Publications by authors named "Lars Lannfelt"

193 Publications

Novel multivalent design of a monoclonal antibody improves binding strength to soluble aggregates of amyloid beta.

Transl Neurodegener 2021 Sep 28;10(1):38. Epub 2021 Sep 28.

Protein Drug Design, Faculty of Pharmacy, Uppsala University, 75124, Uppsala, Sweden.

Background: Amyloid-β (Aβ) immunotherapy is a promising therapeutic strategy in the fight against Alzheimer's disease (AD). A number of monoclonal antibodies have entered clinical trials for AD. Some of them have failed due to the lack of efficacy or side-effects, two antibodies are currently in phase 3, and one has been approved by FDA. The soluble intermediate aggregated species of Aβ, termed oligomers and protofibrils, are believed to be key pathogenic forms, responsible for synaptic and neuronal degeneration in AD. Therefore, antibodies that can strongly and selectively bind to these soluble intermediate aggregates are of great diagnostic and therapeutic interest.

Methods: We designed and recombinantly produced a hexavalent antibody based on mAb158, an Aβ protofibril-selective antibody. The humanized version of mAb158, lecanemab (BAN2401), is currently in phase 3 clinical trials for the treatment of AD. The new designs involved recombinantly fusing single-chain fragment variables to the N-terminal ends of mAb158 antibody. Real-time interaction analysis with LigandTracer and surface plasmon resonance were used to evaluate the kinetic binding properties of the generated antibodies to Aβ protofibrils. Different ELISA setups were applied to demonstrate the binding strength of the hexavalent antibody to Aβ aggregates of different sizes. Finally, the ability of the antibodies to protect cells from Aβ-induced effects was evaluated by MTT assay.

Results: Using real-time interaction analysis with LigandTracer, the hexavalent design promoted a 40-times enhanced binding with avidity to protofibrils, and most of the added binding strength was attributed to the reduced rate of dissociation. Furthermore, ELISA experiments demonstrated that the hexavalent design also had strong binding to small oligomers, while retaining weak and intermediate binding to monomers and insoluble fibrils. The hexavalent antibody also reduced cell death induced by a mixture of soluble Aβ aggregates.

Conclusion: We provide a new antibody design with increased valency to promote binding avidity to an enhanced range of sizes of Aβ aggregates. This approach should be general and work for any aggregated protein or repetitive target.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s40035-021-00258-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8477473PMC
September 2021

The Amyloid-β Pathway in Alzheimer's Disease.

Mol Psychiatry 2021 Aug 30. Epub 2021 Aug 30.

Eisai Inc., Neurology Business Group, Woodcliff Lake, NJ, USA.

Breakthroughs in molecular medicine have positioned the amyloid-β (Aβ) pathway at the center of Alzheimer's disease (AD) pathophysiology. While the detailed molecular mechanisms of the pathway and the spatial-temporal dynamics leading to synaptic failure, neurodegeneration, and clinical onset are still under intense investigation, the established biochemical alterations of the Aβ cycle remain the core biological hallmark of AD and are promising targets for the development of disease-modifying therapies. Here, we systematically review and update the vast state-of-the-art literature of Aβ science with evidence from basic research studies to human genetic and multi-modal biomarker investigations, which supports a crucial role of Aβ pathway dyshomeostasis in AD pathophysiological dynamics. We discuss the evidence highlighting a differentiated interaction of distinct Aβ species with other AD-related biological mechanisms, such as tau-mediated, neuroimmune and inflammatory changes, as well as a neurochemical imbalance. Through the lens of the latest development of multimodal in vivo biomarkers of AD, this cross-disciplinary review examines the compelling hypothesis- and data-driven rationale for Aβ-targeting therapeutic strategies in development for the early treatment of AD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41380-021-01249-0DOI Listing
August 2021

The deletion causes early onset autosomal dominant Alzheimer's disease by altering APP processing and increasing amyloid β fibril formation.

Sci Transl Med 2021 08;13(606)

Department of Public Health and Caring Sciences, Geriatrics, Uppsala University, 75185 Uppsala, Sweden.

Point mutations in the amyloid precursor protein gene () cause familial Alzheimer's disease (AD) by increasing generation or altering conformation of amyloid β (Aβ). Here, we describe the mutation (Δ690-695), the first reported deletion causing autosomal dominant AD. Affected individuals have an age at symptom onset in their early forties and suffer from a rapidly progressing disease course. Symptoms and biomarkers are typical of AD, with the exception of normal cerebrospinal fluid (CSF) Aβ42 and only slightly pathological amyloid-positron emission tomography signals. Mass spectrometry and Western blot analyses of patient CSF and media from experimental cell cultures indicate that the mutation alters APP processing by increasing β-secretase cleavage and affecting α-secretase cleavage. Furthermore, in vitro aggregation studies and analyses of patient brain tissue samples indicate that the longer form of mutated Aβ, AβUpp1-42, accelerates the formation of fibrils with unique polymorphs and their deposition into amyloid plaques in the affected brain.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/scitranslmed.abc6184DOI Listing
August 2021

Elevated soluble amyloid beta protofibrils in Down syndrome and Alzheimer's disease.

Mol Cell Neurosci 2021 07 4;114:103641. Epub 2021 Jun 4.

BioArctic AB, Warfvinges väg 35, SE-112 51 Stockholm, Sweden; Dept. of Public Health/Geriatrics, Uppsala University, SE-751 22 Uppsala, Sweden.

