WYMM Tour: Toronto | Oncology + Rare Disease

Despite advances in sequencing technologies, a significant portion of the human genome - and biologically relevant variation - remains inaccessible using traditional short-read approaches. Oxford Nanopore sequencing addresses these gaps by enabling comprehensive, real-time analysis of DNA and RNA from a single platform. By directly sequencing native molecules without amplification, nanopore technology captures single nucleotide variants, structural variants, repeat expansions, and epigenetic modifications simultaneously, providing a more complete genomic picture.

We will explore how long-read whole genome sequencing enables more comprehensive characterization of genomic variation, including complex regions often missed by standard methods. Additionally, we will showcase adaptive sampling as a powerful, software-driven enrichment approach for targeted sequencing without additional wet-lab steps, with applications such as immune-genomic profiling in transplantation research.

Through real-world workflows, including rapid whole genome sequencing and multiomic applications, this talk demonstrates how nanopore sequencing delivers scalable, end-to-end solutions. Ultimately, we emphasize that “what you’re missing matters,” and how more complete genomic insight can support advances in precision medicine and genomic research.

The NeuRo Genomics Initiative (NRGI), a project within the Canadian Precision Health Initiative (CPHI), aims to advance our understanding of rare and common neurological diseases by identifying genetic factors contributing to their onset, progression and response to medication. As part of NRGI, we investigated a cohort of patients affected with cerebellar ataxia-neuropathy-vestibular areflexia syndrome (CANVAS) caused by a biallelic repeat expansions in an intron of the RFC1 gene encoding the Replication Factor C subunit 1. Whereas the reference allele contains eleven repeats of the pentanucleotide AAAAG11, the pathogenic expansion usually comprises hundreds of AAGGG repeats. Additional pathogenic conformations and biallelic combinations have now been confirmed in several studies. Moreover, several studies are expanding the phenotypic spectrum associated with RFC1 intronic repeat expansion, highlighting a heterogeneity that goes beyond the repeat size. We performed whole-genome and transcriptome long-read sequencing to characterize the repeat expansion (size and sequence) in our cohort and investigate expression and/or splicing changes. This project consolidate the potential of LRS in genetics and functional studies. Regarding CANVAS, results should provide new insights into pathogenic mechanisms, offering the potential of identifying molecular signatures associated to disease penetrance. Such findings not only support more personalized patient care but also assist the search for new therapeutic targets.

Timely and comprehensive molecular classification is essential for therapeutic decision-making in pediatric oncology, yet current diagnostic workflows rely on multi-step, time- and resource-intensive testing. We present a standardized Oxford Nanopore Technologies’ Whole-Genome Sequencing approach using Adaptive Sampling (AS-WGS), optimized for pediatric cancers, that enables rapid and comprehensive molecular profiling in a single assay.

AS-WGS was applied to 63 pediatric cancer samples (42 leukemias, 21 solid tumors) and compared with extensive clinical testing including cytogenetics, exome, and RNA sequencing. AS-WGS achieved somatic-grade coverage (mean 167X) across 380 clinically relevant loci while maintaining sufficient pan-genomic coverage (18X) for genome-wide analyses. AS-WGS reliably detected all classes of clinically actionable alterations—including copy number changes, gene fusions, and single-nucleotide variants—capturing subclonal variants down to 5% allelic frequency, comparable to clinical sequencing. Additionally, four methylation classifiers (Malin, Alma, CrossNN and Sturgeon) were successfully applied, providing cancer subtype confirmation and resolution of complex cases.

Most clonal alterations were confidently identified within 24 hours of sequencing, with some, such as methylation-based classification, detected within the first hour, enabling same-day insights. We developed an open-source bioinformatic pipeline, nf-core-oncoseq, to enable on-the-fly analysis and data interpretation during sequencing with automated reporting.

Together, this integrated platform consolidates complex molecular diagnostics into a single, rapid assay, establishing AS-WGS as a transformative approach for pediatric cancer diagnostics with the potential to accelerate molecular classification and expand timely access to precision therapies.

RapidOmics 2.0 is a clinical diagnostic research project in British Columbia that seeks to improve care for acutely ill children at BC Children’s Hospital, and for pregnancies at high-risk for rare genetic disorders at BC Women’s Hospital. Co-funded by Genome BC, Genome Canada and the Provincial Health Services Authority, RapidOmics 2.0 is recruiting 100 parent-child trios who require access to rapid and thorough genetic diagnosis. In BC, the current standard of care for urgent genome-wide sequencing is trio short-read exome sequencing sent out-of-province. We are comparing the current standard of care head-to-head with rapid trio ONT long-read whole genome sequencing done at the Michael Smith Genome Sciences Centre at BC Cancer (“the GSC”). One hundred trios within this study should provide statistical power that is more than sufficient to quantify a difference between the trio ES and trio long-read GS, based on a one-year follow-up period.

Notably, we are not just comparing two sequencing technologies: We are comparing two different clinical approaches to urgent genome-wide sequencing. Our current standard is primary evaluation by the patient’s treating specialist in consultation with a clinical geneticist, sending the samples to a clinical lab in the US where the trio exome sequencing and analysis are done, and then sharing the results with the family. Regardless of what is found, the patient and family continue to be followed by the treating medical specialist - if a genetic disease is diagnosed, the family is usually seen again by the geneticist and a genetic counsellor. If the result is nondiagnostic of a genetic disease, genetic follow-up is more variable, and the exome data are not routinely re-analysed.

In contrast, our clinical research pathway includes rapid ONT LRWGS at the GSC, clinical-molecular interpretation of the genomic data by a multidisciplinary team, continuing research analysis using advanced bioinformatics methods, and detailed clinica.