Tau imaging: early progress and future directions.

Use of selective in-vivo tau imaging will enable improved understanding of tau aggregation in the brain, facilitating research into causes, diagnosis, and treatment of major tauopathies such as Alzheimer's disease, progressive supranuclear palsy, corticobasal syndrome, chronic traumatic encephalopathy, and some variants of frontotemporal lobar degeneration. Neuropathological studies of Alzheimer's disease show a strong association between tau deposits, decreased cognitive function, and neurodegenerative changes. Selective tau imaging will allow the in-vivo exploration of such associations and measure the global and regional changes in tau deposits over time. Such imaging studies will comprise non-invasive assessment of the spatial and temporal pattern of tau deposition over time, providing insight into the role tau plays in ageing and helping to establish the relation between cognition, genotype, neurodegeneration, and other biomarkers. Once validated, selective tau imaging might be useful as a diagnostic, prognostic, and progression biomarker, and a surrogate marker for the monitoring of efficacy and patient recruitment for anti-tau therapeutic trials.

Non-invasive assessment of Alzheimer's disease neurofibrillary pathology using 18F-THK5105 PET.

Non-invasive imaging of tau pathology in the living brain would be useful for accurately diagnosing Alzheimer's disease, tracking disease progression, and evaluating the treatment efficacy of disease-specific therapeutics. In this study, we evaluated the clinical usefulness of a novel tau-imaging positron emission tomography tracer 18F-THK5105 in 16 human subjects including eight patients with Alzheimer's disease (three male and five females, 66-82 years) and eight healthy elderly controls (three male and five females, 63-76 years). All participants underwent neuropsychological examination and 3D magnetic resonance imaging, as well as both 18F-THK5105 and 11C-Pittsburgh compound B positron emission tomography scans. Standard uptake value ratios at 90-100 min and 40-70 min post-injection were calculated for 18F-THK5105 and 11C-Pittsburgh compound B, respectively, using the cerebellar cortex as the reference region. As a result, significantly higher 18F-THK5105 retention was observed in the temporal, parietal, posterior cingulate, frontal and mesial temporal cortices of patients with Alzheimer's disease compared with healthy control subjects. In patients with Alzheimer's disease, the inferior temporal cortex, which is an area known to contain high densities of neurofibrillary tangles in the Alzheimer's disease brain, showed prominent 18F-THK5105 retention. Compared with high frequency (100%) of 18F-THK5105 retention in the temporal cortex of patients with Alzheimer's disease, frontal 18F-THK5105 retention was less frequent (37.5%) and was only observed in cases with moderate-to-severe Alzheimer's disease. In contrast, 11C-Pittsburgh compound B retention was highest in the posterior cingulate cortex, followed by the ventrolateral prefrontal, anterior cingulate, and superior temporal cortices, and did not correlate with 18F-THK5105 retention in the neocortex. In healthy control subjects, 18F-THK5105 retention was ∼10% higher in the mesial temporal cortex than in the neocortex. Notably, unlike 11C-Pittsburgh compound B, 18F-THK5105 retention was significantly correlated with cognitive parameters, hippocampal and whole brain grey matter volumes, which was consistent with findings from previous post-mortem studies showing significant correlations of neurofibrillary tangle density with dementia severity or neuronal loss. From these results, 18F-THK5105 positron emission tomography is considered to be useful for the non-invasive assessment of tau pathology in the living brain. This technique would be applicable to the longitudinal evaluation of tau deposition and allow a better understanding of the pathophysiology of Alzheimer's disease.

Assessing THK523 selectivity for tau deposits in Alzheimer's disease and non-Alzheimer's disease tauopathies.

