Phase 1 Trial of Antibody NI006 for Depletion of Cardiac Transthyretin Amyloid.

BACKGROUND: Transthyretin amyloid (ATTR) cardiomyopathy is a progressive and fatal disease caused by misfolded transthyretin. Despite advances in slowing disease progression, there is no available treatment that depletes ATTR from the heart for the amelioration of cardiac dysfunction. NI006 is a recombinant human anti-ATTR antibody that was developed for the removal of ATTR by phagocytic immune cells. METHODS: In this phase 1, double-blind trial, we randomly assigned (in a 2:1 ratio) 40 patients with wild-type or variant ATTR cardiomyopathy and chronic heart failure to receive intravenous infusions of either NI006 or placebo every 4 weeks for 4 months. Patients were sequentially enrolled in six cohorts that received ascending doses (ranging from 0.3 to 60 mg per kilogram of body weight). After four infusions, patients were enrolled in an open-label extension phase in which they received eight infusions of NI006 with stepwise increases in the dose. The safety and pharmacokinetic profiles of NI006 were assessed, and cardiac imaging studies were performed. RESULTS: The use of NI006 was associated with no apparent drug-related serious adverse events. The pharmacokinetic profile of NI006 was consistent with that of an IgG antibody, and no antidrug antibodies were detected. At doses of at least 10 mg per kilogram, cardiac tracer uptake on scintigraphy and extracellular volume on cardiac magnetic resonance imaging, both of which are imaging-based surrogate markers of cardiac amyloid load, appeared to be reduced over a period of 12 months. The median N-terminal pro-B-type natriuretic peptide and troponin T levels also seemed to be reduced. CONCLUSIONS: In this phase 1 trial of the recombinant human antibody NI006 for the treatment of patients with ATTR cardiomyopathy and heart failure, the use of NI006 was associated with no apparent drug-related serious adverse events. (Funded by Neurimmune; NI006-101 ClinicalTrials.gov number, NCT04360434.).

Comprehensive evaluation of human-derived anti-poly-GA antibodies in cellular and animal models of C9orf72 disease.

Hexanucleotide G4C2 repeat expansions in the C9orf72 gene are the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. Dipeptide repeat proteins (DPRs) generated by translation of repeat-containing RNAs show toxic effects in vivo as well as in vitro and are key targets for therapeutic intervention. We generated human antibodies that bind DPRs with high affinity and specificity. Anti-GA antibodies engaged extra- and intra-cellular poly-GA and reduced aggregate formation in a poly-GA overexpressing human cell line. However, antibody treatment in human neuronal cultures synthesizing exogenous poly-GA resulted in the formation of large extracellular immune complexes and did not affect accumulation of intracellular poly-GA aggregates. Treatment with antibodies was also shown to directly alter the morphological and biochemical properties of poly-GA and to shift poly-GA/antibody complexes to more rapidly sedimenting ones. These alterations were not observed with poly-GP and have important implications for accurate measurement of poly-GA levels including the need to evaluate all centrifugation fractions and disrupt the interaction between treatment antibodies and poly-GA by denaturation. Targeting poly-GA and poly-GP in two mouse models expressing G4C2 repeats by systemic antibody delivery for up to 16 mo was well-tolerated and led to measurable brain penetration of antibodies. Long-term treatment with anti-GA antibodies produced improvement in an open-field movement test in aged C9orf72450 mice. However, chronic administration of anti-GA antibodies in AAV-(G4C2)149 mice was associated with increased levels of poly-GA detected by immunoassay and did not significantly reduce poly-GA aggregates or alleviate disease progression in this model.

Efficient characterization of multiple binding sites of small molecule imaging ligands on amyloid-beta, tau and alpha-synuclein.