Down syndrome (DS) is caused by trisomy of chromosome 21, which leads to a propensity to develop amyloid β (Aβ) brain pathology in early adulthood followed later by cognitive and behavioral deterioration. Characterization of the Aβ pathology is important to better understand the clinical deterioration of DS individuals and to identify interventive strategies. Brain samples from people with DS and Alzheimer's disease (AD), as well as non-demented controls (NDC), were analyzed with respect to different Aβ species. Immunohistochemical staining using antibodies towards Aβ was also performed. Elevated levels of soluble Aβ protofibrils and insoluble Aβx-40 and Aβx-42 in formic acid brain extracts, and elevated immunohistochemical staining of Aβ deposits were demonstrated with the antibody BAN2401 (lecanemab) in DS and AD compared with NDC. These data and the promising data in a large phase 2 CE clinical trial with lecanemab suggest that lecanemab may have the potential to preserve cognitive capacity in DS. Lecanemab is currently in a phase 3 CE clinical trial.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.mcn.2021.103641DOI Listing
July 2021

Heparanase overexpression impedes perivascular clearance of amyloid-β from murine brain: relevance to Alzheimer's disease.

Acta Neuropathol Commun 2021 05 10;9(1):84. Epub 2021 May 10.

Department of Medical Biochemistry and Microbiology, SciLifeLab Uppsala, University of Uppsala, The Biomedical Center Husargatan 3, Box 582, 751 23, Uppsala, Sweden.

Defective amyloid-β (Aβ) clearance from the brain is a major contributing factor to the pathophysiology of Alzheimer's disease (AD). Aβ clearance is mediated by macrophages, enzymatic degradation, perivascular drainage along the vascular basement membrane (VBM) and transcytosis across the blood-brain barrier (BBB). AD pathology is typically associated with cerebral amyloid angiopathy due to perivascular accumulation of Aβ. Heparan sulfate (HS) is an important component of the VBM, thought to fulfill multiple roles in AD pathology. We previously showed that macrophage-mediated clearance of intracortically injected Aβ was impaired in the brains of transgenic mice overexpressing heparanase (Hpa-tg). This study revealed that perivascular drainage was impeded in the Hpa-tg brain, evidenced by perivascular accumulation of the injected Aβ in the thalamus of Hpa-tg mice. Furthermore, endogenous Aβ accumulated at the perivasculature of Hpa-tg thalamus, but not in control thalamus. This perivascular clearance defect was confirmed following intracortical injection of dextran that was largely retained in the perivasculature of Hpa-tg brains, compared to control brains. Hpa-tg brains presented with thicker VBMs and swollen perivascular astrocyte endfeet, as well as elevated expression of the BBB-associated water-pump protein aquaporin 4 (AQP4). Elevated levels of both heparanase and AQP4 were also detected in human AD brain. These findings indicate that elevated heparanase levels alter the organization and composition of the BBB, likely through increased fragmentation of BBB-associated HS, resulting in defective perivascular drainage. This defect contributes to perivascular accumulation of Aβ in the Hpa-tg brain, highlighting a potential role for heparanase in the pathogenesis of AD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s40478-021-01182-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8111754PMC
May 2021

A randomized, double-blind, phase 2b proof-of-concept clinical trial in early Alzheimer's disease with lecanemab, an anti-Aβ protofibril antibody.

Alzheimers Res Ther 2021 04 17;13(1):80. Epub 2021 Apr 17.

Chambers-Grundy Center for Transformative Neuroscience, Department of Brain Health, School of Integrated Health Sciences, University of Nevada Las Vegas, Las Vegas, NV, USA.

Background: Lecanemab (BAN2401), an IgG1 monoclonal antibody, preferentially targets soluble aggregated amyloid beta (Aβ), with activity across oligomers, protofibrils, and insoluble fibrils. BAN2401-G000-201, a randomized double-blind clinical trial, utilized a Bayesian design with response-adaptive randomization to assess 3 doses across 2 regimens of lecanemab versus placebo in early Alzheimer's disease, mild cognitive impairment due to Alzheimer's disease (AD) and mild AD dementia.

Methods: BAN2401-G000-201 aimed to establish the effective dose 90% (ED90), defined as the simplest dose that achieves ≥90% of the maximum treatment effect. The primary endpoint was Bayesian analysis of 12-month clinical change on the Alzheimer's Disease Composite Score (ADCOMS) for the ED90 dose, which required an 80% probability of ≥25% clinical reduction in decline versus placebo. Key secondary endpoints included 18-month Bayesian and frequentist analyses of brain amyloid reduction using positron emission tomography; clinical decline on ADCOMS, Clinical Dementia Rating-Sum-of-Boxes (CDR-SB), and Alzheimer's Disease Assessment Scale-Cognitive Subscale (ADAS-Cog14); changes in CSF core biomarkers; and total hippocampal volume (HV) using volumetric magnetic resonance imaging.

Results: A total of 854 randomized subjects were treated (lecanemab, 609; placebo, 245). At 12 months, the 10-mg/kg biweekly ED90 dose showed a 64% probability to be better than placebo by 25% on ADCOMS, which missed the 80% threshold for the primary outcome. At 18 months, 10-mg/kg biweekly lecanemab reduced brain amyloid (-0.306 SUVr units) while showing a drug-placebo difference in favor of active treatment by 27% and 30% on ADCOMS, 56% and 47% on ADAS-Cog14, and 33% and 26% on CDR-SB versus placebo according to Bayesian and frequentist analyses, respectively. CSF biomarkers were supportive of a treatment effect. Lecanemab was well-tolerated with 9.9% incidence of amyloid-related imaging abnormalities-edema/effusion at 10 mg/kg biweekly.

