Immunohistochemical COX-2 overexpression correlates with HER-2/neu overexpression in invasive breast carcinomas: a pilot study

Cyclooxygenase-2 (COX-2) is a prostaglandin synthase that catalyzes the synthesis of prostaglandin G2 and H2. It has been shown that COX-2 plays an important role in tumorigenesis of different tumor types and it is thought to take part in breast carcinogenesis. In the present study, we aimed to investigate the relationship of immunohistochemical COX-2 expression with clinicopathological parameters, including HER-2/neu overexpression in invasive breast carcinoma (IBC).Our study population comprised 10 normal breasts, 25 ductal carcinomas in situ (DCIS), and 51 invasive breast carcinomas. Immunohistochemical overexpressions of COX-2 and HER-2/neu were investigated in sections of formalin-fixed, paraffin-embedded blocks by 3 observers.In normal breast, DCIS and IBC, the COX-2 overexpression rate was 0%, 84%, and 58.8%, respectively. In IBC, COX-2 overexpression had a significant relationship with HER-2/neu overexpression (p = 0.026) and a high histological grade (p = 0.026). COX-2 expression in both DCIS (n = 25) and IBC (n = 51) was significantly higher than in normal breast tissue (p < 0.0001). In addition, the COX-2 expression rate was significantly higher in DCIS than in IBC (p = 0.042).Our results indicated that COX-2 overexpression correlates with aggressive phenotypic features, such as HER-2/neu overexpression and high histological grade in IBC. Increased expression of COX-2 in both DCIS and IBC in comparison to normal breast could indicate a role in breast carcinogenesis. COX-2 overexpression may provide a clinically useful biomarker for estimating tumor aggressiveness.     

The status of cyclooxygenase-2 expression in ductal carcinoma in situ lesions and invasive breast cancer correlates to cyclooxygenase-2 expression in normal breast tissue

OBJECTIVES: There is a paucity of data on cyclooxygenase (COX)-2 expression in normal breast tissue and on the changes in COX-2 expression from normal tissue via ductal carcinoma in situ (DCIS) lesions to invasive cancer. The aim of this study, therefore, was to investigate COX-2 protein expression in normal breast tissue, DCIS, and invasive breast cancer in samples from the same patients. METHODS: In 39 patients, we investigated and compared COX-2 expression in paired samples of invasive cancer and normal adjacent breast epithelium by immunohistochemistry with a monoclonal COX-2 antibody. Furthermore, in 29 of these cases, we also analyzed a concomitant DCIS lesion. RESULTS: Patients without COX-2 expression in normal breast tissue also do not express COX-2 in invasive breast cancer and in DCIS lesions, respectively. Conversely, COX-2 expression in normal breast tissue was an indicator for COX-2 expression in the paired breast tumors. There was no significant correlation between COX-2 expression and pathologic tumor stage, nodal status, hormone receptor status, tumor size, grading, and lymphovascular space involvement. CONCLUSIONS: This is the largest study to date investigating COX-2 in paired samples of breast tumors and normal adjacent breast tissue. Our data are consistent with the hypothesis that COX-2 overexpression is an early event in breast carcinogenesis.     

Expression of EGFR family and steroid hormone receptors in ductal carcinoma in situ of the breast.

The expression of EGFR family and steroid hormone receptors was examined in a series of 40 cases of pure ductal carcinoma in situ (DCIS) of the breast by immunohistochemical staining of paraffin-embedded sections. Hematoxylin and eosin-stained sections were used to classify the tumors according to the published criteria by Holland et al. (Holland R, Peterse JL, Millis RR, et al. Semin Diagn Pathol. 1994;1 1:167-180). Of the tumors 48% were immunoreactive for EGFR, 63% for c-erbB-2, 78% for c-erbB-3, 95% for c-erbB-4, 88% for estrogen receptor (ER) and 80% for progesterone receptor (PR). Statistically significant association between histological grade (differentiation) and c-erbB-2 protein expression was seen (p <.001). In addition, expression of c-erbB-4 protein was associated with c-erbB-2 (p=.004), c-erbB-3 (p=.058), ER (p=.002) and PR (p=.004). It is concluded that c-erbB-2 expression in DCIS is associated with high-grade pathological features, and a higher c-erbB-2 expression is seen in DCIS than in invasive breast carcinomas. A possible association between extensive expression of c-erbB-4 and steroid hormone receptors in proliferative and premalignant breast epithelial cells and the c-erbB-2 expression in DCIS and invasive breast carcinomas is discussed.     

