The Fundamental Role of BARD1 Mutations and Their Applications as a Prognostic Biomarker for Cancer Treatment

Abstract

BRCA1-associated RING domain 1 (BARD1) constitutes a heterodimeric complex with BRAC1 that triggers several essential biological functions that regulate gene transcription and DNA double-stranded break repair mechanism. BARD1 gene was discovered in 1996 to interact with BRCA1 directly and encodes a 777-aa protein. Interestingly, the BARD1 has a dual role in breast cancer development and progression. It acts as a tumor suppressor and oncogene; therefore, it is included on panels of clinical genes as a prognostic marker. Structurally, BARD1 has homologous domains to BRCA1 that aid their heterodimer interaction to inhibit the progression of different cancers, including breast and ovarian cancers. In addition to the BRCA1-independent pathway, other pathways are involved in tumor suppression, such as the TP53-dependent apoptotic signaling pathway. However, there are abundant BARD1 isoforms that are different from full-length BARD1 due to nonsense and frameshift mutations and deletions associated with susceptibility to cancer, such as neuroblastoma, lung cancer, cervical cancer, and breast cancer. In the current chapter, we shed light on the spectrum of BARD1 full-length genes and isoform mutations and their associated risk with breast cancer. The chapter also highlights the role of BARD1 as an oncogene in breast cancer patients and its uses as a prognostic biomarker for cancer susceptibility testing and treatment

“Our genomes carry the story of evolution, written in DNA, the language of molecular genetics, and the narrative is unmistakable.”

Kenneth R. Miller

July 14, 1948

1. Introduction

1.1 A glance on The BRAC1/BARD1

In recent decades, cell biology and molecular genetics have revolutionized our understanding of cancer in general and breast cancer in particular. In this book, we focused on the BRCA1 and BRCA2 mutations. BRCA1-associated RING domain 1 (BARD1) is the name of a protein that Wu et al. in 1996 found as a BRCA1 (BReast CAncer type 1) binding partner [1]. Here, we shifted our focus to the BARD1 as a potential prognostic biomarker for breast cancer.

Generally, the BRAC1/BARD1 constitutes a heterodimeric complex that mediates numerous fundamental biological functions, specifically in regulating gene transcription and DNA double-stranded break repair mechanism [2, 3]. Furthermore, BRCA1 and its partner BARD1 protein are essential in other cellular processes involving chromatin remodeling, telomere regulation, replication fork maintenance, cell cycle progression, apoptosis, and tumor inhibition [4]. BRAC1/BARD1 possesses an enzymatic activity through its E3 ubiquitin ligase capacity that assists in regulating the biological processes and controlling the activity and transcription of other protein complexes [2]. BRAC1/BARD1 acts as a nucleosome reader and writer to scan and correct the DNA breaks by following the homologous recombination pathway. The C-terminal domain of BARD1 serves as a reader/scanner player, while the N-terminal domain exhibits the writer/corrector capacity. Both domains interact with a nucleosome in a wrapping fashion, activating the Ub ligase function [2]. One study has identified a negative regulator of BARD1, DCAF8L2 (a DDB1-Cullin-associated factor (DCAF) associated with CRL4 E3 ligase). The interaction between BARD1 and DCAF8L2 resulted in the degradation and ubiquitination of BARD1 with subsequent disassembly and uncoupling of the BRCA1/BARD1 complex. Additionally, DCAF8L2 expression was upregulated in breast cancer cells suggesting an oncogenic function via disrupting the BRCA1/BARD1 complex stability [3]. The BARD1 gene plays two distinct roles in cancer progression, specifically breast cancer. Thus, several biological researchers have made important discoveries about the BARD1 gene’s role in cancer evolution and its potential applications as a prognostic biomarker for breast tumors, or at the very least, to consider it a possible candidate for targeted breast cancer therapy [5].

