Detection of Spinal Muscular Atrophy Patients Using Dried Saliva Spots

Abstract

Spinal muscular atrophy (SMA) is a lower motor neuron disease, once considered incurable. The main symptoms are muscle weakness and muscular atrophy. More than 90% of cases of SMA are caused by homozygous deletion of survival motor neuron 1 (SMN1). Emerging treatments, such as splicing modulation of SMN2 and SMN gene replacement therapy, have improved the prognoses and motor functions of patients. However, confirmed diagnosis by SMN1 testing is often delayed, suggesting the presence of diagnosis-delayed or undiagnosed cases. To enable patients to access the right treatments, a screening system for SMA is essential. Even so, the current newborn screening system using dried blood spots is still invasive and cumbersome. Here, we developed a completely non-invasive screening system using dried saliva spots (DSS) as an alternative DNA source to detect SMN1 deletion. In this study, 60 DSS (40 SMA patients and 20 controls) were tested. The combination of modified competitive oligonucleotide priming-polymerase chain reaction and melting peak analysis clearly distinguished DSS samples with and without SMN1. In conclusion, these results suggest that our system with DSS is applicable to SMA patient detection in the real world.

A new line method; A direct test in spinal muscular atrophy screening for DBS

“…In the literature, it is seen that methods requiring consecutive two‐step PCR (Shinohara et al, 2019 ) or post‐PCR processes such as melting curve/agarose gel electrophoresis (Takeuchi et al, 2019 ; Wijaya, Nishio, Niba, Okamoto, et al, 2021 ; Wijaya, Nishio, Niba, Shiroshita, et al, 2021 ) are applied in direct PCR studies for SMA newborn screening. These methods have a low potential for application in SMA newborn screening programs due to involving high risk of contamination and operational challenges.…”

The importance of SMA screening tests in newborns – review

“…The isolated DNA is then subjected to PCR analysis, as in the case of using dry blood drop. However, the DSS method has some limitations -the amount of amplifying DNA obtained from liquid saliva is lower than in blood, and PCR inhibitors contained in saliva may distort the results (Wijaya et al -1/61 test was failure out of DSS samples -1.6%; using DBS showed no failure of the test) [11]. The use of the new Intelligent Ratio (IR) method using FII as a reference gene in the analysis by Cavdarli et al allowed the user and the amount of DNA used in the extraction to provide quick and accurate results, completely consistent with the MLPA analysis [12].…”