Gene methylation of oestrogen and epidermal growth factor receptors in neoplastic and perineoplastic breast tissues.

Oestrogen receptor (ER) and epidermal growth factor receptor (EGFR) gene methylation was evaluated in neoplastic and perineoplastic breast tissues from 20 patients. In both tissues, ER gene methylation was inversely correlated with protein levels, while EGFR gene methylation was not. A preferential ER gene hypomethylation was found in neoplastic tissues, suggesting a significant role in neoplastic transformation.     

Epidermal growth factor receptors in human breast cancer

Summary The capacity for specific binding of¹²⁵I-epidermal growth factor (EGF) was studied in crude membrane fractions from 95 human breast carcinomas. About 42% of the samples showed saturable, high affinity, specific binding of EGF. In 21% of the tumors we were able to demostrate high (above 10 fmoles/mg protein) binding capacity. Moreover, high EGF receptor values were associated with a low content of estradiol receptor. These studies are related to the definition of new biochemical markers in human breast cancer.     

Epidermal growth factor receptors and effect of epidermal growth factor on growth of human breast cancer cells in long-term tissue culture

Epidermal growth factor (EGF) may be important in regulating the proliferation of mammary epithelial cells. In the present study, we examined EGF binding and effect on growth in nine human mammary cell lines. The T-47D, MCF-7, SK-Br-3, AIAb 496, BT-20, and BT-474 tumor cell lines and a cell line (HBL-100) derived from milk exhibited EGF binding; both high (Ka 10(10) M-1)- and low (Ka 10(9) M-1)-affinity sites were detected. The total number of EGF receptors per cell of different cell lines varied from 1.6 X 10(3) sites/cell (for AIAb 496) to 1.5 X 10(6) sites/cell (for BT-20). The two floating cell lines, DU4475 and Lev III, had no detectable EGF binding. Effect of EGF on growth was studied by monitoring cell number and the incorporation of [3H]thymidine into DNA of cells maintained in Dulbecco's modified Eagle's medium supplemented with 0.1% fetal bovine serum. Using these procedures, only T-47D cells were stimulated by EGF at low concentrations (0.1 to 1 ng/ml). At concentrations higher than 10 ng/ml, EGF was inhibitory to varying degrees in most cell lines that contained EGF receptors. The growth of the two floating cell lines that had no detectable EGF binding was unaffected by EGF. Our results show that EGF receptors are not present in all human breast cancer cell lines. There is no apparent correlation between EGF binding and its mitogenic activity in cell lines with EGF receptors. EGF may have biological roles in human breast cancer other than growth regulation.     

Activity of ixabepilone in oestrogen receptor-negative and oestrogen receptor-progesterone receptor-human epidermal growth factor receptor 2-negative metastatic breast cancer

Oestrogen receptor (ER)-negative breast cancer, including oestrogen receptor-, progesterone receptor- and human epidermal growth factor receptor 2-negative (ER/PR/HER2-negative) breast cancer, is more aggressive than ER-positive disease. A major limitation in the treatment of ER-negative disease subtypes is the inherent insensitivity to hormonal agents (tamoxifen, aromatase inhibitors) that are widely used in the treatment of breast cancer. Thus, therapeutic options for poor prognosis patients with ER-negative breast cancer are limited to a handful of chemotherapeutic agents, and new agents are needed to improve the treatment of this disease.Ixabepilone, a novel epothilone B analogue with low susceptibility to cellular mechanisms that confer resistance to taxanes and other chemotherapeutic agents, has demonstrated potent preclinical antitumour activity in multiple models, including those with primary or acquired drug resistance. This review summarises the results of a prospective subset analysis from a phase III clinical trial evaluating ixabepilone for the treatment of metastatic breast cancer (MBC), in which efficacy and safety were evaluated in patients with ER-negative and ER/PR/HER2-negative disease.     