INTRODUCTION: The introduction of tau imaging agents such as (18)F-THK523 offers new hope for the in vivo assessment of tau deposition in tauopathies such as Alzheimer's disease (AD), where preliminary (18)F-THK523-PET studies have demonstrated significantly higher cortical retention of (18)F-THK523 in AD compared to age-matched healthy individuals. In addition to AD, tau imaging with PET may also be of value in assessing non-AD tauopathies, such as corticobasal degeneration (CBD), progressive supranuclear palsy (PSP) and Pick's disease (PiD). METHODS: To further investigate the ability of THK523 to recognize tau lesions, we undertook immunohistochemical and fluorescence studies in serial brain sections taken from individuals with AD (n = 3), CBD (n = 2), PSP (n = 1), PiD (n = 2) and Parkinson's disease (PD; n = 2). In addition to the neuropathological analysis, one PSP patient had undergone a (18)F-THK523 PET scan 5 months before death. RESULTS: Although THK523 labelled tau-containing lesions such as neurofibrillary tangles and neuropil threads in the hippocampus and frontal regions of AD brains, it failed to label tau-containing lesions in non-AD tauopathies. Furthermore, though THK523 faintly labelled dense-cored amyloid-ß plaques in the AD frontal cortex, it failed to label α-synuclein-containing Lewy bodies in PD brain sections. CONCLUSION: The results of this study suggest that (18)F-THK523 selectively binds to paired helical filament tau in AD brains but does not bind to tau lesions in non-AD tauopathies, or to α-synuclein in PD brains.

In vivo evaluation of a novel tau imaging tracer for Alzheimer's disease.

PURPOSE: Diagnosis of tauopathies such as Alzheimer's disease (AD) still relies on post-mortem examination of the human brain. A non-invasive method of determining brain tau burden in vivo would allow a better understanding of the pathophysiology of tauopathies. The purpose of the study was to evaluate (18)F-THK523 as a potential tau imaging tracer. METHODS: Ten healthy elderly controls, three semantic dementia (SD) and ten AD patients underwent neuropsychological examination, MRI as well as (18)F-THK523 and (11)C-Pittsburgh compound B (PIB) positron emission tomography (PET) scans. Composite memory and non-memory scores, global and hippocampal brain volume, and partial volume-corrected tissue ratios for (18)F-THK523 and (11)C-PIB were estimated for all participants. Correlational analyses were performed between global and regional (18)F-THK523, (11)C-PIB, cognition and brain volumetrics. RESULTS: (18)F-THK523 presented with fast reversible kinetics. Significantly higher (18)F-THK523 retention was observed in the temporal, parietal, orbitofrontal and hippocampi of AD patients when compared to healthy controls and SD patients. White matter retention was significantly higher than grey matter retention in all participants. The pattern of cortical (18)F-THK523 retention did not correlate with Aß distribution as assessed by (11)C-PIB and followed the known distribution of tau in the AD brain, being higher in temporal and parietal areas than in the frontal region. Unlike (11)C-PIB, hippocampal (18)F-THK523 retention was correlated with several cognitive parameters and with hippocampal atrophy. CONCLUSION: (18)F-THK523 does not bind to Aß in vivo, while following the known distribution of paired helical filaments (PHF)-tau in the brain. Significantly higher cortical (18)F-THK523 retention in AD patients as well as the association of hippocampal (18)F-THK523 retention with cognitive parameters and hippocampal volume suggests (18)F-THK523 selectively binds to tau in AD patients. Unfortunately, the very high (18)F-THK523 retention in white matter precludes simple visual inspection of the images, preventing its use in research or clinical settings.

SERF protein is a direct modifier of amyloid fiber assembly.

The inherent cytotoxicity of aberrantly folded protein aggregates contributes substantially to the pathogenesis of amyloid diseases. It was recently shown that a class of evolutionary conserved proteins, called MOAG-4/SERF, profoundly alter amyloid toxicity via an autonomous but yet unexplained mode. We show that the biological function of human SERF1a originates from its atypical ability to specifically distinguish between amyloid and nonamyloid aggregation. This inherently unstructured protein directly affected the aggregation kinetics of a broad range of amyloidogenic proteins in vitro, while being inactive against nonamyloid aggregation. A representative biophysical analysis of the SERF1a:α-synuclein (aSyn) complex revealed that the amyloid-promoting activity resulted from an early and transient interaction, which was sufficient to provoke a massive increase of soluble aSyn amyloid nucleation templates. Therefore, the autonomous amyloid-modifying activity of SERF1a observed in living organisms relies on a direct and dedicated manipulation of the early stages in the amyloid aggregation pathway.

