Background

Prognostic factors in breast cancer have exploded over the past several years. Pathologists have played a major role in identifying different histological and immunohistochemical markers that have a direct bearing on both the treatment and behavior of breast cancer.

OUTLINE

GENERAL	Activated Akt Kinase Age Angiogenesis Beta-Catenin Bilateral Cancers Biopsy effect CD105 Chromosomal abnormalities Cyclin D1 Cyclooxygenase-2 DNA Ploidy DCIS associated Epidermal Growth Factor Receptor Fibrotic focus Glutathione S-transferase Grading Hormone receptor status Ipsilateral Breast Failure (IBF) Ki-67 LCIS associated Lymph node status Lymphovascular invasion Mast Cells Microarray Gene Expression Mitotic figure count Mucin Neuroendocrine differentiation Nuclear features Oncogenes p53 Pregnancy PTEN Race Second primary breast cancer Size of tumor Skin Involvement Stromelysin-3 Topoisomerases Treatment inadequacies Tumor associated antigens Vascular endothelial growth factor
COMMONLY USED TERMS
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PROGNOSIS	CHARACTERIZATION
GENERAL BACKGROUND	The prognosis of breast cancer is influenced by a number of factors
Size of primary tumor	Tumors <1 cm with negative nodes have 98% survival at 5 yrs Tumors <2 cm with negative nodes have 96% survival
Lymph node involvement	Single most important factor in determining prognosis for early breast cancer 10 yr disease free survival is 70-80% with negative nodes One to three positive nodes 35-40% Ten or more positive nodes 10-15% Micrometastasis <0.2 cm are associated with better prognosis NOTE: 20-30% of node-negative women will have a recurrence and die within 10 yrs
Histologic type and grade of tumor	Better for tubular, papillary, colloid, medullary versus NOS types >80% of women with Grade I tumors survive 16 yrs
Estrogen and progesterone receptor status	50-85% of tumors have estrogen receptors 70% of tumors with estrogen receptors regress after hormone treatment Only 5% of receptor negative tumors respond
Proliferative rate of tumor	Higher proliferation rates have poorer prognosis High S-phase fraction by flow cytometry
Amplified oncogenes	See her2-neu
Degree of angiogenesis	High correlation with increased vessel density and metastasis
Lymphovascular invasion	Strong association with lymph node metastasis Dermal lymphatic invasion (inflammatory carcinoma) has 3 yr survival of 3-10%
GENERAL
Histopathologic types of benign breast lesions and the risk of breast cancer: case-control study. Shaaban AM, Sloane JP, West CR, Moore FR, Jarvis C, Williams EM, Foster CS. Department of Cellular and Molecular Pathology (A.M.S., J.P.S., F.R.M., C.J., C.S.F.), Department of Public Health (C.R.W.), Merseyside and Cheshire Cancer Registry Office (E.M.W.), University of Liverpool, Liverpool, U.K.	Am J Surg Pathol 2002 Apr;26(4):421-30 Abstract quote The hypothetical multistep model of carcinogenesis indicates that breast cancer develops via a series of intermediate hyperplastic lesions through in situ to invasive carcinoma. To identify the risk inherent within the different morphologic lesions, we have analyzed the data from 674 benign biopsy specimens comprising 120 cases who subsequently developed breast cancer and 382 controls (matched for age and date of biopsy) spanning a period up to 20 years of follow-up (mean 66.95 months). In this series we have confirmed an increased risk associated with certain types of benign breast lesions. Atypical lobular hyperplasia was the most significant risk factor for breast cancer with more unfavorable outcome in patients <50 years of age (p = 0.003) and a relative risk (RR) of 4.55 (confidence interval [CI] 1.77-11.69). Hyperplasia of usual type showed an RR of 1.53 (CL 1.10-2.13) with a statistically worse probability of survival (cancer-free time) for patients >50 years. For atypical ductal hyperplasia the RR was 2.03 (CI 0.80-1.39). Blunt duct adenosis was significantly more common in cases progressing to breast cancer compared with controls, showing an RR of 2.08 (CI 1.12-2.85). We describe in detail the criteria of morphologic changes observed in blunt duct adenosis and define, for the first time, the level of risk associated with each of its six subtypes. Improved knowledge of breast carcinogenesis will provide insight for defining high-risk groups thus resulting in improved screening and management regimens.
ACTIVATED Akt KINASE
Prognostic relevance of activated Akt kinase in node-negative breast cancer: a clinicopathological study of 99 cases. Schmitz KJ, Otterbach F, Callies R, Levkau B, Holscher M, Hoffmann O, Grabellus F, Kimmig R, Schmid KW, Baba HA. 1Institute of Pathology, University of Essen, Germany.	Mod Pathol. 2004 Jan;17(1):15-21 Abstract quote.   Patients with lymphnode-negative breast cancer show a 10-year tumor recurrence rate of approximately 30%. Therefore, it is important to identify high-risk patients who would benefit from further adjuvant therapy. For this purpose, we examined the activation state of two kinases important in the regulation of cell proliferation and apoptosis in a series of 99 node-negative breast cancer cases with a mean follow-up of 10 years: Akt and extracellular regulated kinase (ERK1/2). The activation of Akt and ERK1/2 was investigated by immunohistochemistry using phospho-specific antibodies. The results were correlated with HER-2/neu expression, histological grading, receptor status, overall survival (OS) as well as with cell proliferation (Ki67 immunoreactivity, mitotic count) and tumor apoptosis assessed by TUNEL staining. Activation of Akt (pAkt) but not activation of ERK1/2 (pERK1/2) correlated with HER-2/neu overexpression (P<0.05) and was related to reduced tumor apoptosis (P<0.05). No association was found between pAkt or pERK1/2 with cell proliferation assessed by Ki67 and mitotic count (MC). Survival analysis of receptor status, HER2/neu expression, histological grading, MC and pAkt immunoexpression showed a significant correlation with decreased OS, but only pAkt reached statistical significance in the multivariate Cox regression analysis (P=0.015). Activation of Akt in node-negative breast cancer may indicate aggressive tumor behavior and may constitute an independent prognostic factor of OS. The determination of pAkt status may be of value in identifying high-risk patients, who would benefit from adjuvant therapy, and gives a rationale to investigate new therapy strategies by specific inhibition of the Akt signaling pathway in breast cancer.
AGE
Pathology and heredity of breast cancer in younger women. Marcus JN, Watson P, Page DL, Lynch HT. Department of Pathology, Creighton University, Omaha, Neb.	J Natl Cancer Inst Monogr 1994;(16):23-34 Abstract quote The pathology of early-age onset breast cancer is considered here from three perspectives: 1) benign proliferative disease, 2) the cancers themselves, and 3) familial and hereditary breast cancer. Hereditary breast cancer, a subset of familial breast cancer featuring a strong autosomal dominant pedigree pattern and multiple primary cancers, has a strong predilection for younger women, accounting for about one half of breast cancers under age 30. With respect to benign proliferative disease, the increased relative risk of breast cancer associated with proliferative disease with atypia, about fourfold to fivefold for all ages, is doubled by the presence of a family history of breast cancer and amplified by young age. With respect to the carcinomas, the relative incidences of medullary carcinoma and ductal carcinoma in situ are increased in young women, while lobular and tubular carcinomas are decreased. Invasive breast cancer is higher grade and more proliferative in younger women, as measured by thymidine-labeling index, DNA flow cytometric S-phase fraction, and proliferation-associated proteins. The increased fraction of ductal carcinoma in situ and higher grade invasive cancers may help to account, respectively, for increased recurrence rates with conservative therapy, and more aggressive natural history in younger women. Familial breast cancers show trends for increased medullary type, but the effect is not independent of age. Weak associations of family history with tubular carcinoma have been reported, but data for associations with lobular carcinoma in situ and invasive lobular carcinoma are conflicting. Hereditary breast cancer as a class has higher tumor proliferation rates, an effect independent of age. Knowledge of the pathology and biomarker characteristics of BRCA1 gene-linked hereditary breast cancers, which account for a substantial fraction of breast cancers in younger women, should shed light on the nature of the responsible gene(s) and guide approaches to therapy and prophylaxis.
The histologic phenotypes of breast carcinoma occurring before age 40 years in women with and without BRCA1 or BRCA2 germline mutations: a population-based study. Armes JE, Egan AJ, Southey MC, Dite GS, McCredie MR, Giles GG, Hopper JL, Venter DJ. Molecular Pathology Laboratory, Victorian Breast Cancer Research Consortium, Peter MacCallum Cancer Institute, East Melbourne, Victoria, Australia.	Cancer 1998 Dec 1;83(11):2335-45 Abstract quote BACKGROUND: Women with breast carcinoma diagnosed before age 40 years have a greater prevalence of germline BRCA1 and BRCA2 mutations than women with breast carcinoma diagnosed at older ages. Several recognizable histologic characteristics have been identified in breast carcinoma from studies of BRCA1/2 mutation carriers who belong to multiple-case families. The authors attempted to determine whether breast carcinoma occurring before age 40 years in BRCA1 or BRCA2 mutation carriers who were not selected for family history could be distinguished histologically from one another and from breast carcinoma in women of a similar age without a germline BRCA1 or BRCA2 mutation. METHODS: The study undertook a histologic assessment of breast carcinomas diagnosed before age 40 years identified from a population-based study. RESULTS: Breast carcinoma in BRCA1 mutation carriers was associated with a distinct histologic appearance; these tumors were high grade, and had exceptionally high mean mitotic counts, a syncytial growth pattern, pushing margins, and confluent necrosis. Atypical medullary carcinoma was overrepresented in BRCA1 mutation carriers. All low grade tumors and tumors with low mitotic rates belonged to the group without BRCA1 or BRCA2 mutations. Pleomorphic lobular carcinomas and extensive intraduct carcinomas were more common in BRCA2 mutation carriers. CONCLUSIONS: Breast carcinoma occurring in women with a germline BRCA1 or BRCA2 mutation have recognizable histologic phenotypes, which may be useful in identifying individuals more likely to carry germline mutations. Histologic examination of breast carcinoma should become an important part of the evaluation of women seeking genetic testing for germline mutations in these breast carcinoma susceptibility genes.
Breast cancer in young women: clinicopathologic correlation. Bertheau P, Steinberg SM, Cowan K, Merino MJ. Laboratory of Surgical Pathology, Saint-Louis Hospital, Paris, France.	Semin Diagn Pathol 1999 Aug;16(3):248-56 Abstract quote It has been suggested that early-onset breast carcinomas may be different from those that occur in older women. The clinicopathologic characteristics of 191 young female patients (under 40 years of age) diagnosed with breast carcinoma (BC) were studied. Clinical history, staging, treatment and outcome were reviewed. Histology was assessed for tumor subtypes, invasive and in situ components, nuclear and histologic grades and lymph node status. Adjacent nontumoral breast tissue was evaluated. Clinically, 11 patients were stage 0, 21 stage I, 94 stage II, 38 stage III, 6 stage IV, and in 21 no information was obtained. Sixty five percent of patients had positive lymph nodes at diagnosis; 102 patients (54%) relapsed at a median of 29 months after diagnosis. Histologically, 180 cases were infiltrating BC, 150 ductal (83%), 19 lobular (11%) and 11 of special types (6%); 11 cases were ductal carcinoma in situ. We found no cases of medullary carcinoma. High nuclear grade and vascular invasions were frequent (68% and 67%, respectively) even in patients who remained disease-free at least 5 years after diagnosis (61% and 60%, respectively). Our study demonstrates that the histologic types of early-onset breast cancer are not different from other BC. However, BC in young women is often associated with histologic features of high-grade malignancy even in patients with better survival. Our results suggest that BCs in young women are different from those that occur in older women.
ANGIOGENESIS
Lymphangiogenesis does not occur in breast cancer. Agarwal B, Saxena R, Morimiya A, Mehrotra S, Badve S. From the Department of Pathology, Indiana University School of Medicine, Indianapolis, IN.	Am J Surg Pathol. 2005 Nov;29(11):1449-55. Abstract quote   Breast cancer metastasis predominantly occurs via lymphatic vessels. However, the study of lymphatic vessels and lymphangiogenesis has been hampered by lack of specific markers. Recently, antibodies directed against M2A (D2-40), Podoplanin, and Prox-1 that specifically mark lymphatic vessels in paraffin-embedded sections have become available. These were used to study lymphangiogenesis in archival paraffin sections of normal breast (n = 23), fibrocystic disease (n = 7), ductal carcinoma in situ (n = 32), invasive ductal carcinoma (n = 50), and invasive lobular carcinoma (n = 5). In addition, endothelial proliferation in lymphatic vessels was analyzed by dual-color immunohistochemistry with D2-40 and proliferating cell nuclear antigen (PCNA). Expression of D2-40, Prox-1, and Podoplanin was seen in lymphatic vessels but not in blood vessels. Lymphatic vessels were seen in the peritumoral area and as "entrapped" intratumoral vessels adjacent to preexisting normal lobules and ducts. Unlike angiogenesis, there was no increase of lymphatic vessel density in association with neoplastic transformation. On the contrary, a marked reduction in intratumoral lymphatic vessel density was seen in comparison to normal breast tissue, fibrocystic disease, and ductal carcinoma in situ (P = 0.0001). There was an increase in peritumoral lymphatic vessel density as compared with normal breast (P = 0.0001). However, the endothelial cells in the "entrapped" or the peritumoral lymphatic vessels did not show any expression of PCNA indicating minimal or no proliferative activity. This was in contrast to the strong expression seen in adjacent tumor cells and blood vessel endothelial cells. Thus, lymphangiogenesis was not evident when studied by lymphatic vessel density or by lymph vessel endothelial proliferation.
Long-term prognostic significance of neoangiogenesis in breast carcinomas: comparison of Tie-2/Tek, CD105, and CD31 immunocytochemical expression. Dales JP, Garcia S, Carpentier S, Andrac L, Ramuz O, Lavaut MN, Allasia C, Bonnier P, Charpin C. Department of Pathology, Centre Hospitalier et Universitaire Nord, Marseille, France.	Hum Pathol. 2004 Feb;35(2):176-83 Abstract quote. The immunocytochemical detection of Tie-2/Tek, CD105, and CD31 was assessed in a large series (n = 905) of breast carcinomas on frozen sections. Results were correlated with patients' long-term outcome (median, 11.7 years) to define the respective prognostic significance of these markers. Univariate (Kaplan-Meier) analysis demonstrated that higher expression of CD31 (P = 0.032), CD105 (P = 0.001), and Tie-2/Tek (P = 0.025) correlated with poorer survival. However, only greater CD105 expression could significantly (P = 0.035) identify node-negative patients with poorer survival. Moreover, in multivariate analysis, CD105 and Tie-2/Tek, but not CD31, expression proved to be independent significant prognostic indicators. Marked expression of CD31 (P = 0.024), CD105 (P = 0.001), and Tie-2/Tek (P = 0.01) also correlated with higher risk of metastases in node-negative patients. It is concluded that CD105 immunoexpression in breast carcinomas is an independent prognostic indicator in node-negative patients, better in terms of overall survival than Tie-2/Tek and CD31. Also, Tie-2/Tek expression proved more sensitive than CD31 expression in terms of prognostic significance. Compared with CD31, CD105 and Tie-2/Tek have more clinical relevance for patient monitoring and also can serve as targets for specific therapy, such as CD105 immunotoxins or Tie-2/Tek pathway blockade, as recently suggested in experimental studies.
BETA CATENIN
Study of phospho-beta-catenin subcellular distribution in invasive breast carcinomas in relation to their phenotype and the clinical outcome. Nakopoulou L, Mylona E, Papadaki I, Kavantzas N, Giannopoulou I, Markaki S, Keramopoulos A. 1Department of Pathology, Attikon Hospital, Athens, Greece.	Mod Pathol. 2006 Apr;19(4):556-63. Abstract quote   beta-Catenin has a crucial role in cell-cell adhesion as well as a signaling role as a member of the Wnt pathway. The aim of this study was to examine the clinicopathological and prognostic value of phosphorylated beta-catenin, as well as its relation to the tumors' phenotype, in breast cancer. Immunohistochemistry was applied on 141 paraffin-embedded breast tissue specimens for the detection of phospho-beta-catenin, ER, PR, c-erbB-2, p53, Ki-67, bcl-2, uPAR and TIMP-1. For each case, a phospho-beta-catenin index was determined by image analysis. Phospho-beta-catenin staining was detected in the cytoplasm and the nucleus of the malignant cells. Cytoplasmic phospho-beta-catenin was statistically higher in carcinomas of smaller tumor size (P=0.030), lower stage (P=0.026), decreased Ki-67 and high c-erbB-2 immunoreactivity (P=0.052 and P=0.037, respectively). Nuclear phospho-beta-catenin showed a parallel correlation with ER and ERbeta (P=0.022 and P=0.043, respectively), bcl-2 (P=0.042), uPAR in cancer cells (P=0.041) and TIMP-1, although the correlation was borderline (P=0.066). Cytoplasmic phospho-beta-catenin was found to be independently correlated with prolonged disease-free and overall survival (P=0.046 and P=0.002, respectively), whereas nuclear localization was correlated with a shortened overall survival (P=0.046). In conclusion, phospho-beta-catenin may have a different involvement in invasive breast carcinomas, according to its subcellular distribution. Nuclear localization seems to be related to an aggressive tumor phenotype, negatively affecting patients' overall survival, whereas cytoplasmic localization is associated with a favorable tumor phenotype and a longer disease-free and overall survival.
