Breast cancer is the most common cancer in most countries, including Thailand, and it has been classified according to gene expression profiling into 5 subtypes [1]. Immunohistochemical (IHC) markers of specific gene expression products were then proposed [1-3] with some modification in the St. Gallen consensus on the primary therapy of early breast cancer [4] to classify breast cancer as a practical basis for treatment, as follows: luminal A (lumA), luminal B (lumB), HER2+ (HER2+), and triple-negative (TN). TN subtype is subgrouped into [1] basal-like TN (TNB) when it is immunophenotypically negative ER, PR, and HER2 with at least any basal cytokeratin (basal CK; CK5, CK14, CK17) or epidermal growth factor receptor (EGFR) expression, and [2] non-basal-like TN (TNN) when it is immunophenotypically negative ER, PR, HER2, basal CK, and EGFR.

Of the 5 subtypes, TN is usually poorly differentiated, has aggressive clinical behavior, and has a poor prognosis. Moreover, therapy for TN subtype is limited due to the lack of hormone receptors, and HER2 expression of the tumor. Several studies have investigated the predictive/ prognostic factors of this breast cancer subtype, especially factors that could lead to targeted therapies.

We previously classified 100 Thai breast cancer patients recruited during 2002-2004 according to the results of IHC study and found a prevalence of TNB and TNN of 15% and 14%, respectively [5]. We found no significant difference in age, tumor size, lymphovascular invasion (LVI), nodal metastasis, microvessel density, expression of vascular endothelial growth factor (VEGF), or survival between subtypes. Moreover, we were unable to find significant clinical difference between TNB and TNN due to our small sample size. Comparison between TN and non-TN revealed significantly higher tumor grade, mitotic count expressed by Ki-67 index, p53, and vimentin expression, and decreased overall survival (OS) in TN subtype compared to non-TN subtype [6, 7].

In the current retrospective study of a larger population, we focused on the identification of TNB and TNN using biomarkers, clinical outcomes, and predictive/prognostic factors in the TN subtype.

The protocol for this study was approved by the Institutional Review Board of the Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand [COA no. R635/2553(EC3)]. We performed a retrospective study of archival formalin-fixed paraffin-embedded (FFPE) breast cancer specimens previously reported ER-, PR-, and HER2- (IHC 0/1+) or equivocal (IHC 2+) during 2007 to 2010. Cases with adequate tissue for study were enrolled, and the histology and prognostic/ predictive factors of the retrieved tumors were reviewed by 3 pathologists, TC, NS and MW.

Construction of tissue microarray (TMA) and IHC study Three cores of 2 mm diameter FFPE tissue from each case and 6 controls (breast cancer with ER+, PR+, HER2+; breast cancer with p53+; fibroadenoma; appendix; lung; placental tissue) were constructed averaging 24 cases per one recipient block. Cases with HER2 IHC2+ were embedded in a separate recipient block. Each block was cut and stained for ER, PR, and HER2, markers of basal-like subtype [three basal CKs (CK5, CK14, and CK17) and EGFR], CD117, p63, p53, vimentin, Ki67, CK7, CK8/18, CK19, BCL2, p16, WT1, and cyclin D1. Dual in situ hybridization (DISH) for HER2 was performed on the block with HER2 IHC2+ by using the Ventana HER2 Dual ISH DNA Probe Cocktail Assay. Details of the antibodies used and the criteria for interpretation are provided in Table 1.

According to the IHC results of ER, PR, HER2, basal CK, and EGFR, there were 121 TNB, 17 TNN, 42 lumB, and 14 HER2+ patients with complete follow-up data. A total of 193 female patients were clinicopathologically assessed. The clinical, histologic, and IHC studies are summarized in Tables 2 and 3.

