Neuronal activity regulates secretion of endogenous tau

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Oral Sessions: O2-01: Basic Science: Biology of Tau

of beta-amyloid, together with the alteration of key anatomical regions could partly explain the specificity of the networks targeted in AD. PLENARY SESSIONS: PL-02 PL-02-01

POLICY, SYSTEMS, CARE AND SUPPORT

Martin Knapp, London School of Economics, London, United Kingdom. Contact e-mail: [email protected] Background: Developments in public health, medicine and economics have combined to create the marvellous achievement of worldwide population aging. But associated with this achievement are a number of challenges, some of them exacerbated by macro-economic difficulties in many countries. In this presentation I will briefly summarise those challenges, most of them familiar to AAIC delegates. I will then turn to what we know about possible solutions. I will particularly want to draw on new evidence from economics research in the field to offer insights and suggest courses of action. Methods: The approach will be first to highlight recent and expected future trends as they relate to dementia; and the personal, familial, social and economic challenges which might result. The second step will be to consider what the response is - in terms of both policy and practice - might be to those challenges, drawing on recent evidence from robust research. Results: The pressures on personal, private and public resources are focusing the minds of decision-makers on how best to meet needs and respond to preferences in ways that are affordable and, in some sense, ‘efficient’. One important topic to be covered in the presentation, therefore, is to be clear as to who those ’decision-makers’ are, ranging from people using the services and their families, to service providers, funding agencies and national government bodies. It is also important to understand what criteria they use in their decision-making and how these can influence outcomes. The presentation will include (but could go beyond) discussion of such ’solutions’ as better targeting of home care, better recognition and response to comorbidities, personalisation (including devolved budgets), telehealth and telecare, better support for family and other care-givers, and the roles of ’community capital’. Conclusions: New research evidence in the dementia field offers promising opportunities. Public and private decision-makers can use this emerging evidence platform - some of it built on economics studies - to improve the lives of people with dementia and their family carers in ways that make better use of available better and public and private resources. PL-02-02

TAU-BASED THERAPEUTICS FOR ALZHEIMER’S DISEASE AND RELATED TAUOPATHIES

Michael Hutton, Eli Lilly United Kingdom, Windlesham, United Kingdom. Contact e-mail: [email protected] Background: Neurofibrillary pathology composed of the microtubule associated protein tau is a characteristic feature of Alzheimer’s Disease as well as multiple rare tauopathies such as FTLD-tau and Progressive Supranuclear Palsy. In each case, the density and distribution of the tau lesions correlates with the extent of neuronal loss and with the clinical progression of the disorder. Moreover multiple mutations in the tau (MAPT) gene have been shown to cause FTD linked to chromosome 17 establishing that tau dysfunction is sufficient to cause neurodegeneration. These mutations impact tau in different ways but significantly all are predicted to enhance tau aggregation suggesting that protein misfolding is a critical component of the neurodegenerative cascade. Results: As a result of these findings, considerable research effort has focused on determining the mechanism of neurofibrillary degeneration and on identifying potential therapies for the treatment of tauopathy. Kinase inhibitors that have been postulated to block tau hyperphosphorylation and toxicity, aggregation inhibitors designed to prevent the formation of filamentous inclusions, and various approaches to enhance the clearance of pathological tau have been proposed and have undergone varying degrees of preclinical development. More recently, therapeutic antibodies designed to block the transmission and spreading of tau pathology have also received considerable attention. However despite the significance of tau and the high level of research activity, only a handful of potential small molecule drugs have progressed into early clinical trials and to date none of these have produced robust evidence of clinical efficacy. Conclu-

sions: Several issues explain this relative lack of success, in particular there remains no consensus on the pathogenic mechanism that underlies taubased neurodegeneration and in addition there is currently a lack of validated, tractable targets for which translatable biomarkers exist to enable clinical development. Despite this there remains grounds for considerable optimism with the recognition that tau likely spreads through the brain in a prion-like manner and the development of the first tau PET imaging agents providing new opportunities for drug discovery. This presentation will therefore review the current state of tau therapeutic development and will discuss progress towards translating preclinical findings into effective treatments for patients with tauopathy. ORAL SESSIONS: O2-01 BASIC SCIENCE: BIOLOGY OF TAU O2-01-01

