Genetic risk and clinical implications of BRCA1 and BRCA2 mutations in Turkish triple-negative breast cancer patients

Abstract

Background

Triple-negative breast cancer (TNBC) lacks expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). Genetic mutations, particularly in BRCA1 and BRCA2, significantly influence its pathogenesis and clinical outcomes. This study evaluated the prevalence of BRCA1 and BRCA2 mutations in Turkish TNBC patients and investigated associated clinical, demographic, and pathological factors.

Materials and methods

Patients with TNBC who presented to the Cancer Genetics Department at Istanbul University Oncology Institute were included. Peripheral blood mononuclear cells were analyzed for BRCA1 and BRCA2 mutations using the Illumina MiSeq Next Generation Sequencing (NGS) platform. Patients were categorized as BRCA1 + or BRCA2+ (mutation carriers) or BRCA1- or BRCA2- (mutation non-carriers). Comparative analyses were conducted to identify differences between groups, and clustering analysis examined mutation patterns.

Results

Among 485 TNBC patients, 119 (24.5%) carried pathogenic variants in BRCA1 and/or BRCA2 genes. Of these 119 carriers, 4 (0.8% of the total) harbored mutations in both genes. Specifically, 101 (20.8%) had BRCA1 + mutations, and 22 (4.5%) had BRCA2 + mutations. The carrier group had higher rates of bilateral breast cancer (BC) (14.3% vs. 4.9%), a family history of breast and ovarian cancer (BC and OC) (51.3% vs. 26%), and increased first-degree relative cancer cases. Bilateral BC was associated with a 2.748-fold increased risk of BRCA1 + mutations, while postmenopausal status reduced risk by 0.350-fold. Each additional first-degree BC case increased BRCA1 + mutation risk by 2.410-fold. Cluster analysis identified two distinct mutation patterns.

Conclusion

Turkish TNBC patients with BRCA1 + or BRCA2 + mutations exhibit unique clinical and familial characteristics, emphasizing the importance of genetic screening and familial risk evaluation in TNBC management. These findings underscore the clinical relevance of BRCA testing in TNBC patients for personalized screening and treatment strategies. Notably, this study provides the largest TNBC cohort (n = 485) reported from Türkiye, highlighting the significant role of BRCA1 in TNBC pathogenesis and offering a roadmap for individualized management.

Supplementary Information

The online version contains supplementary material available at 10.1007/s12672-025-03470-7.

Introduction

Breast cancer (BC) remains a significant global health challenge, affecting millions of women worldwide and contributing substantially to morbidity and mortality [1]. As research has advanced, various subtypes of BC have been identified, each with special biological and clinical characteristics. One such subtype that has drawn significant attention is triple-negative breast cancer (TNBC), which is distinct due to its lack of expression of the estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) [2]. Despite representing only 10–15% of all BC cases [3], TNBC exerts a significant clinical burden that is disproportionate to its prevalence, due to its aggressive nature and limited treatment options [4, 5].

Genetic predisposition, particularly mutations in the BRCA1 and BRCA2 genes, plays a pivotal role in the development and prognosis of TNBC [6]. These mutations are key drivers of tumorigenesis in TNBC, linking genetic alterations to its aggressive nature and clinical outcomes. Women carrying BRCA1 mutations have an 80% lifetime risk of developing BC, while BRCA2 mutation carriers face a 69% risk [7]. These mutations are strongly associated with early-onset BC, TNBC, and familial cancer syndromes [8, 9]. However, while extensive research has focused on Western populations, there remains a paucity of data regarding the correlation between BRCA1 and BRCA2 mutations and the clinical and pathological features of TNBC within other populations, such as the Turkish population. Therefore, understanding the genetic and clinical characteristics of TNBC, particularly in the context of BRCA1 or BRCA2 mutations, is crucial for developing personalized treatment strategies and prognostic assessments in this region.

In Türkiye, recent studies have highlighted distinct genetic patterns in BC patients [10]. BRCA1 mutations appear to be more prevalent than BRCA2 mutations [11], with frequencies ranging from 19 to 37% among high-risk BC patients [12]. Interestingly, sporadic cases of BC without familial histories have shown a higher prevalence of BRCA2 mutations [13]. Specific to TNBC, recent research has identified associations between BRCA1 and BRCA2 mutations and high Ki-67 indices, bilateral BC, and a significant familial cancer history [14]. These findings underscore the importance of population-specific studies, as genetic predispositions can vary significantly across different ethnic and geographic groups. Despite this emerging evidence, comprehensive data on BRCA1 and BRCA2 mutation prevalence and associated clinical factors in Turkish TNBC patients are still limited. Addressing this gap is critical for advancing personalized medicine approaches, optimizing screening programs, and improving therapeutic strategies tailored to the unique genetic and clinical characteristics of the population.

This study aims to investigate the prevalence of BRCA1 + and BRCA2 + mutations in a large cohort of 485 Turkish TNBC patients and to explore the demographic, clinical, and pathological features associated with these mutations. By elucidating the genetic landscape of TNBC in Türkiye, the study seeks to contribute valuable insights to the growing body of evidence on the role of BRCA1 + and BRCA2 + mutations in BC and to inform population-specific strategies for risk assessment and management.

Materials and methods

Study population and sample collection

This study included 485 TNBC patients who applied to the Cancer Genetics Department of Istanbul University Oncology Institute, Istanbul, Türkiye. The study protocol was approved by the Ethics Committee of Istanbul University (approval number 2023/500), and all procedures adhered to the principles of the Declaration of Helsinki [15]. Informed written consent was obtained from all participants. Peripheral blood mononuclear cells were collected, and the BRCA1 and BRCA2 mutation status was analyzed using Next-Generation Sequencing (NGS). All methods were carried out in accordance with relevant guidelines and regulations.

Patient demographics and clinical data

The study cohort consisted of 485 patients with confirmed TNBC. A flowchart summarizing the study design is shown in Fig. 1.

Fig. 1.

Study Flowchart: This diagram illustrates the patient inclusion process, from breast cancer diagnosis to BRCA1+ or BRCA2+ NGS testing and hormone receptor status confirmation (ER, PR, HER2) for study eligibility

Data collection

Clinical and pathological data, including BRCA1 and BRCA2 genetic testing results, age at diagnosis, family history of BC and OC, tumor grade, histological features, tumor size (< 2 cm or ≥ 2 cm), metastasis status, lymph node involvement, menopausal status, treatment modalities (chemotherapy and radiotherapy), and current status (alive or deceased), were collected from medical records. The receptor status for ER, PR, and HER2 was confirmed based on immunohistochemical staining, with positivity defined as staining in at least 1% of cells [16] from the patient’s record.

Detection of BRCA1+ or BRCA2 + germline mutations by Next Generation Sequencing

Deoxyribonucleic acid (DNA) was extracted from peripheral blood mononuclear cells using the GeneAll^® Exgene™ kit(GeneAll Biotechnology, Seoul, Korea). DNA quantification was performed using the NanoDrop 2000c spectrophotometer (Thermo Fisher Scientific, USA). All coding exons and adjacent exon-intron boundaries of BRCA1 and BRCA2 were sequenced using the Illumina MiSeq^® platform and Sophia Genetics (Illumina, San Diego, CA, USA) kits. Targeted sequencing was performed using the Sophia Genetics BRCA Hereditary Cancer Solution v2.0 (HCSv2.0), RUO (Research Use Only), designed for Illumina platforms (Catalog No: 000407_20181204_01) according to the manufacturer’s protocol. Sequencing libraries were prepared using a multiplex reaction with 100 ng of genomic DNA, followed by a second PCR to incorporate molecular barcodes and adapter sequences. Libraries were normalized and sequenced using the MiSeq Reagent Kit v2 (300 cycles), generating paired-end reads with an average coverage depth of 1200x. Variants identified by NGS were validated through Sanger sequencing for point mutations and by Multiplex Ligation-dependent Probe Amplification (MLPA) for large deletions and duplications.

