Gender-Specific Genetic Predisposition to Breast Cancer: BRCA Genes and Beyond

Simple Summary The role of gender in oncology is an issue that is starting to be recognized as being of extreme importance in the last few years. While breast cancer is commonly perceived as a female-only disease, it does also occur in men, although rarely, thus opening relevant gender issues. Breast cancer in men is much less studied, with most knowledge coming from research on female breast cancer; however, several crucial differences have begun to be discovered between male and female patients. For example, the gender-specific impact and magnitude of risks conferred by breast cancer genetic risk factors are emerging, and they should be taken into consideration for a proper personalized clinical management. Overall, addressing all the challenges and the open issues regarding breast cancer genetic predisposition, including gender, will have an important clinical impact on the management of patients of both sexes. Abstract Among neoplastic diseases, breast cancer (BC) is one of the most influenced by gender. Despite common misconceptions associating BC as a women-only disease, BC can also occur in men. Additionally, transgender individuals may also experience BC. Genetic risk factors play a relevant role in BC predisposition, with important implications in precision prevention and treatment. The genetic architecture of BC susceptibility is similar in women and men, with high-, moderate-, and low-penetrance risk variants; however, some sex-specific features have emerged. Inherited high-penetrance pathogenic variants (PVs) in BRCA1 and BRCA2 genes are the strongest BC genetic risk factor. BRCA1 and BRCA2 PVs are more commonly associated with increased risk of female and male BC, respectively. Notably, BRCA-associated BCs are characterized by sex-specific pathologic features. Recently, next-generation sequencing technologies have helped to provide more insights on the role of moderate-penetrance BC risk variants, particularly in PALB2, CHEK2, and ATM genes, while international collaborative genome-wide association studies have contributed evidence on common low-penetrance BC risk variants, on their combined effect in polygenic models, and on their role as risk modulators in BRCA1/2 PV carriers. Overall, all these studies suggested that the genetic basis of male BC, although similar, may differ from female BC. Evaluating the genetic component of male BC as a distinct entity from female BC is the first step to improve both personalized risk assessment and therapeutic choices of patients of both sexes in order to reach gender equality in BC care. In this review, we summarize the latest research in the field of BC genetic predisposition with a particular focus on similarities and differences in male and female BC, and we also discuss the implications, challenges, and open issues that surround the establishment of a gender-oriented clinical management for BC.


Introduction
Gender-sensitive medicine is an innovative approach that takes into consideration the impact of differences in biological sex or gender identity on health and disease status, with the goal of improving prevention, screening, diagnosis, and treatment of all individuals [1].
The importance of gender in oncology is starting to be recognized, but this issue is still undervalued if compared to other medical disciplines [1].The interplay between genetic mediators, hormonal mediators, including estrogens, progesterone and androgens, age, and reproductive status may modulate both local determinants of carcinogenesis, such as cancer-initiating cells and the tumor microenvironment, and systemic ones, such as cell metabolism and the immune system [2].
It is noteworthy that all these aforementioned features are largely involved in the pathogenesis of breast cancer (BC).Indeed, among neoplastic diseases, BC is one of the most influenced by gender.Although often mistakenly considered as a female-only disease, BC may also occur in men (referring to cisgender men) [3,4].
Moreover, BC may also occur in transgender individuals, both female-to-male (transgender men) and male-to-female (transgender women), although with a lower incidence compared with women (referring to cisgender women) [5].
Cancer arising in female breast tissue (female breast cancer, FBC) is the most common cancer and one of the leading causes of death among women globally [6], while cancer arising in male breast tissue (male breast cancer, MBC) is a rare disease, representing less than 1% of all cancers in men and of all BCs [3].However, MBC incidence has been increasing over the last 30 years, and morbidity in MBC patients is a serious concern [7,8].
The rarity of MBC has precluded the development of ad hoc clinical trials, and currently, clinical management and therapeutic options of MBC patients is informed almost entirely by FBC research [9].Notably, although an improvement in BC survival was observed in the last decades, mortality after cancer diagnosis is higher among male patients with BC compared with their female counterparts, even after accounting for possible prognostic factors, suggesting that such disparity may be due to factors yet to be identified [10,11].Indeed, although MBC is thought to resemble post-menopausal FBC, increasing evidence indicates that MBC may be different, with unique molecular features, suggesting sex-specific differences in terms of biological and clinical behavior [12][13][14].
BC in both sexes is likely to be caused by the concurrent effects of different risk factors, including advanced age, BC family history, increased levels of estrogens (i.e., estradiol), and environmental exposures to carcinogens [15][16][17].Clearly, the molecular and epidemiological determinants of MBC are not confounded by reproductive-history-related variables as in FBC [3,8].
Overall, it is estimated that 10-20% of BC cases are associated with hereditary factors [18].Exploration into the complex BC genetic predisposition might be facilitated more by MBC, unencumbered by the many confounding factors that make FBC a heterogeneous disease.Nevertheless, sex-specific differences in the impact and magnitude of risks conferred by BC genetic risk factors are emerging, and they should be taken into consideration for a proper personalized clinical management.
This review covers all the main genes and genetic variants associated with BC risk, highlighting differences and similarities between BC genetic predisposition in the two sexes, as well as addressing the implications, challenges, and open issues of establishing a gender-oriented BC clinical management.