In vitro characterization of [18F]-florbetaben, an Aß imaging radiotracer.

PURPOSE: Amyloid-ß (Aß) plaques are a major pathological hallmark of Alzheimer's disease (AD). The noninvasive detection of Aß plaques may increase the accuracy of clinical diagnosis as well as monitor therapeutic interventions. While [(11)C]-PiB is the most widely used Aß positron emission tomography (PET) radiotracer, due to the short half-life of (11)C (20 min), its application is limited to centers with an on-site cyclotron and (11)C radiochemistry expertise. Therefore, novel [(18)F] (half-life 110 min)-labeled Aß PET tracers have been developed. We have demonstrated that [(18)F]-florbetaben-PET can differentiate individuals diagnosed with AD from healthy elderly, Parkinson's disease and frontotemporal lobe dementia (FTLD-tau) patients. While [(18)F]-florbetaben-PET retention matched the reported postmortem distribution of Aß plaques, the nature of [(18)F]-florbetaben binding to other pathological lesions comprising misfolded proteins needs further assessment. The objective of this study was to determine whether Florbetaben selectively binds to Aß plaques in postmortem tissue specimens containing mixed pathological hallmarks (i.e., tau and α-synuclein aggregates). METHOD: Human AD, FTLD-tau and dementia with Lewy bodies (DLB) brain sections were analyzed by [(18)F]-florbetaben autoradiography and [(3)H]-florbetaben high-resolution emulsion autoradiography and [(19)F]-florbetaben fluorescence microscopy. RESULTS: Both autoradiographical analyses demonstrated that Florbetaben exclusively bound Aß plaques in AD brain sections at low nanomolar concentrations. Furthermore, at concentrations thousand-folds higher than those during a PET scan, [(19)F]-florbetaben did not bind to α-synuclein or tau aggregates in DLB and FTLD-tau brain sections, respectively. Detection of [(19)F]-florbetaben staining by fluorescence microscopy in several AD brain regions demonstrated that Florbetaben identified Aß plaques in all brain regions examined. CONCLUSION: This study provides further evidence that [(18)F]-florbetaben-PET is a highly selective radiotracer to assess Aß plaque deposition in the brain.

Amyloid-ß: the seeds of darkness.

Whilst the amyloid-ß (Aß) hypothesis/centric theory continues to evolve, genetic, biochemical and pathological evidence still suggests that Aß is central to the etiology of Alzheimer's disease (AD). In particular, Aß-oligomers/soluble Aß, may be an earlier determinant of Alzheimer's disease and better correlative of cognitive impairment. Whilst there are a number of Aß-oligomeric species in existence (making therapeutic and diagnostic biomarker choice cumbersome), their existence is in equilibrium with Aß-fibrils, the main constituent of cored plaques. Although Alzheimer's disease remains incurable, improvements to Aß immunotherapies and strategies to target Aß continue to evolve, with the reliance upon Aß imaging to shed light on the outcome of therapeutics proving very useful.

18F-THK523: a novel in vivo tau imaging ligand for Alzheimer's disease.

While considerable effort has focused on developing positron emission tomography ß-amyloid imaging radiotracers for the early diagnosis of Alzheimer's disease, no radiotracer is available for the non-invasive quantification of tau. In this study, we detail the characterization of (18)F-THK523 as a novel tau imaging radiotracer. In vitro binding studies demonstrated that (18)F-THK523 binds with higher affinity to a greater number of binding sites on recombinant tau (K18Δ280K) compared with ß-amyloid(1-42) fibrils. Autoradiographic and histofluorescence analysis of human hippocampal serial sections with Alzheimer's disease exhibited positive THK523 binding that co-localized with immunoreactive tau pathology, but failed to highlight ß-amyloid plaques. Micro-positron emission tomography analysis demonstrated significantly higher retention of (18)F-THK523 (48%; P < 0.007) in tau transgenic mice brains compared with their wild-type littermates or APP/PS1 mice. The preclinical examination of THK523 has demonstrated its high affinity and selectivity for tau pathology both in vitro and in vivo, indicating that (18)F-THK523 fulfils ligand criteria for human imaging trials.

