Clinicopathologic significance of the basal-like subtype of breast cancer: a comparison with hormone receptor and Her2/neu-overexpressing phenotypes

DNA microarray profiling studies have led to the classification of invasive breast carcinoma into luminal/estrogen receptor-positive, normal breast-like, Her2/neu-overexpressing, and basal-like types. Among these groups, the basal-like subtype is associated with the poorest clinical outcome in Western countries. To date, the clinicopathologic characteristics of the basal-like carcinomas, compared with other subtypes, have not been described in the Korean population. In this study, we used tissue microarray to examine the expression of basal cytokeratins (CK) (CK5 and CK14) and luminal CK (CK8/18), epidermal growth factor receptor, c-kit, hormone receptors (HRs), p53, and Her2/neu in 776 consecutive patients diagnosed with invasive breast carcinoma from January 1993 to December 1998 and categorized these cases into 5 subgroups (basal-like, HR-expressing, Her2/neu-overexpressing, HR and Her2/neu-expressing, and null subtypes negative for all markers), based on the immunohistochemical data. We identified cases of 114 (14.7%) basal-like, 345 (44.5%) HR-expressing, 133 (17.1%) Her2/neu-overexpressing, 61 (7.8%) HR and Her2/neu-expressing, and 123 (15.9%) null subtypes. Histologically, most basal-like breast cancers were invasive ductal carcinoma, not otherwise specified (98 cases, 86.0%), with high nuclear and/or histologic grades, and most metaplastic carcinomas (6 [75.0%] of 8 cases) were the basal-like subtype. Both basal-like and Her2/neu-overexpressing subtypes were associated with larger tumor sizes (mean, 3.6 and 3.3 cm, respectively) than the HR-expressing group (mean, 2.8 cm) (P = .001 and P = .036, respectively). Nodal stage of Her2/neu-overexpressing subtype was higher than that of basal-like subtype; however, overall stage was not different between the 2 groups (P = .010 and .123, respectively). Distant metastasis was most frequently observed in the Her2/neu-overexpressing subtype (33.8%), which was prognostically the worst subgroup of breast cancers. In contrast to previous findings from Western countries, our analyses reveal that the Her2/neu status is the most important prognostic factor of breast cancers.     

Osteoclast-like giant cells in invasive breast cancer predominantly possess M2-macrophage phenotype

Breast carcinoma with osteoclast-like giant cells (OGCs) is a rare tumor; however, their clinicopathological aspects remain unclear. We described the clinicopathological characteristics of 8 patients with breast carcinoma with OGCs. Immuno-phenotypes of the OGCs were comparatively examined with that of foreign body giant cells (FBGCs) in 4 cases of granulomatous reaction (GR) without cancerous elements. In most cancers, tumors displayed cribriform and tubular growth patterns. Three cases showed moderate to high nuclear grade, while all the other tumors had low nuclear grade. Six patients were estrogen receptor (ER) positive, while triple negative phenotype was identified in 2 patients. During the follow-up period, 1 patient had local recurrence of the tumor, and all the patients remained alive. All OGCs and FBGCs expressed CD68, a pan-macrophage marker. OGCs in all the breast cancers showed moderate to high expression of CD163 — a marker of M2-macrophage with pro-tumoral function — whereas its expression in FBGCs was low to moderate (p = 0.04). CD86 — a marker of M1-macrophage with a tumoricidal activity — was positive in the OGCs of 3 breast cancers, and in the FBGCs of 3 GR cases (p = 0.15). The expression of CD163 was significantly higher than that of CD86 in the OGCs of breast cancer (p < 0.001), whereas they were comparable in the FBGCs of GR (p = 0.79). In summary, we found that breast carcinoma with OGCs mostly exhibited cribriform and tubular growth pattern, ER positivity, and predominantly possessed the M2-macrophage phenotype. However, the clinical significance of OGCs in breast cancer needs to be elucidated in further studies involving a larger number of cases.     

Frequency of the basal-like phenotype in African breast cancer

Basal-like breast carcinoma has been recognized as a subtype with specific prognostic implications. However, there is a lack of reports about this category of breast tumors in African women. The aim of this study was to explore the basal-like phenotype in breast cancer patients in an African population, and a registry-based series was included from the well-defined Kyadondo County in Uganda (1.7 millions). We studied a total of 65 archival paraffin blocks of invasive breast cancer using antibodies against cytokeratin 5/6 and P-cadherin, and these markers were expressed in 34% of all cases and in 52% of ER (estrogen receptor)-negative tumors. All basal-like tumors were ER negative (p<0.0005) and PR (progesterone receptor) negative (p=0.002). Basal-like breast carcinomas were of a higher histologic grade (p=0.001), had high mitotic counts (p=0.002), and marked nuclear pleomorphism (p=0.002). P-cadherin-positive tumors had a high Ki-67 proliferative rate (p=0.039). In conclusion, the basal-like phenotype is frequent in this African series of breast cancer and is strongly associated with poor prognostic factors. Our findings might be significant in relation to clinical management of these patients, including novel targeted therapy.     

