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Open AccessArticle

Male Breast Cancer: Results of the Application of Multigene Panel Testing to an Italian Cohort of Patients

1
Biosciences Laboratory, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, 47014 Meldola, Italy
2
Biostatistics and Clinical Trials Unit, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, 47014 Meldola, Italy
3
Romagna Cancer Registry, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, 47014 Meldola, Italy
4
Department of Medical Oncology, Ospedale Infermi, 47923 Rimini, Italy
5
Department of Medical Oncology, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, 47014 Meldola, Italy
*
Authors to whom correspondence should be addressed.
Diagnostics 2020, 10(5), 269; https://doi.org/10.3390/diagnostics10050269
Received: 31 March 2020 / Revised: 21 April 2020 / Accepted: 28 April 2020 / Published: 30 April 2020
(This article belongs to the Section Pathology and Molecular Diagnostics)

Abstract

Male breast cancer (MBC) is a rare tumor, accounting for less than 1% of all breast cancers. In MBC, genetic predisposition plays an important role; however, only a few studies have investigated in depth the role of genes other than BRCA1 and BRCA2. We performed a Next-Generation Sequencing (NGS) analysis with a panel of 94 cancer predisposition genes on germline DNA from an Italian case series of 70 patients with MBC. Moreover, we searched for large deletions/duplications of BRCA1/2 genes through the Multiplex Ligation-dependent Probe Amplification (MLPA) technique. Through the combination of NGS and MLPA, we identified three pathogenic variants in the BRCA1 gene and six in the BRCA2 gene. Besides these alterations, we found six additional pathogenic/likely-pathogenic variants in PALB2, CHEK2, ATM, RAD51C, BAP1 and EGFR genes. From our study, BRCA1 and BRCA2 emerge as the main genes associated with MBC risk, but also other genes seem to be associated with the disease. Indeed, some of these genes have already been implicated in female breast cancer predisposition, but others are known to be involved in other types of cancer. Consequently, our results suggest that novel genes could be involved in MBC susceptibility, shedding new light on their role in cancer development.
Keywords: male breast cancer; next-generation sequencing; cancer predisposition; BRCA1/2 genes; hereditary cancer; multigene panel testing; multiplex ligation-dependent probe amplification male breast cancer; next-generation sequencing; cancer predisposition; BRCA1/2 genes; hereditary cancer; multigene panel testing; multiplex ligation-dependent probe amplification

1. Introduction

Breast cancer (BC) is the first tumor for incidence and mortality in women [1], but it can also affect men. Indeed, male breast cancer (MBC) represents less than 1% of all BCs [2]. However, being a rare and poorly known disease, it is often diagnosed at later stages with the consequence of a poor prognosis [3].
Similarly to female breast cancer (FBC), genetic predisposition is an important risk factor in MBC [4,5]. BRCA1 [6] and BRCA2 [7] are the genes best known for their involvement in BC predisposition in both females and males [8], with an MBC risk that has been estimated of 1.2% and 6.8% by the age of 70 for carriers of variants in BRCA1 and BRCA2, respectively [9].
In recent years, thanks to the wide use of Next-Generation Sequencing (NGS), the number of genes suspected to be involved in cancer predisposition has dramatically increased [10]. This is true especially for cancers with a strong hereditary component such as FBC, in which several studies have investigated the role of genes other than BRCA1/2 [11,12,13,14,15,16,17,18,19,20]. Some studies have highlighted the role of genes such as PALB2, ATM, CHEK2, FANCM, PTEN, APC and MUTYH in MBC [21,22,23,24,25,26,27,28], thus excluding others, such as BRIP1 and RAD51C [29,30,31].
Within this complex scenario, the aim of our study was the application of multigene panel testing (MGPT) in a series of patients with MBC, in order to assess the presence of BRCA1/2 variants and identify new genes involved in the predisposition to MBC.

2. Materials and Methods

2.1. Ethics Statement

The study was conducted in accordance with ethical standards, the Declaration of Helsinki and national and international guidelines, and was approved by our local ethics committee (CE IRST IRCCS-AVR, protocol 2207/2012). All the patients enrolled in the study have signed an informed consent for the genetic analyses and for the use of the results for research purposes.

2.2. Patients’ Selection

The patients have been selected through an appropriate oncologic genetic counseling according to the Hereditary Breast Cancer Protocol of the Emilia-Romagna region [32]. During the genetic counseling, the personal and family history of cancer was collected and verified by our medical geneticists. According to the FONCaM guidelines used in the protocol [33], a diagnosis of MBC at any age is sufficient to proceed with the BRCA1/2 genetic test, regardless of family history of breast and ovarian cancers (BC/OC). Through this procedure, we selected 70 Italian patients with a diagnosis of MBC.

2.3. Sample Collection and DNA Extraction

Peripheral blood samples were collected from the 70 MBC patients and stored at −80 °C until the genetic analyses. Genomic DNA was extracted from blood using QIAamp DNA Mini Kit (Qiagen, Hilden, Germany) and quantified using Qubit fluorometer and Qubit dsDNA BR Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA).

2.4. Next-Generation Sequencing (NGS)

We used 50 ng of DNA from each sample to create libraries following the Trusight Rapid Capture protocol (Illumina, San Diego, CA, USA). Libraries were enriched for the regions of interest with the Trusight Cancer panel (Illumina), which contains probes targeting the coding regions of 94 genes involved in hereditary cancer (Table 1).
Libraries were sequenced on the MiSeq platform (Illumina) with MiSeq Reagent Kit v2 configured 2 × 150 cycles, according to the manufacturer’s instructions.
All deleterious variants (classes 4–5) identified in BRCA1/2 genes were confirmed through Sanger sequencing following a custom protocol previously described [15]. The same procedure was performed for the BRCA1/2 coding regions with a coverage < 50×. The deleterious variants identified in other genes were confirmed through a second NGS analysis with the same kit and protocol.

2.5. Bioinformatics Analysis

The bioinformatics analysis of NGS results was performed with a customized pipeline described in our previous studies [15,34]. The fastq files have been aligned with BWA software [35] to the reference genome, version UCSC-Build37/hg19. Reads were remapped against the Trusight Cancer Panel reference, removed from duplicates and then realigned for germline indels using GATK indelRealigner. Finally, variants were called using Unified Genotyper of GATK software, version 3.2.2 [36]. Genomic and functional annotations of detected variants were made by Annovar [37].

2.6. Multiplex Ligation-Dependent Probe Amplification (MLPA)

The presence of large deletions/duplications of BRCA1/2 genes, not detectable with the Trusight protocol, was assessed by Multiplex Ligation-dependent Probe Amplification (MLPA) technique with the P002-BRCA1 and P045-BRCA2/CHEK2 kits (MRC Holland, Amsterdam, The Netherlands). The large BRCA1 deletion identified by MLPA was confirmed through the P087-BRCA1 confirmation kit (MRC Holland). Coffalyser software (MRC Holland) was used for the quantitative analysis of the electropherograms.

2.7. Variant Classification

All the genetic variants identified were divided into five classes according to the International Agency for Research on Cancer (IARC) [38]: Benign Variant (BV-class 1), Likely-Benign Variant (LBV-class 2), Variant of Uncertain Significance (VUS-class 3), Likely-Pathogenic Variant (LPV-class 4) and Pathogenic Variant (PV-class 5). The process of variant classification was performed in accordance with the guidelines of the American College of Medical Genetics (ACMG) [39].
In particular, for BRCA1/2 variants, we used different BRCA-specific databases [40,41,42,43,44,45]. The variants of other genes were classified using comprehensive variant databases [46,47,48].

3. Results

3.1. Patient Characteristics

All 70 patients selected for the study had MBC between 36 and 87 years of age, with an average age at cancer onset of 63.8 years. Among the patients, 2/70 (2.9%) had a second contralateral MBC and 16/70 (22.9%) had a second non-BC malignancy.
Regarding the histopathological classification [49], 56/70 MBCs (80.0%) were infiltrating ductal carcinomas (IDC-8500/3), 9/70 (12.9%) were ductal carcinomas in situ (DCIS-8500/2), 3/70 (4.3%) were infiltrating papillary carcinomas (IPC-8503/3) and 2/70 (2.9%) were papillary carcinomas in situ (PCIS-8503/2).
As far as it concerns the family history of cancer, 24/70 patients (34.3%) had I- and/or II-degree relatives with BC/OC and 17/70 patients (24.3%) had I- and/or II-degree relatives with other cancers. In particular, 3/70 patients (4.3%) had a family history of MBC among I-degree relatives.