Methods
To summarize and describe the latest research in the field of BC genetic predisposition with particular focus on similarities and differences in male and female BC, we conducted a systematic literature search by using PubMed (http://www.ncbi.nlm.nih.gov/pubmed/accessed up to 20 December 2023).We selected original articles and reviews in English, published up to December 2023.The following search key words were used to query the PubMed website: 'breast cancer and gender', 'breast cancer and genetic predisposition', 'breast cancer and BRCA', 'male breast cancer and genetic predisposition', 'male breast cancer and BRCA', 'breast cancer genetic predisposition and transgender', and 'breast cancer management'.The abstracts resulting from these queries were individually assessed, and inclusion in the review was evaluated based on relevance.Exclusion criteria were papers written in non-English languages, case-reports, low-quality studies, and articles evaluated as out of the review's scope.

The Genetic Architecture of BC Predisposition
The genetic architecture of BC predisposition may be explained by genetic risk factors that, based on their frequency and the magnitude of their impact on BC susceptibility, can be classified as high-, moderate-, and low-penetrance.
About 30 years ago, linkage analyses performed in families affected by multiple cases of FBC allowed for the identification of BRCA1, the first high-penetrance gene associated with BC susceptibility [19].Shortly after, BRCA2 was identified, analyzing families affected by both female and male BC [20].
Most cases of hereditary breast and ovarian cancer (HBOC) syndrome are linked to BRCA1 or BRCA2 genes.Germline pathogenic and likely pathogenic variants (herein called pathogenic variants, PVs) in these genes can be found in about 25% of families with HBOC [21].
The role of BRCA1 and BRCA2 PVs in BC susceptibility is significantly different in the two sexes, with BRCA1 mainly involved in females while BRCA2 in males.PVs in BRCA2 are often found in patients with MBC who have multiple cases of BC/ovarian cancer (OC) in their family, but they have also been found in patients with MBC without family history [3].Notably, a high percentage of BC patients without BRCA1/2 PVs was shown to have a positive family history of BC, suggesting the existence of other susceptibility factors [22].
Direct sequencing of candidate genes involved in BRCA1/2-associated DNA damage repair pathways led to the identification of other BC susceptibility genes, including PALB2, CHEK2, and ATM.In the last years, next-generation sequencing (NGS) analyses performed in BC by multigene panel testing have given the opportunity to identify PVs in a large number of candidate BC susceptibility genes, as well as to clarify their role and impact on BC predisposition [23,24].These genes were generally classified as moderate-penetrance genes since their PVs confer a smaller risk of BC than BRCA1/2 PVs.On the other hand, a small but increasing number of studies applied multigene panel testing to investigate additional genes associated with MBC predisposition [25][26][27][28][29][30][31][32].Collaborative studies are starting to provide reliable gender-specific cancer risk estimates for BC susceptibility genes [32][33][34].
BCs unaccounted by PVs in currently known BC susceptibility genes can be explained by the occurrence of low-penetrance risk variants [35].A polygenic model, in which many variants that confer low risk individually act in combination to confer much larger risk in the population, has been suggested for susceptibility to several types of cancer, including BC.This hypothesis has been confirmed for both female and male BC by international multigroup collaborations working in genome-wide association studies (GWAS) [36][37][38][39][40][41][42][43][44].Overall, the genetic architecture of BC predisposition is similar in both sexes; however, some differences in the impact of the risk conferred by specific genes have emerged (Table 1).

BRCA1
BRCA1, the first gene identified in 1994 as responsible for HBOC syndrome [19], is located on the long arm of chromosome 17 and encodes for an 1863 amino acid protein.This protein, expressed in a wide range of tissues, is critical in the DNA damage repair mechanism, cell cycle regulation, and other functions, such as transcriptional regulation and protein degradation by ubiquitination [55].
Germline PVs of BRCA1 are reported in 1-7% of FBC patients unselected for family history or age at onset [56].The largest available prospective study showed that women with germline BRCA1 PVs have a cumulative BC risk to age 80 years of 72%, compared with the 13% of the general female population [45].Risk assessment based on familial cases provided risk estimates higher than 80% [46], whereas recent population-based studies provided a lifetime risk estimate around 50% [23,24] (Table 1).
Prospective studies are not available for male BRCA1 PV carriers; however, recent risk assessment, based on pedigree data, showed that the cumulative BC risk at age 80 years is around 0.4% for male BRCA1 PV carriers, fourfold higher than the risk of 0.1% reported for the general male population [34].Overall, compared with FBC, BRCA1 PVs are quite rare in MBC cases, accounting for up to 4% of unselected MBC cases, but are more frequent in specific populations in which a founder effect is known to occur, representing 10-16% of MBC cases [46,57].
A large number of loss-of-function PVs, such as nonsense, small insertions or deletions, or splice or large rearrangements, occurring in functional domains are described along the whole BRCA1 gene and three BC cluster regions (BCCRs), located at c.179 to c.505, c.4328 to c.4945, and c. 5261 to c.5563, were identified [58].Large-scale rearrangements, including insertions, deletions, or duplications of more than 500 kb of DNA, have been also identified in BC cases [59][60][61].
Specific BRCA1 PVs show high frequency in specific countries or ethnic groups, particularly in genetically isolated populations, and are in part responsible for the variability in BC incidence rates among countries [62].For example, the founder BRCA1 PVs c.68_69delAG and c.5266dupC in the Ashkenazi Jewish female and male BC patients account for a significant portion of all familial BC in this population [63,64], and several other founder BRCA1 PVs have been observed in different populations worldwide [65,66].
It is now well established that BRCA1-related FBCs represent a subgroup of tumors characterized by a peculiar phenotype characterized by lack of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) and defined as triple negative BC (TNBC) [67]; in addition, these tumors are more frequently of high grade compared with sporadic tumors [68].
By contrast, the phenotypic correlation between BRCA1 PVs and TNBC observed in women was not observed in men.It was shown that MBCs with germline PVs in the BRCA1 gene were more likely to be ER positive; PR positive; and, overall, non-TNBC compared with BRCA1 FBCs [12].Drivers of hypoxia and related proteins associated with BRCA1 FBCs were shown to have a marginal role in MBCs [69].
In addition to BC, BRCA1 is a well-established OC susceptibility gene, with a cumulative risk to age 80 years of 44% for female BRCA1 PV carriers [45]; moreover, BRCA1 PVs were recently associated with a twofold increased risk of pancreatic and stomach cancers in both sexes [34].

