Intended use: The SALSA MLPA probemix P165 HSP mix-1 is an in vitro diagnostic (IVD)¹ or a research use only (RUO) assay for the detection of deletions or duplications in the human ATL1 and SPAST genes, in order to confirm a potential cause and clinical diagnosis for spastic paraplegia (SPG) type 3A and 4, respectively. This assay is for use with human DNA extracted from peripheral blood. This product can also be used for molecular genetic testing of at-risk family members.

Deletions or duplications obtained with the P165 HSP mix-1 probemix must be confirmed by another technique. In particular, deletions or duplications detected by only a single probe always require conformation by another method. Most defects in ATL1 and SPAST genes are point mutations, none of which will be detected by MLPA. It is therefore recommended to use this SALSA MLPA probemix in combination with sequence analysis of the aforementioned genes. This assay is not intended to be used as a standalone assay for clinical decisions. The results of this test must be interpreted by a clinical molecular geneticist or equivalent.

¹ Please note that this probemix is for In Vitro Diagnostic use (IVD) in the countries specified at the end of this product description. In all other countries, the product is for Research Use Only (RUO).

Clinical background: Hereditary spastic paraplegias (HSP) are genetically heterogeneous neurodegenerative disorders characterised by progressive spasticity and weakness of the lower limbs due to
axonal degeneration in the pyramidal tract. To date, more than 80 genetic types of HSP have been defined
by genetic linkage analysis and identification of HSP-related gene variants.

Spastic paraplegia type 3A (SPG3A) is caused by pathogenic mutations in the ATL1 gene and accounts for approximately 10% of all autosomal dominant HSP. Currently, there are more than 60 different ATL1 mutations described for SPG3A patients, which are divided into five broad groups: 91.5% missense mutations, 4% small insertions, 2.8% small deletions, 0.7% splice site mutation, and one (0.7%) whole exon deletion (exon 4) (Sulek et al. 2013).

Mutations in the SPAST gene are responsible for both autosomal dominant HSP (40-50%) and sporadic cases (10-15%), causing spastic paraplegia type 4 (SPG4). The most common type of SPAST mutations are point mutations (75-80%), and the frequency of large rearrangements in SPAST, such as exon deletions, has been estimated at 20% (Beetz et al. 2006, Depienne et al. 2006).
More information on HSP is available on https://www.ncbi.nlm.nih.gov/books/NBK1509/, https://www.ncbi.nlm.nih.gov/books/NBK1160/, https://www.ncbi.nlm.nih.gov/books/NBK45978/.

Probemix content: The SALSA MLPA Probemix P165-C3 contains 47 probes with amplification products between 130 nt and 481 nt. This includes 16 probes for the ATL1 gene and 20 probes for the SPAST gene. In addition, 11 reference probes are included that detect autosomal chromosomal locations. Complete probe sequences and the identity of the genes detected by the reference probes are available online (www.mlpa.com).

This probemix contains nine quality control fragments generating amplification products between 64 and 105 nt: four DNA Quantity fragments (Q-fragments), two DNA Denaturation fragments (D-fragments), one Benchmark fragment, and one chromosome X and one chromosome Y-specific fragment. More information on how to interpret observations on these control fragments can be found in the MLPA General Protocol and online at www.mlpa.com.