Receptors for epidermal growth factor and insulin-like growth factor I and their relation to steroid receptors in human breast cancer

Levels of epidermal growth factor receptor (EGF-R) and insulin-like growth factor receptor (IGF-R) in breast cancer tissue were evaluated. The binding of growth factors was compared to the content of estrogen receptors (ER) and progesterone receptors (PgR). EGF-R correlated negatively to the ER and PgR (Kendall correlation, P less than 0.001), whereas the IGF-R correlated positively to ER and PgR (analysis of variance, P less than 0.001). In contrast, no correlation was found between EGF-R and IGF-R. IGF-R binding was higher in tumor tissues than in adjacent normal tissues (Wilcoxon rank test, P less than 0.001), whereas the EGF-R binding in normal tissue did not differ from that in cancer tissue. The degree of differentiation in ductal breast cancer correlated to EGF-R (chi 2 test, P = 0.018), but not to IGF-R. The bindings of both growth factors were the same in metastases and primary breast tumors. Our results show that EGF-R and IGF-R are present in normal breast tissue and breast cancer tissue. The growth factor receptors are related to steroid receptor content and their presence is associated with malignant transformation of breast cells and dedifferentiation of breast cancer.     

Dual human epidermal growth factor receptor 2 blockade for the treatment of HER2-positive breast cancer

The introduction of trastuzumab into clinical practice changed the natural course of HER2-positive breast cancer. Currently, treatment with trastuzumab represents the standard of care for HER2-positive breast cancer and this treatment has been approved in the adjuvant, neoadjuvant, and metastatic settings. Besides trastuzumab, two other anti-HER2 agents—lapatinib and pertuzumab—have been approved for the treatment of HER2-positive advanced breast cancer. Strong biologic data support the concept of dual HER2 blockade, with different anti-HER2 agents demonstrating complementary mechanisms of action. Several neoadjuvant and metastatic studies performed in HER2-positive breast cancer using dual HER2 blockade have been proven to outperform anti-HER2 monotherapies. These dual combinations of agents represent a promising therapeutic strategy that is now reaching clinical practice. In this review we describe the results of studies utilizing dual blockade in patients with HER2-positive breast cancer.     

Receptors for epidermal growth factor and steroid hormones in human breast cancer

Epidermal growth factor (EGF) seems to play an important role in regulating the proliferation of human breast cancer. Fifty-five primary breast tumors and 7 lymph node metastases were simultaneously assayed for the presence of EGF receptors (EGFR), estrogen receptors (ER), and progesterone receptors (PR). Overall, 42% (23/55) of the tumors were EGFR positive. EGFR were more frequently present in ER- and PR-negative than in ER- and PR-positive tumors. In particular, a negative correlation between EGFR and PR (chi 2 = 6.8; p greater than 0.01) was observed. All metastatic tumors were EGFR negative, and in all cases but 1 the levels of EGFR were higher in metastatic than in primary tumors. Our results suggest the presence of a subclass of breast tumors, the growth of which is primarily regulated by EGF or EGF-like substances rather than by steroid hormones. In this group, not amenable to endocrine therapy, EGF receptors should represent a target for therapeutic intervention.     

Obesity increases the incidence of distant metastases in oestrogen receptor-negative human epidermal growth factor receptor 2-positive breast cancer patients

BackgroundObesity is a major negative determinant of breast cancer outcome. However, there are contrasting data on the differential impact of obesity on specific breast cancer subtypes. In particular, very little is known on human epidermal growth factor receptor 2-positive (HER2+) tumours.Patients and methodsWe assessed the prognostic role of increased body mass index (BMI) on a consecutive series of non-metastatic HER2+ patients treated at our institution before the introduction of adjuvant Trastuzumab. We separately analysed oestrogen receptor-positive (ER+) and -negative (ER?) HER2+ cases.ResultsIn ER?/HER2+ tumours we observed a significantly worse overall survival (Hazard ratio (HR) 1.79, p-value 0.041) and cumulative incidence of distant metastases (HR 2.03, p-value 0.019) in obese (BMI > 30) versus normal/underweight (BMI < 25) patients. Local relapses appeared to be non-significantly reduced in obese patients, masking the overall effect on disease-free survival. Outcome in ER+ tumours, instead, was not significantly different between BMI groups.ConclusionsObesity significantly correlates with worse overall survival and cumulative incidence of distant metastases in ER?/HER2 positive breast cancer. Differences in the biology of breast tumours may determine individual susceptibility to obesity. The biology of the underlying tumour should be taken into account in the design of dietary intervention trials in breast cancer.     

bcl-2 in normal human breast and carcinoma, association with oestrogen receptor-positive, epidermal growth factor receptor-negative tumours and in situ cancer.