Variant analysis and classification

Sequence alignment and variant calling were performed using SOPHiA DDM software (v5.10.6) (Saint-Sulpice, France) against the reference genome (GRCh37/hg19). Variants were annotated and filtered using multiple databases, including dbSNP [17], ExAC [18], GnomAD [19], SIFT [20], POLYPHEN2 [21], MUTATION TASTER [22], ClinVar [23], and HGMD [24]. Variants were classified following the guidelines of the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP), with a focus on pathogenicity associated with loss-of-function variants [25]. In patients harboring both BRCA1 + and BRCA2+, each variant was individually confirmed as pathogenic or likely pathogenic according to ClinVar and included in the analysis only if consistent with ACMG/AMP classification criteria.

Statistical analysis

Descriptive statistics were presented as frequencies and percentages for categorical variables and as mean, standard deviation, median, minimum, and maximum for continuous variables. The Pearson chi-square and Fisher’s exact tests were used for categorical variable comparisons, with Bonferroni correction applied for multiple comparisons. The Kolmogorov-Smirnov test was used to assess the normality of continuous variables. The Mann-Whitney U test compared the medians of two independent groups, while multivariate binary logistic regression was employed to evaluate factors associated with BRCA1 or BRCA2 mutation status. Variables with a p-value < 0.25 in univariate analysis were included in the regression model using the enter method. Model fit was verified with the Hosmer-Lemeshow test. Cluster analysis, using the two-step clustering method, categorized patients based on BRCA1 or BRCA2 mutation status, family history, and clinical features. Key variables influencing cluster formation were identified, and differences in mortality rates between clusters were assessed. Statistical significance was set at p < 0.05. Analyses were performed using SPSS Statistics (Version 26.0, IBM Corp., Armonk, NY, USA).

Kaplan-Meier survival analysis

Kaplan-Meier survival curves were generated to evaluate overall survival as a function of BRCA1 + mutation status. Survival time was calculated as the time (in months) from the date of diagnosis to the date of last follow-up. Patients who were alive at the last follow-up were treated as censored, and censoring was indicated in the survival charts by checking a box. Although only a total of 26 patients (8 BRCA1 + patients) had died at the time of analysis, many patients had not yet experienced the event during the follow-up period, resulting in a high number of censored observations on the Kaplan-Meier curves. Survival distributions were compared using the log-rank test (p = 0.234). All p-values were two-sided, and a p-value of less than 0.05 was considered statistically significant.

Results

General characteristics of TNBC patients

This study included 485 patients diagnosed with TNBC. Figure 2 summarizes the demographic and clinical characteristics, including diagnostic profiles, BRCA1 or BRCA2 mutation status, age distribution, tumor characteristics, treatment modalities, and family cancer history. The detailed table can be seen in supplementary material 1. Among the 485 TNBC patients, 92.7% (n = 450) had unilateral BC, whereas 7.2% (n = 35) were diagnosed with bilateral BC. The total number of patients with BRCA1 + or BRCA2 + mutations was determined to be 119 (%24.5). Among these 119 patients, 4 carry both BRCA1 or BRCA2 + mutations. Thus, 101 (20.8%) patients carry only the BRCA1 + mutation, 22 (4.5%) patients carry only the BRCA2 + mutation, and 4 patients constitute a group with both mutations.

Fig. 2.

Clinical and Diagnostic Characteristics of TNBC Patients were investigated in this study. IDC Invasive ductal carcinoma, ILC Invasive lobular carcinoma

Notably, 4 patients (0.8%) were found to carry pathogenic variants in both genes simultaneously, indicating that while BRCA1 or BRCA2 + mutations are generally mutually exclusive due to their distinct pathogenic mechanisms, rare cases of co-occurrence can occur within the same individual. The age distribution revealed that nearly half of the patients (46.5%) were ≤ 40 years at diagnosis, and 69.5% (n = 337) were ≤ 45 years. Grade 3 tumors predominated (74.0%, n = 359), followed by Grade 2 (24.3%) and Grade 1 (1.6%). Most TNBC cases (93.2%) were classified as invasive ductal carcinoma (IDC), with invasive lobular carcinoma (ILC) representing 5.2%. Tumors measuring ≥ 2 cm accounted for 66.6% of cases. Metastasis was observed in 22.5%, and lymph node involvement occurred in 48.7%. Regarding treatment, 84.1% of patients underwent surgery, 90.1% received chemotherapy, and 54.8% had radiotherapy (all 485 TNBC patients were included). A significant proportion (77.9%) were diagnosed premenopausally. Regarding family history, 67.2% of patients reported < 2 occurrences of BC and OC in their families, while 32.2% reported ≥ 2 occurrences.

Comparison of general characteristics between BRCA1+ or BRCA2+ (mutation carriers) or BRCA1- or BRCA2- (mutation non-carriers) in TNBC patients

We evaluated the clinical and familial characteristics of TNBC patients with BRCA1 + or BRCA2 + mutations (mutation carriers, n = 119) and without BRCA1- or BRCA2- mutations (mutation non-carriers,366). Table 1 shows the comparative analysis of TNBC patients based on BRCA1 or BRCA2 mutation status. The study identified significant differences between the two groups in terms of diagnosis patterns, family cancer history, and other key clinical factors.

Table 1.

General characteristics of the carrier (BRCA1 + or BRCA2+) and Non-Carrier (BRCA1– or BRCA2–) TNBC patients investigated in this study