Identification of a basal-like subtype of breast ductal carcinoma in situ

Microarray profiling of invasive breast carcinomas has identified subtypes including luminal A, luminal B, HER2-overexpressing, and basal-like. The poor-prognosis, basal-like tumors have been immunohistochemically characterized as estrogen receptor (ER)-negative, HER2/neu-negative, and cytokeratin 5/6-positive and/or epidermal growth factor receptor (EGFR)-positive. The aim of this study was to determine the prevalence of basal-like ductal carcinoma in situ in a population-based series of cases using immunohistochemical surrogates. A total of 245 pure ductal carcinoma in situ cases from a population-based, case-control study were evaluated for histologic characteristics and immunostained for ER, HER2/neu, EGFR, cytokeratin 5/6, p53, and Ki-67. The subtypes were defined as: luminal A (ER+, HER2-), luminal B (ER+, HER2+), HER2 positive (ER-, HER2+), and basal-like (ER-, HER2-, EGFR+, and/or cytokeratin 5/6+). The prevalence of breast cancer subtypes was basal-like (n = 19 [8%]); luminal A, n = 149 (61%); luminal B, n = 23 (9%); and HER2+/ER-, n = 38 (16%). Sixteen tumors (6%) were unclassified (negative for all 4 defining markers). The basal-like subtype was associated with unfavorable prognostic variables including high-grade nuclei (P < .0001), p53 overexpression (P < .0001), and elevated Ki-67 index (P < .0001). These studies demonstrate the presence of a basal-like in situ carcinoma, a potential precursor lesion to invasive basal-like carcinoma.     

AGR2 associates with HER2 expression predicting poor outcome in subset of estrogen receptor negative breast cancer patients

Expression of the AGR2 oncogene was shown to be associated with estrogen receptor positive tumors. This gene contributes to enhanced cellular proliferation, drug resistance, metastasis development and may also serve as a predictor of poor prognosis. However, our analysis of AGR2 expression in a subset of estrogen-receptor negative tumors revealed that AGR2 could also be upregulated in hormone-independent manner. AGR2 expression was shown to be significantly increased in HER2 positive breast tumors on both the mRNA and the protein level. Moreover, in a subset of estrogen- and progesterone-receptor negative and simultaneously HER2-positive cases, increased AGR2 expression significantly correlated with worse patient prognosis. Subsequent analysis of independent data sets either collected in our institute or obtained from Oncomine cancer microarray database confirmed all these findings.     

p16INK4a expression in basal-like breast carcinoma

BLBC represents a distinctive group of invasive breast carcinomas with specific genotype and immunopro-file. BLBC is usually defined by gene expression profiling and is currently associated with poor outcome. BLBCs are estrogen receptor (ER) negative, progesterone receptor (PgR) negative, HER2 negative, and usually show a variable expression of basal cytokeratins (CKs), EGFR and CD117. p16 ^INK4a is a tumor suppressor protein, encoded by the CDKN2A gene, which regulates cell cycle. The reported association of abnormalities in the p16/Rb pathway with increased risk of malignancy prompted us to determine the expression of p16^INK4a in a group of BLBC; the results were compared with a group of high-grade invasive carcinoma (HG-IC) of breast. Tissue microarrays (TMA) were constructed in triplicate including 18 BLBC and 18 HG-IC. All BLBC cases were ER-/PgR-/HER2-. Seventeen (94%) BLBC were CK 5/6+/CK 14+; 14 (78%) BLCB showed EGFR expression and 13 (72%) were CD117 positive. BLBCs showed a strong positive reaction with p16 ^INK4a antibody in 16 of 18 (89%) cases. Although the significance of p16 ^INK4a expression in breast cancer is not fully understood, we have shown that p16^INK4a is strongly expressed in breast cancers with basal-like phenotype. Since it is known that p16^INK4a is associated with aggressive behavior in human carcinomas, these data suggest that p16^INK4a play a role in the poor prognosis of BLBC.     

