NEW YORK (GenomeWeb) – Exome sequencing may outperform standard-of-care testing as a first-tier diagnostic for infants with a suspected monogenic disorder, according to researchers from the University of Melbourne.
The researchers reported the results of a prospective study, which compared exome sequencing with a standard diagnostic workup, in Genetics in Medicine this week.
The team enrolled 80 infants from newborns up to two years of age in the study, who all likely had a genetic disorder. They sequenced their exomes and concurrently conducted standard diagnostic testing, as ordered by the patients' physicians. Testing included both single-gene Sanger testing as well as next-generation sequencing-based gene panels.
Of the 80 infants, 46, or 57.5 percent, were diagnosed through exome sequencing, compared to 11, or 13.75 percent, who were diagnosed via standard methods.
The study "provides strong evidence for increased diagnostic and clinical utility of singleton [exome] sequencing as a first-tier sequencing test for infants with a suspected monogenic disorder," the authors wrote.
The researchers enrolled a cohort of patients who had well-defined phenotypes and they chose to use singleton exome sequencing as opposed to a gene panel, parent-child trio exome sequencing, or whole-genome sequencing, in order to both reduce the costs as well as to maximize the number of genes tested.
They used the Nextera Rapid Exome Capture Kit and Illumina TruSeq Rapid chemistry on the HiSeq 2500 and sequenced each sample to an average of about 145-fold.
The researchers applied a variety of informatics tools and databases, both internally developed and publicly available ones, and assessed only potential variants that were related to the patient's phenotype.
A multidisciplinary team reviewed the variant classifications, and all pathogenic and likely pathogenic variants were confirmed with Sanger sequencing. When possible, the researchers also tested the parents with Sanger sequencing to determine segregation.
Exome sequencing was able to diagnose 46 out of 80 patients, identifying 48 different genetic conditions. The median turnaround time from sample to receiving a report was 134 days.
The standard-of-care process identified molecular causes in 11 patients. Physicians ordered multiple tests for 21 patients, including Sanger sequencing of single genes, NGS panels, methylation studies, and mitochondrial mutation panels. The clinicians considered an additional 20 tests, but ultimately did not order them due to cost or because certain panels did not have all the genes they wanted to analyze. If those tests had been ordered, an additional 11 diagnoses would likely have been made. So theoretically, a maximum of 22 patients could have been diagnosed using standard of care, although patients would have been subjected to multiple tests.
For the patients diagnosed by the standard methods, the exome sequencing test diagnosed all but one.
The diagnosis changed clinical management for 15 participants, 11 of whom would not have been diagnosed without exome sequencing. Three were able to start a new treatment, one patient stopped a treatment, four patients had their treatment modified, nine had surveillance started for known complications of their conditions, while one patient had surveillance stopped since the exome test clarified a previous erroneous clinical diagnosis.
The researchers found that exome sequencing also had an impact on patients' families. For instance, 12 relatives of infants diagnosed via exome sequencing also received a diagnosis, seven of whom would not have done so under the standard-of-care methods. In addition, 28 couples were identified as being at high risk of having another affected child if they became pregnant. Standard testing methods would only have identified that risk in 13 couples. Exome sequencing also identified one couple as having a risk for two separate disorders, only one of which would have been found through standard testing.
The authors wrote that while currently, most patients receive exome sequencing only after undergoing substantial prior testing, "the decreasing cost and increasing availability of [exome sequencing] present an opportunity to consider the optimal timing of [it] relative to established diagnostic pathways."
The researchers noted that their diagnostic rate of 57.5 percent for exome sequencing was higher than previous studies of exome sequencing, which have had diagnostic rates between 20 percent and 30 percent. Baylor College of Medicine has attempted to improve on this by boosting coverage in some genes and is now testing sequencing some regions with long reads generated by Pacific Biosciences' technology.
The authors wrote that there may be a few reasons why their diagnostic rate was so much higher. They ran the test earlier in the diagnostic process, so weren't just evaluating patients who had already exhausted all other options. In addition, their cohort was well-phenotyped and only included infants, as opposed to a broad spectrum of both pediatric and adult patients.
The authors noted that additional analyses of exome testing should be done, including its cost-effectiveness, reproductive outcomes in families, and the psychosocial impact on families.
Nevertheless, the study provides evidence that exome sequencing as a first-line test for infants can "considerably shorten and simplify the diagnostic process" as well as "provide a higher diagnostic yield" than standard testing, which "maximizes opportunities to improve clinical outcome for patients and families," the authors wrote.