The tumor suppressor gene TP53 is localized on chromosome 17 in the region 17p13.1 and encodes the p53 protein of the size of 393 aminoacids. It plays a key role in the regulation of many genes affecting proliferation, apoptosis, DNA reparation or angiogenesis. Mutations in this gene lead to disturbing the balance of these processes. The most mutations which inactivate p53 cause a loss of function of the protein to bind to its target DNA sequences and so preventing the transcriptional activation of corresponding genes.
p53 searches damaged areas on the DNA . If such a place is found it triggers the transcription of p21 gene which stops the cell proliferation until the damaged area is repaired.
Germline mutations of this gene are associated with a familial incidence of Li-Fraumeni syndrome characteristic by the autosomal dominant transmitted predisposition to a broad spectrum of tumors, such as soft tissue and bone sarcomas, breast tumors, brain tumors, adrenocortical carcinomas and others.
The importance of the TP53 gene confirms also the fact that it is mutated almost in a half of all human tumors.
Examination
In our laboratory, we perform the mutation analysis of all coding sequences of the TP53 gene including exon - intron joints using PCR amplification and direct sequencing. Material for the examination is a tumor tissue and peripheral blood. It is possible to freeze the tumor tissue in an empty tube till the date of transport or to fix it by loading into a 10 % buffered formalin (our laboratory delivers for free on request). 0.5 – 1 ml of the peripheral blood collected in a tube with EDTA must be stored in a fridge under the temperature of 2 -8 °C till the date of transport (max. 5 days). Transport from the ambulance to the laboratory is provided by the laboratory.
Clinical sensitivity
Germline mutations of the gene TP53 are found in cca 80 % of patients who meet diagnostic criteria for Li-Fraumeni syndrome.
Sequence analysis of the coding exons and exon-intron joints captures about 98 % of TP53 mutations listed in the publicly available version of mutation databases HGMD (Human Gene Mutation The Database).
MLPA (Multiple-Ligation Probe Amplification) analysis of large-scale mutation (amplification / deletion) captures cca 1 % of TP53 gene mutations listed in the publicly available version of HGMD mutation database.
The analytical sensitivity and specificity of sequencing: 99 %.
Limitations:
Mutations deeply in intron and regulatory sequences are not captured. Rare polymorphisms in the annealing site of primers or probes could cause a diagnostic mistake.
Mutations will not be detected, in the case of somatic mutations analysis by sequencing, if the altered cell line will not represent at least 20 %.