New Genetic Analysis Method Aids SMA ‘Silent Carrier’ Detection

Marisa Wexler MS avatar

by Marisa Wexler MS |

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An illustration shows a DNA strand.

A new computational tool developed by Pacific Biosciences can identify genetic mutations that cause spinal muscular atrophy (SMA) with high accuracy and could help identify “silent carriers” who aren’t detected by current tests, a new study reports.

The study, “Comprehensive SMN1 and SMN2 profiling for spinal muscular atrophy analysis using long-read PacBio HiFi sequencing,” was published in the American Journal of Human Genetics.

SMA is caused by mutations in the gene SMN1, which provides instructions for making the protein SMN. A second gene called SMN2 has a virtually identical sequence to SMN1, but due to a change in a single nucleotide (one of the building blocks in DNA), it produces much less SMN protein — about 10% of the total.

Both SMN1 and SMN2 are on chromosome 5, and most people inherit two copies of each, one from each parent. However, a person can inherit additional copies of both genes. Extra copies of SMN2 are generally linked to less severe disease, as it can partly compensate for mutated SMN1.

But extra copies of SMN1 can complicate SMA genetic testing because of so-called “silent carriers” — people with two healthy copies of SMN1 on one copy of chromosome 5, but no working versions of the gene on the chromosome’s other copy.

Technically, these people produce as much SMN protein as a healthy person without any SMN1 mutations. But because the copy of chromosome 5 without any functional SMN1 can be passed to their children, they’re considered carriers for SMA.

Current genetic tests won’t accurately identify these silent carriers because they’ll only show two healthy copies of SMN1, without differentiating chromosome location. Silent carriers are particularly common among Africans, accounting for more than a quarter of African SMA carriers.

The only way to identify silent carriers is through pedigree analysis, looking at patterns in family trees. A particular genetic variation called g.27134T>G has been proposed as a marker of two SMN1 copies on the same chromosome, but it’s not very sensitive because it’s also often present in people with one SMN1 copy on each chromosome.

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Seeking out ‘silent carriers’ of SMA

To better identify silent carriers, a research team led by scientists at Pacific Biosciences developed a computational tool called Paraphase to analyze SMN1 and SMN2 in greater detail using genetic sequencing data. In these datasets, a person’s genetic code is present in small sections known as “reads.”

The algorithm aligns reads to the sequence of either SMN1 or SMN2, with specific considerations to tell the difference between these two genes and for identifying SMA-causing mutations in SMN1. The program also predicts the number of copies of these genes based on calculations of the number of reads that align to each sequence.

The researchers analyzed genetic data from 438 people across five ethnic groups.  Most was from people of European descent, but there was also data from admixed American, African, and South and East Asian groups.

Paraphase could identify with more than 99% accuracy for each gene the copy number of SMN1 and SMN2. Plus, all the SMA patients and carriers were correctly identified in the algorithm, with no false-negatives or false-positives.

Combining haplotype, pedigree data

Because it determined sequences of the SMN1 and SMN2 genes, the algorithm could also identify their haplotypes, a set of closely linked DNA regions that tend to be inherited together.

The haplotype data were combined with pedigree data to look for markers of two SMN1 copies on the same chromosome, with a particular focus on African populations, where this variation is most common.

Results showed that a particular combination of haplotypes, dubbed S1-8+S1-9d, was found on two-thirds (21 of 31) of chromosomes with two SMN1 copies among African patients. These haplotypes were rarely present when there was only one SMN1 copy.

“We identified a common two-copy SMN1 allele that comprises 67.7% of two-copy SMN1 alleles in Africans,” the researchers wrote.

Taking into account the known prevalence of chromosomes without any functional SMN1, this means the likelihood an African carrying the S1-8+S1-9d haplotype is a silent carrier is about 88%.

The haplotype markedly outperformed the g.27134T>G marker at identifying double SMN1 copies, as those with this marker have only about a 3% risk of being silent carriers.

“Based on our limited sample of 87 African alleles, we estimate that testing positive for these two haplotypes in an individual with two copies of SMN1 gives a silent carrier risk of 88.5%, which is significantly higher than the previously found marker [variant] g.27134T>G,” the researchers said, noting their study is limited by the relatively small number of samples, particularly for non-European populations. They said more research is needed to validate and expand these results.

“For years, researchers have known that SMN1 and SMN2 play roles in the onset and severity of SMA but have been unable to answer exactly why and how people develop more or less severe forms of the disease,” Pacific Biosciences wrote in a company blog post. “With Paraphase, there is now the opportunity to answer some of those long-standing questions and enable more accurate carrier screening.”

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