3.2. Pathogenic and Likely-Pathogenic Variants in BRCA1/2 Genes

Since BRCA1 and BRCA2 variants are the most common alterations in BC patients, we initially focused on these genes.
The NGS analysis allowed us to detect eight pathogenic variants (PVs) in BRCA1/2 genes. In particular, we identified the c.4964_4982del and the c.5266dupC variants in BRCA1 gene, both already reported [46] and classified as pathogenic [47]. In BRCA2 gene, we identified five PVs in six patients, c.1238delT, c.1813delA, c.3195_3198delTAAT, c.5073dupA in single patients and c.6039delA in two unrelated patients. In addition, these variants were already reported [46] and classified as pathogenic [47].
The MLPA analysis revealed an additional PV in BRCA1 gene, a large deletion covering exons 1 and 2. However, the MLPA technique does not give information about the exact location of the breakpoints and, consequently, the variant has been classified as c.-113-?_80+?del. This variant is not reported in the databases, but the deletion of the transcription start site and the first two exons certainly has a deleterious effect on the gene transcription. Moreover, large deletions involving these two exons have been previously described as pathogenic in BC/OC patients [50,51,52,53,54,55,56,57].
Overall, we identified 9/70 patients (12.9%) with BRCA1/2 PVs (Figure 1). All the BRCA1/2 PVs identified in our cohort are reported in Figure 2 and Table 2 with the corresponding patient characteristics. In particular, 8/9 patients had an IDC and 1/9 had a DCIS. The average age at MBC onset was 62.5 years. Regarding the family history of cancer, 5/9 patients (55.6%) had I- and/or II-degree relatives with BC/OC and 3/9 patients (33.3%) had I- and/or II-degree relatives with other cancers.

3.3. Pathogenic and Likely-Pathogenic Variants in other Genes

After the assessment of the BRCA1/2 mutation status of our patients, we searched for PVs and likely-pathogenic variants (LPVs) in the other 92 genes of the panel. We detected six additional PVs/LPVs of six different genes in 6/70 patients (8.6%) (Figure 1). In particular, we identified the c.8319_8323dupTGTCC variant in ATM gene, the c.1110_1116delCATGCAG variant in BAP1 gene, the c.1100delC variant in CHEK2 gene, the c.3538_3541delGAAG in the EGFR gene, the c.73A>T variant in PALB2 gene and the c.181_182delCT in RAD51C gene. The variants in ATM, CHEK2, PALB2 and RAD51C genes were already reported [46] and classified as pathogenic [47]. On the opposite, the variant in EGFR gene was reported only in dbSNP without classification [46] and the variant in BAP1 gene was novel. According to the guidelines [39], we classified these two alterations as LPVs.
All the PVs/LPVs identified in genes other than BRCA1/2 are reported in Table 3 with the corresponding patient characteristics. In particular, 5/6 patients had an IDC and 1/6 had a DCIS. The average age at MBC onset was 55.8 years. Regarding the family history of cancer, 2/6 patients (33.3%) had I- and/or II-degree relatives with BC/OC and 2/6 patients (33.3%) had I- and/or II-degree relatives with other cancers.

3.4. Patients without Pathogenic and Likely-Pathogenic Variants

In the remainder of our cohort, i.e., 55 patients (78.6%), we did not find any PV/LPV neither in BRCA1/2 genes nor in the other 92 genes of the panel (Figure 1).
All these patients had MBC with an average age at cancer onset of 64.9 years. In particular, 43/55 patients had an IDC, 7/55 had a DCIS, 3/55 had an IPC and 2/55 had a PCIS. Regarding the family history of cancer, 17/55 patients (30.9%) had I- and/or II-degree relatives with BC/OC and 12/55 patients (21.8%) had I- and/or II-degree relatives with other cancers.

4. Discussion

MBC is a rare disease but deserves attention in terms of diagnosis and treatment as clinicians tend to follow the guidelines that have been developed for FBC management [58]. In this scenario, the research on the risk factors associated with MBC, in particular the genetic predisposition, is a key element in the prevention and early diagnosis of the disease.
In the present study, we performed an NGS analysis on an Italian cohort of 70 patients with MBC with a panel of 94 genes. At the same time, we conducted an MLPA analysis on BRCA1/2 genes in order to identify also variants that are not easily detectable by NGS.
Among 70 cases of MBC, we detected BRCA1/2 PVs in nine patients. BRCA1 and BRCA2 genes are known since a long time for their involvement in high risk of BC, also in male variant carriers. In particular, germline PVs/LPVs in the BRCA1 gene are associated with a 57%–65% and 1.2% risk of developing BC in females and males, respectively, by the age of 70 [9,59,60,61]. In addition, BRCA1 alterations have been associated with an increased risk of colon cancer [62], prostate cancer [63] and pancreatic cancer [64,65]. On the contrary, germline PVs/LPVs in the BRCA2 gene are associated with a 45%–55% and 6.8% risk of developing BC for females and males, respectively, by the age of 70 [9,59,60,61]. In addition, BRCA2 alterations have been associated with an increased risk of prostate cancer [66], pancreatic cancer [65,67], and uveal melanoma [68,69]. For these reasons, men with BRCA1/2 PVs/LPVs should undergo clinical breast examination every 6–12 months, starting at the age of 35 [70], and annual prostate cancer screening, starting at the age of 40 (in particular in BRCA2 variant carriers) [71], whereas screening for melanoma and pancreatic cancer should be evaluated on the basis of family history [70]. Overall, we detected a 12.9% of MBC patients with BRCA1/2 alterations with an average age at onset of 62.5 years. This result is in accordance with the findings of other authors [24,72,73,74] and confirms further the high risk of BC also for male carriers, in particular BRCA2 variant carriers. As expected, the majority of these cases have a positive family history for BC/OC. Indeed, the genetic test on the consenting relatives allowed us to detect several carriers of BRCA1/2 alterations, that have been addressed to a surveillance protocol according to the guidelines [32,33].
In addition to BRCA1/2 variants, we identified four PVs in PALB2, ATM, CHEK2 and RAD51C, classified as moderate penetrance genes for the risk of BC and/or OC [5] and two LPVs in unexpected genes, such as BAP1 and EGFR. The patients in which we detected PVs/LPVs in genes other than BRCA1/2 have an average age at MBC onset (55.8 years), that is lower than the age of MBC patients without PVs/LPVs (64.9 years) and also than the age of MBC patients with BRCA1/2 PVs (62.5 years). This result suggests that, besides BRCA1 and BRCA2, the genetic predisposition to MBC is associated with a plethora of genes, some of which are known to be linked to BC risk and others are novel in the field of hereditary BC.
In particular, the PALB2 gene, encoding a protein that interacts with BRCA2 during the homologous recombination, is the most promising gene that emerges from NGS studies on BC/OC predisposition [15,75,76,77]. PALB2 alterations have also been reported recurrently in MBC patients [23,24,27]. Regarding the penetrance, a recent study, conducted on 524 families with PALB2 variants, reported a BC risk of 53% for females and 1% for males by the age of 80 [21]. The same study also reported an increased risk for OC and pancreatic cancer, which has been estimated at 5% and 2%–3%, respectively, by the age of 80. Our patient with the PALB2 alteration had an IDC at 75 years old and reported a family history of cancer, with the mother deceased for BC at 60 years old. The genetic analysis on the relatives showed that the variant is carried also by his sister, who is healthy and now under a surveillance program for BC risk. Our result confirms further the important role of PALB2 gene in MBC predisposition and highlights the importance of developing a surveillance protocol for male carriers, since the current guidelines give suggestions only for the management of female carriers [70].
The CHEK2 gene encodes a tumor suppressor protein involved in DNA damage repair and its germline alterations are associated with an increased BC risk for female carriers [78,79,80,81], which is estimated to be 20%–44% during lifetime [82,83,84]. In addition to BC, CHEK2 PVs/LPVs have also been associated with other cancers [85], including prostate [86,87,88], colorectal [89], and gastric cancers [90]. In particular, the CHEK2 variant c.1100delC, detected in one of our MBC patients, is associated with a two- to three-fold increase in BC risk in women and a ten-fold increase of risk in men [91,92,93]. The patient had an IDC at 36 years old and reported a family history of cancer, in particular, the father with prostate cancer at 70 years and the paternal grandfather with non-Hodgkin lymphoma at 85 years. Unexpectedly, the genetic test on the relatives showed that the variant was harbored by his mother and also by his brother, who are both healthy. This result confirms the moderate penetrance of this CHEK2 variant but, at the same time, suggests further that CHEK2 alterations are associated with BC risk also in men.
We identified also a PV in the ATM gene, encoding a protein involved in DNA repair and cell cycle control, whose germline alterations are associated with an increased BC risk for women, which is estimated to be 15%–60% during lifetime [94,95,96,97,98,99]. ATM alterations have been previously reported in MBC [100,101]; however, some studies detected no increased risk of MBC for ATM variant carriers [24,72]. Our patient who carried the ATM variant had an IDC at 38 years and reported a family history of cancer, in particular, the father deceased for melanoma at 65 years. The very young age at cancer onset of our case and the recurrent detection of ATM deleterious alterations in MBC patients suggest that the involvement of this gene in the predisposition to the disease should be further investigated in larger case series.
We also found a PV in RAD51C gene, encoding a protein involved in homologous recombination. Germline alterations of the RAD51C gene have been associated with an increased OC risk, whereas the BC risk for variant carriers is controversial [102,103,104,105]. Regarding MBC, RAD51C variants were initially excluded for the BC risk for males [31] but, more recently, a study on a large cohort of MBC patients detected PVs in RAD51C and RAD51D genes, encoding proteins of the same complex [24]. Our patient carrying the RAD51C alteration had an IDC at 59 years old and had a sister deceased for BC at 55 years. Our finding reinforces a possible role of RAD51C gene alterations in BC predisposition, even in male carriers, and confirms the rationale of including this gene in a panel for the assessment of BC risk.
Moreover, we identified the c.1110_1116delCATGCAG in the BAP1 gene, encoding a nuclear ubiquitin carboxy-terminal hydrolase that contains binding domains for BRCA1, BARD1 and HCFC1 proteins [106]. Germline deleterious variants in the BAP1 gene are associated with the BAP1 tumor predisposition syndrome, an autosomal dominant condition characterized by an increased risk for atypical Spitz tumors and other types of cancer, such as uveal melanoma, malignant mesothelioma, cutaneous melanoma, clear cell renal cell carcinoma, and basal cell carcinoma [107]. However, BAP1 germline alterations have also been reported in patients with BC and seem to be associated with an increased risk of BC [108,109,110,111,112], even if further studies are needed to confirm this link. Besides this, BAP1 somatic mutations have been identified in sporadic BCs [106,113] and the involvement of BAP1 protein in the BRCA pathway supports the role of this protein in BC development. Indeed, the variant identified in our MBC patient is a frameshift deletion that generates a premature stop codon in the middle of the transcript, destroying completely the BRCA1 binding domain. The patient had an IDC at 65 years, he had no family history of cancer in I- and II-degree relatives but reported a female cousin deceased for BC before the age of 40. Although we were not able to verify the segregation of the disease with the genetic alteration, taking into account all the above considerations, we can conclude that BAP1 is an emerging gene in the predisposition to BC and that also male carriers seem to have an increased BC risk.
The most unexpected result we obtained was the identification of a germline frameshift deletion in EGFR gene, encoding the Epidermal Growth Factor Receptor. It is well known that, in cancer, somatic mutations in EGFR lead to its constant activation with the result of an uncontrolled cell division [114]. On the opposite, germline EGFR missense variants, such as T790M, V834L and V843I, have been associated with rare cases of familial lung adenocarcinoma [115,116,117,118,119,120,121]. Additionally, a germline loss of function variant, affecting the extracellular domain of EGFR, has been described, but it was associated with a severe and lethal form of epithelial inflammation [122,123]. The c.3538_3541delGAAG variant in the EGFR gene, identified in one of our MBC patients, is located in the last exon, encoding the cytoplasmic domain with autophosphorylation function, but almost immediately after the last tyrosine (Y1173) subjected to phosphorylation [124]. The alteration determines the incorporation of 17 wrong amino acids, starting from position 1180, and a premature stop codon at position 1197. With the available information, it is not easy to predict the effect of this variant on the structure and function of the EGFR protein, but the variant has been already reported (rs781064539) [46] and is predicted to be deleterious according to the guidelines [39,48]. Of note, the variant carrier had a DCIS at 62 years old and also a non-Hodgkin lymphoma at 67 years old. The patient had no family history of cancer, so it was impossible to verify the segregation of the variant with the disease in his relatives. Consequently, further studies are needed to assess the real effect of the variant on the protein function and if it can have a role in cancer predisposition.
Finally, we did not identify any PV/LPV in 78.6% of the MBC patients subjected to the genetic test. This finding is compatible with the results obtained by other studies on MBC and, more in general, on genetic predisposition to cancer. We cannot exclude the presence of genetic alterations in regulatory regions or in other genes not analyzed in our panel. MBC, like other tumors, is a multifactorial disease and environmental/behavioral factors play a pivotal role in cancer development. Consequently, many of the MBC cases in which we did not find any genetic alteration could be the result of risk factors not linked to genetic predisposition.