1.2 BARD1 structure, locations, and isoforms

The human BARD1 gene has 11 exons that code for a 777 aa protein with a molecular weight of 87 kDa. BARD1 was discovered in 1996 and reported to interact with BRCA1 directly through their homologous N-terminal RING domains n chromosome 2 (2q34–35) [6]. Bard1 protein is structurally made up of a RING-finger domain at the N-terminal region, three repeating Ankyrin (ANK) domains in between, and two tandems of BRCA1 domains at the C-terminal area (BRCT) [6]. Interestingly, the BRCT repeats are essential for controlling how other partners’ proteins interact with one another in a phosphorylation-based manner. These interactor proteins are crucial to mediate crucial cellular processes, including DNA damage checkpoints, DNA repair machinery, and cell cycle regulation [7, 8]. Notably, the RING-finger domain and BRCT repeats are essential for the BRCA1-BARD1 complex’s ability to suppress cancer [9, 10]. The presence of several exomes in full-length BARD1 (FL-BARD1, resulted in different isoforms) Figure 1 represents a scaled diagram depicting the comparison of protein structures of BARD1, BRCA1, and BRCA2

Figure 1.

A scaled diagram depicting the comparison of protein structures of BARD1, BRCA1, and BRCA2. The diagram illustrates the different protein structures of BARD1, BRCA1, and BRCA2. BRCT (dark red), RING (green), and ankyrin (dark blue) domains. The putative nuclear localization signals (NLS, light blue) and the nuclear export signal (NES, yellow) are shown. The location of BARD1’s third NLS, at amino acid residue 321, is crucial for nuclear localization. With eight copies of a 70 amino acid motif known as the BRC repeats and a conserved transactivation domain (TD), BRCA2 is entirely unrelated to either BARD1 or BRCA1.

There are many BARD1 isoforms with skipped exons and various molecular weights [11]. The isoforms are more frequently found in association with cancerous cells [12, 13]. For instance, isoform α has skipped exon 2. While the isoform β skipped exons 2 and 3, which causes the open reading frame (ORF) to a frameshift, resulting in the translation of shorter proteins (758 aa (85 kDa) and 680 aa (75 kDa), respectively. However, isoform γ is typically interrupted by the deletion of exon 4. Isoforms φ and δ skipped exons 2−6 and 3−6 to produce 326aa (37 kDa) and 307 aa (35 kDa) proteins, respectively. Isoform ε skipped exons 4−9, resulting in a protein with a molecular weight of 30 kDa (264 aa), while the skipping of exons 1, 10, and 11 leads to isoform η. Another splicing from exons 1 to 10 is worthwhile since it interrupts the ORF. As a result, additional alternative ORFs may host the translation’s start codon, producing a short protein with 167 amino acids (19 kDa). Surprisingly, most of these isoforms had agonistic cancer susceptibility potential because they lack the RING finger and ankyrin repeats, which are essential for the full-length BARD1’s tumor suppressor capabilities [14, 15]. Table 1 summarizes the different BARD1 isoforms.

No.	BARD1 isoform	Amino acid	Molecular weights	Corresponding exons skipping	Associated cancer phenotypes
1	isoform α	758 aa	85 kDa	lacks exon 2
2	β isoform	680 aa	75 kDa	lacks exons 2 and 3
3	isoform γ			exon four deletion
4	isoforms φ	326aa	37 kDa	missing exons 2–6	HeLa and ovarian cancer cells
5	isoforms δ	307 aa	35 kDa	missing exons 3–6	HeLa and ovarian cancer cells
6	isoform ε	264 aa	30 kDa	lack of exons 4–9
7	isoform η	167 aa	19 kDa	lack of exons 1, 10, and 11

Table 1.

Summarizes the different BARD1 isoforms.

Since we highlighted some important aspects of the BARD1 structure, it is worth highlighting the function of the Bard1 protein.

1.3 BARD1 as a tumor-suppressor gene and oncogene

The Bard1 protein performs a tumor suppressor function in BRCA1-dependent and -independent pathways. Due to their homologous domains, the BRCA1/BARD1 heterodimer can be configured by the N-terminal RING-finger domains, altering the ubiquitin ligase’s activity, which controls the cell cycle chromatin structure and hormone signaling pathways as well as DNA damage response pathways [16, 17]. BRCA1-BARD1 heterodimers are disrupted by mutations in cancer cells, which results in the degradation of both proteins [1].