Antibody		Total no.	TNB	TNN	LumB	HER2	P	P	P	P
		(193, 100%)	no.of cases(120, 62.2%)	no.of cases(17, 8.8%)	no.of cases(42, 21.8%)	no.of cases(14, 7.3%)	All subtypes	(TNB, TNNand lumB)	(TN vslumB)	(TN vs Non-TN)
ER	Negative		120 (100)	17 (100)	25 (59.5)	14 (100)
	Positive		0	0	17 (40.5)	0
PR	Negative		120 (100)	17 (100)	10 (23.8)	14 (100)
	Positive		0	0	32 (76.2)	0
HER2	Negative		120 (100)	17 (100)	38 (90.5)	0
	Positive		0	0	4 (9.5)	14(100)
Ki-67		45.7	46.9	41.61	49.11	30.13
(95%CI, SD)		(43.05-48.35,18.66)	(43.41-50.38,19.28)	(33.72-49.49,15.34)	(43.63-54.59,17.58)	(23.24-37.02,11.94)	0.005	0.373	0.394	0.528
Ki-67	≤14	3 (1.6)	2 (1.7)	0	0 (2.4)	1 (7.1)
	>14-30	41 (21.2)	23 (19.2)	6 (35.3)	5 (11.9)	7 (50.0)
	>30-50	72 (38.9)	44 (36.7)	5 (29.4)	18 (42.9)	5 (35.7)	0.038	0.524	0.499	0.879
	>50	77 (39.9)	51 (42.5)	6 (35.3)	19 (45.2)	1 (7.1)
CK5/6	Negative	82 (42.5)	37 (30.8)	17 (100)	16 (38.1)	12 (85.7)
	Positive	111 (57.5)	83 (69.2)	0	26 (61.9)	2 (14.3)			0.945	0.177
CK14	Negative	110 (57.0)	62 (51.7)	17 (100)	19 (45.2)	12 (85.7)
	Positive	83 (43.0)	58 (48.3)	0	23 (54.8)	2 (14.3)			0.157	0.769
CK17	Negative	93 (48.4)	48 (40.0)	16 (100)	20 (47.6)	9 (64.3)
	Positive	99 (51.6)	72 (60.0)	0	22 (52.4)	5 (35.7)			0.949	0.551
EGFR	Negative	67 (34.7)	31 (25.8)	17 (100)	12 (28.6))	8 (57.1)
	Positive	126 (65.3)	89 (74.2)	0	30 (71.4)	6 (42.9)			0.437	0.852
CD117	Negative	124 (64.2)	66 (55.0)	13 (76.5)	31 (73.8)	14 (100)
	Positive	69 (35.8)	54 (45.0)	4 (23.5)	11 (26.2)	0	0.002	0.04	0.06	0.003
p63	Negative	153 (79.3)	100 (83.3)	17 (100)	26 (61.9)	10 (71.4)
	Positive	40 (20.7)	20 (16.7)	0	16 (38.1)	4 (28.6)	0.003	0.001	0.001	0.001
p53	Negative	92 (47.7)	63 (52.5)	7 (41.2)	16 (38.1)	6(42.9)
	Positive	101 (52.3)	57 (47.5)	10 (58.8)	26 (61.9)	8 (57.1)	0.379	0.23	0.14	0.136
Vimentin	Negative	119 (61.7)	74 (61.7)	12 (70.6)	20 (47.6)	13 (92.9)
	Positive	74 (38.3)	46 (38.3)	5 (29.4)	22 (52.4)	1 (7.1)	0.02	0.151	0.066	0.618
CK7	Negative	19 (9.9)	12 (10.0)	2 (11.8)	4 (9.8)	1 (7.1)
# score ≥3	Positive	173 (90.1)	108 (90.0)	15 (88.2)	37 (90.2)	13 (92.9)	0.98	0.971	0.931	0.813
CK8/18	Negative	13 (6.8)	8 (6.7)	4 (23.5)	1 (2.4)	0 (0.0)
# score ≥3	Positive	179 (93.2)	112 (93.3)	13 (76.5)	40 (97.6)	14 (100.0)	0.02	0.017	0.172	0.084
BCL2	Negative	121 (63)	70 (58.3)	16 (94.1)	21 (51.2)	14 (100.0)
# score >0	Positive	71 (37)	50 (41.7)	1 (5.9)	20 (48.8)	0 (0.0)	<0.001	0.008	0.185	0.911
p16	Negative	28 (14.6)	14 (11.7)	4 (23.5)	6 (14.6)	4 (28.6)
# score >0	Positive	164 (85.4)	106 (88.3)	13 (76.5)	35 (85.4)	10 (71.4)	0.25	0.395	0.806	0.371
# score 3-6		83 (52.9)	58 (56.3)	7 (58.3)	11 (32.4)	7 (87.5)
# score 7-8		74 (47.1)	45 (43.7)	5 (41.7)	23 (67.6)	1 (12.5)	0.017	0.046	0.013	0.129
WT1	Negative	86 (44.8)	48 (40.0)	7 (41.2)	19 (46.3)	12 (85.7)
# score ≥3	Positive	106 (55.2)	72 (60.0)	10 (58.8)	22 (53.7)	2 (14.3)	0.013	0.776	0.48	0.041
WT1	Negative	188 (97.9)	116 (96.7)	17 (100.0)	41 (100.0)	14 (100.0)
nuclear stain	Positive	4 (2.1)	4 (3.3)	0 (0.0)	0 (0.0)	0 (0.0)	0.484	0.372	0.268	0.2
Cyclin D1
# score 0		47 (24.4)	28 (23.3)	9 (52.9)	9 (21.4)	1 (7.1)
# score >0		145 (75.1)	92 (76.7)	8 (47.1)	32 (76.2)	13 (92.9)	0.019	0.027	0.516	0.198

Abbreviations: TNB, triple negative basal-like; TNN, triple negative non-basal-like; TN, triple negative breast cancer; LumB, luminal B. #score=(intensity, 1-3)+(quantity,1-5); intensity, 1=mild, 2=intermediate, 3=strong; quantity, 0=no positive cell, 1=<1, 2=≥1-10, 3=>10-33, 4=>33-66, 5=>66-100% positive cells

