P4-332

S614

Poster P4:: Wednesday Posters

whether anti-A␤1-11 polyclonal antibodies exhibit a protective effect on A␤42-induced neurotoxicity and reduce A␤ deposits in the brain of 3xTg-AD mice. Polyclonal as well as 6E10 monoclonal antibodies inhibit neurotoxicity induced by A␤ fibrils, but only the former is relatively potent in reducing toxicity induced by A␤ oligomers. Both antibodies significantly reduce the A␤ plaque burden in 3xTg-AD mice after a single intrahippocampal injection. Conclusions: Thus, anti-A␤1-11 antibodies are effective against all pathological forms of the A␤ peptide, including most toxic oligomers, which suggests the potential utility of our novel AD vaccine. P4-330

SUBSTRATE CONCENTRATION DEPENDENT INHIBITION OF GAMMA SECRETASE BY NOVEL INHIBITORS

Susanna S. Hemphill, Zhao Ren, Martin L. Neitzel, Jennifer L. Marugg, Jacek Jagodzinski, Lee H. Latimer, Michael A. Pleiss, Stephen B. Freedman, Sukanto Sinha, I. Paul Shapiro, Guriqbal S. Basi, Elan Pharmaceuticals, S. San Francisco, CA, USA. Contact e-mail: [email protected] Accumulation of A␤ peptides in the brain parenchyma and cerebral vasculature are pathologic features of Alzheimer’s disease (AD). A␤ peptides are produced from amyloid precursor protein (APP) by sequential proteolytic cleavages catalyzed by ␤- and ␥-secretases. Inhibition of these proteases is considered a leading strategy for therapeutic intervention in AD. A series of novel inhibitors have been identified (see accompanying poster by Neitzel et al) and characterized by dose-response analysis in cell-free and cellular assays and in vivo (see accompanying poster by Brigham et al.). We observed that the potency of these inhibitors varied in cells expressing different amounts of holoAPP. Experiments with cells expressing different levels of an APP C99 construct (with signal sequence) indicated that the cellular potency of these inhibitors was a function of the level of C99, the immediate ␥-secretase substrate. The dependence of inhibitory potency for these compounds on C99 levels was confirmed in cells expressing constant levels of holoAPP along with variable amounts of BACE1. These compounds were also shown to possess an uncompetitivemixed kinetic mode of inhibition in a cell-free ␥-secretase enzyme assay. As most gamma secretase inhibitors have been shown to exhibit noncompetitive kinetics in cell-free assays, this series of gamma secretase inhibitors and related inhibitors appear novel with regard to substrate concentration effects and kinetic mechanism of inhibition. P4-331

MELATONIN PARTIALLY REVERSES CALYCULIN A-INDUCED AXONAL TRANSPORT DEFECTS IN N2A CELLS

Xifei Yang1, Ying Yang1, Qing Tian1, Xiaochuan Wang1, Hong-Lian Li1, Qun Wang1, Jianzhi Wang1, Geng Li2, E.S. Yang2, 1 Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China; 2The Jockey Club MRI Centre, The University of Hong Kong, Hong Kong, Hong Kong. Contact e-mail: [email protected] Background: Abnormal neuronal aggregation of cytoplasm proteins and loss of the transport function as well as axonal atrophy are the histopathological characteristics of neurodegenerative diseases including Alzheimer’s disease (AD). Objectives: Investigate the effect of protein phosphatase-2A (PP-2A) and protein phosphatase-1 (PP-1) on axonal transport of neurofilament and search for a potential therapeutic agent. Methods: Cell culture, fluorescence labeling, total internal reflective fluorescence microscopy (TIRFM). Conclusions: (i) Suppression of PP-2A/PP-1 leading to axonal transport defects might be at least one of the underlying mechanisms for the abnormal aggregation of cytoplasm proteins in AD; (ii) Melatonin might have potential therapeutic benefit in AD by preventing the axonal transport impairment.