Expression of EGFR Family and Steroid Hormone Receptors in Ductal Carcinoma in situ of the Breast

The expression of EGFR family and steroid hormone receptors was examined in a series of 40 cases of pure ductal carcinoma in situ (DCIS) of the breast by immunohistochemical staining of paraffin-embedded sections. Hematoxylin and eosin-stained sections were used to classify the tumors according to the published criteria by Holland et al. (Holland R, Peterse JL, Millis RR, et al. Semin Diagn Pathol. 1994;11:167-180). Of the tumors 48% were immunoreactive for EGFR, 63% for c-erbB-2, 78% for c-erbB-3, 95% for c-erbB-4, 88%for estrogen receptor (ER) and 80% for progesterone receptor (PR). Statistically significant association between histological grade (differentiation) and c-erbB-2 protein expression was seen ( p <.001). In addition, expression of c-erbB-4 protein was associated with c-erbB-2 ( p =.004), c-erbB-3 ( p =.058), ER ( p =.002) and PR ( p =.004). It is concluded that c-erbB-2 expression in DCIS is associated with high-grade pathological features, and a higher c-erbB-2 expression is seen in DCIS than in invasive breast carcinomas. A possible association between extensive expression of c-erbB-4 and steroid hormone receptors in proliferative and premalignant breast epithelial cells and the c-erbB-2 expression in DCIS and invasive breast carcinomas is discussed.     

Immunohistochemical distribution of c-erbB-2 in in situ breast carcinoma--a detailed morphological analysis

An immunohistochemical study of c-erbB-2 expression was carried out on in situ (non-invasive) breast carcinoma, using antibody 21N, raised to the intracytoplasmic domain of the c-erbB-2 oncogene product. Strong membrane staining was observed in 44 out of 74 (59 per cent) cases of ductal carcinoma in situ (DCIS), but none of 48 lobular carcinoma in situ (LCIS) lesions. A detailed comparative morphological evaluation using several different parameters, including histological subtypes, was performed within the DCIS group. The results showed that there was a significant correlation between c-erbB-2 expression and the presence of large cell size, periductal lymphoid cell infiltration, marked nuclear pleomorphism, multinucleation, and a high mitotic rate. Of these, cell size appears to be the most important predictor of c-erbB-2 status, followed by the presence of periductal lymphoid cell infiltration. These results indicate, firstly, that LCIS and DCIS are biologically (as well as histologically) different and, secondly, that a subgroup of DCIS, which is associated with c-erbB-2 over-expression, exists and appears to have distinct histological features. The subgroup of DCIS cases which over-express c-erbB-2 may be a biologically definable category with prognostic importance. These results may therefore have relevance to breast screening programmes, but a larger study incorporating clinical data would be necessary to correlate these findings with clinical outcome.     

Physiological COX-2 Expression in Breast Epithelium Associates with COX-2 Levels in Ductal Carcinoma in Situ and Invasive Breast Cancer in Young Women