BILATERAL CANCERS
Bilateral carcinomas of the breast with local recurrence: analysis of genetic relationship of the tumors. Regitnig P, Ploner F, Maderbacher M, Lax SF. Department of Pathology, Division of Medical Oncology, Medical University of Graz, Austria.	Mod Pathol. 2004 May;17(5):597-602. Abstract quote   Local recurrence of bilateral breast carcinomas is rare, but of biological interest, since it is unclear as to which tumor the local recurrence is related to, the ipsilateral or the contralateral, or whether it is an independent neoplasm. The aim of this study was to investigate the genetic relationship of bilateral breast carcinomas to each other and to their local recurrences. Eight cases of bilateral breast carcinomas, five with and three without local recurrence were analyzed using a microsatellite assay for 13 microsatellite loci. The presence of loss of heterozygosity and microsatellite instability in the various tumors was used for clonal analysis. All eight bilateral breast carcinomas showed divergent alterations in at least two microsatellite loci, which ruled out a genetic relationship. Four of five local recurrences were genetically related to the ipsilateral tumor and unrelated to the contralateral tumor. Only one local recurrence that occurred 11.8 years after the surgery of an infiltrative lobular carcinoma simultaneously with distant metastases was genetically related to the contralateral breast carcinoma. Although the number of cases in our study is limited, there is evidence that local recurrence of bilateral breast carcinoma frequently arises from the ipsilateral tumor.
Multicentricity and bilaterality in invasive breast carcinoma. Lesser ML, Rosen PP, Kinne DW.	Surgery 1982 Feb;91(2):234-40 Abstract quote Multicentricity and bilaterality are well-established characteristics of breast carcinoma, but little is known about the relationship of these variables with each other. This question was explored by analyzing the data pertaining to 880 women with invasive breast carcinoma. Patients with multicentric carcinoma had bilateral disease more often than those whose carcinoma was apparently limited to a single quadrant. Among women who had lobular carcinoma in situ coexisting with infiltrating duct carcinoma or infiltrating lobular carcinoma, bilaterality and multicentricity were significantly more common than they were among patients whose only lesion was infiltrating duct or medullary cancer. Other variables associated with bilaterality and multicentricity were degree of ductal differentiation, tumor size, nodal status, type of tumor margin, intensity of lymphoid infiltrate, and menstrual status. Age at diagnosis and estrogen receptors were related to bilaterality but not to multicentricity. The following variables proved to be unrelated to bilaterality and multicentricity: family history of breast carcinoma, height, weight, and parity. The data obtained in this study tend to support a conclusion that multicentricity and bilaterality are manifestations of similar factors involved in the neoplastic transformation of mammary gland epithelium leading to the development of breast cancer.
Women with breast cancer: histologic findings in the contralateral breast. Roubidoux MA, Helvie MA, Wilson TE, Lai NE, Paramagul C. Department of Radiology, University of Michigan Medical Center, Ann Arbor 48109-0326, USA.	Radiology 1997 Jun;203(3):691-4 Abstract quote PURPOSE: To investigate contralateral breast biopsy histologic findings in women with breast cancer. MATERIALS AND METHODS: Histologic findings in 237 patients with breast cancer who underwent contralateral breast biopsy for clinically or mammographically detected abnormalities were retrospectively reviewed. Malignant findings were categorized by histologic type. Benign findings were categorized by risk of breast cancer. Comparison was made with mammographically guided breast biopsy results in 1,294 patients without breast cancer. RESULTS: Of the 237 patients, 168 (70.9%) had either malignancy or high-risk histologic findings. One hundred thirty-nine patients (58.6%) had malignant findings; 98 (41.4%) had benign findings. Of the 98 with benign findings, 29 (30%) had high-risk histologic findings. Thirty (33%) of the 91 patients with invasive cancer had invasive lobular carcinoma. Forty-seven (45.6%) of the 103 patients with malignant lesions at mammographically guided biopsies had ductal carcinoma in situ alone. CONCLUSION: Compared with biopsy in women without breast cancer, contralateral biopsy in women with breast cancer was more likely to show malignancy, invasive lobular carcinoma, or ductal carcinoma in situ alone (P < .001) or to show high-risk histologic benign findings (P < .001). Mammographic and clinical findings in the contralateral breast should be regarded as more suspicious than those in patients without known breast cancer.
A case-control study of unilateral and bilateral breast carcinoma patients. Newman LA, Sahin AA, Bondy ML, Mirza NQ, Vlastos GS, Whitman GJ, Brown H, Buchholz TA, Lee MH, Singletary SE. Department of Surgical Oncology, The University of Texas M. D. Anderson Cancer Center, Houston 77030, USA.	Cancer 2001 May 15;91(10):1845-53 Abstract quote BACKGROUND: Women with unilateral breast carcinoma are at increased risk for developing contralateral disease. The clinical significance of bilateral breast carcinoma has not been fully defined, and the subset of patients who may benefit from medical or surgical risk-reduction intervention has not yet been characterized. The purpose of this study was to evaluate risk factors and outcomes for bilateral breast carcinoma. METHODS: A subject group of 70 bilateral breast carcinoma patients (62% metachronous) was matched by age and survival interval with a control group of 70 unilateral breast carcinoma patients. Median follow-up was 103 months. RESULTS: Eighty-two percent of the unilateral patients and 80% of the bilateral patients had Stage I or II disease at diagnosis. Median age at presentation was 53 years. In the bilateral group, the contralateral cancer was diagnosed at the same or earlier stage than the first cancer in 87% of cases. Bilateral patients were significantly more likely to have multicentric disease and to have a positive family history for breast carcinoma compared with the unilateral group. There were no significant differences regarding history of exogenous hormone exposure, lobular histology, hormone-receptor status, or HER-2/neu expression. Five-year disease-free survival was 94% for the unilateral breast carcinoma patients and 91% for the bilateral breast carcinoma patients (P = 0.16). CONCLUSIONS: Survival for patients with bilateral breast carcinoma is similar to that of patients with unilateral disease; however, prophylactic risk-reduction intervention for the contralateral breast should be considered in patients who have multicentric unilateral disease or a positive family history for breast carcinoma.
BIOPSY, PRIOR
Effect of time interval on residual disease in breast cancer. Wiley EL, Diaz LK, Badve S, Morrow M.	Am J Surg Pathol 2003 Feb;27(2):194-8 Abstract quote The histologic responses of breast tissue to injury are limited. Needle core biopsies of the breast are associated with displacement of tumor cells, and the incidence of tumor displacement decreases as the time interval between needle core biopsy and subsequent excision increases. This suggests that displaced tumor cells are destroyed by reparative processes induced by tissue injury. Residual tumor in a lumpectomy site may also be subjected to the same destructive processes associated with tissue repair. A total of 259 consecutive cases of infiltrating ductal carcinoma with margin-positive lumpectomies and their associated reexcision specimens obtained over a 7-year period were analyzed for the presence, type, and quantity of residual disease. The overall incidence of residual disease was 69%. Residual infiltrating ductal carcinoma was present in 35% of cases, and residual ductal carcinoma in situ was present in 50%. An increased time interval between lumpectomy and reexcision was associated with a decreased incidence of residual infiltrating carcinoma (p <0.0043); this decrease was not found associated with ductal carcinoma in situ. These findings suggest that the host response to injury may destroy residual infiltrating carcinoma cells in some margin-positive cases.
BRAIN METASTASES
Breast Cancers With Brain Metastases are More Likely to be Estrogen Receptor Negative, Express the Basal Cytokeratin CK5/6, and Overexpress HER2 or EGFR. Hicks DG, Short SM, Prescott NL, Tarr SM, Coleman KA, Yoder BJ, Crowe JP, Choueiri TK, Dawson AE, Budd GT, Tubbs RR, Casey G, Weil RJ. *Clinical and Anatomic Pathology daggerBreast Center double daggerTaussig Cancer Center section signLerner Research Institute paragraph signBrain Tumor Institute, Cleveland Clinic Foundation, Cleveland, OH 44195.	Am J Surg Pathol. 2006 Sep;30(9):1097-1104. Abstract quote Brain metastases (BM) from breast cancer are associated with significant morbidity and mortality. In the current study, we have examined a cohort of breast cancer patients who went on to develop BM for clinical-pathologic features and predictive markers that identify this high-risk subgroup of patients at the time of diagnosis. The primary tumors from 55 patients who developed BM were used to construct a tissue microarray. The clinical and pathologic features were recorded and the tissue microarray was stained for estrogen receptor, human epidermal growth factor receptor 2, cytokeratin 5/6, and epidermal growth factor receptor by immunohistochemistry . This cohort of patients was compared against a group of 254 patients who remain free of metastases (67 mo mean follow-up), and another cohort of 40 patients who developed mixed visceral and bone metastatic disease without brain recurrence over a similar period of time. Breast cancer patients who went on to develop BM were more likely to be <50 years old (P<0.001), and the primary tumors were more likely to be estrogen receptor negative (P<0.001) and high grade (P=0.002). The primary tumors were also more likely to express cytokeratin 5/6 (P<0.001) and epidermal growth factor receptor (P=0.001), and to overexpress human epidermal growth factor receptor 2 (P=0.001). The data presented above suggest a profile for breast cancer patients at increased risk for developing BM. Predictive factors to help identify patients with metastatic breast cancer who are at an increased risk for developing central nervous system recurrence might allow for screening of this population for early detection and treatment or for the development of targeted strategies for prevention.
CD105
CD105 Expression Is a Marker of High Metastatic Risk and Poor Outcome in Breast Carcinomas Correlations Between Immunohistochemical Analysis and Long-Term Follow-up in a Series of 929 Patients Jean-Philippe Dales, MD Stephane Garcia, MD Pascal Bonnier, MD Florence Duffaud, MD Lucile Andrac-Meyer, MD Olivier Ramuz, MD Marie-Noëlle Lavaut, MD Claude Allasia, PhD Colette Charpin, MD	Am J Clin Pathol 2003;119:374-380 Abstract quote CD105 (endoglin) is expressed significantly in activated endothelial cells in culture and in tumor microvessels. Quantification of CD105 immunocytochemical expression that may be clinically relevant has not been accurately evaluated. We studied CD105 expression on frozen tissue sections by using immunohistochemical assays in a series of 929 patients and correlated the findings with long-term follow-up (median, 11.3 years). Univariate (Kaplan-Meier) analysis showed that the number of CD105+ microvessels (cutoff, 15 vessels) correlated significantly with poor overall survival among all patients (P = .001). This correlation was less significant in node-negative patients (P = .035). Marked CD105 expression also correlated with a high risk for metastasis among all patients (P = .006) and among node-negative patients (P = .001). Multivariate analysis (Cox model) identified CD105 immunodetection as an independent prognostic indicator. Our results suggest that immunohistochemical expression of CD105 has practical clinical relevance for identifying node-negative patients with a poor prognosis. Moreover, immunodetection of CD105 also may be considered a potential tool for selecting patients who could benefit from specific antiangiogenic therapy, using anti-CD105 conjugates.
CHROMOSOMAL ABNORMALITIES
Simultaneous chromosome 1q gain and 16q loss is associated with steroid receptor presence and low proliferation in breast carcinoma. Farabegoli F, Hermsen MA, Ceccarelli C, Santini D, Weiss MM, Meijer GA, van Diest PJ. Department of Experimental Pathology, University of Bologna, Bologna, Italy.	Mod Pathol. 2004 Apr;17(4):449-55. Abstract quote We applied comparative genomic hybridization (CGH) to 46 breast carcinoma samples, collected from 1993 to 1995, in order to detect chromosome 1q gains and 16q losses and to define whether samples showing both these alterations had distinct biopathologic features and different clinical outcome. A total of 22 samples (48%) had simultaneous chromosome 1q gain and 16q loss, which was always associated with other genetic changes. In total, 23 samples had various chromosome imbalances (including chromosome 1q gain independent of chromosome 16q loss and vice versa) and one sample did not show detectable alterations. Samples having chromosome 1q gain/16q loss were compared to the other samples with regard to neoplasm size, lymph-node status, histologic and nuclear grade, estrogen and progesterone receptor presence, Ki-67, pRB, Cyclin D1, Cyclin A, p53, p21 and p27 expression as detected by immunohistochemistry. The samples showing chromosome 1q gain/16q loss had high steroid hormone receptor expression (P=0.02), low cell growth fraction (Ki-67, P=0.03) and high p27 expression (P<0.001). No statistical correlation with disease-free survival and overall survival or response to hormonal therapy was found. We conclude that simultaneous chromosome 1q gain/16q loss is a frequent event in invasive breast cancer, which occurs in a subset of both intermediate- and high-grade breast carcinomas. Although the final chromosome 1q and 16q imbalances might have originated from different chromosome alterations in low- and high-grade samples, the gene-dosage effect might be important in conferring peculiar biopathologic characteristics to this subset of samples. The cytogenetic and molecular mechanisms underlying these chromosome changes deserve further investigations.
Aneuploidy of chromosome 20 in invasive breast cancer correlates with poor outcome. Nakopoulou L, Tsirmpa I, Giannopoulou I, Trafalis D, Katsarou S, Davaris P. Department of Pathology, Medical School, The National and Kapodistrian University of Athens, Greece.	Cancer Genet Cytogenet 2002 Apr 15;134(2):127-32 Abstract quote Breast carcinoma is a genetically and phenotypically heterogeneous disease and is frequently associated with nonrandom chromosomal alterations. The aim of this study was to investigate the numerical aberrations of chromosome 20 in breast cancer. The observed chromosome-specific numerical abnormalities were evaluated along with the established clinicopathological parameters, the immunohistochemical expression of ER, PR, p53, c-erbB-2, Ki-67 and patients' survival. Nonisotopic in situ hybridization was applied to interphase cell nuclei on paraffin embedded tissue sections. Polysomy of chromosome 20 was the prevalent alteration in 45 of 50 (90%), monosomy in 2 of 50 (4%) and disomy in 3 of 50 (6%) cases. Invasive ductal carcinomas displayed a higher percentage of polysomy than lobular ones. A statistical significant association was demonstrated between Ki-67 immunohistochemical expression and polysomy of chromosome 20. Disomy was inversely correlated with Ki-67, while monosomy was suggestively associated with PR positive expression. Among the patients, those with the highest levels of polysomy showed the worst survival. In conclusion, the gain of chromosome 20 is the prevalent aberration in patients with breast carcinomas and may be useful prognostic marker in breast cancer.
CYCLIN D1
Bio-pathologic Characteristics Related to Chromosome 11 Aneusomy and Cyclin D1 Gene Status in Surgically Resected Stage I and II Breast Cancer: Identification of an Adverse Prognostic Profile. Mottolese M, Orlandi G, Sperduti I, Merola R, Buglioni S, Di Benedetto A, Pinnaro P, Perracchio L, Venturo I, Cognetti F, Cianciulli A. Departments of *Pathology daggerClinical Pathology (Cytogenetic Unit) double daggerBiostatistic Unit section signRadiotherapy paragraph signOncology B parallelOncology A, Regina Elena Cancer Institute, Rome, Italy.	Am J Surg Pathol. 2007 Feb;31(2):247-254. Abstract quote We aimed at developing a more detailed understanding of cyclin D1 in early stage human breast cancer and defining the biologic profiles with different prognostic value correlating cyclin D1 gene amplification and chromosome 11 aneusomy with bio-pathologic variables of known clinical importance. Cyclin D1 gene amplification and chromosome 11 aneusomy were investigated using fluorescence in situ hybridization whereas cyclin D1, PgR, HER-2, Bcl2, p53, and Ki-67 expressions were analyzed by immunohistochemistry in 121 stage I or II breast cancer patients uniformly treated with cyclophosphamide/metotrexate/5-fluorouracil-based chemotherapy. Cyclin D1 was amplified in 6.6% and overexpressed in 32.2% of cases. Amplification was not associated with any selected bio-pathologic variables, whereas the chromosome 11 aneusomy level significantly increased in tumors with higher histologic grade (P<0.01), higher tumor size (P<0.003), p53 nuclear accumulation (P<0.04), and ERalpha negativity (P<0.049). Multiple correspondence analysis showed 4 different biologic tumor profiles. The first, characterized by high Ki-67 score, p53+, cyclin D1+, HER-2+, aneusomy level>30%, ratio (cyclin D1 gene/CEP11)>2, was associated with tumor relapse defining the most unfavorable biologic profile. Kaplan-Meier's method showed significantly shorter disease-free survival in patients with at least 3 variables positive out of the 6 detected by multiple correspondence analysis. In multivariate analysis, the identified biologic profile emerged as the only significant prognostic indicator. Our findings are of particular clinical interest for early stage breast cancer patients, because the assessment of biologic factors predictive of tumor aggressiveness may influence postoperative therapeutic strategies.
CYCLOOXYGENASE-2
Overexpression of cyclooxygenase-2 is associated with breast carcinoma and its poor prognostic factors. Shim JY, An HJ, Lee YH, Kim SK, Lee KP, Lee KS. Department of Pathology, Bundang CHA Hospital, Pochon CHA University, Sungnam, Korea.	Mod Pathol. 2003 Dec;16(12):1199-204. Abstract quote   Cyclooxygenase is the rate-limiting enzyme that catalyzes the conversion of arachidonic acid to prostaglandins. The inducible form, cyclooxygenase-2, is known to be overexpressed in various human cancers including the colon, stomach, and urinary bladder. In this study, we evaluated the overexpression of cyclooxygenase-2 in 64 cases of breast cancer and correlated the results with clinicopathologic parameters. Immunohistochemical staining for cyclooxygenase-2 demonstrated positivity of the tumor cells in 46 of 64 cases (72%). Cyclooxygenase-2 overexpression was significantly correlated with larger tumor size and advanced clinical stage. Cyclooxygenase-2 overexpression tended to be more frequently observed in cases with presence of lymph node metastasis and in cases without expression of estrogen and progesterone; however, there was no significant correlation statistically. Nuclear and histologic grade were not well correlated with cyclooxygenase-2 overexpression. When ductal carcinoma in situ was considered separately, 32 of 42 cases (76%) were positive for cyclooxygenase-2. We conclude that cyclooxygenase-2 is up-regulated in a high proportion of breast cancers. The overexpression of cyclooxygenase-2 was associated with larger tumor size and advanced clinical stage, although lymph node status, estrogen and progesterone expression, and nuclear and histologic grade were not significantly correlated. Therefore, cyclooxygenase-2 overexpression may be a feature of the aggressive phenotype and may be useful as a prognostic indicator in breast cancer.