The mean age at diagnosis was 52.3±13.1 years (range: 26-88 years, median: 51 years). There were 184 (95.3%) invasive ductal carcinomas, not otherwise specified (IDC, NOS), 1 invasive lobular (ILC), 5 metaplastic, 1 adenosquamous, 1 medullary-like, and 1 mucinous carcinoma with micropapillary-like feature. High-grade tumor accounted for 71.5%. Tumor size ranged from 0.5 to 19 cm (mean: 3.22±2.4 cm). Modified radical mastectomy was performed in 89 (46.1%) patients. Total mastectomy with sentinel lymph node (SNL) biopsy with or without axillary node dissection was performed in 59 (30.6%) patients. SLN was performed in 55.4%, and 9.3% were positive. Among 193 patients, 32, 103, 6, and 52 patients had stage 1, 2, 3, and 4 at diagnosis, respectively. Positive axillary nodes were found in 37.8% of patients, of which 50.8% had perinodal invasion. LVI was observed in 37.3% of all patients. Neoadjuvant chemotherapy was given to 11.4% of patients, and 81.9% received postoperative chemotherapy. Less than half of the patients (46.5%) received radiation therapy. The follow-up time ranged from 0.53 to 96.53 months (median follow-up: 62.93 months). Of those, 10.9% and 30.3% had locoregional recurrence and distant metastasis, respectively; 61.7% were alive and disease-free; 31.1% had relapse or metastasis during the course of their disease; and, 34.7% died of their disease.

Patients with TNN breast cancer had significantly older age than those with lumB breast cancer (p=0.013).

No significant difference in tumor size was found among the TNB, TNN, and lumB subtypes. LumB was associated with the lowest proportion of more than 3 axillary node involvement, while TNN was associated with the highest proportion (p=0.005), and so did TN when compared with lumB (p=0.003) (Table 2). Forty-eight patients had metastasis at the time of diagnosis. TNN had higher stages than TNB or lumB (p=0.028), and so did TN versus lumB (p=0.020).

One-third of TNN and 8.3% of TNB received neoadjuvant treatment, whereas no HER2+ received neoadjuvant chemotherapy. Of 174 patients with known history of chemotherapy treatment, 90.8% received neoadjuvant and/or postoperative adjuvant chemotherapy, including 92.5% of TNB, 82.4% of TNN, 86.8% of lumB, and all HER2+. Of those, 81.6%, 78.6%, 84.8%, and 84.6% received anthracyclines either singly or in combination with others, respectively. TN had a significantly higher recurrence rate than non-TN.

IHC of biomarkers according to the 4 reclassified subtypes are summarized in Table 3. ER, PR, and HER2 of lumB were expressed in 40.5%, 76.2%, and 9.5%, respectively.

Basal CKs: Both TNB and lumB contained basal CKs (CK 5/6, CK14, or CK17) in a significant number. They were found in 35.7% of HER2+. Among these, CK5/6 had greater sensitivity (70.0%) for detecting basal-like subtype when compared with other CKs. When all three were used, 81.7% of TNB could be diagnosed.

EGFR: This protein was detected in nearly equal proportions in TNB and lumB (74.2% and 71.4%, respectively), and in less than half of HER2+. It was insignificantly more prevalent than CK5/6 (18.3% of TNB and 9.5% of lumB had positive EGFR alone). With the use of CK5/6 and EGFR, 98.3% of TNB could be detected.

CD117: The distribution of CD117 expressions was significantly different with no expression in HER2+, low frequency in TNN (23.5%), and higher frequency in lumB and TNB (26.2% and 45.0%, respectively).

p63: This protein had low sensitivity (15.8% positive) when compared with basal CKs for detecting TNB. In addition, it was not expressed in TNN. It was present in all 5 metaplastic and adenosquamous carcinomas, and in one-third of lumB; however, it declined significantly when TN (TNB and TNN) subtype was compared with lumB (13.9% vs. 38.1%, respectively).

Vimentin: This marker was expressed in only 10% of HER2 (p=0.020) while nearly 30-50% of tumors in the three remaining subtypes.

p53: This protein was expressed in more than 50% of tumors in all subtypes without significant difference in distribution.

Ki-67 index: Prolonged storage of tissue caused diminished staining, so repeat staining with increased concentration was performed. Ki67 index was high in all subtypes with a median value of 45%, except for the HER2+ subtype, which had a significantly lower Ki67 index (30.13%, p=0.005).

CD34: There was no expression of this protein in tumor cells, including spindle cells in metaplastic carcinoma. No stromal cell staining was observed.

CK7: Only 5 patients had no expression of this marker. Expression was similar in all subtypes with approximately 90% positive when using a cutoff score of 3.

CK8/18: Only 3 patients had no CK8/18 expression. Expression among subtypes (score of at least 3) was significantly different with highest expression of 100% in HER2+, and lowest expression of 76.5% in TNN subtypes. Three patients had no CK7 expression, but had CK8/18 expression.