PURPOSE: There is an unmet need for compounds to detect fibrillar forms of alpha-synuclein (αSyn) and 4-repeat tau, which are critical in many neurodegenerative diseases. Here, we aim to develop an efficient surface plasmon resonance (SPR)-based assay to facilitate the characterization of small molecules that can bind these fibrils. METHODS: SPR measurements were conducted to characterize the binding properties of fluorescent ligands/compounds toward recombinant amyloid-beta (Aß)42, K18-tau, full-length 2N4R-tau and αSyn fibrils. In silico modeling was performed to examine the binding pockets of ligands on αSyn fibrils. Immunofluorescence staining of postmortem brain tissue slices from Parkinson's disease patients and mouse models was performed with fluorescence ligands and specific antibodies. RESULTS: We optimized the protocol for the immobilization of Aß42, K18-tau, full-length 2N4R-tau and αSyn fibrils in a controlled aggregation state on SPR-sensor chips and for assessing their binding to ligands. The SPR results from the analysis of binding kinetics suggested the presence of at least two binding sites for all fibrils, including luminescent conjugated oligothiophenes, benzothiazole derivatives, nonfluorescent methylene blue and lansoprazole. In silico modeling studies for αSyn (6H6B) revealed four binding sites with a preference for one site on the surface. Immunofluorescence staining validated the detection of pS129-αSyn positivity in the brains of Parkinson's disease patients and αSyn preformed-fibril injected mice, 6E10-positive Aß in arcAß mice, and AT-8/AT-100-positivity in pR5 mice. CONCLUSION: SPR measurements of small molecules binding to Aß42, K18/full-length 2N4R-tau and αSyn fibrils suggested the existence of multiple binding sites. This approach may provide efficient characterization of compounds for neurodegenerative disease-relevant proteinopathies.

Results from the long-term extension of PRIME: A randomized Phase 1b trial of aducanumab.

INTRODUCTION: Aducanumab selectively targets aggregated forms of amyloid beta (Aß), a neuropathological hallmark of Alzheimer's disease (AD). METHODS: PRIME was a Phase 1b, double-blind, randomized clinical trial of aducanumab. During the 12-month placebo-controlled period, participants with prodromal AD or mild AD dementia were randomized to receive aducanumab or placebo. At week 56, participants could enroll in a long-term extension (LTE), in which all participants received aducanumab. The primary endpoint was safety and tolerability. RESULTS: Amyloid-related imaging abnormalities-edema (ARIA-E) were the most common adverse event. Dose titration was associated with a decrease in the incidence of ARIA-E. Over 48 months, aducanumab decreased brain amyloid levels in a dose- and time-dependent manner. Exploratory endpoints suggested a continued benefit in the reduction of clinical decline over 48 months. DISCUSSION: The safety profile of aducanumab remained unchanged in the LTE of PRIME. Amyloid plaque levels continued to decrease in participants treated with aducanumab. HIGHLIGHTS: PRIME was a Phase 1b, double-blind, randomized clinical trial of aducanumab. We report cumulative safety and 48-month efficacy results from PRIME. Amyloid-related imaging abnormalities-edema (ARIA-E) were the most common adverse event (AE); 61% of participants with ARIA-E were asymptomatic. Dose titration was associated with a decrease in the incidence of ARIA-E. Aducanumab decreased levels of amyloid beta (Aß) in a dose- and time-dependent manner.

Lifestyle Affects Amyloid Burden and Cognition Differently in Men and Women.

OBJECTIVE: Evidence on associations of lifestyle factors with Alzheimer's pathology and cognition are ambiguous, potentially because they rarely addressed inter-relationships of factors and sex effects. While considering these aspects, we examined the relationships of lifestyle factors with brain amyloid burden and cognition. METHODS: We studied 178 cognitively normal individuals (women, 49%; 65.0 [7.6] years) and 54 individuals with mild cognitive impairment (women, 35%; 71.3 [8.3] years) enrolled in a prospective study of volunteers who completed 18 F-Flutemetamol amyloid positron emission tomography. Using structural equation modeling, we examined associations between latent constructs representing metabolic/vascular risk, physical activity, and cognitive activity with global amyloid burden and cognitive performance. Furthermore, we investigated the influence of sex in this model. RESULTS: Overall, higher cognitive activity was associated with better cognitive performance and higher physical activity was associated with lower amyloid burden. The latter association was weakened to a nonsignificant level after excluding multivariate outliers. Examination of the moderating effect of sex in the model revealed an inverse association of metabolic/vascular risk with cognition in men, whereas in women metabolic/vascular risk trended toward increased amyloid burden. Furthermore, a significant inverse association between physical activity and amyloid burden was found only in men. Inheritance of an APOE4 allele was associated with higher amyloid burden only in women. INTERPRETATION: Sex modifies effects of certain lifestyle-related factors on amyloid burden and cognition. Notably, our results suggest that the negative impact of metabolic/vascular risk influences the risk of cognitive decline and Alzheimer's disease through distinct paths in women and men. ANN NEUROL 2022;92:451-463.