Cyclooxygenase-2 (COX-2) overexpression is implicated in increased risk and poorer outcomes in breast cancer in young women. We investigated COX-2 regulation in normal premenopausal breast tissue and its relationship to malignancy in young women. Quantitative COX-2 immunohistochemistry was performed on adjacent normal and breast cancer tissues from 96 premenopausal women with known clinical reproductive histories, and on rat mammary glands with distinct ovarian hormone exposures. COX-2 expression in the normal breast epithelium varied more than 40-fold between women and was associated with COX-2 expression levels in ductal carcinoma in situ and invasive cancer. Normal breast COX-2 expression was independent of known breast cancer prognostic indicators, including tumor stage and clinical subtype, indicating that factors regulating physiological COX-2 expression may be the primary drivers of COX-2 expression in breast cancer. Ovarian hormones, particularly at pregnancy levels, were identified as modulators of COX-2 in normal mammary epithelium. However, serial breast biopsy analysis in nonpregnant premenopausal women suggested relatively stable baseline levels of COX-2 expression, which persisted independent of menstrual cycling. These data provide impetus to investigate how baseline COX-2 expression is regulated in premenopausal breast tissue because COX-2 levels in normal breast epithelium may prove to be an indicator of breast cancer risk in young women, and predict the chemopreventive and therapeutic efficacy of COX-2 inhibitors in this population.In 2010, approximately 13% of all breast cancers in the United States were diagnosed in women age 45 and younger, accounting for nearly 18,600 cases of invasive breast cancer and 6500 cases of ductal carcinoma in situ (DCIS).¹ Furthermore, the proportion of advanced breast cancers diagnosed in young American women is increasing at a rate of 2% per year, making young women''s breast cancer an emerging concern.² Compared with breast cancer in older women, young breast cancer patients have increased recurrence and lower survival rates.^3–8 Although a delayed diagnosis can contribute to poorer survival in some young patients,^6,9 the primary factor driving poor prognosis is tumor biology. Young women''s breast cancer has increased hormone-receptor negativity, tumor cell proliferation, and lymphovascular invasion compared with postmenopausal cases.^4,6,10 Moreover, young age at the time of breast cancer diagnosis is an independent poor prognostic factor.^4,6,7,11,12 These data provide compelling arguments to develop novel strategies to reduce breast cancer incidence and poor outcomes in young women.One potential target for young women''s breast cancer is cyclooxygenase-2 (COX-2),¹³ a key enzyme in the synthesis of homeostatic and proinflammatory prostanoids.¹⁴ In rodent breast cancer models, COX-2 overexpression induces mammary tumorigenesis and is associated with multiple tumor-promotional effects including increased angiogenesis, enhanced tumor cell migration and invasion, and reduced antitumor immunity.^15–20 Conversely, COX-2 inhibition or loss in rodent models reduces mammary tumorigenesis and metastasis.^17,21–23 Clinical data are consistent with similar roles for COX-2 in human breast cancer because COX-2 overexpression in breast cancer is associated with decreased disease-free and overall survival.^24,25 In addition, regular use of nonsteroidal anti-inflammatory drugs (NSAIDs), which inhibit the COX family of enzymes, can reduce overall breast cancer risk.^26,27 To date, the function of COX-2 in young women''s breast cancer has not been addressed. In a single study, high COX-2 expression in combination with increased collagen I is reported as a poor prognostic indicator in young-onset breast cancer patients (age, <45 years).¹⁶ One mechanism by which COX-2 may contribute to young-onset breast cancers is through its role in normal breast tissue remodeling. Importantly, windows of active breast tissue remodeling specific to young women, such as those associated with puberty, menstrual cycling, pregnancy, and postpartum breast involution, correlate with an increased risk for incidence and progression of breast cancer.^28,29 Support for this has been shown in rodent models in which postpartum mammary gland involution promotes tissue remodeling, tumor progression, and metastasis, all of which are mitigated by anti–COX-2 treatment.^16,30 Furthermore, COX-2 up-regulation has been observed in rat mammary glands after treatment with the ovarian hormones estrogen and progesterone,³¹ which is consistent with a role for COX-2 in physiological breast tissue remodeling associated with pregnancy and the menstrual cycle.We hypothesized that if COX-2 is involved in breast tissue remodeling, then COX-2 inhibition may represent a particularly efficacious chemoprevention strategy for young women. One important step in addressing this hypothesis is to evaluate COX-2 expression in young women''s breast tissue. We used human and rodent mammary tissues to investigate the effect of pregnancy and ovarian hormones on COX-2 expression in normal tissue as well as to explore the link between COX-2 expression in histologically normal adjacent breast tissue, DCIS, and invasive ductal carcinoma (IDC) in young-onset cases. We found that COX-2 expression primarily was epithelial and varied greatly between individual women, with evidence of modulation by ovarian hormones. In addition, analysis of COX-2 expression in paired normal adjacent breast epithelium, DCIS, and IDC within breast tissue from the same woman showed that COX-2 expression in the normal epithelium was associated with COX-2 expression in DCIS and IDC. Altogether, these data suggest that factors regulating COX-2 expression in normal breast epithelium influence COX-2 levels in breast cancer, and indicate that further research is warranted into whether women with high COX-2 expression may benefit preferentially from COX-2 inhibition strategies.     

COX-2 induction by heparanase in the progression of breast cancer

Breast cancer confined within the lactiferous duct or lobule, without invading the stroma, is called ductal carcinoma in situ (DCIS), whereas breast cancer that has invaded the stroma through the basal membrane is called invasive cancer. Heparanase, an endo-beta-D-glucuronidase that specifically degrades heparan sulfate proteoglycans (HSPGs) in the extracellular matrix (ECM), plays an important role when breast cancer cells breach the basal membrane. Recently, we have reported that heparanase is involved in angiogenesis through direct induction of cyclo-oxygenase-2 (COX-2). COX-2 induces vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) and is thus involved in neovascularization. The present study was undertaken to analyze surgically resected breast cancer specimens for heparanase and COX-2 expression, using specimens from 59 patients with invasive cancer and 85 patients with DCIS (including 41 cases of DCIS adjacent to invasive cancer). This study yielded the following results: a) the distribution of heparanase within tumor tissue was identical to that of COX-2; b) heparanase expression was more frequent in invasive cancer than in non-invasive cancer; c) a close positive correlation was noted between heparanase and COX-2 expression (this correlation was particularly strong in cases of invasive cancer); and d) COX-2 expression was always seen in cases positive for heparanase expression. Our results indicate that heparanase expression increases during the progression of breast cancer into invasive cancer, and that this change is accompanied by increased COX-2 expression. They also suggest that heparanase may play a novel role for COX-2 mediated tumor angiogenesis in breast-cancer progression.     