Conclusions: BAN2401-G000-201 did not meet the 12-month primary endpoint. However, prespecified 18-month Bayesian and frequentist analyses demonstrated reduction in brain amyloid accompanied by a consistent reduction of clinical decline across several clinical and biomarker endpoints. A phase 3 study (Clarity AD) in early Alzheimer's disease is underway.

Trial Registration: Clinical Trials.gov NCT01767311 .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13195-021-00813-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8053280PMC
April 2021

Characterization of monomeric and soluble aggregated Aβ in Down's syndrome and Alzheimer's disease brains.

Neurosci Lett 2021 05 10;754:135894. Epub 2021 Apr 10.

Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden; Clinical Neurochemistry Laboratory, Sahlgrenska University Hospital, Mölndal, Sweden.

The major characteristics of Alzheimer's disease (AD) are amyloid plaques, consisting of aggregated beta amyloid (Aβ) peptides, together with tau pathology (tangles, neuropil treads and dystrophic neurites surrounding the plaques), in the brain. Down's syndrome (DS) individuals are at increased risk to develop AD-type pathology; most DS individuals have developed substantial pathology already at the age of 40. DS individuals have an extra copy of chromosome 21, harbouring the amyloid precursor protein gene (APP). Our aim was to investigate the Aβ peptide pattern in DS and AD brains to investigate differences in their amyloid deposition and aggregation, respectively. Cortical tissue from patients with DS (with amyloid pathology), sporadic AD and controls were homogenized and fractionated into TBS (water soluble) and formic acid (water insoluble) fractions. Immunoprecipitation (IP) was performed using a variety of antibodies targeting different Aβ species including oligomeric Aβ. Mass spectrometry was then used to evaluate the presence of Aβ species in the different patient groups. A large number of Aβ peptides were identified including Aβ1-X, 2-X, 3-X, 4-X, 5-X, 11-X, and Aβ peptides extended N terminally of the BACE1 cleavage site and ending at amino 15 in the Aβ sequence APP/Aβ(-X to 15), as well as peptides post-translationally modified by pyroglutamate formation. Most Aβ peptides had higher abundance in AD and DS compared to controls, except the APP/Aβ(-X to 15) peptides which were most abundant in DS followed by controls and AD. Furthermore, the abundancies of AβX-40 and AβX-34 were increased in DS compared with AD. Aβ1-40, Aβ1-42, and Aβ4-42 were identified as the main constitutes of protofibrils (IP'd using mAb158) and higher relative Aβ1-42 signals were obtained compared with samples IP'd with 6E10 + 4G8, indicating that the protofibrils/oligomers were enriched with peptides ending at amino acid 42. All Aβ peptides found in AD were also present in DS indicating similar pathways of Aβ peptide production, degradation and accumulation, except for APP/Aβ(-X to 15). Likewise, the Aβ peptides forming protofibrils/oligomers in both AD and DS were similar, implying the possibility that treatment with clinical benefit in sporadic AD might also be beneficial for subjects with DS.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.neulet.2021.135894DOI Listing
May 2021

Fluorine-18-Labeled Antibody Ligands for PET Imaging of Amyloid-β in Brain.

ACS Chem Neurosci 2020 12 25;11(24):4460-4468. Epub 2020 Nov 25.

Department of Public Health and Caring Sciences, Rudbeck Laboratory, Uppsala University, Uppsala 751 85, Sweden.

Antibodies are attractive as radioligands due to their outstanding specificity and high affinity, but their inability to cross the blood-brain barrier (BBB) limits their use for CNS targets. To enhance brain distribution, amyloid-β (Aβ) antibodies were fused to a transferrin receptor (TfR) antibody fragment, enabling receptor mediated transport across the BBB. The aim of this study was to label these bispecific antibodies with fluorine-18 and use them for Aβ PET imaging. Bispecific antibody ligands RmAb158-scFv8D3 and Tribody A2, both targeting Aβ and TfR, were functionalized with -cyclooctene (TCO) groups and conjugated with F-labeled tetrazines through an inverse electron demand Diels-Alder reaction performed at ambient temperature. F-labeling did not affect antibody binding , and initial brain uptake was high. Conjugates with the first tetrazine variant ([F]T1) displayed high uptake in bone, indicating extensive defluorination, a problem that was resolved with the second and third tetrazine variants ([F]T2 and [F]T3). Although the antibody ligands' half-life in blood was too long to optimally match the physical half-life of fluorine-18 ( = 110 min), [F]T3-Tribody A2 PET seemed to discriminate transgenic mice (tg-ArcSwe) with Aβ deposits from wild-type mice 12 h after injection. This study demonstrates that F-labeling of bispecific, brain penetrating antibodies is feasible and, with further optimization, could be used for CNS PET imaging.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acschemneuro.0c00652DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7747219PMC
December 2020

Discovery of rare variants associated with blood pressure regulation through meta-analysis of 1.3 million individuals.

Nat Genet 2020 12 23;52(12):1314-1332. Epub 2020 Nov 23.

Department of Clinical Biochemistry, Herlev and Gentofte Hospital, Copenhagen University Hospital, Herlev, Denmark.