CK19: Six patients had no CK19 expression. One of them had negative CK7 expression, and two had neither CK7 nor CK8/18 expression. All patients with negative CK19 also had negative BCL2. (data not shown in Table 3) BCL2: There was no BCL2 expression in 63%, and less than 10% expression in 20.8% of patients. HER2+ subtype had no expression regardless of cutoff score. When using a cutoff score of less than 10%, TNN subtype also had no expression.

p16: There was no expression in 14.6%, and diffuse strong expression in 47.1% of all patients. There was no significant difference in high p16 expression between TNB and TNN. Diffuse strong expression was less in HER2+ subtype, and was significantly increased in lumB when compared with those with non-diffuse staining (Table 3, p16 score 3-6 vs. score 7-8).

Cyclin D1: All types with any positive tumor cell ranged from 47.1% in TNN to 92.9% in HER2+ subtype. Only 2.1% of all cases were strongly positive.

WT1: HER2+ subtype had the lowest cytoplasmic expression of WT1 (14.3% when using score 3 as the cutoff), whereas TNB, TNN, and lumB subtypes had a similar proportion of positive WT1. Localization of WT1 in the nucleus was observed in only 4 patients belonging to the TNB group, and all of them died of their disease. Negative BCL2 was found in these 4 patients, and 3 of them had negative p16, luminal CKs, or cyclin D1. There was no association between cytoplasmic/cell membrane staining intensity or percentage of staining and nuclear staining of WT1.

Ki67 index was not significantly different between the TN and non-TN groups. Increased CD117 expression was significantly found in TN, while p63 expression was more common in non-TN.

There were 10 patients with distant metastasis at the time of diagnosis, so only 116 TNB, 14 TNN, 40 lumB, and 13 HER2 were analyzed for DFS. The mean follow- up time was 72.44±2.61 months. The median follow-up time was 62.93 months (range: 0.53-96.5 months). Loco- regional recurrence occurred in 21 patients, and 53 patients had distant metastasis. Sixty-six deaths occurred during the follow-up period (Table 2).

The clinical parameters significantly associated with lower DFS and OS were tumor size larger than 5 cm, grade 3 tumor, and presence of axillary nodal involvement. Furthermore, patients with higher stage had significantly lower OS (Table 4).

Of 45 patients, 10 of 37 TNB, 1 of 2 TNN, 0 of 5 lumB, and 1 HER2+ had recurrence within 5 years, and 9 of 37 TNB, 1 of 2 TNN, 0 of 5 lumB, and 1 HER2+ died within 5 years (p=0.191 and p=0.019, respectively). However, the results of this analysis should be interpreted with caution due to the small number of subjects.

Of 145 patients with exclusion of stage 4 breast cancer, 37.8 % of 82 TNB, 69.2% of 13 TNN, 37.8% of 37 lumB, and 46.2% of 13 HER2+ had recurrence within 5 years, and of 148 patients, 33.7% of 83 TNB, 66.3% of 15 TNN, 32.4% of 37 lumB, and 46.2% of 13 HER2+ died within 5 years. TNN had significantly lower DFS and OS when compared with TNB or lumB (p=0.027 and p=0.021, respectively). No significant difference in DFS or OS was observed between TNB and lumB.

TN patients and overall patients in our cohort who received chemotherapy had significantly increased DFS, but not significantly increased OS (all patients p=0.028 and p=0.121, respectively, TN patients p<0.001 and p=0114, respectively). Among the overall cohort, anthracycline-based alone or in combination with taxane was used in 81% of patients who received neoadjuvant chemotherapy, in 77.8% who received postoperative chemotherapy, and in 82.3% who received neoadjuvant and postoperative chemotherapy (81.6% of TNB, and 78.6% of TNN) (Tables 2 and 4).

Patients who received radiotherapy had significantly lower DFS and OS (p=0.03 and p=0.011, respectively). In TN subtype, receiving radiotherapy did not result in significantly different DFS or OS. However, patients with TN breast cancer who underwent lumpectomy and received subsequent radiotherapy had significantly increased DFS (p=0.029). In multivariate analysis, patients who received radiotherapy had borderline significantly increased DFS, but not OS (adjusted for age, types of surgery, and stages).

Overall, multivariate analysis by Cox linear regression model showed that among clinicopathological parameters, tumor size and axillary nodal involvement were the independent prognostic factors for DFS (hazard ratio [HR]: 1.154, 95% confidence interval [CI]: 1.030-1.293, p=0.013, and HR: 2.497, 95% CI: 1.411-4.420, p=0.002, respectively), and for OS (HR: 1.162, 95% CI: 1.084- 1.245, p<0.001, and HR: 2.628, 95% CI: 1.488-4.643, p=0.001, respectively). In TNN subtype, CK8/18 was the independent predictor for better DFS and OS (HR: 0.040, 95% CI: 0.002-0.758, p=0.032, and HR: 0.090, 95% CI: 0.011-0.717, p=0.023, respectively). In TN subtype, positive CK8/18 (score >0) and positive EGFR (score >0) were the independent predictors for DFS (HR: 0.045, 95% CI: 0.005-0.405, p=0.006, and HR: 0.491, 95% CI: 0.262- 0.921, p=0.027, respectively). CK8/18 (score >0) and positive EGFR (score >0) were the independent predictors for OS (HR: 0.060, 95% CI: 0.007-0.512, p=0.010, and HR: 0.489, 95% CI: 0.263-0.909, p=0.024, respectively).

