Immunohistochemical expression of extracellular matrix components tenascin,fibronectin, collagen type IV and laminin in breast cancer: their prognostic value and role in tumour invasion and progression

The immunohistochemical expression of the extracellular matrix (ECM) components tenascin (TN), fibronectin (FN), collagen type IV (Coll) and laminin (LN), and their possible relationships were studied in a series of 134 operable breast cancer cases. Their expression was also compared with the expression of the proteolytic enzyme cathepsin D (CD), the adhesion molecule CD44 standard form (CD44s) and other known factors to clarify the prognostic value and role of these molecules in tumour progression and metastasis. TN expression in the tumour stroma was positively correlated with tumour grade and size, CD44s expression, tumour and stromal CD expression as well as with FN, laminin and Coll expression in the same areas. TN expression was inverse correlated with ER status. Its expression at the invasion front was only positively correlated with the lymph node status. Survival analysis showed an increased mortality risk associated with high levels of TN expression. In multivariate analysis, among the ECM proteins, only TN expression was independently correlated with patients' survival. FN expression was positively correlated with lymph node involvement, with the proliferation-associated index Ki-67 and stromal CD expression. Survival analysis showed an increased mortality risk associated with a high level of FN expression. Coll expression was positively correlated with the tumour size and LN expression. An inverse relationship of Coll expression with ER and PgR receptor status was also found. LN expression was positively correlated with tumour and stromal CD expression, with the proliferation-associated index Ki-67 and inversely with ER receptor status. The observed alterations in the expression of ECM proteins in breast cancer tissue and their correlations with the proteolytic enzyme CD and the adhesion molecule CD44s, suggest an involvement in cancer progression. In addition, overexpression of stromal TN and FN seems to have negative prognostic value in breast cancer patients.     

Regulation of stromal versican expression by breast cancer cells and importance to relapse-free survival in patients with node-negative primary breast cancer.

PURPOSE: Determination of meaningful prognostic indicesremains a high priority for women diagnosed with node-negative primary breast cancer. Currently, 30% of these women relapse, and there is no reliable means of predicting this group of patients. This study investigates whether the level of expression of versican, an anticell adhesive proteoglycan, in the peritumoral stromal tissue of women with node-negative, primary breast cancer predicts relapse-free survival. This study also examines whether breast cancer cells regulate the secretion of versican by mammary fibroblasts. EXPERIMENTAL DESIGN: Immunoreactive versican was measured in breast cancer tissue sections of 58 node-negative patients by video image analysis. Primary isolates of mammary fibroblasts were cultured in medium conditioned by the breast cancer cell lines ZR-75-1, MCF-7, BT-20, and MB231. Changes in versican secretion were measured by immunoblotting and enhanced chemiluminescence. RESULTS: Cox analyses indicated that peritumoral versican level was the sole predictor of relapse-free survival. The relapse rate in patients with low versican levels was lower than in patients with high versican levels (Kaplan-Meier: 83% relapse free at 5 years for versican mean integrated absorbance <14 versus 33% for > or = 14, P = 0.0006). Accumulation of versican in medium of mammary fibroblasts was increased after culture in conditioned medium from breast cancer cell lines. CONCLUSIONS: Relapse in women with node-negative breast cancer is related to the level of versican deposited in peritumoral stroma by mammary fibroblasts. Versican secretion appears to be regulated by breast cancer cell mediators. Neoplastic remodeling of extracellular matrix through increased versican deposition may facilitate local invasion and metastasis.     

基质金属蛋白酶-2在淋巴结阴性乳腺癌中的表达及临床意义

目的 探讨淋巴结阴性乳腺癌组织中基质金属蛋白酶-2(MMP-2)的表达水平、与其他因子的关系及临床意义。方法：采用免疫组织化学ABC法，检测270例淋巴结阴性乳腺癌石蜡标本中MMP-2的表达水平：结果：270例淋巴结阴性乳腺癌标本中MMP-2阳性表达为56．7％。MMP-2阳性表达与肿瘤大小和分级有关，与雌、孕激素受体(ER、PR)等因子无关；与无复发生存率(RFS)有关，而与总生存率(OS)无关：结论：MMP-2蛋白在淋巴结阴性乳腺癌转移复发中起着重要作用，是重要的预后因子：     