A copper radiopharmaceutical for diagnostic imaging of Alzheimer's disease: a bis(thiosemicarbazonato)copper(II) complex that binds to amyloid-beta plaques.

A bis(thiosemicarbazonato)copper(ii) complex with an appended stilbene functional group binds to amyloid-beta plaques that are associated with Alzheimer's disease. The complex has the potential to be of use as a copper-64 radiopharmaceutical to assist in the diagnosis of Alzheimer's disease by positron emission tomography.

Bis(thiosemicarbazonato) Cu-64 complexes for positron emission tomography imaging of Alzheimer's disease.

A bis (thiosemicarbazonato) complex radiolabeled with positron emitting Cu-64 can be used for a new and alternative method for the non-invasive diagnosis of Alzheimer's disease using positron emission tomography (PET). Most imaging agents being investigated for the diagnosis of Alzheimer's disease target amyloid-beta plaque burden but our new approach highlights altered copper homeostasis. This approach has the potential to offer complementary information to other diagnostic procedures that elucidate plaque burden.

In vitro characterisation of BF227 binding to alpha-synuclein/Lewy bodies.

Amyloid-beta (Abeta) plaques are a pathological hallmark of Alzheimer's disease and a current target for positron emission tomography (PET) imaging agents. Whilst [(11)C]-PiB is currently the most widely used PET ligand in clinic, a novel family of benzoxazole compounds have shown promise as Abeta imaging agents; particularly BF227. We characterised the in vitro binding of [(18)F]-BF227 toward alpha-synuclein to address its selectivity for Abeta pathology, to establish whether [(18)F]-BF227 binds to alpha-synuclein/Lewy bodies, in addition to Abeta plaques. In vitro [(18)F]-BF227 saturation studies were conducted with 200 nM alpha-synuclein or Abeta(1-42) fibrils or 100 microg of Alzheimer's disease, pure dementia with Lewy bodies or control brain homogenates. Non-specific binding was established with PiB (1 microM). In vitro binding studies indicated that [(18)F]-BF227 binds with high affinity to two binding sites on Abeta(1-42) fibrils (K(D1) = 1.31 and K(D2) = 80 nM, respectively) and to one class of binding sites on alpha-synuclein fibrils (K(D) = 9.63 nM). [(18)F]-BF227 bound to Abeta-containing Alzheimer's disease brain (K(D) = 25 +/- 0.5 nM), but failed to bind to Abeta-free dementia with Lewy bodies or age-matched control homogenates. Moreover, BF227 labelled both Abeta plaques and Lewy bodies in immunohistochemical/fluorescence analysis of human Alzheimer's disease and Parkinson's disease brain sections, respectively. This study suggests that [(18)F]-BF227 is not Abeta-selective. Evaluation of BF227 as a potential biomarker for Parkinson's disease is warranted.

Formation of dopamine-mediated alpha-synuclein-soluble oligomers requires methionine oxidation.

alpha-Synuclein is the major component of the intracellular Lewy body inclusions present in Parkinson disease (PD) neurons. PD involves the loss of dopaminergic neurons in the substantia nigra and the subsequent depletion of dopamine (DA) in the striatum. DA can inhibit alpha-synuclein fibrillization in vitro and promote alpha-synuclein aggregation into soluble oligomers. We have studied the mechanism by which DA mediates alpha-synuclein aggregation into soluble oligomers. Reacting alpha-synuclein with DA increased the mass of alpha-synuclein by 64 Da. NMR showed that all four methionine residues were oxidized by DA, consistent with the addition of 64 Da. Substituting all four methionines to alanine significantly reduced the formation of DA-mediated soluble oligomers. The (125)YEMPS(129) motif in alpha-synuclein can modulate DA inhibition of alpha-synuclein fibrillization. However, alpha-synuclein ending before the (125)YEMPS(129) motif (residues 1-124) could still form soluble oligomers. The addition of exogenous synthetic YEMPS peptide inhibited the formation of soluble oligomers and resulted in the YEMPS peptide being oxidized. Therefore, the (125)YEMPS(129) acts as an antioxidant rather than interacting directly with DA. Our study defines methionine oxidation as the dominant mechanism by which DA generates soluble alpha-synuclein oligomers and highlights the potential role for oxidative stress in modulating alpha-synuclein aggregation.