		Non-Carrier (n = 366)	Carrier (n = 119)	p-Value
Diagnosis	Unilateral Breast Cancer	294 (80.3%)	78 (65.5%)	< 0.001
	Bilateral Breast Cancer	19 (4.9%)	17 (14.3%)
	Breast and Ovarian Cancer Together	2 (0.5%)	6 (5%)
	Breast and Other Cancers Together	51 (13.9%)	18 (15.1%)
Age (Years)	≤40	179 (48.9%)	68 (57.1%)	0.261
	41–49	106 (29.0%)	31 (26.1%)
	≥50	81 (22.1%)	20 (16.8%)
Age 45	≤45	246 (67.2%)	91 (76.5)	0.057
	> 45	120 (32.8%)	28 (23.5%)
Age 40	≤40	179 (48.9%)	68 (57.1%)	0.118
	> 40	187 (51.1%)	51 (42.9%)
Pathological Stage	Stage 1	32 (8.7%)	12 (10.1%)	0.145
	Stage 2	122 (33.3%)	36 (30.3%)
	Stage 3	189 (51.6%)	56 (47.1%)
	Stage 4	23 (6.3%)	15 (12.6%)
Tumor Grade (Nottingham)	Grade 1	4 (1.1%)	4 (3.4%)	0.132
	Grade 2	94 (25.7%)	24 (20.2%)
	Grade 3	268 (73.2%)	91 (76.5%)
Histological Type	IDC	325 (88.8%)	109 (91.6%)	0.572
	ILC	6 (1.6%)	1 (0.8%)
	IDC + ILC	15 (4.1%)	2 (1.7%)
	Other	20 (5.5%)	7 (5.9%)
IDC	No	25 (6.8%)	8 (6.7%)	0.968
	Yes	341 (93.2%)	111 (93.3%)
ILC	No	345 (94.3%)	115 (96.6%)	0.308
	Yes	21 (5.7%)	4 (3.4%)
Tumor Size	< 2 cm	121 (33.1%)	41 (34.5%)	0.823
	≥2 cm	245 (66.9%)	78 (65.5%)
Metastasis Status	No	286 (78.1%)	90 (75.6%)	0.569
	Yes	80 (21.9%)	29 (24.4%)
Lymph Node Involvement	No	191 (52.2%)	58 (48.7%)	0.513
	Yes	175 (47.8%)	61 (51.3%)
Chemotherapy	No	41 (11.2%)	7 (5.9%)	0.091
	Yes	325 (88.8%)	112 (94.1%)
Radiotherapy	No	164 (44.8%)	55 (46.2%)	0.788
	Yes	202 (55.2%)	64 (53.8%)
Surgery	No	62 (16.9%)	16 (13.4%)	0.367
	Yes	304 (83.1%)	103 (86.6%)
Last Status	Deceased	18 (4.9%)	8 (6.7%)	0.448
	Alive	348 (95.1%)	111 (93.3%)
The Number of BC/OC in the family	< 2 BC and OC	271 (74.0%)	58 (48.7%)	< 0.001
	≥2 BC and OC	95 (26.0%)	61 (51.3%)
Menopausal Status	Pre-menopausal	279 (76.2%)	99 (83.2%)	0.112
	Pos-tmenopausal	87 (23.8%)	20 (16.8%)
Number of Breast Cancer Cases in the Family		0.6858 ± 1.0479 0 (0–8)	1.3697 ± 1.5230 1 (0–6)	< 0.001
Number of Ovarian Cancer Cases in the Family		0.1202 ± 0.4210 0 (0–4)	0.4622 ± 0.7896 0 (0–5)	< 0.001
Number of Other Tumor Cases in the Family		1.9399 ± 1.80765 1 (0–8)	2.395 ± 2.14404 2 (0–11)	0.045
Number of Total Cancer Cases in the Family		2.6913 ± 2.9956 2 (0–15)	4.1092 ± 2.93947 4 (0–15)	< 0.001
Average Number of First-Degree Relatives with Breast Cancer		0.17 ± 0.419 0 (0–2)	0.38 ± 0.638 0 (0–2)	< 0.001
Average Number of Second-Degree Relatives with Breast Cancer		0.20 ± 0.469 0 (0–2)	0.45 ± 0.778 0 (0–4)	< 0.001
Average Number of Third-Degree Relatives with Breast Cancer		0.17 ± 0.476 0 (0–4)	0.39 ± 0.714 0 (0–4)	< 0.001
Average Number of Fourth-Degree Relatives with Breast Cancer		0.14 ± 0.565 0 (0–7)	0.16 ± 0.638 0 (0–5)	0.700

IDC Invaziv Ductal Carcinoma, ILC Invaziv Lobular Carcinoma, BC Breast Cancer, OC Ovarian Cancer

Diagnosis patterns

Significant differences were observed in the distribution of cancer diagnoses. Bilateral BC is significantly more common among carriers compared to non-carriers (14.3% vs. 4.9%, p < 0.001). Additionally, the occurrence of concurrent BC and OC was higher in carriers (5% vs. 0.5%), further emphasizing the heightened risk for OC in BRCA1 + or BRCA2 + mutation carriers.

Family cancer history

Carriers had significantly more family members affected by BC and OC. The number of BC and OC cases in the family was notably higher among carriers, with 51.3% having two or more cases compared to 26.0% of non-carriers (p < 0.001). Carriers also exhibited significantly higher average numbers of BC cases among first-degree (0.38 ± 0.638 vs. 0.17 ± 0.419, p < 0.001), second-degree (0.45 ± 0.778 vs. 0.20 ± 0.469, p < 0.001), and third-degree relatives (0.39 ± 0.714 vs. 0.17 ± 0.476, p < 0.001).

Clinical and pathological features

While the average age at diagnosis did not differ significantly between carriers and non-carriers, carriers were more frequently diagnosed at a younger age (≤ 40 years: 57.1% vs. 48.9%, p = 0.261). The pathological stage, tumor grade, histological type, and metastasis status were comparable between the two groups. Most tumors were IDC (carriers: 91.6%; non-carriers: 88.8%, p = 0.572) and high grade (Grade 3: carriers 76.5%, non-carriers 73.2%, p = 0.132). Tumor size also did not significantly differ, with most patients presenting with tumors ≥ 2 cm (carriers: 65.5%, non-carriers: 66.9%, p = 0.823).

Treatment and outcomes

Treatment modalities, including surgery, chemotherapy, and radiotherapy, were similar between the groups. However, a slightly higher proportion of carriers received chemotherapy (94.1% vs. 88.8%, p = 0.091). Survival outcomes were also comparable, with 93.3% of carriers and 95.1% of non-carriers alive at the last follow-up (p = 0.448). These findings highlight distinct familial and clinical patterns in BRCA1 + or BRCA2 + mutation carriers among TNBC patients. The significant association between BRCA1 + or BRCA2 + mutations and a family history of BC and OC underscores the importance of comprehensive family history in genetic risk assessment. Additionally, the higher prevalence of bilateral BC and concurrent BC and OC in carriers supports the inclusion of these clinical factors in BRCA1 + and BRCA2 + mutation screening criteria.

The age-specific prevalence of BRCA1 + or BRCA2 + mutations is visualized in Fig. 3. Patients aged ≤ 40 years demonstrated the highest mutation prevalence (BRCA1: 56.4%; BRCA2: 63.6%; BRCA1 + or BRCA2+: 50.9%), declining with increasing age (p < 0.001).

Fig. 3.

The proportion of TNBC patients with positive BRCA1+ or BRCA2+ testing based on age at diagnosis. The proportion of patients testing positive for BRCA1 by age group were: ≤40 y: 56.4 % ; 41–49y:27.7%; ≥50y:15.8%; for BRCA2: ≤40: 63.6%; 41–49:18.2%; ≥50:18.2%; either BRCA1+ and BRCA2+: ≤40: 50.9%; 41–49:28.2%; ≥50:20.8%)

Spectrum and frequency of BRCA1 + or BRCA2 +  mutations in Turkish TNBC patients

Among 485 TNBC patients, 119 (24.5%) harbored BRCA1 + and/or BRCA2 + mutations, comprising 101 (20.8%) with BRCA1 + mutations and 22 (4.5%) with BRCA2 + mutations; notably, 4 patients had mutations in both genes, explaining the difference between the summed counts and the total number of unique carriers (Fig. 4). Comparison of clinical and demographic profiles between TNBC Patients with and without BRCA1 or BRCA2 mutations is shown in Table 2.

Table 2.