The role of bcl-2 expression in solid tumours is as yet undefined. It was, therefore, the purpose of this study to investigate expression of bcl-2 protein in 111 human breast carcinomas using immunohistochemistry and the monoclonal antibody bcl-2 124. Expression was then compared with the established indicators of prognosis and biological behaviour in malignant breast disease. No relationship could be observed between bcl-2 and node status, tumour size, differentiation, type or age at excision. However, a strong positive relationship was seen between bcl-2 and oestrogen receptor (ER), with 70 of 88 (80%) bcl-2-positive tumours being ER positive also, compared with seven of 23 (30%) bcl-2-negative tumours being ER positive (P < 0.0001). The converse was found when bcl-2 was compared with epidermal growth factor receptor (EGFR). A strong negative relationship was observed, with 26 of 88 (30%) bcl-2-positive tumours being EGFR positive, compared with 16 of 23 (70%) bcl-2-negative tumours being EGFR positive (P = 0.001), raising the possibility that bcl-2 is an ER-regulated gene. An inverse relationship was also found between bcl-2 and the oncogenes c-erbB-2 and p53. Thus, loss of bcl-2 expression in breast cancer is associated with a range of molecular markers of poor prognosis and may define part of an ER-negative, EGFR-positive phenotype.     

Comparison between immunocytochemical and polymerase chain reaction techniques for detection of oestrogen receptor and transforming growth factor beta in breast cancer.

The utility of the polymerase chain reaction (PCR) as a technique for determining the expression of transforming growth factor beta (TGF-beta) and of the oestrogen receptor (ER) in clinical breast cancer tissue was examined. PCR analysis was compared with immunocytochemical assays for TGF-beta and for ER. Seventy confirmed breast carcinoma samples were analysed for ER using both techniques with a statistically highly significant concordance (P < 0.001) between the two methods. Nineteen samples were observed to be ER positive and 46 samples were found to be ER negative by both techniques. Forty-eight samples were analysed for TGF-beta using both PCR and immunocytochemistry. Of the 24 samples observed to be positive for TGF-beta by immunocytochemistry, all were found to be positive for TGF-beta mRNA (PCR). Similarly, the 24 samples observed to be TGF-beta negative by immunocytochemistry were also negative for TGF-beta mRNA, indicating 100% specificity and 100% sensitivity of the PCR technique. PCR is therefore considered a viable technique for analysis of both ER and TGF-beta in small samples such as fine-needle aspirates.     

Ectopic expression of epidermal growth factor receptors induces hormone independence in ZR-75-1 human breast cancer cells.

Epidermal growth factor (EGF) receptor is inversely related to expression of estrogen receptor (ER) and progesterone receptor in primary breast tumors and is a negative predictor for response to endocrine therapy. To investigate a possible causal role of EGF receptor expression in breast cancer progression to hormone independence, we have created an experimental cell system. Epidermal growth factor receptor complementary DNA was introduced in estrogen-dependent ZR-75-1 breast cancer cells, and the resulting ZR/HERc cells exhibited a mitogenic response to epidermal growth factor, thus bypassing estrogen dependence. This EGF-induced proliferation could not be inhibited by antiestrogens. In addition, we noted changes in cell morphology and keratin expression of EGF-stimulated ZR/HERc cells, suggestive of an altered differentiation state. Furthermore, intolerance of functional ER and EGF receptor signal transduction pathways in ZR/HERc cells was observed during simultaneous activation, which possibly explains the inverse relationship of ER and EGF receptor expression in primary tumors. In contrast to the parental cells, ZR/HERc cells rapidly progressed to a stable ER-negative phenotype when cultured in the presence of the antiestrogen hydroxy-tamoxifen. These results suggest a possible role for EGF receptor in progression of breast cancer to hormone independence.     

