A gene encoding a protein related to the serpin family of protease inhibitors was identified as a candidate tumor suppressor gene that may play a role in human breast cancer. The gene product, called maspin, is expressed in normal mammary epithelial cells but not in most mammary carcinoma cell lines. Transfection of MDA-MB-435 mammary carcinoma cells with the maspin gene did not alter the cells' growth properties in vitro, but reduced the cells' ability to induce tumors and metastasize in nude mice and to invade through a basement membrane matrix in vitro. Analysis of human breast cancer specimens revealed that loss of maspin expression occurred most frequently in advanced cancers. These results support the hypothesis that maspin functions as a tumor suppressor.

This revised staging system will be officially adopted for use in tumor registries in January 2003.

A B S T R A C T PurposeNo biomarkers have been identified to predict outcome with the use of an antiangiogenesis agent for cancer. Vascular endothelial growth factor (VEGF) genetic variability has been associated with altered risk of breast cancer and variable promoter activity. Therefore, we evaluated the association of VEGF genotype with efficacy and toxicity in E2100, a phase III study comparing paclitaxel versus paclitaxel plus bevacizumab as initial chemotherapy for metastatic breast cancer. Patients and MethodsDNA was extracted from tumor blocks of patients from E2100. Three hundred sixty-three samples were available to evaluate associations between genotype and outcome. Genotyping was performed for selected polymorphisms in VEGF and VEGF receptor 2. Testing for associations between each polymorphism and efficacy and toxicity was performed. ResultsThe VEGF-2578 AA genotype was associated with a superior median overall survival (OS) in the combination arm when compared with the alternate genotypes combined (hazard ratio ϭ 0.58; 95% CI, 0.36 to 0.93; P ϭ .023). The VEGF-1154 A allele also demonstrated a superior median OS with an additive effect of each active allele in the combination arm but not the control arm (hazard ratio ϭ 0.62; 95% CI, 0.46 to 0.83; P ϭ .001). Two additional genotypes, VEGF-634 CC and VEGF-1498 TT, were associated with significantly less grade 3 or 4 hypertension in the combination arm when compared with the alternate genotypes combined (P ϭ .005 and P ϭ .022, respectively). ConclusionOur data support an association between VEGF genotype and median OS as well as grade 3 or 4 hypertension when using bevacizumab in metastatic breast cancer.

There is a significant dose-response effect of adjuvant chemotherapy with cyclophosphamide, doxorubicin, and fluorouracil in patients with overexpression of c-erbB-2 but not in patients with no c-erbB-2 expression or minimal c-erbB-2 expression. Overexpression of c-erbB-2 may be a useful marker to identify the patients who are most likely to benefit from high doses of adjuvant chemotherapy.

Introduction: The androgen receptor (AR) is widely expressed in breast cancers and has been proposed as a therapeutic target in estrogen receptor alpha (ER) negative breast cancers that retain AR. However, controversy exists regarding the role of AR, particularly in ER + tumors. Enzalutamide, an AR inhibitor that impairs nuclear localization of AR, was used to elucidate the role of AR in preclinical models of ER positive and negative breast cancer.

The anti-diabetic drug metformin reduces human cancer incidence and improves the survival of cancer patients, including those with breast cancer. We studied the activity of metformin against diverse molecular subtypes of breast cancer cell lines in vitro. Metformin showed biological activity against all estrogen receptor (ER) positive and negative, erbB2 normal and abnormal breast cancer cell lines tested. It inhibited cellular proliferation, reduced colony formation and caused partial cell cycle arrest at the G(1) checkpoint. Metformin did not induce apoptosis (as measured by DNA fragmentation and PARP cleavage) in luminal A, B or erbB2 subtype breast cancer cell lines. At the molecular level, metformin treatment was associated with a reduction of cyclin D1 and E2F1 expression with no changes in p27(kip1) or p21(waf1). It inhibited mitogen activated protein kinase (MAPK) and Akt activity, as well as the mammalian target of rapamycin (mTOR) in both ER positive and negative, erbB2-overexpressing and erbB2-normal expressing breast cancer cells. In erbB2-overexpressing breast cancer cell lines, metformin reduced erbB2 expression at higher concentrations, and at lower concentrations within the therapeutic range, it inhibited erbB2 tyrosine kinase activity evidenced by a reduction of phosphorylated erbB2 (P-erbB2) at both auto- and Src- phosphorylation sites. These data suggest that metformin may have potential therapeutic utility against ER positive and negative, erbB2-overexpressing and erbB2-normal expressing breast cancer cells.