BRCA2
The BRCA2 gene, identified soon after BRCA1 in 1995, is located on the long arm of chromosome 13 and encodes for a 3418 amino acid protein [20].This protein is expressed in response to cell proliferation, and its expression is initiated before DNA synthesis [70].
Both BRCA1 and BRCA2 are involved in maintaining genome integrity by engaging in DNA repair, cell cycle checkpoint control, and the regulation of key mitotic or cell division steps [55].While BRCA1 is a pleiotropic DNA damage response protein that functions in both checkpoint activation and DNA repair, BRCA2 is a mediator of the core mechanism of homologous recombination (HR) [70].
Germline PVs of BRCA2 are reported in 1-3% of FBC patients unselected for family history or age at onset [56].The largest available prospective study showed that women with germline BRCA1 PVs have a cumulative BC risk to age 80 years of 69% [45], whereas recent population-based studies provided a lifetime risk estimate around 50%, similar to the risk reported for BRCA1 PVs [23,24] (Table 1).
Initial studies reported that the majority (81%) of the HBOC families were due to BRCA1 PVs; conversely, most families with both male and female BCs were due to BRCA2 (76%) [46].Indeed, inherited PVs in BRCA2 are the strongest genetic risk factor for MBC [71].The estimated lifetime risk of MBC in BRCA2 PV carriers is about 4%, more than 40-fold higher than the risk of 0.1% reported in the general male population [34].Notably, male BRCA2 PVs carriers are significantly more likely to develop cancer, particularly BC, and are at increased risk of developing second breast and non-breast tumors, compared to male BRCA1 PVs carriers [72].
As in BRCA1, loss-of-function BRCA2 PVs are identified along the whole gene, although several putative BCCRs have been defined, particularly at the 3 ′ end of the gene [58].Like BRCA1, specific founder BRCA2 PVs are also present in genetically isolated population groups, as for example, the Ashkenazi Jewish founder BRCA2 c.5964delT [62].In Icelanders, the predominant BRCA2 c.771_775del accounts for a high proportion of BC families and was detected in up to 40% of MBC cases [73].Overall, in high-risk families, or in populations in which a founder effect was observed, PVs in the BRCA2 gene are estimated to be responsible for 60-76% of MBCs [74].Interestingly, large genomic rearrangements in BRCA2 are more frequent in families with MBC [61,[75][76][77], and, on the other hand, BRCA2 rearrangements seem to be infrequent in MBC cases unselected for family history [78].
It has been shown that FBC associated with BRCA2 PVs are more similar to sporadic tumors, exhibiting a luminal phenotype characterized by ER and PR overexpression, and they are often HER2 negative [67,79].By contrast, MBCs associated with BRCA2 PVs display specific pathologic features suggestive of an aggressive phenotype, such as higher histologic grade, compared both with FBC in BRCA2 PV carriers and with MBC in the general population [12,80,81].In particular, high histologic grade breast tumors are more frequent among male BRCA2 PV carriers diagnosed at younger ages (<50 years) than among those diagnosed at older ages [12].Moreover, BRCA2-associated MBCs displayed a higher TNM status, an over-representation of invasive micropapillary, and a lower representation of lobular morphologies, compared with BRCA2-associated FBCs [81,82].
Overall, BRCA2 has been associated with a more heterogeneous cancer spectrum, compared with BRCA1.Similar to BRCA1, BRCA2 is also an OC susceptibility gene, with a cumulative risk to age 80 years of 17% for female BRCA2 PV carriers [45].Moreover, BRCA2 PVs were associated with three-to fourfold increased risks of pancreatic and stomach cancers in both sexes [34], as well as with a twofold increased risk of prostate cancer, with a cumulative risk to age 80 years of 27% for male BRCA2 PV carriers [34].