腋窝淋巴结阴性浸润性乳腺癌分子分型的表达及预后分析

  目的   探讨腋窝淋巴结阴性浸润性乳腺癌不同分子分型的分布,临床病理特征以及和预后的关系。   方法   回顾性分析183例该类乳腺癌患者的临床病理资料,对管腔上皮（Luminal）型、基底样（Basal-1ike）型和HER-2过表达（over-expression）型乳腺癌患者在年龄、肿瘤大小、临床分期和无瘤生存率（disease-free survival,DFS）、总生存率（overall survival,OS）方面进行统计分析。   结果   不同分子亚型乳腺癌在年龄、肿瘤大小、临床分期方面无显著性差异。Luminal、Basal-like、HER-2过表达型的复发率分别为3.9%（4/102）、20.4%（10/49）、6.3%（2/32）（P=0.002）;死亡率分别为2.0%（2/102）、6.1%（3/49）、3.1%（1/32）（P>0.05）。Ka-plan-Meier分析显示Basal-like型的DFS最低（P=0.002）,Basal-like型的OS较低（P=0.39）。Cox多因素比例风险模型分析显示分子亚型对DFS存在显著影响（P=0.001）。   结论   在腋窝淋巴结阴性浸润性乳腺癌患者中,不同分子亚型在年龄、肿瘤大小及临床分期方面无显著性差异,Basal-like型预后最差,分子分型是其独立预后指标。       

细胞外基质蛋白1在乳腺癌组织和细胞株中的表达及其意义

目的:探讨细胞外基质蛋白1(extracellular matrix protein 1,ECM1)在乳腺癌组织和细胞株中的表达及其意义.方法:应用免疫组织化学和免疫细胞化学EnVision法检测乳腺癌手术切除标本中乳腺组织(正常组织和癌组织)和恶性程度不同的乳腺癌细胞株(MCF-7、Sk-Br-3和MDA-435)中ECM1的表达,并比较正常乳腺组织和乳腺癌组织、淋巴结转移和未转移乳腺癌之间的ECM1表达差异.Western blotting检测确定ECM1亚型蛋白.结果:ECM1的表达在正常乳腺组织中阳性率为8.7%(2/23),在乳腺癌组织中阳性率为60.4%(29/48);其中乳腺癌淋巴结未转移者为38.5% (10/26),淋巴结转移者为86.4% (19/22).统计分析表明,ECM1的表达在乳腺癌组织中高于正常组织(P<0.01),在转移性乳腺癌中高于非转移性乳腺癌(P<0.01).恶性程度低的乳腺癌细胞株MCF-7、Sk-Br-3中不表达ECM1,而恶性程度较高的MDA-435细胞株中高表达ECM1.乳腺癌细胞中高表达的ECM1蛋白主要是相对分子质量为68 000的ECM1a亚型.结论:ECM1(主要为ECM1a亚型)在乳腺癌组织和恶性程度高的乳腺癌细胞中过表达,并且与乳腺癌的淋巴结转移相关.     

Inflammation and breast cancer. Metalloproteinases as common effectors of inflammation and extracellular matrix breakdown in breast cancer

Two rapidly evolving fields are converging to impact breast cancer: one has identified novel substrates of metalloproteinases that alter immune cell function, and the other has revealed a role for inflammation in human cancers. Evidence now shows that the mechanisms underlying these two fields interact in the context of breast cancer, providing new opportunities to understand this disease and uncover novel therapeutic strategies. The metalloproteinase class of enzymes is well studied in mammary gland development and physiology, but mostly in the context of extracellular matrix modification. Aberrant metalloproteinase expression has also been implicated in breast cancer progression, where these genes act as tumor modifiers. Here, we review how the metalloproteinase axis impacts mammary physiology and tumorigenesis and is associated with inflammatory cell influx in human breast cancer, and evaluate its potential as a regulator of inflammation in the mammary gland.     

Reduced expression of the small leucine-rich proteoglycans, lumican, and decorin is associated with poor outcome in node-negative invasive breast cancer.