Coregistered transcranial optoacoustic and magnetic resonance angiography of the human brain.

Imaging modalities capable of visualizing the human brain have led to major advances in neurology and brain research. Multi-spectral optoacoustic tomography (MSOT) has gained importance for studying cerebral function in rodent models due to its unique capability to map changes in multiple hemodynamic parameters and to directly visualize neural activity within the brain. The technique further provides molecular imaging capabilities that can facilitate early disease diagnosis and treatment monitoring. However, transcranial imaging of the human brain is hampered by acoustic attenuation and other distortions introduced by the skull. Here, we demonstrate non-invasive transcranial MSOT angiography of pial veins through the temporal bone of an adult healthy volunteer. Time-of-flight (TOF) magnetic resonance angiography (MRA) and T1-weighted structural magnetic resonance imaging (MRI) were further acquired to facilitate anatomical registration and interpretation. The superior middle cerebral vein in the temporal cortex was identified in the MSOT images, matching its location observed in the TOF-MRA images. These initial results pave the way toward the application of MSOT in clinical brain imaging.

Addendum: The antibody aducanumab reduces Aß plaques in Alzheimer's disease.

This corrects the article DOI: 10.1038/nature21361.

Early ß-amyloid accumulation in the brain is associated with peripheral T cell alterations.

INTRODUCTION: Fast and minimally invasive approaches for early diagnosis of Alzheimer's disease (AD) are highly anticipated. Evidence of adaptive immune cells responding to cerebral ß-amyloidosis has raised the question of whether immune markers could be used as proxies for ß-amyloid accumulation in the brain. METHODS: Here, we apply multidimensional mass-cytometry combined with unbiased machine-learning techniques to immunophenotype peripheral blood mononuclear cells from a total of 251 participants in cross-sectional and longitudinal studies. RESULTS: We show that increases in antigen-experienced adaptive immune cells in the blood, particularly CD45RA-reactivated T effector memory (TEMRA) cells, are associated with early accumulation of brain ß-amyloid and with changes in plasma AD biomarkers in still cognitively healthy subjects. DISCUSSION: Our results suggest that preclinical AD pathology is linked to systemic alterations of the adaptive immune system. These immunophenotype changes may help identify and develop novel diagnostic tools for early AD assessment and better understand clinical outcomes.

Fe65 is the sole member of its family that mediates transcription regulated by the amyloid precursor protein.

The amyloid precursor protein (APP), a central molecule in Alzheimer's disease (AD), has physiological roles in cell adhesion and signaling, migration, neurite outgrowth and synaptogenesis. Intracellular adapter proteins mediate the function of transmembrane proteins. Fe65 (also known as APBB1) is a major APP-binding protein. Regulated intramembrane proteolysis (RIP) by γ-secretase releases the APP intracellular domain (AICD), together with the interacting proteins, from the membrane. We studied the impact of the Fe65 family (Fe65, and its homologs Fe65L1 and Fe65L2, also known as APBB2 and APBB3, respectively) on the nuclear signaling function of the AICD. All Fe65 family members increased amyloidogenic processing of APP, generating higher levels of ß-cleaved APP stubs and AICD. However, Fe65 was the only family member supporting AICD translocation to nuclear spots and its transcriptional activity. Using a recently established transcription assay, we dissected the transcriptional activity of Fe65 and provide strong evidence that Fe65 represents a transcription factor. We show that Fe65 relies on the lysine acetyltransferase Tip60 (also known as KAT5) for nuclear translocation. Furthermore, inhibition of APP cleavage reduces nuclear Tip60 levels, but this does not occur in Fe65-knockout cells. The rate of APP cleavage therefore regulates the nuclear translocation of AICD-Fe65-Tip60 (AFT) complexes, to promote transcription by Fe65.

Xeno-free induced pluripotent stem cell-derived neural progenitor cells for in vivo applications.

