Epidemiological Study of Triple-Negative Breast Cancer Patients in North Indian Population: a Hospital-Based Study

Abstract

Triple-negative breast cancer (TNBC) accounts for 10–25% of all breast tumors. This makes it more difficult to treat, so triple-negative cancers often require targeted therapies. We studied the prevalence of TNBC in a hospital-based study and compared the clinicopathological characteristics of triple-negative and non-triple-negative breast tumors. One hundred three patients were included in the study that underwent modified radical mastectomy. The procedure of immunostaining was performed using formalin-fixed, paraffin-embedded tissue sections. These sections were stained immunohistochemically for ER, PR, and HER2 neu by using ready-to-use monoclonal antibody detection system with 3′-3′ diaminobenzidine hydrochloride (DAB) as chromogen. Of all 103 patients, 35 (34%) were triple negative. The average age of patients of TNBC and non-TNBC group was found as 44.16 and 40.73 years, respectively. Patients of post-menopausal state were higher than premenopausal in TNBC (22/35; 62%) and non-TNBC groups (45/68; 66%). Further, TNBC patients reported at clinically early stages I and II (18/35; 51.4%) while non-TNBC patients predominantly reported at later stages III and IV (44/68; 64.7%). It was also observed that breast tumor size in majority of the patients in both groups lies between 2 to 5 cm (TNBC = 23/35; 65.7% and non-TNBC = 35/68; 51.5%). Lymph node metastases were present in 51.5% (18/35) cases in TNBC patients and 64.7% (44/68) cases in non-TNBC group. Despite the limitation of less number of breast cancer cases, we analyzed that TNBC tumors have aggressive clinical values than non-TNBC, though having no statistically significant difference between the prognostic clinical parameters of two groups.

Introduction

Breast cancer is the most common cancer among women. Its incidence is 25% of all cancers. (Ferlay J et al. 2015) [1]. Molecular profiling has provided biological evidence for heterogeneity of breast cancer through identification of intrinsic subtypes. These subtypes consist of estrogen receptor (ER) positive types (Luminal A and Luminal B) and ER negative types (HER 2 expressing, basal-like and normal breast-like) [2, 3]. These are shown to have distinct clinical presentation, different clinical outcomes, and differing prognosis. As per study, there is estimated rise of 100,000 new breast cancer patients in India and with this rise, India has become the country with largest number of death worldwide due to breast cancer [4–6]. In a population-based cancer registry data, Bangalore ranks the top most position in India in patients of breast cancer [7].

Two different studies explained that complementary DNA microassay profiling has been identified and explained the various subtypes of breast cancer [2, 8]. These are based on consideration that two distinct types of epithelial cells are found in human mammary glands: basal (and/or myoepithelial) cells and luminal epithelial cells. The term “basal-like cancer” and triple-negative cancer have been used interchangeably by many authors; however, they are not synonymous, approximately 75% of basal-like cancer is considered as triple negative but 25% of them may express HER-2 neu or HRs [2]. Triple-negative breast cancer (TNBC) refers to any breast cancer which does not show the expression of estrogen receptor (ER), progesterone receptor (PR), and HER2/neu. TNBC accounts for 10–25% of all breast carcinomas [9, 10]. This makes it more difficult to treat since most chemotherapy targets one of these receptors, so triple-negative cancers often require targeted therapies. Treatment decisions in this are based mainly on prognostic and predictive factors which include age of patient, status of hormone receptors (HR), human epidermal growth receptor factor, and involvement of lymphatic and lymphovascular invasion. The main characteristics of TNBC are as follows: they are frequent in early aged women (<50 years), they show weak association between tumor size and lymph node metastases, and highly aggressive and proliferative, having higher chances of brain metastases and shorter survival rates of the patients when compared with other subtypes of breast cancer [8]. Moreover, there is consistent trend across studies conforming unfavorable clinical outcomes associated to the triple-negative (TN) phenotype or basal-like breast cancer (BLBC). Previous studies shows that TNBC has aggressive clinical and pathologic features including onset at young age, diagnosis at advance age, high histopathologic and nuclear grade, high mitotic index, higher frequency of unfavorable histopathology, and most distant recurrence [10]. In Asian population, the clinical data on TNBC are limited.

This study is being done to investigate the prevalence of TNBC in study of North Indian population and further we aimed to compare various clinicopathological characteristics of TNBC with non-TNBC.

Materials and Methods

The study was carried out in the Department of General Surgery in collaboration with the Department of Pathology, Institute of Medical Sciences, Banaras Hindu University, Varanasi,

India, from January 2013 to December 2015. Informed consent was taken from all patients and the study was approved by the Institute’s Ethics Committee.

This is a prospective study carried out on all proven cases of carcinoma breast attending a single surgical unit during the study period. One hundred three patients were included in the study that underwent modified radical mastectomy and specimen was sent for histopathology. The demographic and clinical variables included age, residential area (rural or urban), oral contraceptive intake, smoking, alcohol and tobacco addiction, menstrual status, parity, pain in breast tumor, laterality, and clinical stage were noted.