Genetic studies of blood pressure (BP) to date have mainly analyzed common variants (minor allele frequency > 0.05). In a meta-analysis of up to ~1.3 million participants, we discovered 106 new BP-associated genomic regions and 87 rare (minor allele frequency ≤ 0.01) variant BP associations (P < 5 × 10), of which 32 were in new BP-associated loci and 55 were independent BP-associated single-nucleotide variants within known BP-associated regions. Average effects of rare variants (44% coding) were ~8 times larger than common variant effects and indicate potential candidate causal genes at new and known loci (for example, GATA5 and PLCB3). BP-associated variants (including rare and common) were enriched in regions of active chromatin in fetal tissues, potentially linking fetal development with BP regulation in later life. Multivariable Mendelian randomization suggested possible inverse effects of elevated systolic and diastolic BP on large artery stroke. Our study demonstrates the utility of rare-variant analyses for identifying candidate genes and the results highlight potential therapeutic targets.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41588-020-00713-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7610439PMC
December 2020

[Alzheimer's disease - the most common cause of dementia].

Lakartidningen 2020 03 9;117. Epub 2020 Mar 9.

Klinisk Neurokemi Lab - Inst. för Neurovetenskap och Fysiologi Mölndal, Sweden Klinisk Neurokemi Lab - Inst. för Neurovetenskap och Fysiologi Mölndal, Sweden.

Alzheimer's disease is the most common cause of dementia. As many as 250,000 people in Sweden will have a dementia disease in 2050. The »amyloid cascade hypothesis« is a common model which explains how β-amyloid affects the function of the nerve cells. Alzheimer's disease has a long-lasting course and can present in typical and atypical forms. CSF analyses for »core AD CSF biomarkers« and synaptic proteins have been available for clinical diagnostics. PET scanning can detect either β-amyloid or tau aggregates in the brain of living humans. Current Alzheimer's disease therapy is based on two classes of cognition-enhancing drugs: acetylcholinesterase inhibitor and NMDA-receptor antagonist, which delays cognitive decline in most patients. The latest clinical development of potential therapy for Alzheimer's is active or passive immunotherapy against brain β-amyloid and tau, where several studies have shown varying but promising treatment effects. Non-pharmacological interventions in patients with AD aim to delay the loss of mental abilities, helping people to be independent in everyday life for as long as possible, and to increase their well-being and quality of life.
View Article and Find Full Text PDF

Download full-text PDF

Source
March 2020

Longitudinal changes in the frequency of mosaic chromosome Y loss in peripheral blood cells of aging men varies profoundly between individuals.

Eur J Hum Genet 2020 03 25;28(3):349-357. Epub 2019 Oct 25.

Department of Immunology, Genetics and Pathology and Science for Life Laboratory, Uppsala University, Uppsala, Sweden.

Mosaic loss of chromosome Y (LOY) is the most common somatic genetic aberration and is associated with increased risk for all-cause mortality, various forms of cancer and Alzheimer's disease, as well as other common human diseases. By tracking LOY frequencies in subjects from which blood samples have been serially collected up to five times during up to 22 years, we observed a pronounced intra-individual variation of changes in the frequency of LOY within individual men over time. We observed that in some individuals the frequency of LOY in blood clearly progressed over time and that in other men, the frequency was constant or showed other types of longitudinal development. The predominant method used for estimating LOY is calculation of the median Log R Ratio of probes located in the male specific part of chromosome Y (mLRRY) from intensity data generated by SNP-arrays, which is difficult to interpret due to its logarithmic and inversed scale. We present here a formula to transform mLRRY-values to percentage of LOY that is a more comprehensible unit. The formula was derived using measurements of LOY from matched samples analysed using SNP-array, whole genome sequencing and a new AMELX/AMELY-based assay for droplet digital PCR. The methods described could be applied for analyses of the vast amount of SNP-array data already generated in the scientific community, allowing further discoveries of LOY associated diseases and outcomes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41431-019-0533-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7028735PMC
March 2020

Longitudinal cognitive decline in autosomal-dominant Alzheimer's disease varies with mutations in APP and PSEN1 genes.

Neurobiol Aging 2019 10 4;82:40-47. Epub 2019 Jul 4.

Division of Neurogeriatrics, Department of Neurobiology Care Sciences and Society, Karolinska Institutet, Stockholm, Sweden.

The purpose was to compare longitudinal cognitive changes in APP and PSEN1 gene mutation carriers and noncarriers from four autosomal-dominant Alzheimer's disease (ADAD) families across preclinical and early clinical stages of disease. Carriers (n = 34) with four different mutations (PSEN1, PSEN1, APP, and APP) and noncarriers (n = 41) were followed up longitudinally with repeated cognitive assessments starting many years before the expected clinical onset. The relationship between cognition and years to expected clinical onset, education, age, and type of mutation was analyzed using mixed-effects models. Results showed an education-dependent and time-related cognitive decline with linear and quadratic predictors in mutation carriers. Cognitive decline began close to the expected clinical onset and was relatively rapid afterward in PSEN1 mutation carriers, whereas decline was slower and started earlier than 10 years before expected clinical onset in APP mutation carriers. In noncarriers, the decline was minimal across time in accordance with normal aging. These results suggest that phenotypes for onset and rate of cognitive decline vary with PSEN1 and APP genes, suggesting a behavioral heterogeneity in ADAD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.neurobiolaging.2019.06.010DOI Listing
October 2019

Aβ and tau prion-like activities decline with longevity in the Alzheimer's disease human brain.

Sci Transl Med 2019 05;11(490)

Institute for Neurodegenerative Diseases, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA 94158, USA.

