HER2-neu Expression Research Articles

Expression of HER2neu in Ductal Carcinoma in situ is Associated with Local Recurrence

AimsDetermination of the risk of recurrence after local excision of ductal carcinoma in situ (DCIS) remains a challenge. Molecular profiling based on immunohistochemical staining to oestrogen receptor (ER), progesterone receptor (PR) and HER2neu improved risk prediction in invasive breast cancer, but few studies have evaluated if molecular classification of DCIS predicts local recurrence. We evaluated the expression of ER, PR and HER2neu in DCIS to determine if molecular classification predicts local recurrence after breast-conserving therapy for DCIS. Materials and methodsWe reviewed the records of patients with DCIS treated between 1987 and 2000, carried out a pathology review and immunohistochemical staining for ER, PR and HER2neu and categorised cases into four molecular phenotypes [luminal A (ER+ and/or PR+, HER2neu–), luminal B (ER+ and/or PR+, HER2neu+), HER2neu subtype (ER–, PR–, HER2neu+), triple negative (ER–, PR–, HER2neu–)]. We evaluated the association between the molecular subtype and the development of local recurrence. ResultsIn total, 180 cases of DCIS were included in the study (luminal A, n=113; luminal B, n=25; HER2neu type, n=29; triple negative, n=13). The median follow-up time was 8.7 years. We observed higher rates of local recurrence among luminal B (40%) and HER2neu type (38%) DCIS compared with luminal A (21%) and triple negative (15%) DCIS. On multivariable analysis, HER2neu overexpression was associated with an increased risk of local recurrence (hazard ratio=1.98; 95% confidence interval: 1.11, 3.53, P=0.02). ConclusionHER2neu expression in DCIS is a significant predictor of local recurrence, whereas luminal A and triple negative phenotypes are associated with relatively low risks of local recurrence.

Predictors of disease progression in ductal carcinoma in situ of the breast and vascular patterns

Breast carcinoma-induced angiogenesis helps meet growing metabolic needs of tumors and progressively increases with malignant transformation of benign ducts to ductal carcinoma in situ (DCIS) and ductal carcinoma in situ to invasive carcinoma. There are conflicting data regarding the difference in angiogenesis in low-, intermediate-, and high-grade ductal carcinoma in situ. If angiogenesis is related to ductal carcinoma in situ progression, the types of ductal carcinoma in situ with more aggressive biologic potential would have different vascular patterns than the less aggressive ones. In this study, we classified 51 cases of ductal carcinoma in situ as low (10-20 years to progression to invasive carcinoma), moderate, or high aggressive (2-5 years to progression to invasive carcinoma), based on criteria outlined by Tsikitis and Chung (Am J Clin Oncol 2006; 29:305), which takes into account nuclear grade, mitotic rate, Ki-67, Her2Neu, P53, estrogen, and progesterone receptor expression. We correlated these 3 groups of ductal carcinoma in situ with the extent of periductal and stromal vascularity and the presence and type of vascular breaks. No association of aggressive biologic behavior of ductal carcinoma in situ with any vascular pattern was found. Moreover, no correlation was found between vascular patterns and classifiers of aggressiveness, microvascular density, or outcome (local recurrence, invasive carcinoma, or metastatic disease). To validate our cohort, we confirmed expected correlations of all measured parameters of aggressiveness by correlating them with each other. In summary, vascular patterns in ductal carcinoma in situ do not correlate with the predictors of aggressive behavior, suggesting that the biologic potential of ductal carcinoma in situ is independent of angiogenesis.

Her-2 neu (Cerb-B2) expression in fine needle aspiration samples of breast carcinoma: A pilot study comparing FISH, CISH and immunocytochemistry

Background:Breast cancers with Her-2 neu gene amplification are recognized as important markers for aggressive disease and targets which respond to therapy with trastuzumab. Her-2 neu testing on histological sections is routinely performed to select patients who may benefit from anti- Her-2 neu therapy. Few reports are available which document Her-2 neu status on fine needle aspirates (FNA).Aim:This pilot study is to document expression of Her-2 neu (Cerb-B2) on cytospin smears from FNA of patients with breast carcinoma.Materials and Methods:Twenty samples of FNA already collected for diagnostic purposes from patients with primary breast carcinoma were studied for demonstration of Her-2 neu expression by immunohistochemistry (IHC), Fluorescent in-situ hybridization (FISH) and chromogenic in-situ hybridization (CISH) on cytospin smears from FNA. Their expression was compared with tissue sections where possible.Results:Good correlation was observed between Her-2 neu protein expression and gene amplification in cytospin smears. Three of five (60%) breast carcinomas cases with 2+ and 3+ staining on IHC showed gene amplification by FISH and CISH. Three of 7 (43%) and 5 of 7 (71%) cases negative/1+ staining on IHC did not show gene amplification by FISH and CISH respectively. Tissue sections from 10 cases with 2+ and 3+ staining for Her2neu by IHC showed gene amplification in 8 cases.Conclusion:Demonstration of Her-2 neu by IHC, FISH or CISH in FNA is possible and may play a role in the management of patients with advanced breast cancer or those cases where surgical resection is not advisable.

