Deep Molecular Characterization of Milder Spinal Muscular Atrophy Patients Carrying the c.859G>C Variant in SMN2

Abstract

Spinal muscular atrophy (SMA) is a severe neuromuscular disorder caused by biallelic loss or pathogenic variants in the SMN1 gene. Copy number and modifier intragenic variants in SMN2, an almost identical paralog gene of SMN1, are known to influence the amount of complete SMN proteins. Therefore, SMN2 is considered the main phenotypic modifier of SMA, although genotype–phenotype correlation is not absolute. We present eleven unrelated SMA patients with milder phenotypes carrying the c.859G>C-positive modifier variant in SMN2. All were studied by a specific NGS method to allow a deep characterization of the entire SMN region. Analysis of two homozygous cases for the variant allowed us to identify a specific haplotype, Smn2-859C.1, in association with c.859G>C. Two other cases with the c.859G>C variant in their two SMN2 copies showed a second haplotype, Smn2-859C.2, in cis with Smn2-859C.1, assembling a more complex allele. We also identified a previously unreported variant in intron 2a exclusively linked to the Smn2-859C.1 haplotype (c.154-1141G>A), further suggesting that this region has been ancestrally conserved. The deep molecular characterization of SMN2 in our cohort highlights the importance of testing c.859G>C, as well as accurately assessing the SMN2 region in SMA patients to gain insight into the complex genotype–phenotype correlations and improve prognostic outcomes.

Recommendations for Interpreting and Reporting Silent Carrier and Disease-Modifying Variants in SMA Testing Workflows

“…For instance, while 90% of individuals with SMA and two copies of SMN2 exon 7 typically have SMA type 1, individuals with SMA that have two copies of SMN2 exon 7 and the c.859G>C variant typically have SMA type 2 or type 3, with no known cases of SMA type 1 in individuals where this variant is present [ 13 , 14 , 33 ]. A similar effect has been observed in patients with three copies of SMN2 exon 7 and the c.859G>C variant, typically resulting in SMA type 3 [ 12 , 33 ]. The number of SMN2 copies with c.859G>C also correlates with phenotype, with multiple copies leading to milder phenotypes [ 12 ].…”

“…A similar effect has been observed in patients with three copies of SMN2 exon 7 and the c.859G>C variant, typically resulting in SMA type 3 [ 12 , 33 ]. The number of SMN2 copies with c.859G>C also correlates with phenotype, with multiple copies leading to milder phenotypes [ 12 ]. While the c.859G>C variant has not been reported in patients with one or four copies of SMN2 , available evidence suggests that any individual with this variant would have a milder phenotype than expected based on SMN2 copy number alone.…”

“…SMA disease severity inversely correlates with SMN2 copy number, meaning the more copies of SMN2 , the less severe the phenotype [ 5 ]. SMN1 and SMN2 differ in 16 paralogue sequence variants (PSVs) [ 12 ]. One PSV, c.840C>T, disrupts a splice enhancer that decreases the number of exon 7 containing mRNAs to 10–20%, which results in a significantly reduced amount of functional SMN protein compared to that made from a functional SMN1 gene.…”

“…Indeed, 44 SMA patients carrying the c.859G>C variant have been described, all of whom presented a milder phenotype than expected according to their SMN2 copies. This variant has been described in various populations, showing a common haplotype that points towards a common ancestral origin [ 12 ]. Thus, SMN1 is associated with molecular SMA diagnosis and carrier status, whereas SMN2 is associated with the severity of the disease.…”

“…In addition to the SMN1 copy number, data has shown that the presence of SMN1 gene duplication variants c.*3+80T>G in intron 7 and c.*211_*212del in exon 8 ( Figure 1 A) can be indicative of the silent carrier (2 + 0) genotype in many ethnicities [ 10 , 11 ]. Several test methods can detect these variants, including MLPA (P-460), NGS, PCR/Sanger, and PCR/CE [ 12 , 18 , 19 , 24 , 26 , 27 ]. Typically, these variants co-occur [ 10 ]; however, individuals with only one of the two variants have been identified [ 11 ].…”