Modulating therapeutic effects of the c-Src inhibitor via oestrogen receptor and human epidermal growth factor receptor 2 in breast cancer cell lines

Purposec-Src is an important adapter protein with oestrogen receptor (ER) and human epidermal growth factor receptor 2 (HER2), which validates it as an attractive target for the treatment of breast cancer. A specific c-Src inhibitor, 4-amino-5-(4-chlorophenyl)-7(t-butyl)pyrazolo[3,4-d]pyrinidine (PP2), was utilised to block c-Src activity to identify targeted vulnerabilities affected by ER and HER2 in a panel of breast cancer cell lines.MethodsER, growth factor receptors and signalling pathways were detected by Western-blot. The DNA content of the cells was determined by using a DNA fluorescence quantitation kit. Cell cycles were analysed by flow cytometry.ResultsThe antiproliferative effect of PP2 closely correlated with the inhibition of c-Src mediated extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) and/or phosphoinositide 3-kinase (PI3K)/Akt growth pathways. Inhibition of c-Src tyrosine kinase predominantly blocked ER negative breast cancer cell growth, particularly the triple (i.e. ER, progesteron receptor (PR), and HER2) negative cells. In contrast, ER negative Sk-Br-3 cells with highest HER2 phosphorylation were resistant to PP2, in which hyper-activated HER2 directly regulated growth pathways. However, blocking c-Src recovered ER expression and down-regulated HER2 which made Sk-Br-3 cells regain responsiveness to 4-hydroxytamoxifen. The majority of ER positive cells were not sensitive to PP2 regardless of wild-type or endocrine resistant cell lines.Conclusionsc-Src mediates the essential role of growth pathways in ER negative breast cancer cells. The ER positive and HER2 over-activation are two important predictive biomarkers for the resistance to a c-Src inhibitor. These data provided an important therapeutic rationale for patient selection in clinical trials with c-Src inhibitors in breast cancer.     

HER-2 amplification, steroid receptors and epidermal growth factor receptor in primary breast cancer

Amplification of HER-2 oncogene was analysed in DNAs obtained from 291 primary human mammary carcinomas. 52/291 (18%) were found to contain amplified HER-2 oncogene. Moderate amplification (2- to 5-fold) was noted in 36/291 (12%). Thirteen tumors (4.5%) had a copy number of 5 to 10. A 10- to 20-fold and greater than 20-fold amplification was observed in 2 and 1 patient, respectively. Sample sizes allowed the determination of estrogen receptor (ER) and progesterone receptor (PgR) levels in 253/291 primary breast cancers. HER-2 gene amplification was noted in 14% of ER+ patients and in 28% of ER- patients, respectively (P = 0.02). Similarly a significantly greater number of PgR- primary mammary carcinoma exhibited an amplification of the HER-2 gene compared to PgR+ cases (22% vs. 16%, P = 0.01). Although statistically not significant, tumors with HER-2 gene amplification were found to have lower levels of ER and PgR. No association of HER-2 amplification with the androgen receptor and EGF receptor was observed. Present data combine to suggest that tumor progression is more stringently controlled by the oncogene upon loss of hormone dependency. Differences found in HER-2 amplification between steroid receptor positive and negative tumors could be helpful to define a specific subset of women to whom adjuvant therapy should be directed.     