BACKGROUND: Currently, there is no regenerative therapy for patients with neurological and neurodegenerative disorders. Cell-therapies have emerged as a potential treatment for numerous brain diseases. Despite recent advances in stem cell technology, major concerns have been raised regarding the feasibility and safety of cell therapies for clinical applications. METHODS: We generated good manufacturing practice (GMP)-compatible neural progenitor cells (NPCs) from transgene- and xeno-free induced pluripotent stem cells (iPSCs) that can be smoothly adapted for clinical applications. NPCs were characterized in vitro for their differentiation potential and in vivo after transplantation into wild type as well as genetically immunosuppressed mice. RESULTS: Generated NPCs had a stable gene-expression over at least 15 passages and could be scaled for up to 1018 cells per initially seeded 106 cells. After withdrawal of growth factors in vitro, cells adapted a neural fate and mainly differentiated into active neurons. To ensure a pure NPC population for in vivo applications, we reduced the risk of iPSC contamination by applying micro RNA-switch technology as a safety checkpoint. Using lentiviral transduction with a fluorescent and bioluminescent dual-reporter construct, combined with non-invasive in vivo bioluminescent imaging, we longitudinally tracked the grafted cells in healthy wild-type and genetically immunosuppressed mice as well as in a mouse model of ischemic stroke. Long term in-depth characterization revealed that transplanted NPCs have the capability to survive and spontaneously differentiate into functional and mature neurons throughout a time course of a month, while no residual pluripotent cells were detectable. CONCLUSION: We describe the generation of transgene- and xeno-free NPCs. This simple differentiation protocol combined with the ability of in vivo cell tracking presents a valuable tool to develop safe and effective cell therapies for various brain injuries.

Non-invasive imaging of tau-targeted probe uptake by whole brain multi-spectral optoacoustic tomography.

PURPOSE: Abnormal tau accumulation within the brain plays an important role in tauopathies such as Alzheimer's disease and frontotemporal dementia. High-resolution imaging of tau deposits at the whole-brain scale in animal disease models is highly desired. METHODS: We approached this challenge by non-invasively imaging the brains of P301L mice of 4-repeat tau with concurrent volumetric multi-spectral optoacoustic tomography (vMSOT) at ~ 115 µm spatial resolution using the tau-targeted pyridinyl-butadienyl-benzothiazole derivative PBB5 (i.v.). In vitro probe characterization, concurrent vMSOT and epi-fluorescence imaging of in vivo PBB5 targeting (i.v.) was performed in P301L and wild-type mice, followed by ex vivo validation using AT-8 antibody for phosphorylated tau. RESULTS: PBB5 showed specific binding to recombinant K18 tau fibrils by fluorescence assay, to post-mortem Alzheimer's disease brain tissue homogenate by competitive binding against [11C]PBB3 and to tau deposits (AT-8 positive) in post-mortem corticobasal degeneration and progressive supranuclear palsy brains. Dose-dependent optoacoustic and fluorescence signal intensities were observed in the mouse brains following i.v. administration of different concentrations of PBB5. In vivo vMSOT brain imaging of P301L mice showed higher retention of PBB5 in the tau-laden cortex and hippocampus compared to wild-type mice, as confirmed by ex vivo vMSOT, epi-fluorescence, multiphoton microscopy, and immunofluorescence staining. CONCLUSIONS: We demonstrated non-invasive whole-brain imaging of tau in P301L mice with vMSOT system using PBB5 at a previously unachieved ~ 115 µm spatial resolution. This platform provides a new tool to study tau spreading and clearance in a tauopathy mouse model, foreseeable in monitoring tau targeting putative therapeutics.

The antibody aducanumab reduces Aß plaques in Alzheimer's disease.

Alzheimer's disease (AD) is characterized by deposition of amyloid-ß (Aß) plaques and neurofibrillary tangles in the brain, accompanied by synaptic dysfunction and neurodegeneration. Antibody-based immunotherapy against Aß to trigger its clearance or mitigate its neurotoxicity has so far been unsuccessful. Here we report the generation of aducanumab, a human monoclonal antibody that selectively targets aggregated Aß. In a transgenic mouse model of AD, aducanumab is shown to enter the brain, bind parenchymal Aß, and reduce soluble and insoluble Aß in a dose-dependent manner. In patients with prodromal or mild AD, one year of monthly intravenous infusions of aducanumab reduces brain Aß in a dose- and time-dependent manner. This is accompanied by a slowing of clinical decline measured by Clinical Dementia Rating-Sum of Boxes and Mini Mental State Examination scores. The main safety and tolerability findings are amyloid-related imaging abnormalities. These results justify further development of aducanumab for the treatment of AD. Should the slowing of clinical decline be confirmed in ongoing phase 3 clinical trials, it would provide compelling support for the amyloid hypothesis.