Technology advancements have made it clear that numerous genes, including BRCA1/2 and BARD1, play a crucial role in hereditary and familial breast cancer and ovarian cancer [18, 19]. Research has identified BARD1’s function in the BRCA1-dependent pathway as an anti-breast cancer agent [20]. Because it activates ubiquitination through E3 ubiquitin ligase activity and starts the degradation process for the damaged proteins, the BARD1-BRCA1 complex is essential to the DNA damage machinery [16]. The involvement of BARD1 and BRCA1 in a homology-directed repair (HDR) of chromosomal breaks that clarifies their presence alongside RAD51 in response to DNA damage was previously discussed by Westermark et al. [21, 22, 23]. Furthermore, through a particular interaction with the poly (ADP-ribose), the BARD1 BRCT domain promotes the early recruitment of the BRCA1/BARD1 heterodimer to DNA damage sites (PAR) [24]. Studies have also demonstrated that disruptive mutations in the phosphate-binding pocket of the BARD1 BRCT domain in mice (S563F and K607A) hinder the recruitment of the BRCA1/BARD1 heterodimer to the stalled replication fork (SRF), which ultimately causes chromosomal instability [25]. Such mutations do not affect recruitment to HDR [25], contrasting with the comparable modification in BRCA1 BRCT (S1598F) [10]. Additionally, BARD1 or BRACA1 mutations linked to the prevalence of breast cancer, such as alterations in the RING finger domain, interfered with the BRCA1/BARD1 heterodimer interaction [26, 27], missense mutations [28, 29, 30], and ANK sequences that are involved in the regulation of transcription [31]. Additionally, the heterodimer prevents inappropriate mRNA polyadenylation at DNA repair sites with cleavage stimulation factor subunit 1 (CSTF1) [32, 33]. Through the ubiquitination pathway, BRCA1/BARD1 also aids in the prevention of tumor growth [34] and BRCA1’s subcellular location [35].

BARD1 also acts as a tumor suppressor in a BRCA1-independent manner by interacting with the repetitive regions of the BCL3 ankyrin domains and altering the transcription factor activities of NFKB in the TP53-dependent apoptotic signaling pathway [36, 37]. Furthermore, a decrease in Bard1 expression has been linked to cellular changes related to a premalignant phenotype [38]. BARD1 has a role in preserving genomic integrity, and BRCA1 null animals were also discovered to have this trait. Early embryonic death was caused by chromosomal instability and BARD1 damage or total deletion [39]. RNA polymerase II was shown to be ubiquitinated by BARD1, and its transcription of damaged DNA was inhibited [34], ubiquitination, beta, and other processes crucial to breast cancer growth [40]. Together, these actions help FL-BARD1 to play a tumor suppressor role, in contrast to reports that BARD1 isoforms such as BARD1 work against this function and accelerate cancer development [41].

More recently, the Exome Sequencing Project and Exome Aggregation Consortium used 1915 patients to link the BARD1 gene and ovarian cancer [42], where the BARD1 gene has a mutation frequency of 0.2%. BARD1 is currently being studied to be included in panels of clinical gene testing for cancer susceptibility due to compelling data linking BARD1 mutations and breast/ovarian cancer susceptibility [43]. Since we mentioned the function of the BARD1 gene as a tumor suppressor, now we should turn to the other vital function of some isoforms of the BARD1 gene as an oncogene.

BARD1 has about 19 distinct expressed isoforms that have been identified so far [12, 44]; several of these isoforms, including BARD1β, BARD1κ, and BARD1π, have been implicated in the development of cancer by an oncogenic role [12, 13]. At the same time, it has been noted that the FL- BARD1, either on its own or in combination with BRCA1, has a tumor suppressor function [41]. However, BARD1β and BARD1δ were previously reported to have an antagonistic effect on full-length BARD1, resulting in oncogenicity and cancer susceptibility [12]. The majority of BARD1 isoforms possess BRCT domains but lack the RING finger domain necessary for the formation of BRCA1 heterodimers. Non-small cell lung cancer (NSCLC), colon cancer, breast cancer, and ovarian cancer all have abnormal BARD1 isoforms, which play a part in cancer progression and carcinogenesis. Additionally, it was revealed that the expression of BARD1 isoforms is significantly linked to a decline in the survival rate of patients with malignancies [12, 15].

BARD1 isoform anomalies result from protein translation from a different open reading frame (ORF). For instance, BARD1 can be translated as a noncontinuous ORF beginning with exon three and then exons 4 through 11. Additionally, it has been demonstrated that BARD1 isoforms inhibit the BRCA1-BARD1 ubiquitin ligase activity necessary to cause the death of cancer cells [12, 15, 40]. Furthermore, the expression of BARD1β has been associated with impaired homologous recombination (HR) and negatively impacted ubiquitin ligase activity in PARPi-sensitive colon cancer cells [45]. The epigenetic has a profound role in BARD1 gene expression and biological consequences. So, the question now is what is this role?