Comparative analysis of clinical and demographic characteristics in TNBC patients with and without BRCA1 + and BRCA2 + Mutations

		BRCA1		p-Value	BRCA2		p-Value
		Negative	Positive		Negative	Positive
Diagnosis	Unilateral Breast Cancer	306 (79.7%)	66 (65.3%)	0.001	357 (77.1%)	15 (68.2%)	0.222
	Bilateral Breast Cancer	22 (5.8%)	14 (13.9%)		32 (6.8%)	4 (18.2%)
	Breast and Ovarian Cancer Together	3 (0.8%)	5 (5.0%)		7 (1.5%)	1 (4.5%)
	Breast and Other Cancers Together	53 (13.8%)	16 (15.8%)		67 (14.5%)	2 (9.1%)
Age (Years)	≤40	190 (49.5%)	57 (56.4%)	0.318	233 (50.3%)	14 (63.6%)	0.442
	41–49	109 (28.4%)	28 (27.7%)		133 (28.7%)	4 (18.2%)
	≥50	85 (22.1%)	16 (15.8%)		97 (21.0%)	4 (18.2%)
Age45	≤45	259 (67.4%)	78 (77.2%)	0.058	320 (69.1%)	17 (77.3%)	0.417
	> 45	125 (32.6%)	23 (22.8%)		143 (30.9%)	5 (22.7%)
Age40	≤40	190 (49.5%)	57 (56.4%)	0.213	233 (50.3%)	14 (63.6%)	0.222
	> 40	194 (50.5%)	44 (43.6%)		230 (49.7%)	8 (36.4%)
Pathological Stages	Stage 1	32 (8.3%)	12 (11.9%)	0.094	43 (9.3%)	1 (4.5%)	0.308
	Stage 2	130 (33.9%)	28 (27.7%)		148 (32%)	10 (45.5%)
	Stage 3	197 (51.3%)	48 (47.5%)		237 (51.2%)	8 (36.4%)
	Stage 4	25 (6.5%)	13 (12.9%)		35 (7.6%)	3 (13.6%)
Tumor Grade (Nottingham)	Grade 1	5 (1.3%)	3 (3.0%)	0.197	7 (1.5%)	1 (4.5%)	0.361
	Grade 2	99 (25.8%)	19 (18.8%)		111 (24.0%)	7 (31.8%)
	Grade 3	280 (72.9%)	79 (78.2%)		345 (74.5%)	14 (63.6%)
Histological Type	IDC	341 (88.8%)	93 (92.1%)	0.455	415 (89.6%)	19 (86.4%)	0.547
	ILC	6 (1.6%)	1 (1.0%)		7 (1.5%)	0
	IDC + ILC	16 (4.2%)	1 (1.0%)		16 (3.5%)	1 (4.5%)
	Other	21 (5.5%)	6 (5.9%)		25 (5.4%)	2 (9.1%)
IDC	No	26 (6.8%)	7 (6.9%)	0.955	31 (6.7%)	2 (9.1%)	0.663
	Yes	358 (93.2%)	94 (93.1%)		432 (93.3%)	20 (90.9%)
ILC	No	362 (94.3%)	98 (97.0%)	0.265	439 (94.8%)	21 (95.5%)	0.895
	Yes	22 (5.7%)	3 (3.0%)		24 (5.2%)	1 (4.5%)
Tumor Size	< 2 cm	126 (32.8%)	36 (35.6%)	0.591	155 (33.5%)	7 (31.8%)	0.872
	≥2 cm	258 (67.2%).	65 (64.4%)		308 (66.5%)	15
Metastasis Status	No	301 (78.4%)	75 (74.3%)	0.377	358 (77.3%)	18 (81.8%)	0.622
	Yes	83 (21.6%)	26 (25.7%)		105 (22.7%)	4 (18.2%)
Lymph Node Involvement	No	197 (51.3%)	52 (51.5%)	0.974	240 (51.8%)	9 (40.9%)	0.316
	Yes	187 (48.7%)	49 (48.5%)		223 (48.2%)	13 (59.1%)
Chemotherapy	No	39 (10.2%)	9 (8.9%)	0.709	48 (10.4%)	0	0.112
	Yes	345 (89.8%)	92 (91.1%)		415 (89.6%)	22 (100.0%)
Radiotherapy	No	168 (43.8%)	51 (50.5%)	0.225	212 (45.8%)	7 (31.8%)	0.198
	Yes	216 (56.3%)	50 (49.5%)		251 (54.2%)	15 (68.2%)
Surgery	No	62 (16.1%)	16 (15.8%)	0.941	77 (16.6%)	1 (4.5%)	0.132
	Yes	322 (83.9%)	85 (84.2%)		386 (83.4%)	21 (95.5%)
Last Status	Deceased	18 (4.7%)	8 (7.9%)	0.199	26 (5.6%)	0	0.253
	Alive	366 (95.3%)	93 (92.1%)		437 (94.4%)	22 (100.0%)
The Number of BC/OC in the family	< 2 BC and OC	284 (74.0%)	45 (44.6%)	< 0.001	315 (68.0%)	14 (63.6%)	0.666
	≥2 BC and OC	100 (26.0%)	56 (55.4%)		148 (32.0%)	8 (36.4%)
Menopausal Status	Pre-menopausal	291 (75.8%)	87 (86.1%)	0.026	362 (78.2%)	16 (72.7%)	0.546
	Post-menopausal	93 (24.2%)	14 (13.9%)		101 (21.8%)	6 (27.3%)
Number of Breast Cancer		0.6875 ± 1.045 0 (0–8)	1.4851 ± 1.5723 1 (0–6)	< 0.001		0.8531 ± 1.2256 0 (0–8)	0.8636 ± 1.0371 0.5 (0–3)	0.711
Number of Ovarian Cancer		0.1224 ± 0.4127 0 (0–4)	0.5149 ± 0.8439 0 (0–5)	< 0.001		0.2030 ± 0.5600 0 (0–5)	0.2273 ± 0.4289	0.403
Number of Other Tumors		1.9297 ± 1.8417 1 (0–9)	2.5149 ± 2.0669 2 (0–11)	< 0.004		2.0605 ± 1.8842 2 (0–11)	1.8636 ± 2.31549 1 (0–9)	0.290
Total Number of Cancer Cases		2.7031 ± 2.3616 2 (0–15)	4.3168 ± 2.8033 4 (0–15)	0.001		3.0389 ± 2.5137 3 (0–15)	3.0455 ± 3.1694 2 (0–12)	0.582
Average Number of First-Degree Relatives With Breast Cancer		0.17 ± 0.421 0 (0–2)	0.43 ± 0.653 0 (0–2)	< 0.001		0.22 ± 0.489 0 (0–2)	0.18 ± 0.501 0 (0–2)	0.549
Average Number of Relatives With Second-Degree Breast Cancer		0.20 ± 0.465 0 (0–2)	0.50 ± 0.820 0 (0–4)	< 0.001		0.26 ± 0.572 0 (0–4)	0.32 ± 0.568 0 (0–2)	0.468
Average Number of Relatives With Third-Degree Breast Cancer		0.17 ± 0.476 0 (0–4)	0.41 ± 0.751 0 (0–4)	0.001		0.22 ± 0.557 0 (0–4)	0.23 ± 0.429 0 (0–1)	0.545
Average Number of Relatives With Fourth-Degree Breast Cancers		0.15 ± 0.564 0 (0–7)	0.15 ± 0.654 0 (0–5)	0.366		0.14 ± 0.587 0 (0–7)	0.18 ± 0.501 0 (0–2)	0.493

IDC Invaziv Ductal Carcinoma, ILC Invaziv Lobular Carcinoma, BC Breast Cancer, OC Ovarian Cancer

Fig. 4.

