridization allowed them to better characterize the variants, particularly a 1.55-megabase chromothripsis event that included stretches of chromosomes 11 and 17. Chromothripsis is a chromosomal rearrangement characterized by many, potentially thousands, of cluster chromosomal rearrangements in a specific region in the genome.
In addition, the analysis of the long-read sequences also led to the detection of a novel complex rearrangement pattern called templated insertion (TI) threads. The TI threads copy and link large numbers of sub-kilobase-sized TIs in both the forward and reverse orientation, which results in amplified sequences that range from a few kilobases to tens of kilobases in size. Originally the TI threads were not detected in whole genome sequencing data from short-read instruments, but after a close examination, the researchers identified common features linked with TI threads used to detect them in cancer genomes built with shorter reads.
“Our study shows the advantage of long-read sequencing in the discovery and characterization of complex somatic rearrangements,” wrote the study authors. They also added that their work shows the benefits of long reads for “refining complex and repetitive rearrangement patterns such as TI threads and telomere-associated SVs, and of integrating these with ASM and expression changes.”
While the study didn’t have a clinical focus, the researchers wanted to show the practicality of using long-read sequencing to characterize cancer genomes and epigenomes, with the potential for clinical use. "Nanopore sequencing has a very rapid turnaround, and reveals both genetic changes and epigenetic changes in tumors," said Kobel. "Therefore, it is likely to contribute to accelerated diagnosis and clinical decision-making in the future.”