This study, which had a larger sample size than our previous study, found 11.9% TN, of which 87.7% were TNB. This is quite different from the previous study that reported 25% TN, of which 56% were TNB [5]. This difference between studies may be explained by the larger size of our study cohort, differences in criteria for evaluating predictors or markers, and different clones of antibodies. The current positive criteria used were for the most part similar to those used in studies of IHC markers for basal-like breast cancer against a gene expression profile gold standard [10-12].

The prevalence of TNB over TNN in the current study was quite high (87.7%). With comparable thresholds of biomarker expression, this was similar to the finding of Taliano, et al. [13] (89.1%), lower (80.6% of 352 TN cases) than the finding of Levva, et al. [12], and lower (81.06% of 391 TN cases) than the finding of Prat, et al. [14]. We could find significant differences in DFS or OS between TNB and TNN, even with increased survival in the former. These findings were in agreement with those from a study by Choi, et al. [15], but contrary to the findings of other studies [16, 17]. The better prognosis of TNB could be related to its good response to chemotherapy [14] and its heterogeneity of disease [18]. This might suggest the need to differentiate the TNN from the TNB subgroup.

TNN also had the poorest prognosis among the four subtypes/subgroups (TNB, TNN, lumB, and HER2+), which was also reported by Choi, et al. [15]. Most lumB in our study had high Ki67 index, and approximately 10% of those had HER2 amplification. These findings may be explained by the archived pathology report diagnosis of TN breast cancer that was then revised to low positive ER or PR after our review. These types of lumB breast cancer have some phenotypes of TN breast cancer (high grade nuclei, high Ki67 index) that are considered to be subgroup with poor prognosis close to the TN cancer.

Regarding clinicopathological factors, the patients with TNN were significantly older than lumB breast cancer patients, and there was a trend toward TNN being older than TNB. Most patients in our cohort had high histologic grade, and all TNN were IDC, NOS whereas non-IDC, NOS including metaplastic and medullary-like carcinomas, were TNB subtype. There was no significant difference in tumor size, LVI, lymph node metastasis, or perinodal invasion among subtypes, except for TNN, which had a higher proportion of more than 3 metastatic axillary lymph nodes compared to lumB, and had the least perinodal extension. TNN also had higher staging (stages 3 and 4) and a higher death rate when compared with others. The higher staging of TNN could be one of the explanations for the observed worse prognosis.

Regarding the clinical prognostic factors, the commonly known adverse prognostic factors, tumor size larger than 5 cm and presence of axillary nodal involvement, were also the independent predictors for lower DFS and/or OS in this study. The presence of LVI or higher stage was associated with decreased OS.

Maximal tumor size was an adverse prognostic factor for both DFS and OS by univariate analysis in TNN if the tumor was more than 5 cm; however, in TNB, it only adversely affected OS.

The authors gratefully acknowledge Chomkwan Atthapoch, Rapeephan Cholkate, and Jiranun Sangsakul for preparation of the tissue microarray; Chomkwan Atthapoch for slide sectioning; Siwaporn Klamkhlai for immunohistochemistry staining; Tiranan Talalag and Phensri Niamyim for manuscript preparation; and, Suthipol Udompunthurak of the Research Group and Research Network Division, Research Department, Faculty of Medicine Siriraj Hospital, Mahidol University for his assistance with some aspects of the statistical analysis.

Conflict of interest declaration

All authors declare no personal or professional conflicts of interest, and no financial support from the companies that produce and/or distribute the drugs, devices, or materials described in this report.

Funding disclosure

This study was supported by a Siriraj Grant for Research Development from the Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand (grant no. COA no. R635/2553(EC3).