NEUROFIBRILLARY TANGLES REMAIN FUNCTIONALLY INTEGRATED IN CORTICAL NETWORKS

Kishore Kuchibhotla1, Susanne Wegmann2, Tara Spires-Jones2, Brian Bacskai2, Bradley Hyman2, 1New York University School of Medicine, New York, New York, United States; 2Harvard Medical School, Massachusetts General Hospital, Charlestown, Massachusetts, United States. Contact e-mail: [email protected] Background: The intracellular deposition of tau protein aggregates in neurofibrillary tangles (NFTs) and the formation of extracellular plaques containing beta-amyloid are hallmarks of Alzheimer’s disease (AD). Whereas beta-amyloid plaques are locally toxic and lead to significant disruptions of network function, the impact of NFTs on cellular and network function remains unclear. Recent work demonstrated that NFTs persist long after caspase activation, an event indicating the initiation of cell death. This suggested that neurons with NFTs can remain structurally integrated in a local neuronal network well after neurodegenerative processes have begun. Here we investigate whether neurons with NFTs can also retain their functional integrity in the local network. Methods: Using in vivo 2-photon calcium imaging in the visual cortex of awake mice we monitor the cell-specific responses of neurons to visual stimuli. Awake calcium imaging allows us to characterize neuronal tuning while limiting the impact of anesthesia on neuronal response dampening and tangle formation. Intracellular calcium transients are recorded through Yellow-Cameleon 3.6 expression induced by intracortical AAV injections in mice (rTg4510) that over-express mutant human tau (P301L) leading to cortical NFT deposition by the age of 7-8 months. Results: We observed that in adult rTg4510 mice with severe NFT deposition, neurons retain both their resting calcium levels as well as their tuning for the orientation of visual stimuli when compared to control mice. Conclusions: It appears that tangle-bearing neurons can remain functionally integrated in local cortical circuits, which questions the cell toxic aspect of tau aggregate deposition in AD and brings up questions about putative roles of tau aggregation e.g., for network plasticity deficits. O2-01-02

WITHDRAWN

O2-01-03

NEURONAL ACTIVITY REGULATES SECRETION OF ENDOGENOUS TAU

Amy Pooler, Emma Phillips, Dawn Lau, Wendy Noble, Diane Hanger, King’s College London, London, United Kingdom. Contact e-mail: amy. [email protected] Background: In Alzheimer’s disease, the cytoskeletal protein tau becomes hyperphosphorylated, and forms into aggregates and intracellular tangles. These neuropathological deposits of the cytoskeletal protein tau spread progressively from the entorhinal cortex to anatomically connected regions of the brain. Propagation of tau pathology is associated with progressive neurodegeneration, but the mechanism underlying trans-synaptic spread of tau has yet to be elucidated. Objective: To determine whether neurons release endogenous tau, and the mechanism by which this release may occur. Methods: We analyzed tau released from cultured rat cortical neurons using western blotting and ELISA. To explore mechanisms regulating tau release,

Oral Sessions: O2-02: Diagnosis and Prognosis: Amyloid Deposition and Normal Cognition we treated neurons with various compounds to stimulate or inhibit neuronal activity. Results: We show that stimulation of neuronal activity by KCl or glutamate, or specific AMPA receptor activation, induces tau release from mature cortical neurons. Further analysis of secreted tau revealed that phosphorylation of extracellular tau appears reduced in comparison to intracellular tau. We also find that AMPA-induced release of tau is calcium-dependent, and is mimicked by treatment of neurons with the ionophore ionomycin. Blocking pre-synaptic vesicle release by tetanus toxin, and inhibiting neuronal activity with the neurotoxin tetrodotoxin, both significantly impair AMPA-stimulated tau release. Analysis of neuronal exosomes revealed barely detectable levels of tau, indicating that tau secretion is unlikely to occur via exosome release. Conclusions: Endogenous release of tau from neurons is a regulatable process, dysregulation of which could lead to the spread of tau pathology in disease.