Lysine acetyltransferase Tip60 acetylates the APP adaptor Fe65 to increase its transcriptional activity.

Proteolytic processing of the amyloid precursor protein (APP) releases the APP intracellular domain (AICD) from the membrane. Bound to the APP adaptor protein Fe65 and the lysine acetyltransferase (KAT) Tip60, AICD translocates to the nucleus. Here, the complex forms spherical condensates at sites of endogenous target genes, termed AFT spots (AICD-Fe65-Tip60). We show that loss of Tip60 KAT activity prevents autoacetylation, reduces binding of Fe65 and abolishes Fe65-mediated stabilization of Tip60. Autoacetylation is a prerequisite for AFT spot formation, with KAT-deficient Tip60 retained together with Fe65 in speckles. We identify lysine residues 204 and 701 of Fe65 as acetylation targets of Tip60. We do not detect acetylation of AICD. Mutation of Fe65 K204 and K701 to glutamine, mimicking acetylation-induced charge neutralization, increases the transcriptional activity of Fe65 whereas Tip60 inhibition reduces it. The lysine deacetylase (KDAC) class III Sirt1 deacetylates Fe65 and pharmacological modulation of Sirt1 activity regulates Fe65 transcriptional activity. A second acetylation/deacetylation cycle, conducted by CBP and class I/II KDACs at different lysine residues, regulates stability of Fe65. This is the first report describing a role for acetylation in the regulation of Fe65 transcriptional activity, with Tip60 being the only KAT tested that supports AFT spot formation.

SWI and phase imaging reveal intracranial calcifications in the P301L mouse model of human tauopathy.

OBJECTIVE: Brain calcifications are associated with several neurodegenerative diseases. Here, we describe the occurrence of intracranial calcifications as a new phenotype in transgenic P301L mice overexpressing four repeat tau, a model of human tauopathy. MATERIALS AND METHODS: Thirty-six P301L mice (Thy1.2) and ten age-matched non-transgenic littermates of different ages were assessed. Gradient echo data were acquired in vivo and ex vivo at 7 T and 9.4 T for susceptibility-weighted imaging (SWI) and phase imaging. In addition, ex vivo micro-computed tomography (µCT) was performed. Histochemistry and immunohistochemistry were used to investigate the nature of the imaging lesions. RESULTS: SW images revealed regional hypointensities in the hippocampus, cortex, caudate nucleus, and thalamus of P301L mice, which in corresponding phase images indicated diamagnetic lesions. Concomitantly, µCT detected hyperdense lesions, though fewer lesions were observed compared to MRI. Diamagnetic susceptibility lesions in the hippocampus increased with age. The immunochemical staining of brain sections revealed osteocalcin-positive deposits. Furthermore, intra-neuronal and vessel-associated osteocalcin-containing nodules co-localized with phosphorylated-tau (AT8 and AT100) in the hippocampus, while vascular osteocalcin-containing nodules were detected in the thalamus in the absence of phosphorylated-tau deposition. DISCUSSION: SWI and phase imaging sensitively detected intracranial calcifications in the P301L mouse model of human tauopathy.

Diffusion Tensor Imaging Reveals Whole-Brain Microstructural Changes in the P301L Mouse Model of Tauopathy.