A meta-analysis of oestrogen receptor,progesterone receptor and human epidermal growth factor receptor 2 discordance between primary breast cancer and metastases

BackgroundThe discordance in oestrogen receptor (ER), progesterone receptor (PgR) and human epidermal growth factor receptor 2 (HER2) status between primary and recurrent breast cancer is being intensively investigated and a large amount of data have been produced. However, results from different studies are heterogeneous and often conflicting. To highlight this issue, a meta-analysis of published data was performed.MethodsA literature search was performed using Medline, and all the studies published from 1983 to 2011 comparing changes in ER, PgR and/or HER2 status in patients with matched breast primary and recurrent tumours were included. We used random-effects models to estimate pooled discordance proportions.ResultsWe selected 48 articles, mostly reporting retrospective studies. Thirty-three, 24 and 31 articles were focused on ER, PgR and HER2 changes, respectively. A total of 4200, 2739 and 2987 tumours were evaluated for ER, PgR and HER2 discordance, respectively. The heterogeneity between study-specific discordance proportions was high for ER (I² = 91%, p < 0.0001), PgR (I² = 79%, p < 0.0001) and HER2 (I² = 77%, p < 0.0001). Pooled discordance proportions were 20% (95% confidence interval (CI): 16–35%) for ER, 33% (95% CI: 29–38%) for PgR and 8% (95% CI: 6–10%) for HER2. Pooled proportions of tumours shifting from positive to negative and from negative to positive were 24% and 14% for ER (p = 0.0183), respectively. The same figures were 46% and 15% for PgR (p < 0.0001), and 13% and 5% for HER2 (p = 0.0004).ConclusionOur findings strengthen the concept that changes in receptor expression may occur during the natural history of breast cancer, suggesting clinical implications and a possible impact on treatment choice.     

Role of epidermal growth factor receptor in breast cancer

Decades of research in molecular oncology have brought about promising new therapies which are designed to target specific molecules which promote tumor growth and survival. The epidermal growth factor receptor (EGFR) is one of the first identified important targets of these novel antitumor agents. Approximately half of cases of triple-negative breast cancer (TNBC) and inflammatory breast cancer (IBC) overexpress EGFR. Thus, EGFR inhibitors for treatment of breast cancer have been evaluated in several studies. However, results so far have been disappointing. One of the reasons for these unexpected results is the lack of biomarkers for predicting which patients are most likely to respond to EGFR inhibitors. Recent studies have shown that EGFR and its downstream pathway regulate epithelial-mesenchymal transition, migration, and tumor invasion and that high EGFR expression is an independent predictor of poor prognosis in IBC. Further, recent studies have shown that targeting EGFR enhances the chemosensitivity of TNBC cells by rewiring apoptotic signaling networks in TNBC. These studies indicate that EGFR-targeted therapy might have a promising role in TNBC and IBC. Further studies of the role of EGFR in TNBC and IBC are needed to better understand the best way to use EGFR-targeted therapy??e.g., as a chemosensitizer or to prevent metastases??to treat these aggressive diseases.     

Expression of estrogen,progesterone and epidermal growth factor receptors in primary and metastatic breast cancer

The prognostic value of epidermal growth factor receptor (EGFR) expression and its biological role in estrogen receptor-positive (ER⁺) and ER-negative (ER^?) primary breast cancer is controversial. In this study, distributions of ER, progesterone receptor and EGFR have been established using immunohisto-chemistry in both primary breast tumors and their matched axillary lymph node metastases of 26 patients or their matched distant metastases of 2 patients. In addition, 5 patients with bilateral breast cancer were studied. ER⁺ tumor cells were detected in 22 (69%) and EGFR⁺ tumor cells were detected in 11 (34%) primary breast carcinomas. Expression of ER and EGFR was inverse regarding the individual tumor cells in both primary tumors and metastases. Relationship of EGFR expression with poorly differentiated and large breast tumors was observed. Furthermore, primary tumors with a predominant lobular component were ER⁺ and, with one exception, EGFR^?. Invasive ductal carcinomas were more frequently EGFR⁺. No apparent differences in receptor expression were observed between primary tumors and lymph node metastases or chro-nously or metachronously occurring bilateral breast cancers. Only one ER⁺ primary tumor showed a switch to EGFR expression in the involved lymph node. Our study shows that a shift in receptor phenotype between primary tumors and lymph node metastases is a rare event and, thus, additional analyses of involved lymph nodes will not likely serve as a better predictor for response to anti-estrogen therapy. We conclude that expression of EGFR is not a prerequisite for development of metastases. © 1995 Wiley-Liss, Inc.