PALB2
PALB2 can be considered as the third most important gene, following BRCA1 and BRCA2, in terms of BC susceptibility.PALB2 is located on the short arm of chromosome 16 and encodes for a 1186 amino acid protein.PALB2 protein has a large number of interactions with other DNA damage response proteins that function in DNA repair by HR [83].
PALB2 PVs were initially found in 1% of families with BCs and were found to be associated with a twofold FBC risk [84,85].Subsequent pedigree-based investigations showed that PALB2 PVs were associated with a sevenfold increased FBC risk and a lifetime risk of about 50% (Table 1) [47].At the population level, PALB2 PVs showed an increased risk of about fourfold, whereas among familial FBC cases at about eightfold [23].For male PALB2 PV carriers, a sevenfold increased risk of BC and a cumulative BC risk of 1% to age 80 years were estimated [32,47].In addition to BC, PALB2 PVs were associated with two-to threefold increased risks of OC in women and pancreatic cancer in both sexes [47].
As expected, studies enriched for high-risk MBC cases (i.e., cases with bilateral BC, and/or early onset BC, and/or a positive family history of BC) showed a higher PALB2 PV frequency.Overall, a higher frequency of PALB2 PVs in high-risk MBC cases than in high-risk FBC cases was observed (4% vs. 1%) [93].
Moreover, PALB2 PVs were frequently observed in families with history of cancers other than BC/OC, including melanoma, pancreatic, prostate, and stomach cancers, suggesting that PALB2-related families may resemble BRCA2-like families, in which MBC and several other cancers may be found in addition to FBC [87,90,91,93,97,98].
There is evidence that PALB2-associated FBCs may display peculiar pathological features, including TNBC status [24].However, no evidence of such association has emerged for MBC yet.

CHEK2
The CHEK2 gene is located on the long arm of chromosome 22 and encodes for a 543 amino acid protein.CHEK2 protein is a tumor-suppressive serine/threonine kinase that is involved in cell cycle progression and DNA structure modification, and it is part of the DNA damage response system activated by genotoxic stress [99].
The protein truncating variant CHEK2 c.1100delC was the first BC genetic risk factor identified after BRCA genes [100,101].At population level, CHEK2 c.1100delC showed an increased risk of about 2.5-fold, whereas among familial FBC cases was about 4.8-fold [23, 24,102,103].
The CHEK2 c.1100delC has been initially shown to confer approximately a 10-fold increase in BC risk in men, and it was estimated to account for 9% of familial high-risk MBC cases [100].However, those results were not replicated in all populations or in unselected cases [27,32].
Notably, the contribution of the CHEK2 c.1100delC PV to BC predisposition in both sexes varies by ethnic group and from country to country, with a founder effect in North-Eastern Europe, and a decreased frequency in North to South orientation [78,100,[104][105][106][107][108].
Both single-gene direct sequencing and the introduction of NGS multigene panels have enabled the identification of additional PVs in the CHEK2 gene, mainly missense variants.Overall, the most recent estimates confirmed that protein truncating variants in CHEK2, including the c.1100delC, are associated with an approximately 2.5-fold FBC risk, whereas CHEK2 pathogenic missense variants are shown to confer a small increase in FBC risk, below twofold (Table 1) [23,24].Data on CHEK2 PVs besides the c.1100delC in MBC cases are still scarce.In recent studies, CHEK2 PV prevalence ranged from 0.4% to 4.1%, and risk estimates ranged from 2.43 to 3.78, based on the population analyzed [25][26][27]29,31,32].Our recent Italian case-control study showed no association between CHEK2 PVs and increased MBC risk, confirming the limited role of CHEK2 PVs in the Italian population in both sexes [32].
Female CHEK2 PV carriers were shown to be more strongly predisposed to ER-positive BCs [23, 24,109].In other reports, CHEK2 PVs were associated with all BC subtypes except for TNBC [24,110].
Although CHEK2 PVs were observed in a wide range of cancer types, there are no reliable additional cancer risks associated with CHEK2 PVs, besides BC.Evidence for association with increased risk of colorectal and prostate cancers is emerging [111,112].

ATM
The ATM gene is located on the long arm of chromosome 11 and encodes for a 3056 amino acid protein.This protein is mainly involved in cell cycle regulation and DNA damage recognition and repair [113].
ATM is involved in ataxia telangiectasia, a rare disease inherited in an autosomal recessive pattern.Individuals homozygous or compound heterozygous for a germline ATM PV develop ataxia telangiectasia.Heterozygous PVs in ATM are relatively common in the population, with a prevalence of about 0.35%, and they are frequently observed in cancer patients; however, the magnitude of cancer risk remains uncertain [33].Many years ago, the seminal study of Swift and colleagues, investigating an excess risk of cancer in a series of 110 ataxia-telangiectasia families, suggested that the relative risk of cancer incidence doubled in men and tripled in women, with BC being the most associated cancer [48].
Studies investigating the role of ATM in FBC reported a PV prevalence ranging from 0.6% in population-based studies to 2.7% in studies enriched for familial cases.Estimates of BC risk in women heterozygous for germline ATM PVs ranged from a two-to fivefold increased risk, compared to women without ATM PVs [33,[48][49][50][51][52][53][54].Population-based risk estimates ranged from two-to threefold, with a cumulative lifetime risk of about 20-60% (Table 1) [50].
In MBC, heterozygous ATM PVs were found with a frequency ranging from 0.5% to 1.96% depending on the population analyzed [25][26][27]32,114].Although larger studies are needed to better estimate the BC risk in men with ATM PVs, recent studies suggested that ATM may be considered as a possible risk gene also in MBC susceptibility with up to about fourfold increased risk [32,33].
Recently, ATM PVs were also associated with moderate-to-high risks (two-to fourfold) of pancreatic, prostatic, and gastric cancers, as well as with low-to-moderate risks (<2-fold) for OC, colorectal cancer, and melanoma [33].