Distribution of Pathogenic BRCA1 or BRCA2+ Mutations Among Turkish TNBC Patients: BRCA1+ mutations were detected in 20.8% (n=101), BRCA2+ mutations in 4.5% (n=22), and concurrent BRCA1 or BRCA2+ mutations in 0.8% (n=4) of patients. Notably, the majority (74.6%, n=366) of patients harbored no pathogenic mutations in either gene)

A comprehensive mutational analysis of BRCA1 or BRCA2 genes in the TNBC cohort revealed a diverse range of pathogenic alterations, including frameshift, nonsense, missense, splice site variants, in-frame deletions, and large copy number variations (CNVs) (Fig. 5). Frameshift mutations represented the most common alteration type (47.9%), predominantly affecting BRCA1. Nonsense (20.3%) and missense (13.0%) mutations were also observed, with missense variants primarily localized to BRCA1. Additionally, splice site variants were identified exclusively in BRCA1, while in-frame deletions were rare and confined to BRCA2. Copy number variants (CNVs) accounted for 8.9% of all pathogenic alterations, with exon-level deletions and duplications observed in both genes. Although BRCA1 or BRCA2 mutations are typically considered mutually exclusive due to their distinct biological functions, four patients (0.8%) were found to harbor concurrent pathogenic variants in both genes. These findings underscore the genetic heterogeneity of BRCA-related TNBC and highlight the importance of comprehensive testing, including full gene sequencing and CNV analysis.

Fig. 5.

Distribution of All Pathogenic BRCA1 or BRCA2+ Mutations and Their Locations. The most prevalent mutation type was frameshift (50%), followed by nonsense (21%), missense (14%), copy number variants (9%), splice site mutations (3%), and in-frame deletions (3%). These findings reflect the mutational heterogeneity observed in BRCA-associated TNBC

Logistic regression analysis

Factors associated with BRCA1, BRCA2, and BRCA1 + or BRCA2 + Gene Mutation Status

The logistic regression analysis, examining factors related to BRCA1, BRCA2, and BRCA1 + or BRCA2 + mutation risks in 485 TNBC patients, is shown in Table 3. The analysis showed significant associations between various clinical and familial factors and the risk of BRCA1 + and combined BRCA1 + or BRCA2 + mutations, but no significant associations were found for BRCA2 + mutations alone. As represented in Table 3, in the logistic regression analysis, factors influencing BRCA1 + mutation risks in TNBC patients includes the presence of bilateral BC (p = 0.018, OR = 2.748, 95% C.I. 1.192–6.335), menopausal status (p = 0.025, OR = 0.350, 95% C.I. 0.140–0.876), the number of OC cases in family/relatives (p < 0.001, OR = 4.449, 95% C.I. 2.174–9.102), and the occurrence of BC in first (p = 0.010, OR = 2.410, 95% C.I. 1.239–4.687), second (p = 0.013, OR = 2.211, 95% C.I. 1.185–4.127), and third (p = 0.037, OR = 1.936, 95% C.I. 1.039–3.608) degree relatives were significantly associated with BRCA1 + mutation risks, suggesting that family clustering of BC across different degrees of relatives is relevant to BRCA1 + mutation risk. The Hosmer-Lemeshow test confirmed the model fit (p = 0.977, p = 0.610, p = 0.771). The risk of BRCA + 1 mutation was found to be 2.748 times higher in individuals with bilateral BC compared to those with unilateral BC (p = 0.018). Postmenopausal status was associated with a 0.350 times lower risk of BRCA1 + mutation (p = 0.025). Moreover, an increase of one BC case in the first-degree relatives of a patient with TNBC resulted in a 2.410 times higher risk of BRCA1 + mutation. Postmenopausal status was associated with a 0.350 times lower risk of carrying the BRCA1 + mutation (p = 0.025). Furthermore, each additional BC case in first-degree relatives of a TNBC patient increased the risk of carrying the BRCA1 + mutation by 2.410 times. No significant factors were identified in the analysis of BRCA2 + mutation status. Therefore, no links were found between clinical or familial factors and the chance of carrying a BRCA2 + mutation in this population. For BRCA1 + or BRCA2 + mutations, the number of OC cases in the family was also significantly linked to mutation risk (p = 0.004, OR = 2.494, 95% CI: 1.350–4.605). Similar to BRCA1+, BC cases in first-degree (p = 0.009, OR = 2.260, 95% CI: 1.222–4.182), second-degree (p = 0.036, OR = 1.843, 95% CI: 1.039–3.268), and third-degree relatives (p = 0.038, OR = 1.830, 95% CI: 1.033–3.242) were significantly associated with BRCA1 + or BRCA2 + mutation risk. Figure 6 also shows the forest plot analysis of factors linked to BRCA1 + or BRCA2 + mutation risk.

Table 3.

Factors associated with BRCA1+, BRCA2+, and BRCA1 + and BRCA2 + Mutation risk in TNBC patients

		p-value	OR	95% C.I.for OR
				Lower	Upper
BRCA1+	Bilateral Breast Cancer	0.018	2.748	1.192	6.335
	Menopausal Status	0.025	0.350	0.140	0.876
	Number of Ovarian Cancer	< 0.001	4.449	2.174	9.102
	Number of Breast Cancers in First-Degree Relatives	0.010	2.410	1.239	4.687
	Number of Breast Cancers in Second-Degree Relatives	0.013	2.211	1.185	4.127
	Number of Breast Cancers in Third-Degree Relatives	0.037	1.936	1.039	3.608
BRCA2+	Diagnosis	0.472	0.575	0.127	2.600
	Age 40	0.182	0.539	0.217	1.337
	Operation	0.754	2.051	0.023	184.882
	Chemotherapy	0.548	1.906	0.233	15.622
	Radiotherapy	0.466	1.439	0.541	3.826
	Surgery	0.867	1.477	0.016	140.124
BRCA1 + and/or BRCA2+	Number of Ovarian Cancer	0.004	2.494	1.350	4.605
	Number of Breast Cancers in First-Degree Relatives	0.009	2.260	1.222	4.182
	Number of Breast Cancers in Second-Degree Relatives	0.036	1.843	1.039	3.268
	Number of Breast Cancers in Third-Degree Relatives	0.038	1.830	1.033	3.242

OR Odds Ratio. (The small number of patients with BRCA2 mutations (n = 22) limited statistical power)

Fig. 6.

Forest plot analysis of factors associated with BRCA1+, BRCA2+ mutation risk, and combined BRCA1+ and BRCA2+ mutation risk. BRCA1+ associated risk factors are shown in blue, and BRCA1+ and BRCA2+ associated risk factors are shown in orange. No significant associations were identified between BRCA2+ mutation status and clinical or familial factors in the study population; hence, these variables were not included in the results. The dashed line indicates an OR of 1.0 (no association).