The hallmarks of Alzheimer's disease (AD) are the accumulation of Aβ plaques and neurofibrillary tangles composed of hyperphosphorylated tau. We developed sensitive cellular assays using human embryonic kidney-293T cells to quantify intracellular self-propagating conformers of Aβ in brain samples from patients with AD or other neurodegenerative diseases. Postmortem brain tissue from patients with AD had measurable amounts of pathological Aβ conformers. Individuals over 80 years of age had the lowest amounts of prion-like Aβ and phosphorylated tau. Unexpectedly, the longevity-dependent decrease in self-propagating tau conformers occurred in spite of increasing amounts of total insoluble tau. When corrected for the abundance of insoluble tau, the ability of postmortem AD brain homogenates to induce misfolded tau in the cellular assays showed an exponential decrease with longevity, with a half-life of about one decade over the age range of 37 to 99 years. Thus, our findings demonstrate an inverse correlation between longevity in patients with AD and the abundance of pathological tau conformers. Our cellular assays can be applied to patient selection for clinical studies and the development of new drugs and diagnostics for AD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/scitranslmed.aat8462DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6640844PMC
May 2019

Genetic meta-analysis of diagnosed Alzheimer's disease identifies new risk loci and implicates Aβ, tau, immunity and lipid processing.

Nat Genet 2019 03 28;51(3):414-430. Epub 2019 Feb 28.

Research Center and Memory Clinic of Fundació ACE, Institut Català de Neurociències Aplicades-Universitat Internacional de Catalunya, Barcelona, Spain.

Risk for late-onset Alzheimer's disease (LOAD), the most prevalent dementia, is partially driven by genetics. To identify LOAD risk loci, we performed a large genome-wide association meta-analysis of clinically diagnosed LOAD (94,437 individuals). We confirm 20 previous LOAD risk loci and identify five new genome-wide loci (IQCK, ACE, ADAM10, ADAMTS1, and WWOX), two of which (ADAM10, ACE) were identified in a recent genome-wide association (GWAS)-by-familial-proxy of Alzheimer's or dementia. Fine-mapping of the human leukocyte antigen (HLA) region confirms the neurological and immune-mediated disease haplotype HLA-DR15 as a risk factor for LOAD. Pathway analysis implicates immunity, lipid metabolism, tau binding proteins, and amyloid precursor protein (APP) metabolism, showing that genetic variants affecting APP and Aβ processing are associated not only with early-onset autosomal dominant Alzheimer's disease but also with LOAD. Analyses of risk genes and pathways show enrichment for rare variants (P = 1.32 × 10), indicating that additional rare variants remain to be identified. We also identify important genetic correlations between LOAD and traits such as family history of dementia and education.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41588-019-0358-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6463297PMC
March 2019

Improved Differential Diagnosis of Alzheimer's Disease by Integrating ELISA and Mass Spectrometry-Based Cerebrospinal Fluid Biomarkers.

J Alzheimers Dis 2019 ;67(2):639-651

Department of Medical Sciences, Clinical Chemistry, Uppsala University, Uppsala, Sweden.

Background: Alzheimer's disease (AD) is diagnosed based on a clinical evaluation as well as analyses of classical biomarkers: Aβ42, total tau (t-tau), and phosphorylated tau (p-tau) in cerebrospinal fluid (CSF). Although the sensitivities and specificities of the classical biomarkers are fairly good for detection of AD, there is still a need to develop novel biochemical markers for early detection of AD.

Objective: We explored if integration of novel proteins with classical biomarkers in CSF can better discriminate AD from non-AD subjects.

Methods: We applied ELISA, mass spectrometry, and multivariate modeling to investigate classical biomarkers and the CSF proteome in subjects (n = 206) with 76 AD patients, 74 mild cognitive impairment (MCI) patients, 11 frontotemporal dementia (FTD) patients, and 45 non-dementia controls. The MCI patients were followed for 4-9 years and 21 of these converted to AD, whereas 53 remained stable.

Results: By combining classical CSF biomarkers with twelve novel markers, the area of the ROC curves (AUROCS) of distinguishing AD and MCI/AD converters from non-AD were 93% and 96%, respectively. The FTDs and non-dementia controls were identified versus all other groups with AUROCS of 96% and 87%, respectively.

Conclusions: Integration of new and classical CSF biomarkers in a model-based approach can improve the identification of AD, FTD, and non-dementia control subjects.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3233/JAD-180855DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6398544PMC
March 2020

Alzheimer's disease pathology propagation by exosomes containing toxic amyloid-beta oligomers.

Acta Neuropathol 2018 07 13;136(1):41-56. Epub 2018 Jun 13.

Department of Pathology, Department of Clinical and Experimental Medicine, Linköping University, Linköping, Sweden.

The gradual deterioration of cognitive functions in Alzheimer's disease is paralleled by a hierarchical progression of amyloid-beta and tau brain pathology. Recent findings indicate that toxic oligomers of amyloid-beta may cause propagation of pathology in a prion-like manner, although the underlying mechanisms are incompletely understood. Here we show that small extracellular vesicles, exosomes, from Alzheimer patients' brains contain increased levels of amyloid-beta oligomers and can act as vehicles for the neuron-to-neuron transfer of such toxic species in recipient neurons in culture. Moreover, blocking the formation, secretion or uptake of exosomes was found to reduce both the spread of oligomers and the related toxicity. Taken together, our results imply that exosomes are centrally involved in Alzheimer's disease and that they could serve as targets for development of new diagnostic and therapeutic principles.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00401-018-1868-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6015111PMC
July 2018

CRISPR/Cas9 Mediated Disruption of the Swedish APP Allele as a Therapeutic Approach for Early-Onset Alzheimer's Disease.

Mol Ther Nucleic Acids 2018 Jun 16;11:429-440. Epub 2018 Mar 16.