1.4 The epigenetic effect on BARD1 gene expression and biological consequences

Exons 6 to 11 (truncated isoforms) of the BARD1 gene were strongly expressed in Acute myeloid leukemia (AML) in vivo blasts compared to the BARD1-FL expression level. Lepore et al. demonstrated that HDACi (Vorinostat) treatment epigenetically controls the expression of BARD1 mRNA in AML cells, MCF-7 breast cancer cell line, and Kelly neuroblastoma cells. An increase in miR-19a and miR-19b levels were observed after vorinostat therapy, and when BARD1 3’UTR expression was targeted, this increased the apoptotic activity of malignant breast cells [46]. Following a similar pattern, estrogen also activated the estrogen response element (ERE) on BARD1’s intron 9, which favorably controlled the protein expression of BARD1 [47].

While BARD1 9’L, a particular mutation of the BARD1 gene, was reported to compete with miRNAs (such as miR-101 and miR-203) on their binding sites of BARD1 3’UTR, it was also identified to act as competing for endogenous RNA (ceRNA) that negatively controls the expression of BARD1 mRNA [16]. The long non-coding RNAs (lncRNAs) display gene regulatory roles that modulate different biological mechanisms. GUARDIAN, a p53-responsive lncRNA, was determined to be involved efficiently in preserving genomic integrity and delivering protection against genotoxic stress. Additionally, GUARDIAN can facilitate the heterodimerization of BRCA1 with its partner interactor, BARD1, by acting as an RNA scaffold. Therefore, the breakdown of the BRCA1-BARD1 complex caused by GUARDIAN suppression increased the adverse effects of genotoxic stress, induced apoptosis, and caused genomic instability [48].

BRCA1 and BARD1 significantly influence the ATM/ATR pathway for DNA repair mechanisms important for a cell’s decision to die. The BARD1 gene’s epigenetic regulation was affected by histone modification of hESC. Splicing process regulation by H3K36 decreased BARD1 expression, suppressing the ATM/ATR signaling pathways that control hESC development [49]. Hepatocellular carcinoma (HCC) patients whose livers had developed cirrhosis were studied, and it was discovered that the BARD1 gene had much lower levels of methylation (13.3 percent) than in healthy controls. Additionally, it was proposed that BARD1 hypomethylation could be a biomarker for predicting aggressive illness in patients who do not have HBV [50]. Before we address the association of the BARD1 variant to breast cancer risk, we should have a breast cancer mutation in general.

1.5 Breast cancer mutations

The multiple risk factors that have been linked to the development of breast cancer include both genetic and environmental elements. To comprehend the pathogenesis and create a treatment plan, it is essential to identify the genetic and hereditary factors [51]. Numerous genes, including PALB2, ATM, BARD1, BRCA1, BRCA2, and CHEK2, have significantly maintained DNA fidelity and genomic integrity [52, 53, 54]. They are also essential in controlling the HR mechanics. As a result, it has been discovered that various mutations in genes are linked to an increased risk of developing many hereditary malignancies [55], including breast, prostate [56], ovarian, and pancreatic cancers [56, 57, 58, 59]. Several vital polymorphisms in the domains of the BRCA1/BARD1 protein-protein interaction (PPI) complex, including M18K, V11G, L22S, and T97R, were identified by a thorough mutational investigation. These mutations also impacted the stability of the BRCA1/BARD1 PPI complex [60]. The development of preventive and therapeutic strategies to thwart the advancement of breast cancer can therefore be facilitated by a thorough understanding and investigation of the interplay between the BRCA1 and BARD1 platforms [4].