Characterization of PiB binding to white matter in Alzheimer disease and other dementias.

UNLABELLED: 11C-Pittsburgh Compound B (11C-PiB) PET has demonstrated significantly higher PiB retention in the gray matter of Alzheimer disease (AD) patients than in healthy controls (HCs). PiB is similarly retained within the white matter of HC and AD brains. Although the specificity of PiB for Abeta plaques in gray matter has been well described, the nature of PiB binding to white matter remains unclear. In this study, we characterized the binding of PiB to human white matter homogenates. METHODS: In vitro binding studies were conducted using 3H-PiB (0.1-500 nM) and white matter brain homogenates (100 microg) from 3 AD patients and 3 HCs. Nonspecific binding was determined using PiB (1 microM). White matter from the same patients was also analyzed by immunofluorescence/immunohistochemistry (IF/IHC) microscopy and Western blotting for Abeta expression. White matter kinetics were also characterized in vivo through 11C-PiB PET studies in 27 HCs and 34 patients with dementia. IF/IHC experiments were conducted on 1 postmortem patient with dementia, to compare with the 11C-PiB distribution volume ratio data acquired 23 mo earlier. RESULTS: In vitro saturation studies indicated that 3H-PiB binds nonspecifically to white matter brain homogenates. PiB fluorescence staining of AD and HC brain sections was consistent with absence of Abeta in IHC staining. Higher gray matter-to-white matter ratios were observed in IHC images than in 11C-PiB PET images. CONCLUSION: These studies suggest that PiB binding to white matter is mainly nonsaturable and nonspecific and that PiB retention in the 11C-PiB PET studies is largely attributable to slower PiB white matter kinetics.

Advances in molecular imaging for the diagnosis of dementia.

BACKGROUND: There is an urgent need for early diagnosis and treatment of dementia to ease caregiver burden and medical costs associated with the increasing number of affected patients. Molecular imaging with target-specific ligands is contributing to the early diagnosis of dementia and the evaluation of anti-dementia therapy. OBJECTIVE: This article reviews recent advances in the molecular imaging field applied to dementia. To illustrate the utility of molecular imaging in the clinical management of dementia, results from recently published papers using new imaging probes are compared with those from conventional imaging strategies. CONCLUSION: The recent development of ß-sheet binding agents including FDDNP, PIB, SB-13, BF-227 and BAY94-9172 enables the non-invasive detection of amyloid deposition in the brain. These agents would be useful for the early and accurate diagnosis of Alzheimer's disease, patient selection for disease-modifying therapeutic trials and monitoring the effect of anti-amyloid therapy. Also, monitoring neurotransmitter function contributes to the differential diagnosis of dementia and refinement of treatment protocols. New targets for molecular imaging are focusing on protein misfolding diseases associated with the neurotoxic deposition of aggregated tau, α-synuclein and prion proteins.

Amyloid imaging in Alzheimer's disease and other dementias.

With the advent of new therapeutic strategies aimed at reducing ß-amyloid (Aß) burden in the brain to potentially prevent or delay functional and irreversible cognitive loss, there is increased interest in developing agents that allow assessment of Aß burden in vivo. Molecular neuroimaging techniques such as positron emission tomography (PET), in conjunction with related biomarkers in plasma and cerebrospinal fluid, are proving valuable in the early and differential diagnosis of Alzheimer's disease (AD). (11)C-PiB PET has proven useful in the discrimination of dementias, showing significantly higher PiB retention in grey matter of AD patients when compared with healthy controls or patients with frontotemporal dementia. (11)C-PiB PET also appears to be more accurate than FDG for the diagnosis of AD. Despite apparently underestimating the Aß burden in the brain, (11)C-PiB PET is an optimal method to differentiate healthy controls from AD, matching histopathological reports in aging and dementia and reflecting the true regional density of Aß plaques in cortical areas. High striatal Aß deposition seems to be typical for carriers of familial forms of AD, whilst ApoE ε4 carriers, independent of diagnosis or disease severity, present with higher Aß burden than non- ε4 carriers. Characterization of the binding properties of PiB has shown that despite binding to other misfolded proteins in vitro, PiB is extremely selective for Aß at the concentrations achieved during a PET scan. Aß burden as assessed by PET does not correlate with measures of cognition or cognitive decline in AD. Approximately 30% of apparently healthy older people, and 50-60% of people with mild cognitive impairment, present with cortical (11)C-PiB retention. In these groups, Aß burden does correlate with episodic memory and rate of memory decline. These observations suggest that Aß deposition is not part of normal ageing, supporting the hypothesis that Αß deposition occurs well before the onset of symptoms and is likely to represent preclinical AD. Further longitudinal observations, coupled with different disease-specific tracers and biomarkers are required not only to confirm this hypothesis, but also to better elucidate the role of Αß deposition in the course of Alzheimer's disease.