Thrombospondin 1 protein expression relates to good prognostic indices in ductal carcinoma in situ of the breast

AIM: Angiogenesis plays an important role in tumour growth and has been shown to occur around both in situ and invasive tumours. The degree of angiogenesis within tumours depends on the balance of pro-angiogenic and anti-angiogenic factors. One such anti-angiogenic factor is thrombospondin 1 (TSP-1). This study investigates the pattern of expression of TSP-1 in ductal carcinoma in situ (DCIS) of the breast and its relation to the surrounding microvessel pattern and density. MATERIALS/METHODS: The expression of TSP-1 was studied in formalin fixed, paraffin wax embedded sections from 58 cases of pure DCIS, using a monoclonal antibody against TSP-1 and the avidin-biotin-diaminobenzidine immunoperoxidase detection system. Vessels were stained with a monoclonal antibody to the endothelial cell marker CD31. Stromal microvessel density was assessed by counting "hot spots" within 500 micro m of the basement membrane of involved ducts using a 25 point Chalkey graticule. RESULTS: TSP-1 staining of the basement membrane around duct spaces with DCIS was seen in 69% of cases. In addition, staining of the stroma between involved duct spaces was seen in 31% of cases, with a fibrillary pattern identical to that seen in invasive breast carcinomas. In 12% of cases no staining for TSP-1 was seen. Two patterns of vascularity were identified. A cuff of vessels immediately adjacent to the basement membrane of ducts with DCIS was seen in 71% of cases. The presence of stromal TSP-1 was significantly associated with DCIS showing no/little necrosis (p = 0.01) and no/little periductal inflammation (p = 0.04). There was a trend between the presence of stromal TSP-1 and tumour cell negativity for p53 (p = 0.087). The stromal microvessel Chalkey point count ranged between 3.33 and 16. An increased stromal microvessel count was associated with high histological grade (p = 0.02), extensive necrosis (p = 0.047), and pronounced periductal inflammation (p = 0.049). There was no association between the presence of stromal TSP-1 and stromal microvessel density. CONCLUSIONS: TSP-1 is expressed in the stroma around DCIS and in the immediately adjacent basement membrane. Expression of stromal TSP-1 is lost in DCIS with more aggressive histological features. The absence of a relation with microvessel density suggests that other angiogenic factors may play an important role in DCIS.     

Immunohistochemical evaluation of c-erbB-2 oncogene expression in ductal carcinoma in situ and atypical ductal hyperplasia of the breast

Cases of ductal carcinoma in situ (DCIS) and atypical ductal hyperplasia (ADH) of the breast were examined for expression of the protein product of the c-erbB-2 (neu, HER-2) oncogene using two different polyclonal antibodies via an avidin-biotin immunoperoxidase method on formalin- or Bouin'-fixed, paraffin-embedded tissue. Fifty-five percent (18/33) of DCIS and 10% (2/21) of ADH were positive. Significant c-erbB-2 expression in DCIS was generally divided on histologic grounds: ten of ten comedocarcinomas showed strong membrane staining, while only one of 14 small cell DCIS cases (micropapillary or cribiform patterns) showed immunostaining (which was weak and basilar in this single case). DCIS cases of mixed histology were strongly positive in areas of comedocarcinoma. In two of three cases of associated Paget's disease strong membrane staining was seen. The two c-erbB-2-positive ADH cases showed weak basilar staining akin to the small cell DCIS cases. Five cases of lobular neoplasia (atypical lobular hyperplasia or lobular carcinoma in situ) associated with DCIS or ADH were negative for c-erbB-2 expression. We conclude that comedocarcinoma in situ and Paget's disease frequently express the c-erbB-2 protein and are both histologically and biochemically distinct from ADH and small cell patterns of DCIS. We advocate precise subclassification of DCIS on histopathologic reports, particularly in view of reports that overexpression of the c-erbB-2 oncogene in infiltrating breast carcinomas may be associated with a poor prognosis.     

