Screening of Neonatal UK Dried Blood Spots Using a Duplex SMN1 Screening Assay

Abstract

Spinal muscular atrophy (SMA) is an autosomal inherited neuromuscular genetic disease caused, in 95% of cases, by homozygous deletions involving the SMN1 gene exon 7. It remains the leading cause of death in children under 2 years of age. New treatments have been developed and adopted for use in many countries, including the UK. Success of these treatments depends on early diagnosis and intervention in newborn babies, and many countries have implemented a newborn screening (NBS) or pilot NBS program to detect SMN1 exon 7 deletions on dried blood spots. In the UK, there is no current NBS program for SMA, and no pilot studies have commenced. For consideration of adoption of NBS for a new condition, numerous criteria must be satisfied, including critical assessment of a working methodology. This study uses a commercially available real-time PCR assay to simultaneously detect two different DNA segments (SMN1 exon 7 and control gene RPP30) using DNA extracted from a dried blood spot. This study was carried out in a routine clinical laboratory to determine the specificity, sensitivity, and feasibility of SMA screening in a UK NBS lab setting. Just under 5000 normal DBSs were used alongside 43 known SMA positive DBSs. Study results demonstrate that NBS for SMA using real-time PCR is feasible within the current UK NBS Laboratory infrastructure using the proposed algorithm.

Keywords: NBS; SMA; SMN1; dried blood spot; newborn screening; spinal muscular atrophy.

Conflict of interest statement

F.M. has received honoraria for presentation at meetings and scientific advisory boards of Biogen, Novartis, and Roche. He also acknowledges a grant from Biogen for the SMA REACH UK registry for patients affected by SMA. M.S. has received honoraria for presentation at meetings and scientific advisory boards of Biogen, Novartis, and Roche. Other authors declare no conflicts of interest.

Figures

Figure 1

(a) Amplification plot for SMN1 controls with C1 and NTC duplicates showing no amplification, and duplicates of C2 and C3 showing successful amplification. (b) Amplification plot for RPP30 controls with NTC duplicates showing no amplification, while duplicates of C1, C2, and C3 show successful amplification.

Figure 2

Sample box plotted in a histogram showing the Cq distribution of the healthy samples (red bars) and the SMA patients (each blue circle representing one known positive sample). One SMA-positive sample (SMA-affected sample, in turquoise) was detected as a normal healthy sample. Later, it was found that the sample was actually derived from a carrier and not from an affected individual. The aim of the kit is not to identify SMA carriers, and therefore, the results were aligned with the intended use of the kit.

Figure 3

Simple, suggested algorithm for SMA NBS.