17 6797 08 747-752 406 2000 10.1038/35021093 Perou C. M. Sørlie T. Eisen M. B. Rijn M. Jeffrey S. S. Rees C. A. Pollack J. R. et al Nature Molecular portraits of human breast tumours 11 19 09 10869-10874 98 2001 10.1073/pnas.191367098 Sørlie T. Perou C. M. Tibshirani R. Aas T. Geisler S. Johnsen H. Hastie T. et al Proceedings of the National Academy of Sciences of the United States of America Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications 02 18 09 10393-10398 100 2003 10.1073/pnas.1732912100 Sotiriou C Neo SY McShane LM Korn EL Long PM Jazaeri A Martiat P et al Proceedings of the National Academy of Sciences of the United States of America Breast cancer classification and prognosis based on gene expression profiles from a population-based study 8 08 1736-1747 22 2011 10.1093/annonc/mdr304 Goldhirsch A. Wood W. C. Coates A. S. Gelber R. D. Thürlimann B. Senn H.-J. Annals of Oncology: Official Journal of the European Society for Medical Oncology Strategies for subtypes--dealing with the diversity of breast cancer: highlights of the St. Gallen International Expert Consensus on the Primary Therapy of Early Breast Cancer 2011 3 1187-1192 15 2014 10.7314/apjcp.2014.15.3.1187 Chuangsuwanich T Pongpruttipan T O-Charoenrat P Komoltri C Watcharahirun S Sa-Nguanraksa D Asian Pacific journal of cancer prevention: APJCP Clinicopathologic features of breast carcinomas classified by biomarkers and correlation with microvessel density and VEGF expression: a study from Thailand 5 05 739-746 139 2013 10.1007/s00432-013-1376-6 Yamashita N Tokunaga E Kitao H Hisamatsu Y Taketani K Akiyoshi S Okada S et al Journal of Cancer Research and Clinical Oncology Vimentin as a poor prognostic factor for triple-negative breast cancer 2 e0172324 12 2017 10.1371/journal.pone.0172324 Pan Y Yuan Y Liu G Wei Y PloS One P53 and Ki-67 as prognostic markers in triple-negative breast cancer patients 01 16 06 2784-2795 28 2010 10.1200/JCO.2009.25.6529 Hammond MEH Hayes DF Dowsett M Allred DC Hagerty KL Badve S Fitzgibbons PL et al Journal of Clinical Oncology: Official Journal of the American Society of Clinical Oncology American Society of Clinical Oncology/College Of American Pathologists guideline recommendations for immunohistochemical testing of estrogen and progesterone receptors in breast cancer 01 31 11 3997-4013 31 2013 10.1200/JCO.2013.50.9984 Wolff AC Hammond MEH Hicks DG Dowsett M McShane LM Allison KH Allred DC et al Journal of Clinical Oncology: Official Journal of the American Society of Clinical Oncology Recommendations for human epidermal growth factor receptor 2 testing in breast cancer: American Society of Clinical Oncology/College of American Pathologists clinical practice guideline update 01 10 04 1684-1691 28 2010 10.1200/JCO.2009.24.9284 Voduc KD Cheang MCU Tyldesley S Gelmon K Nielsen TO Kennecke H Journal of Clinical Oncology: Official Journal of the American Society of Clinical Oncology Breast cancer subtypes and the risk of local and regional relapse 11 11 1438-1450 26 2013 10.1038/modpathol.2013.97 Won JR Gao D Chow C Cheng J Lau SYH Ellis MJ Perou CM Bernard PS Nielsen TO Modern Pathology: An Official Journal of the United States and Canadian Academy of Pathology, Inc A survey of immunohistochemical biomarkers for basal-like breast cancer against a gene expression profile gold standard 03 3 05 181-195 14 2017 10.21873/cgp.20030 Levva S Kotoula V Kostopoulos I Manosou K Papadimitriou C Paradopoulou K Lakis S et al Cancer Genomics & Proteomics Prognostic Evaluation of Epidermal Growth Factor Receptor (EGFR) Genotype and Phenotype Parameters in Triple-negative Breast Cancers 12 12 2743-2750 44 2013 10.1016/j.humpath.2013.07.021 Taliano RJ Lu S Singh K Mangray S Tavares R Noble L Resnick MB Yakirevich E Human Pathology Calretinin expression in high-grade invasive ductal carcinoma of the breast is associated with basal-like subtype and unfavorable prognosis 18 12 303 13 2015 10.1186/s12916-015-0540-z Prat A Fan C Fernández A Hoadley KA Martinello R Vidal M Viladot M et al BMC medicine Response and survival of breast cancer intrinsic subtypes following multi-agent neoadjuvant chemotherapy 23 09 507 10 2010 10.1186/1471-2407-10-507 Choi YL Oh E Park S Kim Y Park Y Song K Cho EY et al BMC cancer Triple-negative, basal-like, and quintuple-negative breast cancers: better prediction model for survival 24 07 134 7 2007 10.1186/1471-2407-7-134 Tischkowitz M Brunet JS Bégin LR Huntsman DG Cheang MCU Akslen LA Nielsen TO Foulkes WD BMC cancer Use of immunohistochemical