Departments of Neurology and Radiology, Massachusetts General Hospital and Center for NeuroDiscovery, Harvard Medical School, Boston, MA, USA; Department of Public Health and Caring Sciences, Geriatrics, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden. Electronic address:

The APPswe (Swedish) mutation in the amyloid precursor protein (APP) gene causes dominantly inherited Alzheimer's disease (AD) as a result of increased β-secretase cleavage of the amyloid-β (Aβ) precursor protein. This leads to abnormally high Aβ levels, not only in brain but also in peripheral tissues of mutation carriers. Here, we selectively disrupted the human mutant APP allele using CRISPR. By applying CRISPR/Cas9 from Streptococcus pyogenes, we generated allele-specific deletions of either APP or APP. As measured by ELISA, conditioned media of targeted patient-derived fibroblasts displayed an approximate 60% reduction in secreted Aβ. Next, coding sequences for the APP-specific guide RNA (gRNA) and Cas9 were packaged into separate adeno-associated viral (AAV) vectors. Site-specific indel formation was achieved both in primary neurons isolated from APP transgenic mouse embryos (Tg2576) and after co-injection of these vectors into hippocampus of adult mice. Taken together, we here present proof-of-concept data that CRISPR/Cas9 can selectively disrupt the APP allele both ex vivo and in vivo-and thereby decrease pathogenic Aβ. Hence, this system may have the potential to be developed as a tool for gene therapy against AD caused by APPswe and other point mutations associated with increased Aβ.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.omtn.2018.03.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5992788PMC
June 2018

Antibody-Based In Vivo PET Imaging Detects Amyloid-β Reduction in Alzheimer Transgenic Mice After BACE-1 Inhibition.

J Nucl Med 2018 12 31;59(12):1885-1891. Epub 2018 May 31.

Department of Public Health and Caring Sciences/Geriatrics, Uppsala University, Uppsala, Sweden

Visualization of amyloid-β (Aβ) pathology with PET has become an important tool for making a specific clinical diagnosis of Alzheimer disease (AD). However, the available amyloid PET radioligands, such as C-Pittsburgh compound B, reflect levels of insoluble Aβ plaques but do not capture soluble and protofibrillar Aβ forms. Furthermore, the plaque load appears to be fairly static during clinical stages of AD and may not be affected by Aβ-reducing treatments. The aim of the present study was to investigate whether a novel PET radioligand based on an antibody directed toward soluble aggregates of Aβ can be used to detect changes in Aβ levels during disease progression and after treatment with a β-secretase (BACE-1) inhibitor. One set of transgenic mice (tg-ArcSwe, a model of Aβ pathology) aged between 7 and 16 mo underwent PET with the Aβ protofibril-selective radioligand I-RmAb158-scFv8D3 (where RmAb is recombinant mouse monoclonal antibody and scFv is single-chain variable fragment) to follow progression of Aβ pathology in the brain. A second set of tg-ArcSwe mice, aged 10 mo, were treated with the BACE-1 inhibitor NB-360 for 3 mo and compared with an untreated control group. A third set of tg-ArcSwe mice, also aged 10 mo, underwent PET as a baseline group. Brain tissue was isolated after PET to determine levels of Aβ by ELISA and immunohistochemistry. The concentration of I-RmAb158-scFv8D3, as measured in vivo with PET, increased with age and corresponded well with the ex vivo autoradiography and Aβ immunohistochemistry results. Mice treated with NB-360 showed significantly lower in vivo PET signals than untreated animals and were similar to the baseline animals. The decreased I-RmAb158-scFv8D3 concentrations in NB-360-treated mice, as quantified with PET, corresponded well with the decreased Aβ levels measured in postmortem brain. Several treatments for AD are in phase 2 and 3 clinical trials, but the possibility of studying treatment effects in vivo on the important, nonfibrillar, forms of Aβ is limited. This study demonstrated the ability of the Aβ protofibril-selective radioligand I-RmAb158-scFv8D3 to follow disease progression and detect treatment effects with PET imaging in tg-ArcSwe mice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2967/jnumed.118.213140DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6278900PMC
December 2018

Efficient clearance of Aβ protofibrils in AβPP-transgenic mice treated with a brain-penetrating bifunctional antibody.

Alzheimers Res Ther 2018 05 24;10(1):49. Epub 2018 May 24.

Department of Public Health and Caring Sciences/Geriatrics, Uppsala University, Rudbeck Laboratory, Dag Hammarskjölds väg 20, 75185, Uppsala, Sweden.

Background: Amyloid-β (Aβ) immunotherapy is one of the most promising disease-modifying strategies for Alzheimer's disease (AD). Despite recent progress targeting aggregated forms of Aβ, low antibody brain penetrance remains a challenge. In the present study, we used transferrin receptor (TfR)-mediated transcytosis to facilitate brain uptake of our previously developed Aβ protofibril-selective mAb158, with the aim of increasing the efficacy of immunotherapy directed toward soluble Aβ protofibrils.

Methods: Aβ protein precursor (AβPP)-transgenic mice (tg-ArcSwe) were given a single dose of mAb158, modified for TfR-mediated transcytosis (RmAb158-scFv8D3), in comparison with an equimolar dose or a tenfold higher dose of unmodified recombinant mAb158 (RmAb158). Soluble Aβ protofibrils and total Aβ in the brain were measured by enzyme-linked immunosorbent assay (ELISA). Brain distribution of radiolabeled antibodies was visualized by positron emission tomography (PET) and ex vivo autoradiography.

