Aims/Context: NGS multigene panels are routinely used to identify germline pathogenic variants in cancer susceptibility genes. In addition, NGS allows the identification of low-level mosaicism events that may not be detectable by conventional Sanger sequencing. We describe two cases of presumed TP53 mosaic variants detected by NGS on blood-derived DNA, and confirmed by ARMS-PCR and Sanger sequencing. Case 1: female, 87 years old, colon cancer at 83 and metachronous breast cancer at 86, no history of familial cancer. Case 2: female, 75 years old, ovarian cancer at 71, local relapse at 74.
Methods: NGS using TruSight® Cancer Sequencing Panel and TruSight® Rapid Capture kit (Illumina) and paired-end sequencing on MiSeq® platform (Illumina). Bioinformatic analysis with MiSeq Reporter, Enrichment, VariantStudio, VEP, Alamut Visual, VarAFT, VarSome and IGV. ARMS-PCR and Sanger sequencing were used to confirm the TP53 variants.
Results and Conclusions: Two cases of presumed TP53 mosaic variants were studied. Case 1: the missense alteration TP53: c.764T>G, p.(Ile255Ser) was detected with a variant allele frequency (VAF) of 26% (39/150 reads). This variant is described in ClinVar as a somatic alteration, classified as likely pathogenic. It is not reported in gnomAD and VarSome software classified it as a variant of uncertain significance. Case 2: missense variant TP53: c.524G>A, p.(Arg175His) detected with a VAF of 15% (10/58 reads). This variant is described as pathogenic in HGMD Professional, LOVD and ClinVar, in association with Li-Fraumeni syndrome. These two cases seem to represent TP53 mosaicism, supported by: i) VAF lower than 30%, ii) detection at the sensitivity limit of Sanger sequencing and iii) confirmation by ARMS-PCR. Confirming this hypothesis by studying tumor and other tissue samples and offspring analysis (underway in both cases), is essential for disease diagnosis, assessing recurrence risk and genetic counseling. The hypothesis of acquired aberrant clonal expansion limited to the hematologic compartment, versus a germline variant should be considered in similar cases, and confirmatory methodologies are mandatory.info:eu-repo/semantics/publishedVersio