markers can refine prognosis in triple negative breast cancer 01 7 04 2302-2310 15 2009 10.1158/1078-0432.CCR-08-2132 Rakha EA Elsheikh SE Aleskandarany MA Habashi HO Green AR Powe DG El-Sayed ME et al Clinical Cancer Research: An Official Journal of the American Association for Cancer Research Triple-negative breast cancer: distinguishing between basal and nonbasal subtypes 7 07 2750-2767 121 2011 10.1172/JCI45014 Lehmann BD Bauer JA Chen X Sanders ME Chakravarthy AB Shyr Y Pietenpol JA The Journal of Clinical Investigation Identification of human triple-negative breast cancer subtypes and preclinical models for selection of targeted therapies 15 16 08 5367-5374 10 2004 10.1158/1078-0432.CCR-04-0220 Nielsen TO Hsu FD Jensen K Cheang M Karaca G Hu Z Hernandez-Boussard T et al Clinical Cancer Research: An Official Journal of the American Association for Cancer Research Immunohistochemical and clinical characterization of the basal-like subtype of invasive breast carcinoma 2 07 317-328 116 2009 10.1007/s10549-008-0206-z Viale G Rotmensz N Maisonneuve P Bottiglieri L Montagna E Luini A Veronesi P et al Breast Cancer Research and Treatment Invasive ductal carcinoma of the breast with the "triple-negative" phenotype: prognostic implications of EGFR immunoreactivity 6 06 4243-4252 36 2015 10.1007/s13277-015-3061-7 Bouchalova K Svoboda M Kharaishvili G Vrbkova J Bouchal J Trojanec R Koudelakova V et al Tumour Biology: The Journal of the International Society for Oncodevelopmental Biology and Medicine BCL2 is an independent predictor of outcome in basal-like triple-negative breast cancers treated with adjuvant anthracycline-based chemotherapy 7 07 1093-1097 33 2009 10.1097/PAS.0b013e31819c1c93 Collins LC Martyniak A Kandel MJ Stadler ZK Masciari S Miron A Richardson AL Schnitt SJ Garber JE The American Journal of Surgical Pathology Basal cytokeratin and epidermal growth factor receptor expression are not predictive of BRCA1 mutation status in women with triple-negative breast cancers 357485 2015 2015 10.1155/2015/357485 Zhang L Fang C Xu X Li A Cai Q Long X BioMed Research International Androgen receptor, EGFR, and BRCA1 as biomarkers in triple-negative breast cancer: a meta-analysis 15 8 04 2670-2674 10 2004 10.1158/1078-0432.ccr-03-0114 Woelfle U Sauter G Santjer S Brakenhoff R Pantel K Clinical Cancer Research: An Official Journal of the American Association for Cancer Research Down-regulated expression of cytokeratin 18 promotes progression of human breast cancer 1 38-46 24 2009 10.1177/172460080902400106 Vora HH Patel NA Rajvik KN Mehta SV Brahmbhatt BV Shah MJ Shukla SN Shah PM The International Journal of Biological Markers Cytokeratin and vimentin expression in breast cancer 404 8 2014 10.3332/ecancer.2014.404 Aiad HA Samaka RM Asaad NY Kandil MA Shehata MA Miligy IM Ecancermedicalscience Relationship of CK8/18 expression pattern to breast cancer immunohistochemical subtyping in Egyptian patients 24 53 11 8590-8607 22 2003 10.1038/sj.onc.1207102 Cory S Huang DCS Adams JM Oncogene The Bcl-2 family: roles in cell survival and oncogenesis 524308 2012 2012 10.1155/2012/524308 Tzifi F Economopoulou C Gourgiotis D Ardavanis A Papageorgiou S Scorilas A Advances in Hematology The Role of BCL2 Family of Apoptosis Regulator Proteins in Acute and Chronic Leukemias 1 02 141-148 150 2015 10.1007/s10549-015-3305-7 Seong MK Lee JY Byeon J Sohn YJ Seol H Lee JK Kim EK Kim HA Noh WC Breast Cancer Research and Treatment Bcl-2 is a highly significant prognostic marker of hormone-receptor-positive, human epidermal growth factor receptor-2-negative breast cancer 1 03 54-64 20 2017 10.4048/jbc.2017.20.1.54 Hwang K Han W Kim J Moon H Oh S Song YS Kim YA Chang MS Noh D Journal of Breast Cancer Prognostic Influence of BCL2 on Molecular Subtypes of Breast Cancer 4 424-432 59 2012 10.4149/neo_2012_055 Kolacinska A. Chalubinska J. Zawlik I. Szymanska B. Borowska-Garganisz E. Nowik M. Fendler W. et al Neoplasma Apoptosis-, proliferation, immune function-, and drug resistance- related genes in ER positive, HER2 positive and triple negative breast cancer 12 12 12255-12263 35 2014 10.1007/s13277-014-2534-4 Choi JE Kang SH Lee SJ Bae YK Tumour Biology: The Journal of the International Society for Oncodevelopmental Biology and Medicine Prognostic significance of Bcl-2 expression in non-basal triple-negative breast cancer patients treated with anthracycline-based chemotherapy 01 11 06 2568-2579 130 2012 10.1002/ijc.26271 Arima Y Hayashi N Hayashi H Sasaki M Kai K Sugihara E Abe E et al International