The ability to diagnose rare genetic diseases through the use of genetic sequencing has improved to a rate of about 40 percent from just about 10 percent through traditional genetic testing.

Despite the improvements, more than half of patients who have symptoms of a rare genetic disease are without a molecular diagnosis.

There are a number of reasons for this, but one that often doesn’t get a lot of attention is that exome sequencing, which is more widely used than whole genome sequencing, looks at less than 2 percent of the entire genome. The problem is that it has limited capacity to detect certain types of variants and focuses on only the parts of the genome that codes for the production of proteins.

In a new study from BC Children’s Hospital, the University of British Columbia, and an international team of researchers published in the New England Journal of Medicine, investigators for the first time identified a DNA mutation underlying an inherited metabolic disorder due to a mutation adjacent to the gene rather than in the gene itself.

The researchers were trying to diagnose three unrelated patients. The children had early-onset delays in both gross and find motor skills and delayed speech. All three patients developed ataxia and became dependent on a wheelchair or walker.

Testing revealed that all three patients had a deficiency of the enzyme glutaminase, which is needed to convert glutamine to glutamate, an essential neurotransmitter. Though the condition is not fully understood, the researchers said it’s likely that either a build-up of glutamine or the lack of glutamate caused the children’s serious developmental delays and disabilities including difficulty with language, speech, balance, and coordination.

But when the researchers zeroed in on the gene that codes for the production of the enzyme glutaminase, it was normal. After further investigations using exome sequencing and whole genome sequencing, the team couldn’t pinpoint the error in the DNA.

Study co-authors Britt Drögemöller and Phillip Richmond discovered and confirmed that the gene responsible for the disorder was intact. They used new bioinformatic tools and a manual approach to discover a repeat expansion error in a part of the genome adjacent to the gene that prevented it from functioning. In the case of these children, the DNA adjacent to the gene  had extended several hundred times its normal length.

“In our search, we focused on variations that would have been hard to discover through exome sequencing” said Drögemöller, UBC postdoctoral fellow at BC Children’s. “After months of experimenting with various different analyses, we finally uncovered this novel genetic variant by using new targeted approaches aimed at identifying DNA repeat expansions.”

Repeat expansion disorders involve pieces of genetic code that repeat over and over. Everyone has parts of their genomes with repeats, but when the repeats become elevated, they can be pathogenic. Certain rare conditions such as myotonic dystrophy and Huntington’s disease are well known examples of repeat expansion disorders. To date, DNA repeat expansions have been linked to approximately 30 different diseases. But this is the first case of a repeat expansion disorder identified in the non-coding portion of the genome.

“To detect this kind of DNA multiplication, you can only use whole genome sequencing and have to search through billions of pieces of DNA. It’s truly a search for the needle in the haystack,” said lead author Clara van Karnebeek, associate professor in the department of pediatrics at UBC and in pediatrics and biochemical genetics at Amsterdam University Medical Centers. “With our new approach we have finally solved our mystery cases, and we now expect to find the genetic cause of other, as of yet unexplained, genetic metabolic diseases.”

The molecular diagnosis now opens the possibility of finding an intervention to address this condition. But it also sheds light on the value of looking at the non-coding portion of the genome and provides some new avenues to pursue for patients still without answers despite having had their genomes sequenced.

Photo: Clara van Karnebeek, associate professor in the department of pediatrics at UBC and in pediatrics and biochemical genetics at Amsterdam University Medical Centers.

X