DNA PLOIDY
DNA ploidy, S-phase, and steroid receptors in more than 127,000 breast cancer patients. Wenger CR, Beardslee S, Owens MA, Pounds G, Oldaker T, Vendely P, Pandian MR, Harrington D, Clark GM, McGuire WL. University of Texas Health Science Center, Department of Medicine/Medical Oncology, San Antonio 78284-7884.	Breast Cancer Res Treat 1993 Oct;28(1):9-20 Abstract quote Several potential prognostic factors are available today for patients with breast cancer, and many more are being identified and studied. To evaluate the clinical utility of these factors, it will be necessary to measure them on a large number of patients, and then follow these patients so that multivariate survival analyses can be performed. The Oncology Research Network was established in 1986 by the University of Texas Health Science Center at San Antonio and Nichols Institute Reference Laboratories in order to evaluate the clinical utility of new prognostic factors for patients with primary breast cancer. The first generation of prognostic factors included steroid receptors, along with DNA ploidy and S-phase fraction determined by flow cytometry. Currently, laboratory results have been obtained from more than 127,000 patients, and follow-up information is available on a subset of more than 25,000 of these patients. S-phase fraction was related to the ploidy status of the tumor. An increased incidence of aneuploidy and higher S-phase fractions were found in estrogen and progesterone receptor negative tumors, tumors from patients with positive axillary lymph nodes, tumors greater than 2 cm in diameter, and patients younger than 35 years of age. Preliminary survival analyses suggest that S-phase fraction and DNA ploidy, in combination with other prognostic factors, are powerful predictors of early disease relapse. The Oncology Research Network provides an important resource for examining the clinical significance of new laboratory assays and for expediting improvements in existing laboratory techniques.
DCIS ASSOCIATED (EXTENSIVE INTRADUCTAL COMPONENT)
An assessment of extensive intraductal component as a risk factor for local recurrence after breast-conserving therapy. Jacquemier J, Kurtz JM, Amalric R, Brandone H, Ayme Y, Spitalier JM. Department of Anatomic Pathology, Marseille Cancer Institute, France.	Br J Cancer 1990 Jun;61(6):873-6 Abstract quote The influence of extensive intraductal component (EIC) on local recurrence risk was studied for 496 patients with stage I-II infiltrating ductal cancers treated by conservative surgery and irradiation. EIC was diagnosed in 65 of 231 (28%) premenopausal and 41 of 265 (15.5%) post-menopausal patients. Local recurrence risk was markedly increased in EIC+ patients (5-year actuarial risk 18% versus 8% without EIC, P less than 0.001), but this effect appeared limited to premenopausal patients. Local recurrence risk increased with increasing degree of EIC. EIC with more than 50% intraductal carcinoma was more prevalent in patients younger than 40, perhaps accounting to some degree for the higher local recurrence rates observed in younger patients. The presence of EIC had no influence on overall survival, on median time to local recurrence, or on short-term survival after local failure. The usefulness of EIC as a risk factor for local recurrence is discussed.
Intraductal carcinoma associated with invasive carcinoma of the breast. A comparison of the two lesions with implications for intraductal carcinoma classification systems. Goldstein NS, Murphy T. Department of Anatomic Pathology, William Beaumont Hospital, Royal Oak, Michigan 48073, USA.	Am J Clin Pathol 1996 Sep;106(3):312-8 Abstract quote Intraductal carcinoma (DCIS) is a useful marker for predicting which women will develop a recurrent breast malignancy. The authors examined 150 consecutive, mammographically detected, T1 invasive carcinomas associated with DCIS to study the DCIS and compare it to its associated invasive carcinoma. Intraductal carcinoma nuclear grades were assigned to each duct on a scale of 1 to 3. The percentage of DCIS ducts that were involved by each grade was quantitated into quartiles for cases with more than one DCIS nuclear grade. The predominant architectural pattern corresponding to each DCIS nuclear grade was recorded. Ninety-two percent of the 150 invasive carcinomas were of ductal type, 4% were tubular, and the remainder were various other subtypes. Nine percent of the DCIS cases were nuclear grade 1. The remaining 91% of cases were almost evenly distributed between mixed DCIS nuclear grades 1 and 2 (19%), pure DCIS nuclear grade 2 (24%), mixed DCIS nuclear grade 2 to 3 (25%), and pure DCIS nuclear grade 3 (22%). Two percent of cases were a mixture of DCIS nuclear grades 1 and 3 or 1, 2, and 3. All pure DCIS nuclear grade 1 or mixed 1 and 2 were associated with well or moderately differentiated invasive carcinomas, whereas the majority (61%) of the pure DCIS nuclear grade 3 cases were associated with poorly differentiated invasive carcinomas. There was no relation between the DCIS architectural pattern and the invasive carcinoma grade. In general, the DCIS nuclear grade correlates with the grade of the invasive carcinoma. Unlike DCIS architecture, nuclear grade heterogeneity within DCIS associated with invasive carcinoma is minimal. DCIS classification systems based on nuclear grade have merit because there is little variation in nuclear grade within a given patient's lesion.
Extensive and predominant in situ component in breast carcinoma: their influence on treatment results after breast-conserving therapy. Sinn HP, Anton HW, Magener A, von Fournier D, Bastert G, Otto HF. Department of Pathology, University of Heidelberg, Germany.	Eur J Cancer 1998 Apr;34(5):646-53 Abstract quote Intramammary tumour recurrence is one of the most important problems in breast-conserving therapy. We reviewed a series of 957 patients treated with breast-conserving therapy for primary invasive breast carcinomas between 1 January 1985 and 31 December 1992 at the University of Heidelberg. All histological slides were re-evaluated for risk factors with special emphasis on the extent and subclassification of the in situ tumour and the margin status. Six parameters were identified as significant risk factors for intramammary recurrence in the univariate analysis, including extensive or predominant in situ component (EIC, with at least twice the greatest dimension of the invasive tumour component), histological grade, angioinvasion, lobular tumour type, involved resection margin and lymph node status. The presence of an EIC was statistically correlated with low tumour grade, tumour at the resection margins and in re-excision specimens and with multifocal tumour invasion. Multivariate logistic regression analysis revealed that EIC (relative risk (RR) = 1.9), tumour grade (RR = 1.76), angioinvasion (RR = 1.34), lobular tumour type (RR = 1.65) and young age (< or = 40 years, RR = 1.39) were independent predictors of local recurrence. When combining these factors in a linear model, the simultaneous presence of at least two of the five risk factors predicted a 5-year risk of intramammary recurrence of 20.9% compared with a risk of only 1-5% when none or one of these risk factors were identifiable. We conclude that the risk of subsequent intramammary recurrence after breast-conserving therapy can be estimated from a scoring system that includes four histological risk factors and the patient's age.
EPIDERMAL GROWTH FACTOR RECEPTOR
Clinical Value of Epidermal Growth Factor Receptor Expression in Primary Breast Cancer. Rampaul RS, Pinder SE, Nicholson RI, Gullick WJ, Robertson JF, Ellis IO. From the *Nottingham Breast Institute, University of Nottingham, Nottingham City Hospital, NHS Trust, Nottingham, U.K.; daggerHistopathology Department, Addenbrooke's NHS Trust, Cambridge, UK; double daggerCardiff University, Cardiff, UK; and section signSchool of Biological Sciences, University of Kent, Kent, UK.	Adv Anat Pathol. 2005 Sep;12(5):271-273. Abstract quote   EGFR expression in primary breast cancer has been extensively investigated for its prognostic and predictive value. However overall there is no consensus on its potential to guide such prognostication. This is largely because of the great heterogeneity in study designs and methods used to assay the EGFR protein. The impetus to standardize such studies is much needed as there are now several tyrosine kinase inhibitors directed against the EGF receptor and phase II trials are showing significant promise.
EGFR gene amplification in breast cancer: correlation with epidermal growth factor receptor mRNA and protein expression and HER-2 status and absence of EGFR-activating mutations. Bhargava R, Gerald WL, Li AR, Pan Q, Lal P, Ladanyi M, Chen B. 1Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, NY, USA.	Mod Pathol. 2005 Aug;18(8):1027-33. Abstract quote   The human epidermal growth factor receptor (HER) family of receptor tyrosine kinase has been extensively studied in breast cancer; however, systematic studies of EGFR gene amplification and protein overexpression in breast carcinoma are lacking. We studied EGFR gene amplification by chromogenic in situ hybridization (CISH) and protein expression by immunohistochemistry in 175 breast carcinomas, using tissue microarrays. Tumors with >5 EGFR gene copies per nucleus were interpreted as positive for gene amplification. Protein overexpression was scored according to standardized criteria originally developed for HER-2. EGFR mRNA levels, as measured by Affymetrix U133 Gene Chip microarray hybridization, were available in 63 of these tumors. HER-2 gene amplification by fluorescence in situ hybridization (FISH) and protein overexpression by immunohistochemistry were also studied. EGFR gene amplification (copy number range: 7-18; median: 12) was detected in 11/175 (6%) tumors, and protein overexpression was found in 13/175 (7%) tumors. Of the 11 tumors, 10 (91%) with gene amplification also showed EGFR protein overexpression (2+ or 3+ by immunohistochemistry). The EGFR mRNA level, based on Affymetrix U133 chip hybridization data, was increased relative to other breast cancer samples in three of the five tumors showing gene amplification. Exons 19 and 21 of EGFR, the sites of hotspot mutations in lung adenocarcinomas, were screened in the 11 EGFR-amplified tumors but no mutations were found. Three of these 11 tumors also showed HER-2 overexpression and gene amplification. Approximately 6% of breast carcinomas show EGFR amplification with EGFR protein overexpression and may be candidates for trials of EGFR-targeted antibodies or small inhibitory molecules.
FIBROTIC FOCUS
Fibrotic focus in invasive ductal carcinoma: an indicator of high tumor aggressiveness. Hasebe T, Tsuda H, Hirohashi S, Shimosato Y, Iwai M, Imoto S, Mukai K. Pathology Division, National Cancer Center Research Institute, East, Chiba, Japan.	Jpn J Cancer Res 1996 Apr;87(4):385-94 Abstract quote Histological examination of invasive ductal carcinoma of the breast often demonstrates the presence of an extensive central fibrotic focus (FF). The clinicopathological significance of the FF, or scar, in primary invasive ductal carcinoma is still ambiguous. One hundred and fifty-three cases of invasive ductal carcinoma (IDC) were classified into two groups, those with and those without FF. The differences in frequency of immunohistochemically detected overexpression of c-erbB-2 protein and nuclear accumulation of p53 protein, and the labeling index of proliferating cell nuclear antigen (PCNA), as well as histopathological parameters were compared between these two groups. IDCs smaller than 50 mm with FF showed a higher frequency of high-grade tumors, a higher frequency of lymph node metastasis, and a significantly higher frequency of c-erbB-2 protein overexpression than those without FF. In tumors of 20 mm or less, the incidence of nuclear accumulation of p53 protein was significantly higher in tumors with than those without FF. Tumors with FF showed a significantly higher PCNA labeling index than those without FF, regardless of tumor size. The present results indicate that the presence of FF is an important clinicopathological parameter associated with a higher degree of malignancy in IDCs, especially those smaller than 50 mm. Therefore, dividing IDCs into those with and those without FF appears to be meaningful clinicopathologically.
Fibrotic focus in invasive ductal carcinoma of the breast: a histopathological prognostic parameter for tumor recurrence and tumor death within three years after the initial operation. Hasebe T, Tsuda H, Tsubono Y, Imoto S, Mukai K. Pathology Division, National Cancer Center Research Institute, East, Tokyo.	Jpn J Cancer Res 1997 Jun;88(6):590-9 Abstract quote We investigated whether the presence of a fibrotic focus (FF) in the primary lesion and in lymph node metastasis is a good predictor of early tumor recurrence or death in patients with invasive ductal carcinoma (IDC). Multivariate relative risk (RR) of tumor recurrence and death according to the presence of FF in the primary tumor was estimated using the Cox proportional hazards regression model with adjustment for other prognostic factors (histologic grade, T classification, nodal status, tumor necrosis, DNA ploidy, c-erbB-2 protein expression, p53 protein expression, and labeling index of proliferating cell nuclear antigen). For the evaluation of the metastatic status in the axillary lymph nodes, RR of multivariate analysis was adjusted for the presence of FF in the metastatic tumor and the number of lymph nodes involved (1-3 and > 3). The presence of FF increased the RR of tumor recurrence significantly for the cases in all stages, and especially for those in stages I and II (RR = 6.9, P < 0.05 and RR = 25.0, P < 0.005, respectively). All cases that died of disease had FF. Among IDCs with FF, 24 cases had FF in lymph node metastasis. Significantly higher RRs of tumor recurrence and death were observed in cases with FF in lymph node metastasis than in those without it (RR = 2.0, P < 0.001 and RR = 5.9, P < 0.05, respectively). It was suggested that the presence of FF is an important predictor of early tumor recurrence or death in patients with IDCs. The presence of FF in lymph node metastatic lesions is also a significant prognostic parameter.
New prognostic histological parameter of invasive ductal carcinoma of the breast: clinicopathological significance of fibrotic focus. Hasebe T, Mukai K, Tsuda H, Ochiai A. Pathology Division, National Cancer Center Research Institute East, Chiba, Japan.	Pathol Int 2000 Apr;50(4):263-72 Abstract quote Immunohistochemistry, DNA ploidy analysis and molecular genetics have made it possible to predict the outcome of breast cancer more precisely than routine histological examination alone. However, in routine practice, it is difficult to incorporate these methodologies in all cases. If certain histological parameters can accurately predict the outcome of patients with breast cancer, they would be more practical for routine use. We showed that the presence of fibrotic focus (FF) in invasive ductal carcinoma (IDC) is closely associated with c-erbB-2 or p53 protein expression, high proliferative activity, and high angiogenesis of the tumors. Furthermore, multivariate analyses with well-known prognostic parameters for IDC demonstrated that the presence of FF is the most useful independent parameter to predict IDC patient outcome. In addition, our data suggested that the interaction between tumor cells and stromal fibroblasts may play an important role in the formation of FF in IDC based on growth factor and growth factor receptor protein expression in the tumor cells and fibroblasts forming FF. Based on the results of our clinicopathological studies, we propose a new prognostic classification scheme for the prediction of IDC patient outcome, which consists of FF, nuclear atypia, and fat invasion. This classification has superior predicting power to existing prognostic classifications.
Prognostic significance of fibrotic focus in invasive ductal carcinoma of the breast: a prospective observational study. Hasebe T, Sasaki S, Imoto S, Mukai K, Yokose T, Ochiai A. Pathology Division (TH, TY, AO).	Mod Pathol 2002 May;15(5):502-16 Abstract quote We have already reported invasive ductal carcinomas (IDCs) with fibrotic focus (FF) to be associated with significantly poorer survival than IDCs without FF. The purpose of this study was to prospectively investigate the effect of the presence of FF on the outcomes of 439 patients with IDCs to confirm the prognostic significance of FF, by the multivariate analysis, employing the Cox proportional hazard regression model, as compared with well-known clinicopathological parameters. We also precisely evaluated the prognostic significance of FF from the viewpoint of FF characteristics. The present study demonstrated that the presence of FF is a very useful parameter predicting tumor recurrence (TR), as well as initial distant organ metastasis (IDOM), in lymph node-negative IDCs (P =.024 and P =.026) and in IDCs positive for either or both estrogen receptor (ER) or progesterone receptor (PR) (P =.007 and P =.015), respectively. In addition, FF of >8 mm in diameter was found to be an independent prognostic parameter for TR and IDOM in lymph node-negative patients and patients with IDC positive for either or both ER or PR (P =.005 and P =.018). We conclude that the presence of FF is a very important histologic prognostic parameter for patients with IDCs of the breast.
GLUTATHIONE S-TRANSFERASE
Prognostic significance of glutathione S-transferase-pi in invasive breast cancer. Huang J, Tan PH, Thiyagarajan J, Bay BH. Department of Medical Oncology, National Cancer Centre, Singapore.	Mod Pathol. 2003 Jun;16(6):558-65. Abstract quote Glutathione S-transferase pi (GST-pi), a Phase II detoxification enzyme, has recently been implicated in protection against apoptosis. Expression of GST-pi and Bcl-2 protein, an established apoptosis marker, was analyzed by immunohistochemistry in 116 cases of infiltrative ductal breast carcinomas in Singapore women. The markers were correlated with apoptosis detected by the TUNEL method and clinico-pathological parameters. There were 67 (58%) GST-pi-positive breast tumors and 43 (37%) Bcl-2-positive tumors. In a large proportion of GST-pi-positive/Bcl-2-positive tumors, there was a distinct accumulation of the GST-pi enzyme within the nucleus of cancer cells when examined by double immunofluorescence labeling under confocal microscopy. GST-pi immunoreactivity was not significantly correlated with any of the traditional histologic factors known to influence prognosis, whereas Bcl-2 overexpression was associated with reduced size of primary tumor (P =.021) and positive estrogen receptor status (P =.001). Univariate analysis revealed that GST-pi-positive, Bcl-2-positive, and lower histological grade tumors had decreased levels of apoptosis (P =.024, P =.011, and P =.029, respectively). However, multivariate analysis showed that histological grade and Bcl-2, but not GST-pi, immunoreactivity were correlated with apoptotic status. The Kaplan-Meier disease-free survival curves showed a significant difference between GST-pi-positive and GST-pi-negative breast cancer cases (P =.002). Disease-free survival in patients with GST-pi-positive tumors was also worse than that in patients with GST-pi-negative tumors in the group who had adjuvant chemotherapy (P =.04). In patients who were lymph node positive, GST-pi immunopositivity was found to influence disease-free survival. Recurrence of tumors was also significantly affected by GST-pi immunoreactivity (relative risk of 8.1). The findings indicate that GST-pi-positive tumors are more aggressive and have a poorer prognosis than do corresponding GST-pi-negative breast cancers.