INTRODUCTION: Increased expression of hyperphosphorylated tau and the formation of neurofibrillary tangles are associated with neuronal loss and white matter damage. Using high-resolution ex vivo diffusion tensor imaging (DTI), we investigated microstructural changes in the white and grey matter in the P301L mouse model of human tauopathy at 8.5 months of age. For unbiased computational analysis, we implemented a pipeline for voxel-based analysis (VBA) and atlas-based analysis (ABA) of DTI mouse brain data. METHODS: Hemizygous and homozygous transgenic P301L mice and non-transgenic littermates were used. DTI data were acquired for generation of fractional anisotropy (FA), mean diffusivity (MD), radial diffusivity (RD), and axial diffusivity (AD) maps. VBA on the entire brain was performed using SPM8 and the SPM Mouse toolbox. Initially, all DTI maps were coregistered with the Allen mouse brain atlas to bring them to one common coordinate space. In VBA, coregistered DTI maps were normalized and smoothed in order to perform two-sample and unpaired t tests with false discovery rate correction to compare hemizygotes with non-transgenic littermates, homozygotes with non-transgenic littermates, and hemizygotes with homozygotes on each DTI parameter map. In ABA, the average values for selected regions of interests were computed with coregistered DTI maps and labels in Allen mouse brain atlas. Afterwards, a Kruskal-Wallis one-way ANOVA on ranks with a Tukey post hoc test was executed on the estimated average values. RESULTS: With VBA, we found pronounced and brain-wide spread changes when comparing homozygous, P301L mice with non-transgenic littermates, which were not seen when comparing hemizygous P301L with non-transgenic animals. Statistical comparison of DTI metrics in selected brain regions by ABA corroborated findings from VBA. FA was found to be decreased in most brain regions, while MD, RD, and AD were increased in homozygotes compared to hemizygotes and non-transgenic littermates. DISCUSSION/CONCLUSION: High-resolution ex vivo DTI demonstrated brain-wide microstructural and gene-dose-dependent changes in the P301L mouse model of human tauopathy. The DTI analysis pipeline may serve for the phenotyping of models of tauopathy and other brain diseases.

Development of an aggregate-selective, human-derived α-synuclein antibody BIIB054 that ameliorates disease phenotypes in Parkinson's disease models.

Aggregation of α-synuclein (α-syn) is neuropathologically and genetically linked to Parkinson's disease (PD). Since stereotypic cell-to-cell spreading of α-syn pathology is believed to contribute to disease progression, immunotherapy with antibodies directed against α-syn is considered a promising therapeutic approach for slowing disease progression. Here we report the identification, binding characteristics, and efficacy in PD mouse models of the human-derived α-syn antibody BIIB054, which is currently under investigation in a Phase 2 clinical trial for PD. BIIB054 was generated by screening human memory B-cell libraries from healthy elderly individuals. Epitope mapping studies conducted using peptide scanning, X-ray crystallography, and mutagenesis show that BIIB054 binds to α-syn residues 1-10. BIIB054 is highly selective for aggregated forms of α-syn with at least an 800-fold higher apparent affinity for fibrillar versus monomeric recombinant α-syn and a strong preference for human PD brain tissue. BIIB054 discriminates between monomers and oligomeric/fibrillar forms of α-syn based on high avidity for aggregates, driven by weak monovalent affinity and fast binding kinetics. In efficacy studies in three different mouse models with intracerebrally inoculated preformed α-syn fibrils, BIIB054 treatment attenuated the spreading of α-syn pathology, rescued motor impairments, and reduced the loss of dopamine transporter density in dopaminergic terminals in striatum. The preclinical data reported here provide a compelling rationale for clinical development of BIIB054 for the treatment and prevention of PD.

A fluorescent protein-readout for transcriptional activity reveals regulation of APP nuclear signaling by phosphorylation sites.

Signaling pathways that originate at the plasma membrane, including regulated intramembrane proteolysis (RIP), enable extracellular cues to control transcription. We modified the yeast Gal4 transcription system to study the nuclear translocation of transcriptionally active complexes using the fluorescent protein citrine (Cit) as a reporter. This enabled highly sensitive quantitative analysis of transcription in situ at the single cell level. The Gal4/UAS-Cit transcription assay displayed a sigmoidal response limited by the number of integrated reporter cassettes. We validated the assay by analyzing nuclear translocation of the amyloid precursor protein (APP) intracellular domain (AICD) and confirmed the requirement of Fe65 for nuclear translocation of AICD. In addition to the strong on-off effects on transcriptional activity, the results of this assay establish that phosphorylation modifies nuclear signaling. The Y682F mutation in APP showed the strongest increase in Cit expression, underscoring its role in regulating Fe65 binding. Together, we established a highly sensitive fluorescent protein-based assay that can monitor transcriptional activity at the single cell level and demonstrate that AICD phosphorylation affects Fe65 nuclear activity. This assay also introduces a platform for future single cell-based drug screening methods for nuclear translocation.

Tau Antibody Targeting Pathological Species Blocks Neuronal Uptake and Interneuron Propagation of Tau in Vitro.