Differential expression of HER2 and downstream proteins in prediction of advanced tumor phenotypes and overall survival of patients with Epstein-Barr virus-positive vs. negative gastric cancers

This study evaluated the associations of HER2 protein, HER2 gene amplification, and positivity for p-AKT, p-ERK, and p-PLCγ proteins with clinicopathological status and overall survival (OS) of patients who had Epstein-Barr virus-associated gastric cancer (EBVaGC; n = 58) or EBV-negative GC (EBVnGC; n = 329). Tissue samples were subjected to immunohistochemistry and fluorescence in situ hybridization (FISH). Results showed that EBVaGC less expressed HER2 and amplified HER2 gene. p-AKT (p = 0.035) and p-ERK (p = 0.001) were inhibited in EBVaGC than in EBVnGC, while p-PLCγ (p = 0.034) was upregulated. Among EBVaGC patients, p-ERK positivity was associated with Lauren classification (p = 0.023), and p-PLCγ positivity was inversely associated with TNM stage (p = 0.041) and lymph node metastasis (p = 0.041). In contrast, among EBVnGC patients, HER2 expression was associated with distant metastasis (p = 0.043) and p-AKT positivity was associated with intestinal subtype (p < 0.004), lymph node metastasis (p = 0.031), distant metastasis (p < 0.001), and elder age (>60y, p < 0.004). Overall analysis showed that EBVaGC patients presented better OS than EBVnGC patients (p = 0.044). Among EBVaGC patients, p-AKT positivity (p = 0.008) was associated with worse OS; as well as, HER2 high expression (p < 0.001), p-AKT positivity (p = 0.010), and p-PLCγ (p < 0.001) were associated with worse OS in EBVnGC patients. Multivariate analysis showed that distant metastasis (95% CI: 1.559 to 4.028, p < 0.001), HER2 high expression (95% CI: 1.058 to 2.454, p = 0.026), and p-PLCγ positivity (95% CI: 1.056 to 2.435, p = 0.027) were independent prognostic predictors of OS in EBVnGC patients. Our results indicated that p-AKT positive patients presented worse OS than p-AKT negative ones in EBVaGC, as well as, HER2, p-AKT, and p-PLCγ are prognostic biomarkers for OS in EBVnGC patients.     

Prognostic factors in patients with metastatic breast cancer at the time of diagnosis

Approximately 90% of breast cancer mortality is due to metastases that are resistant to adjuvant therapies. Thus, assessment of factors associated with clinical outcomes in patients with advanced breast cancer is of significant importance. Despite the recent improvement in early detection, between 5 and 10% of breast cancer patients are diagnosed with metastasis at initial presentation or, rarely, before the primary breast cancer has been identified. These patients typically have poorer survival outcomes compared to those who develop distant metastasis subsequently. Yet, the prognostic relevance in these patients has not been intensively explored. In this study, we analyzed breast cancer patients with distant metastasis at the time of diagnosis between 1997 and 2010 (n = 194) to identify the clinicopathological factors significant for overall survival. By univariate analysis, race, estrogen receptor (ER) and progesterone receptor status were significantly associated with overall survival, while race and ER remained independent factors in multivariate analysis. Being Caucasian and overexpressing of ER both showed a significantly decreased hazard of death (P = 0.015 and 0.017, respectively). Reflecting these findings, the overall survival differed significantly between breast subtypes, with the luminal subtype and triple negative disease being associated with the longest and worst survival, respectively. Further, multi-organ involvement was associated with a worse prognosis than those with single organ metastasis, whereas no significant difference in survival was found between the different anatomic sites (bone, viscera and brain). Our findings suggest that it is predominantly the intrinsic nature of the tumor along with the genetic makeup of the patient that predicts the prognostic outcome in those patients with advanced disease at presentation.     

Preserved Axin expression is associated with an aggressive phenotype in invasive breast carcinomas

Reduced Axin expression has been associated with an aggressive behavior in lung and esophageal squamous cell carcinomas. Its role in breast cancer has not been defined. The aim of our study was to investigate the expression pattern of Axin protein in invasive breast carcinomas, in relation to the behavior and prognosis of the disease. Immunohistochemistry was performed for the detection of Axin expression in 232 breast cancer tissues. Univariate and multivariate statistical analyses were used to assess the relation of Axin expression with classic clinicopathological parameters, patients' survival and various biologic markers Human Epidermal Factor‐2 (HER‐2), Ki‐67, topoIIα, glycogen synthase kinase‐3β (GSK‐3β)]. Preserved cytoplasmic Axin expression was positively correlated to lymph node invasion, HER‐2 and GSK‐3β and inversely to Ki‐67 and topoIIα. Nuclear Axin was positively associated with tumor size. Stromal Axin showed a parallel association with lymph node status and HER‐2. In the subgroup of lobular breast carcinomas, preserved Axin was found to exert an unfavorable impact on patients' overall survival. Our findings indicate, for the first time, that in invasive breast cancer preserved Axin expression is associated with a more aggressive phenotype and that in the discrete subtype of lobular breast carcinomas Axin negatively influences patients' overall survival.     

