There is a growing body of evidence that HER2 status can change during disease recurrence or progression in breast cancer patients. In this context, re-evaluation of HER2 status by assessment of HER2 expression on circulating tumor cells (CTCs) is a strategy with potential clinical application. The aim of this trial was to determine the HER2 status of CTCs in metastatic breast cancer patients comparing two CTC assays. A total of 254 patients with metastatic breast cancer from nine German university breast cancer centers were enrolled in this prospective study. HER2 status of CTCs was assessed using both the FDA-approved CellSearch® assay and AdnaTest BreastCancer™. Using the CellSearch assay, 122 of 245 (50%) patients had ≥5 CTCs, and HER2-positive CTCs were observed in 50 (41%) of these patients. Ninety of 229 (39%) patients were CTC positive using AdnaTest BreastCancer, and HER2 positivity rate was 47% (42 of 90). The rate of breast cancer patients with HER2-negative primary tumors but HER2-positive CTCs was 32% (25 of 78) and 49% (28 of 57) using the CellSearch assay and AdnaTest BreastCancer, respectively. Considering only those patients who had CTCs on both tests (n = 62), concordant results regarding HER2 positivity were obtained in 50% of the patients (31/62) (P = 0.96, κ = -0.006). HER2-positive CTCs can be detected in a relevant number of patients with HER2 negative primary tumors. Therefore, it will be mandatory to correlate the assay-dependent HER2 status of CTCs to the clinical response on HER2-targeted therapies.

Abstract-Excessive deformation of vascular smooth muscle cells (SMCs) caused by a prolonged increase in blood pressure (eg, in hypertension) results in an adaptive remodeling of the vessel wall that is characterized by SMC hypertrophy or hyperplasia and contributes to fixation of the increase in blood pressure. The onset of this process is characterized by a unique change in gene expression in the SMC. However, thus far, no transcription factor has been identified that specifically mediates mechanosensitive gene expression in these cells. Therefore, the role of a putative mechanotransducer, the cytoskeletal protein zyxin, was investigated in rat aortic cultured SMCs. Immunofluorescence and Western blot analysis revealed that on exposure to cyclic stretch, but not to osmotic stress or treatment with proinflammatory cytokines, zyxin dissociates from focal adhesions and accumulates in the nucleus. Unlike zyxin, vinculin, another focal adhesion-associated protein, did not translocate. Moreover, antisense oligonucleotide downregulation of zyxin protein abundance suggested that zyxin accumulation in the nucleus is a prerequisite for mechanosensitive gene expression in these cells. Thus, stretch-induced endothelin B receptor expression, for example, was attenuated, whereas that of tenascin-C was augmented after zyxin suppression. The data are consistent with a role for zyxin in transducing mechanical stimuli from the cell membrane to the nucleus in vascular SMCs and in controlling the expression of mechanosensitive genes that have been implicated in hypertension-induced arterial remodeling. y virtue of the resulting changes in circumferential wall stress and the degree of stretch that the cells are exposed to, blood pressure variations have a major impact on the phenotype of smooth muscle cells (SMCs) in conduit and resistance-size arteries. Although transient increases in blood pressure result in a compensatory increase in SMC tone, as exemplified by the myogenic response, pressure overload triggers a profound change in gene expression in these cells that ultimately leads to an adaptive remodeling of the vessel wall. 1 This is characterized by SMC hypertrophy (conduit arteries) or SMC hyperplasia (resistance-size arteries), resulting in an increase in wall thickness that offsets the blood pressure-induced rise in wall stress. However, this structural adaptation causes an increase in vascular resistance and, although aimed at regaining local blood flow control, eventually contributes to fixation of the elevated blood pressure as, eg, in hypertension. 2 Arterial hypertension is one of the primary risk factors for atherosclerosis, hence myocardial infarction, stroke, and dilated cardiomyopathy. Therefore, pressure-dependent arterial remodeling is a major contributor to the pathogenesis of these diseases. 3 Although the aforementioned interrelation is well established in the clinic, little is known about SMC mechanotransduction, namely the activation of transcription factors that specifically link deformation of the cell ...

Estrogen receptor (ER) b1 and its splice variants are expressed both in ovary and ovarian cancer. We studied the role of ERb1 and two of its splice variants in regulation of gene expression, cellular proliferation, apoptosis, and migration of an ovarian cancer cell line. In this study, we transfected SK-OV-3 ovarian cancer cells with vectors coding for ERb1 or its splice variants ERb-d125 and ERb-d1256, and tested their response to estrogen and tamoxifen in comparison with the untransfected cells. Heterologous expression of ERb1, but not of the exon-deleted ERb variants resulted in notably slower cell growth of SK-OV-3 ovarian cancer cells, an effect accompanied by more than tenfold increase of cyclindependent kinase inhibitor p21(WAF1) transcript levels and a significant reduction of cyclin A2 mRNA levels. SK-OV-3 cells stably overexpressing ERb1 ligand independently also exhibited an increased apoptosis rate and a significantly decreased motility, an effect accompanied by upregulation of fibulin 1c. Our data demonstrate that ERb1, but not the exon-deleted isoforms tested exerts multiple antitumoral effects on SK-OV-3 ovarian cancer cells even in the absence of estradiol or functional ERa.

Estrogen receptor b (ERb) is widely expressed in mammary epithelium. ERb expression is reported to decline during carcinogenesis of the breast and other tissues. In this study, we examined the consequences of a loss of ERb expression in mammary epithelial cells. We knocked down ERb transcript levels in human mammary epithelial MCF-10A cells and in MCF-7 breast cancer cells by means of stable transfection with a specific shRNA plasmid. ERb knockdown resulted in a significant growth increase of both cell types in a ligand-independent manner. This effect was accompanied by elevated cyclin A2 expression in MCF-10A cells and by decreased expression of growth-inhibitory p21/ WAF and epithelial cell marker cytokeratine 8 in both cell lines. Transfection of ERb shRNA did not alter the absent proliferative estrogen response of MCF-10A cells, but conferred sensitivity to selective estrogen receptor modulator tamoxifen to this cell line. In contrast, ERb knockdown diminished estrogen responsiveness of MCF-7 breast cancer cells and also weakened the effect of tamoxifen on this cell line. These ligand-dependent effects only observed in MCF-7 cells exhibiting a high ERa/b ratio were accompanied by smaller estrogenic repression of p21/WAF expression, an impaired tamoxifen-triggered induction of this gene and by relative downregulation of ERa and cyclin A2 transcript levels. Our data suggest that ERb exerts antiproliferative effects both on MCF-10A and MCF-7 cells in a ligand-and ERa-independent manner by regulation of p21/WAF or cyclin A2 gene expression. Knockdown of ERb in both cell types was sufficient to significantly decrease transcript levels of epithelial cell marker cytokeratin 8. The results of this study support the hypothesis that ERb acts as a tumor suppressor in mammary epithelium.