Other Genes
The involvement of BRCA1 and BRCA2 in the HR pathway promoted mutation screening of other DNA repair genes functionally linked to these two genes [115].Most of these candidate BC genes have been included in multigene panels used for BC patients of both sexes.Overall, to date, BRCA1, BRCA2, PALB2, CHEK2, and ATM are the only well-established BC risk genes at the population level [18,116,117].However, PVs in other genes may also increase the risk of BC, particularly for specific subtypes.
PVs in FANCM, a gene encoding a protein involved in the Fanconi anemia (FA) molecular pathway, have been associated with increased BC risk [118,119].Specifically, FANCM PVs have been associated with a 2-4-fold increased BC risk in case-control studies conducted in different European populations [120,121].Overall, a high frequency of proteintruncating variants in the FANCM gene (about 2%) was identified in BRCA1/2-negative BCs [122].
Notably, recent evidence has shown peculiar genotype-phenotype correlations for FANCM, as PVs seem to be moderate risk factors for ER-negative and TNBC.In particular, PVs within the more 5 ′ region of the FANCM gene may have a stronger effect on the risk for ER-negative BC.Conversely, the risk effects of FANCM PVs within the more 3 ′ region are probably lower [123].
FANCM PVs were also found in MBC at a frequency of 0.5%, which was raised up to 1% when only cases at increased genetic risk for BC were considered; however, risk estimates are not available yet [27,32,124].
PVs in OC risk genes, proposed also as candidate BC predisposing genes, such as BARD1, RAD51C, and RAD51D, were associated with moderate risk of ER-negative BC in women [23,125].At present, there is no evidence that BARD1, RAD51C, and RAD51D PVs may contribute to BC susceptibility in men, probably due to the low frequency of the ER-negative subtype in MBC [25,27,28,32,126].
PVs in the BRIP1 gene were originally associated with low-penetrance BC risk [127]; however, some studies indicated that BRIP1 PV carriers have a higher risk for postmenopausal OC rather than BC [128][129][130].Consistently, no evidence was found that germline variants in BRIP1 might contribute to MBC predisposition [25,27,28,32,131].
Other genes initially linked to BC susceptibility, but whose association was not replicated in recent large case-control studies, include NBN, BLM, GEN1, FA genes other than FANCM, mismatch repair genes, and RAD50, as well as genes identified by whole exome sequencing analysis, such as RECQL, RINT1, and XRCC2 [18,23,24,132,133].

Syndromic Genes
PVs in genes associated with hereditary multi-cancer syndromes have been implicated in hereditary BC, although they accounted for a very small percentage of BC cases in both sexes.
High-penetrance PVs in TP53, PTEN, and STK11 genes, involved in Li-Fraumeni, Cowden, and Peutz-Jeghers syndromes, respectively, are known to confer a high risk of childhood and adult cancers, including BC, with lifetime risks ranging from 40 to 80% [134][135][136][137][138], although these estimates may be overestimated due to the rarity of the conditions.
PVs in CDH1, the hereditary diffuse gastric cancer gene, have been associated with increased BC risk, particularly of lobular histology and ER-positive status [139].
In addition, PVs in NF1, the gene associated with neurofibromatosis 1, may also moderately increase BC risk both in women and men [27,140].
Overall, it is unlikely that these PVs would account for a relevant proportion of BC in the absence of their respective syndromes [141,142].Recent studies confirmed that PVs in these syndromic genes are rarely detected by multigene panel testing performed at the population level due to the rarity of the variants, early single-gene testing at younger age, and/or early death.Although these genes, particularly TP53, are considered bona fide BC risk genes, the debate on whether to include these genes in clinical multigene panels for use in BC patients is still open [18,143].
Notably, the relative risk associated with several of the loci identified shows BC subtype specificity, specifically defined by hormonal receptors status [150,153].Associations with most of the susceptibility loci are stronger for ER-positive rather than for ER-negative BCs, probably because the majority of BC cases are ER positive [151,154].Studies focused on ER-negative BC and TNBC identified about 20 specific loci associated with ER-negative disease, accounting for approximately 14% of familial relative risk for ER-negative BC [150,155].
A few studies addressed the role of low-penetrance variants in MBC susceptibility through GWAS or a candidate SNP genotyping approach [41,42,44,[156][157][158][159][160][161].Overall, five loci were associated with increased MBC risk.These loci were also associated with increased FBC; however, they seem to confer greater risks of BC in men than in women [41,44].
Although the relative risk associated with SNPs is low, they are likely to be responsible for a substantial percentage of hereditary and sporadic BCs due to their high frequency at the population level and to polygenic effect [35].When summarizing all SNPs in a polygenic risk score (PRS), the cumulative risk can be substantial.Risk profiling based on a combined SNP effect can identify individuals at substantially increased or reduced BC risk, providing the basis for targeted prevention.Thus, the PRS can aid in stratifying patients into different risk categories of developing BC [169][170][171][172].
Recently, a PRS including 313 risk SNPs has been constructed and validated for the prediction of BC risk in women of European ancestry, showing that women with high PRS have the same risk as ten-year-older women with average PRS [169].
PRS models developed based on associations from FBC GWAS were also evaluated in MBC risk [44].The recently developed 313-SNPs PRS, identified in FBC [169], was also associated with MBC risk, and its distribution in male cases was similar to that of FBC cases, suggesting a shared genetic architecture of BC in the two sexes [44].
The 313-SNP PRS was associated with both female and male BC risk in BRCA PV carriers, suggesting that risk profiling on the basis of PRS may provide a further individual cancer risk stratification for carriers of BRCA1/2 PVs, with implications for their clinical management [173,174].Notably, distribution in male BRCA PV carriers was similar to female BRCA PV carriers but lower than in cases from the general population, possibly reflecting a general attenuation of the effect sizes of common variants on genetic risk in the presence of a PV in a high-risk gene.
It was recently demonstrated that PRS created meaningful risk gradients among female carriers of PVs in cancer-predisposing genes other than BRCA, including ATM, CHEK2, PALB2, BARD1, BRIP1, CDH1, and NF1.In particular, PRS may help differentiate BC risk among carriers of PVs in well-established moderate-penetrance genes, such as CHEK2 and ATM, enabling more informed decisions about screening practices and more personalized risk management approaches [175].Currently, no data on the possible joint effect of PVs in moderate-penetrance BC predisposition genes and PRS with MBC are available.