GRADING
Breast carcinoma malignancy grading by Bloom-Richardson system vs proliferation index: reproducibility of grade and advantages of proliferation index. Meyer JS, Alvarez C, Milikowski C, Olson N, Russo I, Russo J, Glass A, Zehnbauer BA, Lister K, Parwaresch R. 1St Luke's Hospital, Chesterfield, MO, USA.	Mod Pathol. 2005 Aug;18(8):1067-78. Abstract quote   Questions of reproducibility and efficacy of histologic malignancy grading relative to alternative proliferation index measurements for outcome prediction remain unanswered. Microsections of specimens from the Cooperative Breast Cancer Tissue Resource (CBCTR) were evaluated by seven pathologists for reproducibility of grade and classification. Nuclear figure classification was assessed using photographs. Grade was assigned by the Bloom-Richardson method, Nottingham modification. Proliferation index was evaluated prospectively by deoxyribose nucleic acid precursor uptake with thymidine (autoradiographic) or bromodeoxyuridine (immunohistochemical) labeling index using fresh tissue from 631 node-negative breast cancer patients accessioned at St Luke's Hospital. A modified Nottingham-Bloom-Richardson grade was derived from histopathologic data. Median post-treatment observation was 6.4 years. Agreement on classification of nuclear figures (N=43) was less than good by kappa statistic (kappa=0.38). Grade was moderately reproducible in four trials (N=10,10,19, 10) with CBCTR specimens (kappa=0.50-0.59). Of components of Bloom-Richardson grade, agreement was least for nuclear pleomorphism (kappa=0.37-0.50), best for tubularity (kappa=0.57-0.83), and intermediate for mitotic count (kappa=0.45-0.64). Bloom-Richardson grade was a univariate predictor of prognosis in node-negative St Luke's patients, and was improved when mitotic count was replaced by labeling index (low, mid, or high). When labeling index was added to a multivariate model containing tumor size and vessel invasion, grade was no longer a significant predictor of tumor-specific relapse-free or overall survival. Mitotic index predicted best when intervals were lowered to 0-2, 3-10, and >10 mitotic figures per ten 0.18 mm(2) high-power fields. We conclude that Nottingham-Bloom-Richardson grades remain only modestly reproducible. Prognostically useful components of grade are mitotic index and tubularity. The Nottingham-Bloom-Richardson system can be improved by lowering cutoffs for mitotic index and by counting 20-30 rather than 10 high-power fields. Measurement of proliferation index by immunohistochemically detectable markers will probably give superior prognostic results in comparison to grade.
HORMONE RECEPTOR STATUS
Progesterone receptor by immunohistochemistry and clinical outcome in breast cancer: a validation study. Mohsin SK, Weiss H, Havighurst T, Clark GM, Berardo M, Roanh le D, To TV, Zho Q, Love RR, Allred DC. [1] 1The Breast Center, Baylor College of Medicine, Houston, TX, USA [2] 2Department of Pathology, The Methodist Hospital, Houston, TX, USA.	Mod Pathol. 2004 Dec;17(12):1545-54. Abstract quote   Progesterone receptor is a surrogate marker of estrogen receptor activity in breast cancer and its utility in helping predict clinical outcome has been established using biochemical assays. However, most laboratories worldwide have switched to immunohistochemistry to assess progesterone receptor, but unfortunately no validated immunohistochemical assay exists for progesterone receptor. The purpose of this study was to develop and validate an immunohistochemical assay for progesterone receptor in breast cancer. The assay was based on monoclonal antibody 1294 (DakoCytomation) and slides were scored microscopically using the 'Allred score' on a scale of 0-8. The assay was compared to ligand-binding assay in 1235 breast cancers, and a subset (n=362) that received only hormonal therapy was used to define a cutoff for progesterone receptor-positive. Clinical utility was validated in an independent set of samples (n=423) from a clinical trial randomizing premenopausal breast cancer patients to tamoxifen+oophorectomy vs observation following surgery. A cutoff of >2 (corresponding to >1% positive cells) dichotomized patients with significantly better or worse clinical outcome (P=0.0014). Progesterone receptor by immunohistochemistry provided significantly better results than progesterone receptor by ligand-binding assay in predicting clinical outcome. In the clinical trial, a positive result in univariate analyses was associated with significantly improved disease-free and overall survival both in untreated (hazard ratios/P=0.656/0.060 and 0.479/0.005, respectively) and hormonally treated patients (hazard ratios/P=0.529/0.017 and 0.451/0.007, respectively). Positive progesterone receptor remained significant for improved disease-free and overall survival (hazard ratios/P=0.666/0.038 and 0.549/0.007, respectively) in multivariate analyses including the standard variables of tumor size, nodal status, treatment, histological grade, and HER-2/neu status. Estrogen and progesterone receptors are codependent variables and progesterone receptor was a weaker predictor of response to endocrine therapy than estrogen receptor when both were included in multivariate analysis. This is the first comprehensive study assessing the clinical usefulness of progesterone receptor by immunohistochemistry in archival tissue in breast cancer. Progesterone receptor assessed by immunohistochemistry provides useful information about clinical outcome and it is better than progesterone receptor measured by ligand-binding assay.
Androgen Receptor Expression in Estrogen Receptor–Negative Breast Cancer Immunohistochemical, Clinical, and Prognostic Associations S. Nicholas Agoff, MD, Paul E. Swanson, MD, Hannah Linden, MD, Stephen E. Hawes, PhD, and Thomas J. Lawton, MD	Am J Clin Pathol 2003;120:725-731 Abstract quote We sought to determine the prevalence of androgen receptor (AR) expression in a predominantly estrogen receptor (ER)-negative subset of breast cancers and delineate the immunohistochemical and clinical associations, including whether AR expression has prognostic significance in ER-negative tumors. We identified 69 ER-negative and 19 ER-positive breast cancer cases with concurrent immunohistochemical prognostic panels (ER, PR, HER-2/neu, Ki-67, and p53); immunohistochemical analysis was performed for AR using standard techniques. Clinical data were extracted from medical records. c 2 tests were used to assess associations between variables. AR was found in 49% (34/69) of ER-negative and 89% (17/19) of ER-positive cases. In ER-negative tumors, AR was associated with increased age ( P = .02), postmenopausal status ( P < .001), tumor grade ( P = .03), tumor size ( P = .03), and HER-2/neu overexpression ( P = .003). In ER-positive tumors, AR was associated with progesterone receptor expression ( P < .03). In univariate analysis of ER-negative tumors, patients with AR-positive tumors had significantly better disease-free survival ( P = .049). AR is expressed in a significant number of ER-negative cases and shows significant associations with important clinical and pathologic prognostic factors.
Morphologic spectrum of estrogen receptor-negative breast carcinoma. Scawn R, Shousha S. Department of Histopathology, Charing Cross Hospital and Imperial College of Science, Technology and Medicine, London, United Kingdom.	Arch Pathol Lab Med 2002 Mar;126(3):325-30 Abstract quote Context.-Estrogen receptor (ER)-negative breast carcinomas are a heterogeneous group of breast cancers that are generally thought to be aggressive. Objective.-To determine the morphologic and immunohistochemical spectrum of a consecutive series of ER-negative breast carcinomas, in an attempt to understand the pathogenesis and behavior of these lesions. Design.-Seventy-four consecutive cases of ER-negative invasive carcinomas were studied. Hematoxylin-eosin-stained sections were reviewed, and new sections were stained for c-erbB-2, p53, vimentin, and androgen and prolactin receptors. The findings were correlated with the axillary lymph node status as a measure of tumor aggressiveness. Setting.-The histopathology department of a tertiary referral teaching hospital. Results.-The tumors included 50 (68%) invasive ductal carcinomas, 21 (28%) medullary/atypical medullary carcinomas, and 1 each of invasive lobular, apocrine, and papillary carcinoma. Some of the invasive ductal cases had distinctive features that are described in this report. Maximum tumor diameter varied between 5 and 100 mm. Sixty tumors (81%) were grade 3, 13 (18%) were grade 2, and 1 (1%) was grade 1. Of the 60 cases in which the axillary node status was known, 34 (57%) had metastases, and 26 did not. Tumors associated with positive nodes were significantly larger than those associated with negative nodes (37.2 vs 17.8 mm, P <.001). A higher percentage of node-negative tumors were c-erbB-2 positive (42% vs 21%, P <.05). There were no differences between the 2 groups with regard to histologic type, tumor grade, or the expression of p53, vimentin, or androgen or prolactin receptors Conclusions.-Many ER-negative breast carcinomas have distinctive microscopic features. Not all ER-negative tumors are aggressive, as judged by the absence of lymph node metastases in 43% of cases in this series. Tumor size is the most important indicator for the likelihood of the presence of lymph node metastases. The wide range of tumor sizes encountered in this series suggests that the ER status of a tumor is determined early in its natural history and supports the existence of 2 separate pathways for the development of ER-negative and ER-positive breast carcinomas.
HYPER-CHOLESTEROLEMIA
Hypercholesterolemia impairs angiogenesis in patients with breast carcinoma and, therefore, lowers the risk of metastases. Ozdemir BH, Akcali Z, Haberal M. Department of Pathology, Baskent University Faculty of Medicine, Ankara, Turkey.	Am J Clin Pathol. 2004 Nov;122(5):696-703. Abstract quote   Our aim was to study the effect of hypercholesterolemia on angiogenesis induced by breast carcinoma. Of 51 patients with invasive ductal carcinoma, 28 had hypercholesterolemia and 23 had normocholesterolemia. The intratumoral microvessel density (MVD) was evaluated by using anti-CD31 antibody. The expression of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) on endothelial and tumor cells was examined and graded semiquantitatively. Patients with normocholesterolemia had a higher MVD (76.4 +/- 8.2) than those with hypercholesterolemia (54.6 +/- 5.1) (P < .01). The risks of recurrence and distant metastasis were higher in patients with normocholesterolemia than in patients with hypercholesterolemia (P < .01). Patients with hypercholesterolemia showed lower expression of endothelial VEGF and bFGF than patients with normocholesterolemia (P < .05 and P < .01, respectively). In addition, tumoral bFGF and VEGF expression showed negative correlation with the presence of hypercholesterolemia (P < .01). We suggest that hypercholesterolemia impairs angiogenesis by suppressing endothelial and tumoral bFGF and VEGF expression and, therefore, lowers the risk of metastases in cases of invasive breast carcinoma.
IPSILATERAL BREAST FAILURE
Molecular Clonality Relationships in Initial Carcinomas, Ipsilateral Breast Failures, and Distant Metastases in Patients Treated With Breast-Conserving Therapy Evidence Suggesting That Some Distant Metastases Are Derived From Ipsilateral Breast Failures and That Metastases Can Metastasize Neal S. Goldstein, MD, etal.	Am J Clin Pathol 2005;124:49-57 Abstract quote We studied the clonality relationships in invasive breast carcinomas, ipsilateral breast failures (IBFs), and distant metastases (DMs) using a polymerase chain reaction–loss of heterozygosity (LOH) clonality assay to determine whether IBFs can be the source of DMs. Six cases of initial carcinomas, IBFs, and DMs were identified. Carcinoma DNA was extracted from paraffin blocks and analyzed with 20 markers. In 2 cases, the LOH pattern suggested the DM directly resulted from the IBF. In 2 cases, the initial carcinoma, IBF, and DM were one progressive, genetically unstable process. Separate subclones in the initial carcinoma gave rise to the IBF and DM in 1 case, and the DM derived from a second IBF in 1 case. The relationships of initial carcinomas, IBFs, and DMs are complex. DMs seem to be the direct result of IBFs in some cases. Some carcinomas seem to be composed of subclones with different and unrelated IBF and DM potential.
Molecular Clonality Determination of Ipsilateral Recurrence of Invasive Breast Carcinomas After Breast-Conserving Therapy Comparison With Clinical and Biologic Factors Neal S. Goldstein, MD, etal.	Am J Clin Pathol 2005;123:679-689 Abstract quote We established clonality relationships between invasive ipsilateral breast failures (IBFs; local recurrences) and initial invasive carcinomas using a molecular polymerase chain reaction loss of heterozygosity (LOH) assay for 26 patients treated with breast-conserving therapy for invasive carcinoma with no distant metastases (DMs) before the IBF. LOH was ± 50% allelic loss. Eighteen IBFs (69%) were related clonally to initial carcinomas; 8 (31%) were clonally distinct, second primary carcinomas. IBFs and initial invasive carcinomas were morphologically similar in 6 (75%) of 8 clonally different cases. Clinical IBF classification and molecular assay results differed in 11 cases (42%). The mean intervals to IBF were 4.7 years in related and 8.7 years in different cases (P = .013). In 6 patients, DMs developed; 5 had related IBFs. In related IBF cases, the mean increase in fractional allelic loss (FAL) of IBFs associated with DMs was 18.9% compared with 7.6% in cases unassociated with DMs (P = .004). Molecular assays can accurately establish the clonality of most IBFs. Morphologic comparison and clinical IBF classification are unreliable methods of determining clonality. Clonally related IBFs occurred sooner than clonally different IBFs. Patients with clonally related IBFs are the main pool in which DMs occur. Not all clonally related IBFs have the same DM association; those with large FAL gains were associated with DMs. Molecular clonality assays may provide a reliable means of identifying patients who might benefit from systemic chemotherapy at the time of IBF.
Ki-67
Immunohistochemical detection of Ki67 in breast cancer correlates with transcriptional regulation of genes related to apoptosis and cell death. Tan PH, Bay BH, Yip G, Selvarajan S, Tan P, Wu J, Lee CH, Li KB. 1Department of Pathology, Singapore General Hospital, Singapore.	Mod Pathol. 2005 Mar;18(3):374-81. Abstract quote   Ki67 is a nuclear protein that is tightly linked to the cell cycle. It is a marker of cell proliferation and has been used to stratify good and poor prognostic categories in invasive breast cancer. Its correlation with gene expression patterns has not been fully elucidated. In this study, Ki67 immunohistochemistry using the MIB-1 antibody was performed on sections cut from 21 formalin-fixed, paraffin-embedded invasive breast cancers. Scoring was determined as nil (no immunostaining), low (10% or less immunopositivity) or high (>10% immunoreactive cells) respectively. The relationship of Ki67 immunohistochemical detection with clinicopathologic parameters was evaluated. Using Affymetrix U133A GeneChips, expression profiles for these tumors were generated and correlated with Ki67 immunohistochemical findings. Analysis of variance was used to define genes that were differentially regulated between the groups. Real-time polymerase chain reaction (PCR) was used to confirm the presence of a downregulated gene. Our results showed high, low and nil Ki67 immunostaining in nine (43%), six (28.5%) and six (28.5%) invasive breast cancers respectively, with increased Ki67 protein expression correlating with high histologic grade (P=0.02), mitotic score (P=0.001) and estrogen receptor immunonegativity (P=0.002). Expression profiling trends of the Ki67 gene mirrored the observed proportions of immunostained cells when the Ki67 immunoscore was >10%. Genes related to apoptosis and cell death (bcl2, MAP2K4, TNF10) were noted to be downregulated in tumors that disclosed >40% Ki67 immunostaining (P<0.001). Downregulation of the bcl2 gene was confirmed at the RNA level by real-time RT-PCR. Differential regulation of these genes, especially bcl2, may contribute to the biological nature of clinically more aggressive and highly proliferative breast cancers.
LOBULAR CARCINOMA IN SITU
The relation between the presence and extent of lobular carcinoma in situ and the risk of local recurrence for patients with infiltrating carcinoma of the breast treated with conservative surgery and radiation therapy Abner AL, Connolly JL, Recht A, Bornstein B, Nixon A, Hetelekidis S, Silver B, Harris JR, Schnitt SJ. Joint Center for Radiation Therapy, Harvard Medical School, Boston, MA 02215, USA	Cancer 2000 Mar 1;88(5):1072-7 Abstract quote BACKGROUND: When found in an otherwise benign biopsy, lobular carcinoma in situ (LCIS) has been associated with an increased risk of development of a subsequent invasive breast carcinoma. However, the association between LCIS and the risk of subsequent local recurrence in patients with infiltrating carcinoma treated with conservative surgery and radiation therapy has received relatively little attention. METHODS: Between 1968 and 1986, 1625 patients with clinical Stage I-II invasive breast carcinoma were treated at the Joint Center for Radiation Therapy at Harvard Medical School with breast-conserving surgery (CS) and radiation therapy (RT) to a total dose to the primary site of > or =60 grays. Analysis was limited to 1181 patients with infiltrating ductal carcinoma, infiltrating lobular carcinoma, or infiltrating carcinoma with mixed ductal and lobular features who, on review of their histologic slides, had sufficient normal tissue adjacent to the tumor to evaluate for the presence of LCIS and also had a minimum potential follow-up time of 8 years. The median follow-up time was 161 months. RESULTS: One hundred thirty-seven patients (12%) had LCIS either within the tumor or in the macroscopically normal adjacent tissue. The 8-year crude risk of recurrence was not significantly increased for patients with LCIS associated with invasive ductal, invasive lobular, or mixed ductal and lobular carcinoma. Among the 119 patients with associated LCIS adjacent to the tumor, the 8-year rate of local recurrence was 13%, compared with 12% for the 1062 patients without associated LCIS. For the 70 patients with moderate or marked LCIS adjacent to the tumor, the 8-year rate of local recurrence was 13%. The extent of LCIS did not affect the risk of recurrence. The risks of contralateral disease and of distant failure were similarly not affected by the presence or extent of LCIS. CONCLUSIONS: Breast-conserving therapy involving limited surgery and radiation therapy is an appropriate method of treating patients with invasive breast carcinoma with or without associated LCIS. Neither the presence nor the extent of LCIS should influence management decisions regarding patients with invasive breast carcinoma.