5. Conclusions

Overall, our results confirmed the high risk of BC also for male carriers of BRCA1/2 germline alterations, reinforced the emerging link between MBC and other genes involved in the predisposition to BC, and highlighted the association of novel genes with MBC.

Author Contributions

Conceptualization, D.C. and G.T.; methodology, G.T., M.T.; validation, G.T.; formal analysis, M.T.; investigation, G.T., V.Z., I.C., F.P. and E.F.; data curation, R.D., V.A. and M.R.; writing—original draft preparation, G.T.; writing—review and editing, D.C. and P.U.; visualization, G.T.; supervision, G.M., F.F., P.U., D.C.; project administration, D.C. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no external funding.

Acknowledgments

We are grateful to all the patients enrolled in this study for providing information about their personal and family history. We would like also to thank Rosa Vattiato of the Romagna Cancer Registry for the collection of clinical information of patients.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Bray, F.; Ferlay, J.; Soerjomataram, I.; Siegel, R.L.; Torre, L.A.; Jemal, A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA. Cancer J. Clin. 2018, 68, 394–424. [Google Scholar] [CrossRef] [PubMed]
  2. Ottini, L. Male breast cancer: A rare disease that might uncover underlying pathways of breast cancer. Nat. Rev. Cancer 2014, 14, 643. [Google Scholar] [CrossRef] [PubMed]
  3. Gnerlich, J.L.; Deshpande, A.D.; Jeffe, D.B.; Seelam, S.; Kimbuende, E.; Margenthaler, J.A. Poorer survival outcomes for male breast cancer compared with female breast cancer may be attributable to in-stage migration. Ann. Surg. Oncol. 2011, 18, 1837–1844. [Google Scholar] [CrossRef] [PubMed]
  4. Rizzolo, P.; Silvestri, V.; Tommasi, S.; Pinto, R.; Danza, K.; Falchetti, M.; Gulino, M.; Frati, P.; Ottini, L. Male breast cancer: Genetics, epigenetics, and ethical aspects. Ann. Oncol. Off. J. Eur. Soc. Med. Oncol. 2013, 24 (Suppl. 8), viii75–viii82. [Google Scholar] [CrossRef] [PubMed]
  5. Angeli, D.; Salvi, S.; Tedaldi, G. Genetic Predisposition to Breast and Ovarian Cancers: How Many and Which Genes to Test? Int. J. Mol. Sci. 2020, 21, 1128. [Google Scholar] [CrossRef] [PubMed]
  6. Miki, Y.; Swensen, J.; Shattuck-Eidens, D.; Futreal, P.A.; Harshman, K.; Tavtigian, S.; Liu, Q.; Cochran, C.; Bennett, L.M.; Ding, W. A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1. Science 1994, 266, 66–71. [Google Scholar] [CrossRef]
  7. Wooster, R.; Bignell, G.; Lancaster, J.; Swift, S.; Seal, S.; Mangion, J.; Collins, N.; Gregory, S.; Gumbs, C.; Micklem, G. Identification of the breast cancer susceptibility gene BRCA2. Nature 1995, 378, 789–792. [Google Scholar] [CrossRef]
  8. Petrucelli, N.; Daly, M.B.; Pal, T. BRCA1- and BRCA2-Associated Hereditary Breast and Ovarian Cancer. In GeneReviews®; University of Washington: Seattle, WA, USA, 1998; (updated 2016). [Google Scholar]
  9. Tai, Y.C.; Domchek, S.; Parmigiani, G.; Chen, S. Breast cancer risk among male BRCA1 and BRCA2 mutation carriers. J. Natl. Cancer Inst. 2007, 99, 1811–1814. [Google Scholar] [CrossRef]
  10. Rahman, N. Realizing the promise of cancer predisposition genes. Nature 2014, 505, 302–308. [Google Scholar] [CrossRef]
  11. Easton, D.F.; Pharoah, P.D.P.; Antoniou, A.C.; Tischkowitz, M.; Tavtigian, S.V.; Nathanson, K.L.; Devilee, P.; Meindl, A.; Couch, F.J.; Southey, M.; et al. Gene-panel sequencing and the prediction of breast-cancer risk. N. Engl. J. Med. 2015, 372, 2243–2257. [Google Scholar] [CrossRef]
  12. Apostolou, P.; Fostira, F. Hereditary breast cancer: The era of new susceptibility genes. Biomed. Res. Int. 2013, 2013, 747318. [Google Scholar] [CrossRef] [PubMed]
  13. Kurian, A.W.; Antoniou, A.C.; Domchek, S.M. Refining Breast Cancer Risk Stratification: Additional Genes, Additional Information. Am. Soc. Clin. Oncol. Educ. book. Am. Soc. Clin. Oncol. Annu. Meet. 2016, 35, 44–56. [Google Scholar] [CrossRef] [PubMed]
  14. Nielsen, F.C.; van Overeem Hansen, T.; Sørensen, C.S. Hereditary breast and ovarian cancer: New genes in confined pathways. Nat. Rev. Cancer 2016, 16, 599–612. [Google Scholar] [CrossRef] [PubMed]
  15. Tedaldi, G.; Tebaldi, M.; Zampiga, V.; Danesi, R.; Arcangeli, V.; Ravegnani, M.; Cangini, I.; Pirini, F.; Petracci, E.; Rocca, A.; et al. Multiple-gene panel analysis in a case series of 255 women with hereditary breast and ovarian cancer. Oncotarget 2017, 8, 47064–47075. [Google Scholar] [CrossRef] [PubMed]
  16. Walsh, T.; Lee, M.K.; Casadei, S.; Thornton, A.M.; Stray, S.M.; Pennil, C.; Nord, A.S.; Mandell, J.B.; Swisher, E.M.; King, M.-C. Detection of inherited mutations for breast and ovarian cancer using genomic capture and massively parallel sequencing. Proc. Natl. Acad. Sci. USA 2010, 107, 12629–12633. [Google Scholar] [CrossRef] [PubMed]
  17. Gracia-Aznarez, F.J.; Fernandez, V.; Pita, G.; Peterlongo, P.; Dominguez, O.; de la Hoya, M.; Duran, M.; Osorio, A.; Moreno, L.; Gonzalez-Neira, A.; et al. Whole exome sequencing suggests much of non-BRCA1/BRCA2 familial breast cancer is due to moderate and low penetrance susceptibility alleles. PLoS ONE 2013, 8, e55681. [Google Scholar] [CrossRef] [PubMed]
  18. Kurian, A.W.; Hare, E.E.; Mills, M.A.; Kingham, K.E.; McPherson, L.; Whittemore, A.S.; McGuire, V.; Ladabaum, U.; Kobayashi, Y.; Lincoln, S.E.; et al. Clinical evaluation of a multiple-gene sequencing panel for hereditary cancer risk assessment. J. Clin. Oncol. 2014, 32, 2001–2009. [Google Scholar] [CrossRef]
  19. Desmond, A.; Kurian, A.W.; Gabree, M.; Mills, M.A.; Anderson, M.J.; Kobayashi, Y.; Horick, N.; Yang, S.; Shannon, K.M.; Tung, N.; et al. Clinical Actionability of Multigene Panel Testing for Hereditary Breast and Ovarian Cancer Risk Assessment. JAMA Oncol. 2015, 1, 943–951. [Google Scholar] [CrossRef]
  20. Kraus, C.; Hoyer, J.; Vasileiou, G.; Wunderle, M.; Lux, M.P.; Fasching, P.A.; Krumbiegel, M.; Uebe, S.; Reuter, M.; Beckmann, M.W.; et al. Gene panel sequencing in familial breast/ovarian cancer patients identifies multiple novel mutations also in genes others than BRCA1/2. Int. J. Cancer 2017, 140, 95–102. [Google Scholar] [CrossRef]
  21. Yang, X.; Leslie, G.; Doroszuk, A.; Schneider, S.; Allen, J.; Decker, B.; Dunning, A.M.; Redman, J.; Scarth, J.; Plaskocinska, I.; et al. Cancer Risks Associated With Germline PALB2 Pathogenic Variants: An International Study of 524 Families. J. Clin. Oncol. 2019, JCO1901907. [Google Scholar] [CrossRef]