Immunohistochemistry

The procedure of immunostaining procedures were performed using formalin-fixed, paraffin-embedded tissue sections. These sections were stained immunohistochemically for estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2 neu) by using ready to use monoclonal antibody and HRP polymer detection system with 3′-3′ diaminobenzidine hydrochloride (DAB) as chromogen. All the cases were analyzed using light microscope for both H&E and IHC slides and intensity of nuclear immunostaining was semi quantitatively assessed.

ER and PR results that were assessed manually were considered positive when >1% of tumor cells showed positive nuclear staining and negative if <1% nuclei were immunostained [11]. Assessment of HER2 neu was scored from 0 to 3+ using Hercept test guideline into following different categories: 0—no immunostaining; 1—weak membranous staining; 2—membranous staining, either weak or non-uniform in at least 10% of tumor cells; 3—intense membranous staining in >10% of tumor cells (0–1+ = negative, 2+ = borderline and 3+ is considered as positive) [12]. The pathological variables included tumor type, modified Bloom–Richardson grade of tumor, lymph node metastasis, and lymphovascular invasion. Patients with histopathology showing benign tumors, incomplete surgical details and those patients in whom no/incomplete information about ER/PR and HER-2 neu status was available were excluded from the present study. Human epidermal growth factor receptor-2 neu score of 3+ was taken as positive by immunohistochemistry method.

Statistical Analysis

The statistical analysis was performed by using SPSS, version 16.0. The percentages and the frequencies of all the variables were computed to compare the characteristics of the TNBC and the non-TNBC of the breast tumors using chi-square test, and univariate logistic regression analysis was done to identify the prognostic factors associated with TNBC with respect to non-TNBC. p < 0.05 was considered statistically significant.

Results

Out of the total patients of breast cancer, 34% patients showed triple negativity. The differences between TNBC and non-TNBC based on clinicopathological parameters have been summarized (Table 1). All the patients from both groups were of invasive ductal carcinoma. The average age of patient for TNBC and non-TNBC group was found as 44.16 and 40.73 years, respectively.

Table 1.

Clinicopathological Features of Triple-negative and Non Triple-negative Breast Cancer

Characteristic		Total patients N = 103		(p value)	OR (95% CI)
		Frequency (%)
		TNBC (N = 35)	Non-TNBC ((N = 68)
Average age (years) (mean ± SD)		45.5 ± 9.6	47 ± 11.31
Age (years)	≤50	25 (71.4)	45 (66.2)	0.588	1(Ref)
	>50	10 (28.6)	23 (33.8)		0.78 (0.32–1.90)
Menstrual history	Pre-menopausal	13 (37.1)	23 (33.8)	0.738	1(Ref)
	Post-menopausal	22 (62.9)	45 (66.2)		1.15 (0.49–2.70)
Laterality	Right	16 (45.7)	23 (33.8)	0.239	1(Ref)
	Left	19 (54.3)	45 (66.2)		1.64 (0.71–3.79)
Parity	≤2	6 (17.1)	23 (33.8)	0.075	1(Ref)
	>2	29 (82.9)	45 (66.2)		0.40 (0.14–1.11)
Addiction	Yes	1 (2.9)	3 (4.4)	0.699	1(Ref)
	No	34 (97.1)	65 (95.6)		0.63 (0.06–6.36)
Past history of illness	Yes	13 (37.1)	26 (38.2)	0.914	1(Ref)
	No	22 (62.9)	42 (61.8)		0.95 (0.41–2.21)
Breast pain	Yes	10 (28.6)	26 (38.2)	0.330	1(Ref)
	No	25 (71.4)	42 (61.8)		0.64 (0.26–1.56)
Chemotherapy	Neo ad.	14 (40)	37 (54.4)	0.166	1(Ref)
	Adjuvant	21 (60)	31 (45.6)		1.79 (0.79–4.06)
LN status (cN)	Negative	11 (31.4)	16 (23.5)	0.388	1(Ref)
	Positive	24 (68.6)	52 (76.5)		1.49 (0.60–3.69)
Tumor size (cm)	0.1–2	1 (2.9)	7 (10.3)	0.250	1(Ref)
	2.1–5	23 (65.7)	35 (51.5)		0.22 (.03–1.48)
	>5	11 (31.4)	26 (38.2)		0.34 (0.05–2.45)
Stage	Early	18 (51.4)	24 (35.3)	0.155	1(Ref)
	Late	17 (48.6)	44 (64.7)		1.94 (0.85–4.42)
Histological grade	I	1 (2.9)	4 (5.9)	0.787	1(Ref)
	II	7 (20)	14 (20.6)		0.50 (0.07–4.25)
	III	27 (77.1)	50 (73.5)		0.46 (0.07–3.30)
LN status (pN)	Negative	17 (48.6)	24 (35.3)	0.192	1(Ref)
	Positive	18 (51.4)	44 (64.7)		1.73 (0.75–3.96)