A patient with a breast cancer diagnosis and a family history of the disease was seen as a significant factor in the hereditary predisposition to the condition. BRCA1, BRCA2, PTEN, TP53, CDH1, and STK11 are rare but highly penetrant genes that account for about 30% of hereditary breast cancer cases. BRCA1 mutations were initially found in families with a similar pedigree in 1990. The BRCA2 gene variations were discovered four years later [61]. Hereditary Breast/Ovarian Cancer (HBOC) syndrome is caused by BRCA1 or BRCA2 mutations, yet some with this syndrome were negative for BRCA1 and BRCA2 mutations. BRCA1/2-mutant tumors have a basal, extremely aggressive character. Additionally, 2%–3% of breast cancer cases were found to have mutations in the uncommon but moderately penetrance genes, including; CHEK2, PALB2, ATM, RAD50, BRIP1, RAD51C, NBN, and MRE11. These genes engaged in DNA repair processes and interacted with BRCA1/2. A small number of SNPs, including the mutations for RAD51D, BARD1, RAD51C, ABRAXAS, NBN, and XRCC2BRIP, were linked to poor penetrance alleles and an increase in the risk of breast cancer in a polygenic manner. Clinical testing for mutations found in the high penetrance gene set was typically done on individuals with suspected genetic risk [62]. In addition, a genome-wide association study (GWAS) of breast cancer reported the discovery of 65 novel loci, including FES, MAP3K11, CLK2, GRK7, USP25, DFFA, PKP1, and ZKSCAN3, that are notably related with a high risk of breast cancer at P < 5 × 10⁻⁸ [63].

1.6 The significance of BARD1 in genetic predisposition to breast cancer

Many comprehensive sequencing studies have discovered several genetic variations among different clinical samples. Whereas the biological roles of BRCA1 have been exceptionally well documented, the functional machinery of BARD1 hasn’t been fully understood. Two BARD1 cis mutations, P24S and R378S, were identified in a hereditary breast and ovarian malignancies report. BARD1 and BRCA1interaction m’s affinity is decreased by the P24S mutation, whereas the R378S variant prevents the BRCA1/BARD2 complex from moving into the nucleus. The simultaneous presence of these two mutations contributed synergistically to tumor progress in vitro and in vivo models. Additionally, these two mutations mutually impair the DNA damage response, imposing genomic stability, although neither mutation alone can have a harmful effect [64]. Seven polymorphisms, including somatic missense mutations and germline modifications, were identified within the coding sequence of BARD1 in mutational research that included a variety of gynecological malignancies, ovarian, breast, and uterine tumors. These mutations caused the loss of the wild-type BARD1 allele, which led to the growth and spread of malignancies. A woman with breast and endometrial cancer presented simultaneously had the BARD1 mutation (Gln564His) [65]. Furthermore, the Gln564His mutation of BARD1 was reported to avoid p53-dependent apoptosis by reducing binding to the polyadenylation cleavage specification complex (CSTF-50) [33, 36].

Three non-synonymous variants in the BARD1 gene (Pro24Ser, Arg378Ser, and Val507Met) were assessed in a case-control analysis of 507 Chinese women with breast cancer and 539 matched controls. These SNPs demonstrated significant reductions in breast cancer risk and limited penetrance effects in the BARD1 gene on breast cancer propensity [66]. On the other hand, a large case-control study was carried out among the European (Polish and Belarusian) population to investigate the impact of the nonsense mutation c.1690C>T (p.Q564X). This nonsense variant was found to have a low/moderate increase in breast cancer risk (OR = 2.30, p = 0.04). The risk was further elevated in breast cancer forms that are more aggressive; TNBCs, bilateral breast cancers, early-onset cancer, and hereditary breast and ovarian cancers are a few examples [67]. According to the European study, the BARD1 mutation, one of the most prevalent non-BRCA1/2 mutations, was identified in 10901 TNBC cases and demonstrated a significant contribution to TNBC propensity with an incidence of 0.5−0.7%. Furthermore, Caucasian PVs American carriers of BARD1 gene pathological variants were at lower risk of TNBC (21%) than African American carriers of BARD1 gene mutations (39%) [41, 68].

To categorize the exon mutation of the BARD1 gene in 60 early-onset breast cancer patients and 240 healthy controls, direct sequencing and SNaPshot analysis were used. BARD1’s rs28997575 site was found to have a deletion mutation, which increased the incidence of breast cancer by 3.4 times (P = 0.013) compared to the unaffected group. On the other hand, it was discovered that a different GC genotype missense mutation at the rs2229571 location of BRDA1 was associated with a 72.6 percent (P = 0.001) decreased risk of breast cancer. Remarkably, compared to the control group, the majority of variant carriers have a long family history of breast cancer. Highlighting the significant contribution of breast cancer-positive family history to the elevated risk of breast cancer due to genetic predisposition, especially in BARD1 polymorphism carriers [69]. Likewise, several pathogenic variants (PVs) of the BARD1 gene were compiled in a sizable pooled analytical research of both breast cancer (48,000 cases) and ovarian cancer (20,800 cases). These BARD1 PVs had a moderate chance of developing breast cancer (odds ratio (OR) = 2.90, 95 percent confidence intervals [CIs]: 2.25–3.75, p 0.0001) but not ovarian cancer (OR = 1.36, CIs: 0.87 to 2.11, p = 0.1733). As a result, the BARD1 gene has been suggested as a diagnostic biomarker for evaluating breast cancer patients [70].