  22. Silvestri, V.; Rizzolo, P.; Zanna, I.; Falchetti, M.; Masala, G.; Bianchi, S.; Papi, L.; Giannini, G.; Palli, D.; Ottini, L. PALB2 mutations in male breast cancer: A population-based study in Central Italy. Breast Cancer Res. Treat. 2010, 122, 299–301. [Google Scholar] [CrossRef] [PubMed]
  23. Silvestri, V.; Zelli, V.; Valentini, V.; Rizzolo, P.; Navazio, A.S.; Coppa, A.; Agata, S.; Oliani, C.; Barana, D.; Castrignanò, T.; et al. Whole-exome sequencing and targeted gene sequencing provide insights into the role of PALB2 as a male breast cancer susceptibility gene. Cancer 2017, 123, 210–218. [Google Scholar] [CrossRef] [PubMed]
  24. Rizzolo, P.; Zelli, V.; Silvestri, V.; Valentini, V.; Zanna, I.; Bianchi, S.; Masala, G.; Spinelli, A.M.; Tibiletti, M.G.; Russo, A.; et al. Insight into genetic susceptibility to male breast cancer by multigene panel testing: Results from a multicenter study in Italy. Int. J. Cancer 2019, 145, 390–400. [Google Scholar] [CrossRef] [PubMed]
  25. Rizzolo, P.; Silvestri, V.; Bucalo, A.; Zelli, V.; Valentini, V.; Catucci, I.; Zanna, I.; Masala, G.; Bianchi, S.; Spinelli, A.M.; et al. Contribution of MUTYH Variants to Male Breast Cancer Risk: Results From a Multicenter Study in Italy. Front. Oncol. 2018, 8, 583. [Google Scholar] [CrossRef] [PubMed]
  26. Silvestri, V.; Rizzolo, P.; Zelli, V.; Valentini, V.; Zanna, I.; Bianchi, S.; Tibiletti, M.G.; Varesco, L.; Russo, A.; Tommasi, S.; et al. A possible role of FANCM mutations in male breast cancer susceptibility: Results from a multicenter study in Italy. Breast 2018, 38, 92–97. [Google Scholar] [CrossRef] [PubMed]
  27. Ding, Y.C.; Steele, L.; Kuan, C.-J.; Greilac, S.; Neuhausen, S.L. Mutations in BRCA2 and PALB2 in male breast cancer cases from the United States. Breast Cancer Res. Treat. 2011, 126, 771–778. [Google Scholar] [CrossRef]
  28. Fackenthal, J.D.; Marsh, D.J.; Richardson, A.L.; Cummings, S.A.; Eng, C.; Robinson, B.G.; Olopade, O.I. Male breast cancer in Cowden syndrome patients with germline PTEN mutations. J. Med. Genet. 2001, 38, 159–164. [Google Scholar] [CrossRef]
  29. Silvestri, V.; Rizzolo, P.; Falchetti, M.; Zanna, I.; Masala, G.; Bianchi, S.; Palli, D.; Ottini, L. Mutation analysis of BRIP1 in male breast cancer cases: A population-based study in Central Italy. Breast Cancer Res. Treat. 2011, 126, 539–543. [Google Scholar] [CrossRef]
  30. Rizzolo, P.; Silvestri, V.; Valentini, V.; Zelli, V.; Bucalo, A.; Zanna, I.; Bianchi, S.; Tibiletti, M.G.; Russo, A.; Varesco, L.; et al. Evaluation of CYP17A1 and CYP1B1 polymorphisms in male breast cancer risk. Endocr. Connect. 2019, 8, 1224–1229. [Google Scholar] [CrossRef]
  31. Silvestri, V.; Rizzolo, P.; Falchetti, M.; Zanna, I.; Masala, G.; Palli, D.; Ottini, L. Mutation screening of RAD51C in male breast cancer patients. Breast Cancer Res. 2011, 13, 404. [Google Scholar] [CrossRef]
  32. Rischio Eredo-familiare di Tumore al Seno. Available online: https://salute.regione.emilia-romagna.it/screening/tumori-femminili/screeningmammografico/rischio-eredo-familiare (accessed on 29 April 2020).
  33. Collegio Italiano dei Senologi Predisposizione Genetica al Tumore Mammario e Geni BRCA1 e BRCA2. Available online: https://www.senologia.it/wp-content/uploads/2019/10/Carcinoma-eredo-familiare-10.19.pdf (accessed on 29 April 2020).
  34. Tedaldi, G.; Pirini, F.; Tebaldi, M.; Zampiga, V.; Cangini, I.; Danesi, R.; Arcangeli, V.; Ravegnani, M.; Abou Khouzam, R.; Molinari, C.; et al. Multigene Panel Testing Increases the Number of Loci Associated with Gastric Cancer Predisposition. Cancers 2019, 11, 1340. [Google Scholar] [CrossRef] [PubMed]
  35. Li, H.; Durbin, R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 2009, 25, 1754–1760. [Google Scholar] [CrossRef] [PubMed]
  36. McKenna, A.; Hanna, M.; Banks, E.; Sivachenko, A.; Cibulskis, K.; Kernytsky, A.; Garimella, K.; Altshuler, D.; Gabriel, S.; Daly, M.; et al. The Genome Analysis Toolkit: A MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res. 2010, 20, 1297–1303. [Google Scholar] [CrossRef] [PubMed]
  37. Wang, K.; Li, M.; Hakonarson, H. ANNOVAR: Functional annotation of genetic variants from high-throughput sequencing data. Nucleic Acids Res. 2010, 38, e164. [Google Scholar] [CrossRef]
  38. Plon, S.E.; Eccles, D.M.; Easton, D.; Foulkes, W.D.; Genuardi, M.; Greenblatt, M.S.; Hogervorst, F.B.L.; Hoogerbrugge, N.; Spurdle, A.B.; Tavtigian, S.V.; et al. Sequence variant classification and reporting: Recommendations for improving the interpretation of cancer susceptibility genetic test results. Hum. Mutat. 2008, 29, 1282–1291. [Google Scholar] [CrossRef] [PubMed]
  39. Richards, S.; Aziz, N.; Bale, S.; Bick, D.; Das, S.; Gastier-Foster, J.; Grody, W.W.; Hegde, M.; Lyon, E.; Spector, E.; et al. Standards and guidelines for the interpretation of sequence variants: A joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet. Med. 2015, 17, 405–424. [Google Scholar] [CrossRef]
  40. BRCA Exchange. Available online: https://brcaexchange.org (accessed on 29 April 2020).
  41. LOVD-BRCA1. Available online: https://databases.lovd.nl/shared/genes/BRCA1 (accessed on 29 April 2020).
  42. LOVD-BRCA2. Available online: https://databases.lovd.nl/shared/genes/BRCA2 (accessed on 29 April 2020).
  43. Breast Cancer Information Core (BIC). Available online: https://research.nhgri.nih.gov/bic/ (accessed on 29 April 2020).
  44. BRCA Share-BRCA1. Available online: http://www.umd.be/BRCA1 (accessed on 29 April 2020).
  45. BRCA Share-BRCA2. Available online: http://www.umd.be/BRCA2 (accessed on 29 April 2020).
  46. dbSNP–NCBI–NIH. Available online: https://www.ncbi.nlm.nih.gov/snp/ (accessed on 29 April 2020).
  47. ClinVar–NCBI–NIH. Available online: https://www.ncbi.nlm.nih.gov/clinvar/ (accessed on 29 April 2020).
  48. Varsome. Available online: https://varsome.com (accessed on 29 April 2020).
  49. Breast Equivalent Terms and Definition. Available online: https://seer.cancer.gov/tools/solidtumor/Breast_STM.pdf (accessed on 29 April 2020).
  50. Swensen, J.; Hoffman, M.; Skolnick, M.H.; Neuhausen, S.L. Identification of a 14 kb deletion involving the promoter region of BRCA1 in a breast cancer family. Hum. Mol. Genet. 1997, 6, 1513–1517. [Google Scholar] [CrossRef]