Observer prediction of HER2 amplification in HercepTest 2+ breast cancers as a potential audit instrument

Going J J
(2011) Histopathology 59 , 333–335 Observer prediction of HER2 amplification in HercepTest 2+ breast cancers as a potential audit instrument Aims: Correct assignment of human epidermal growth factor receptor 2 (HER2) status in breast cancer is important. Indeterminate (2+) HER2 immunohistochemistry (IHC) is usually resolved by FISH for HER2 gene amplification. It was hypothesised that predicting HER2 amplification in IHC 2+ cases could improve audit of HER2 IHC and its interpretation. Methods and results: One observer (J. J. G.) interpreted 4343 assessable HercepTests on consecutive breast cancers from the West of Scotland over 45 consecutive months during 2007–2010, with 2+ cases classified prospectively as ‘probably amplified’, ‘possibly amplified’ or ‘probably not amplified’ prior to FISH. A HER2 to chromosome 17 FISH ratio >2 was taken to define HER2 amplification. There were 265 3+ cases (6.1%) and 883 2+ cases (20.3%). Of 187 ‘probably amplified’ 2+ cases, 166 (88.8%) were amplified, as were 90 (37.8%) of 238 ‘possibly amplified’ 2+ cases. Of 458 ‘probably not amplified’ but still 2+ cases, 59 (12.8%) were in fact amplified (overall χ² 333.89, df 2, P < 0.0001). In total, 580 of 4343 (13.4%) cancers were HER2‐positive (265 3+ by IHC and 315 2+ and HER2 amplified). Conclusions: Breast cancers HER2‐indeterminate (2+) by HercepTest IHC can be strongly separated into those probably HER2 amplified, a core indeterminate group and those probably not HER2 amplified. The percentage of HER2 amplified cases in each category is proposed as an instrument for comparison of HER2 IHC and its interpretation between laboratories and observers.     

Tissue microarray technology in breast cancer HER2 diagnostics

Tissue microarrays (TMAs) as current medical research tools significantly lower the costs of immunohistochemical examinations (IHC) and fluorescence in situ hybridization (FISH) while enabling high levels of standardization and reliability. Taking HER2 testing of breast cancer into consideration, we assessed the routine applicability of TMAs. A hundred and seventy-four consecutive samples of invasive breast cancer cases were selected. TMAs were constructed in order to conduct double HER2 immunohistochemical analysis and FISH abreast using the conventional slide by slide method. Comparing the immunohistochemical data obtained from TMAs with the routinely processed large sections, we found a 94.5%/92.7%, 85.7%/88.9% and 91.2%/90% concordance at immunohistochemically HER2-negative, HER2 2+ and 3+ cases using the CB11/HercepTest, respectively. FISH performed on TMAs helped to determine Herceptin therapy suitability in all cases, and when discordance was found, we controlled FISH on "large sections". Being able to conduct FISH examinations at a reasonable price with or without prior immunohistochemical analysis, departments confronted with a certain frequency of breast cancer cases might extensively use the type of TMAs applied in our study. This is a relieve not only with regard to diagnostic work using microarrays, but this also allows to take new directions in research by shedding light on certain unusual cases.     

乳腺浸润性导管癌HER2免疫组化检测结果判定的重复性研究

目的 探讨病理医师对乳腺浸润性导管癌HER2免疫组化(immunohistochemistry,IHC)检测结果判定的重复性及其影响因素,寻求提高判读重复性和准确性的措施.方法 参照<乳腺癌HER2检测指南>,2位乳腺专科病理医师共同读片选取56例浸润性导管癌的HER2 IHC切片作为研究对象.4位非专科病理医师独立盲法完成对该组病例IHC切片的前后两轮结果判定.对第一轮4位观察者间的判读重复性和观察者自身的判读重复性进行Kappa分析.同时将2位专科医师共同确定的判读结果作为HER2判读参考分级,4位非专科医师第一轮的判读结果分别与之进行配对Kappa分析.结果 观察者间的判读重复性为中等(K=0.593 9),重复性最高的级别是HER2为0(K=0.811 4),最低的是HER2为2 (K=0.473 6).判读级别简化为阳性和阴性后,观察者间的重复性明显提高(K=0.723 0).4位观察者自身的判读重复性极好(Kw=0.821 9～0.916 5),判读的准确性则与经验有关.结论 采用标准化的检测手段、深入学习和实践现有的判读标准以及联合判读等措施应有助于提高HER2 IHC判读的重复性和准确性.     