O2-01-04

INFLAMMATORY PATHWAYS LINK AMYLOID AND TAU PATHOLOGY

Bruce Lamb, Sungho Lee, Guixiang Xu, Richard Ransohoff, Cleveland Clinic, Cleveland, Ohio, United States. Contact e-mail: [email protected] Background: Alzheimer’s disease (AD) and non-AD tauopathies are a major cause of dementia, disability and death in the elderly. Neuropathological characteristics of AD include; intracellular aggregates of hyperphosphorylated microtubule associated protein tau (MAPT) in neurofibrillary tangles and extracellular deposits of the beta-amyloid peptide in senile plaques along with marked neuroinflammation, while non-AD tauopathies have both MAPT aggregates and neuroinflammation. While these pathological alterations are invariably observed within the diseased brain the exact relationship between neuroinflammatory processes, beta-amyloid, and MAPT pathologies remains unclear. Increasing evidence suggests that inflammatory processes may contribute to the pathopysiology of AD as well as non-AD tauopathies. First, there are increased numbers of activated astrocytes and microglia as well as increased expression of numerous pro-inflammatory molecules within the brain. Second, retrospective epidemiological studies demonstrated that long-term use of non-steroidal anti-inflammatory drugs substantially reduces AD risk. Third, recent studies utilizing genetically modified mouse models have implicated several different inflammatory pathways in AD-like pathologies. Methods: Neuronal-microglial communication through the chemokine fractalkine (CX3CL1) and its cognate receptor, CX3CR1, plays a critical role in neuroinflammation and neuroprotection. Notably, CX3CL1 is highly expressed by neurons while CX3CR1 is only expressed by microglia within the CNS. In the current studies, we provide evidence that membrane-tethered neuronal CX3CL1 signals to microglial CX3CR1 and in the process regulates both beta-amyloid and MAPT pathologies in opposing directions. Briefly, animals that were deficient for CX3CR1, CX3CL1 or the membrane anchored form of CX3CL1 were mated to the APPPS1 mouse model of beta-amyloid deposition and the htau mouse model of MAPT pathology and aged to determine the effects of the development of these two hallmark AD pathologies. Results: Our results demonstrate that loss of either total or membrane anchored CX3CL1 as well as CX3CR1 leads to reduced beta-amyloid pathology, but enhanced MAPT pathology. Current studies are focused on identifying alterations in microglia that could explain the potential disease mechanisms linking CX3CL1-CX3CR1 signaling to both beta-amyloid and MAPT pathologies. Conclusions: In summary, our results suggest that a specific inflammatory pathway between neurons and microglia links beta-amyloid and MAPT pathologies that could serve as a potential therapeutic target in future translational studies.

O2-01-05

THE RELATIONSHIP AMONG MTOR, BETAAMYLOID AND TAU: THERAPEUTIC IMPLICATIONS FOR ALZHEIMER’S DISEASE

Salvatore Oddo, University of Texas Health Science Center at San Antonio, San Antonio, Texas, United States. Contact e-mail: [email protected]