CD24 expression in ductal carcinoma in situ and invasive ductal carcinoma of breast: an immunohistochemistry-based pilot study

CD24 is a small, heavily glycosylated cell surface protein, that is expressed in a large variety of solid tumors. It is considered to play an important role in tumor progression and metastasis. We aimed to evaluate CD24 expression in invasive ductal carcinomas (IDCa), ductal carcinoma in situ (DCIS) and non-tumorous breast tissues, and to investigate the relationship between histopathological parameters, estrogen and progesterone receptors, and c-erbB2 expressions. The study included 34 IDCa, 25 DCIS, and 13 non-tumorous breast tissues. All cases were reevaluated histopathologically, and immunohistochemistry was performed with monoclonal CD24 antibody. The results clearly demonstrated that CD24 expression, including membranous and cytoplasmic staining, was significantly higher in DCIS and IDCa than in the non-tumorous breast (p=0.001, p=0.000, and p=0.035, p=0.000, respectively). Cytoplasmic staining was detected predominantly in neoplastic tissues and was significantly increased in high grade DCIS (p=0.013). In invasive carcinomas, although the level of membranous staining was significantly positively correlated with tumor grade (p=0.040), there was no such an association with the cytoplasmic level. However, it showed a trend towards pT (p=0.089). In conclusion, our results suggest that higher CD24 expression may be associated with malignant transformation and progression in breast cancer biology. Furthermore, higher membranous expression and, in particular, cytoplasmic staining seem to predict malignant transformation, and different patterns of CD24 expression may be associated with different pathological features in breast tumors.     

Postmenopausal status, hypertension and obesity as risk factors for malignant transformation in endometrial polyps

OBJECTIVES: We analyzed clinical data and pathological features of six cases of malignant endometrial polyps, to compare these with other examples reported in literature and to define the features of endometrial cancer arising in polyps. Moreover, to clarify the mechanisms of carcinogenesis in malignant endometrial polyps we examined the expression of cyclooxygenase-2 (COX-2), P53 and Ki 67 and their relationships with clinicopathologic characteristics. METHODS: The surgical pathology files of the Pathology Department of Parma University were searched for cases of endometrial polyps with nests of endometrial carcinomas, from the years 2002-2005. Clinical records, histological slides of endometrial curetting, hysterectomy with salpingo-oophorectomy specimens and pelvic lymph nodes were reviewed in each case. The main pathological features analyzed were histological types of endometrial cancer and the stage of development of neoplasm. The presence of other malignancies in the genital tract were also considered. Immunohistochemical staining was done using antibodies COX-2, p53 and Ki 67. RESULTS: In our study, all malignant endometrial polyps had been detected in postmenopausal women. The majority of our patients with malignant endometrial polyps had risk factors for the development of endometrial carcinoma such as hypertension, obesity and unopposed estrogen therapy. Unlike other studies, no patients had a history of previous breast carcinoma and Tamoxifen treatment. The most common subtypes of endometrial carcinoma in malignant polyps are endometrioid carcinoma and serous papillary carcinoma. Endometrial carcinoma arising in endometrial polyps is an early endometrial carcinoma with good prognosis, except for papillary serous carcinoma, which can be associated with multiple omental involvement, despite low stage of development in the uterus. Immunohistochemical study showed that COX-2 expression was found in cytoplasm of tumor cells and this was elevated in all cases, independently of the grade and the stage of development of the malignancy, histological subtype and deep invasion of myometrium. P53 and Ki 67 expression, detected in the nuclei of neoplastic cells, was not correlated with COX-2 immunoreactivity, but these markers were associated with more advanced stage, grading, and histologic subtypes of tumor. CONCLUSIONS: Postmenopausal status, hypertension, obesity could all be considered as risk factors for carcinomatous transformation within endometrial polyps in women without a history of breast carcinoma and Tamoxifen treatment. However, our series is small (only six cases considered) and further studies are necessary to confirm this hypothesis. In the current study, immunohistochemical data reveal that COX-2 expression may be associated with the carcinogenesis in endometrial carcinomas arising in endometrial polyps, but this antibody is not correlated with tumor aggressiveness, P53 and Ki 67 expression. P53 and Ki 67 overexpression, instead, are associated with advanced stage, histologic subtype and deep myometrial invasion of neoplasm.     