The clinical progression of Alzheimer disease (AD) is associated with the accumulation of tau neurofibrillary tangles, which may spread throughout the cortex by interneuronal tau transfer. If so, targeting extracellular tau species may slow the spreading of tau pathology and possibly cognitive decline. To identify suitable target epitopes, we tested the effects of a panel of tau antibodies on neuronal uptake and aggregation in vitro. Immunodepletion was performed on brain extract from tau-transgenic mice and postmortem AD brain and added to a sensitive fluorescence resonance energy transfer-based tau uptake assay to assess blocking efficacy. The antibodies reduced tau uptake in an epitope-dependent manner: N-terminal (Tau13) and middomain (6C5 and HT7) antibodies successfully prevented uptake of tau species, whereas the distal C-terminal-specific antibody (Tau46) had little effect. Phosphorylation-dependent (40E8 and p396) and C-terminal half (4E4) tau antibodies also reduced tau uptake despite removing less total tau by immunodepletion, suggesting specific interactions with species involved in uptake. Among the seven antibodies evaluated, 6C5 most efficiently blocked uptake and subsequent aggregation. More important, 6C5 also blocked neuron-to-neuron spreading of tau in a unique three-chamber microfluidic device. Furthermore, 6C5 slowed down the progression of tau aggregation even after uptake had begun. Our results imply that not all antibodies/epitopes are equally robust in terms of blocking tau uptake of human AD-derived tau species.

Extravascular CD3+ T Cells in Brains of Alzheimer Disease Patients Correlate with Tau but Not with Amyloid Pathology: An Immunohistochemical Study.

BACKGROUND: Strong genetic and epidemiological evidence points to a crucial role of the immune system in the development of Alzheimer disease (AD). CD3+ T lymphocytes have been described in brains of postmortem AD patients and in transgenic models of AD-like cerebral amyloidosis and tau pathology. However, the occurrence of T cells in AD brains is still controversial; furthermore, the relationship between T cells and hallmarks of AD pathology (amyloid plaques and neurofibrillary tangles) remains to be established. OBJECTIVES: We have studied the occurrence of T cells in postmortem hippocampi and mid frontal gyrus (MFG) samples of AD patients (Braak stage V-VI) and nondemented control subjects and correlated it with amyloid and tau pathology burden. METHODS: Confocal microscopy and bright-field immunohistochemistry were used to identify brain-associated T cells. Extravascular CD3+ T cells were quantified and compared to nondemented controls. In addition, numbers of extravascular CD3+ T cells were correlated with amyloid (6E10 staining) and tau pathology (AT8 staining) in the same sections. RESULTS: Several CD3+, extravascular T cells were observed in the brains of AD patients, mostly of the CD8+ subtype. AD hippocampi harbored significantly increased numbers of extravascular CD3+ T cells compared to nondemented controls. CD3+ T cells significantly correlated with tau pathology but not with amyloid plaques in AD samples. CONCLUSIONS: Our data support the notion of T-cell occurrence in AD brains and suggest that, in advanced stages of AD, T-cell extravasation is driven by tau-related neurodegenerative changes rather than by cerebral amyloidosis. T cells could be crucial for driving the amyloid-independent phase of the AD pathology.

Hippocampal shape alterations are associated with regional Aß load in cognitively normal elderly individuals.

Aß deposition is a driving force of Alzheimer's disease pathology and can be detected early by amyloid positron emission tomography. Identifying presymptomatic structural brain changes associated with Aß deposition might lead to a better understanding of its consequences and provide early diagnostic information. In this respect we analyzed measures of cortical thickness and subcortical volumes along with hippocampal, thalamic and striatal shape and surface area by applying novel analysis strategies for structural magnetic resonance imaging. We included 69 cognitively normal elderly subjects after careful clinical and neuropsychological workup. Standardized uptake value ratios (cerebellar reference) for uptake of 11-C-Pittsburgh Compound B (PiB) were calculated from positron emission tomographic data for a cortical measurement and for bilateral hippocampus, thalamus and striatum. Associations to shape, surface area, volume and cortical thickness were tested using regression models that included significant predictors as covariates. Left anterior hippocampal shape was associated with regional PiB uptake (P < 0.05, FDR corrected), whereas volumes of the hippocampi and their subregions were not associated with cortical or regional PiB uptake (all P > 0.05, FDR corrected). Within the entorhinal cortical region of both hemispheres, thickness was negatively associated with cortical PiB uptake (P < 0.05, FDR corrected). Hence, localized shape measures and cortical thickness may be potential biomarkers of presymptomatic Alzheimer's disease.