Abstract LB-393: Characterization of triple-negative breast cancer patients among Hispanics in Puerto Rico

Abstract Introduction: Hormone receptor status had been correlated with prognosis in breast cancer patients. Breast cancer patients with no expression of estrogen receptor (ER), progesterone receptor (PR) and her-2 neu (her-2) have been grouped in the Triple Negative Breast Cancer (TNBC) category. At present, these patients have very limited adjuvant therapy alternatives. TNBC has been correlated with poor prognosis among African-American and Hispanic populations. However, there is very few data regarding the prevalence and characterization of TNBC among Hispanics groups (e.g. Puerto Ricans, Cubans, etc.). The objective of this study is to characterize the TNBC population among the Puerto Rican population and to assess whether expression of ER change the characteristics of the disease in this group. We hypothesize that expression of ER+ in her-2 negative breast cancer tumors will change the initial presentation of the disease. Methods: This cross-sectional study analyzed data from female patients with breast cancer diagnosed between 2000 and 2005, at the I. Gonzalez Martinez Hospital and the Auxilio Mutuo Hospital (n=1,082) in San Juan, Puerto Rico. Information on Her-2 status and other clinical characteristics was retrieved from the hospital's cancer registries and from medical record review. This study was approved by the Institutional Review Boards of the participating hospitals. Logistic regression models were used to evaluate the associations between relevant clinical characteristics and TNBC. We also evaluated whether there were any interaction between age, tumor size and receptor status. Results: The prevalence of TNBC in our study was 16.3% and 58.7% for the Her-2-ER+s phenotype. Patients in the TNBC group have younger age (<50 yr.) at diagnosis (24.3% vs 75.7%), bigger tumor size (>2cm) (70.3% vs 45.6%), invasive ductal histology (87.3% vs 68.7%), and higher tumor grades (III-IV) (49.7% vs 13.2%) compared to Her-2-ERs+. Using multinomial models, we found that, compared to TNBC, women with Her-2-ER+PR- and those with Her-2-ER+PR+ were 1.99 (CI95%= 1.15,3.44) and 1.66 (CI95%= 1.11, 2.46) times more likely to have > 50 years at diagnosis as compared to those with <50 years. This relationship is not seen in women with Her-2-ER-PR+ tumors. Furthermore, compared to TNBC, women with Her-2-ER+PR-are 0.4 (IC95%=0.24-0.67) times more likely to have tumors > 2cm. There was no interaction between receptor status, age and tumor size (p=0.582). Conclusions: TNBC in Hispanics from Puerto Rican origin showed the same prevalence of Hispanic women in California. Furthermore, disease characteristics (early age at diagnosis, tumor size, and histology and tumor grade) were also similar, suggesting that race has a significant effect in the presentation of TNBC in Hispanic women. In addition, the expression of ER, but not PR in Her-2 negative patients dramatically changes the initial presentation of the disease. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-393.

A First Report of Incidence Rates (Not Prevalence) by Breast Cancer Subtypes.

Abstract BackgroundWe and others have recently repeatedly documented that among women diagnosed with invasive breast cancer in the U.S., the majority of cancers (based on clinical evaluation of the Estrogen Receptor (ER), Progesterone Receptor (PR) and Her2-neu (HER2) expression) classify as ER positive(+)/PR+HER2 negative(-), (50-70%). HER2+ breast cancers represent another 10-20% of all diagnosed breast cancers and Triple Negative (ER-PR-HER2-) cancers 12-20%. It is also now widely accepted that these prevalences vary by race and age, with young and black women most likely to be diagnosed with Triple Negative tumors—reasons for which remain unexplained. However, to our knowledge, no one has reported actual incidence rates for the breast cancers by the subtypes described above. We report the first incidence rates for breast cancer subtypes overall (and by race and age groups) and extrapolite new annual case burdens.MethodsAfter identifying invasive breast cancer cases through the Surveillance Epidemiology End Results (SEER) cancer registry, case medical records and cancer registry abstracts were investigated to determine ER, PR, and HER2 status among 1842 female residents of two large metropolitan Atlanta counties (Fulton and Dekalb) diagnosed with breast cancer during 2003-2004 (52.5% White, 44.2% Black, 1.6% Hispanic, and 1.7% 'Other'). Four triple subtypes were defined, based on positive (+) or negative (-) expression of ER, PR, and HER2: ER-PR-HER2- (Triple negative), ER-PR-HER2+, ER+/PR+HER2-, and ER+/PR+HER2+. SEER prep and SEER stat were used to calculate age adjusted incidence rates per 100,000, based on the 2000 U.S. census data representing our study population.ResultsTable 1. Age-adjusted Incidence Rates* of Women Diagnosed with Primary Invasive Breast Cancer by Triple Subtypes, Atlanta, GA 2003-2004 All breast cancersTriple NegativeER-PR- HER2+ER+/PR+ HER2+ER+/PR+ HER2-Unknown SubtypeAll women176.027.88.912.298.928.2 White179.319.47.512.6110.729.0Age<5066.38.73.27.637.79.0Age50+357.936.414.420.5226.160.5 Black171.236.310.012.186.726.1Age<5069.615.75.77.428.112.7Age50+331.869.016.919.5179.247.2 *Rates per 100,000 and adjusted to the 2000 U.S. population census for Fulton-Dekalb counties, Atlanta, GA.ConclusionTriple negative incidence rates differ markedly with rates for black women nearly double those for white women, regardless of age, and at the expense of the more treatable ER+/PR+Her2- tumors. Incidence rates, though, are highest for ER+/PR+ HER2- breast cancer among both black women and white women ages 50 and over. Our overall incidence rates representing two large metropolitan Atlanta counties are comparable to the SEER 9 area U.S. rates for the same time period among women ages 50+. However our rates are higher for women under age 50. Thus, the triple negative incidence rates we report can be cautiously extrapolated to an annual burden of new U.S. triple negative cases of 5,000 for black women and 16,000 for white women. Variations in actual incidence rates of breast cancer subtypes warrant in-depth population-based explorations. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 3065.

