The development of acquired resistance to antihormonal agents in breast cancer is a major therapeutic problem. We have developed a tamoxifen-resistant (TAM-R) MCF-7 breast cancer cell line to investigate the mechanisms behind this condition. Both epidermal growth factor receptor (EGFR) and c-erbB2 mRNA and protein expression were increased in TAM-R compared with wild-type MCF-7 cells, whereas comparable levels of c-erbB3 mRNA and protein were expressed in both cell lines. Under basal conditions, phosphorylated EGFR/c-erbB2, EGFR/c-erbB3 but not c-erbB2/c-erbB3 receptor heterodimers were detected in TAM-R cells in association with increased levels of phosphorylated extracellular-signal regulated kinase 1/2 (ERK1/2). Both cell lines were capable of generating a range of EGFR-specific ligands and increased expression of transforming growth factor alpha was observed in TAM-R cells. Treatment of TAM-R cells with ZD1839 (Iressa) or trastuzumab (Herceptin) blocked c-erbB receptor heterodimer formation and phosphorylation, reduced ERK1/2 activity, and strongly inhibited cell growth. The MAPK kinase inhibitor PD098059 specifically reduced phosphorylated ERK1/2 levels and inhibited TAM-R growth. All three agents abolished ERK1/2 activity in wild-type cells but caused only small reductions in cell proliferation. These results demonstrate that TAM-R MCF-7 cell growth is mediated by the autocrine release and action of an EGFR-specific ligand inducing preferential EGFR/c-erbB2 dimerization and downstream activation of the ERK pathway.

The function of the massive feedback projection from visual cortex to its thalamic relay nucleus has so far eluded any clear overview. This feedback exerts a range of effects, including an increase in the inhibition elicited by moving contours, but the functional logic of the direct connections to the thalamic cells that relay the retinal input to the cortex remains largely unknown. In contrast to its thalamic nucleus, the visual cortex is characterized by cells that are strongly sensitive to the orientation of moving contours. Here we report that when driven by moving oriented visual stimuli the cortical feedback induces correlated firing in relay cells. This cortically induced correlation of relay cell activity produces coherent firing in those groups of relay cells with receptive field alignments appropriate to signalling the particular orientation of the moving contour to the cortex. Synchronization of relay cell firing means that they will elicit temporally overlapping excitatory postsynaptic potentials in their cortical target cells, thus increasing the chance that the cortical cells will fire. Effectively this increases the gain of the input for feature-linked events detected by the cortex. We propose that this feedback loop serves to lock or focus the appropriate circuitry onto the stimulus feature.

Apoptotic nerve cell death is implicated in the pathogenesis of several devastating neurodegenerative conditions, including glaucoma and Alzheimer's and Parkinson's diseases. We have devised a noninvasive real-time imaging technique using confocal laserscanning ophthalmoscopy to visualize single nerve cell apoptosis in vivo, which allows longitudinal study of disease processes that has not previously been possible. Our method utilizes the unique optical properties of the eye, which allow direct microscopic observation of nerve cells in the retina. We have been able to image changes occurring in nerve cell apoptosis over hours, days, and months and show that effects depend on the magnitude of the initial apoptotic inducer in several models of neurodegenerative disease in rat and primate. This technology enables the direct observation of single nerve cell apoptosis in experimental neurodegeneration, providing the opportunity for detailed investigation of fundamental disease mechanisms and the evaluation of interventions with potential clinical applications, together with the possibility of taking this method through to patients.A poptosis is an orchestrated form of cell ''death by suicide.'' It is essential in both the development and normal maintenance of tissue function. It is also implicated in the pathology of a number of severe neurodegenerative disorders such as glaucoma, motor neuron, and Alzheimer's, Parkinson's, and Huntington's diseases (1). There is evidence that a similar pathogenesis may contribute to neurodegenerative processes in these conditions (2), and that the extent of nerve cell loss is correlated with functional deficit (3-6). If we could directly visualize this process, it would facilitate a much more precise diagnosis and critically enable accurate tracking of the disease state and the action of therapy. However, until now, it has not been possible to detect nerve cell apoptosis in vivo (1, 7-10).Annexin 5 is a protein that, in the presence of Ca 2ϩ , has a high affinity for phosphatidylserine (PS), an anionic phospholipid that is enriched in the inner leaflet of plasma membranes. The externalization of PS from the inner leaflet to the outer layer of the cell membrane is an invariant early feature in the apoptotic process that occurs before DNA fragmentation and nuclear condensation. Because of its properties, FITC-annexin 5 has become widely used in the cytological detection of cells undergoing apoptosis (11). More recently, annexin 5 has been shown to be effective in the identification of apoptosis in vivo by using radiological and macroscopic fluorescent techniques (7,8,10,12,13).However, existing in vivo techniques using annexin 5 either have been unable to resolve the process to a single cellular level (7-9) or require an invasive method performed under terminal anesthesia (10). Imaging the eye, compared with the rest of the body, offers a unique opportunity because of the presence of clear optical media allowing direct visualization of labeled disease processes as they occur. This mea...

Drugs that target the insulin-like growth factor-I receptor (IGF-IR) and/or insulin receptor (IR) are currently under investigation for a variety of malignancies including breast cancer. Although we have previously reported that IGF-IR expression in primary breast tumors is common, the activation status of this receptor has not been examined in relation to survival. Phosphorylated IGF-IR/IR (P-IGF-IR/IR) and its downstream signaling partner phospho-S6 (P-S6) were evaluated immunohistochemically in tumor tissue microarrays representing 438 cases of invasive breast cancer. P-IGF-IR/IR (n = 114; P = 0.046) and total levels of IR (n = 122; P = 0.009) were indicative of poor survival, whereas total IGF-IR (n = 112; P = 0.304) was not. P-IGF-IR/IR and P-S6 were coordinately expressed in primary breast tumors (likelihood ratio, 11.57; P = 6.70 Â 10 À4