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. 2019 Jun 3;19(1):535.
doi: 10.1186/s12885-019-5756-4.

Analysis of hereditary cancer syndromes by using a panel of genes: novel and multiple pathogenic mutations

Georgios N Tsaousis  1 , Eirini Papadopoulou  2 , Angela Apessos  2 , Konstantinos Agiannitopoulos  2 , Georgia Pepe  2 , Stavroula Kampouri  2 , Nikolaos Diamantopoulos  3 , Theofanis Floros  4 , Rodoniki Iosifidou  3 , Ourania Katopodi  5 , Anna Koumarianou  6 , Christos Markopoulos  7 , Konstantinos Papazisis  8 , Vasileios Venizelos  9 , Ioannis Xanthakis  10 , Grigorios Xepapadakis  11 , Eugeniu Banu  12 , Dan Tudor Eniu  13 , Serban Negru  14 , Dana Lucia Stanculeanu  15 , Andrei Ungureanu  16 , Vahit Ozmen  17 , Sualp Tansan  18 , Mehmet Tekinel  19 , Suayib Yalcin  20 , George Nasioulas  2
Affiliations

Analysis of hereditary cancer syndromes by using a panel of genes: novel and multiple pathogenic mutations

Georgios N Tsaousis et al. BMC Cancer. .

Abstract

Background: Hereditary cancer predisposition syndromes are responsible for approximately 5-10% of all diagnosed cancer cases. In the past, single-gene analysis of specific high risk genes was used for the determination of the genetic cause of cancer heritability in certain families. The application of Next Generation Sequencing (NGS) technology has facilitated multigene panel analysis and is widely used in clinical practice, for the identification of individuals with cancer predisposing gene variants. The purpose of this study was to investigate the extent and nature of variants in genes implicated in hereditary cancer predisposition in individuals referred for testing in our laboratory.

Methods: In total, 1197 individuals from Greece, Romania and Turkey were referred to our laboratory for genetic testing in the past 4 years. The majority of referrals included individuals with personal of family history of breast and/or ovarian cancer. The analysis of genes involved in hereditary cancer predisposition was performed using a NGS approach. Genomic DNA was enriched for targeted regions of 36 genes and sequencing was carried out using the Illumina NGS technology. The presence of large genomic rearrangements (LGRs) was investigated by computational analysis and Multiplex Ligation-dependent Probe Amplification (MLPA).

Results: A pathogenic variant was identified in 264 of 1197 individuals (22.1%) analyzed while a variant of uncertain significance (VUS) was identified in 34.8% of cases. Clinically significant variants were identified in 29 of the 36 genes analyzed. Concerning the mutation distribution among individuals with positive findings, 43.6% were located in the BRCA1/2 genes whereas 21.6, 19.9, and 15.0% in other high, moderate and low risk genes respectively. Notably, 25 of the 264 positive individuals (9.5%) carried clinically significant variants in two different genes and 6.1% had a LGR.

Conclusions: In our cohort, analysis of all the genes in the panel allowed the identification of 4.3 and 8.1% additional pathogenic variants in other high or moderate/low risk genes, respectively, enabling personalized management decisions for these individuals and supporting the clinical significance of multigene panel analysis in hereditary cancer predisposition.

Keywords: BRCA1 & BRCA2; Breast cancer; Cancer susceptibility genes; Genetic testing; Hereditary cancer; Large genomic rearrangement; Multigene panels; Next generation sequencing; Pathogenic variant.

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Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
Panel testing outcomes and positive results for the 1197 individuals tested grouped by gene and gene category based on risk for any cancer type (Table 1). a. Outcomes of panel testing for the 1197 individuals tested. Positive results refer to the cases where a pathogenic/likely pathogenic variant was identified b. Percentage of pathogenic/likely pathogenic findings identified in each gene c. Pathogenic/likely pathogenic findings stratified by gene risk category for any cancer type
Fig. 2
Fig. 2
mRNA analysis of the c.49-1G > A variant in PALB2 a. Chromatograms of sequencing analysis of genomic DNA of a patient carrying the c.49-1G > A variant in PALB2. b. RT-PCR products on 3% agarose gel. Lanes 1 and 2: the sample of the patient with the variant, Lane 3: normal sample, Lane 4: negative control, Lane 5: 100 bp DNA Ladder (New England Biolabs). c. Chromatograms of sequencing analysis of cDNA from the same patient showing that this splicing variant leads to the in-frame deletion of two amino acid residues, p.Leu17_Lys18 (bottom panel) compared to the sequencing analysis of a wild type sample (top panel)
Fig. 3
Fig. 3
mRNA analysis of the c.1166G > T in BMPR1A a. Chromatograms of sequencing analysis of genomic DNA of a patient carrying the c.1166G > T in BMPR1A. b. RT-PCR products on 3% agarose gel. Lane 1: 100 bp DNA Ladder (New England Biolabs), Lanes 2 and 3: the sample of the patient with the variant, Lane 4: normal sample, Lane 5: negative control. c. Chromatograms of sequencing analysis of cDNA A from the same patient. The c.1166 T variant is not present, indicating instability of the aberrantly spliced transcript (top panel: wild type sample, bottom panel: sample of the patient with the variant)
Fig. 4
Fig. 4
Pedigree of a family with strong breast cancer history
Fig. 5
Fig. 5
Pedigree of the family of a 67-year old CRC patient
Fig. 6
Fig. 6
Apportionment of positive results of genetic testing for the 768 individuals with personal history of Breast cancer using 4 different testing scenarios; that of testing the BRCA1 and BRCA2 genes only and the three scenarios of using gene panels that include other high-risk, moderate-risk and low-risk genes for breast cancer (Table 1). The percentage in each case corresponds to the number of individuals identified with positive findings
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