Current data of targeted therapies for the treatment of triple-negative advanced breast cancer: empiricism or evidence-based?

Approximately 10 – 15% of breast carcinomas (BCs) are known to be ‘triple–negative (TN) receptor’ (i.e., not expressing ER or PR and not exhibiting overexpression and/or gene amplification of HER2-neu). Triple-negative BCs comprise approximately 85% of all basal-type tumours. Classically, basal-like BCs have been characterised by low expression of ER, PR, and HER2 neu and high expression of CK5, CK14, caveolin-1, CAIX, p63, and EGFR (HER1), which reflects the mammary gland basal/myoepithelial cell component. Although there is no standard first-line chemotherapy regimen for metastatic TN BCs, anthracycline- and taxane-containing regimens are acceptable treatments. A large number of agents, including DNA-damaging agents, EGFR inhibitors, antiangiogenic agents and novel taxane formulations are currently being tested in clinical trials for first-line and pretreated patients. Limited experiences with platinum salts, poly(ADP-ribose) polymerase (PARP) inhibitors, cetuximab, bevacizumab and ixabepilone have been published in recent years and will be reported. Novel immunohistochemistry analysis for identification of basal like/TN phenotype are awaited to correctly select this population. The clinical trials investigating new agents have to be designed for a specific (and possibly large) subset of patients with BC. In the future, a gene array platform with greater sensitivity for distinguishing the various BC subtypes, as well as having the power to predict the molecular biology of the disease, will be an indispensible tool for treatment selection. Currently, treatment of TN BC is more empirical than evidence-based. The cornerstone of treatment is chemotherapy, but in the near future, novel target agents will emerge as possible partners.

Glycolytic phenotype in breast cancer: activation of Akt, up-regulation of GLUT1, TKTL1 and down-regulation of M2PK

Metabolic dependence on glucose utilisation has been described for different tumours characterised by activation of Akt, upregulation of GLUT1, M2PK and TKTL1. To date, however, little is known about glucose metabolism in breast cancer tissue. We analysed 55 breast cancer specimens, 26 adjacent ductal carcinomas in situ (DCIS) and 23 adjacent normal breast tissues for expression of glycolytic markers by immunohistochemistry. We found expression of pAkt in 49%, GLUT1 in 25%, M2PK in 68% and TKTL1 in 31% of the tumours investigated. Expression of pAkt and Her2neu are positively correlated with borderline significance (P = 0.055). Expression of pAkt, GLUT1 and TKTL1 were higher in breast cancer and DCIS than in normal tissue. Surprisingly, M2PK expression was highest in normal breast tissue. We found a glycolytic phenotype in a high percentage of breast cancer samples. Inhibition of glycolysis might evolve as a future option for breast cancer therapy.

Chemokine receptors in advanced breast cancer: differential expression in metastatic disease sites with diagnostic and therapeutic implications

Background: We investigated the expression of CXCR4, CCR7, estrogen receptor (ER), progesterone receptor (PR) and HER2-neu in human metastatic breast cancers to determine whether these biological biomarkers were preferentially expressed in any organ-specific metastases.Materials and methods: CXCR4, CCR7, ER, PR and HER2-neu expression levels were evaluated by immunohistochemical staining using paraffin-embedded tissue sections of metastatic breast cancers (n=41) obtained by either diagnostic biopsy or surgical resection.Results: The metastatic sites included the following: bone (n=15), brain (n=14), lung (n=6), liver (n=2), and omental metastases (n=2). CXCR4 was expressed in 41% of cases, CCR7 expression was demonstrated in 10%, and HER2-neu overexpression was present in 27%. CXCR4 was more likely to be expressed in bone metastases than visceral metastases (67% versus 26%, P=0.020). Visceral sites demonstrated a lower rate of CXCR4 positivity (33% and 23%, respectively, for lung and brain metastases). Similarly, CCR7 was more likely to be found in bone metastases than visceral sites (27% versus 0%, P=0.037).Conclusions: These results indicate that CXCR4 can contribute to the homing of breast cancer cells to the bone. This finding might have important clinical implications since patients with metastatic bone disease may achieve the highest benefit from a CXCR4-targeted therapy.