More recently, three BARD1 inherited missense mutations were found in the RING domain (Cys53Trp, Cys71Tyr, and Cys83Arg) in a family diagnosed with breast malignancy. However, according to the study, the mutant BARD1/BRCA1 complex was unable resulting nucleosomes and resulted in a loss of H2A ubiquitylation. Mutant BARD1 could heterodimerize with BRCA1 due to its mutations. These mutations also activate a defect in transcriptional repression of the BRCA1-regulated estrogen metabolism genes CYP1A1 and CYP3A4, which are usually controlled by the H2A ubiquitylation pathway [71]. 76 BARD1 cancer-associated missense and truncation variants were effectively identified in a whole-exome sequencing analysis on 10,000 cancer samples from 33 cancer types. Significantly, just two known benign mutations were found to be connected to HDR, whereas four pathogenic mutations are not linked to HDR. DNA damaging agents were more sensitive to BARD1 mutant cells [72]. BARD1 is believed to be a gene predisposing to triple-negative breast cancer and a breast cancer susceptibility gene [68]. With an incidence of 0.5–0.7%, BARD1 was statistically substantially related to a moderate to high risk of TNBC. A rare missense mutation of BARD1 gene c.403G>A or p.Asp135Asn was noticed in TNBC patients. This mutation was reported to increase the response of breast cancer cells to PARPi therapy [73]. While additional BARD1 isoforms are highly expressed in several types of cancer, their common pathogenic effect is owing to the expression of the oncogenic dominant-negative form and alternative splicing (Table 2) [15, 74].

No.	Exon	Nt change	Effect on protein	Frequency for heterozygotes	Previously reported in reference
1	4	1126G → C	Thr351Thr	17.3% (9/52)	[28, 75]
2	4	1145del21	7 aa deletion	1.9% (1/52)	[28, 30]
3	4	1207G → C	Arg378Ser	40.4% (21/52)	[29, 75]
4	6	1591C → T	His506His	7.7% (4/52)	[30, 75]
5	6	1592G → A	Val507Met	50% (26/52)	[28, 30, 75]
6	7	1743G → C	Cys557Ser	1.9% (1/52)	[28, 75]
7	10	2045C → T	Arg658Cys	1.9% (1/52)	[28, 75]

Table 2.

BARD1 variants in breast cancer predisposition.

BARD1 gene polymorphism was found in cases of neuroblastoma and breast cancer cases. The probability of developing neuroblastoma was strongly correlated with three BARD1 gene polymorphisms (rs7585356 GNA, rs6435862 TNG, and rs3768716 ANG). Using the TaqMan approach on 145 cases and 531 controls, only the rs7585356 GNA polymorphism demonstrated notable findings in relation to higher vulnerability to nephroblastoma (odds ratio (OR) = 1.78, 95 percent confidence interval (CI) = 1.01–3.12) with stage I + II clinically [76]. The impact of eleven BARD1 SNPs on NB development has been studied in a Chinese publication. Seven out of eleven BARD1 SNPs revealed an increased risk of high stage (III/IV) NB occurrence. One SNP in the 5’-UTR (rs17489363 G > A), two SNPs in exon (rs2229571 G > C and rs3738888 C > T), and four SNPs in intron (rs3768716 A > G, rs6435862 T > G, rs3768707 C > T, and rs17487792 C > T), were among the eleven BARD1 SNPs [77]. According to reports, the variant (rs17489363 G > A) in the BARD1 gene, which is tied to NB and linked to a decrease in BARD1-FL transcription, is the most common SNP in the gene [50].