Loss of heterozygosity (LOH) was detected in morphologically normal lobules adjacent to breast cancers. The most frequent aberration was at chromosome 3p22-25; of ten cases with this LOH in the carcinoma, six displayed the same LOH in adjacent normal lobules. This suggests that in a subset of sporadic breast cancers, a tumor suppresser gene at 3p22-25 may be important in initiation or early progression of tumorigenesis. Among sixteen breast cancers with LOH at 17p13.1 and five breast cancers with LOH at 11p15.5, one case each displayed the same LOH in adjacent normal lobules. Thus the molecular heterogeneity that characterizes invasive breast cancers may occur at the earliest detectable stages of progression.The mature breast contains lobules, clusters of closed glandular spaces that produce milk during lactation. These lobules are connected to the nipple-areolar complex by a system of branching ducts that are surrounded by varying amounts of fat and connective tissue. Breast cancer is thought to develop within a terminal ductal-lobular unit (TDLU), which includes the lobule and its most proximal ducts (1).Breast cancer evolves by clonal selection of cells that acquire multiple molecular changes. One model suggests that breast cancer, like colon cancer (2), develops through a defined progression of morphologically distinguishable stages beginning with benign hyperplasia, which progresses to atypical hyperplasia, then to in situ carcinoma, and finally to invasive cancer (1). This sequential progression may not be the only way that breast cancers develop, however. Many small invasive cancers do not have atypical components, which suggests that they may have developed directly from morphologically normal epithelium. If this were true, one might expect to find evidence of a "field effect" in which at least some of the genetic aberrations found in invasive cancers are also present in the morphologically normal epithelium.To test this hypothesis, we carefully microdissected hematoxylin-eosin-stained sections of breast cancers so as to isolate morphologically discrete regions (Fig. 1A). DNA was prepared from malignant areas of the section and from adjacent normal TDLUs. As a control for each case, DNA was also prepared from normal breast skin (usually from a separate section) that had been similarly microdissected.We studied LOH at chromosome 3p24, 11p15.5, 13q13, and 17p13.1 because these loci show LOH in a high percentage (ϳ30 to 60%) of invasive ductal breast cancers (3, 4). For the carcinomatous regions, the frequency of LOH at 3p24 (48%) and 11p15.5 (29%) was similar to that previously reported (4). The frequency of LOH in the invasive components was higher than the literature values for 13q13 (64% here versus ϳ40%) and for 17p13.1 (80% here versus ϳ60%). These discrepancies may be due to random variation because our sample size was small.In 8 of 30 cases we detected LOH in the adjacent morphologically normal TDLUs (Table 1). In all eight cases, the same allele was missing in the adjacent carcinoma (Fig. 2, A an...

Triple negative (TN) breast cancer is more frequent in women who are obese or have type II diabetes, as well as young women of color. These cancers do not express receptors for the steroid hormones estrogen or progesterone, or the type II receptor tyrosine kinase (RTK) Her-2 but do have upregulation of basal cytokeratins and the epidermal growth factor receptor (EGFR). These data suggest that aberrations of glucose and fatty acid metabolism, signaling through EGFR and genetic factors may promote the development of TN cancers. The anti-type II diabetes drug metformin has been associated with a decreased incidence of breast cancer, although the specific molecular subtypes that may be reduced by metformin have not been reported. Our data indicates that metformin has unique anti-TN breast cancer effects both in vitro and in vivo. It inhibits cell proliferation (with partial S phase arrest), colony formation and induces apoptosis via activation of the intrinsic and extrinsic signaling pathways only in TN breast cancer cell lines. At the molecular level, metformin increases P-AMPK, reduces P-EGFR, EGFR, P-MAPK, P-Src, cyclin D1 and cyclin E (but not cyclin A or B, p27 or p21), and induces PARP cleavage in a dose- and time-dependent manner. These data are in stark contrast to our previously published biological and molecular effects of metformin on luminal A and B, or Her-2 type breast cancer cells. Nude mice bearing tumor xenografts of the TN line MDA-MB-231, treated with metformin, show significant reductions in tumor growth (p = 0.0066) and cell proliferation (p = 0.0021) as compared to untreated controls. Metformin pre-treatment, before injection of MDA-MB-231 cells, results in a significant decrease in tumor outgrowth and incidence. Given the unique anti-cancer activity of metformin against TN disease, both in vitro and in vivo, it should be explored as a therapeutic agent against this aggressive form of breast cancer.