  51. Engert, S.; Wappenschmidt, B.; Betz, B.; Kast, K.; Kutsche, M.; Hellebrand, H.; Goecke, T.O.; Kiechle, M.; Niederacher, D.; Schmutzler, R.K.; et al. MLPA screening in the BRCA1 gene from 1,506 German hereditary breast cancer cases: Novel deletions, frequent involvement of exon 17, and occurrence in single early-onset cases. Hum. Mutat. 2008, 29, 948–958. [Google Scholar] [CrossRef]
  52. Yassaee, V.R.; Emamalizadeh, B.; Omrani, M.D. Screening for genomic rearrangements at BRCA1 locus in Iranian women with breast cancer using multiplex ligation-dependent probe amplification. J. Genet. 2013, 92, 131–134. [Google Scholar] [CrossRef]
  53. Puget, N.; Stoppa-Lyonnet, D.; Sinilnikova, O.M.; Pagès, S.; Lynch, H.T.; Lenoir, G.M.; Mazoyer, S. Screening for germ-line rearrangements and regulatory mutations in BRCA1 led to the identification of four new deletions. Cancer Res. 1999, 59, 455–461. [Google Scholar]
  54. Stegel, V.; Krajc, M.; Zgajnar, J.; Teugels, E.; De Grève, J.; Hočevar, M.; Novaković, S. The occurrence of germline BRCA1 and BRCA2 sequence alterations in Slovenian population. BMC Med. Genet. 2011, 12, 9. [Google Scholar] [CrossRef] [PubMed]
  55. Caux-Moncoutier, V.; Castéra, L.; Tirapo, C.; Michaux, D.; Rémon, M.-A.; Laugé, A.; Rouleau, E.; De Pauw, A.; Buecher, B.; Gauthier-Villars, M.; et al. EMMA, a cost- and time-effective diagnostic method for simultaneous detection of point mutations and large-scale genomic rearrangements: Application to BRCA1 and BRCA2 in 1,525 patients. Hum. Mutat. 2011, 32, 325–334. [Google Scholar] [CrossRef] [PubMed]
  56. Iyevleva, A.G.; Suspitsin, E.N.; Kroeze, K.; Gorodnova, T.V.; Sokolenko, A.P.; Buslov, K.G.; Voskresenskiy, D.A.; Togo, A.V.; Kovalenko, S.P.; van der Stoep, N.; et al. Non-founder BRCA1 mutations in Russian breast cancer patients. Cancer Lett. 2010, 298, 258–263. [Google Scholar] [CrossRef] [PubMed]
  57. Arnold, A.G.; Otegbeye, E.; Fleischut, M.H.; Glogowski, E.A.; Siegel, B.; Boyar, S.R.; Salo-Mullen, E.; Amoroso, K.; Sheehan, M.; Berliner, J.L.; et al. Assessment of individuals with BRCA1 and BRCA2 large rearrangements in high-risk breast and ovarian cancer families. Breast Cancer Res. Treat. 2014, 145, 625–634. [Google Scholar] [CrossRef] [PubMed]
  58. Mangone, L.; Ferrari, F.; Mancuso, P.; Carrozzi, G.; Michiara, M.; Falcini, F.; Piffer, S.; Filiberti, R.A.; Caldarella, A.; Vitale, F.; et al. Epidemiology and biological characteristics of male breast cancer in Italy. Breast Cancer 2020. [Google Scholar] [CrossRef] [PubMed]
  59. Antoniou, A.; Pharoah, P.D.P.; Narod, S.; Risch, H.A.; Eyfjord, J.E.; Hopper, J.L.; Loman, N.; Olsson, H.; Johannsson, O.; Borg, A.; et al. Average risks of breast and ovarian cancer associated with BRCA1 or BRCA2 mutations detected in case Series unselected for family history: A combined analysis of 22 studies. Am. J. Hum. Genet. 2003, 72, 1117–1130. [Google Scholar] [CrossRef] [PubMed]
  60. Chen, S.; Parmigiani, G. Meta-analysis of BRCA1 and BRCA2 penetrance. J. Clin. Oncol. 2007, 25, 1329–1333. [Google Scholar] [CrossRef]
  61. Mavaddat, N.; Peock, S.; Frost, D.; Ellis, S.; Platte, R.; Fineberg, E.; Evans, D.G.; Izatt, L.; Eeles, R.A.; Adlard, J.; et al. Cancer risks for BRCA1 and BRCA2 mutation carriers: Results from prospective analysis of EMBRACE. J. Natl. Cancer Inst. 2013, 105, 812–822. [Google Scholar] [CrossRef]
  62. Ford, D.; Easton, D.F.; Bishop, D.T.; Narod, S.A.; Goldgar, D.E. Risks of cancer in BRCA1-mutation carriers. Breast Cancer Linkage Consortium. Lancet 1994, 343, 692–695. [Google Scholar] [CrossRef]
  63. Leongamornlert, D.; Mahmud, N.; Tymrakiewicz, M.; Saunders, E.; Dadaev, T.; Castro, E.; Goh, C.; Govindasami, K.; Guy, M.; O’Brien, L.; et al. Germline BRCA1 mutations increase prostate cancer risk. Br. J. Cancer 2012, 106, 1697–1701. [Google Scholar] [CrossRef]
  64. Thompson, D.; Easton, D.F. Breast Cancer Linkage Consortium Cancer Incidence in BRCA1 mutation carriers. J. Natl. Cancer Inst. 2002, 94, 1358–1365. [Google Scholar] [CrossRef] [PubMed]
  65. Ghiorzo, P.; Pensotti, V.; Fornarini, G.; Sciallero, S.; Battistuzzi, L.; Belli, F.; Bonelli, L.; Borgonovo, G.; Bruno, W.; Gozza, A.; et al. Contribution of germline mutations in the BRCA and PALB2 genes to pancreatic cancer in Italy. Fam. Cancer 2012, 11, 41–47. [Google Scholar] [CrossRef] [PubMed]
  66. Kote-Jarai, Z.; Leongamornlert, D.; Saunders, E.; Tymrakiewicz, M.; Castro, E.; Mahmud, N.; Guy, M.; Edwards, S.; O’Brien, L.; Sawyer, E.; et al. BRCA2 is a moderate penetrance gene contributing to young-onset prostate cancer: Implications for genetic testing in prostate cancer patients. Br. J. Cancer 2011, 105, 1230–1234. [Google Scholar] [CrossRef] [PubMed]
  67. Iqbal, J.; Ragone, A.; Lubinski, J.; Lynch, H.T.; Moller, P.; Ghadirian, P.; Foulkes, W.D.; Armel, S.; Eisen, A.; Neuhausen, S.L.; et al. The incidence of pancreatic cancer in BRCA1 and BRCA2 mutation carriers. Br. J. Cancer 2012, 107, 2005–2009. [Google Scholar] [CrossRef] [PubMed]
  68. Moran, A.; O’Hara, C.; Khan, S.; Shack, L.; Woodward, E.; Maher, E.R.; Lalloo, F.; Evans, D.G.R. Risk of cancer other than breast or ovarian in individuals with BRCA1 and BRCA2 mutations. Fam. Cancer 2012, 11, 235–242. [Google Scholar] [CrossRef] [PubMed]
  69. Breast Cancer Linkage Consortium. Cancer risks in BRCA2 mutation carriers. J. Natl. Cancer Inst. 1999, 91, 1310–1316. [Google Scholar] [CrossRef]
  70. National Comprehensive Cancer Network (NCCN). Guidelines: Genetic/Familial High-Risk Assessment: Breast, Ovarian and Pancreatic. Version 1.2020. Available online: https://www.nccn.org/professionals/physician_gls/pdf/genetics_bop.pdf (accessed on 29 April 2020).
  71. National Comprehensive Cancer Network (NCCN). Guidelines: Prostate Cancer Early Detection. Version 2.2019. Available online: https://www.nccn.org/professionals/physician_gls/pdf/prostate_detection.pdf (accessed on 29 April 2020).
  72. Pritzlaff, M.; Summerour, P.; McFarland, R.; Li, S.; Reineke, P.; Dolinsky, J.S.; Goldgar, D.E.; Shimelis, H.; Couch, F.J.; Chao, E.C.; et al. Male breast cancer in a multi-gene panel testing cohort: Insights and unexpected results. Breast Cancer Res. Treat. 2017, 161, 575–586. [Google Scholar] [CrossRef]