Association of RB/p16-pathway perturbations with DCIS recurrence: dependence on tumor versus tissue microenvironment

The prevalence of ductal carcinoma in situ (DCIS) diagnoses has significantly increased as a result of active radiographic screening. Surgical resection and hormone and radiation therapies are effective treatments, but not all DCIS will progress to invasive breast cancer. Therefore, markers are needed to define tumors at low risk of recurrence and progression that can be treated by surgery alone rather than by adjuvant therapies. Initial analyses indicate that retinoblastoma (RB)-pathway perturbations occur at high frequency in DCIS and mirror the molecular alterations observed in invasive breast cancer. Particularly, the elevated expression of p16ink4a in DCIS was associated with loss of RB function and estrogen receptor-negative biology. Furthermore, high expression of p16ink4a in conjunction with Ki-67 was associated with increased risk of DCIS recurrence and progression to invasive disease in multivariate analyses. These data are consistent with a functional role for RB in modulating the invasive behavior of mammary epithelial cells. The tissue microenvironment is particularly relevant to the behavior of DCIS, and, surprisingly, elevated expression of p16ink4a in nonproliferative stroma was observed in a substantial fraction of cases. In this tissue compartment, p16ink4a expression was strongly associated with disease recurrence, independent of standard histopathologic features. Together, the data herein describe dual aspects of RB-pathway biology that are associated with disease recurrence through the epithelial or stromal compartment of DCIS.     

乳腺交界性病变不典型囊性导管的临床病理学特征

目的 探讨乳腺的一种新的交界性病变,即不典型囊性导管(ACD)的临床病理学特征及其意义。方法 对200例术前没有做活检取材(仅经细胞学或细针穿刺活检诊断)的乳腺癌乳房切除材料作全乳腺切片检查,观察ACD的临床病理学特征和免疫组织化学特性。结果 ACD的发生率为22％(44／200),多发于绝经前女性(P=0.001),存在部位以乳腺癌巢周边多见;连续切片(间隔50张切片,150μm距离)可观察到ACD向非浸润性导管癌的移行;36％(16／44)伴有ACD的乳腺癌病例,癌组织p53染色阳性,其中12／16伴有的ACD也同时呈现阳性改变(P=0．001);α-平滑肌肌动蛋白染色显示ACD的肌上皮与非浸润性导管癌的肌上皮相似呈明显的受压变薄;伴有ACD的乳腺癌和不伴有ACD的乳腺癌两组在雌激素受体(ER),雄激素受体(PR),p53、c-erbB-2和Ki-67蛋白,肿瘤大小以及淋巴结转移方面没有明显的区别;44例ACD c-erbB-2染色呈阴性,Ki67标记指数也明显低于其所伴有的乳腺癌。结论 鉴于ACD与癌巢间的连续性及与癌组织p53蛋白的共同表达特性,将ACD作为乳腺的一种交界性病变具有一定的意义。     

DEK overexpression is correlated with the clinical features of breast cancer

To investigate the clinicopathological significance of DEK overexpression in breast cancers, a total of 196 cases, including 20 of normal tissues, 12 of intraductal hyperplasia, 31 of ductal carcinoma in situ (DCIS) and 133 of invasive ductal carcinoma of the breast, were selected from the Department of Pathology, Yanbian Tumor Hospital for immunohistochemical staining of DEK, estrogen (ER), progesterone (PR) and Ki-67 proteins. In results, DEK protein had higher positivity in DCIS, compared with the adjacent normal breast tissues. Also, DEK protein was strongly positive in invasive ductal carcinoma of the breast on immunohistochemistry, which was significantly higher than normal breast tissues. However, only two (2/12) cases of intraductal hyperplasia of the breast showed positive staining for DEK protein. Additionally, DEK overexpression was significantly correlated with the increased proliferating index of Ki-67. For the histological grade, DEK positive rate was only 39.6% in G1 breast cancers, but significantly higher in G2 (92.3%) and G3 (97.0%) cases (P<0.05). Also, a strongly positive rate of DEK was lower in Stage-0 (21.4%) and Stage-I (40.9%) compared with Stage-IIa (87.5%), Stage-IIb (89.7%) and Stage-IIIa (92.3%) (P<0.05). And DEK protein showed higher expression level in < 3 years disease free survival breast cancers than it did in ≥ 3 years disease free survival cases (P<0.05). However, no statistically difference was found among DEK expression, lymph node metastasis, and ER and PR expressions. In conclusion, DEK overexpression appears to be associated with breast cancer progression and DEK may potentially be used as a breast cancer biomarker for the early diagnosis, prognostic evaluation and therapeutic target for breast cancer.     