Results: ELISA analysis of Tris-buffered saline brain extracts demonstrated a 40% reduction of soluble Aβ protofibrils in both RmAb158-scFv8D3- and high-dose RmAb158-treated mice, whereas there was no Aβ protofibril reduction in mice treated with a low dose of RmAb158. Further, ex vivo autoradiography and PET imaging revealed different brain distribution patterns of RmAb158-scFv8D3 and RmAb158, suggesting that these antibodies may affect Aβ levels by different mechanisms.

Conclusions: With a combination of biochemical and imaging analyses, this study demonstrates that antibodies engineered to be transported across the blood-brain barrier can be used to increase the efficacy of Aβ immunotherapy. This strategy may allow for decreased antibody doses and thereby reduced side effects and treatment costs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13195-018-0377-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5968497PMC
May 2018

The Aβ protofibril selective antibody mAb158 prevents accumulation of Aβ in astrocytes and rescues neurons from Aβ-induced cell death.

J Neuroinflammation 2018 Mar 28;15(1):98. Epub 2018 Mar 28.

Department of Public Health and Caring Sciences, Molecular Geriatrics, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden.

Background: Currently, several amyloid beta (Aβ) antibodies, including the protofibril selective antibody BAN2401, are in clinical trials. The murine version of BAN2401, mAb158, has previously been shown to lower the levels of pathogenic Aβ and prevent Aβ deposition in animal models of Alzheimer's disease (AD). However, the cellular mechanisms of the antibody's action remain unknown. We have recently shown that astrocytes effectively engulf Aβ protofibrils, but store rather than degrade the ingested Aβ aggregates. In a co-culture set-up, the incomplete degradation of Aβ protofibrils by astrocytes results in increased neuronal cell death, due to the release of extracellular vesicles, containing N-truncated, neurotoxic Aβ.

Methods: The aim of the present study was to investigate if the accumulation of Aβ in astrocytes can be affected by the Aβ protofibril selective antibody mAb158. Co-cultures of astrocytes, neurons, and oligodendrocytes, derived from embryonic mouse cortex, were exposed to Aβ protofibrils in the presence or absence of mAb158.

Results: Our results demonstrate that the presence of mAb158 almost abolished Aβ accumulation in astrocytes. Consequently, mAb158 treatment rescued neurons from Aβ-induced cell death.

Conclusion: Based on these findings, we conclude that astrocytes may play a central mechanistic role in anti-Aβ immunotherapy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12974-018-1134-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5875007PMC
March 2018

Structural heterogeneity and intersubject variability of Aβ in familial and sporadic Alzheimer's disease.

Proc Natl Acad Sci U S A 2018 01 8;115(4):E782-E791. Epub 2018 Jan 8.

Institute for Neurodegenerative Diseases, Weill Institute for Neurosciences, University of California, San Francisco, CA 94158;

Point mutations in the amyloid-β (Aβ) coding region produce a combination of mutant and WT Aβ isoforms that yield unique clinicopathologies in familial Alzheimer's disease (fAD) and cerebral amyloid angiopathy (fCAA) patients. Here, we report a method to investigate the structural variability of amyloid deposits found in fAD, fCAA, and sporadic AD (sAD). Using this approach, we demonstrate that mutant Aβ determines WT Aβ conformation through prion template-directed misfolding. Using principal component analysis of multiple structure-sensitive fluorescent amyloid-binding dyes, we assessed the conformational variability of Aβ deposits in fAD, fCAA, and sAD patients. Comparing many deposits from a given patient with the overall population, we found that intrapatient variability is much lower than interpatient variability for both disease types. In a given brain, we observed one or two structurally distinct forms. When two forms coexist, they segregate between the parenchyma and cerebrovasculature, particularly in fAD patients. Compared with sAD samples, deposits from fAD patients show less intersubject variability, and little overlap exists between fAD and sAD deposits. Finally, we examined whether E22G (Arctic) or E22Q (Dutch) mutants direct the misfolding of WT Aβ, leading to fAD-like plaques in vivo. Intracerebrally injecting mutant Aβ40 fibrils into transgenic mice expressing only WT Aβ induced the deposition of plaques with many biochemical hallmarks of fAD. Thus, mutant Aβ40 prions induce a conformation of WT Aβ similar to that found in fAD deposits. These findings indicate that diverse AD phenotypes likely arise from one or more initial Aβ prion conformations, which kinetically dominate the spread of prions in the brain.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1714966115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5789926PMC
January 2018

Sequence data and association statistics from 12,940 type 2 diabetes cases and controls.

Sci Data 2017 12 19;4:170179. Epub 2017 Dec 19.

Wellcome Trust Centre for Human Genetics, Nuffield Department of Medicine, University of Oxford, Oxford, UK.

To investigate the genetic basis of type 2 diabetes (T2D) to high resolution, the GoT2D and T2D-GENES consortia catalogued variation from whole-genome sequencing of 2,657 European individuals and exome sequencing of 12,940 individuals of multiple ancestries. Over 27M SNPs, indels, and structural variants were identified, including 99% of low-frequency (minor allele frequency [MAF] 0.1-5%) non-coding variants in the whole-genome sequenced individuals and 99.7% of low-frequency coding variants in the whole-exome sequenced individuals. Each variant was tested for association with T2D in the sequenced individuals, and, to increase power, most were tested in larger numbers of individuals (>80% of low-frequency coding variants in ~82 K Europeans via the exome chip, and ~90% of low-frequency non-coding variants in ~44 K Europeans via genotype imputation). The variants, genotypes, and association statistics from these analyses provide the largest reference to date of human genetic information relevant to T2D, for use in activities such as T2D-focused genotype imputation, functional characterization of variants or genes, and other novel analyses to detect associations between sequence variation and T2D.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/sdata.2017.179DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5735917PMC
December 2017

Alpha-synuclein oligomer-selective antibodies reduce intracellular accumulation and mitochondrial impairment in alpha-synuclein exposed astrocytes.