  73. Wolpert, N.; Warner, E.; Seminsky, M.F.; Futreal, A.; Narod, S.A. Prevalence of BRCA1 and BRCA2 mutations in male breast cancer patients in Canada. Clin. Breast Cancer 2000, 1, 57–63. [Google Scholar] [CrossRef]
  74. Silvestri, V.; Barrowdale, D.; Mulligan, A.M.; Neuhausen, S.L.; Fox, S.; Karlan, B.Y.; Mitchell, G.; James, P.; Thull, D.L.; Zorn, K.K.; et al. Male breast cancer in BRCA1 and BRCA2 mutation carriers: Pathology data from the Consortium of Investigators of Modifiers of BRCA1/2. Breast Cancer Res. 2016, 18, 15. [Google Scholar] [CrossRef]
  75. Antoniou, A.C.; Casadei, S.; Heikkinen, T.; Barrowdale, D.; Pylkäs, K.; Roberts, J.; Lee, A.; Subramanian, D.; De Leeneer, K.; Fostira, F.; et al. Breast-cancer risk in families with mutations in PALB2. N. Engl. J. Med. 2014, 371, 497–506. [Google Scholar] [CrossRef]
  76. Antoniou, A.C.; Foulkes, W.D.; Tischkowitz, M. Breast-cancer risk in families with mutations in PALB2. N. Engl. J. Med. 2014, 371, 1651–1652. [Google Scholar] [CrossRef] [PubMed]
  77. Casadei, S.; Norquist, B.M.; Walsh, T.; Stray, S.; Mandell, J.B.; Lee, M.K.; Stamatoyannopoulos, J.A.; King, M.-C. Contribution of inherited mutations in the BRCA2-interacting protein PALB2 to familial breast cancer. Cancer Res. 2011, 71, 2222–2229. [Google Scholar] [CrossRef] [PubMed]
  78. Adank, M.A.; Jonker, M.A.; Kluijt, I.; van Mil, S.E.; Oldenburg, R.A.; Mooi, W.J.; Hogervorst, F.B.L.; van den Ouweland, A.M.W.; Gille, J.J.P.; Schmidt, M.K.; et al. CHEK2*1100delC homozygosity is associated with a high breast cancer risk in women. J. Med. Genet. 2011, 48, 860–863. [Google Scholar] [CrossRef] [PubMed]
  79. Walsh, T.; Casadei, S.; Coats, K.H.; Swisher, E.; Stray, S.M.; Higgins, J.; Roach, K.C.; Mandell, J.; Lee, M.K.; Ciernikova, S.; et al. Spectrum of mutations in BRCA1, BRCA2, CHEK2, and TP53 in families at high risk of breast cancer. JAMA 2006, 295, 1379–1388. [Google Scholar] [CrossRef] [PubMed]
  80. Desrichard, A.; Bidet, Y.; Uhrhammer, N.; Bignon, Y.-J. CHEK2 contribution to hereditary breast cancer in non-BRCA families. Breast Cancer Res. 2011, 13, R119. [Google Scholar] [CrossRef] [PubMed]
  81. Tedaldi, G.; Danesi, R.; Zampiga, V.; Tebaldi, M.; Bedei, L.; Zoli, W.; Amadori, D.; Falcini, F.; Calistri, D. First evidence of a large CHEK2 duplication involved in cancer predisposition in an Italian family with hereditary breast cancer. BMC Cancer 2014, 14, 478. [Google Scholar] [CrossRef]
  82. Meijers-Heijboer, H.; van den Ouweland, A.; Klijn, J.; Wasielewski, M.; de Snoo, A.; Oldenburg, R.; Hollestelle, A.; Houben, M.; Crepin, E.; van Veghel-Plandsoen, M.; et al. Low-penetrance susceptibility to breast cancer due to CHEK2(*)1100delC in noncarriers of BRCA1 or BRCA2 mutations. Nat. Genet. 2002, 31, 55–59. [Google Scholar]
  83. Weischer, M.; Bojesen, S.E.; Ellervik, C.; Tybjaerg-Hansen, A.; Nordestgaard, B.G. CHEK2*1100delC genotyping for clinical assessment of breast cancer risk: Meta-analyses of 26,000 patient cases and 27,000 controls. J. Clin. Oncol. 2008, 26, 542–548. [Google Scholar] [CrossRef]
  84. Cybulski, C.; Wokołorczyk, D.; Jakubowska, A.; Huzarski, T.; Byrski, T.; Gronwald, J.; Masojć, B.; Deebniak, T.; Górski, B.; Blecharz, P.; et al. Risk of breast cancer in women with a CHEK2 mutation with and without a family history of breast cancer. J. Clin. Oncol. 2011, 29, 3747–3752. [Google Scholar] [CrossRef]
  85. Cybulski, C.; Górski, B.; Huzarski, T.; Masojć, B.; Mierzejewski, M.; Debniak, T.; Teodorczyk, U.; Byrski, T.; Gronwald, J.; Matyjasik, J.; et al. CHEK2 is a multiorgan cancer susceptibility gene. Am. J. Hum. Genet. 2004, 75, 1131–1135. [Google Scholar] [CrossRef]
  86. Dong, X.; Wang, L.; Taniguchi, K.; Wang, X.; Cunningham, J.M.; McDonnell, S.K.; Qian, C.; Marks, A.F.; Slager, S.L.; Peterson, B.J.; et al. Mutations in CHEK2 associated with prostate cancer risk. Am. J. Hum. Genet. 2003, 72, 270–280. [Google Scholar] [CrossRef] [PubMed]
  87. Cybulski, C.; Huzarski, T.; Górski, B.; Masojć, B.; Mierzejewski, M.; Debniak, T.; Gliniewicz, B.; Matyjasik, J.; Złowocka, E.; Kurzawski, G.; et al. A novel founder CHEK2 mutation is associated with increased prostate cancer risk. Cancer Res. 2004, 64, 2677–2679. [Google Scholar] [CrossRef] [PubMed]
  88. Cybulski, C.; Wokołorczyk, D.; Huzarski, T.; Byrski, T.; Gronwald, J.; Górski, B.; Debniak, T.; Masojć, B.; Jakubowska, A.; Gliniewicz, B.; et al. A large germline deletion in the Chek2 kinase gene is associated with an increased risk of prostate cancer. J. Med. Genet. 2006, 43, 863–866. [Google Scholar] [CrossRef] [PubMed]
  89. Meijers-Heijboer, H.; Wijnen, J.; Vasen, H.; Wasielewski, M.; Wagner, A.; Hollestelle, A.; Elstrodt, F.; van den Bos, R.; de Snoo, A.; Fat, G.T.A.; et al. The CHEK2 1100delC mutation identifies families with a hereditary breast and colorectal cancer phenotype. Am. J. Hum. Genet. 2003, 72, 1308–1314. [Google Scholar] [CrossRef] [PubMed]
  90. Teodorczyk, U.; Cybulski, C.; Wokołorczyk, D.; Jakubowska, A.; Starzyńska, T.; Lawniczak, M.; Domagała, P.; Ferenc, K.; Marlicz, K.; Banaszkiewicz, Z.; et al. The risk of gastric cancer in carriers of CHEK2 mutations. Fam. Cancer 2013, 12, 473–478. [Google Scholar] [CrossRef] [PubMed]
  91. CHEK2 Breast Cancer Case-Control Consortium. CHEK2*1100delC and susceptibility to breast cancer: A collaborative analysis involving 10,860 breast cancer cases and 9,065 controls from 10 studies. Am. J. Hum. Genet. 2004, 74, 1175–1182. [Google Scholar] [CrossRef] [PubMed]
  92. Bernstein, J.L.; Teraoka, S.N.; John, E.M.; Andrulis, I.L.; Knight, J.A.; Lapinski, R.; Olson, E.R.; Wolitzer, A.L.; Seminara, D.; Whittemore, A.S.; et al. The CHEK2*1100delC allelic variant and risk of breast cancer: Screening results from the Breast Cancer Family Registry. Cancer Epidemiol. Biomark. Prev. 2006, 15, 348–352. [Google Scholar] [CrossRef]
  93. Weischer, M.; Bojesen, S.E.; Tybjaerg-Hansen, A.; Axelsson, C.K.; Nordestgaard, B.G. Increased risk of breast cancer associated with CHEK2*1100delC. J. Clin. Oncol. 2007, 25, 57–63. [Google Scholar] [CrossRef]
  94. Thompson, D.; Duedal, S.; Kirner, J.; McGuffog, L.; Last, J.; Reiman, A.; Byrd, P.; Taylor, M.; Easton, D.F. Cancer risks and mortality in heterozygous ATM mutation carriers. J. Natl. Cancer Inst. 2005, 97, 813–822. [Google Scholar] [CrossRef]