Cluster analysis of TNBC patients

In our cluster analysis, we identified several variables that contributed most significantly to differentiating the two patient groups. To improve clarity and consistency, we refined the clinical definitions of these groups, ensuring that the classification accurately reflects both familial burden and survival differences. Cluster 1 was characterized by a higher familial burden of BC and OC, reflected by significantly increased numbers of first- and second-degree relatives with BC, greater total cancer cases in the family, and elevated rates of BRCA1 + mutations. Accordingly, we now define Cluster 1 as the “High Familial Burden” group. In contrast, Cluster 2 demonstrated a lower familial cancer burden, fewer BRCA1 + mutations, and overall better survival outcomes; we define this group as the “Sporadic/Lower Familial Risk” cluster. Notably, this refined classification resolves previous inconsistencies between advanced stage presentation and improved survival observed in Cluster 2, by highlighting the greater genetic and familial predisposition—and associated clinical aggressiveness—within Cluster 1. As shown in Fig. 7, factors such as family history of BC and OC, the total number of cancer cases in the family, the number of BC cases among first-degree relatives, and BRCA1 + mutation status exhibited the highest relative importance in defining cluster membership. These findings are consistent with the statistical results presented in Table 4, which demonstrated significant differences between clusters for these variables (p < 0.001). Collectively, these results indicate that familial cancer burden and BRCA1 + mutation status play a critical role in defining distinct clinical subgroups among TNBC patients. Initially, BRCA2 + mutation status was also included in the clustering analysis. However, statistical testing revealed that BRCA2 + was not significantly associated with any of the examined clinical or demographic variables (all p-values > 0.05, shown in Table 2). Therefore, to maintain clarity and focus on meaningful associations, BRCA2 + was excluded from the final clustering model. Our analysis revealed that BRCA1 + mutation positivity was substantially higher in Class 1 (36.9%) compared to Class 2 (6.9%, p < 0.001), with MLPA positivity following a similar pattern (41.8% vs. 8.1%, p < 0.001). Pathological staging differed significantly between groups, with Stage 3 disease more common in Class 2 (60.4%) than in Class 1 (39.1%, p < 0.001). Although IDC was the predominant histological type in both clusters, it was slightly more prevalent in Class 2 (91.9%) than in Class 1 (86.7%, p < 0.001). Family history patterns also showed marked differences. Class 1 patients had a higher mean number of total cancers among relatives (4.15 ± 2.749) compared to Class 2 (2.08 ± 1.883, p < 0.001). Treatment patterns varied as well: Class 2 patients were more likely to receive radiotherapy (51.5% vs. 37.8%, p = 0.003) but less likely to undergo surgery (27.3% vs. 96.9%, p < 0.001). Importantly, survival outcomes were better in Class 2, with 98.8% of patients alive compared to 90.0% in Class 1 (p < 0.001). No significant differences were observed between the groups regarding menopausal status (p = 0.826) or age distribution (p = 0.375). Overall, these findings highlight the heterogeneity of TNBC, where Class 1 is characterized by a higher familial cancer burden, more advanced pathological stages, and poorer survival outcomes. In contrast, Class 2 exhibits less familial aggregation, improved treatment adherence, and better survival outcomes. Together, these results suggest that patients in Cluster 1—with a higher prevalence of BRCA1 + mutations, advanced disease stages, and strong family cancer histories—may benefit from more intensive treatment strategies and closer clinical monitoring. Key variables contributing to the differentiation of these clusters included family history of BC and OC, number of BC cases in the family, presence of BRCA1 + mutation, total number of cancers in the family, MLPA positivity, diagnosis type, number of first- and second-degree BC cases, clinical stage, and metastasis status, as depicted in Fig. 7.

Table 4.

Comparison of clinical, pathological, and familial variables between the two clusters identified in the analysis

		TwoStep Cluster Number		P-Value
		Class 1 N (%)	Class 2 N (%)
BRCA1	Negative Positive	142 (63.1%)	242 (93.1%)	< 0.001
		83 (36.9%)	18 (6.9%)
MLPA Positivity	Negative BRCA1 Positive	131 (58.2%)	(91.9%)	< 0.001
		94 (41.8%)	21 (8.1%)
Age 45	≤45 > 45	161(71.6%)	176 (67.7%)	0.375
		64 (28.4%	84 (32.3%)
Pathological Stages	Stage 1 Stage 2 Stage 3 Stage 4	26 (11.6%)	18 (6.9%)	< 0.001
		74 (32.9%)	84 (32.3%)
		88 (39.1%)	157 (60.4%)
		37 (16.4%)	1 (0.4%)
Histological Grades	Grade 1 Grade 2 Grade 3	3 (1.3%)	5 (1.9%)	0.494
		60 (26.7%)	58 (22.3%)
		162 (72.0%)	197 (75.8%)
Histological Type	IDC ILC IDC + ILC Other	195 (86.7%)	239 (91.9%)	< 0.001
		3 (1.3%)	4 (1.5%)
		16 (7.1%)	1 (0.4%)
		11 (4.9%)	16 (6.2%)
IDC	No Yes < 2 cm ≥2 cm	14 (6.2%)	19 (7.3%)	0.719
		(93.8%)	241 (92.7%)
Tumor Size Status		76 (33.8%)	(33.1%)	0.923
		149 (66.2%)	174 (66.9%)
Lymph Node Involvement	No Yes	102 (45.3%)	(56.5%)	0.014
		123 (54.7%)	113 (43.5%)
Chemotherapy	No Yes	14 (6.2%)	(13.1%)	0.014
		211 (93.8%)	226 (86.9%)
Radiotherapy	No Yes	(37.8%)	(51.5%)	0.003
		140 (62.2%)	126 (48.5%)
Surgery	No Yes	7 (3.1%)	71 (27.3%)	< 0.001
		(96.9%)	(72.7%)
The Number of Breast Cancers in First-Degree Relatives	0 1 2	142 (63.1%)	251 (96.5%)	< 0.001
		67 (29.8%)	9 (3.5%)
		16 (7.1%)	0 (0.0%)
The Number of BC and OC in the family	< 2 Breast Cancer and Ovarian Cancer ≥2 Breast Cancer and Ovarian Cancer	75 (33.3%)	254 (97.7%)	< 0.001
		150 (66.7%)	6 (2.3%)
Menopausal Status	Pre-menopausal Post-menopausal	174 (77.3%)	204 (78.5%)	0.826
		51 (22.7%)	56 (21.5%)
Last status	Alive Deceased	208 (90.0%)	251 (98.8%)	< 0.001
		23 (10.0%)	3 (1.2%)
		Mean ± SD	Mean ± SD
Breast Cancer in a First-Degree Relative		0.44 ± 0.625	0.03 ± 0.183	< 0.001
Breast Cancer in a Second-Degree Relative		0.46 ± 0.726	0.09 ± 0.298	< 0.001
Breast Cancer in a Third-Degree Relative		0.38 ± 0.704	0.08 ± 0.313	< 0.001
Breast Cancer in a Fourth-Degree Relative		0.24 ± 0.805	0.07 ± 0.248	< 0.001
Number of Breast Cancer		1.52 ± 1.433	0.27 ± 0.511	< 0.001
Number of Ovarian Cancer		0.37 ± 0.734	0.05 ± 0.243	< 0.001
Number of Cancer		4.15 ± 2.749	2.08 ± 1.883	< 0.001

N Number, IDC Invaziv Ductal Carcinoma, ILC Invaziv Lobular Carcinoma, BC Breast Cancer, OC Ovarian Cancer, SD Standard Deviation

Fig. 7.