Exon 5 is frequently skipped due to the mutation c.1361C>T, which interferes with ANK repeat domains, a critical component of the splicing factor SC35 that regulates apoptosis in the ovarian cancer cell line. NuTu-19 [78]. The NuTu-19 cell line was resistant to the induction of apoptosis. Still, after exogenous expression of the entire gene BARD1, it became susceptible to apoptosis, indicating that the absence of exon 5 results in abnormal isoforms that have lost their capacity to suppress tumors and affect the apoptosis pathway [79]. The BARD1 mutations c.1977A > G, p.Gln715Ter, c.2148delCA, and p.Thr716fs*12 have also been associated with other gynecological cancers, including fallopian tube, ovarian, and cervical cancers [79, 80, 81].

These findings from mounting data have collectively led to the conclusion that there is a context-dependent high/moderate risk of breast cancer associated with specific BARD1 SNPs. Therefore, additional practical and experimental studies are required to validate the aforementioned facts further.

1.7 Correlation between The Cys557Ser BARD1 mutation and risk of breast cancer

One meaningful known change to BARD1 is a missense mutation that causes the amino acid cysteine to be swapped out for the amino acid serine at position 557 (Cys557Ser) [75]. The 126 Finnish breast and ovarian cancer cases were used in a mutational analysis study to examine the possible impact of BARD1 alterations on tumor formation. Breast cancer cases were more likely to have the Cys557Ser missense mutation than healthy controls (7.4 vs. 1.4 percent, p = 0.001). To alter the transcriptional and apoptotic machinery, this variation is required. Intriguingly, the index cases were negative for BRCA1 and BRCA2 mutations highlighting that the occurrence of this mutation in familial predisposition to breast cancer is sufficient to cause the disease on its own [75]. In a study after this one, Stacey et al. and his colleague investigated the relationship between BARD1 Cys557Ser mutation and a familial group of breast cancer using a dataset of 1,090 Icelandic breast cancer patients with invasive type and 703 controls. Carriers of this variant are more likely than non-carriers to develop lobular and medullary breast carcinomas as single or multiple primary breast cancers. Additionally, this risk increased to 0.047 among individuals with the BARD1 Cys557Ser mutation and the BRCA2 999del5 mutation (OR 14 3.11, 95 percent CI 1.16–8.40, p 14 0.046) [82]. A case-control study of the Spanish and South American populations supported past investigations. Despite having a strong family history of breast cancer, the selected individuals have intact BRCA1/2 genes with no mutations. Examining the C-terminal of BARD1 Cys557Ser revealed a substantial increase in the risk of breast cancer (P = 0.04, OR = 3.4 [95 percent CI 1.2–10.2]). This likelihood was further elevated in patients with a family history of breast and ovarian cancer who were also found to have the BARD1 Cys557Ser joint mutation and the XRCC3 241Met variant (P = 0.02, OR = 5.01 [95 percent CI 1.36–18.5]) among patients with a family history of breast and ovarian cancer [83].

Contrarily, a study of Australian patients with a family history of breast cancer revealed that the frequency of the BARD1 Cys557Ser variant was not substantially different from case-control cases (P0.3) and was not linked to an increased risk of breast cancer [84]. Similar to the Australian findings, numerous additional studies have been unable to establish a direct connection between the BADR1 variation and the development of breast cancer [85, 86, 87]. In a cohort of 5,546 BRCA1 and 2,865 BRCA2 mutation carriers, the function of the BARD1 Cys557Ser variation or BARD1 haplotypes as modifiers of BRCA1/2 linked with breast cancer risk was further evaluated. In both BRCA1 and BRCA2 mutation carriers, with a combined expected effect of 0.90 and 0.87, respectively, there was no evidence of either BARD1 mutation to indicate a significant connection with breast cancer risk [88]. Another team of researchers employed DHPLC analysis to identify nine BARD1 coding mutations, including two novel variants, in 210 breast cancer families of Australian descent (129 of which do not have BRCA1 or BRCA2 mutations) (Thr598Ile and Ile692Thr). Yet none of these mutations harbor a pathogenic impact based on their segregation, distribution, and frequency among the selected cases. In addition, non-pathogenic polymorphisms were found in the three variants (1139del21, G1756C, and A2285G) connected to breast cancer in other populations. Therefore, it was not advised in the Australian population to use BARD1 mutations or polymorphisms as a high penetrance susceptibility gene in the progression of familial breast cancer [87].