Lobular carcinoma in situ increases the risk of local recurrence in selected patients with stages I and II breast carcinoma treated with conservative surgery and radiation. Sasson AR, Fowble B, Hanlon AL, Torosian MH, Freedman G, Boraas M, Sigurdson ER, Hoffman JP, Eisenberg BL, Patchefsky A. Department of Surgical Oncology, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19104, USA	Cancer 2001 May 15;91(10):1862-9 Abstract quote BACKGROUND: Lobular carcinoma in situ (LCIS) is a known risk factor for the development of invasive breast carcinoma. However, little is known regarding the impact of LCIS in association with an invasive carcinoma on the risk of an ipsilateral breast tumor recurrence (IBTR) in patients who are treated with conservative surgery (CS) and radiation therapy (RT). The purpose of this study was to examine the influence of LCIS on the local recurrence rate in patients with early stage breast carcinoma after breast-conserving therapy. METHODS: Between 1979 and 1995, 1274 patients with Stage I or Stage II invasive breast carcinoma were treated with CS and RT. The median follow-up time was 6.3 years. RESULTS: LCIS was present in 65 of 1274 patients (5%) in the study population. LCIS was more likely to be associated with an invasive lobular carcinoma (30 of 59 patients; 51%) than with invasive ductal carcinoma (26 of 1125 patients; 2%). Ipsilateral breast tumor recurrence (IBTR) occurred in 57 of 1209 patients (5%) without LCIS compared with 10 of 65 patients (15%) with LCIS (P = 0.001). The 10-year cumulative incidence rate of IBTR was 6% in women without LCIS compared with 29% in women with LCIS (P = 0.0003). In both groups, the majority of recurrences were invasive. The 10-year cumulative incidence rate of IBTR in patients who received tamoxifen was 8% when LCIS was present compared with 6% when LCIS was absent (P = 0.46). Subsets of patients in which the presence of LCIS was associated with an increased risk of breast recurrence included tumor size < 2 cm (T1), age < 50 years, invasive ductal carcinoma, negative lymph node status, and the absence of any adjuvant systemic treatment (chemotherapy or hormonal therapy) (P < 0.001). LCIS margin status, invasive lobular carcinoma histology, T2 tumor size, and positive axillary lymph nodes were not associated with an increased risk of breast recurrence in these women. CONCLUSIONS: The authors conclude that the presence of LCIS significantly increases the risk of an ipsilateral breast tumor recurrence in certain subsets of patients who are treated with breast-conserving therapy. The risk of local recurrence appears to be modified by the use of tamoxifen. Further studies are needed to address this issue.
LYMPH NODE STATUS
Prognostic features in patients with stage T1 breast carcinoma and a 0.5-cm or less lymph node metastasis. Significance of lymph node hilar tissue invasion. Goldstein NS, Mani A, Vicini F, Ingold J. Department of Anatomic Pathology, William Beaumont Hospital, Royal Oak, Michigan 48073, USA.	Am J Clin Pathol 1999 Jan;111(1):21-8 Abstract quote Trends in surgical practice suggest that pathologists will encounter increased numbers of patients with small invasive ductal adenocarcinomas; small, if any, metastatic deposits in axillary lymph nodes (ALNs); and possibly fewer ALN specimens to examine. New prognostic histologic features may be needed in this environment. We studied histologic features of primary breast carcinoma and ALN metastasis from 86 patients who had stage T1 ductal carcinomas with only 1 ALN metastasis that was 0.5 cm or less and correlated these features with the development of distant metastases to evaluate their potential usefulness as prognostic indicators. The median follow-up period was 5.3 years. Distant metastases developed in 12 patients. Features significantly associated with 10-year distant metastases-free survival were lymph node hilar tissue invasion (HTI) and ALN metastasis size (stage N1a vs N1b). Tumor grade 1 vs grades 2 or 3 approached significance. The presence of HTI also was related significantly to a decreased 10-year distant metastases-free survival in the stage N1b group. Our study suggests that HTI, along with other well-known parameters, is a useful prognostic feature. In addition, it supports the opinion that ALN dissection may provide limited additional information for patients with grade 1, stage Tla, invasive ductal carcinomas. Additional studies are needed to confirm our findings.
The risk of nodal metastases in breast cancer patients with clinically negative lymph nodes: a population-based analysis. Voogd AC, Coebergh JW, Repelaer van Driel OJ, Roumen RM, van Beek MW, Vreugdenhil A, Crommelin MA. Comprehensive Cancer Center South, Eindhoven, The Netherlands.	Breast Cancer Res Treat 2000 Jul;62(1):63-9 Abstract quote A population-based study was performed to assess the likelihood of axillary lymph node metastases in patients with clinically negative lymph nodes, according to patient age, tumor size and site, estrogen receptor status, histologic type and mode of detection. Data were obtained from the population-based Eindhoven Cancer Registry. During the period 1984-1997, 7680 patients with invasive breast cancer were documented, 6663 of whom underwent axillary dissection. Of the 5125 patients who were known to have clinically negative lymph nodes and underwent axillary dissection, 1748 (34%) had positive lymph nodes at pathological examination. After multivariate analysis, histologic type, tumor size, tumor site and the number of lymph nodes in the axillary specimen remained as independent predictors of the risk of nodal involvement (P < 0.001). Lower risks were found for patients with medullary or tubular carcinoma, smaller tumors, a tumor in the medial part of the breast and patients with less than 16 nodes examined. This study gives reliable estimates of the risk of finding positive lymph nodes in patients with a clinically negative axilla. Such information is useful when considering the need for axillary dissection and to predict the risk of a false-negative result when performing sentinel lymph node biopsy.
LYMPHOVASCULAR INVASION
Improved Methods of Detection of Lymphovascular Invasion Demonstrate That It is the Predominant Method of Vascular Invasion in Breast Cancer and has Important Clinical Consequences. Mohammed RA, Martin SG, Gill MS, Green AR, Paish EC, Ellis IO. *Clinical Oncology †Histopathology Departments, University of Nottingham, University Hospitals, City Hospital Campus ‡Division of Pathology, School of Molecular Medical Sciences, University of Nottingham Queenʼs Medical Centre, Nottingham, UK.	Am J Surg Pathol. 2007 Dec;31(12):1825-1833. Abstract quote The presence of vascular invasion (VI), encompassing both lymphovascular invasion (LVI) and blood vascular invasion (BVI), in breast cancer has been found to be a poor prognostic factor. It is not clear, however, which type of VI plays the major role in metastasis. The aims of this study were to use an endothelial subtype specific immunohistochemical approach to distinguish between LVI and BVI by comparing the differential expression of blood vascular (CD34 and CD31) and lymphatic markers (podoplanin/D2-40) to determine their prognostic role in a well-characterized group of breast cancer patients with known long-term follow-up. Sections from177 consecutive paraffin-embedded archival specimens of primary invasive breast cancer were stained for expression of podoplanin, D2-40, CD31, and CD34. BVI and LVI were identified and results were correlated with clinicopathologic criteria and patient survival. VI was detected in 56/177 specimens (31.6%); 54 (96.4%) were LVI and 2 (3.5%) were BVI. The presence of LVI was significantly associated with the presence of lymph node metastasis, larger tumor size, development of distant metastasis, regional recurrence and worse disease-free interval and overall survival. In multivariate analysis, LVI retained significance association with decreased disease-free interval and overall survival. In conclusion, VI in breast cancer is predominantly of lymph vessels and is a powerful independent prognostic factor, which is associated with risk of recurrence and death from the disease. The use of immunohistochemical staining with a lymphendothelial specific marker such as podoplanin/D2-40 increases the accuracy of identification of patients with tumor associated LVI.
Significance of lymph vessel invasion identified by the endothelial lymphatic marker D2-40 in node negative breast cancer. Arnaout-Alkarain A, Kahn HJ, Narod SA, Sun PA, Marks AN. 1Department of Surgery, Sunnybrook Health Sciences Centre and Women's College Hospital, University of Toronto, Toronto, ON, Canada.	Mod Pathol. 2007 Jan 5; Abstract quot Monoclonal antibody D2-40, a marker of lymphatic endothelium, identifies tumor emboli in lymph vessels. The aim of the study was to assess whether D2-40+ lymph vessel invasion (LVI) correlates with clinicopathologic factors including lymphovascular invasion (LVI) as assessed by haematoxylin and eosin-stained sections (H&#38;E+ or H&#38;E-) and to assess the prognostic significance in node-negative breast cancer. The study group consisted of 303 node-negative breast cancer patients that had a median follow-up of 7.6 years. Clinical and pathological data were retrieved from the Henrietta Banting database. Immunohistochemical staining was performed on formalin-fixed, paraffin-embedded tissue sections of the primary invasive carcinoma using D2-40. Immunostaining with CD31 was performed on the discordant cases that were H&#38;E+/D2-40-. D2-40+ lymph vessel invasion was detected in 82/303 (27%) cases. The foci of lymphatic invasion occurred predominantly at the invasive front of the tumor. The absence of D2-40 and CD31 in 13/17 discordant cases was suggestive of retraction artefact. D2-40+ lymph vessel invasion correlated significantly with age (P=0.0003), tumor size (P=0.005), histological grade (P=0.0001), H&#38;E+ (P=<0.0001) and estrogen receptor status (P=0.005) but not with histological type or progesterone receptor status. Multivariate analysis revealed that D2-40+ lymph vessel invasion was the only significant predictor of distant recurrence. There was no significant association between D2-40 status and local recurrence (P=0.752) or regional recurrence (P=0.13). Both D2-40+lymph vessel invasion (P=0.009) and H&#38;E+LVI cases (P=0.02) were associated with overall shorter survival in univariate analysis. These data indicate that D2-40 identifies lymphatic invasion in breast tumors and is a significant predictor of outcome in breast cancer.
Extensive Retraction Artifact Correlates With Lymphatic Invasion and Nodal Metastasis and Predicts Poor Outcome in Early Stage Breast Carcinoma. Acs G, Dumoff KL, Solin LJ, Pasha T, Xu X, Zhang PJ. *Department of Pathology and Laboratory Medicine daggerDepartment of Radiation Oncology, University of Pennsylvania School of Medicine, Philadelphia, PA.	Am J Surg Pathol. 2007 Jan;31(1):129-140. Abstract quote Retraction artifact resulting in clear spaces around tumor cell nests is frequently seen in histologic material and may present difficulty in their differentiation from lymphovascular invasion. We noticed that retraction artifact seemed to be more common around groups of breast cancer cells compared with benign acini, and when extensively present, metastasis to axillary lymph nodes was often seen. Thus, we performed a study of 304 cases of stage pT1 and pT2 breast carcinomas to test our hypothesis that extensive retraction artifact in tumors correlates with lymphatic spread and outcome. Tumors were evaluated to determine the presence and extent of retraction artifact around tumor cell nests and the presence of lymphatic invasion. Lymphatic invasion was confirmed by D2-40 immunostaining. The extent of retraction artifact in tumors was correlated with clinicopathologic tumor features and patient outcome. Variable degree of retraction artifact was present in 183 of 304 (60%) invasive carcinomas, with its extent ranging from 0% to 90% (median 5%). The extent of retraction artifact showed a significant correlation with tumor size, histologic type, histologic grade, presence of lymphovascular invasion, and nodal metastasis. Further, extensive retraction artifact was significantly associated with poor overall and disease-free survival in both univariate and multivariate analyses. We propose that the apparent retraction of the stroma from cells of invasive breast carcinoma on routine histologic sections is not a phenomenon merely due to inadequate fixation as currently believed. Rather, it likely signifies important biologic changes that alter tumor-stromal interactions and contribute to lymphatic spread and tumor progression.
Pathologic Features of Breast Cancer Associated With Complete Response to Neoadjuvant Chemotherapy: Importance of Tumor Necrosis. Pu RT, Schott AF, Sturtz DE, Griffith KA, Kleer CG. From the Departments of *Pathology, and daggerInternal Medicine/Oncology, double daggerBiostatistics Unit, Cancer Center, University of Michigan, Ann Arbor, MI.	Am J Surg Pathol. 2005 Mar;29(3):354-358. Abstract quote   Breast cancer patients with a complete pathologic response after neoadjuvant chemotherapy have a better prognosis than incomplete responders. The predictive value of the histologic characteristics of the tumor prior to neoadjuvant treatment has not been well defined, and there are no guidelines for reporting tumor characteristics in the core biopsy report. Histologic and nuclear grades, presence of tumor necrosis and angiolymphatic invasion (ALI), and estrogen receptor (ER), progesterone receptor (PR), and HER-2/neu expression were assessed in core biopsies of 55 patients with invasive carcinomas. Patients were then uniformly treated with four cycles of doxorubicin/docetaxel followed by excisions and lymph node dissections. Complete pathologic response (pCR) was defined as having no invasive carcinoma at excision. Noncomplete pathologic response was defined as having invasive carcinoma at excision. Five of the 55 patients (9%) achieved pCR. Of the 5 complete responders, 4 (80%) had tumor necrosis in the core biopsy specimens, while only 8 of the 46 (17%) noncomplete responders (pNR) had this feature (P = 0.0086). Higher histologic and nuclear grades, ER, PR status, and HER-2/neu overexpression were not associated with pCR. The presence of ALI in the core biopsy, post-therapy excision, or both was associated with axillary lymph node metastases (P = 0.0062, P = 0.0249, and P = 0.0021, respectively). Although preliminary, our study suggests that the presence of tumor necrosis and ALI in the core biopsy may be important features to be included in the standard report.
Histological characteristics of tumor in vessels and lymph nodes are significant predictors of progression of invasive ductal carcinoma of the breast: A prospective study. Hasebe T, Sasaki S, Imoto S, Ochiai A.	Hum Pathol 2004;35:298-308 Abstract quote Invasive ductal carcinomas (IDCs) of the breast are composed of primary invasive tumors as well as tumor cells in blood vessels and lymph nodes. The purpose of this study was to determine whether the histological characteristics of tumor in the vessels and nodes are significantly associated with outcome. In a series of 393 patients, multivariate analyses showed that in IDCs without nodal metastasis and with fibrotic focus dimension, lymph vessel tumor emboli with >6 apoptotic figures and those invading >3 mm from the tumor margin had significantly higher hazard rates (HRs) for recurrence (P <0.05). In IDCs with 1 to 3 nodal metastases, >2 apoptotic figures in tumor emboli in blood vessels and >5 invaded lymph vessels were associated with significantly higher HRs for tumor recurrence and death (P <0.005). In IDCs with 4 or more nodal metastases, nodal tumors with >5 mitotic figures and >5 nodes with extranodal extension were associated with significantly higher HRs for tumor recurrence or death (P <0.05). We conclude that several histological characteristics of tumors in vessels and nodes have significant implications for the progression of IDCs.
Characteristics of tumors in lymph vessels play an important role in the tumor progression of invasive ductal carcinoma of the breast: a prospective study. Hasebe T, Sasaki S, Imoto S, Ochiai A. Pathology Division (TH, AO) and Epidemiology and Biostatistics Division (SS), National Cancer Center Research Institute East.	Mod Pathol 2002 Sep;15(9):904-13 Abstract quote It is unknown whether the characteristics of tumor cells in lymph vessels play an important role in the tumor progression of invasive ductal carcinoma (IDC) of the breast. The purpose of this study was to investigate the significance of the characteristics of tumor cells in the lymph vessels in relation to the tumor progression in 393 IDC patients in comparison with well-known histological parameters. The dimensions of lymph vessel tumor emboli were measured, and their structural features, nuclear atypia, and numbers of mitotic and apoptotic figures were also assessed. Multiple regression analysis showed the dimension, the distance, the number of mitotic figures, the number of apoptotic figures, and papillary features of lymph vessel tumor emboli to be significantly associated with the increased number of cells invading the lymph vessels (P<.05). The Cox proportional hazard multivariate analyses showed that more than six apoptotic figures in lymph vessel tumor emboli significantly increased the hazard rates (HRs) of tumor recurrence and death in IDCs without nodal metastasis and that more than four mitotic figures in lymph vessel tumor emboli significantly increased the HRs of tumor recurrence and death in IDCs with nodal metastasis (P <.05). The present study showed that the histological characteristics of tumor cells in lymph vessels play a very important role in the tumor progression of IDCs.
MAST CELLS
The presence of stromal mast cells identifies a subset of invasive breast cancers with a favorable prognosis. Dabiri S, Huntsman D, Makretsov N, Cheang M, Gilks B, Badjik C, Gelmon K, Chia S, Hayes M. 1Department of Pathology, British Columbia Cancer Agency, Vancouver, BC, Canada.	Mod Pathol. 2004 Jun;17(6):690-5. Abstract quote   Tissue microarrays containing 348 cases of invasive breast carcinoma were studied by immunohistochemical staining for CD-117, CD-3, CD-20, CD-68, Her2, estrogen receptor protein, and progesterone receptor protein, and results were correlated with patient outcome. Hormone receptor status (both estrogen receptor and progesterone receptor) correlated with a good outcome while Her2 overexpression was associated with a poor outcome. The presence of mast cells in the stroma, as demonstrated by positive c-kit (CD-117) staining, correlated with a good prognosis (P=0.0036). On subset analysis, this association between the presence of mast cells and favorable prognosis was present in the node-negative patients (P=0.018). The presence of mast cells showed an inverse correlation with the presence of CD-68 positive macrophages. No correlation was observed between the presence of mast cells and either B-cells (CD20-positive) or T-cells (CD3-positive). The presence of stromal mast cells was of prognostic significance independent of nodal status and tumor size (P=0.02). When the multivariate analysis was expanded to include tumor grade, estrogen receptor status and Her2 status, as well as tumor size and nodal status, the presence of stromal mast cells approached significance as an independent prognostic indicator.