P4-332

OVEREXPRESSION OF CARBOXYL TERMINUS OF HSC70 INTERACTING PROTEIN PROMOTES TAU DEGRADATION IN VIVO

Yong-Jie Zhang, Ya-Fei Xu, Jian-Zhi Wang, Department of Pathophysiology, Key Laboratory of Neurological Disease of Hubei Province, Tongji Medical College of HUST, Wuhan, China. Contact e-mail: [email protected] Background: Neurofibrillary tangle (NFT) is one of the hallmark lesions of Alzheimer disease (AD) and the major protein component of NFT is the hyperphosphorylated tau. Recently, it has been reported that carboxyl terminus of Hsc70 interacting protein (CHIP) is the specific tau E3 ligase and can mediate tau degradation in vitro. However, the nature of CHIP on in vivo tau degradation is not known. Objective(s): In the present study, we injected CHIP plasmid into rat hippocampus bilaterally and studied the degradation of tau in rat brain. Methods: After the injection, the levels of mRNA and protein of CHIP were increased in a time-dependant manner in 24 h. Concomitantly, the levels of the phosphorylated tau at PHF-1 epitope, non-phosphorylated tau at tau-1 epitope and total tau probed by tau-5 were decreased significantly and this decrease was inversely correlated with the increase of CHIP. CHIP injection affected neither tau mRNA expression nor the proteasome activity. By co-immunoprecipitation, a complex of tau with Hsc70 and dimmeric CHIP was observed. These results indicate that CHIP, in collaborating with Hsc70, regulates tau degradation regardless of its phosphorylation state in normal rats. Then, we tested whether CHIP could degrade hyperphosphorylated tau induced by ventricular infusion of wortmannin and GFX. We found that the level of the hyperphosphorylated tau at PHF-1 epitope was decreased significantly upon CHIP overexpression. CHIP did not restore wortmannin/GFXinduced suppression of Ser9-phosphorylated GSK-3␤, infusion of wortmannin, GFX and CHIP did not affect tau mRNA and proteasome activity. These results indicate that overexpression of CHIP promotes degradation of hyperphosphorylated tau in the brain. Conclusions: Taken together, our findings suggest that CHIP is a critical mediator of tau degradation both in physiological and pathological conditions and overexpression of CHIP may be a potential therapeutic target for AD. P4-333

NAP REDUCES PHOSPHORYLATED TAU IN AD TRANSGENIC MICE

Yasuji Matsuoka1, Audrey J. Gray1, Hui-Fang Li1, Chiho Hirata1, Emmanuel Planel2, Karen Duff2, Mark P. Mattson3, Illana Gozes4, Paul S. Aisen1, 1Georgetown University Medical Center, Washington, DC, USA; 2Nathan Kline Institute, New York University, Orangeburg, NY, USA; 3National Institute on Aging, Baltimore, MD, USA; 4Tel Aviv University, Tel Aviv, Israel. Contact e-mail: [email protected] Background: The octapeptide NAP (NAPVSIPQ) is a highly active fragment of activity-dependent neuroprotective protein (ADNP), which is an essential for brain development. NAP shows neuroprotective effects in numerous in vitro and in vivo models of neurodegenerative disorders. When administered intranasally, the peptide enters the brain at therapeutic levels in mice and other experimental animals. Previous tissue culture studies have demonstrated that NAP binds tubulin, prevents zinc-induced microtubule rearrangement and transiently stabilizes microtubules. Tubulin interacts with tau in physiological environments, and tubulin-binding compounds may stabilize tau, reducing tau hyperphosphorylation and associated pathology. Recent in vitro studies have demonstrated that NAP may be associated with reduced tau hyperphosphorylation. NAP also inhibits Abeta aggregation in vitro. Objective(s): We aimed to determine whether NAP influences tau and Abeta pathology in a transgenic mouse model of AD in vivo. Methods: Triple transgenic mice expressing mutant amyloid precursor protein (APP, Swedish), tau (P301L) and presenilin-1 (M146V) at the early-to-moderate pathological stage were treated with NAP (intranasal, 2 ␮g/day, Monday through Friday) for 3 months. Mouse brains were isolated, and