COX-2 expression in DCIS: correlation with VEGF, HER-2/neu, prognostic molecular markers and clinicopathological features

AIMS: There is considerable evidence that links COX-2 to the development of cancer. The aim of our study was to assess, by immunohistochemistry, COX-2 expression in ductal carcinoma in situ (DCIS) and its possible correlation with HER-2/neu, vascular endothelial growth factor (VEGF) expression and other common immunohistochemical parameters (p53, ER, PGR, Ki67). METHODS AND RESULTS: Tissue samples of 49 archival cases of DCIS without any invasive component were analysed for COX-2, HER-2/neu, VEGF, oestrogen and progesterone receptors, Ki67 and p53 by immunohistochemistry using specific antibodies. COX-2 expression was detected in 43 (87.8%) tissue samples, of which 12 (24.5%) were graded as weak, 22 (44.9%) as moderate and nine (8.4%) as high expression. Only six (12.2%) lesions were negative for COX-2 expression. VEGF expression was detected in 93.8% of samples; 66.7% of lesions were found to be positive for HER-2/neu expression. Furthermore, COX-2 expression was significantly correlated with VEGF expression (P = 0.003). A significant positive correlation was also observed between COX-2 and HER-2/neu expression (P < 0.0001). CONCLUSIONS: Our results suggest that COX-2 is highly expressed in DCIS and takes part in the molecular pathway implicated in progression of breast cancer and may provide a rationale for targeting COX-2 in preinvasive breast cancer therapy.     

Histological markers that predict clinical recurrence in ductal carcinoma in situ of the breast: an Australian population-based study

AIMS: The incidence of ductal carcinoma in situ (DCIS) has increased substantially since the advent of widespread screening mammography. Identification of histological markers that predict recurrent disease is essential for optimal treatment management. To assist the clinico-pathological stratification of DCIS, we sought to determine histological markers of recurrence in DCIS in an Australian population-based series. METHODS: In a study of all DCIS reported in Victoria between 1988 and 1992, managed by breast conserving therapy (wide local excision or subtotal mastectomy) with or without adjuvant radiotherapy and/or hormonal therapy, the histological features of DCIS lesions with subsequent ipsilateral recurrence as in situ or invasive breast cancer were compared with a similarly managed control group of DCIS without recurrence. RESULTS: Large lesion size, presence of nuclear pleomorphism, absence of cellular polarisation and extensive necrosis were all significant predictors of recurrence (P<0.05). Primary and recurrent DCIS lesions had similar morphological features, and invasive recurrence was characterised by ductal type with high nuclear grade. CONCLUSION: This study identifies histological markers in DCIS associated with recurrence in an Australian population, and demonstrates similar histological appearances between primary and secondary lesions. These histological characteristics may be used to stratify DCIS subtypes and facilitate the future optimisation of disease management.     

Loss of bcl-2 expression in ductal carcinoma in situ of the breast relates to poor histological differentiation and to expression of p53 and c-erbB-2 proteins

Aim : This study (1) investigates the incidence of bcl-2 protein expression in a series of 108 cases of ductal carcinoma in situ (DCIS), including 25 with early invasive carcinoma, and (2) evaluates the relationship of bcl-2 expression to the histological grade of DCIS and to the expression of oestrogen receptor (ER), c-erbB-2 and p53 proteins.  

Methods and results

The expression of bcl-2, oestrogen receptor (ER), c-erbB-2 and p53 proteins was determined immunohistochemically. Cases were regarded as positive for individual antibodies when at least 10% of the DCIS cells showed positive staining. DCIS was graded histologically as well ( n  = 9), intermediately ( n  = 24), or poorly differentiated ( n  = 75). bcl-2 expression was documented in 57 cases (53%) and was strongly associated with the histological grade of DCIS ( P  < 0.0001). All cases of well-differentiated DCIS were bcl-2 positive and loss of bcl-2 expression was almost exclusively confined to poorly differentiated DCIS lesions. bcl-2 expression was also closely associated with positive ER status ( P  < 0.0001). Forty-seven of 57 (82%) bcl-2 positive cases were ER positive while 49/51 (96%) bcl-2 negative cases were ER negative. There was a significant inverse correlation between bcl-2 expression and both p53 protein expression ( P  = 0.0004) and c-erbB-2 expression ( P  < 0.0001). Nineteen of 24 (79%) p53 positive cases and 38/45 (84%) c-erbB-2 positive cases showed loss of bcl-2.  

Conclusions

Loss of bcl-2 expression occurs in poorly differentiated DCIS and is related to negative ER status and to positive p53 and c-erbB-2 status. This pattern of bcl-2 expression and its association with other biological markers in DCIS is similar to that reported in invasive breast carcinoma.     