Pulmonary metastases of breast cancer. When is resection indicated?

While resection of pulmonary metastases is a common treatment in other primaries, the role of breast cancer metastasectomy is still unclear. The objective of the present study was to investigate the clinical outcome of our operated patients with pulmonary breast cancer metastases and discuss the different indications for metastasectomy. From January 1998 to December 2007 we retrospectively analysed 47 patients with histologically proven pulmonary metastases from breast cancer. The mean age of the 47 female patients was 56.2 years, the median disease-free interval (DFI) was 3.66 (0-25.8) years and the median follow-up was 20.6 months (3.2-110). The grading of the metastases was higher than the primary tumour in 12 of 45 (26.7%) and lower in 6 of 45 (13.3%) patients. R0, R1 and R2 resections were achieved in 27, 6 and 14 cases. The oestrogen receptor status of the metastases differed from the primary tumour in 11 out of 39 (28.2%) tested cases. Her2-neu receptor status differed in 4 out of 16 tested patients. The histological reports described a tumour spread around the metastasis in lymph or blood vessels in at least one resection specimen in 25 out of 47 (53.2%) patients. The rate of major complications was 5.8%. The overall survival from the first pulmonary metastasectomy was 32 months with a 5-year survival of 36%. The main prognostic factor was the oestrogen receptor status with a 5-year survival for receptor positive patients of 76% and 12.1% for receptor negative ones (p=0.002). A similar survival difference was found for the status of Her2-neu receptor (p=0.037). No prognostic influence could be demonstrated for age, number of metastases, initial tumour stage, complete versus incomplete resection, lymphatic spread, lymph node or parietal pleural involvement. The gain in life expectancy in breast cancer patients with pulmonary metastases is based on chemotherapy and antihormone treatment. Tissue of the lung metastasis is needed to adjust medical therapy to oestrogen and Her2-neu expression and to reliably rule out primary lung cancer. In case of proved pulmonary metastases, the level of evidence for a curative approach is low but some patients might benefit.

Predictors of clinical outcome in triple negative breast cancer: An institutional review

22180 Background: Triple negative (−) breast cancer (BC) is defined as lack of expression of estrogen, pregesterone and Her2 neu, being considered a high risk group. We study the pathological characteristics and the clinical outcome of a cohort of 64 consecutive triple negative(−) BC patients (pts) diagnosed at our institution. Methods: between january 1999 and june 2007, 64 (stage I to III) triple(−) BC pts. were retrospectively analyzed. All the Pts. had their receptor status and staining for Her neu by a single pathologist at the Institute of Oncology “Ángel H.Roffo”, (UBA). Univariate and multivariate analysis for the risk factors associated with relapse, and the log rank test for disease free survival (DFS), and overall survival (OS) was performed. Results: The median age was 52 years (range, 28–84 years).The median follow up time was 32 months (range, 3.−106). Overall, relapse rate is 42% (n=27), and 56 Pts (87.5%) are alive. Median DFS was 23 months (range, 3–106 months). Stage at the time of presentation was: 14 (22%) stage I; 29 (47%) stage II; 21 (31%) stage III. Pre-menopausal status was 46.6%, and 41% were limph node positive. Median OS: Stage I, II, and III was 33 (3–106), 30 ( 6–87), and 20 months (6–53) respectively, (p=0.0186). Median DFS: stage I, II and III was 33 (3–106), 27 (6–87), and 16 months (5–38) respectively, (p=0.0011). Pathological parameters analyzed ( nuclear grade, histologic grade, mitotic index and vascular invasion) were not significant predictors for DFS and OS by multivariate analysis, however vasculo-lymphatic invasion showed significance for both clinical outcomes in the univariate analysis, (p value 0.04 and 0.02, respectively). Conclusions: Triple (−) BC has an aggressive clinical course with significant risk of either local or systemic relapse and subsequent increased risk of death on short term follow up. In this triple (−) population only stage of disease was the only independent prognostic factor for DFS and OS. Between the pathological features analyzed only vasculo-lymphatic invasion seems to have prognostic significance. No significant financial relationships to disclose.