Contribution of key variables to the clustering of TNBC patients. (The bar plot shows the relative importance of each variable in defining cluster membership, with family history of breast and ovarian cancer (BC and OC), number of BC cases in the family, and presence of BRCA1+ mutation emerging as the most influential factors in differentiating patient groups)

In the survival analysis using Kaplan-Meier curves (Fig. 8), BRCA1 + mutation patients demonstrated poorer overall survival compared to BRCA1- patients. At the time of analysis, a total of 18 patients (4.9%) had died, while 348 patients (95.1%) were still alive. In the BRCA1 + mutation group, 8 patients (6.7%) had died, and 111 patients (93.3%) were still alive. These results suggest that the majority of patients had not experienced the event (death) during the follow-up period, contributing to the high censoring rate observed in the Kaplan-Meier survival analysis. Although the Kaplan-Meier curves suggested that BRCA1 + mutation patients had poorer overall survival compared to BRCA1- patients, the difference was not statistically significant (log-rank test, p = 0.234). However, this trend was in line with the clustering results, which also indicated worse outcomes in the group enriched for BRCA1 + mutations. This finding is consistent with the cluster analysis results, where the cluster characterized by higher BRCA1 + mutation, advanced pathological stages, and greater familial cancer burden also exhibited poorer survival outcomes. Together, these analyses highlight the prognostic impact of BRCA1 + mutations in TNBC patients and underscore the importance of integrating genetic and clinical variables in patient stratification and management strategies.

Fig. 8.

Kaplan-Meier survival curves comparing overall survival between BRCA1+ and BRCA1– TNBC patients. (Although BRCA1+ mutation patients showed a trend toward poorer survival, the difference was not statistically significant (log-rank test, p = 0.234). Censored data points are indicated with cross marks)

Discussion

This study is among the few that specifically assess the prevalence of BRCA1 + or BRCA2 + mutations in Turkish patients diagnosed with TNBC. While earlier studies have broadly explored the spectrum of BRCA1 + or BRCA2 + mutations in BC patients in Türkiye, our work emphasizes this aggressive TNBC subtype. Furthermore, we aimed to identify clinical and demographic factors associated with these mutations to improve screening and management strategies. Using next-generation sequencing (NGS), we analyzed peripheral blood mononuclear cells (PBMCs) from 485 TNBC patients. Key clinical parameters—including age at diagnosis, tumor size, lymph node involvement, and metastasis status—were compared between mutation carriers and non-carriers. Among 485 TNBC patients, 119 (24.5%) harbored BRCA1 and/or BRCA2 + mutations, comprising 101 (20.8%) with BRCA1 + mutations and 22 (4.5%) with BRCA2 + mutations; notably, 4 patients had mutations in both genes, explaining the difference between the summed counts and the total number of unique carriers. This finding aligns with global studies reporting a predominance of BRCA1 + mutations in TNBC patients [26, 27].

BRCA1 and BRCA2 are key drivers of TNBC through the disruption of homologous recombination repair, promoting genomic instability and aggressive tumor phenotypes [28]. While these mutations are typically considered mutually exclusive due to their distinct molecular roles [29] such co-occurrence has been reported. For instance, studies have reported patients harboring both BRCA1 or BRCA2 + mutations, implying that simultaneous inactivation of both alleles may contribute to tumorigenesis [30]. Investigations into familial BC and OC have indeed revealed instances of patients harboring mutations in both BRCA1 or BRCA2+, suggesting that the accumulation of mutations contributes to the complexity of cancer predisposition [31, 32]. These rare cases challenge the prevailing understanding of the interactions between these two genes and suggest a more complex relationship in certain patient populations. In our cohort, we identified 101 patients (20.8%) with BRCA1 + mutations, 22 patients (4.5%) with BRCA2 + mutations, and, notably, four patients (0.8%) carrying pathogenic variants in both genes. This finding underscores the importance of comprehensive genetic screening and highlights the need for further investigation into the biological mechanisms, clinical behavior, and therapeutic responses of TNBC patients harboring concurrent BRCA1 + or BRCA2 + mutations, particularly in the Turkish population.

The rare co-occurrence of pathogenic BRCA1 or BRCA2 + mutations, as observed in four patients in our cohort, represents a form of trans-heterozygosity that carries distinct clinical and therapeutic implications. Studies have shown that such dual mutation status may lead to earlier disease onset and more aggressive tumor behavior, particularly in TNBC [33–35]. Mechanistically, the presence of deleterious variants in both genes may amplify genomic instability and influence cellular phenotypes, thereby affecting tumor progression and treatment response [36, 37]. This genotype may also confer heightened sensitivity to DNA-damaging agents such as cisplatin, carboplatin, and PARP inhibitors, underscoring the needfor precision treatment approaches [34, 38, 39]. All four trans-heterozygous patients harbored pathogenic variants in both genes per ACMG/AMP criteria, suggesting additive risk implications for cascade genetic testing. Given the complexity and rarity of BRCA1 + or BRCA2 + trans-heterozygosity, future studies should investigate the prognostic and therapeutic consequences of this genotype more thoroughly [40–42].

Turkish TNBC patients harboring BRCA1 + mutations exhibited distinct clinical characteristics, including younger age at diagnosis, premenopausal status, higher tumor grade, and an increased likelihood of bilateral BC. These observations are consistent with previous research emphasizing the aggressive nature of the disease in BRCA1 + mutation carriers [43, 44]. Moreover, BRCA1 + mutation carriers demonstrated a stronger family history of BC and OC, underscoring the role of genetic predisposition. The association of BRCA1 + mutations with a family history of these cancers highlights the importance of genetic screening in high-risk populations, particularly among younger patients [45, 46]. In light of these findings, the prevalence of BRCA1 + mutations observed in our Turkish TNBC cohort (20.8%) appears notably higher than the rates reported in many Western populations, which generally range between 10% and 15% [47, 48]. For instance, a large-scale study by Young et al. documented a BRCA1 + mutation frequency of 12.4% among TNBC patients in a Western population [49]. Similarly, Zhang et al. found BRCA1 + mutation frequencies ranging from 9.4 to 11.2% in unselected TNBC cohorts, reinforcing the consistent association between BRCA1 + and TNBC pathogenesis [48]. Manning et al. further emphasized that approximately 70% of BRCA1 + BCs are of the TNBC subtype, suggesting a strong genetic predisposition [50]. These differences raise the possibility that BRCA1 may play a more prominent role in TNBC tumorigenesis in the Turkish population, potentially due to regional founder effects, consanguinity patterns, or population-specific risk factors. In support of this hypothesis, our previous national study involving a Turkish TNBC cohort also demonstrated a similarly elevated BRCA1 + mutation prevalence among TNBC patients [51]. Taken together, these findings suggest that the relatively high mutation frequency observed in our cohort may reflect underlying genetic structures unique to Türkiye and underscore the need for population-tailored genetic screening and counseling protocols. In contrast, BRCA2 + mutations in our cohort did not show significant associations with clinical or pathological parameters. This lack of correlation suggests that BRCA1 focused screening strategies may be more effective in Turkish TNBC patients. BRCA2 testing, on the other hand, might be more appropriately reserved for individuals with a strong family history of BC and OC, where its clinical utility remains relevant, so BRCA2 screening in Turkish TNBC patients should be prioritized only in the presence of a strong family history.

The clinical profiles of BRCA1 + mutation carriers underline the necessity of tailored genetic counseling and testing. Identifying at-risk individuals can facilitate enhanced surveillance and preventive measures [52]. Our analysis further revealed a significant association between BRCA1 + mutations and specific clinical factors, including a family history of OC. In contrast, no significant clinical associations were observed for BRCA2 + mutations, highlighting BRCA1 + mutations as a potentially more reliable marker for identifying TNBC patients with a higher genetic predisposition [53]. These findings are particularly significant considering the poorer survival outcomes observed in BRCA1 + mutation carriers, who are more likely to present with advanced disease stages and greater lymph node involvement [54].