Obesity is associated with a poorer prognosis in women with hormone receptor positive breast cancer

Objective

Whether moderate to severe obesity (body mass index (BMI) ≥ 30 to <40 kg/m²) contributes to breast cancer recurrence and mortality remains uncertain.

Subjects and methods

1199 women, recruited within 12 months of their diagnosis of hormone receptor positive (HR+), human epidermal growth factor receptor 2 negative (HER2−) invasive breast cancer completed an enrolment questionnaire and an annual follow-up questionnaire every 12 months for another 5 years. The impact of obesity on time to either local or distant recurrence or new breast cancer, or death due to breast cancer was determined by Cox regression. Women in the most extreme categories of BMI (<18.5 and ≥40) were excluded from the analysis.

Results

Of the 1155 included women, mean age, 58.4 ± 11.6 years, 53.8% had Stage 1 disease and 88.9% received oral adjuvant endocrine therapy (OAET) within 2 years of diagnosis. The likelihood of an event was significantly associated with moderate to severe obesity (HR = 1.71, 95%CI, 1.12–2.62, p = 0.014), disease beyond Stage 1 (HR = 2.87, 95% CI 1.73–4.75, p < 0.001), OAET (HR = 0.26, 95%CI 0.14–0.46, p < 0.001), mastectomy (HR = 3.28, 95%CI 1.98–5.44, p < 0.001) and radiotherapy (HR = 2.12, 95%CI 1.24–3.63, p = 0.006). For Stage 1 disease, only moderate to severe obesity (HR 3.23, 95%CI 1.48–7.03, p = 0.003) and OAET use (HR 0.41, 95%CI 0.17–0.98, p = 0.046) were significantly associated with an event.

Conclusion

Moderate to severe obesity is associated with a poorer invasive breast cancer prognosis; this is also true for women with Stage 1 disease, and is independent of age and treatment.     

Impact of chromosome 17 centromere region assessment on HER2 status reported in breast cancer

Recent studies have indicated that polysomy 17 is a rare event in breast cancer, and polysomy is usually mimicked in FISH analysis by gain or amplification of the centromere covered by the chromosome 17 centromere probe. To estimate the impact of chromosome 17 centromere assessment on routine practice, we conducted a retrospective re-classification study.Four hundred and five consecutive cases were selected. The original molecular pathology reports were available. Centromere 17 copy counts were ignored in the reassessment.Altogether, nineteen (4.69%) discrepant cases were found, from which five (1.23%) were considered originally non-amplified but had an HER2 copy number >6. Therefore, we reclassified them as HER2-amplified, while fourteen (3.46%) cases were originally considered amplified with 6 or fewer HER2 signals/cell.The discrepant cases found in our reassessment study would require further high-resolution genetic analysis to resolve the disagreement. On the other hand, our result also highlights that for the vast majority of breast cancer cases traditional FISH examination is still adequate to reach the correct diagnosis. This diagnostic gap must be filled by more sophisticated genetic examinations. Moreover, upcoming HER2 guidelines should consider the aid that high-resolution karyotyping can give to the diagnostic algorithm.     

The prognostic value of tumor budding in invasive breast cancer

We investigated the prognostic value of tumor budding in 160 cases of operable invasive ductal carcinoma, not otherwise specified (IDC-NOS). The number of buds was counted in H&E slides with a maximal invasive margin in a 0.950 mm² field of vision (200×). According to a cut-off score selected by ROC analysis, the cohort was dichotomized into a low (0–7 budding foci, 107 cases, 66.9%) and a high-grade budding group (8 or more budding foci, 53 cases, 33.1%). The inter-observer test showed a good reproducibility with 0.717 as the К value. High-grade budding was significantly associated with the presence of lymphovascular invasion (LVI) (P = 0.001), larger tumor size (P = 0.014), and worse clinical outcome (P < 0.001). By immunohistochemical staining, budded cells at the margin displayed epithelial mesenchymal transition (EMT)-like molecular phenotype and decreased proliferative activity. Survival analyses revealed that tumor budding (HR 4.275, P < 0.001) together with tumor size (HR 2.468, P = 0.002), node status (HR 2.362, P < 0.001), and LVI status (HR 1.910, P = 0.035) was the independent prognostic factor in IDC-NOS. In conclusion, tumor budding is a reproducible, significant, and independent histopathological prognostic factor in IDC-NOS.