SAN FRANCISCO (GenomeWeb) – The Garvan Institute of Medical Research has been investigating how to best implement whole-genome sequencing in clinical care, analyzing various genetic diseases to determine when it makes the most sense as a first-line diagnostic.
The Sydney, Australia-based institution recently published two studies comparing whole-genome sequencing's ability to diagnose two hereditary cardiovascular disorders with more targeted approaches, finding that the whole-genome approach identified all the pathogenic variants called by targeted approaches plus some variants that the panels missed.
The studies "certainly make the case that whole-genome sequencing could be used as a first-line test," said Mark Cowley, head of translational genomics at the Garvan Institute. "It found everything that targeted sequencing found and more."
The next step, Cowley said, is to do a cost-effectiveness study, which would help make the case that testing should be covered by health insurance. Currently, he said, patients primarily pay for genomic testing out of pocket, and in some cases, testing is covered through the hospital's medical department testing budget. For a test to be reimbursed through the Australian health system, the researchers would need to demonstrate evidence that it is "both analytically valid and cost effective," Cowley said. A whole-genome test typically runs around A$5,500 ($4,061), which Cowley said was approximately double a gene panel.
Much of Garvan's clinical and translational research ends up being validated for diagnostic and clinical testing by Genome.One, a startup wholly owned by Garvan. For instance, based on the recent studies, Genome.One has commercialized a whole-genome cardiac test, which uses whole-genome sequencing but focuses the analysis on a set of validated genes. Despite analyzing a subset of genes, Cowley said that whole-genome sequencing is still able to identify more pathogenic variants because there is no capture bias or regions that are unable to be captured. In addition, the whole-genome approach enables the flexibility to easily analyze additional genes.
Genome.One also offers a whole-genome test for polycystic kidney disease, as well as a whole-genome or exome test for unknown genetic conditions and a whole-genome test for healthy individuals who want to know their risk of some diseases. For each of its tests, patients also have the option to obtain secondary findings. In addition, in one of the recent studies, Cowley's team developed a structural variant calling bioinformatics tool, which Genome.One has since validated for clinical purposes to call copy number variants.
The Garvan Institute has been focusing for a while on developing whole-genome sequencing for clinical use. Its whole-genome sequencing pipeline on the Illumina HiSeq X Ten has clinical accreditation and it plans to continue to perform clinical genome sequencing on that instrument. It did, however, also invest in the Illumina NovaSeq platform. One of its major focuses over the last several years has been to sequence patient cohorts of various genetic disorders in collaboration with clinical colleagues. For the two recent studies, it focused on cardiovascular disorders.
Similar to those two studies, Garvan has been comparing whole-genome sequencing with other types of genetic testing, such as gene panels, exome sequencing, and and other conventional means of genetic testing like single-gene testing, for individuals with intellectual disability, epilepsy, movement disorders, and mitochondrial disorders.
Previously, Garvan had said that it was achieving nearly a 90 percent diagnostic rate using whole-genome sequencing for polycystic kidney disease. In a study published in the European Journal of Human Genetics in 2016, its researchers described whole-genome sequencing's advantage over targeted approaches at being able to identify pathogenic variants in the gene PKD1, which cause polycystic kidney disease. The gene has six pseudogenes that are over 95 percent homologous, and targeted sequencing typically struggles with capturing the correct gene. Based on that work, Cowley said, there is now a whole-genome sequencing diagnostic test for that condition.
The research on disease-specific patient cohorts with clinicians has been key to translating whole-genome sequencing from the research realm to clinical care.
"There was an assumption from our clinical colleagues that targeted sequencing would be better because it provides overall higher coverage," Cowley said. "But that wasn't matching our experience," and indeed, when the team compared the two approaches directly, it determined that as expected, whole-genome sequencing resulted in more complete coverage. Importantly, there were several cases where the targeted panel missed a diagnosis even though the pathogenic variant was on the panel because it was just poorly covered. Whole-genome sequencing picked up those cases.
In the study on familial dilated cardiomyopathy, published this month in Genetics in Medicine, the researchers analyzed 42 patients with familial dilated cardiomyopathy using both targeted panels and whole-genome sequencing. The panel test included 67 genes associated with familial DCM. Average sequencing depth was 486-fold, but only around 91 percent of the targeted regions were covered. By contrast, whole-genome sequencing had an average sequencing depth of just 34-fold, but covered 99 percent of the regions assessed by the panel, 98 percent of all exons, and 97 percent of the genome.
The panel identified pathogenic or likely pathogenic variants in 21 out of the 42 individuals, while whole-genome sequencing identified the same 21 variants, plus pathogenic variants in an additional three individuals.
The additional three cases included one for whom the panel test failed to call the variant due to poor coverage, another for whom the variant was located in a region outside the targeted panel, and a third case whose disease was due to a complicated structural variant that couldn't be detected by targeted sequencing.
Those three cases illustrate three important advantages of whole-genome sequencing. "Important bases in some genes are still poorly covered by panel sequencing, even at 400- to 500-fold depth," Cowley said. In addition, he said, whole-genome sequencing enables a more flexible approach for including more genes in the analysis as evidence becomes available. And finally, although structural variants can sometimes be identified through panel testing, often they go undetected. In this case, Cowley said, the structural variant was an overlapping deletion/duplication that did not change the copy number, so it wasn't picked up by the gene panel, but was nonetheless confirmed as pathogenic.
For structural variant detection, the researchers developed an in-house pipeline called ClinSV, which they plan to publish in a peer-reviewed journal. As part of developing that pipeline, Cowley said, they used Pacific Biosciences' long-read sequencing technology to assess its performance on the short-read data from Illumina's HiSeq X Ten instrument.
Whole-genome sequencing also enabled the researchers to generate secondary findings, which could have important implications for patients and their family members. In the study, the researchers found three significant secondary findings from an evaluation of the 57 genes that the American College of Medical Genetics and Genomics recommends testing. Those findings included a variant associated with Fabry disease as well as two cases of MUTYH variants, which are associated with Lynch syndrome and an increased cancer risk.
Cowley said that being able to look for secondary findings is a big advantage of whole-genome sequencing, adding that the option is included for patients who order testing through Genome.One.
In the second study, on hypertrophic cardiomyopathy, which was published this month in the Journal of the American College of Cardiology, the researchers analyzed the genomes of 58 patients, including 46 who had previously undergone either panel or exome testing, with negative results. In nine of those patients, they identified pathogenic variants.
Three had variants in genes not previously analyzed in those patients, one had a variant that had been filtered out but turned out to be pathogenic, and five had pathogenic variants in noncoding regions.
Cowley said that the identification of those noncoding variants was especially interesting. Some were "deep intronic variants that ended up changing the length of the exon, causing it to be spliced either sooner or later than typical, resulting in damage to the protein. We're just starting to come to grips with those types of mutations," he said. And in all five cases, the researchers were able to confirm the mutations and validate that they were indeed pathogenic. In addition, in five out of 12 individuals, or 42 percent, who had no previous genetic testing, whole-genome sequencing identified the pathogenic mutations.
Genome.One has now commercialized a whole-genome cardio test, which analyzes targeted gene sets from whole-genome sequencing data, and will develop additional disease-focused tests based on Garvan's research of disease cohorts, Cowley said.