The ART of loss: Abeta imaging in the evaluation of Alzheimer's disease and other dementias.

Molecular neuroimaging based on annihilation radiation tomographic (ART) techniques such as positron emission tomography (PET), in conjunction with related biomarkers in plasma and cerebrospinal fluid (CSF), are proving valuable in the early and differential diagnosis of Alzheimer's disease (AD). With the advent of new therapeutic strategies aimed at reducing beta-amyloid (Abeta) burden in the brain to potentially prevent or delay functional and irreversible cognitive loss, there is increased interest in developing agents that allow assessment of Abeta burden in vivo. Abeta burden as assessed by molecular imaging matches histopathological reports of Abeta plaque distribution in aging and dementia and appears more accurate than FDG for the diagnosis of AD. Abeta imaging is also a very powerful tool in the differential diagnosis of AD from fronto-temporal dementia (FTD). Although Abeta burden as assessed by PET does not correlate with measures of cognitive decline in AD, it does correlate with memory impairment and rate of memory decline in mild cognitive impairment (MCI) and healthy older subjects. Approximately 30% of asymptomatic controls present cortical (11)C-PiB retention. These observations suggest that Abeta deposition is not part of normal ageing, supporting the hypothesis that Abeta deposition occurs well before the onset of symptoms and is likely to represent preclinical AD. Further longitudinal observations are required to confirm this hypothesis and to better elucidate the role of Abeta deposition in the course of Alzheimer's disease.

Amyloid-beta-anti-amyloid-beta complex structure reveals an extended conformation in the immunodominant B-cell epitope.

Alzheimer's disease (AD) is the most common form of dementia. Amyloid-beta (A beta) peptide, generated by proteolytic cleavage of the amyloid precursor protein, is central to AD pathogenesis. Most pharmaceutical activity in AD research has focused on A beta, its generation and clearance from the brain. In particular, there is much interest in immunotherapy approaches with a number of anti-A beta antibodies in clinical trials. We have developed a monoclonal antibody, called WO2, which recognises the A beta peptide. To this end, we have determined the three-dimensional structure, to near atomic resolution, of both the antibody and the complex with its antigen, the A beta peptide. The structures reveal the molecular basis for WO2 recognition and binding of A beta. The A beta peptide adopts an extended, coil-like conformation across its major immunodominant B-cell epitope between residues 2 and 8. We have also studied the antibody-bound A beta peptide in the presence of metals known to affect its aggregation state and show that WO2 inhibits these interactions. Thus, antibodies that target the N-terminal region of A beta, such as WO2, hold promise for therapeutic development.

In vitro characterization of Pittsburgh compound-B binding to Lewy bodies.