Implication of Gender-Oriented Genetic Testing and BC Management 4.1. Preventative Strategies
Individuals with suspected BC predisposition should be offered a genetic screening to identify PVs in established HBOC genes, mainly BRCA1/2 [176,177].
However, currently, genetic testing remains somewhat restricted for BC patients.Based on referral criteria from genetic testing from the current available guidelines, women with TNBC, bilateral disease, or young-onset disease might be offered a genetic test at diagnosis, but most will be offered testing only if they also have a family history of the disease [177].
On the other hand, the National Comprehensive Cancer Network (NCCN) recommendations indicate that all men diagnosed with BC should be routinely screened at least for BRCA1 and BRCA2 PVs, regardless of age or family history, which could prove to contribute invaluable genetic information to unaffected family members [177].
Nevertheless, the assessment of the a priori probability of identifying a PV is an important component of pre-test counseling.Risk assessment models to estimate the risk of carrying a BRCA PV, such as BRCAPRO, have been validated for use in patients of both sexes [178][179][180].
Both men and women have the same risk of inheriting a BRCA PV, but men are ten times less likely to get tested [181].However, more than half of male BRCA PV carriers reported that they underwent genotyping for the sake of their children or family, rather than to learn about their own cancer risk [182].In most genotyping programs, less than 10% of the individuals tested are men [183].
It has been reported that male BRCA1/2 PV carriers may be under-informed about their personal cancer risk [184], thus suggesting the need of increasing awareness of possible risks of cancers, including not only BC but also common male cancers like prostate cancer, as well as rare but lethal cancers such as pancreatic and stomach cancers, for male BRCA PV carriers [34].
In this context, it was recently proposed to rename the HBOC syndrome with a name perceived as more gender-neutral and inclusive of the broad cancer spectrum observed in BRCA1/2 PV carriers [185].
Through cascade testing within high-risk families, genetic counseling and testing should be offered to healthy women and men.PVs have a large effect on lifetime risk of BC and their identification enables personalized surveillance or even risk-reducing strategies therapy for high-risk individuals (Table 2).
Table 2. BC management for female and male carriers of PVs in BC risk genes involved in both sexes.