Investigation of immunohistochemical ERα, ERβ and ERβcx expressions in normal and neoplastic breast tissues

Estrogen receptor alpha (ERα) is a well-established prognostic marker in breast cancer. The role of estrogen receptor beta (ERβ) in breast cancers is still under investigation. We aimed to investigate the clinicopathological significance and immunohistochemical expression patterns of ERα, total ERβ (ERβ) and its spliced variant ERβcx in normal breast, ductal carcinoma in situ (DCIS) and invasive ductal carcinoma (IDC). Our study population comprised 10 normal breasts, 26 DCISs and 44 IDCs. Immunohistochemical expression of these markers was investigated in sections of formalin-fixed, paraffin-embedded blocks by 2 observers. In invasive ductal carcinomas, ERβ expression had a significant positive correlation with ERα expression (p=0.013), while ERβcx expression was significantly associated with the presence of lymphovascular invasion (p=0.046). There was a significant relationship between ERα expression and low histological grade (p<0.0001). Similarly, ERα+/ERβ+ tumors (p=0.004) and ERα+/ERβcx+ tumors (p=0.008) were significantly associated with low histological grade, too. ERα expression (p=0.009), ERβcx expression (p=0.048) and ERα+/ERβ+ coexpression (p=0.002) increased significantly in progression from normal breast to invasive ductal carcinoma. Expression of ERα correlates with less aggressive phenotypic features, and ERβ expression is positively correlated with ERα expression in breast cancer. ERβcx is associated with aggressive features and can take part in the progression of invasive carcinoma. Increase in ERα+/ERβ+ coexpression, ERα expression and ERβcx expression in breast cancer progression indicates an enhancement in ER expressions or an alteration in expression patterns of different ER variants during mammary carcinogenesis.     

Cystic duct dilatations and proliferative epithelial lesions in mouse mammary glands upon keratin 5 promoter-driven overexpression of cyclooxygenase-2

Expression and pharmacological studies support a contribution of cyclooxygenase (COX)-2 to mammary gland tumorigenesis. In a recent transgenic study, mouse mammary tumor virus promoter-driven COX-2 expression in mouse mammary glands was shown to result in alveolar hyperplasia, dysplasia, and carcinomas after multiple rounds of pregnancy and lactation. In the study presented here, the effects of constitutive COX-2 overexpression in keratin 5-positive myoepithelial and luminal cells, driven by the keratin 5 promoter in a hormone-independent manner, was investigated. In nulliparous female mice, aberrant COX-2 overexpression correlated with increased prostaglandin (PG) E(2) levels and caused cystic duct dilatations, adenosis, and fibrosis whereas carcinomas developed rarely. This phenotype depended on COX-2-mediated PGE(2) synthesis and correlated with increased expression of proliferation-associated Ki67 in epithelial cells. No changes in the expression of apoptosis-related Bcl-2, caspase 3, or p53 were observed. Hyperproliferation of the mammary gland epithelial cells was associated with increased aromatase mRNA levels in this tissue. The spontaneous pathologies bear analogies to the human breast with fibrocystic changes. Intriguingly, strong COX-2 expression was observed in fibrocystic changes, as compared to low expression in normal breast epithelium. These results show for the first time that aberrant COX-2 expression contributes to the development of fibrocystic changes (FC), indicating that COX-2 and COX-2-mediated PG synthesis represent potential targets for the therapy of this most frequent benign disorder of the human breast.     

Tenascin patterns of expression in duct carcinoma in situ of the breast

Immunohistochemical methods were used to study tenascin (TN) expression in duct carcinoma in situ (DCIS) of the breast of different histologic types. We evaluated 82 lumpectomy specimens of DCIS. There were 5 cases of comedo type, 19 cases of noncomedo type, and 58 cases of mixed comedo and noncomedo type. In 44 cases, the intraductal carcinomas were associated with infiltrating (invasive) duct carcinoma. TN expression was studied by immunohistochemical methods using monoclonal mouse anti-human tenascin (DAKO-TN2M636; 1:50 dilution). Positivity was recorded on a scale of 0 to 2+ for presence of TN staining around tumor ducts and thickness of TN-stained fibrous bands. TN showed positive correlation between thick bands around comedo DCIS and thin bands around noncomedo DCIS. The TN score had statistically significant positive association with high nuclear grade (p 0.004), periductal inflammatory infiltrate of DCIS (p 0.0006), associated extensive central necrosis of DCIS (p 0.0005), and comedo DCIS (p 0.0004). TN expression in the stroma was positively associated with tumor size (p 0.00002), extensive central necrosis (p 0.02), comedo DCIS (p 0.0005), and associated invasive carcinoma (p 0.006). The TN score did not correlate with duct size, multicentric carcinoma, or associated microcalcification. These results demonstrate the different biological nature of DCIS comedo type and suggest its preinvasive potential.