METALLOPROTEINASE
Correlation of Tissue Inhibitor of Metalloproteinase-2 with Proliferative Activity and Patients’ Survival in Breast Cancer Lydia Nakopoulou, M.D., Sophia Katsarou, M.D., Ioanna Giannopoulou, Paraskevi Alexandrou, M.D., Ioanna Tsirmpa, Effie Panayotopoulou, Johnny Mavrommatis and Antonios Keramopoulos, M.D. Department of Pathology (LN, SK, IG, PA, IT, EP, JM) and First Department of Gynaecology and Obstetrics (AK), Medical School, The National and Kapodistrian University of Athens, Athens, Greece	Mod Pathol 2002;15:26-34 Abstract quote Tissue inhibitors of metalloproteinases (TIMPs) are endogenous regulators of matrix metalloproteinases (MMPs). They are believed to possess several distinct cellular functions, particularly the contradictory activities of inhibiting MMPs and promoting tumor cell growth. Immunohistochemistry was performed to detect TIMP-2 protein in 136 infiltrative breast carcinomas. TIMP-2 protein was analyzed in parallel with clinicopathologic features (tumor size, histologic type, nuclear and histologic grade, stage), patients’ overall survival and ER, PR, Ki-67, topo II, c-erbB-2, p53 and bcl-2 proteins. Statistical analysis was performed using univariate and multivariate models analysis. Immunoreactivity for TIMP-2 was observed in cancer cells and stromal fibroblasts in 106 (77.94%) and 104 (76.47%) of 136 cases, respectively. TIMP-2 protein expression in stromal fibroblasts showed a statistically significant inverse correlation with tumor size (P = .014). An inverse correlation was also observed between TIMP-2 epithelial immunoreactivity and nuclear and histologic grade (P = .036 and P = .007, respectively). TIMP-2 protein reactivity showed statistically significant positive associations with topo II and bcl-2 in stromal and cancer cells, respectively (P = .032 and P = .001, respectively). TIMP-2 protein expression in cancer and stromal cells was associated with better patients’ overall survival (P = .002 and P = .038, respectively). When evaluated by the Cox’s proportional hazard regression model, this association was further established, but only as far as TIMP-2 expression in tumor epithelium was concerned (P = .019). Our results support the multifunctional potential of TIMP-2 through its correlation on the one hand to a favorable outcome, due to its MMP inhibitory activity and on the other to topo II contributing to its growth factor activity.
MICROARRAY GENE EXPRESSION
Utilizing Nottingham Prognostic Index in microarray gene expression profiling of breast carcinomas. Miller DV, Leontovich AA, Lingle WL, Suman VJ, Mertens ML, Lillie J, Ingalls KA, Perez EA, Ingle JN, Couch FJ, Visscher DW. Department of Anatomic Pathology, Mayo Clinic, Rochester, MN 55906, USA.	Mod Pathol. 2004 Jul;17(7):756-64. Abstract quote   We report a novel approach to gene expression profiling using the Nottingham Prognostic Index (NPI) to stratify 26 patients with invasive breast carcinoma. As an aggregate index of parameters reflecting metastatic potential, growth rate, and genetic instability the NPI has distinct advantages over other clinicopathologic features used to segregate breast cancer patients. As a continuous variable it offers a responsive and sensitive means of modeling a continuum of clinical aggressiveness. Using RNA extracted from 26 tumors and cDNA microarrays with 23 343 unique genetic elements, 84 genes and expressed sequence tags were identified whose expression patterns correlated with NPI. Differential expression by immunohistochemistry (IHC) was also observed for two of three genes evaluated by this method. Correlation was determined by the Spearman rank correlation method with null distribution analysis. Among the 84 genetic elements were seven previously implicated in neoplastic progression (including the two demonstrating differential expression by IHC), 11 without specific cancer association but localized to chromosomal sites whose loss or gain has been identified in cytogenetic studies of breast carcinoma, and 73 not previously associated with breast carcinoma. Collectively, the expression patterns of these 84 elements have potential to distinguish high and low NPI patient samples. These data add support to the assertion that prognostic groups of breast carcinoma are reflected in distinguishable expression profiles of a limited set of genes.
MITOTIC FIGURE COUNT
Prognostic relevance of histological grade and its components in node-negative breast cancer patients. Volpi A, Bacci F, Paradiso A, Saragoni L, Scarpi E, Ricci M, Aldi M, Bianchi S, Muretto P, Nuzzo F, Simone G, Mangia A, Schittulli F, Amadori D. Division of Oncology and Diagnostics, Pierantoni Hospital, Forli, Italy.	Mod Pathol. 2004 Sep;17(9):1038-44. Abstract quote   Available results highlight the lack of good level of evidence studies on the pure prognostic value of histological grade. In the present study, the prognostic relevance of histological grade and of its three components, tubule formation, nuclear pleomorphism and mitotic count, was analyzed in a series of 372 patients with node-negative breast cancer treated with locoregional therapy alone until early relapse. Histological grade was determined blindly by two observers and discordance between evaluations was resolved after joint review using a multihead microscope. No relation was observed between histological grade and any of its three components and disease-free survival. Conversely, a significant relation was observed between histological grade and distant metastasis-free survival (at 6 years, 94, 86 and 76% for grades 1, 2 and 3, respectively, P=0.013) as well as overall survival (98, 90 and 86%, P=0.001). A breakdown analysis as a function of the three components showed that neither tubule formation nor nuclear pleomorphism was associated with prognosis, and only mitotic count strongly influenced both distant metastasis-free survival (91, 82 and 74%, P=0.014) and overall survival (97, 87 and 85%, P=0.011). Histological grade suffers from a much higher subjectivity than any other microscopic evaluation of biomarkers as it is the sum of three different morphological features. Within the Italian Network for Quality Assessment of Tumor Biomarkers program we observed that histological grade is an independent prognostic variable, but also that this role is ascribable only to the number of mitotic figures. In conclusion, due to the ever smaller size of diagnosed breast cancers, resulting in less cancer tissue for biofunctional and molecular analysis, mitotic count evaluated under strict quality control conditions seems to be an accurate and feasible prognostic variable.
Prognostic relevance of mitotic activity in patients with node-negative breast cancer. Medri L, Volpi A, Nanni O, Vecci AM, Mangia A, Schittulli F, Padovani F, Giunchi DC, Vito A, Amadori D, Paradiso A, Silvestrini R. Pathology Unit, Division of Oncology and Diagnostics, Pierantoni Hospital, Forli, Italy.	Mod Pathol. 2003 Nov;16(11):1067-75 Abstract quote.   The prognostic relevance of mitotic activity was analyzed in a series of 306 patients with node-negative breast cancer treated with locoregional therapy alone, until early relapse. Mitotic activity was evaluated as the number of mitotic figures per 10 high-power fields (mitotic activity index) or per 1000 tumor cells (mitotic index). Counting was carried out blindly by two observers. A high correlation was observed between the two determinations (r(s) =.96, P <.001). For clinical analysis, three mitotic activity index subgroups (mitotic figures/field </= 9, 10-19 and more than 19, according to grading criteria) and three mitotic index subgroups (percentage of mitotic figures less than 0.10, 0.11-0.50 and more than 0.50, according to tertile criteria) were considered. No relation was observed between mitotic variables and 6-year disease-free survival, whereas distant disease-free survival was strongly related to mitotic figures per 10 fields (85%, 89% and 70%, P =.012) and to the percentage of mitotic figures out of a total 1000 tumor cells (87%, 86% and 75%, P =.017). Similarly, both mitotic indices were significantly related to 6-year overall survival (99%, 95% and 77%, P <.001, for mitotic figures per 10 fields and 99%, 93% and 82%, P <.001, for the percentage of mitotic figures). These findings were particularly evident in patients with tumors of 1-2 cm. In conclusion, a high number of mitotic figures is associated with a higher probability of developing distant metastases and a shorter survival. The critical point remains the standardization of the preanalytical and analytical steps within quality control programs.
MUCIN
Expression of mucins (MUC1, MUC2, MUC3, MUC4, MUC5AC and MUC6) and their prognostic significance in human breast cancer. Rakha EA, Boyce RW, Abd El-Rehim D, Kurien T, Green AR, Paish EC, Robertson JF, Ellis IO. 1Department of Histopathology, The Breast Unit, Nottingham City Hospital, University of Nottingham, Nottingham, UK.	Mod Pathol. 2005 Oct;18(10):1295-304. Abstract quote   Mucins are a large family of glycoproteins expressed by many epithelial cells and their malignant counterparts. Much interest has been focused on expression of its members in breast cancer because of their potential role as prognostic indicators and their involvement in cancer therapy. We have examined 1447 cases of invasive breast carcinoma with a long-term follow-up, using tissue microarray (TMA) technology and immunohistochemistry to evaluate the expression profiles of several mucins (MUC1, MUC2, MUC3, MUC4, MUC5AC and MUC6) and to assess their prognostic value. We detected MUC1 expression in 91% of tumours. MUC1 overexpression was associated with a lower grade, smaller tumour size, a higher oestrogen receptor (ER)-positive phenotype and absence of both regional recurrence and distance metastasis. The subcellular localization but not the level of expression had a prognostic value in predicting outcome. The aberrant cytoplasmic and membranous localization of MUC1 was associated with poor outcome compared with apical localization, which is the normal physiological site of expression. MUC2 expression was noticed in only 8.3% of all cases and was restricted to the cytoplasm of the tumour cells. An inverse trend was identified between MUC2 expression and lymph node stage and vascular invasion status. On excluding cases of mucinous carcinoma from the analysis, the inverse association with vascular invasion was still defined and in addition an inverse association with ER status emerged. MUC3 expression was detected in 91% of cases and its expression was associated with increased local recurrence, and lymph node stage. The membranous expression of MUC3 was found to be a potentially poor prognostic feature, with higher grade and poorer Nottingham Prognostic Index (NPI), and negative ER expression. MUC4, MUC5AC and MUC6 were expressed in 95, 37 and 20% of cases, respectively. Apart from an association between MUC4 expression and tumour grade and between MUC6 and ER-negative tumours, no other associations with any clinicopathological variables were found. Apart from the higher expression of MUC2 and MUC6 in mucinous carcinomas, no association was found between the expression of different mucins and tumour type. No association between the level of expression of any of the studied mucins and patient outcomes has been identified. In conclusion, most breast carcinomas express MUC1, MUC3 and MUC4. Among the various mucins expressed in breast cancer, MUC1 and MUC3 are potential prognostic indicators, MUC1 having the strongest relationship with patient outcome.
MYXOID CHANGE
Breast cancer stromal myxoid changes are associated with tumor invasion and metastasis: a central role for hyaluronan. Wernicke M, Pineiro LC, Caramutti D, Dorn VG, Raffo MM, Guixa HG, Telenta M, Morandi AA. Department of Pathology (MW, LCP, DC, VGD, MMLR, MT, AAM) and Breast Cancer Service (HGG), Hospital Italiano, Buenos Aires, Argentina.	Mod Pathol 2003 Feb;16(2):99-107 Abstract quote The interplay between a tumor and its environment is exemplified by the morphological changes observed in the stroma of human breast cancer. These changes are evident as stromal myxoid changes. Hyaluronan, an extracellular polysaccharide that has been implicated in invasion, is one of the major constituents of the stromal myxoid changes. This study evaluated the association of these stromal changes with axillary node status, tumor grade, and mortality. The prognostic value of the stromal myxoid changes was evaluated in patients with negative axillary nodes with 10 years of follow-up. Our results showed a high level of reproducibility of our stromal myxoid changes grading system (overall kappa = 0.68). Image analysis semiquantification showed marked correlation of a strong stromal hyaluronan signal with high-grade stromal myxoid changes. In a multiple logistic regression analysis, positive nodes were associated with stromal myxoid changes, tumor size, desmoplasia, lymphocytic infiltration, high tumor grade, tumor emboli, and multifocality. Stromal myxoid changes were also associated with young age and lymphatic embolizations (P <.001). Overall, there is a weak correlation between mortality and stromal myxoid changes (P <.01). Mortality was more evident with high stromal myxoid changes grades and tumor size >2 cm (P <.008). However Cox multivariate analysis fail to show stromal myxoid changes as an independent prognostic factor. In conclusion, stromal myxoid changes with high hyaluronan concentration are strongly associated with positive nodes, tumor grade, and lymphatic emboli, thereby identifying high-risk group and reinforcing the role of hyaluronan in invasion and metastasis.
NEUROENDOCRINE DIFFERENTIATION
Tissue microarray analysis of neuroendocrine differentiation and its prognostic significance in breast cancer. Makretsov N, Gilks CB, Coldman AJ, Hayes M, Huntsman D.	Hum Pathol. 2003 Oct;34(10):1001-8. Abstract quote   The aim of this study was to detect neuroendocrine differentiation (NE), to determine its association with major clinicopathological parameters of breast cancer, and to study the prognostic significance of NE differentiation in a group of breast carcinomas by using tissue microarray (TMA) methodology. NE differentiation was studied by using 3 markers, synaptophysin (Syn), chromogranin A (ChA), and neuron-specific enolase (NSE) in a group of 334 patients with breast carcinoma. TMA blocks were made by using duplicate 0.6-mm-diameter tissue cores from each paraffin block. Results of immunostaining were scored on a 4-point scale, that is, as negative, weak, intermediate, and strong immunoreactivity. Positive staining of breast cancers for any of the 3 NE markers was detected in 19.5% of cases. Expression of a single marker was present in 16.2% of cases, and expression of 2 or 3 markers in combination was detected in 3.3% of cases. There was no statistically significant correlation of NE phenotype with tumor morphology, except mucinous carcinoma (3 of 6 cases positive), estrogen receptor, progesterone receptor, or nodal status. A weak correlation was noted between synaptophysin staining and higher tumor grade (P = 0.029). Analysis of disease-specific and overall survival based on up to 20 years of follow-up data showed a correlation between NSE expression and improved disease-specific (P = 0.043) and overall survival (P = 0.03) in univariate but not in multivariate analysis. The expression of Syn and ChA, as well as coexpression of multiple NE markers, had no prognostic significance.
Neuroendocrine differentiation and prognosis in breast adenocarcinoma. Miremadi A, Pinder SE, Lee AH, Bell JA, Paish EC, Wencyk P, Elston CW, Nicholson RI, Blamey RW, Robertson JF, Ellis IO. Department of Histopathology, Nottingham City Hospital, Nottingham, UK.	Histopathology 2002 Mar;40(3):215-22 Abstract quote Aims: Neuroendocrine differentiation has been detected, and its prognostic value studied, in a number of common human carcinomas. To date there are few detailed studies examining its relevance in breast carcinoma. In this study we evaluate the frequency and prognostic importance of neuroendocrine differentiation in breast adenocarcinoma.Methods and results: The presence of neuroendocrine differentiation, defined as positive reactivity for three markers, neuron-specific enolase (NSE), chromogranin A and/or synaptophysin, has been evaluated in 99 patients with primary operable breast cancer using standard immunocytochemical techniques. A consecutive cohort of patients were selected from the Nottingham/Tenovus series. Comprehensive patient and tumour records have been maintained, and patients were followed up according to a defined protocol. Eighteen cases were positive for NSE, 10 for chromogranin A and 13 for synaptophysin. Eleven percent were positive with more than one neuroendocrine marker. No significant association was found between neuroendocrine differentiation and tumour size, grade, stage or the prevalence of vascular invasion. There was no significant difference in either overall or disease-free survival between patients with or without neuroendocrine differentiation. Conclusions: In this study we confirm that neuroendocrine differentiation can be identified in a subset (10-18%) of human breast carcinomas. This phenomenon appears to have no relationship to established prognostic factors or patient outcome.
NUCLEAR FEATURES
Importance of nuclear morphology in breast cancer prognosis. Wolberg WH, Street WN, Mangasarian OL. Department of Surgery, University of Wisconsin, Madison 53792, USA	Clin Cancer Res 1999 Nov;5(11):3542-8 Abstract quote The purpose of this study is to define prognostic relationships between computer-derived nuclear morphological features, lymph node status, and tumor size in breast cancer. Computer-derived nuclear size, shape, and texture features were determined in fine-needle aspirates obtained at the time of diagnosis from 253 consecutive patients with invasive breast cancer. Tumor size and lymph node status were determined at the time of surgery. Median follow-up time was 61.5 months for patients without distant recurrence. In univariate analysis, tumor size, nuclear features, and the number of metastatic nodes were of decreasing significance for distant disease-free survival. Nuclear features, tumor size, and the number of metastatic nodes were of decreasing significance for overall survival. In multivariate analysis, the morphological size feature, largest perimeter, was more predictive of disease-free and overall survival than were either tumor size or the number of axillary lymph node metastases. This morphological feature, when combined with tumor size, identified more patients at both the good and poor ends of the prognostic spectrum than did the combination of tumor size and axillary lymph node status. Our data indicate that computer analysis of nuclear features has the potential to replace axillary lymph node status for staging of breast cancer. If confirmed by others, axillary dissection for breast cancer staging, estimating prognosis, and selecting patients for adjunctive therapy could be eliminated.
Computer-generated nuclear features compared with axillary lymph node status and tumor size as indicators of breast cancer survival. Wolberg WH, Street WN. Department of Surgery, University of Wisconsin, Madison, WI and the Management Sciences Department, University of Iowa, Iowa City, IA.	Hum Pathol 2002 Nov;33(11):1086-91 Abstract quote The extent to which malignant cells deviate from normal is generally accepted to be a prognostic indicator. However, assessing the degree of deviation has been subjective and poorly reproducible. Our goal is to develop a computer program for objectively measuring nuclear size, shape, and texture from histologic slides and to make the program available on the Internet. We used this program to analyze 353 histologic sections obtained from patients with invasive breast cancer who were diagnosed and treated from 1981 through 1995 and who had determinable outcomes. The median follow-up was 8.3 years. We compared the relationship of survival with our computer-derived nuclear features versus axillary lymph node status and tumor size. We believe that our results are generally applicable because our patient survival, when stratified by lymph node status, was similar to that of the 24,000 breast cancer patients in the National Cancer Institute's Surveillance, Epidemiology, and End Results program. In multivariate analysis, the strongest prognostic factor was the largest nuclear area, followed by tumor size and the extent of axillary lymph node involvement. The mean area of the 3 largest nuclei when combined with tumor size identified 30% of all breast cancer patients who had an 87% 15-year breast cancer-specific survival. Inclusion of lymph node status added little to this 2-factor model. Routine axillary lymph node surgery for prognostic purposes may become unnecessary, because nuclear features may provide sufficient information.