Dementia with Lewy bodies (DLB) is pathologically characterized by the presence of alpha-synuclein-containing Lewy bodies within the neocortical, limbic, and paralimbic regions. Like Alzheimer's disease (AD), Abeta plaques are also present in most DLB cases. The contribution of Abeta to the development of DLB is unclear. [11C]-Pittsburgh compound B ([11C]-PIB) is a thioflavin-T derivative that has allowed in vivo Abeta burden to be quantified using positron emission tomography (PET). [11C]-PIB PET studies have shown similar high cortical [11C]-PIB binding in AD and DLB subjects. To establish the potential binding of PIB to alpha-synuclein in DLB patients, we characterized the in vitro binding of PIB to recombinant human alpha-synuclein and DLB brain homogenates. Analysis of the in vitro binding studies indicated that [3H]-PIB binds to alpha-synuclein fibrils but with lower affinity than that demonstrated/reported for Abeta(1-42) fibrils. Furthermore, [3H]-PIB was observed to bind to Abeta plaque-containing DLB brain homogenates but failed to bind to DLB homogenates that were Abeta plaque-free ("pure DLB"). Positive PIB fluorescence staining of DLB brain sections colocalized with immunoreactive Abeta plaques but failed to stain Lewy bodies. Moreover, image quantification analysis suggested that given the small size and low density of Lewy bodies within the brains of DLB subjects, any contribution of Lewy bodies to the [11C]-PIB PET signal would be negligible. These studies indicate that PIB retention observed within the cortical gray matter regions of DLB subjects in [11C]-PIB PET studies is largely attributable to PIB binding to Abeta plaques and not Lewy bodies.

Concentration dependent Cu2+ induced aggregation and dityrosine formation of the Alzheimer's disease amyloid-beta peptide.

The Amyloid beta peptide (Abeta) of Alzheimer's diseases (AD) is closely linked to the progressive cognitive decline associated with the disease. Cu2+ ions can induce the de novo aggregation of the Abeta peptide into non-amyloidogenic aggregates and the production of a toxic species. The mechanism by which Cu2+ mediates the change from amyloid material toward Cu2+ induced aggregates is poorly defined. Here we demonstrate that the aggregation state of Abeta1-42 at neutral pH is governed by the Cu2+:peptide molar ratio. By probing amyloid content and total aggregation, we observed a distinct Cu2+ switching effect centered at equimolar Cu2+:peptide ratios. At sub-equimolar Cu2+:peptide molar ratios, Abeta1-42 forms thioflavin-T reactive amyloid; conversely, at supra-equimolar Cu2+:peptide molar ratios, Abeta1-42 forms both small spherical oligomers approximately 10-20 nm in size and large amorphous aggregates. We demonstrate that these insoluble aggregates form spontaneously via a soluble species without the presence of an observable lag phase. In seeding experiments, the Cu2+ induced aggregates were unable to influence fibril formation or convert into fibrillar material. Aged Cu2+ induced aggregates are toxic when compared to Abeta1-42 aged in the absence of Cu2+. Importantly, the formation of dityrosine crosslinked Abeta, by the oxidative modification of the peptide, only occurs at equimolar molar ratios and above. The formation of dityrosine adducts occurs following the initiation of aggregation and hence does not drive the formation of the Cu2+ induced aggregates. These results define the role Cu2+ plays in modulating the aggregation state and toxicity of Abeta1-42.

Protein tyrosine phosphatase hPTPN20a is targeted to sites of actin polymerization.

The human genome encodes 38 classical tyrosine-specific PTPs (protein tyrosine phosphatases). Many PTPs have been shown to regulate fundamental cellular processes and several are mutated in human diseases. We report that the product of the PTPN20 gene at the chromosome locus 10q11.2 is alternatively spliced to generate 16 possible variants of the classical human non-transmembrane PTP 20 (hPTPN20). One of these variants, hPTPN20a, was expressed in a wide range of both normal and transformed cell lines. The catalytic domain of hPTPN20 exhibited catalytic activity towards tyrosyl phosphorylated substrates, confirming that it is a bona fide PTP. In serum-starved COS1 cells, hPTPN20a was targeted to the nucleus and the microtubule network, colocalizing with the microtubule-organizing centre and intracellular membrane compartments, including the endoplasmic reticulum and the Golgi apparatus. Stimulation of cells with epidermal growth factor, osmotic shock, pervanadate, or integrin ligation targeted hPTPN20a to actin-rich structures that included membrane ruffles. The present study identifies hPTPN20a as a novel and widely expressed phosphatase with a dynamic subcellular distribution that is targeted to sites of actin polymerization.