Cerebral metastases (CM) in breast cancer (BC) with focus on triple-negative tumors

1010 Background: We analysed the impact of a specific biologic subtype— triple-negative breast cancer (TN-BC)—characterized by lacking expression of steroid receptors and Her2neu (ER-/PR-/Her2neu-) on the occurrence of CM. Methods: Retrospective analysis of in 3,193 BC pts. treated consecutively between 1989 and 2006 in our EUSOMA breast unit. All pts. developing CM were included. Results: 338 pts. suffered from TN-BC (10.58%) and 80 pts. (2.5%) developed CM within a median observation period of 51 months (mos). 19 pts (23.75%) of all pts with CM had TN-BC and 5,6% of all pts with TN-BC had CM. Multivariate analysis indicated the highest risk for developing CM for TN-BC compared with all other types of BC (OR:4.16; 95%CI:2.26–7.64; p<0.001). Further independent risk factors were: ER-/PR-/Her2neu+ (OR:3.43; 95%CI:1.46–8.05; p=0.005), age ≤ 50 years (OR:2.02; 95%CI:1.17–3.49; p=0.012), UICC-stages III/IV (OR:2.49; 95%CI:1.19–5.22; p=0,016) and positive nodal-status (OR:2.37; 95%CI:1.10- 5.10; p=0.028). In addition, TN-BC showed inferior PFS and OS with 23 and 31.5 mos compared to 49.5 and 72 mos in other BC types (PFS:OR:3.2; 95%CI:2.1–18.1; p<0.0001/OS:OR 3.3;95%CI:2.7–18.8; p<0.0001), respectively. The median interval between primary diagnosis and occurrence of CM was 22 mos in TN- BC and 51 mos in other BC (OR:2.7; 95%CI:2.4–11.4; p<0.0001). OS after occurrence of CM was 4 mos in TN-BC and 8 mos on other BC, this trend was not significant. Conclusions: Patients with TN-BC have a higher risk for developing CM compared to other types of BC. This adds to a generally poorer prognosis in these pts. Even in pts. with CM, TN-BC represent a poor prognostic subgroup and CM occurred earlier and showed a trend for poorer outcome. Specific treatment regimens for this poor prognostic subgroup are warranted. Furthermore, the relative high incidence of CM in TN-BC questions if extended diagnostics (e.g. routine CNS MRI) should be applied to these pts. No significant financial relationships to disclose.

Genomic activation of the EGFR and HER2-neu genes in a significant proportion of invasive epithelial ovarian cancers

BackgroundThe status of the EGFR and HER2-neu genes has not been fully defined in ovarian cancer. An integrated analysis of both genes could help define the proportion of patients that would potentially benefit from targeted therapies.MethodsWe determined the tumour mutation status of the entire tyrosine kinase (TK) domain of the EGFR and HER2-neu genes in a cohort of 52 patients with invasive epithelial ovarian cancer as well as the gene copy number and protein expression of both genes in 31 of these patients by DGGE and direct sequecing, immunohistochemistry and Fluorescent in Situ Hybridisation (FISH).ResultsThe EGFR was expressed in 59% of the cases, with a 2+/3+ staining intensity in 38%. HER2-neu expression was found in 35%, with a 2/3+ staining in 18%. No mutations were found in exons 18–24 of the TK domains of EGFR and HER2-neu. High polysomy of the EGFR gene was observed in 13% of the invasive epthelial cancers and amplification of the HER2-neu gene was found in 10% and correlated with a high expression level by immunohistochemistry.Mutations within the tyrosine kinase domain were not found in the entire TK domain of both genes, but have been found in very rare cases by others.ConclusionGenomic alteration of the HER2-neu and EGFR genes is frequent (25%) in ovarian cancer. EGFR/HER2-neu targeted therapies should be investigated prospectively and specifically in that subset of patients.

Development of real-time quantitative reverse transcription-PCR for Her2 detection in peripheral blood from patients with breast cancer

BackgroundAmplification of the Her2neu oncogene is a well known indicator of poor prognosis in breast cancer patients. The implementation into the clinical setting of therapeutical strategies directly targeting the Her2neu gene product, has create the need for the development of non-invasive analytical techniques in order to monitoring minimal residual disease and response to the treatment. MethodsTo detect the expression of Her2neu mRNA in peripheral blood, we developed a specific real-time quantitative reverse transcription-PCR (QPCR) assay. The analyses were performed on blood samples obtained from 30 breast cancer patients positive for Her2neu overexpression by immunohistochemical analysis (IHC), 10 breast cancer patients negative for Her2neu overexpression, and 24 healthy controls. ResultsHer2neu positive tumors showed a significant increase in mRNA transcripts as compared with both healthy controls (n=24) and Her2neu negative patients (n=10). After establishing a cut-off value, 18 out of the 30 Her2neu positive patient scored positive for Her2neu expression, whereas only 1 out of the 10 Her2neu negative patients was weakly positive. ConclusionsHer2neu QPCR is suitable method for Her2neu overexpression detection in peripheral blood from clinical samples of breast cancer patients. QPCR could be used to identify breast cancer patients with poor prognosis and for monitoring response to the therapy.