The frequency was higher in total population of TNBC in age group <50 years (71.4%) than that of other group (66.2%). TNBC tumors are more aggressive as compared to non-TNBC. In both TNBC and non-TNBC groups, majority of the patients were post-menopausal. When both the groups were compared, TNBC patients having more than two children were frequent (82.9%) than the non-TNBC which was statistically insignificant (p = 0.075). There was no significant association observed in patients having breast pain with any past history of surgery. Addiction to smoking, alcohol, and tobacco was not found to be associated with the disease in both the groups. Those patients who received adjuvant chemotherapy were higher in TNBC patients (60%) whereas in the other group, neoadjuvant-treated patients were higher (54.4%). It was observed that the majority of the patients in both the groups were in between 2 and 5 cm of tumor size. In TNBC patients, the frequency of early stage of cancer was higher (51%) whereas in non-TNBC patients, it was in the late stage (64.7%). In both groups, majority of the patient were of histological grade III (TNBC 77.1% and non-TNBC 73.5%). Pathological lymph node metastasis was noted in 51.5% cases of TNBC and 64.7% in non-TNBC group.

The number of patients with ER-positive, PR-positive and HER2 neu-positive was 35, 31.1, and 44.7%, respectively (Table 2). Further, we also analyzed the association of receptor expression with menopausal status as summarized in Table 3. In postmenopausal versus premenopausal women, the ratio of ER positivity is 32.8% versus 38.9%, PR positivity is 28.4 versus 36.1% and HER positivity is 46.3 versus 41.7%.

Table 2.

Receptor expression pattern in all patients

Tumor subtype	N (%)
ER positive	36 (35.0)
PR positive	32 (31.1)
HER-2 positive	46 (44.7)
Triple negative	35 (34.0)

Table 3.

Receptor expression in different menopausal groups

Receptors	Menopausal status n = 103(%)
	Pre (n = 36)	Post (n = 67)
ER
Positive	14 (38.9)	22 (32.8)
Negative	22 (61.1)	45 (67.2)
PR
Positive	13 (36.1)	19 (28.4)
Negative	23 (63.9)	48 (71.6)
HER2 neu
Positive	15 (41.7)	31 (46.3)
Negative	21 (58.3)	36 (53.7)
Triple negative
Yes	13 (36.1)	22 (32.8)
No	23 (63.9)	45 (67.2)

Discussion

At the present time, TNBC lacks any specific targeted therapy. Combined chemotherapy is the standard treatment like anthracycline, taxanes, ixabepitine, and platinum agents. This study was designed to see the difference between clinical outcomes of TNBC and non-TNBC along with incidence of TNBC patients in North Indian population. In our study, TNBC comprised 34% of the total breast cancer cases which is comparable to the study showing the prevalence of TNBC as 34.4 and 32.4% in Indian population, respectively [13, 14]. In contrast to this outcome, various studies showed different incidence of triple negativity as 19.9, 18, and 11.8%, respectively [10, 15, 16].Our study showed that in TNBC, postmenopausal patients were higher than premenopausal women which support the other studies who showed higher incidence of TNBC in postmenopausal women [17–19]. No significant difference in age was found in both two groups in TNBC and non-TNBC patients, which also reported in a previous study [20]. There are conflicting reports in literature regarding tumor size in TNBC. Various study reported the larger tumor size in TNBC than in non-TNBC patients [7, 17, 21, 22] while our study shows higher frequency of tumor size lying between 2.1–5 cm in both the groups. The most common clinical stage was stage II in TNBC which is similar to previous findings [15]. In our study, higher rate of lymph node positivity was found in TNBC (51.4%) and in non-TNBC group (64.7%), as compared with some publications which shows the same [7, 17, 21]. Although the difference was not statistically significant, some of the studies also showed a higher propensity for lymph node involvement in TNBC (54.4 and 71.3%), respectively [21, 22]. In our study, patients with TNBC mostly lies in histological grade III tumor with higher percentage than non-TNBC patients (77.1 versus 73.5%, respectively). Other studies also reported the similar results [8, 13, 16, 17, 20–24]. In the present study, the association between prognostic parameters was statistically not significant, which also agree with few other studies [15, 24, 25]. Our limited study population may be the reason behind nonsignificant results.

In conclusion, despite the limitation of less number of breast cancer cases in the present study, we analyzed that TNBC tumors are more aggressive as compared to non-TNBC tumors, though having no statistically significant difference between the prognostic clinical parameters of two groups. However, our findings highlight the need to further studies in this field which could lead to investigate new molecular biomarkers which help to develop targeted therapy for triple-negative breast cancer.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest. All procedures performed in studies involving human participants were in accordance with the Ethical Standards of the Institutional Research Committee.