  95. Renwick, A.; Thompson, D.; Seal, S.; Kelly, P.; Chagtai, T.; Ahmed, M.; North, B.; Jayatilake, H.; Barfoot, R.; Spanova, K.; et al. ATM mutations that cause ataxia-telangiectasia are breast cancer susceptibility alleles. Nat. Genet. 2006, 38, 873–875. [Google Scholar] [CrossRef]
  96. Goldgar, D.E.; Healey, S.; Dowty, J.G.; Da Silva, L.; Chen, X.; Spurdle, A.B.; Terry, M.B.; Daly, M.J.; Buys, S.M.; Southey, M.C.; et al. Rare variants in the ATM gene and risk of breast cancer. Breast Cancer Res. 2011, 13, R73. [Google Scholar] [CrossRef] [PubMed]
  97. Marabelli, M.; Cheng, S.-C.; Parmigiani, G. Penetrance of ATM Gene Mutations in Breast Cancer: A Meta-Analysis of Different Measures of Risk. Genet. Epidemiol. 2016, 40, 425–431. [Google Scholar] [CrossRef] [PubMed]
  98. Van Os, N.J.H.; Roeleveld, N.; Weemaes, C.M.R.; Jongmans, M.C.J.; Janssens, G.O.; Taylor, A.M.R.; Hoogerbrugge, N.; Willemsen, M.A.A.P. Health risks for ataxia-telangiectasia mutated heterozygotes: A systematic review, meta-analysis and evidence-based guideline. Clin. Genet. 2016, 90, 105–117. [Google Scholar] [CrossRef] [PubMed]
  99. Tavtigian, S.V.; Oefner, P.J.; Babikyan, D.; Hartmann, A.; Healey, S.; Le Calvez-Kelm, F.; Lesueur, F.; Byrnes, G.B.; Chuang, S.-C.; Forey, N.; et al. Rare, evolutionarily unlikely missense substitutions in ATM confer increased risk of breast cancer. Am. J. Hum. Genet. 2009, 85, 427–446. [Google Scholar] [CrossRef] [PubMed]
  100. Fostira, F.; Saloustros, E.; Apostolou, P.; Vagena, A.; Kalfakakou, D.; Mauri, D.; Tryfonopoulos, D.; Georgoulias, V.; Yannoukakos, D.; Fountzilas, G.; et al. Germline deleterious mutations in genes other than BRCA2 are infrequent in male breast cancer. Breast Cancer Res. Treat. 2018, 169, 105–113. [Google Scholar] [CrossRef]
  101. Tung, N.; Battelli, C.; Allen, B.; Kaldate, R.; Bhatnagar, S.; Bowles, K.; Timms, K.; Garber, J.E.; Herold, C.; Ellisen, L.; et al. Frequency of mutations in individuals with breast cancer referred for BRCA1 and BRCA2 testing using next-generation sequencing with a 25-gene panel. Cancer 2015, 121, 25–33. [Google Scholar] [CrossRef]
  102. Loveday, C.; Turnbull, C.; Ruark, E.; Xicola, R.M.M.; Ramsay, E.; Hughes, D.; Warren-Perry, M.; Snape, K.; Breast Cancer Susceptibility Collaboration (UK); Eccles, D.; et al. Germline RAD51C mutations confer susceptibility to ovarian cancer. Nat. Genet. 2012, 44, 475–476. [Google Scholar] [CrossRef]
  103. Song, H.; Dicks, E.; Ramus, S.J.; Tyrer, J.P.; Intermaggio, M.P.; Hayward, J.; Edlund, C.K.; Conti, D.; Harrington, P.; Fraser, L.; et al. Contribution of Germline Mutations in the RAD51B, RAD51C, and RAD51D Genes to Ovarian Cancer in the Population. J. Clin. Oncol. 2015, 33, 2901–2907. [Google Scholar] [CrossRef]
  104. Akbari, M.R.; Tonin, P.; Foulkes, W.D.; Ghadirian, P.; Tischkowitz, M.; Narod, S.A. RAD51C germline mutations in breast and ovarian cancer patients. Breast Cancer Res. 2010, 12, 404. [Google Scholar] [CrossRef]
  105. Meindl, A.; Hellebrand, H.; Wiek, C.; Erven, V.; Wappenschmidt, B.; Niederacher, D.; Freund, M.; Lichtner, P.; Hartmann, L.; Schaal, H.; et al. Germline mutations in breast and ovarian cancer pedigrees establish RAD51C as a human cancer susceptibility gene. Nat. Genet. 2010, 42, 410–414. [Google Scholar] [CrossRef]
  106. Jensen, D.E.; Proctor, M.; Marquis, S.T.; Gardner, H.P.; Ha, S.I.; Chodosh, L.A.; Ishov, A.M.; Tommerup, N.; Vissing, H.; Sekido, Y.; et al. BAP1: A novel ubiquitin hydrolase which binds to the BRCA1 RING finger and enhances BRCA1-mediated cell growth suppression. Oncogene 1998, 16, 1097–1112. [Google Scholar] [CrossRef] [PubMed]
  107. Pilarski, R.; Rai, K.; Cebulla, C.; Abdel-Rahman, M. BAP1 Tumor Predisposition Syndrome. In GeneReviews®; University of Washington: Seattle, WA, USA, 2016; (updated 2020). [Google Scholar]
  108. Testa, J.R.; Cheung, M.; Pei, J.; Below, J.E.; Tan, Y.; Sementino, E.; Cox, N.J.; Dogan, A.U.; Pass, H.I.; Trusa, S.; et al. Germline BAP1 mutations predispose to malignant mesothelioma. Nat. Genet. 2011, 43, 1022–1025. [Google Scholar] [CrossRef] [PubMed]
  109. Njauw, C.-N.J.; Kim, I.; Piris, A.; Gabree, M.; Taylor, M.; Lane, A.M.; DeAngelis, M.M.; Gragoudas, E.; Duncan, L.M.; Tsao, H. Germline BAP1 inactivation is preferentially associated with metastatic ocular melanoma and cutaneous-ocular melanoma families. PLoS ONE 2012, 7, e35295. [Google Scholar] [CrossRef] [PubMed]
  110. Popova, T.; Hebert, L.; Jacquemin, V.; Gad, S.; Caux-Moncoutier, V.; Dubois-d’Enghien, C.; Richaudeau, B.; Renaudin, X.; Sellers, J.; Nicolas, A.; et al. Germline BAP1 mutations predispose to renal cell carcinomas. Am. J. Hum. Genet. 2013, 92, 974–980. [Google Scholar] [CrossRef] [PubMed]
  111. Pilarski, R.; Cebulla, C.M.; Massengill, J.B.; Rai, K.; Rich, T.; Strong, L.; McGillivray, B.; Asrat, M.-J.; Davidorf, F.H.; Abdel-Rahman, M.H. Expanding the clinical phenotype of hereditary BAP1 cancer predisposition syndrome, reporting three new cases. Genes. Chromosomes Cancer 2014, 53, 177–182. [Google Scholar] [CrossRef]
  112. Coupier, I.; Cousin, P.-Y.; Hughes, D.; Legoix-Né, P.; Trehin, A.; Sinilnikova, O.M.; Stoppa-Lyonnet, D. BAP1 and breast cancer risk. Fam. Cancer 2005, 4, 273–277. [Google Scholar] [CrossRef]
  113. Wood, L.D.; Parsons, D.W.; Jones, S.; Lin, J.; Sjöblom, T.; Leary, R.J.; Shen, D.; Boca, S.M.; Barber, T.; Ptak, J.; et al. The genomic landscapes of human breast and colorectal cancers. Science 2007, 318, 1108–1113. [Google Scholar] [CrossRef]
  114. Lynch, T.J.; Bell, D.W.; Sordella, R.; Gurubhagavatula, S.; Okimoto, R.A.; Brannigan, B.W.; Harris, P.L.; Haserlat, S.M.; Supko, J.G.; Haluska, F.G.; et al. Activating mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib. N. Engl. J. Med. 2004, 350, 2129–2139. [Google Scholar] [CrossRef]
  115. Bell, D.W.; Gore, I.; Okimoto, R.A.; Godin-Heymann, N.; Sordella, R.; Mulloy, R.; Sharma, S.V.; Brannigan, B.W.; Mohapatra, G.; Settleman, J.; et al. Inherited susceptibility to lung cancer may be associated with the T790M drug resistance mutation in EGFR. Nat. Genet. 2005, 37, 1315–1316. [Google Scholar] [CrossRef]
  116. Van der Leest, C.; Wagner, A.; Pedrosa, R.M.; Aerts, J.G.; Dinjens, W.N.M.; Dubbink, H.J. Novel EGFR V834L Germline Mutation Associated With Familial Lung Adenocarcinoma. JCO Precis. Oncol. 2018, 2, 1–5. [Google Scholar] [CrossRef]