PURPOSE: To examine the prognostic significance of lumican and decorin, two abundant small leucine-rich proteoglycans in breast tissue stroma. EXPERIMENTAL DESIGN: Lumican and decorin expression was examined in a cohort of 140 invasive breast carcinomas by Western blot analysis. All cases were axillary lymph node-negative and treated by adjuvant endocrine therapy. RESULTS: Lumican and decorin expression was highly correlated (r = 0.45, P < 0.0001), but although low levels of lumican were associated with large tumor size (P = 0.0496), negative estrogen receptor (P = 0.0024) and progesterone receptor status (P = 0.0116), and increased host inflammatory response (P = 0.0077), low decorin levels were associated only with large tumor size (P = 0.0496). However, using univariate analysis, low levels of lumican and decorin were both associated with a shorter time to progression (P = 0.0013 and 0.0262) and poorer survival (P = 0.001 and 0.0076). In multivariate analysis using the Cox regression model, low decorin was also shown to be an independent predictive factor for recurrence (hazard ratio 2.25: 95% confidence interval 1-5, P = 0.047) and survival (hazard ratio 3.39: 95% confidence interval 1.2-9.6, P = 0.021). CONCLUSIONS: These results suggest that low levels of small leucine-rich proteoglycans in breast tumors may be associated with a worse prognosis in lymph node-negative invasive breast carcinomas and warrant further study with larger patient cohorts.     

Influence of internal mammary node irradiation on long-term outcome and contralateral breast cancer incidence in node-negative breast cancer patients

Background and purpose

There is no general consensus concerning irradiation (RT) of internal mammary nodes (IMN) in axillary node-negative breast cancer. Based on a large series of patients treated in a single institute and followed up for a long period of time, we looked at the influence of IMN RT on late outcome of these patients as well as the development of contralateral breast cancer (CBC).

Patients and methods

The study was based on 1630 node-negative breast cancer patients treated in our institution between 1975 and 2008 with primary conservative surgery and axillary dissection or sentinel node examination. All patients received post-operative breast RT. IMN RT was more frequent in inner or central tumours. Kaplan–Meier (K–M) overall survival (OS), cancer-specific survival (CSS), and disease-free survival (DFS) according to IMN RT were calculated for all patients and for patients with inner/central tumours. The K–M rate of contralateral breast cancer (CBC) was also analysed and correlated with IMN RT.

Results

Prognostic variables such as tumour size, histological grade, and hormone receptors were not significantly different in the groups having received IMN RT or not. Considering all patients, OS was strictly comparable in the 2 groups: 10-year values were 85% (IMN RT) and 86% (no IMN RT), respective values at 20 years were 66.6% and 61.0% (p = 0.95). However, in patients presenting with inner/central tumours, OS was significantly improved in the IMN RT group with respective values of 92.5% and 87.2% at 10 years, and 80.2% and 63.3% at 20 years: Hazard ratio (HR) = 0.56 (0.37–0.85); p = 0.0052. Again, CSS was improved in patients with inner/central tumours having received IMN RT, with 20-year rates of 89.5% versus 79.1% in patients not receiving IMN RT (p = 0.047). No difference in DFS was noticed. The actuarial rate of CBC development was comparable between patients having received IMN RT and other patients. However, considering only patients alive 10 years after primary breast surgery, the K–M rate of CBC at 20 years was 5.3% in patients without IMN RT and 7.2% in patients with such RT; HR = 2.47 (1.23–4.95); p = 0.008.

Conclusions

IMN RT in node-negative tumours was associated with increase in OS and CSS in patients with inner or centrally located lesions. An increase in CBC development was also noticed in long-survivors of IMN RT patients; however, these findings have to be interpreted with caution because of the difference in follow-up between the 2 groups. Further studies are warranted to investigate the potential role of IMN irradiation in the development of CBC.     

Swiss adjuvant trials in women with node-negative breast cancer. OSAKO.

Women with node-negative breast cancer have a 30% chance of relapse 5 years after mastectomy. If it is possible to prevent or defer recurrent disease with adjuvant systemic therapy, node-negative patients, with their low tumor burden, should theoretically benefit most from such treatment. In 1974 we started a randomized adjuvant trial in eastern Switzerland, using a subjectively less toxic regimen [chlorambucil, methotrexate, and fluorouracil (LMF)]. Two hundred fifty-four patients were randomly assigned after standardized modified radical mastectomy to observation only or to treatment with oral LMF for 6 months followed by BCG skin scarifications monthly for up to 2 years. While we find no significant statistical difference between the control group and the treated group in terms of relapse-free survival, there is a strong and consistent trend toward prolongation of overall survival within the treated group.     

A model of chemotherapy in node-negative breast cancer.