Logistic Regression Analysis of the Association Between Bilateral Breast Cancer, BRCA + 1 Mutations, and Clinical Risk Factors:

The relationship between bilateral BC and BRCA1 + mutations, as well as the influence of family history and menopausal status on mutation risk, plays a pivotal role in understanding the genetic predisposition to BC. Extensive research has established that bilateral BC significantly increases the likelihood of BRCA1 + mutations. Some studies report mutation frequencies as high as 12.5% among patients with bilateral BC [8]. These findings are further supported by the WECARE study, which indicates that BRCA1 + or BRCA2 + mutation carriers face a significantly higher risk of developing contralateral BC [55]. Moreover, the link between a positive family history of BC and BRCA1 + mutations is well-established, with numerous studies confirming that a family history of BC significantly increases the likelihood of carrying these mutations [56]. Our analysis further indicates that bilateral BC is significantly correlated with an increased risk of BRCA1 + mutations, with a 2.748-fold higher risk in patients diagnosed with bilateral BC. Conversely, postmenopausal status appears to be associated with a decreased risk of BRCA1 + mutations, with a 0.350-fold reduced likelihood of carrying the mutation in postmenopausal women. This finding is particularly noteworthy, as studies have shown that postmenopausal women have a reduced risk of BRCA1 + mutations, likely due to hormonal changes influencing BC development [57]. Such findings align with evidence suggesting that hormonal factors significantly contribute to the carcinogenesis of BRCA1 + associated BC [58]. Furthermore, our analysis reveals that each additional case of BC in first-degree relatives remarkably increases the risk of BRCA1 + mutations, with a 2.410-fold higher likelihood. Altogether, the results highlight the importance of family history, menopausal status, and bilateral BC as key clinical risk factors influencing the presence of BRCA1 + mutations. These insights hold significant value for enhancing genetic counseling, early detection efforts, and tailored risk management strategies for BC.

Cluster analysis

Cluster analysis provided valuable insights into the genetic and clinical heterogeneity within the Turkish TNBC cohort. Our findings revealed distinct mutation patterns, supported by evidence emphasizing the heterogeneity of BRCA1 + or BRCA2 + mutations across populations and clinical presentations [59]. Here in this study, Cluster 1 was characterized by a higher prevalence of BRCA1 + mutations, MLPA positivity, advanced cancer stages, and a strong familial cancer history, which collectively contributed to poorer survival outcomes compared to Cluster 2. This observation was further supported by Kaplan-Meier survival analysis, which demonstrated a trend toward reduced overall survival in BRCA1 + patients. Although the survival difference did not reach statistical significance (log-rank p = 0.234), the higher mortality rate in the BRCA1 + mutant group and the clustering of advanced disease characteristics suggest a clinically relevant impact. These findings underscore the potential prognostic value of BRCA1 + mutations in TNBC and highlight the importance of integrating genetic and clinical variables in patient stratification. Future studies with larger cohorts and longer follow-up periods are needed to validate these associations. These results highlight the critical role of genetic predisposition in influencing prognosisfor TNBC patients [60, 61]. Also, despite similar age distributions between the clusters, patients in Cluster 1 also exhibited higher rates of lymph node involvement and mortality, underscoring the aggressive nature of the disease in this subgroup. Specifically, for patients in Cluster 1, characterized by BRCA1 + mutations and strong familial cancer history, aggressive interventions such as adjuvant treatment with PARP inhibitors or prophylactic salpingo-oophorectomy may confer greater clinical benefit compared to patients in Cluster 2. The identification of such clusters has important clinical implications, offering a pathway to more informed decision-making and the development of personalized treatment strategies, particularly for TNBC patients with BRCA1 + mutations. However, it is important to note that the statistical power of the survival analysis may have been limited due to the high proportion of censored cases, as the majority of patients had not experienced the event (death) by the end of the follow-up period. Also, the low event rate (26 deaths) limits the statistical power; longer follow-up is needed for robust survival analysis. This result is limited by the short follow-up period and the small number of deaths (n = 26). The prognostic role of BRCA1 + should be confirmed by long-term follow-up studies.

Clinical implications

Our findings have several important implications. First, the strong association of BRCA1 + or BRCA2 + mutations with early-onset BC highlights the value of genetic screening in high-risk individuals, particularly younger patients. Second, the higher prevalence of BRCA1 + mutations in patients with a family history of BC emphasizes the importance of routine genetic testing in this subgroup. Third, the predominance of BRCA1 + mutations in Turkish TNBC patients suggests that this genetic alteration may play a pivotal role in the pathogenesis and clinical outcomes of TNBC, highlighting the need for tailored therapeutic strategies and enhanced genetic counseling. Based on our findings, we propose refined BRCA screening criteria tailored to Turkish TNBC patients: (1) diagnosis at ≤ 45 years of age, (2) presence of bilateral BC, or (3) a first-degree relative with ≥ 2 cases of BC or OC. Among patients meeting these criteria, the BRCA1 + mutation rate reaches 33.3%, according to our cohort data (see Table 1), suggesting that such targeted testing may improve cost-effectiveness while enhancing the identification of high-risk individuals for genetic counseling and preventive interventions. Also, Platinum-based chemotherapy or PARP inhibitors should be considered first in double-mutation carriers because of the DNA repair defect.

Limitations

This study has several limitations. First, the analysis was limited to Turkish TNBC patients, which may restrict the generalizability of findings to other populations. Second, as a cross-sectional study, it cannot establish causal relationships between BRCA1 + and BRCA2 + mutations and clinical outcomes. Third, although major clinical and demographic factors were considered, the potential influence of unmeasured variables on the prevalence and impact of BRCA1 + or BRCA2 + mutations cannot be ruled out. Fourth, while we observed significant associations between various clinical and familial factors and the risk of BRCA1 + and combined BRCA1 + and BRCA2 + mutations, no such significant associations were detected for BRCA2 + mutations alone. This finding may reflect the limited sample size of BRCA2 + cases in our cohort and highlights the need for larger studies to characterize potential clinical correlates of BRCA2 + mutations more robustly.

Conclusion

In conclusion, this is the largest TNBC cohort reported from Türkiye, and our findings highlight the significant role of BRCA1 + in TNBC pathogenesis in this population, providing a roadmap for personalized screening and treatment approaches. These findings underscore the crucial role of genetic screening, particularly for individuals with BRCA1 + mutations, in identifying high-risk individuals and informing personalized treatment. Future research involving larger, more diverse cohorts is essential to validate these results and further investigate the therapeutic potential of targeting BRCA1 + mutations in TNBC.

Supplementary Information

Below is the link to the electronic supplementary material.

Author contributions

B.C.D. Conceptualization, Investigation, Writing – original draft S. K. E. Data curation, Formal analysis, Writing – review and editingO.S. E. Writing – Review and editing Z. Y. K. Software O.P. Formal Analysis M. C. Ö. Investigation S. B. T. Corresponding Author, Supervision, Writing – review and editing.

Funding

This work was supported by the Istanbul University Oncology Institute&Hospital.

Data availability

The datasets generated and/or analysed during the current study are available in the [TNBC] repository, [https://zenodo.org/records/16902853].

Declarations

Ethics approval and consent to participate

This study was approved by the Ethics Committee of Istanbul University (Approval Number: 2023/500). Informed consent was obtained from all individual participants included in the study.

Consent for publication

The authors affirm that human research participants provided informed consent for the publication of anonymized data included in this manuscript.

Competing interests

The authors declare no competing interests.