Female PV Carriers
Male PV Carriers

BRCA1/ BRCA2
Education regarding signs and symptoms of cancer(s), especially those associated with BRCA PVs.Women with germline PVs in BRCA genes may opt to an enhanced BC screening or undergo prophylactic bilateral mastectomy; primary chemoprophylaxis with tamoxifen or other selective ER modulators has also been recommended [177].Thus far, the management guidelines for men harboring BRCA PVs are still often based on low-level evidence and/or expert opinion [177,186,187].
For men carrying BRCA PVs, the management plan involves starting breast self-exam training and education at the age of 35.As part of this approach, clinical breast exams are recommended every 12 months, starting from the age of 35.Recent studies have shown that mammography may be beneficial in selective screening of men at high risk for developing BC depicting clinically occult early stage malignancy in this population [188][189][190].Based on this evidence, it is suggested to consider an annual mammogram for men with BRCA2 PVs.The ideal starting age for mammograms is either 50 years or 10 years before the earliest known occurrence of MBC in the family.
Once individuals reach the age of 40, it is recommended to start prostate cancer screening for BRCA2 carriers.For those with BRCA1 PVs, it is advised to consider prostate cancer screening as well.Additionally, there should be contemplation of pancreatic cancer screening from the age of 50 (or 10 years younger than the earliest exocrine pancreatic cancer diagnosis in the family, whichever comes earlier).This is especially relevant for individuals with exocrine pancreatic cancer in ≥1 first-or second-degree relatives from the same side of the family (or presumed to be from the same side of the family) as the identified PVs [177].MBC cases found to carry BRCA1/2 PVs are at an increased risk for developing contralateral breast and non-breast second malignancies [34,72], highlighting the need of a specific surveillance scheme for these men [191].Notably, more than half of male BRCA PV carriers did not adhere to the screening guidelines recommended after disclosure of genetic test results.For example, only 25% of male BRCA2 PV carriers reported to have annual clinical breast examination and/or perform breast self-examination [182].
In the last years, gene panels evaluating simultaneously a variable number of BCassociated genes or multiple cancer-associated genes have begun to be routinely used in clinical practice for familial BC cases, including male patients, although for most of the genes included in the multigene panels, robust evidence of association with BC risk are currently unavailable.Recommended guidelines for early detection and cancer risk reduction in women with PVs in moderate-penetrance risk genes increasing lifetime cumulative risk over 20%, including PALB2, ATM, and CHEK2, are starting to become available.This is not the case for male carriers of PVs in the same genes.The recent MBC risk estimates provided for PALB2 have been instrumental in establishing guidelines for the clinical management of male carriers of PALB2 PVs [32,34,47].PALB2 risk estimates for MBC are comparable, if not higher, to those recently reported for BRCA1 [34].In light of these results, the most recent NCCN guidelines suggested that it is reasonable to consider for male PALB2 PV carriers a BC screening similar to that for male carriers of BRCA1 PVs [177].
Notably, the latest NCCN Guidelines includes a new section for transgender and gender diverse people who have a hereditary predisposition to cancer, including BC [177].Overall, given the lack of prospective data, recommendations on appropriate cancer risk reduction and/or screening options must be made on a case-by-case basis.
Regarding transgender men, gender-affirming hormone therapy with testosterone might alter the risk of BC in individuals with a hereditary susceptibility to BC, but data are still limited.Transgender men with a germline PV in a BC gene may want to consider risk-reducing mastectomy instead of gender-affirming breast surgery (top surgery), which typically retains some breast tissue and the nipple areolar complex [192].For transgender men with a PV in a BC gene who have had top surgery, or no breast surgery, BC screening is recommended to begin at an earlier age and may include mammography and breast MRI.For individuals with a personal or family history suggestive of hereditary BC, it is recommended that genetic testing be performed prior to breast surgery to inform the type of surgery [177].
Regarding transgender women, gender-affirming hormone therapy with estrogens and anti-androgens increases breast tissue and may increase BC risk.Nonetheless, such therapy is not contraindicated, even in the presence of a PV in a BC gene.While there are limited data on BC surveillance in transgender women, NCCN guidelines suggest a BC screening like that for cisgender males at increased hereditary risk [177].
Overall, less than 10% of all BC cases are attributable to the monogenic causes sought in clinical practice.Women with familial BC for whom genetic testing did not detect a PV should be offered a personalized risk assessment based on their detailed family history, demographic risk factors, and polygenic risk, using risk assessment tools such as CanRisk [193].
Risk stratification based on PRS is about to enter clinical practice in order to further improve screening and prevention strategies for all women [194,195].Thus, PRSs are becoming indispensable in identifying high-risk individuals who could benefit, with greater advantages, from targeted strategies according to current clinical guidelines.An integrated model, implemented in risk prediction models and including classical risk factors, family history, and PRS, would provide the highest level of risk stratification [170].
Clinical implementation of PRS may be also particularly useful to stratify risk in cohorts where there is a higher prior probability of disease, for example in carriers of PVs in BC risk genes, to personalize cancer screening.For example, the NCCN recommends magnetic resonance imaging screening for women with a lifetime BC risk > 20% [177].
Female BRCA1 and BRCA2 PV carriers are above the 20% threshold, and thus PRS information is unlikely to change clinical recommendations for these women.On the other hand, incorporating PRS into BC risk estimation may help identify 30% of CHEK2 and nearly half of ATM carriers below the 20% lifetime risk threshold, suggesting the addition of PRS may prevent over screening and enable more personalized risk management approaches [175].
Because the age-standardized incidence of MBC is only 1/100,000 person-year with a lifetime risk of about 1/1000, there is no role for risk assessment and BC screening in the general male population.However, PRS may be applied to high-risk men, especially male BRCA PV carriers, which may benefit from a more refined stratification of the individual cancer risk to inform clinical management.Polygenic risk may identify male carriers of BRCA PVs, at both sufficiently reduced or increased risks of BC, in order to aid prevention and screening decisions [42,174].

Personalized Therapeutic Management
The development of NGS technologies has produced a large amount of genomic data in a wide variety of cancers, including BC.These data allowed for the identification of molecular alterations that are potential predictive biomarkers or therapeutic targets to guide personalized treatment in women with BC [196].
To date, approaches to treating men with BC have been extrapolated largely from research conducted in women with BC, although recent genomic and transcriptomic studies showed that MBCs and FBCs, including those associated with germline BRCA1/2 PVs, are different in the somatic landscape [71].
It is well established that FBC can be classified into molecular subtypes based on gene expression, with BRCA1-mutated tumors showing a prevalence in the basal-like subtype, while BRCA2-mutated FBC may show gene expression patterns that resemble those found in luminal epithelial cells [205][206][207].Recent transcriptome data on MBC indicate that germline mutational status could impact also on MBC transcriptome profiles, defining subgroups that may be driven by different underlying molecular pathways [208].Specifically, male breast tumors arising in patients with germline PVs in BRCA and PALB2 were characterized by the activation of the cell cycle pathway, suggesting a possible use of CDK4/6 inhibitors in this setting.In addition, these tumors were characterized by a high HER2 score signaling, suggesting that they might benefit from treatment with trastuzumab [209,210].
Overall, MBC patients show a worse prognosis when compared with FBC patients [11].Significantly reduced survival rates were also registered in the subgroup of MBC patients with BRCA1/2 PVs, while survival rates of FBC patients do not seem to be largely affected by BRCA1/2 PVs [68,211,212].This disparity in prognosis and survival between male and female BC patients may be due to factors yet to be identified and may reflect the lack of specific management strategies in MBC.
The management of hormone-receptor-positive MBC aligns with treatment modalities utilized in female counterparts.Aromatase inhibitor (AI) combined with gonadotropinreleasing hormone agonist (GnRH-a) has become a valid treatment in FBC.However, the available data on the efficacy of this treatment in men is currently confined to case series, and there is an absence of comparative data on their efficacy [213,214].
Based on real-world data and limited studies, it was considered reasonable to extrapolate the use of additional treatment options for men with BC.These options include CDK4/6 inhibitors, mTOR inhibitors, and PI3K inhibitors, employed in conjunction with endocrine therapy.Also, in the context of metastatic BC, the use of chemotherapy, HER2targeted therapy, and immunotherapy in men is currently guided by treatment principles analogous to those applied in women [9,215].
In the adjuvant setting with a high risk of recurrence as well as in the metastatic setting, PARPi are approved for heterozygous germline or somatic BRCA PV carriers with HER2-negative BC [216].PVs in PALB2, CHEK2, and ATM and genomic instability scores have also been tested as additional predictive biomarkers of increased sensitivity to PARPi treatment [217].Data suggest that BC associated with PALB2 PVs are highly sensitive to PARPi, while no responses were observed with CHEK2 or ATM PVs.The potential effectiveness of PARPi is demonstrated also in other types of BRCA-associated tumors including prostate cancer and OC [218][219][220][221][222].However, little is known about the use of PARPi for the optimal management of BC in men, thus further highlighting the need to extend clinical trials to male patients, as recently suggested by the Food and Drug Administration (FDA) [9,223,224].
Immunotherapy, a promising therapeutic strategy, initially used to treat metastatic BCs and TNBCs, has a limited use in BC in general because it is considered an immuno-logically 'cold' tumor [225].Notably, higher tumor-infiltrating lymphocyte levels have been reported in BRCA-associated BCs compared with sporadic BCs [226].It was recently shown that BRCA2 PVs may affect the tumor microenvironment differently than BRCA1 PVs, and that BRCA2-associated tumors might respond better to checkpoint blockade immunotherapy [227].Immune response recently emerged as the most relevant process able to discriminate MBC subgroups at the transcriptional level, particularly in BRCA-associated MBCs [208], opening for further investigation of immune-related features in a subgroup of male breast tumors.

