Urokinase plasminogen activator receptor (uPAR) is a multidomain protein that plays important roles in the growth, invasion, and metastasis of a number of cancers. In the present study, we examined the effects of administration of a monoclonal anti-uPAR antibody (ATN-658) on prostate cancer progression in vitro and in vivo. We examined the effect of treatment of ATN-658 on human prostate cancer cell invasion, migration, proliferation, and regulation of intracellular signaling pathways. For in vivo studies, PC-3 cells (1 x 10(6)) were inoculated into the right flank of male Balb C nu/nu mice through subcutaneous or through intratibial route (2 x 10(5)) of male Fox Chase severe combined immunodeficient mice to monitor the effect on tumor growth and skeletal metastasis. Treatment with ATN-658 resulted in a significant dose-dependent decrease in PC-3 cell invasion and migration without affecting cell doubling time. Western blot analysis showed that ATN-658 treatment decreased the phosphorylation of serine/threonine protein kinase B (AKT), mitogen-activated protein kinase (MAPK), and focal adhesion kinase (FAK) without affecting AKT, MAPK, and FAK total protein expression. In in vivo studies, ATN-658 caused a significant decrease in tumor volume and a marked reduction in skeletal lesions as determined by Faxitron x-ray and micro-computed tomography. Immunohistochemical analysis of subcutaneous and tibial tumors showed a marked decrease in the levels of expression of pAKT, pMAPK, and pFAK, consistent with the in vitro observations. Results from these studies provide compelling evidence for the continued development of ATN-658 as a potential therapeutic agent for the treatment of prostate and other cancers expressing uPAR.

We report an autopsy case of tuberous sclerosis complex (TSC) in a 20-week gestational age female fetus. The brain showed lesions suggestive of early cortical tubers and subependymal hamartomatous nodules. The large cells within these nodular clusters were variably immunoreactive for glial fibrillary acidic protein (GFAP) and vimentin and negative for synaptophysin and neurofilament. Subependymal radial glia expressed both vimentin and GFAP, but subpial radial glia either did not express these markers (in contrast to an age-matched control) or were absent. Tuberin expression was noted in heterotopic neurons in the white matter and brain cells consistent with Cajal Retzius cells in the neocortical molecular layer, very weakly in superficial cortical neurons, neurons in the basal ganglia, Purkinje cells and external granular cells of cerebellum, cranial nerve nuclei neurons, occasional germinal matrix cells, ependymal cells, choroid plexus epithelium, and pituitary gland neuroendocrine cells; it was not seen within the cells of subependymal nodules. The pattern of tuberin immunoreactivity was similar to that which we have observed in older TSC patients. Proliferating cell labeling indexes were comparable in the germinal matrix of the TSC patient and an age-matched control. Abnormal subpial radial glia may be responsible for some of the neuronal migration abnormalities that appear to result in neocortical tubers.

Forty-six advanced-stage human breast carcinoma specimens were evaluated by immunohistochemistry for PKC alpha expression and compared with 25 samples of normal adjacent breast tissue. For normal tissue, the median staining of ductal epithelia was of moderate intensity. No staining was observed for 67% of tumor specimens, and only 4% showed intensities greater than the median observed in normal tissue. Faint to moderate PKC alpha staining was observed in the stroma, inflammatory cells, and fibroblasts of tumors but was absent in normal tissue. These findings demonstrate that downregulation of PKC alpha protein occurs in epithelial cells of advanced breast tumors (p<0.001).

Tuberous sclerosis (TSC) is a bigenic autosomal dominant disease caused by mutations in one of two tumor-suppressor genes, TSC1 and TSC2, resulting in benign hamartomas and low grade neoplasms in multiple organs including brain, heart, kidney, and skin. We report the results of an immunohistochemical study of the expression of the TSC gene products, tuberin and hamartin, in multiple tissues obtained at autopsy from 12 non-TSC affected patients ranging in age from 20 weeks gestation to 8 years, and surgical specimens from some organs. Tuberin and hamartin are expressed and are colocalized in most tissues. Contrary to a previous report, immunostaining with our antisera detected hamartin in liver, small and large intestine, prostate, and testes. We did not detect significant developmental differences in tuberin or hamartin expression in comparable tissues from patients of different ages. Although tuberin and hamartin colocalize in most tissues and cell types, we provide data that hamartin is more abundantly expressed than tuberin in cells within some tissues including the distal nephron and a population of cells of the endocrine pancreas. These data support the hypothesis that hamartin and tuberin interact and may function together in many tissues where they are co-expressed, but also suggest that hamartin has a discrete and specialized function in certain cell types.

Cerebral cortical dysplasia (CD) is a common cause of intractable childhood epilepsy. Five cases of CD were analyzed for GABA_A receptor subunit β (GABA_Aβ), glutamate decarboxylase, AMPA receptor subunit 1 (GluR1) and subunit 2/3 (GluR2/3), and NMDA receptor 2 (NMDAR2) immunoreactivity. Antisera to the highly polysialylated neural cell adhesion molecule (PSA-NCAM) and human unc-33-like phosphoprotein 1 (hUlip 1) were used to identify neurons with ‘developmentally immature’ characteristics. Differences between CD and comparison tissue (n = 3) included: (1) prominent GABA_Aβ immunoreactivity of the cytoplasm of dysmorphic neurons in the subcortical white matter and cortex in 1 CD case; (2) increased immunolabeling with anti-GluR1 and GluR2/3 antisera in dysmorphic neurons compared with more normal-appearing adjacent neurons and neurons from nondysplastic cortex; (3) varying numbers of cortical dysmorphic neurons stained for NMDAR2 in all 5 CD cases, in contrast to a complete lack of cellular immunoreactivity in 2/3 of the cases of nondysplastic cortex; (4) PSA-NCAM and hUlip 1 expression (usually observed only in populations of neurons that undergo axonal growth) was observed in CD tissue, but not in normal brain tissue. In summary, dysmorphic neurons in cases of CD have increased immunoreactivity for several excitatory neurotransmitter receptor subunits, show variable immunoreactivity for GABA_Aβ and show expression of several proteins that are normally expressed only in immature neurons or those with the potential for synaptic plasticity.