  117. Ohtsuka, K.; Ohnishi, H.; Fujiwara, M.; Morii, T.; Matsushima, S.; Ogura, W.; Yamasaki, S.; Kishino, T.; Tanaka, R.; Watanabe, T. Predisposition to Lung Adenocarcinoma in a Family Harboring the Germline EGFR V843I Mutation. JCO Precis. Oncol. 2019, 3, 1–4. [Google Scholar] [CrossRef]
  118. Oxnard, G.R.; Nguyen, K.-S.H.; Costa, D.B. Germline mutations in driver oncogenes and inherited lung cancer risk independent of smoking history. J. Natl. Cancer Inst. 2014, 106, djt361. [Google Scholar] [CrossRef] [PubMed]
  119. Ikeda, K.; Nomori, H.; Mori, T.; Sasaki, J.; Kobayashi, T. Novel germline mutation: EGFR V843I in patient with multiple lung adenocarcinomas and family members with lung cancer. Ann. Thorac. Surg. 2008, 85, 1430–1432. [Google Scholar] [CrossRef] [PubMed]
  120. Ohtsuka, K.; Ohnishi, H.; Kurai, D.; Matsushima, S.; Morishita, Y.; Shinonaga, M.; Goto, H.; Watanabe, T. Familial lung adenocarcinoma caused by the EGFR V843I germ-line mutation. J. Clin. Oncol. 2011, 29, e191–e192. [Google Scholar] [CrossRef]
  121. Demierre, N.; Zoete, V.; Michielin, O.; Stauffer, E.; Zimmermann, D.R.; Betticher, D.C.; Peters, S. A dramatic lung cancer course in a patient with a rare EGFR germline mutation exon 21 V843I: Is EGFR TKI resistance predictable? Lung Cancer 2013, 80, 81–84. [Google Scholar] [CrossRef]
  122. Campbell, P.; Morton, P.E.; Takeichi, T.; Salam, A.; Roberts, N.; Proudfoot, L.E.; Mellerio, J.E.; Aminu, K.; Wellington, C.; Patil, S.N.; et al. Epithelial inflammation resulting from an inherited loss-of-function mutation in EGFR. J. Investig. Dermatol. 2014, 134, 2570–2578. [Google Scholar] [CrossRef]
  123. Ganetzky, R.; Finn, E.; Bagchi, A.; Zollo, O.; Conlin, L.; Deardorff, M.; Harr, M.; Simpson, M.A.; McGrath, J.A.; Zackai, E.; et al. EGFR mutations cause a lethal syndrome of epithelial dysfunction with progeroid features. Mol. Genet. Genom. Med. 2015, 3, 452–458. [Google Scholar] [CrossRef]
  124. Downward, J.; Parker, P.; Waterfield, M.D. Autophosphorylation sites on the epidermal growth factor receptor. Nature 1984, 311, 483–485. [Google Scholar] [CrossRef]
Figure 1. Pie chart showing the fraction of cases with/without PVs/LPVs; the number of variant carriers is reported between brackets.
Figure 1. Pie chart showing the fraction of cases with/without PVs/LPVs; the number of variant carriers is reported between brackets.
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Figure 2. (a) Schematic representation of the BRCA1 gene (transcript NM_007294) and localization of the three PVs identified in our cohort; (b) schematic representation of the BRCA2 gene (transcript NM_000059) and localization of the six PVs identified in our cohort. Exons are represented with grey boxes for non-coding exons and with blue and orange boxes for coding exons of BRCA1 and BRCA2, respectively; the exon numbering has been reported above the exons (the traditional exon numbering of BRCA1 gene lacks exon four that has not been represented).
Figure 2. (a) Schematic representation of the BRCA1 gene (transcript NM_007294) and localization of the three PVs identified in our cohort; (b) schematic representation of the BRCA2 gene (transcript NM_000059) and localization of the six PVs identified in our cohort. Exons are represented with grey boxes for non-coding exons and with blue and orange boxes for coding exons of BRCA1 and BRCA2, respectively; the exon numbering has been reported above the exons (the traditional exon numbering of BRCA1 gene lacks exon four that has not been represented).
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Table 1. List of the 94 cancer predisposition genes included in the Trusight Cancer panel.
Table 1. List of the 94 cancer predisposition genes included in the Trusight Cancer panel.
Genes
AIPALKAPCATMBAP1BLMBMPR1ABRCA1BRCA2BRIP1
BUB1BCDC73CDH1CDK4CDKN1CCDKN2ACEBPACEP57CHEK2CYLD
DDB2DICER1DIS3L2EGFREPCAMERCC2ERCC3ERCC4ERCC5EXT1
EXT2EZH2FANCAFANCBFANCCFANCD2FANCEFANCFFANCGFANCI
FANCLFANCMFHFLCNGATA2GPC3HNF1AHRASKITMAX
MEN1METMLH1MSH2MSH6MUTYHNBNNF1NF2NSD1
PALB2PHOX2BPMS1PMS2PRF1PRKAR1APTCH1PTENRAD51CRAD51D
RB1RECQL4RETRHBDF2RUNX1SBDSSDHAF2SDHBSDHCSDHD
SLX4SMAD4SMARCB1STK11SUFUTMEM127TP53TSC1TSC2VHL
WRNWT1XPAXPC
Table 2. Carriers of BRCA1/2 pathogenic and likely-pathogenic variants.
Table 2. Carriers of BRCA1/2 pathogenic and likely-pathogenic variants.
Patient IDCancerAge at OnsetGeneChrcDNA (Transcript)ProteinVariant TypeIARC Class [38]dbSNP [46]ClinVar [47]
A142IDC55yBRCA117q21.31c.-113-?_80+?del (NM_007294)p.?large deletion5
A774IDC69yBRCA117q21.31c.4964_4982del (NM_007294)p.Ser1655Tyrfs*16frameshift deletion5rs80359876pathogenic
TR140IDC57yBRCA117q21.31c.5266dupC (NM_007294)p.Gln1756Profs*74frameshift duplication5rs80357906pathogenic
A379IDC58yBRCA213q13.1c.1238delT (NM_000059)p.Leu413Hisfs*17frameshift deletion5rs80359271pathogenic
A581IDC77yBRCA213q13.1c.1813delA (NM_000059)p.Ile605Tyrfs*9frameshift deletion5rs80359306pathogenic
T096DCIS68yBRCA213q13.1c.3195_3198delTAAT (NM_000059)p.Asn1066Leufs*10frameshift deletion5rs80359375pathogenic
B156IDC64yBRCA213q13.1c.5073dupA (NM_000059)p.Trp1692Metfs*3frameshift duplication5rs80359479pathogenic
A933IDC59yBRCA213q13.1c.6039delA (NM_000059)p.Val2014Tyrfs*26frameshift deletion5rs876660637pathogenic
A98IDC56yBRCA213q13.1c.6039delA (NM_000059)p.Val2014Tyrfs*26frameshift deletion5rs876660637pathogenic
IDC: infiltrating ductal carcinoma; DCIS: ductal carcinoma in situ; chr: chromosomal locus.
Table 3. Carriers of pathogenic and likely-pathogenic variants in genes other than BRCA1/2.
Table 3. Carriers of pathogenic and likely-pathogenic variants in genes other than BRCA1/2.
Patient IDCancerAge at OnsetGeneChrcDNAProteinVariant TypeIARC Class [38]dbSNP [46]ClinVar [47]
A841IDC38yATM11q22.3c.8319_8323dupTGTCC (NM_000051)p.Pro2775Leufs*33frameshift duplication5rs1555135596pathogenic
A625IDC65yBAP13p21.1c.1110_1116delCATGCAG (NM_004656)p.Met371Argfs*57frameshift deletion4
A512IDC36yCHEK222q12.1c.1100delC (NM_007194)p.Thr367Metfs*15frameshift deletion5rs555607708pathogenic
A225DCIS62yEGFR7p11.2c.3538_3541delGAAG (NM_005228)p.Glu1180Profs*18frameshift deletion4rs781064539
B887IDC75yPALB216p12.2c.73A>T (NM_024675)p.Lys25*nonsense variant5rs1248579792pathogenic
A334IDC59yRAD51C17q22c.181_182delCT (NM_058216)p.Leu61Alafs*11frameshift deletion5rs786203945pathogenic
IDC: infiltrating ductal carcinoma; DCIS: ductal carcinoma in situ; chr: chromosomal locus.
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