Does HER2/neu expression affect adjuvant treatment choices in routine clinical practice?

10667 Background: HER2/neu (HER2) amplification is a recognized independent prognostic and predictive factor. We retrospectively examined the correlation between HER2 expression and adjuvant therapy in patients (pts) with early breast cancer (EBC) at our Institution (IOSI). Materials and Methods: From January 2003, 326 consecutive pts with EBC have been treated at IOSI and all clinical and pathological data have been prospectively collected into a specifically designed data base (CARMA). HER2 expression was assessed by the c-erbB-2 oncoprotein antibody (NCL-L-CB11) in 309 pts (95%). FISH was not routinely performed. Endocrine responsiveness (ER+) is defined as Er and/or PgR expression by immunohistochemistry. Results: Overall, 58 pts (19%) had HER2+ disease. FISH was performed in 15 pts (24%) with doubtful antibody overexpression (2+) and did not confirm HER2 amplification in 4 cases (27%). Nodal involvement was not assessed in 1 HER2+ (2%) and in 12 HER2- pts (5%) due to advanced age. Menopausal status did not differ in HER2 positive and negative pts (29% and 27% premenopausal pts, respectively). The distribution of adjuvant chemotherapy (CT) according to tumour characteristics in HER2 positive and negative pts was as follows: An anthracycline-containing regimen was chosen in 88% of HER2+ pts receiving adjuvant CT as compared to 70% of HER2- pts. Outside of a clinical trial adjuvant Tamoxifen was given to 57% of HER2+/ER+ pts as compared to 75% of HER2−/ER+ pts. Conclusions: Overall, HER2+ pts showed less endocrine responsiveness (64% versus 88%) and more extensive nodal involvement (43% versus 30% with &gt;3 positive nodes) as compared to HER2− pts. A greater proportion of HER2+ women received adjuvant CT, irrespective of other prognostic and predictive factors. In particular, a significant higher proportion of HER2+ pts with minimal nodal involvement (1–3 positive nodes) and ER+ disease received CT as compared to the equivalent HER2- subset (45% versus 26%, respectively). [Table: see text] No significant financial relationships to disclose.

Immunophenotyping of solid tumors, a new approach for molecular profiling solid tumors

10057 Methods of determining prognosis in breast, ovary, and colon cancer includes detection of molecular markers such as the estrogen receptor (ER), progesterone receptor (PR), and HER-2/neu. These markers are routinely assessed via immunohistochemistry (IHC) and Fluorescent In-situ hybridization (FISH). Flow cytometry (FC) has not yet been established for routine detection of these markers in tumors. We performed FC on cells from fourteen ovary, five breast, and four colon cancer tumors. FC was used to detect the presence of ER, PR, HER-2/neu, Epidermal Growth Factor Receptor (EGFR), E-cadherin, RAS, ERK and TUJ1. Cells obtained directly from tumoral tissue were simultaneous analysized by cell cycle stage (G0, G2M, and &gt; G2M) and presence of antigenic markers. In ovarian tumors, 7 of 14 samples demonstrated &gt;50% of cells with her-2/neu expression in aneuploid fractions (&gt;G2M), 9 of 14 in G2M phase, and only 2 of 14 in G0 phase. With respect to ER, 7 of 14 ovarian tumors exhibited &gt;19% of cells in G2M and &gt;G2M fractions, while only 1 of 14 samples had &gt;19% at G0. The PR receptor was expressed in &gt;19% of cells in 8 of 14 (&gt;G2M), 7 of 14 (G2M), and 2 of 14 (G0). In breast tumors, 4 of 5 tumors demonstrated &gt;50% expression of HER-2/neu in G2M and &gt;G2M cell phases, and only 2 of 5 demonstrated &gt;50% her2neu expression in G0. Three of five tumors had &gt;19% cellular expression of ER in the &gt;G2M and G2M phases, while only one had &gt;19% in G0. With PR, 3 of 5 breast tumors had &gt;19% cellular expression in G0, G2M, and &gt;G2M phases. In colon tumors, 2 of 4 had &gt;50% cellular expression of her2neu in &gt;G2M fractions, 4 of 4 in G2M, and only 1 of 4 in G0. Interestingly, cell fractions containing G2M and &gt; G2M DNA had &gt; 50% cellular expression for RAS and ERK in 2 of 4 colon tumors. No colon tumors expressed RAS and ERK significantly in G0 cells. We found that ER, PR, and HER-2/neu marker expression were consistent with established expression patterns. Additionally, these data point to specific expression patterns that are associated with the cell cycle stage. In particular, we note high expression levels of Her-2/nue, ER, and PR in aneuploid (&gt;G2M) cell populations. These results suggest that patterns of differential expression can be readily assessed in tumor tissues using flow cytometry, and has the potential for broad application to the study of solid tumors. No significant financial relationships to disclose.