Should all women with node-negative breast cancer receive chemotherapy? Acceptance of such a broad recommendation has been controversial due to the relatively good prognosis of these women and the lack of evidence of an improvement from chemotherapy in overall survival. We studied the question with a decision analytic model, which simulates a clinical trial of a large number of women over several decades, to calculate quality-adjusted life expectancy and the cost-effectiveness of chemotherapy in 45-year-old and 60-year-old women. The model considered a variety of scenarios about the long-term benefits of chemotherapy. The initial analysis found a lifetime benefit of 4 to 5 quality months (range, 2 to 14 months) at a cost of $15,400 to $18,800 per quality year (range, $5,100 to $56,800). If a risk-stratification protocol could identify a low-risk group, then benefit drops to about 1 quality month at a cost of $65,000 to $90,000 per year. For the average woman, chemotherapy increases the quality-adjusted life expectancy by a substantial amount at a generally acceptable cost. However, given the uncertainty of the duration of benefit and current potential for risk-stratification, a broad recommendation for chemotherapy is inappropriate.     

Correlation between c-erbB-2 amplification and risk of recurrent disease in node-negative breast cancer.

Drawing upon the comprehensive population-based Northern Alberta Breast Cancer Registry containing 704 patients with histologically negative axillary lymph nodes who have been followed for 5-16 years, we have undertaken a retrospective case-control study to evaluate the utility of genomic amplification of specific protooncogenes [c-erbB-2 (nee HER-2/neu), c-erbA, c-myc, int-2, and hst-1] as predictive indicators of clinical outcome in node-negative disease. To this end, 115 women with node-negative breast cancer who had recurred at any time up to 16 years posttreatment (cases) were matched pairwise for appropriate clinicopathological variables (size of primary tumor, menopausal state, estrogen receptor status, anniversary year of treatment, and patient age) with a second group of 115 women (controls) selected from a cohort of 502 node-negative patients who had not relapsed during long-term follow-up. Tumor DNA extracted from archival formalin-fixed, paraffin-embedded tissue blocks were analyzed for protooncogene copy number by slot-blot hybridization. Taking a gene copy number of 3 as the cutoff, 27 of the 230 tumor samples examined contained from 3- to 22-fold elevation in c-erbB-2 genomic equivalents. Twenty-one of the 27 tumors amplified for c-erbB-2 were derived from cases and 6 from controls, signifying that 18% of the node-negative patients who had relapsed harbored excessive copies of the protooncogene in their malignant tissue compared to only 5% for the patients who had remained in remission. Accordingly, the occurrence of amplification of c-erbB-2 proved to be a statistically significant predictor of poor prognosis, especially disease-free interval (P = 0.006). Moreover, this genetic alteration appeared to be independent of and to have greater predictive power than most commonly used prognostic factors. Our findings also indicated that as a clinical test, measurement of c-erbB-2 amplification suffers from low sensitivity; however, when greater than 6 gene copies are present, the test has a positive predictive value for recurrence of 70%. Concurrent analysis of tumor DNA blots with probes for the other four protooncogenes examined revealed that their amplification, which others have reported to arise often, especially in node-positive disease, was seldom found even in our high-risk case group (2-3%). In short, our data strongly suggest that amplification of c-erbB-2 may contribute to the pathogenesis of some forms of node-negative breast cancer and thus may serve as a useful genetic marker to identify a subset of high-risk patients.     

Clinicopathological study of the expression of syndecan-1 in invasive breast carcinomas. correlation with extracellular matrix components

Syndecan-1, a cell surface proteoglycan found predominantly on epithelia of mature tissues, binds both extracellular matrix (ECM) components and basic fibroblast growth factor (bFGF) and is implicated in the restriction of growth and invasiveness of neoplastic cells, as it induces the adhesion capacity of neoplastic cells with the stroma. In this study we investigated breast carcinomas for the immunohistochemical expression of syndecan-1 protein and these results were assessed in relation to clinicopathological parameters, in order to clarify its prognostic value. The possible relationship with hormone receptors content, p53, cell proliferation markers, and extracellular matrix components was also estimated. Tissue sections from 102 breast carcinomas were used and immunostainings were performed on formalin-fixed, paraffin-embedded tissue sections by the labelled streptavidin avidin biotin (LSAB) method. High expression levels were observed, as 75/102 (73.5%) cases expressed immunoreactivity in more than 80% of neoplastic cells, while 67/102 (65.7%) exhibited high staining intensity. The survival analysis showed an increased mortality risk associated with high syndecan-1 staining intensity with borderline significance (p=0.041). In addition, there was a strong negative correlation between syndecan-1 protein expression and ECM, specifically collagen IV (p=0.026) and tenascin (p=0.0067). The results of the present study show the implication of this protein in the remodeling of breast cancer tissue, through the interaction with other extracellular matrix components, probably influences the tumour progression.     

