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Attractive Therapeutic Strategy (attractive + therapeutic_strategy)
Selected AbstractsRapid purification of active ,-secretase, an intramembrane protease implicated in Alzheimer's diseaseJOURNAL OF NEUROCHEMISTRY, Issue 1 2008Matthias Cacquevel Abstract ,-Secretase is an unconventional aspartyl protease that processes many type 1 membrane proteins within the lipid bilayer. Because its cleavage of amyloid-, precursor protein generates the amyloid-, protein (A,) of Alzheimer's disease, partially inhibiting ,-secretase is an attractive therapeutic strategy, but the structure of the protease remains poorly understood. We recently used electron microscopy and single particle image analysis on the purified enzyme to generate the first 3D reconstruction of ,-secretase, but at low resolution (15 Å). The limited amount of purified ,-secretase that can be produced using currently available cell lines and procedures has prevented the achievement of a high resolution crystal structure by X-ray crystallography or 2D crystallization. We report here the generation and characterization of a new mammalian cell line (S-20) that overexpresses strikingly high levels of all four ,-secretase components (presenilin, nicastrin, Aph-1 and Pen-2). We then used these cells to develop a rapid protocol for the high-grade purification of proteolytically active ,-secretase. The cells and purification methods detailed here provide a key step towards crystallographic studies of this ubiquitous enzyme. [source] Interaction of a ,-sheet breaker peptide with lipid membranesJOURNAL OF PEPTIDE SCIENCE, Issue 2 2010Giuseppe Vitiello Abstract Aggregation of ,-amyloid peptides into senile plaques has been identified as one of the hallmarks of Alzheimer's disease. An attractive therapeutic strategy for Alzheimer's disease is the inhibition of the soluble ,-amyloid aggregation using synthetic ,-sheet breaker peptides that are capable of binding A, but are unable to become part of a ,-sheet structure. As the early stages of the A, aggregation process are supposed to occur close to the neuronal membrane, it is strategic to define the ,-sheet breaker peptide positioning with respect to lipid bilayers. In this work, we have focused on the interaction between the ,-sheet breaker peptide acetyl-LPFFD-amide, iA,5p, and lipid membranes, studied by ESR spectroscopy, using either peptides alternatively labeled at the C- and at the N-terminus or phospholipids spin-labeled in different positions of the acyl chain. Our results show that iA,5p interacts directly with membranes formed by the zwitterionic phospholipid dioleoyl phosphatidylcholine and this interaction is modulated by inclusion of cholesterol in the lipid bilayer formulation, in terms of both peptide partition coefficient and the solubilization site. In particular, cholesterol decreases the peptide partition coefficient between the membrane and the aqueous medium. Moreover, in the absence of cholesterol, iA,5p is located between the outer part of the hydrophobic core and the external hydrophilic layer of the membrane, while in the presence of cholesterol it penetrates more deeply into the lipid bilayer. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd. [source] Selective gene transfer into neurons via Na,K-ATPase ,1.THE JOURNAL OF GENE MEDICINE, Issue 6 2008Targeting gene transfer with monoclonal antibody, adenovirus vector Abstract Background Neuron-selective gene transfer is an attractive therapeutic strategy for neurological disorders. However, optimal targets and gene delivery systems remain to be determined. Methods Following immunization of mice with PC12 cells, hybridomas were screened by ,-Gal reporter gene assay using FZ33 fiber-modified adenovirus vectors. Subsequently, the efficacy and specificity of monoclonal antibody (mAb)-mediated gene transfer via FZ33 and FdZ adenovirus vectors were evaluated by flow cytometry, chemiluminescent ,-Gal reporter gene assay, and immunocytochemistry. Finally, the antigen recognized by the mAb was identified by mass spectrometry and transfection analysis. Results A hybridoma clone 6E3 producing monoclonal antibody, mAb6E3, was screened. Flow cytometry, chemiluminescent ,-Gal reporter gene assay, and immunocytochemistry with mAb6E3 and the fiber mutant adenovirus demonstrated efficient gene transfer into the PC12 cells. Treatment of neuron,glia cocultures with mAb6E3 and FdZ adenovirus resulted in neuron-selective gene transfer. Immunohistochemical images of rat spinal cord tissue showed that mAb6E3 reacts specifically with neurons. Finally, Na,K-ATPase ,1 was identified as the antigen of mAb6E3. Conclusions Hybridoma screening using FZ33 fiber-modified adenovirus vectors serves as an efficient approach to detect antigens in mAb-targeted gene transfer. Neuronal tropism in the central nervous system through mAb6E3 represents an important initial step towards neuron-selective gene transfer in the treatment of local neurological disorders, such as spinal cord injury. Copyright © 2008 John Wiley & Sons, Ltd. [source] Orally available compound prevents deficits in memory caused by the Alzheimer amyloid-, oligomersANNALS OF NEUROLOGY, Issue 6 2006Matthew Townsend PhD Objective Despite progress in defining a pathogenic role for amyloid , protein (A,) in Alzheimer's disease, orally bioavailable compounds that prevent its effects on hippocampal synaptic plasticity and cognitive function have not yet emerged. A particularly attractive therapeutic strategy is to selectively neutralize small, soluble A, oligomers that have recently been shown to mediate synaptic dysfunction. Methods Using electrophysiological, biochemical, and behavioral assays, we studied how scyllo -inositol (AZD-103; molecular weight, 180) neutralizes the acutely toxic effects of A, on synaptic function and memory recall. Results Scyllo -inositol, but not its stereoisomer, chiro -inositol, dose-dependently rescued long-term potentiation in mouse hippocampus from the inhibitory effects of soluble oligomers of cell-derived human A,. Cerebroventricular injection into rats of the soluble A, oligomers interfered with learned performance on a complex lever-pressing task, but administration of scyllo -inositol via the drinking water fully prevented oligomer-induced errors. Interpretation A small, orally available natural product penetrates into the brain in vivo to rescue the memory impairment produced by soluble A, oligomers through a mechanism that restores hippocampal synaptic plasticity. Ann Neurol 2006;60:668,676 [source] | ||||||||||