Collectively, studies linking this BARD1 Cys557Ser mutation to breast cancer incidence have been conducted in Iceland, Finland, Spain/South America, and Italy; however, other studies involving Yoruba, Chinese, Japanese, Australians, and African-Americans have shown different results [30, 65]. These contradictory results regarding the BARD1 Cys557Ser variant’s relationship to familial breast cancer susceptibility raise the possibility that this mutation is restricted to a particular geographic substructure of the European population (as a result of regional migration) rather than being a de novo variant [82]. There are several reported BARD1 mutations, and it is worthwhile to highlight their impact on cancer predisposition risk.

1.8 BARD1 gene as a potential target of new anticancer therapies including sensitivity to chemotherapy with a focus on breast cancer

BARD1 has the potential to be a new target for the therapy of breast cancer, according to several research. According to Zhu Y et al. [14], tamoxifen-resistant breast cancer cells exhibit considerably greater BARD1 and BRCA1 expression levels, which confers resistance to treatment that causes DNA damage such as cisplatin and Adriamycin but not paclitaxel [89, 90, 91, 92]. Watanabe et al. used bisulfite-pyrosequencing to study the aberrant DNA methylation status of the BARD1 gene in 30 TNBC core biopsy specimens from patients with pathologic complete response (noninvasive cancer) and noncomplete response after neoadjuvant chemotherapy (NACT). Even though BRCA1 gene hypermethylation is linked to the TNBC subtype and may affect chemosensitivity and progression under NACT, BARD1 gene hypermethylation only showed a low-to-moderate impact on these procedures [93]. Contrarily, the González-Rivera and his colleagues 2016 underline the low incidence and uncertain clinical implications of gene mutations other than BRCA1/2 (including BARD1) and the associated unfavorable outcomes for patients with breast cancer undergoing NACT [94]. Yet more recent research revealed that tamoxifen-resistant breast cancer cells had increased BRCA1 and its related protein BARD1, making them resistant to treatment that damages DNA [95]. Neoadjuvant chemotherapy now contributes significantly to breast cancer chemotherapy and is a transitional step to adjuvant regimens and other treatments [96]. It is crucial and beneficial to increase research into BARD1’s role in chemotherapy in women who are scheduled for NACT. Both ovarian and breast cancer patients with BRCA1 mutations who initially responded to platinum and PARPi therapy eventually developed resistance to both drugs [97]. In some populations, especially those with evidence of a higher occurrence of BARD1 gene mutations, it is fair to test BARD1 gene isoforms. Additionally, this method would need to be studied for its applicability to outcomes, survival rates, quality of life, influence on treatment choices, and cost-effectiveness for all patients with breast cancer.

2. Concluding remarks and perspectives

This chapter looks at the BRAC1/BARD1, a heterodimeric complex that mediates several biological functions regulating gene transcription and DNA double-stranded break repair mechanism. Then the authors moved to address the BARD1 gene structure, locations, and different isoforms. The authors also focused on the dual function of the BARD1 gene as a tumor-suppressor gene and oncogene. The authors highlighted the epigenetic effect on BARD1 gene expression and biological consequences before turning into breast cancer mutations. We emphasize the significance of BARD1 in genetic predisposition to breast cancer. We also focused on the correlation between the Cys557Ser BARD1 mutation and the risk of breast cancer. Finally, we addressed the BARD1 gene as a potential target of new anticancer therapies, including sensitivity to chemotherapy focused on breast cancer. According to our analysis of the BARD1 gene’s structure and activities, this gene may be crucial to the pathogenesis of breast cancer and the mechanisms underlying cancer cells’ chemo-resistance. In some populations, especially those with evidence of a higher occurrence of BARD1 gene mutations, it is fair to test BARD1 gene isoforms. Additionally, this method would need to be studied for its applicability to outcomes, survival rates, quality of life, influence on treatment choices, and cost-effectiveness for all patients with breast cancer, despite the fact that data on individuals undergoing NACT for breast cancer who have BARD1 gene polymorphism are scarce. Nevertheless, changes in gene expression following NACT may provide insight into the pathophysiology of this complex disease. Regardless of technological advances, there are still some future challenges in including the BARD1 in routine screening—theses challenges including the cost, the technologies sensitivities, and diversity of populations. More work is needed to discover more isoforms for the BARD1. However, limitations are currently present in employing a BARD1 mutation detection panel for breast cancer, such as the lacunae or lack of strong correlation of BARD1 polymorphisms in genetic predisposition to various types of cancer.

Acknowledgments

The King Faisal Specialist Hospital and Research Center in Jeddah and Taif University are gratefully acknowledged by the authors for their help and support.