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Background: Accumulation of beta-amyloid and tau is an invariant feature of Alzheimer’s disease (AD). The upstream role of beta-amyloid accumulation in disease pathogenesis is widely accepted, and there is strong evidence showing that beta-amyloid accumulation causes cognitive impairments. However, the molecular mechanisms linking beta-amyloid to cognitive decline remain to be elucidated. Several lines of evidence indicate that deregulation in the balance between protein synthesis and degradation, which is crucial for maintaining neuronal function, may play a role in the disease pathogenesis. The mammalian target of rapamycin (mTOR) plays a key role in maintaining protein homeostasis as it regulates both protein degradation (via autophagy) and protein synthesis. Methods: Using several mouse models of AD and related disorders, we employ multidisciplinary approaches to dissect the molecular mechanisms linking beta-amyloid, tau and mTOR signaling. Results: We will show that the buildup of beta-amyloid increases the mammalian target of rapamycin (mTOR) signaling, while decreasing mTOR signaling reduces beta-amyloid levels, thereby highlighting an interrelation between mTOR signaling and beta-amyloid. Also, we will show that genetically increasing mTOR activity, significantly increase tau levels and phosphorylation. Further, using complementary animal models of AD, we will show that pharmacologically restoring mTOR signalling with rapamycin rescues cognitive deficits and ameliorates beta-amyloid and tau pathology by increasing autophagy. Conclusions: The results presented here provide a molecular basis for the beta-amyloid-induced cognitive deficits and show that mTOR dysregulation may be a valid terapeutic target for AD and related disorders. Indeed, we show that rapamycin, an FDA approved drug, improves learning and memory and reduces beta-amyloid and tau pathology.

O2-01-06

THE NUCLEOTIDE BINDING DOMAIN OF HEAT SHOCK COGNATE PROTEIN 70 (HSC70) REGULATES TAU CLEARANCE AND STABILITY

Sarah Fontaine1, Umesh Jinwal1, Andrea Thompson2, Jason Gestwicki2, Chad Dickey1, 1University of South Florida, Tampa, Florida, United States; 2 University of Michigan, Ann Arbor, Michigan, United States. Contact e-mail: [email protected] Background: Alzheimer’s disease (AD) is characterized by deposition of pathological protein inclusions of beta-amyloid (amyloid plaques) and tau (neurofibrillary tangles). The accumulation of the intrinsically disordered tau into conformations that lead intracellular tangles is linked to neuronal loss and cognitive dysfunction, which are hallmarks of AD. These structural changes in tau are mediated by Hsp70 and Hsp90 families chaperone proteins. We have previously demonstrated that Hsp70 proteins can interact with both normal and pathological tau and can clear tau via proteasome-mediated and autophagic mechanisms. In addition to degradation, Hsp70s can alter the stability and phosphorylation state of tau. To that end, we recently discovered that drugs targeting the nuclear binding domain (NBD) of the constitutively expressed heat shock cognate protein 70 (Hsc70) inhibited Hsc70 ATPase activity and potently reduced tau levels in a proteasome-dependent manner. To understand the mechanism of this drug, we introduced point mutations in this region to gain insight into how the nucleotide-bound status of Hsc70 impacted tau stability and clearance. Methods: Purified recombinant Hsc70 with or without point mutations in the NBD were subjected to ATPase and luciferase refolding assays. Hsc70-mediated effects on tau stability and clearance were analyzed in neuronal cells overexpressing wt or mutant Hsc70. Results: Point mutants of Hsc70 had distinct stimulatory or inhibitory effects on ATPase activity and luciferase refolding activity. In particular, one mutant inhibited both activities (E175S). Overexpression of the E175S mutant in neuronal cells resulted in a reduction in tau levels, decreased tau co-localization with tubulin and decreased microtubule stability. In contrast, the Hsc70 mutant F68L, which stimulated both ATPase activity and refolding activity, did not impact tau levels. E175S expression increased levels of polyubiquitnated tau, and inhibition of the proteasome restored tau levels in E175S-expressing cells to that of wt Hsc70, thereby suggesting E175S stimulates tau clearance via the proteasome. Conclusions: These data indicate regulation of the conformation of the NBD of Hsc70 may be a potential for therapeutic intervention for the treatment of AD.