Journal of Cancer Loss of p16 expression is associated with the stem cell characteristics of surface markers and therapeutic resistance in estrogen receptor-negative breast cancer 2 02 204-213 27 2014 10.1038/modpathol.2013.137 Bogina GS Lunardi G Marcolini L Brunelli M Bortesi L Marconi M Coati F et al Modern Pathology: An Official Journal of the United States and Canadian Academy of Pathology, Inc P16 but not retinoblastoma expression is related to clinical outcome in no-special-type triple-negative breast carcinomas 16 17 08 8022-8026 91 1994 10.1073/pnas.91.17.8022 Musgrove E. A. Lee C. S. Buckley M. F. Sutherland R. L. Proceedings of the National Academy of Sciences of the United States of America Cyclin D1 induction in breast cancer cells shortens G1 and is sufficient for cells arrested in G1 to complete the cell cycle 20 18 06 4215-4224 23 2005 10.1200/JCO.2005.05.064 Arnold A Papanikolaou A Journal of Clinical Oncology: Official Journal of the American Society of Clinical Oncology Cyclin D1 in breast cancer pathogenesis 11 e0188068 12 2017 10.1371/journal.pone.0188068 Ortiz AB Garcia D Vicente Y Palka M Bellas C Martin P PloS One Prognostic significance of cyclin D1 protein expression and gene amplification in invasive breast carcinoma 10 18656-18664 8 2015 Guo L Liu S Jakulin A Yilamu D Wang B Yan J International Journal of Clinical and Experimental Medicine Positive expression of cyclin D1 is an indicator for the evaluation of the prognosis of breast cancer 3 08 667-678 164 2017 10.1007/s10549-017-4294-5 Ahlin C Lundgren C Embretsén-Varro E Jirström K Blomqvist C Fjällskog M Breast Cancer Research and Treatment High expression of cyclin D1 is associated to high proliferation rate and increased risk of mortality in women with ER-positive but not in ER-negative breast cancers 20 3 03 415-418 98 2002 10.1002/ijc.10151 Umekita Y Ohi Y Sagara Y Yoshida H International Journal of Cancer Overexpression of cyclinD1 predicts for poor prognosis in estrogen receptor-negative breast cancer patients 3 02 472-480 62 2013 10.1111/his.12013 Mylona E Tzelepis K Theohari I Giannopoulou I Papadimitriou C Nakopoulou L Histopathology Cyclin D1 in invasive breast carcinoma: favourable prognostic significance in unselected patients and within subgroups with an aggressive phenotype 4 10 1231-1236 28 2012 10.3892/or.2012.1906 Qi X Zhang F Yang X Fan L Zhang Y Liang Y Ren L et al Oncology Reports High Wilms' tumor 1 mRNA expression correlates with basal-like and ERBB2 molecular subtypes and poor prognosis of breast cancer 27 03 45255 7 2017 10.1038/srep45255 Artibani M Sims AH Slight J Aitken S Thornburn A Muir M Brunton VG et al Scientific Reports WT1 expression in breast cancer disrupts the epithelial/mesenchymal balance of tumour cells and correlates with the metabolic response to docetaxel 5 05 745-750 117 2002 10.1309/QLV6-HH0H-UCTF-WEF6 Lee BH Hecht JL Pinkus JL Pinkus GS American Journal of Clinical Pathology WT1, estrogen receptor, and progesterone receptor as markers for breast or ovarian primary sites in metastatic adenocarcinoma to body fluids 27 12 314 11 2013 10.1186/1477-7819-11-314 Choi EJ Yun JS Jeon EK Won HS Ko YH Kim SY World Journal of Surgical Oncology Prognostic significance of RSPO1, WNT1, P16, WT1, and SDC1 expressions in invasive ductal carcinoma of the breast 5 05 1167-1171 8 2002 Miyoshi Y Ando A Egawa C Taguchi T Tamaki Y Tamaki H Sugiyama H Noguchi S Clinical Cancer Research: An Official Journal of the American Association for Cancer Research High expression of Wilms' tumor suppressor gene predicts poor prognosis in breast cancer patients 18 12 3355-3377 48 2012 10.1016/j.ejca.2012.10.004 Cardoso F Loibl S Pagani O Graziottin A Panizza P Martincich L Gentilini O et al European Journal of Cancer (Oxford, England: 1990) The European Society of Breast Cancer Specialists recommendations for the management of young women with breast cancer 01 8 08 1634-1657 29 2018 10.1093/annonc/mdy192 Cardoso F. Senkus E. Costa A. Papadopoulos E. Aapro M. André F. Harbeck N. et al Annals of Oncology: Official Journal of the European Society for Medical Oncology 4th ESO-ESMO International Consensus Guidelines for Advanced Breast Cancer (ABC 4)† 11 20 11 1938-1948 363 2010 10.1056/NEJMra1001389 Foulkes WD Smith IE Reis-Filho JS The New England Journal of Medicine Triple-negative breast cancer 67 72 17 2011 10.5455/GMJ-30-2011-34 Celaletdin C Kalender ME Paydas S et al Gaziantep Med J Prognostic significance of Wilms Tumor 1 (WT1) protein expression in breast cancer