细胞外基质蛋白1与乳腺癌的研究进展

乳腺癌是威胁女性健康的恶性肿瘤之一。随着治疗方法的改进,乳腺癌患者的预后已得到明显改善;但其浸润转移的机制未明,患者治疗后的复发和转移难题仍无法攻克。细胞外基质（extracellular matrix,ECM）是肿瘤细胞与周围微环境相互作用的分子基础,与多种恶性肿瘤的侵袭和转移相关。     

Skip lesions in the axilla in breast cancer,and their association with micrometastases

The term skip lesion, referring to a tumour-bearing node in the axilla in breast cancer, presupposes that nodal colonization usually occurs in sequence with the path of the lymph flow. Skip lesions are thus out of step with the system. The present study of axillary nodes from 73 node-positive patients demonstrates that skip lesions have much in common with lone micrometastases found at any level, and are thus but a variant of the pattern found in early colonization of the axilla. This in turn indicates that in contrast to a progressive build up of tumour growth first at the lower and later at the middle level, early colonization may be a random process. This again stresses the importance of nodal dissection versus biopsy in the management of breast cancer, particularly in view of recent reports on the prognostic significance of micrometastases.This material formed part of Dr. Msuya's thesis for the degree of M.Phil., University of Bergen.     

The association of plasma DHEA and DHEA sulfate with breast cancer risk in predominantly premenopausal women.

Concentrations of adrenal androgens are positively associated with postmenopausal breast cancer risk; however, results in premenopausal women are conflicting. Therefore, we conducted a prospective nested case-control study within the Nurses' Health Study II cohort to examine the relationship of DHEA and DHEA sulfate (DHEAS) with breast cancer risk in predominantly premenopausal women. Blood samples were collected from 1996 to 1999. The analysis included 317 cases of breast cancer diagnosed after blood collection and before June 1, 2003; for each case, two controls were matched on age, fasting status, time of day and month of blood collection, race/ethnicity, and timing of blood draw within the menstrual cycle. No associations were observed between DHEA or DHEAS levels and breast cancer risk overall [in situ and invasive; DHEA relative risk (RR), top versus bottom quartile, 1.2; 95% confidence interval (95% CI), 0.8-1.8, P(trend) = 0.53; DHEAS RR, 1.3; 95% CI, 0.9-2.0; P(trend) = 0.07]. However, both DHEA and DHEAS were positively associated with estrogen receptor-positive/progesterone receptor-positive breast cancer (DHEA RR, 1.6; 95% CI, 0.9-2.8, P(trend) = 0.09; DHEAS RR, 1.9; 95% CI, 1.1-3.3, P(trend) = 0.02). We observed a significant interaction by age, with an RR for DHEAS of 0.8 (95% CI, 0.4-1.5, P(trend) = 0.62) for women <45 years old and 2.0 (95% CI, 1.2-3.5, P(trend) = 0.003) for women >/=45 years old; results were similar for DHEA. Our results suggest that adrenal androgens are positively associated with breast cancer among predominately premenopausal women, especially for estrogen receptor-positive/progesterone receptor-positive tumors and among women over age 45 years.     

Sequencing of chemotherapy and radiation in lymph node-negative breast cancer.

PURPOSE: To conduct a retrospective analysis of chemotherapy and radiation sequencing in lymph node-negative breast cancer patients treated with breast-conserving surgery. PATIENTS AND METHODS: Between February 1982 and January 1996, 124 patients with lymph node-negative breast cancer underwent breast-conserving surgery with axillary dissection followed by chemotherapy and radiation therapy. The outcome of 68 patients who received chemotherapy first was compared with that of 56 patients who received radiation first. The two groups were balanced with respect to patient age, tumor stage, margin status, and estrogen and progesterone receptor status. Sixty-two percent of the patients had T1 primary disease. The median follow-up among surviving patients was 44 months for the chemotherapy-first group and 61 months for the radiation-first group. RESULTS: There were no statistically significant differences in local control, disease-free survival, or overall survival between the two groups. Five-year actuarial rates for local control for the chemotherapy-first and the radiation-first groups were 100% and 94%, respectively. Five-year recurrence-free rates for the chemotherapy-first and radiation-first groups were 92% and 77%, respectively. The 5-year overall survival rate was 89% for both groups. DISCUSSION: Giving chemotherapy before radiation in lymph node-negative breast cancer did not compromise local control. Given the concerns about increased distant metastases if radiation is given first, the chemotherapy-radiation sequence is recommended.