Gene polimorphisms and biological factors as possible predictors for patients with pleural metastatic breast cancer

10671 Background: Pronosis of breast cancer varies widely in patients with similar clinical features. Therefore, we investigated other factors with possible prognostic value. The CYP17 gene has a polimorphism related to a increased susceptibility to breast cancer (single T; A1 allele to C; A2 allele). ( Mitrunen et al. Cancer Epidemiol Biomarkers Prev 2000,9;12;1343–8). On the other hand, it has been found in the MTHFR gene a common C-T substitution at nucleotide 677 that is associated with reduced enzyme activity and increased risk of breast cancer (Capmbell et al., Breast Cancer Research 2002;4,6:R14). Other genes such as GSTT1 and GSTM1 variant genotypes have a controversial association to breast cancer. We have analyzed the genotype and biological factors in a group of patients with pleural metastatic cancer of the breast and compared it to a control group of women with no cancer. Methods: Forty-six patients with breast cancer and forty women as controls were includes in the study. Age at diagnosis of the primary tumor, TNM staging, interval free of disease, estrogen and progesteron receptors, Ki67, p53 and Her-2neu expression were recorded, as well as survival from the time of diagnosis and polimorphism of CYP17, MTHFR, GSTT1 and GSTM1 genes. Results: Mean survival differed significantly between patients having pleural effusion as the first site of relapse as compared to those with other sites of relapse (28’9 vs 11’9 months, p = 0’004). A high Ki67 expression correlated best with longer survival after talc pleurodesis. We detected a high frequency in the CYP17 A2 variant genotype in patients with metastatic breast cancer compared to the control group. As for MTFHR gene, we found a high frequency for the T/T genotypes variant in breast cancer group. No significant differences between control and cancer group were found in either GSTT1 and GSTM1 gene variations. Conclusions: Genotype and biological factors, such as factors investigated in this study, play an important role as predictors for patients with pleural metastatic breast cancer. No significant financial relationships to disclose.

Understanding resistance to hormonal therapy in estrogen avid breast cancer

566 Background: The estrogen receptor (ER) is an essential target for endocrine therapy in breast cancer. We, and others, have tested an in vivo assay of ER function, [F-18]-16α-fluoroestradiol (FES) PET, which allows assessment of all sites of tumor spread and predicts response to hormonal therapy. However, not all patients with tumors bearing functional ER respond to therapy, suggesting the presence of mechanisms of resistance other than ER loss. Methods: 65 postmenopausal female patients with known ER+ metastatic (mostly bone and soft tissue dominant) breast cancer treated with aromatase inhibitors underwent PET FDG and FES imaging on an institutional imaging protocol (majority in second-line therapy). Hormone levels were assayed at the time of imaging, and biopsy specimens from either the primary tumor or metastasis were analyzed for expression of ER, PR, AR, PSA, HER2neu, and TopoIIα. Results: 49 (75%) patients had at least modest levels of retained ER expression, indicated by an average FES SUV &gt; 1.5 at the tumor sites. We found serum estradiol, or estrone, levels &gt;30pg/ml in 30% of patients. We found the majority of patients had PR, or AR expression, but few had EGFR or TopoIIα expression. FES SUV &gt; 1.5 predicted response to therapy (p=.0006); no patient with an FES SUV &lt; 1.5 had an objective response. Only 18/49 patients with SUV &gt;1.5 responded to AI therapy. In the subset with FES SUV &gt; 1.5, factors predicting lack of response to AI included PR negativity (p=.038) and/or detectable serum estradiol levels (p=.05). Conclusions: Quantitative PET FES predicts response to hormonal therapy. Estrogen levels may not be completely suppressed in patients on long-term AI therapy. Estradiol levels &gt; 30pg/ml and lack of PR expression predict resistance to endocrine therapy in patients with tumors bearing functional ER expression. Efforts to monitor and reduce estradiol levels may offer therapeutic benefit to patients. No significant financial relationships to disclose.

Clinical and pathological correlations in male breast cancer (MBC): Intratumoral aromatase expression (ITA) via tissue microarray (TMA)