Open Challenges and Future Directions
Current data suggest that the genetic architecture of BC predisposition, including high-, moderate-, and low-penetrance variants, is similar in both sexes; however, differences in the impact of the risk conferred by specific genetic factors, including PVs in BRCA genes, in PALB2, and in SNPs, emerged.These differences suggest that the heritable influence on BC susceptibility may be context dependent, perhaps influenced by non-genetic factors present in the breast microenvironment, including hormones, that may differently induce tumors in a mutant background [228].However, the molecular basis of gender-specific genetic susceptibility of BC is still unknown and deserves to be investigated.
Although FBC has been the guide for MBC research to date, exploration into complex biological processes and pathogenetic mechanisms of BC might be facilitated by MBC, being rare and unencumbered by the many confounding factors that exist in FBC.These MBC characteristics offer a unique opportunity to uncover many of the interrelationships that are inherent to the disease in both genders.MBC may help uncover underlying features of BC genetics in general, as well as those shared with other BRCA-associated cancers, including prostate and pancreatic cancers.
In this context, the need for large-sample-size studies is quite compelling in MBC, a rare disease.To date, international studies on BC have put special efforts to include and analyze also male patients, underlining important discoveries in terms of genetic susceptibility and risk assessment, as well as making the expansion of the analysis to larger independent case series promising [174].For this purpose, the large international Confluence project (https://confluence.cancer.gov,accessed on 20 December 2023) has been established, with the aim to study BC genetic susceptibility in women and men of multiple ancestries by integrating existing and new genome-wide genetic data across several BC consortia, including the BC Association Consortium (BCAC), the Consortium of Investigators of Modifiers of BrcA1/2 (CIMBA), and the newly established MBC genetics consortium (MERGE).
To provide a proper gender-oriented BC management, we need to address the peculiar needs of transgender and gender diverse individuals, particularly those at increased BC risk due to hereditary PVs.To date, limited scientific literature and recommendations exist on specific medical management strategies for high-risk transgender individuals [177].The identification of a PV in a BC susceptibility gene may impact a transgender person's decisions regarding hormonal and/or surgical transition; however, the effect of genderaffirming treatments on BC risk remains largely unexplored [229,230].Thus, there is a need for additional research to elucidate the impact of these treatments and their interactions with BC hereditary susceptibility.Furthermore, exploring optimal preventative strategies tailored to these specific populations is essential [231].

Conclusions
Implementation in the clinical practice is the final phase of the investigation of BC susceptibility.As personalized approaches for genetic testing, risk assessment, and cancer screening are ready to enter clinical practice, current practice presents still unresolved questions and challenges, including the use of referral criteria for testing vs. population screening in women with BC, the choice of the genes to be included in gene panels, the implementation of PRS in cancer risk assessment models, the risk of tumors other than BC, germline vs. tumor testing, and the clinical management options for PV carriers [232,233].In addition to this, evaluating the genetic component of BC from the perspective of gender medicine is mandatory to successfully implement personalized strategies.Moreover, the characterization of both germline and somatic genomic landscape may concur to establish, with greater precision, which BC patients of both sexes can benefit from targeted therapeutic strategies.Improving individualized preventative and therapeutic strategies with up-todate genomics tools will ensure the centrality of each individual in innovative BC precision management, taking into consideration gender-specific characteristics.Overall, addressing all the challenges and the open issues regarding BC genetic predisposition will have an important clinical impact on the management of all patients towards reaching gender equality in BC care delivery.

Table 1 .
BC risks conferred by PVs in established BC susceptibility genes in both sexes.Estimates may vary according to study population and/or study design.