The SALSA MLPA probemix P044 NF2 is an in vitro diagnostic (IVD)1
or a research use only (RUO) assay for the detection of deletions or duplications in the NF2
gene in order to confirm a potential cause and clinical diagnosis for Neurofibromatosis type 2 (NF2). This product can also be used for molecular genetic testing of at-risk family members.
This assay is for use with human DNA extracted from peripheral blood. In a research setting this assay can be used on DNA derived from fresh or formalin-fixed paraffin-embedded (FFPE) tumour tissue. Deletions or duplications detected with the P044 NF2 probemix should be verified by another technique. In particular, deletions or duplications detected by only a single probe always require validation by another method. Most defects in the aforementioned gene 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 this gene. 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. NF2 has a high incidence of mosaicism: ~15% of NF2 patients are mosaic. Mosaic mutations may not be detectable in blood or other healthy tissues.
Please note that this probemix is for In Vitro Diagnostic (IVD) use in the countries specified at the end of this product description. In all other countries, the product is for Research Use Only (RUO).
Neurofibromatosis type 2 (NF2) is an autosomal dominant cancer syndrome that is characterized by the development of bilateral vestibular schwannomas (BVSs) in almost all patients. This disease is caused by inactivating mutations of the NF2
tumour-suppressor gene. BVSs result in hearing loss, tinnitus and balance dysfunction. The age of onset is between 18 and 24 years. Patients also suffer from schwannomas of other cranial and peripheral nerves, meningiomas, ependymomas, and rarely, astrocytomas. Although most of these tumours are not malignant, their anatomic location and multiplicity lead to high morbidity and mortality at a low age: the average age of death is 36 years. NF2 occurs in approximately 1 in 25,000-40,000 live births and the estimated prevalence in the general population is 1 in 60,000 without any known ethnic or racial bias. For known pathogenic mutations the penetrance is close to 100% (Asthagiri et al. 2009).
Mutational analysis of the NF2
gene in typical NF2 patients has demonstrated causative mutations in ~70%. The NF2
gene behaves as a typical tumour-suppressor gene, with first hits detectable in both constitutional and tumour specimens and second hits detectable only in tumours. Approximately 50% of NF2
mutation positive patients inherit a germline mutation from an affected parent and the remaining half are sporadic cases due to de novo mutations. Large alterations affecting the NF2
gene account for 15-20% of all known NF2
mutations (Abo-Dalo et al. 2010; Halliday et al. 2017; Kluwe et al. 2005; Smith et al. 2016). Combined with the 70% NF2
mutation detection rate in patients this means that 10-15% of NF2 patients have large deletions or duplications. A high level of mosaicism is observed in NF2, which can complicate mutation detection. More than 30% of the de novo cases are mosaic for NF2
mutations, which may result in subclinical symptoms and/or difficulties with mutation detection, resulting in a false negative diagnosis (Evans et al. 2007).
Mutations in NF2
are also frequently found in the DNA of sporadic schwannomas and meningiomas (Lassaletta et al. 2013; Mohyuddin et al. 2002; Pathmanaban et al. 2017). Both NF2 syndromic tumours and such sporadic tumours have often lost a large part of chromosome 22 resulting in loss of heterozygosity (LOH) of NF2
. These large chromosomal deletions frequently include loss of SMARCB1
, which are also recognized as tumour suppressor genes associated with an NF2-related disorder: schwannomatosis.
More information on NF2 can be found at https://www.ncbi.nlm.nih.gov/books/NBK1201/
The SALSA MLPA Probemix P044-C1 NF2 contains 43 MLPA probes with amplification products between 129 and 472 nucleotides (nt). This includes 21 probes for the NF2
gene; 5 probes on chromosome 22q upstream of NF2
, 4 of which target SMARCB1
; and 4 probes on chromosome 22q downstream of NF2
. In addition, 13 reference probes are included that detect autosomal chromosomal targets that have stable copy numbers in the general population and have relatively stable copy numbers in various cancer types including schwannomas and meningiomas. 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 (see table below). 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