ONCOGENES
Concurrent overexpression of p53 and c-erbB-2 correlates with accelerated cycling and concomitant poor prognosis in node-negative breast cancer Pierre Rudolph, etal.	Hum Pathol 2001;32:311-319 (Abstract quote) Simultaneous overexpression of c-erbB-2 and p53 has been reported to be prognostically unfavorable in breast cancer. Herein, we show that concurrent overexpression of these 2 proteins is associated with a marked reduction in the relative fraction of cells in G1 phase of the cell cycle, indicating an accelerated cell cycle progression. Using an immunohistochemical approach, we examined 261 cases of node-negative infiltrating ductal carcinomas of the breast with respect to c-erbB-2 and p53 expression and to the proliferative activity measured by the Ki-67 index. By means of a novel monoclonal antibody, Ki-S2, which exclusively recognizes proliferating cells in the S, G2, and M phases of the reproductive cycle, we were further able to calculate the relative fraction of the cells having passed the restriction point at the G1/S boundary, thus defining a cycling ratio (CR). The results were correlated with clinical outcome; median follow-up time was 96 months. Tumors that simultaneously overexpressed c-erbB-2 and p53 had a high median CR and followed an unfavorable course. However, increased CRs were also observed independently of c-erbB-2 and p53 overexpression, suggesting that other molecular mechanisms may contribute to acceleration of cell cycle progression. In a multivariate analysis that included patient age, tumor size, hormone receptor status, c-erbB-2 and p53 expression, and the Ki-67 index, CR emerged as the most significant independent predictor of overall and disease-free survival (P < .0001). It is concluded that the CR is a gauge of cell cycle deregulation and therefore may be a powerful indicator of the biologic behavior of cancers.
Loss of Expression of the PTEN Gene Protein Product Is Associated with Poor Outcome in Breast Cancer Peter L. Depowski, M.D., Seth I. Rosenthal, M.D. and Jeffrey S. Ross, M.D. Department of Pathology and Laboratory MedicineAlbany Medical College, Albany, New York	Mod Pathol 2001;14:672-676 Abstract quote The PTEN gene, a candidate tumor suppressor, is localized to chromosome 10q23 and shares extensive homology with cytoskeletal proteins auxilin and tensin. A high frequency of mutations at the PTEN locus has been described in a variety of neoplasms including breast cancer. However, the role of PTEN alternations and its association with outcome variables in breast neoplasia is not well established. DESIGN: Formalin-fixed paraffin embedded tissues from 151 women (mean age 62 years, range 26–98) with primary diagnosis of invasive breast cancer were evaluated for PTEN protein expression by automated immunohistochemical methods. Slides were scored semi-quantitatively based on staining intensity and distribution, and results were compared with clinical pathologic parameters. The mean follow-up was 56 months (range 1–169). RESULTS: Seventy-three (48%) of 151 breast tumors had loss of PTEN protein expression. On univariate analysis, loss of PTEN expression (P = .034), stage (P < .0001), node positive (P < .0001), and tumor grade (P = .002) were associated with disease-related death. Loss of PTEN expression also predicted lymph node metastasis (P < .0001), and correlated with loss of estrogen receptor staining (P = .040). Loss of PTEN did not correlate with stage, tumor grade, disease recurrence, or loss of progesterone receptor [although a trend was seen (P = .092). On multivariate analysis, stage (P < .0001), lymph node metastasis (P < .0001), and tumor grade (P = .002) correlated with survival. CONCLUSION: Loss of PTEN protein expression occurs commonly in breast cancer and correlates with disease related death, lymph node metastasis, and loss of estrogen receptor staining. Our results support the proposed role of PTEN as a candidate tumor suppressor in breast cancer and suggest a need for further study of this marker.
Amplification of c-myc by Fluorescence In Situ Hybridization in a Population-Based Breast Cancer Tissue Array Jaana K. Rummukainen, M.D., Tiina Salminen, M.D., Johan Lundin, M.D., Ph.D., Soili Kytölä, Ph.D., Heikki Joensuu, MD Ph.D. and Jorma J. Isola, MD Ph.D. Laboratory of Cancer Genetics (JKRSK, JJI), Institute of Medical Technology, University of Tampere and Tampere University Hospital, Tampere Finland; and Department of Oncology (TS, JL, HJ), Helsinki University Central Hospital, Helsinki, Finland	Mod Pathol 2001;14:1030-1035 Abstract quote A total of 261 primary breast carcinomas were analyzed for amplification of the c-myc oncogene by fluorescence in situ hybridization performed on tumor tissue array samples. Results were compared with individual clinicopathologic and follow-up data. Thirty-eight (14.6%) of the tumors showed c-myc gene amplification (defined as two or more additional copies of c-myc gene in relation to the number of chromosome 8 centromere). The reproducibility of fluorescence in situ hybridization assay (defined by hybridization with two different myc probes) was good (kappa coefficient 0.402). Statistically significant associations were found between c-myc amplification and DNA aneuploidy (P = .0011), and progesterone receptor negativity (P = .0071), and c-myc amplification also tended to be associated with high histologic grade (P = .064), positive axillary nodal status (P = .080), and a high S-phase fraction (P = .052). c-myc amplification was not significantly associated with overall survival of patients with invasive cancer (P = .32). These data from a population-based tumor material suggest that c-myc amplification is a feature of aggressive breast cancers, but that it is unlikely to be a clinically useful prognostic factor.
p53
p53 in node-negative breast carcinoma: an immunohistochemical study of epidemiologic risk factors, histologic features, and prognosis. Rosen PP, Lesser ML, Arroyo CD, Cranor M, Borgen P, Norton L. Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.	J Clin Oncol 1995 Apr;13(4):821-30 Abstract quote PURPOSE: The present study explores p53 in relation to the following four aspects of node-negative breast carcinoma: epidemiologic risk factors, tumor histopathology, prognosis, and HER2/neu (HER) expression. MATERIALS AND METHODS: Immunohistochemical (IH) staining for p53 was performed on formaldehyde-fixed, paraffin-embedded primary invasive carcinomas from 440 node-negative patients with a median follow-up duration of 119 months. RESULTS: The IH expression, or lack thereof, of p53 separately or in combination with HER did not prove to be prognostically significant and there was no consistent association of p53 with epidemiologic risk factors. p53 was expressed in 68% of medullary carcinomas (MEDs), which is a significantly higher frequency (P < .001) than in lobular (9%) and duct (23%) carcinomas. p53 was not found in some types of low-grade carcinomas (tubular and papillary), and was observed in a minority of mucinous carcinomas. p53 was present significantly more often in carcinomas with high-grade or poorly differentiated nuclear grade than in low- or intermediate-grade tumors. There was an inverse statistically significant relationship between estrogen receptor (ER) positivity and p53 expression. Tumors with the p53(+)/HER(-) immunophenotype tended to be MEDs or duct carcinomas with a marked lymphoplasmacytic reaction. Infiltrating lobular carcinomas (IFLCs) were largely p53(-)/HER(-). p53(+)/HER(+) carcinomas had the best prognosis. The poorest outcome was associated with the p53(-)/HER(+) immunophenotype. This trend was statistically significant for recurrence-free and overall survival in patients with T1NOMO infiltrating duct carcinoma (IFDC). CONCLUSION: The IH demonstration of p53 was not a reliable prognostic indicator in the node-negative breast carcinoma patients studied and it was not associated with major epidemiologic risk factors. The combined immunophenotypic expression of p53 and HER was significantly associated with some histologic types of breast carcinoma and with prognosis in T1NOMO breast carcinoma.
Validation of p53 Immunohistochemistry as a Prognostic Factor in Breast Cancer in Clinical Practice Angela N. Bartley, MBS and Dennis W. Ross, MD, PhD From Wake Forest University School of Medicine (Ms Bartley); and the Department of Pathology, Forsyth Medical Center, Winston-Salem, NC (Dr Ross)	Arch Pathol Lab Med 2002;Vol. 126, No. 4, pp. 456–458. Abstract quote Context.—Abnormal p53 tumor suppressor gene expression as detected by immunohistochemistry is a possible prognostic factor in breast cancer. The difference in techniques used to evaluate the expression of mutated p53 protein is under intense scrutiny, as well as its uses either independently or in conjunction with other prognostic factors in breast cancer. Objective.—To determine whether p53 immunohistochemistry can be used as a reliable indicator of the presence of mutated nuclear p53 protein, and whether this method can be performed reliably in a community hospital's clinical practice. Design.—One hundred twenty-two cases of breast carcinoma were stained and analyzed for the presence of p53 protein using DO-7 (Dako Corporation, Carpinteria, Calif) p53 antibody. Results.—Of the 122 cases of invasive carcinoma studied, 23 (18.7%) were positive for p53, and 16 (16.3%) of 98 cases with coexisting ductal carcinoma in situ were positive for p53. This finding is in agreement with comparable published studies. Conclusions.—Based on the results of this study, we conclude that p53 immunohistochemistry qualifies as a diagnostic technique suitable for clinical practice in a community hospital. Its detection may be particularly promising in clinical trials of new molecular therapies directed at the p53 tumor suppressor gene.
PREGNANCY ASSOCIATED
Breast carcinoma presenting during or shortly after pregnancy and lactation. Shousha S. Department of Histopathology, Imperial College School of Medicine and Charing Cross Hospital, London, England.	Arch Pathol Lab Med 2000 Jul;124(7):1053-60 Abstract quote CONTEXT: Much has been written about the clinical management and prognosis of breast carcinomas presenting during pregnancy and lactation, yet little is known about the detailed histopathology of these tumors. OBJECTIVE: To determine whether these carcinomas have any specific diagnostic features. DESIGN: A detailed histologic and immunohistochemical study of 14 cases of breast carcinoma presenting during or shortly after pregnancy or lactation was conducted. The findings were compared with a control group of 13 tumors developing in age-matched women with no recent history of pregnancy or lactation. SETTING: The histopathology department of a tertiary referral teaching hospital. RESULTS: Tumors in the pregnancy/lactation group had a significantly higher incidence of cancerization of lobules (79% vs 15%) and of grade III invasive ductal carcinomas (80% vs 33%). Tumors occurring during lactation were either totally or partly mucinous and were MUC2 positive. Tumors occurring during pregnancy, but not during lactation, were mostly estrogen and progesterone receptor negative (4/5 and 5/5, respectively). All tumors occurring during pregnancy and lactation that were tested for c-erbB-2 overexpression were negative, whereas all 4 tumors tested that occurred shortly after delivery or cessation of lactation were positive for c-erbB-2 overexpression. The incidence of axillary lymph node metastasis was high in both the study and control groups, although it was slightly higher in the control group (78% and 90%, respectively). CONCLUSIONS: Although breast carcinomas diagnosed during or shortly after pregnancy and lactation have features in common with those developing in women of similar ages, particularly with respect to a high incidence of lymph node metastasis, the findings of this study suggest that they may also have distinct morphologic and immunohistochemical features of their own. Such features may vary according to whether the patient was pregnant, lactating, or had recently terminated her pregnancy or lactation at the time of surgical excision. Examination of more cases would help confirm these findings.
Pregnancy-associated breast cancer: a case-control study in a young population with a high-fertility rate. Ibrahim EM, Ezzat AA, Baloush A, Hussain ZH, Mohammed GH. Department of Oncology, King Faisal Specialist Hospital and Research Centre, Kingdom of Saudi Arabia.	Med Oncol 2000 Nov;17(4):293-300 Abstract quote Pregnancy-associated breast cancer (PABC) is not a rare event. The association frequently imposes a management challenge. We intended to review the clinical features, therapy, and outcome of patients with PABC seen at a single institution over a five-year period and to compare those with that seen in a matched control group. Data of all patients with PABC diagnosed during pregnancy were retrospectively reviewed (Group I). For each patient in Group I, three matched controls with breast cancer without pregnancy were identified (matched for age, stage, and year of diagnosis, Group II). 72 patients in Group I and 216 in Group II were identified. Their median age was similar (34 vs 35 y, respectively). The median number of prior pregnancies for patients in Groups I and II was 5. Patients had shorter duration of symptoms prior to diagnosis as compared with their controls (5.6 vs 9.4 months, P < 0.0001). 3%, 31%, 40%, and 26% of patients had Stage I to IV, respectively. A pattern that was similar to that seen in our breast cancer population. Pregnancy was terminated in 34 patients (47%), while 38 (53%) had normal spontaneous vaginal delivery. 47 patients in Group I had surgery; 37 (52%) had modified radical mastectomy and 10 (14%) had conservative surgery. In 37 patients surgery was performed after termination of pregnancy and 10 had surgery performed during pregnancy. The median number of positive lymph nodes in Group I was 4 as compared with 2 for patients in Group II. No patients in Group I had systemic chemotherapy during first trimester, while only 4 (6%) and 3 (4%) received adjuvant or neoadjuvant during second and third trimester, respectively. No congenital malformation in the newborns was diagnosed. None of the patients in Group I received radiotherapy during pregnancy. Over a median of 47.5 months, 48 (67%) patients in Group I were alive as compared to 126 (58%) in Group II, with no difference in the median survival (P= 0.79). Comparing overall survival (OS) between the two groups stage for stage also showed no significant difference. Also there was no difference in progression-free survival between the two groups. Cox proportional hazard model identified advanced stage as the only independent adverse prognostic variable that influenced OS in Group I. Despite that this series included a relatively young population with a high fertility rate, the study confirmed the lack of a survival difference between patients with PABC and their matched controls.
PREGNANCY, CONCURRENT
Chemotherapy for breast carcinoma during pregnancy: A French national survey. Giacalone PL, Laffargue F, Benos P. Department of Obstetrics and Gynecology, Hopital Arnaud de Villeneuve, Montpellier, France.	Cancer 1999 Dec 1;86(11):2266-72 Abstract quote BACKGROUND: During pregnancy, the need for maternal chemotherapy for breast carcinoma must be balanced against the fetal risk because modification of cancer therapy to assure the birth of a healthy infant may affect maternal prognosis adversely. To the authors' knowledge few studies have documented the oncologic and obstetric management of this association. METHODS: A retrospective nationwide survey was used to identify women treated with chemotherapy for breast carcinoma during pregnancy. Each member of the Societe Francaise d'Oncologie Gynecologique and the Societe Francaise de Senologie et de Pathologie Mammaire completed a postal questionnaire regarding cancer staging, oncologic treatment, obstetric details, pregnancy outcome, fetal behavior, and postdelivery follow-up. Twenty women were accrued to the study. RESULTS: The mean gestational age at the first cycle of treatment was 26 weeks. A total of 38 cycles were administered during pregnancy, with a median of 2 cycles. Delivery was performed at a mean of 34.7 weeks. Two pregnancies that were exposed to chemotherapy during the first trimester resulted in spontaneous abortion. One pregnancy exposed in the second trimester resulted in intrauterine death. The remaining 17 pregnancies resulted in live births, although 3 women had complications related to chemotherapy (anemia, leukopenia, and fetal growth retardation) and 1 newborn died 8 days after birth without apparent etiology. Two newborns had complications related to prematurity (transient respiratory distress). At a mean follow-up of 42.3 months, all live infants were reported to have reached normal developmental milestones. CONCLUSIONS: The current study found that even when chemotherapy was initiated after the first trimester, 95% of the pregnancies resulted in live births with low related morbidity in the newborns.
PREGNANCY, SUBSEQUENT, IN BREAST CANCER SURVIVORS
Childbearing issues in breast carcinoma survivors. Surbone A, Petrek JA. Department of Surgery and Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.	Cancer 1997 Apr 1;79(7):1271-8 Abstract quote BACKGROUND: The issue of the subsequent pregnancy after breast carcinoma treatment is of paramount importance to young survivors and their oncologists. Matters related to having children, whether biologic or not, are analyzed. METHODS: Available evidence on the role of estrogen in the carcinogenesis and promotion of breast carcinoma is summarized. The scanty literature on pregnancy in breast carcinoma survivors is reviewed and evaluated. With reference to infertility as the result of adjuvant treatment, studies on therapy-induced amenorrhea are cited. RESULTS: The survival of women with breast carcinoma is not decreased by subsequent pregnancy in any of the published series. Nevertheless, several biases may be present, making the results less than conclusive; no prospective studies exist. Theoretic concern of tumor promotion may be justified when considering the long term exposure to intense gestational hormones in the presence of established breast carcinoma with possible micrometastases. As a separate issue, the common situation of chemotherapy-induced amenorrhea may not permit pregnancy. Information for the breast carcinoma survivor on assisted conception and adoption is limited. CONCLUSIONS: Further research on the safety of subsequent pregnancy after breast carcinoma treatment is needed; the authors report that they are initiating a multicenter prospective study to address these issues.
Pregnancy after breast carcinoma: outcomes and influence on mortality. Velentgas P, Daling JR, Malone KE, Weiss NS, Williams MA, Self SG, Mueller BA. Department of Biostatistics, University of Washington, Seattle, USA.	Cancer 1999 Jun 1;85(11):2424-32 Abstract quote BACKGROUND: To the authors' knowledge, no previous studies have identified an adverse effect of pregnancy on patient survival after breast carcinoma. However, results are difficult to interpret because of failure to control for stage of disease at the time the pregnancy occurred. METHODS: Study participants were women diagnosed with invasive breast carcinoma between 1983-1992 who previously had participated in a population-based case-control study or, if deceased, proxy respondents. Information regarding subsequent pregnancies was obtained by self-administered questionnaire or telephone interview. Information regarding breast carcinoma recurrences was obtained by questionnaire and from cancer registry abstracts. Women who became pregnant after a diagnosis of breast carcinoma (n = 53) were matched with women without subsequent pregnancies based on stage of disease at diagnosis and a recurrence free survival time in the comparison women greater than or equal to the interval between breast carcinoma diagnosis and onset of pregnancy in the women with a subsequent pregnancy. RESULTS: Sixty-eight percent of women who became pregnant after being diagnosed with breast carcinoma delivered one or more live-born infants. Miscarriages occurred in 24% of the patients who became pregnant compared with 18% of the controls (women without breast carcinoma) of similar ages from the case-control study. Five of the 53 women who had been pregnant after breast carcinoma died of the disease. The age-adjusted relative risk (RR) of death associated with any subsequent pregnancy was 0.8 (95% confidence interval [95% CI], 0.3-2.3). All five deaths occurred among the 36 women who had a live birth (age-adjusted RR = 1.1; 95% CI, 0.4-3.7). CONCLUSIONS: The findings of the current study are based on a small number of deaths but do not suggest that pregnancy after a diagnosis of breast carcinoma has an adverse effect on survival.
