Dovizio M.,University of Chieti Pescara |
Maier T.J.,Frankfurt University |
Alberti S.,University of Chieti Pescara |
Francesco L.D.,University of Chieti Pescara |
And 7 more authors.
Molecular Pharmacology | Year: 2013
Cyclooxygenase (COX)-2-derived prostanoids can influence several processes that are linked to carcinogenesis. We aimed to address the hypothesis that platelets contribute to aberrant COX-2 expression in HT29 colon carcinoma cells and to reveal the role of platelet-induced COX-2 on the expression of proteins involved in malignancy and marker genes of epithelialmesenchymal transition (EMT). Human platelets cocultured with HT29 cells rapidly adhered to cancer cells and induced COX-2 mRNA expression, but not protein synthesis, which required the late release of platelet-derived growth factor and COX-2 mRNA stabilization. Platelet-induced COX-2-dependent prostaglandin E2 (PGE2) synthesis in HT29 cells was involved in the downregulation of p21WAF1/CIP1 and the upregulation of cyclinB1 since these effects were prevented by rofecoxib (a selective COX-2 inhibitor) and rescued by exogenous PGE2. Galectin-3, which is highly expressed in HT29 cells, is unique among galectins because it contains a collagen-like domain. Thus, we studied the role of galectin-3 and platelet collagen receptors in plateletinduced COX-2 overexpression. Inhibitors of galectin-3 function (b-lactose, a dominant-negative form of galectin-3, Gal-3C, and anti-galectin-3 antibody M3/38) or collagen receptor-mediated platelet adhesion (revacept, a dimeric platelet collagen receptor GPVI-Fc) prevented aberrant COX-2 expression. Inhibition of platelet-cancer cell interaction by revacept was more effective than rofecoxib in preventing platelet-induced mRNA changes of EMT markers, suggesting that direct cell-cell contact and aberrant COX-2 expression synergistically induced gene expression modifications associated with EMT. In conclusion, our findings provide the rationale for testing blockers of collagen binding sites, such as revacept, and galectin-3 inhibitors in the prevention of colon cancer metastasis in animal models, followed by studies in patients. © 2013 by The American Society for Pharmacology.
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 255.20K | Year: 2007
DESCRIPTION (provided by applicant): The overall goal of the Phase I research is to develop a three-dimensional model of the blood-testis barrier (BTB) consisting of primary human Sertoli cells cultured on permeable membrane supports. The human Sertoli cells have been isolated from normal human testis tissue by MandalMed scientists using a novel patent-pending methodology. To our knowledge, due to difficulty in their isolation human Sertoli cells have never been readily available before for in vitro studies. Applications of the product include studies of the distribution and toxicology of drugs and industrial chemicals, infertility research, development of male contraceptive agents, and eventually for creation of a model of human spermatogenesis. There is a great need for more accurate, less costly and time- consuming methods of assessing effects on drug distribution in the testis, reproductive function and toxicity. Three Specific Aims have been identified for the Phase I research as described below. I. Develop a transwell three-dimensional culture system for primary human Sertoli cells as a model of the blood-testis barrier (BTB). II. Assess the ability of model compounds to affect permeability of the model BTB. III. Assess the toxicity of compounds for the model BTB. Areas of focus that are envisioned for Phase II are identification of genetic or proteomic markers for human Sertoli cell function, including the formation or disruption of junctions, transporters, immunomodulation, comparisons with animal studies of distribution of drugs for the testis, male reproductive toxicity, and the standardization as a method for quantifying human BTB permeability and Sertoli cell toxicity. In Phase II we will develop more complex, multicellular model systems. The overall goal of the Phase I research is to develop a three-dimensional model of the blood-testis barrier (BTB) consisting of primary human Sertoli cells cultured on permeable membrane supports. Applications of the product include studies of the distribution and toxicology of drugs and industrial chemicals, infertility research, development of male contraceptive agents, and eventually for creation of a model of human spermatogenesis. There is a great need for more accurate, less costly and time- consuming methods of assessing effects on reproductive function and toxicity.