10553 Background: Most cases of MBC are hormone-receptor (HR) positive and there is anecdotal evidence of disease response to aromatase inhibitors in the metastatic setting. Study objectives were to (i) assess clinical-pathologic characteristics in a cohort of MBC, (ii) evaluate intratumoral aromatase expression and (iii) assess its’ prognostic impact. Methods: A retrospective chart review was conducted for all cases of MBC seen at the Nova Scotia Cancer Centre between 1985–2005. Data abstracted included disease stage, tumor grade, HR status (ER/PR), HER2-neu and ITA expression as well as dates of disease recurrence and death. Tumor blocks were incorporated into 3 TMA’s with control specimens (gynecomastia, benign breast tissue, liver). Four 1.0 mm cores were taken of each tumor. Immunohistochemistry (IHC) was performed with the following antibodies; 6F11 ER primary Ab (Ventana, USA), 1A6 PR primary Ab (Ventana, USA), TAB250 monoclonal mouse anti-HER2 (Zymed, USA), DAKO 0185 rabbit anti-human c-erbB2 (DAKO, Canada), mouse anti-human p450 aromatase Ab (Cedarlane, Canada). ITA staining intensity and distribution was compared to benign hepatic tissue and if &gt; or = to liver was called ’strong’ while &lt; liver was called ’weak’. The log-rank test was used for survival comparisons. Kaplan-Meyer curves were used to estimate 5year disease-free (DFS) and overall (OS) survival probabilities. Results: 54 cases were identified with median age at diagnosis of 64 (31–85 yrs). Median tumor size was 2.6 cms (0.9–8cms) and 22 (41%) had nodal metastases. 45 cases had tissue available for IHC. Of these, 40 (89%) were ER and 33 (73%) were PR positive. HER2-neu was overexpressed in 4/42 (9.5%) and 12/45 (27%) had strong ITA expression. 5 yr DFS was 71% and OS was 64%. In univariate analysis strong (vs weak) ITA expression was associated with improved 5 yr OS (92% vs 49%, p = 0.038) but not DFS (82% vs 76%, p = 0.44) rates. Conclusion: The suggestion of improved OS but not DFS for those tumors with strong ITA expression may imply that these were more responsive to hormonal therapy in the metastatic setting or had a more indolent pace of progression. It is suggested that further investigation of ITA and response to hormonal therapy in MBC is warranted. No significant financial relationships to disclose.

EGFR expression and HER2/neu overexpression/amplification in endometrial carcinosarcoma

Objective. Endometrial carcinosarcomas are aggressive biphasic neoplasms traditionally treated as a high-grade uterine sarcoma. Epidermal growth factor receptor (EGFR) and HER2/neu (HER2) tyrosine kinases have been implicated in the development and progression of several human cancers and are targets for therapeutic intervention. The aim of this study was to evaluate for HER2 and EGFR expression in cases of endometrial carcinosarcoma. Methods. Formalin-fixed, paraffin-embedded sections from 55 cases of confirmed endometrial carcinosarcoma were immunostained with commercially available antibodies to EGFR and HER2. Fluorescent in situ hybridization for HER2 gene amplification was performed on all cases showing 2+ or 3+ HER2 staining by immunohistochemistry. HER2 gene amplification and EGFR expression were correlated with several prognostic variables. Results. EGFR expression was identified in the majority of tumors (45/55, 82%). HER2 overexpression (3+) was seen in 14/55 (25%) cases and HER2 gene amplification was seen in 11 (20%) cases. EGFR expression and HER2 gene amplification did not show significant correlation with disease progression, disease-free survival or overall survival. The carcinomatous component of tumors more frequently showed HER2 overexpression as compared to the sarcomatous component (25% vs. 4%, P = 0.008). The sarcomatous component of tumors more frequently showed EGFR overexpression as compared to the carcinomatous component (44% vs. 24%, P = 0.04). Conclusions. EGFR and HER2 appear to play a role in the carcinogenesis of endometrial carcinosarcomas. The carcinomatous and sarcomatous elements of these tumors showed consistent differences in HER2 and EGFR expression patterns supporting biologic differences between these components. Studies evaluating the clinical utility of HER2 or EGFR targeted therapy in these tumors appear warranted.

Demographic difference in elderly patients with breast cancer

872 Background: The elderly population in the United States is growing. There is enough evidence to suggest that elderly patients with cancer, who has good performance status, should be treated equally aggressively as younger patients. Based on these facts a retrospective analysis was done to look at the demographic difference in elderly patients with breast cancer. Methods: Records of 120 patients with breast cancer diagnosed and treated in rural Kentucky were analyzed for their demographics. Subsequently a comparison was done between patient groups of age younger than 70 years (Group-I) and age 70 years and older (Group-II). Results: Ninety-six patients were in Group-I and thirty-four patients were in Group-II. The oldest patient seen was 89 years of age. The important findings were as follows. An interesting observation was seen in six patients who were age 80 years or over. Four had stage II disease, four were hormone receptor negative, of which three had 2–3+ over expression of her2neu by immunohistochemistry. Conclusions:High number of elderly patients still had self /exam detected breast cancer, in spite of the recommendation for routine annual mammogram. Stage II breast cancer was 45%, similar to younger patients. Bilateral breast cancer even though uncommon was seen in 6%. Very elderly women (more than 80 years of age), showed bad prognostic factors like hormone negative status and her2neu over expression in more than 50% of the time. The above findings suggest that the trend is different in elderly population especially at age of 70 years and over. Further it supports the recent study from the University of Chicago that behavior of breast cancer is not more indolent in older patients compared to younger patients. Further attention and studies are recommended in this increasingly common very elderly sub-population to target treatments accordingly and aggressively. No significant financial relationships to disclose.