HiFi sequencing IDs mutations in difficult-to-detect rare diseases
Long-read method detected 93% of mutations in 100 patient samples
PacBio’s high-fidelity (HiFi) sequencing, a new technology for detecting disease-causing DNA mutations in rare diseases, such as AADC deficiency, detected 93% of 145 mutations in 100 samples of patients who had difficult-to-detect rare diseases, a study shows.
While diagnosing these patients previously required multiple tests with traditional methods that read short stretches of DNA, PacBio’s HiFi sequencing, which can read long stretches of DNA with a high level of accuracy, identified nearly all the mutations, or variants, using one test.
“Most importantly, we show the potential to use a single technology to accurately identify all types of clinically relevant variants,” the researchers wrote. The study, “HiFi long-read genomes for difficult-to-detect, clinically relevant variants,” was published in the American Journal of Human Genetics.
“The diagnostic capabilities demonstrated in this study represent a watershed moment for the potential of genomic medicine,” Christian Henry, president and CEO of PacBio, said in a press release.
AADC deficiency is an ultra-rare genetic disorder caused by mutations in the DDC gene, which carries the instructions to make the aromatic L-amino acid decarboxylase (AADC) enzyme, an important protein for producing dopamine and serotonin, two chemical messengers known as neurotransmitters used in nerve cell communication throughout the brain.
DDC mutations lead to too little AADC enzyme being produced and/or one that doesn’t work properly, leaving patients with reduced levels of dopamine and serotonin, as well as other neurotransmitters and resulting in impairments in nerve cell communication. This causes an array of symptoms, including muscle weakness, seizures, and delays in reaching developmental milestones, like walking and talking.
An AADC patient’s diagnostic journey often includes genetic testing where doctors read, or sequence, a DNA sample. This can involve sequencing either short portions of the gene or the entire gene to detect a particular mutation.
Accurate, long-read sequencing
Here, researchers examined DNA from 100 samples of patients with rare diseases caused by genetic mutations located in complex regions of the genome, that is, the entire set of a person’s genetic material. These mutations are difficult to detect with traditional tests that read short stretches of DNA, which is an accurate method for detecting mutations, but generates incomplete results. Long-read sequencing methods provide more information, but are prone to more errors than HiFi, which can read long stretches of DNA with 99.9% accuracy, according to the company.
“Technical limitations associated with short reads challenge their use for detection of disease-associated variation in complex regions of the genome. Long-read sequencing [such as HiFi] technologies may overcome these challenges, potentially qualifying as a first-tier test for all rare diseases,” wrote the researchers, who were able to identify 93% of the disease-causing mutations in their dataset.
Moreover, HiFi technology also enabled mutations that were missed by short-read approaches to be detected. The new technology identified complex mutations in the DNA structure, along with abnormalities in DNA methylation, which is a chemical process that adds a small molecule known as a methyl group to DNA, which can turn genes on and off. DNA methylation is a form of epigenetics, that is, a way for cells to control gene expression.
A subset of the 42 samples with 70 known genetic mutations that were previously tested using short-read sequencing techniques were also tested using HiFi. Short-read sequencing techniques automatically detected 29 of the mutations, but Hifi detected 62, or 89%, using its long-read sequencing techniques.
“Although prospective studies are still needed, our results show that [long-read sequencing] has the potential to be implemented as a first-tier diagnostic workflow for [genetic] testing, potentially replacing all current tests for diagnosing individuals with rare disease,” the researchers wrote.
PacBio is collaborating with RadBound University Medical Center (Radboudumc) in the Netherlands to bring the technology into clinical practice. Recent milestones include processing nearly 1,000 DNA samples from rare disease patients, improving operations for greater throughput in sequencing, and streamlining workflows. Radboudumc also implemented a new technique known as SPRQ chemistry, which is enabled by using a new enzyme and different optimized chemical components that improve the efficiency of the HiFi technique.
“PacBio is honored to partner with Radboudumc and other leading centers by delivering SPRQ chemistry for 5,000 genomes as we work together to study ways to simplify diagnostics, increase accuracy, and improve patient outcomes. These metrics reflect our shared commitment to advancing clinical genomics with scalable, practical solutions,” Henry said.