PTEN
Reduced PTEN expression predicts relapse in patients with breast carcinoma treated by tamoxifen. Shoman N, Klassen S, McFadden A, Bickis MG, Torlakovic E, Chibbar R. 1Department of Pathology, University of Saskatchewan, Saskatoon, SK, Canada.	Mod Pathol. 2005;18:250-259 Abstract quote Tamoxifen treatment substantially improves the 10-year survival of women with estrogen-receptor (ER)-alpha-positive tumors. However, approximately one-third of all breast cancer patients with ER-alpha-positive tumors progress on antiestrogen therapy. The molecular mechanism(s) involved in antiestrogen-resistant phenotype of breast carcinoma is not completely understood. The PTEN (phosphatase and tensin homolog deleted on chromosome Ten) gene is a novel candidate tumor suppressor that plays an important role in cell cycle regulation and apoptosis by regulating Protein kinase-B/Akt activity. Previous studies have shown that PTEN downregulation in breast cancer is associated with high-grade tumor, distant metastases and poorer disease-free survival. Decreased PTEN and/or increased protein kinase B/Akt activity in breast cancer cells has recently been associated with resistance to tamoxifen-induced apoptosis. In this study, we have evaluated PTEN expression by immunohistochemistry in 100 tamoxifen-treated ER-alpha-positive breast cancer patients. Reduced PTEN protein expression was associated with shorter relapse-free survival. When stage I patients were analyzed separately, reduced PTEN expression was a strong predictor of both, shorter relapse-free survival and shorter disease-specific survival. An association of reduced PTEN expression with shorter relapse-free survival and disease-specific survival in stage I patients was still observed after stratification by stage, axillary lymph node status, tumor size, grade, and expression of ER-alpha, progesterone receptor, and Her-2/neu. In summary, our results showed a strong association between downregulation of PTEN expression in ER-alpha-positive tumors and failure to tamoxifen treatment.
RACE
Racial differences in breast carcinoma survival. \Joslyn SA, West MM. University of Northern Iowa, Cedar Falls, Iowa 50614-0241, USA.	Cancer 2000 Jan 1;88(1):114-23 Abstract quote BACKGROUND: Survival after breast carcinoma diagnosis is significantly worse among African American women for reasons unknown. The purpose of this study was to update reports on the National Surveillance, Epidemiology, and End Results Program and to examine the effect of race on breast carcinoma survival. METHODS: Subjects were 135,424 women diagnosed with primary breast carcinoma between 1988-1995. Patient age, tumor stage at the time of diagnosis, hormone receptor status, tumor histology, menopausal status, and survival were compared by race category. RESULTS: African American women diagnosed with breast carcinoma (n = 11,159) had a significantly increased risk of death from breast carcinoma and from all cancers compared with white women (n = 124,265), independent of the effects of other predictor variables. African American women were significantly younger at the time of diagnosis, with approximately 33% of the population age CONCLUSIONS: The results of the current study show that race is an independent predictor of survival from breast carcinoma. These findings are consistent with other large, population-based studies of racial differences in breast carcinoma survival and have been comported by studies of racial differences in the molecular biology of breast carcinoma, thus providing support for the epidemiologic credibility of the independence of the association.
SECOND PRIMARY BREAST CANCERS
Breast carcinoma after cancer at another site: method of detection, tumor characteristics, and surgical treatment. Nissen MJ, Lazovich D, Jolitz G. Oncology Research, Park Nicollet Institute, Minneapolis, Minnesota, USA.	Cancer 2000 Nov 1;89(9):1999-2005Abstract quote BACKGROUND: As the number of cancer survivors increases, so will the number of second primary cancers, including breast carcinoma after cancer at another site. Limited information is available regarding the clinical characteristics of breast carcinoma after a primary at another site. METHODS: TUMORS (The Upper Midwest Oncology Registry Services) was used to identify 937 women with breast carcinoma occurring as a second primary after a first primary at a known site other than the breast. They were compared with a sample of 1874 women with first primary breast carcinoma, frequency-matched by age to the second primary group, for method of detection, tumor characteristics, and type of surgery. RESULTS: Women with breast carcinoma after cancer at another site tended to have smaller tumors and less extensive disease than women with first primary breast carcinoma and were somewhat more likely than first primary cases to have had their breast carcinoma detected by mammogram or clinical breast exam rather than detecting it themselves. Differences in method of detection accounted for differences in tumor size and extent. Second primary breast carcinoma was less likely to be lobular or mixed ductolobular carcinoma compared with first primary breast carcinoma. Surgical treatment (mastectomy vs. breast-conserving surgery) did not differ for first and second primary breast carcinoma. CONCLUSIONS: Clinical characteristics of breast carcinoma after cancer at another site were by and large similar to those of first primary breast carcinoma. The more favorable prognostic characteristics among women with a history of cancer were accounted for by increased medical surveillance.
SIZE OF TUMOR
Axillary lymph node metastases in patients with small carcinomas of the breast: is accurate prediction possible? Anan K, Mitsuyama S, Tamae K, Nishihara K, Iwashita T, Abe Y, Ihara T, Toyoshima S. Department of Surgery, Kitakyushu Municipal Medical Centre, Japan.	Eur J Surg 2000 Aug;166(8):610-5 Abstract quote OBJECTIVES: To find out whether macroscopic classification of the tumour margin is predictive of axillary lymph node metastases and to identify a combination of clinical and pathological findings by which axillary node status can be predicted accurately in small carcinomas (T1) of the breast. DESIGN: Retrospective study. SETTING: Municipal referral centre, Japan. SUBJECTS: All 1003 patients with T1 invasive carcinoma of the breast who had axillary lymph node dissection between January 1970 and December 1996 as part of their treatment. MAIN OUTCOME MEASURES: The association between the incidence of axillary lymph node metastases and 10 clinical and pathological factors (age, palpability and size of tumour, macroscopic classification of tumour margin, clinical axillary status, radiating spiculation on a mammogram, histological type, lymphatic invasion, oestrogen and progesterone receptor status) were analysed. RESULTS: Clinical axillary node status, macroscopic classification of tumour margin, lymphatic invasion, and age of the patient were significant predictors of axillary lymph node metastases (p < 0.01 in each case). Among 47 patients aged 65 or more whose tumours had well-defined margins and with a clinical N0 status in the axillae, the incidence of histological axillary lymph node metastasis was only 6% (n = 3) whereas it was 65% in 57 patients with tumours of ill-defined margins whose axillae were N1 or N2. CONCLUSIONS: Macroscopic classification of tumour margins is an independent predictor of axillary lymph node metastases for patients with small carcinomas of the breast. However, even with combinations of the examined predictors of axillary node metastases, the subgroup of patients at minimal risk of metastasis was less than 5% in T1 breast cancer, whereas three-quarters of the patients had clear axillary lymph nodes. Most patients with T1 breast cancer will need surgical staging of the axillae by methods such as sentinel node biopsy.
SKIN INVOLVEMENT
Breast carcinoma with noninflammatory skin involvement (T4b): time to abandon an historic relic from the TNM classification. Guth U, Wight E, Schotzau A, Langer I, Dieterich H, Rochlitz C, Herberich L, Holzgreve W, Singer G. Department of Gynecology and Obstetrics, University Hospital Basel, Basel, Switzerland.	Cancer. 2005 Nov 1;104(9):1862-70. Abstract quote   BACKGROUND: In this study, the authors evaluated the clinical presentation of patients with T4b breast carcinoma, analyzed the impact of noninflammatory skin involvement on long-term survival, and addressed the question whether the T4 tumor category still has any justification. METHODS: The clinical course of a study group of 119 patients with skin involvement was compared with the outcome of a control group of 299 consecutive patients who had tumors of the same size but without skin involvement. All tumors were stratified into 1 of 4 subsets according to greatest tumor dimension, as follows: Group A, < or = 3.0 cm; Group B, 3.1-5.0 cm; Group C, 5.1-10.0 cm; and Group D, > 10.0 cm. RESULTS: The study group distribution of patients within the size subsets were as follows: Group A, 26.1%; Group B, 24.3%; Group C, 26.1%; and Group D, 23.5%. Differences in disease-specific survival between the tumor size subsets were significant (Groups A and B vs. Groups C and D; P < 0.0001). In contrast to large tumors (> 5.0 cm), carcinomas < or = 5.0 cm showed no statistical significant differences in disease-specific survival between study group patients and control group patients (Group A: P = 0.17; Group B: P = 0.31). CONCLUSIONS: There is a broad range of clinicopathologic breast carcinoma entities within the T4b category. For > 50% of patients with T4b breast carcinoma, the feature noninflammatory skin involvement had no significant prognostic impact. Approximately 25% of patients had an extent of disease similar to that observed in patients with Stage I-II disease and, thus, falsely were considered to have more advanced disease. Heterogeneity and a lack of prognostic significance suggest that a revision of the T4 category, a relic of historic tumor classifications, is necessary.
STROMELYSIN-3
Stromelysin-3 protein expression in invasive breast cancer: relation to proliferation, cell survival and patients' outcome. Nakopoulou L, Panayotopoulou EG, Giannopoulou I, Alexandrou P, Katsarou S, Athanassiadou P, Keramopoulos A. Department of Pathology, Medical School, The National and Kapodistrian University of Athens, and Department of Pathology, "Alexandra's Hospital," Athens, Greece.	Mod Pathol 2002 Nov;15(11):1154-61 Abstract quote Matrix metalloproteinases constitute one of the major extracellular matrix degrading enzymic families implicated in cancer development. Stromelysin-3 in particular, a member of the matrix metalloproteinases belonging to the stromelysins' subgroup, seems to be closely related to invasiveness and tumor progression. In this study, we proceeded to the evaluation of stromelysin-3 protein's expression in paraffin sections of 133 cases of invasive breast carcinomas and statistically estimated its relations with known clinicopathological prognostic parameters and patients' survival, proliferation markers Ki-67 and TopoIIalpha and the antiapoptotic protein bcl-2. Presence of stromelysin-3 was immunodetected, in the 73% of our cases, in stromal cells (65%) and in epithelial tumor cells (26.26%). Stromelysin-3 epithelial positivity presented statistically significant correlations with TopoIIalpha and Ki-67 proliferation indices (P =.042 and P =.031, respectively) and worse disease outcome through multivariate statistics (P =.014). Stromelysin-3 fibroblastic expression was significantly associated with nuclear grade (P =.024), ductal histological type (P =.037), TopoIIalpha (P =.001) and Ki-67 (P =.019), inversely with bcl-2 protein (P =.027) and with adverse overall survival through univariate analysis (P =.017). The subgroup of patients with stromelysin-3 co-expression in stromal and malignant epithelial cells showed statistically significant associations with Ki-67 and TopoIIalpha (P =.019, P <.0001, respectively), an inverse one with bcl-2 protein (P =.027) and furthermore with impaired survival (P =.002) through multivariate analysis. In conclusion, stromelysin-3 protein expression correlated with proliferation indices TopoIIalpha and Ki-67 and the anti-apoptotic protein bcl-2, data confirming stromelysin-3's contribution to breast cancer progression. Moreover its expression was shown to have a direct negative effect on patients' survival, especially in the subgroup of patients with simultaneous epithelial and stromal expression.
TOPOISOMERASES
Topoisomerase II-alpha Expression in Different Cell Cycle Phases in Fresh Human Breast Carcinomas. Villman K, Stahl E, Liljegren G, Tidefelt U, Karlsson MG. Departments of Oncology (KV), Pathology (ES, MGK), Surgery (GL), and Medicine (UT), Orebro University Hospital, Orebro, Sweden.	Mod Pathol 2002 May;15(5):486-91 Abstract quote Topoisomerase II-alpha (topo IIalpha) is the key target enzyme for the topoisomerase inhibitor class of anti-cancer drugs. In normal cells, topo IIalpha is expressed predominantly in the S/G2/M phase of the cell cycle. In malignant cells, in vitro studies have indicated that the expression of topo IIalpha is both higher and less dependent on proliferation state in the cell. We studied fresh specimens from 50 cases of primary breast cancer. The expression of topo IIalpha in different cell cycle phases was analyzed with two-parameter flow cytometry using the monoclonal antibody SWT3D1 and propidium iodide staining. The expression of topo IIalpha was significantly higher in the S/G2/M phase of the cell cycle than in the G0/G1 phase in both DNA diploid and DNA nondiploid tumors. In 18 of 21 diploid tumors, and in 25 of 29 nondiploid tumors, >50% of the topo IIalpha-positive cells were in the G0/G1 phase. This significant expression of topo IIalpha in the G0/G1 phase of the cell cycle may have clinically important implications for treatment efficacy of topoisomerase II inhibitors.
TREAMENT INADEQUACIES
The effect of less than definitive care on breast carcinoma recurrence and mortality. Lash TL, Silliman RA, Guadagnoli E, Mor V. Department of Epidemiology and Biostatistics, Boston University School of Public Health, Boston, Massachusetts, USA.	Cancer 2000 Oct 15;89(8):1739-47 Abstract quote BACKGROUND: Risk factors for breast carcinoma offer few opportunities for prevention; thus, the reduction of morbidity and mortality among breast carcinoma patients must remain a priority. The objective of this study was to measure the effects of less than definitive care for patients with breast carcinoma on disease recurrence and mortality. METHODS: The prognostic evaluation and treatment received by an inception cohort of 494 women was characterized. Three hundred ninety women ages 45-90 years with local or regional breast carcinoma who were diagnosed between 1984 and 1986 and were treated at one of eight Rhode Island hospitals comprised the final cohort. Disease recurrence and mortality were ascertained through December 31, 1996. Candidate determinants of outcomes were a less than definitive prognostic evaluation and less than definitive primary therapy-adjusted for confounding by patient age, extent of disease, and comorbid diseases. RESULTS: During the first 5 years of follow-up, patients who received a less than definitive prognostic evaluation had an adjusted relative hazard of recurrence of 1.7 (95% confidence interval, 1.0-2.7) and an adjusted relative hazard for breast carcinoma mortality of 2.2 (95% confidence interval, 1.2-3.9). Patients who received less than definitive therapy had an adjusted relative hazard of recurrence of 1.6 (95% confidence interval, 1.0-2.5), and an adjusted relative hazard of breast carcinoma mortality of 1.7 (95% confidence interval, 1.0-2.8). CONCLUSIONS: Breast carcinoma patients who receive less than definitive care are at excess risk for disease recurrence and mortality. Women with early stage breast carcinoma should be treated in accordance with existing guidelines.
TUMOR ASSOCIATED ANTIGENS
Expression of MAGE tumour-associated antigens is inversely correlated with tumour differentiation in invasive ductal breast cancers: an immunohistochemical study. Kavalar R, Sarcevic B, Spagnoli GC, Separovic V, Samija M, Terracciano L, Heberer M, Juretic A. Maribor General Hospital, Slovenia.	Virchows Arch 2001 Aug;439(2):127-31 Abstract quote MAGE (Melanoma antigen E) family gene products encompass tumour-associated antigens (TAAs) recognised by human leukocyte antigen (HLA)-restricted specific T-cells. Agents inducing DNA demethylation, an event typically detectable in cellular de-differentiation processes, were shown to induce the expression of MAGE genes. By using a monoclonal antibody specific for MAGE family gene products, we have studied the expression of these TAAs in a group of 144 patients with invasive ductal breast cancers. Immunohistochemical data were correlated with tumour differentiation, lymphatic vessel invasion, oestrogen receptor expression, intratumoural necrosis, lymphocytic infiltration, perineural invasion, tumour microcalcifications and axillary lymph node metastases. MAGE immunoreactivity was undetectable in non-neoplastic cells. In poorly differentiated cancers positive staining was observed in 30/63 cases (47.6%) as compared with 13/51 (25.4%) and 5/30 (16.6%) in moderately and well-differentiated tumours, respectively (P<0.05). In addition, MAGE immunoreactivity was significantly correlated with lymphatic vessel invasion and intratumoural necrosis. Moreover, a significant inverse relationship with oestrogen receptor expression was also observed. However, no significant correlation could be established between MAGE immunoreactivity and defined phenotypic characteristics of tumour infiltrating lymphocytes, including expression of CD3, CD4, CD8, CD20 or granzyme B. Thus, expression of MAGE family gene products in invasive ductal breast cancers appears to be associated with poorly differentiated histological phenotypes. These data support the concept of specific immunotherapy in highly aggressive forms of breast neoplasms. Furthermore, they suggest that MAGE immunoreactivity could represent a tumour marker of potential prognostic relevance.
VASCULAR ENDOTHELIAL GROWTH FACTOR
Clinicopathological significance of vascular endothelial growth factor-C in breast carcinoma with long-term follow-up. Nakamura Y, Yasuoka H, Tsujimoto M, Yang Q, Tsukiyama A, Imabun S, Nakahara M, Nakao K, Nakamura M, Mori I, Kakudo K. Second Department of Pathology, Wakayama Medical University, Wakayama City (YN, HY, QY, MN, IM, KK).	Mod Pathol 2003 Apr;16(4):309-14 Abstract quote Expression of angiogenic and lymphangiogenic factors by tumors may influence the route of metastatic spread. The angiogenic factor vascular growth factor-C (VEGF-C) is implicated in the development of lymphatic vessels and promotion of lymphatic metastasis. The purpose of this study was to determine whether VEGF-C correlates with lymph node metastasis or prognosis. We assessed VEGF-C expression using immunohistochemistry in 123 invasive breast carcinomas with long-term follow-up. The relationship between VEGF-C expression and lymph node status and other established clinicopathological parameters was assessed. Whether VEGF-C expression plays a prognostic role in breast cancer was also investigated. VEGF-C expression was identified in 103 cases (83.7%). Positive VEGF-C was significantly correlated with lymph node metastasis (P = 0.0131). Survival curves determined by the Kaplan-Meier method and univariate analysis demonstrated that positive VEGF-C was associated with both disease-free survival (P = 0.0165) and overall survival (P = 0.0175). On the basis of our findings, VEGF-C plays a crucial role in lymph node metastasis and may be a significant prognostic factor for long-term survival in breast cancer.

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