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 155.69K | Year: 2002
DESCRIPTION (provided by applicant): The goal of the Phase I research is to develop ex vivo gene therapy that will achieve long-term, localized delivery of human neurotrophin-3 (hNT-3) in the rat contusion model of spinal cord injury. Our overall goal is to develop cell-based delivery of neurotrophins and other proteins as therapy for acute spinal cord injury. The following are the Phase I specific aims: Aim 1: To test functional recovery in response to NT-3 in rat spinal cord injury model. Aim 2: To test the effect of methylprednisotone on NT-3 based recovery of injured rats. In Phase II research we will develop a stereotaxic injection method to introduce cells into the injured spinal cords of rats, and study the effect of delaying implantation (24 to 48 hours post-injury) on cell survival, protein production, and functional recovery. Besides NT-3, we also will transduce cells with virus that produces brain-derived neurotrophic factor (BDNF), and look at its effect alone and in combination with NT-3 on functional recovery in animal models. Based on the results in Phase I we will continue working with methylprednisolone. We will test the system in a primate model, and develop human cells towards beginning Phase I clinical trials. PROPOSED COMMERCIAL APPLICATION: NOT AVAILABLE
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase II | Award Amount: 749.98K | Year: 2003
DESCRIPTION (provided by applicant): There are approximately 10,000 traumatic spinal cord injuries in the United States each year. The average age of injury is 31 years. The majority (90%) of individuals survive and live near-normal life spans. Cost of treating these injuries is very high and does not lead to complete recovery. Spinal cord injury can result in progressive axonal degeneration, variable amounts of neuronal loss, and functional impairment. Effective therapies have to be designed to not only minimize the loss of tissue due to secondary processes but also to promote axonal growth and functional regeneration. We are developing cell-based delivery of proteins (neurotrophic factors) that help in regeneration of neurons. Towards testing this therapy we propose the following specific aims: Specific Aim 1: To perform dose response studies with Sertoli cells. Our current data suggest that Sertoli cells alone had a significant effect on functional recovery and rate of locomotor recovery of spinal cord injured animals. This effect was observed at early time points and plateaued thereafter. One interpretation is that there simply are not enough cells to sustain the effect overtime. To address that possibility we will inject varying amounts of Sertoli cells into the acutely injured spinal cord. Sertoli cells will be implanted into white matter caudal to the site of injury, within 30 min of the injury, using a stereotaxic device. Lesion development will be monitored by in vivo magnetic resonance imaging (MRI). The intent is to correlate the temporal profile of lesion progression with locomotor recovery, as defined by a battery of neurologic tests that evaluate coordinated locomotion, hind limb paw position during locomotion, and performance on an inclined grid. To our knowledge this is the first study to use MRI to determine the extent to which lesion progression correlates with neurologic impairment. Animals will be euthanized at 42 days post injury and the spinal cords will be subject to histological assessment of white matter and quantification of NT-3. Specific Aim 2: To determine if delayed implantation of cells following injury results in improved cell survival and functional recovery. In the above studies we have implanted cells in the acutely traumatized spinal cord. The acutely injured spinal cord is characterized by prominent oxidative injury and pronounced proteolytic activity. Thus, survival of Sertoli cells may be challenged by these adverse conditions. We therefore propose to delay the time of cell implantation to 3 days and 7 days post injury. These time points coincide with wound healing including angiogenesis, events that may define an environment that is more receptive to implantation of cells. Lesion development will be monitored by in vivo magnetic resonance imaging (MRI) and correlated with motor function. Animals will be euthanized at 42 days post injury. The cords will be subjected to immuno- and histochemistry and assessment of NT-3 activity. Emphasis will be on quantifying the extent of residual white matter, glial scar formation, and assessment of the long descending serotonergic fiber tracts. Together, these anatomical and behavioral outcomes will determine the extent to which treatment alters wound healing and promotes plasticity and functional recovery. Specific Aim 3: To isolate and characterize human Sertoli cells for therapeutic development as delivery agents. Human Sertoli cells will be isolated from tissues, propagated in culture, and carefully characterized with regard to their expression of surface proteins including follicle stimulating hormone receptor (FSHr) and androgen binding protein (ABP). Gamma-glutamyl transpeptidase (GGT); a marker for Sertoli cells, we be quantified using a standard enzyme assay. Once characterized, cells will be infected with recombinant adenovirus expressing NT-3 and the levels of NT-3 production will be determined by ELISA. Mixed lymphocyte reactions will be performed in the presence of virally infected and uninfected human Sertoli cells, to determine their immunosuppressive ability.
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 134.00K | Year: 2005
DESCRIPTION (provided by applicant): Vascular disease is the principal cause of morbidity and mortality in patients with diabetes, leading to nephropathy, retinopathy, neuropathy, and ischemic disease. Hyperglycemic damage to vascular endothelial cells is a major cause of these vascular complications of diabetes. This endothelial dysfunction is caused by oxidative stress induced by hyperglycemia, resulting in both the increased production of reactive oxygen species (ROS) and decreased bioavailability of nitric oxide (NO). Hyperglycemia also causes similar oxidative stress in renal mesangial cells, which leads to mesangial matrix expansion, glomerosclerosis, and renal functional impairment. We have developed a novel drug for treatment of hyperglycemia induced endothelial and mesangial cell dysfunction. OX029 is a bifunctional NO-donor/SOD-mimetic which is a derivative of lipoic acid, The major goals of this proposal are to (1) determine the efficacy of OX029 to prevent endothelial dysfunction and renal injury in the diabetic streptozotocin-treated rat, and (2) design and synthesize 20 novel analogs of OX029 and test their efficacy in isolated blood vessels and cultured renal mesangial cells exposed to hyperglycemia. If OX029 is efficacious in preventing renal injury in STZ-diabetic rats, we plan to submit a Phase II grant to: (1) test one or two OX029 analogs for prevention of diabetic nephropathy in vivo, (2) expand the in vivo studies in STZ-diabetic rats to a long term study (6-9 months) to include other endpoints of renal pathology, such as histology and morphometry, glucose tolerance and insulin sensitivity, and renal function, and (3) expand the in vivo studies to include another animal model of diabetes, the db/db mouse which spontaneously develops type II diabetes. The long-term commercial goal is to develop an orally active drug which can be given to both type I and type II diabetics to attenuate the renal damage caused by chronic hyperglycemia, and also attenuate other vascular complications of diabetes.