Whole Genome Sequencing Hits the Clinic for Diagnosis in Charcot-Marie-Tooth Neuropathy

Emma Hitt, PhD

March 12, 2010

March 11, 2010 — A whole genome sequencing study has identified novel and clinically relevant genetic variants that appear to be associated with Charcot-Marie-Tooth (CMT) neuropathy.

Their work was published online March 10 in the New England Journal of Medicine.

Richard A. Gibbs, PhD, with the Department of Molecular and Human Genetics at Baylor College of Medicine in Houston, Texas, and colleagues applied “next-generation” sequencing methods to identify the cause of inherited neuropathy in a family diagnosed as having the disorder.

Members of the family had screened negative for common CMT neuropathy, which is associated with mutations in several genes, including PMP22, MPZ, PRX, GDAP1, and EGR2.

“The most important aspect of this study is that it helps demonstrate the clinical applications of whole genome sequencing,” Dr. Gibbs told Medscape Neurology. “This technique is extremely valuable because it provides many insights and does not require that you know exactly what you are looking for at the outset,” he said.

He added, it is important to consider completeness when using this approach. "You don’t want to miss anything.”

Novel CMT Mutation

The family studied consisted of 4 affected and 4 unaffected siblings, as well as a mother and father who were both unaffected.

The 4 affected family members were diagnosed as having CMT type 1 disease on the basis of a physical examination, which showed distal muscle weakness and wasting, highly arched feet, and lack of deep tendon reflexes. Electrophysiological and neurophysiological studies were also performed.

DNA sequencing was conducted with a next-generation sequencing platform (the SOLiD System by Applied Biosystems) and involved ligation-based sequencing and a 2-base encoding method in which 4 fluorescent dyes were used to tag various combinations of dinucleotides.

The researchers examined 40 genes in the family known to be linked to neuropathic conditions, including 3148 single nucleotide polymorphisms (SNPs) containing 54 coding SNPs. Of those, 2 mutations were located in the SH3TC2 gene. One SNP was a missense mutation and the other was a nonsense mutation, with the latter mutation being novel. Each of these mutations was linked to clinically distinct phenotypes in the family studied.

Of note, no copy number variations, several of which are known to be associated with CMT neuropathy, were identified in the family’s analysis.

According to the researchers, whole genome sequencing can rapidly identify alleles that cause disease and can be used to identify all possible DNA changes that are responsible for a disorder.

The researchers noted that the sequencing technology is advancing rapidly, evolving even during the 6-month course of the study. “Sequence read lengths doubled (from 25 bp to 50 bp), the density of samples on the sequencing slide increased, and mapping technology improved,” Dr. Gibbs and colleagues wrote in the discussion. “Overall, the sequence yield increased by a factor of 3, with no appreciable increase in expense.”

They estimated the cost of the study at $50,000.

A "Glimpse of the Future"

In a related editorial, Richard P. Lifton, MD, PhD, chairman of the Department of Genetics at Yale University, New Haven, Connecticut, noted that costs of whole genome sequencing are rapidly decreasing and approaching a “threshold at which DNA sequencing will become a routine part of the diagnostic armamentarium.”

These researchers provide a “glimpse of the future for which we need to prepare,” Dr. Lifton writes.

According to Dr. Lifton, remaining unanswered questions associated with whole genome sequencing include patient selection, how to deal with variants of uncertain clinical significance, and how to convey results of tests without rousing unnecessary anxiety in patients.

This investigator-initiated study was supported by grants from the National Human Genome Research Institute and the National Institute of Neurological Disorders and Stroke. Dr. Gibbs reports grants to his institution from the National Institutes of Health and supply of reagents from Life Technology. Disclosures for the other study authors are available at www.nejm.org. Dr. Lifton has disclosed no relevant financial relationships.

N Engl J Med. Published online March 10, 2010.

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