Association between missense variants of uncertain significance in the CHEK2 gene and hereditary breast cancer: a cosegregation and bioinformatics analysis

Abstract

Inherited mutations in the CHEK2 gene have been associated with an increased lifetime risk of developing breast cancer (BC). We aim to identify in the study population the prevalence of mutations in the CHEK2 gene in diagnosed BC patients, evaluate the phenotypic characteristics of the tumor and family history, and predict the deleteriousness of the variants of uncertain significance (VUS). A genetic study was performed, from May 2016 to April 2020, in 396 patients diagnosed with BC at the University Hospital Lozano Blesa of Zaragoza, Spain. Patients with a genetic variant in the CHEK2 gene were selected for the study. We performed a descriptive analysis of the clinical variables, a bibliographic review of the variants, and a cosegregation study when possible. Moreover, an in-depth bioinformatics analysis of CHEK2 VUS was carried out. We identified nine genetic variants in the CHEK2 gene in 10 patients (two pathogenic variants and seven VUS). This supposes a prevalence of 0.75% and 1.77%, respectively. In all cases, there was a family history of BC in first- and/or second-degree relatives. We carried out a cosegregation study in two families, being positive in one of them. The bioinformatics analyses predicted the pathogenicity of six of the VUS. In conclusion, CHEK2 mutations have been associated with an increased risk for BC. This risk is well-established for foundation variants. However, the risk assessment for other variants is unclear. The incorporation of bioinformatics analysis provided supporting evidence of the pathogenicity of VUS.

Keywords: CHEK2; bioinformatics analysis; breast cancer; cancer genetics; genetic testing.

FIGURE 1

Pedigrees of families 1, 2, and 3.

FIGURE 2

Pedigrees of families 8 and 9.

FIGURE 3

Sequence logo from multiple alignments of CHEK2 homolog sequences. Sequence conservation represented through a sequence logo obtained from the top 500 (best-scored) hits aligned to the CHEK2 sequence (minimum sequence identity of 64.8%). Only those fragments containing the mutation sites associated with the VUS identified in the BC patients are depicted (residues indicated in red font above the position). Sequence numbering is not ordered based on the sequence (UniProtKB O96017, isoform 1) but on the alignment.

FIGURE 4

Chk2 structure, domains, and main features. (A) Chk2 structure (cartoon) with the highest coverage (homodimeric biological unit) released in PDB 3I6U (Grossman et al., 2016). (B) FHA domain (PDB 1GCX (Li et al., 2002), cartoon) with the mutation spots of the VUS highlighted in orange (sticks) and the flexible long loop highlighted in magenta. The simulation box below indicates the way in which this domain has been simulated (monomeric). (C) Kinase domain representation (cartoon) including one monomer and the activation of the T-loop from the second monomer, as solved in PDB 2CN5 (Oliver et al., 2006). The mutation spots of the VUS are highlighted (sticks in orange), as well as the main conserved motifs and residues in the catalytic site and the T-loop. One molecule of substrate ADP (sticks in magenta) and two magnesium ions (green spheres) located at the catalytic site—as per 2CN5—are also depicted. The simulation box below indicates the way in which this domain has been simulated (homodimeric, with Mg²⁺ ions and the ADP substrate).