J Neuroinflammation 2017 Dec 11;14(1):241. Epub 2017 Dec 11.

Molecular Geriatrics, Department of Public Health and Caring Sciences, Rudbeck Laboratory, Uppsala University, SE-751 85, Uppsala, Sweden.

Background: Due to its neurotoxic properties, oligomeric alpha-synuclein (α-syn) has been suggested as an attractive target for passive immunization against Parkinson's disease (PD). In mouse models of PD, antibody treatment has been shown to lower the levels of pathogenic α-syn species, including oligomers, although the mechanisms of action remain unknown. We have previously shown that astrocytes rapidly engulf α-syn oligomers that are intracellularly stored, rather than degraded, resulting in impaired mitochondria.

Methods: The aim of the present study was to investigate if the accumulation of α-syn in astrocytes can be affected by α-syn oligomer-selective antibodies. Co-cultures of astrocytes, neurons, and oligodendrocytes were derived from embryonic mouse cortex and exposed to α-syn oligomers or oligomers pre-incubated with oligomer-selective antibodies.

Results: In the presence of antibodies, the astrocytes displayed an increased clearance of the exogenously added α-syn, and consequently, the α-syn accumulation in the culture was markedly reduced. Moreover, the addition of antibodies rescued the astrocytes from the oligomer-induced mitochondrial impairment.

Conclusions: Our results demonstrate that oligomer-selective antibodies can prevent α-syn accumulation and mitochondrial dysfunction in cultured astrocytes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12974-017-1018-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5725978PMC
December 2017

Identification of a novel proinsulin-associated SNP and demonstration that proinsulin is unlikely to be a causal factor in subclinical vascular remodelling using Mendelian randomisation.

Atherosclerosis 2017 Nov 28;266:196-204. Epub 2017 Sep 28.

Cardiovascular Medicine Unit, Department of Medicine Solna, Karolinska Institutet, Stockholm, Sweden.

Background And Aims: Increased proinsulin relative to insulin levels have been associated with subclinical atherosclerosis (measured by carotid intima-media thickness (cIMT)) and are predictive of future cardiovascular disease (CVD), independently of established risk factors. The mechanisms linking proinsulin to atherosclerosis and CVD are unclear. A genome-wide meta-analysis has identified nine loci associated with circulating proinsulin levels. Using proinsulin-associated SNPs, we set out to use a Mendelian randomisation approach to test the hypothesis that proinsulin plays a causal role in subclinical vascular remodelling.

Methods: We studied the high CVD-risk IMPROVE cohort (n = 3345), which has detailed biochemical phenotyping and repeated, state-of-the-art, high-resolution carotid ultrasound examinations. Genotyping was performed using Illumina Cardio-Metabo and Immuno arrays, which include reported proinsulin-associated loci. Participants with type 2 diabetes (n = 904) were omitted from the analysis. Linear regression was used to identify proinsulin-associated genetic variants.

Results: We identified a proinsulin locus on chromosome 15 (rs8029765) and replicated it in data from 20,003 additional individuals. An 11-SNP score, including the previously identified and the chromosome 15 proinsulin-associated loci, was significantly and negatively associated with baseline IMT and IMT (the primary cIMT phenotypes) but not with progression measures. However, MR-Eggers refuted any significant effect of the proinsulin-associated 11-SNP score, and a non-pleiotropic SNP score of three variants (including rs8029765) demonstrated no effect on baseline or progression cIMT measures.

Conclusions: We identified a novel proinsulin-associated locus and demonstrated that whilst proinsulin levels are associated with cIMT measures, proinsulin per se is unlikely to have a causative effect on cIMT.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.atherosclerosis.2017.09.031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5679136PMC
November 2017

Efficient and inexpensive transient expression of multispecific multivalent antibodies in Expi293 cells.

Biol Proced Online 2017 15;19:11. Epub 2017 Sep 15.

Department of Public Health and Caring Sciences, Uppsala University, Rudbeck laboratory, Dag Hammarskjöldsväg 20, 751 85 Uppsala, Sweden.

Background: Immunotherapy is a very fast expanding field within drug discovery and, hence, rapid and inexpensive expression of antibodies would be extremely valuable. Antibodies are, however, difficult to express. Multifunctional antibodies with additional binding domains further complicate the expression. Only few protocols describe the production of tetravalent bispecific antibodies and all with limited expression levels..

Methods: Here, we describe a protocol that can produce functional tetravalent, bispecific antibodies at around 22 mg protein/l to a low cost. The expression system is based on the Expi293 cells, which have been adapted to grow in denser cultures than HEK293 cells and gives higher expression yields. The new protocol transfects the Expi293 cells with PEI (which has a negligible cost).

Results: The protocol has been used to generate multiple variants of tetra- and hexavalent bispecific antibodies with yields of around 22 mg protein/l within 10 days. All materials are commercially available and the implementation of the protocol is inexpensive and straightforward. The bispecific antibodies generated in our lab were capable of binding to all antigens with similar affinity as the original antibody. Two of the bispecific antibodies have also been used in transgenic mice as positron emission tomography (PET) ligands to successfully detect amyloid-beta (Aβ) aggregates .

Conclusions: This protocol is the first describing transfection of the human Expi293 cells with PEI. It can be used to generate functional multi-specific antibodies in high amounts. The use of biological drugs, and in particular multispecific antibodies, is rapidly increasing, hence improved protocols such as the one presented here are highly valuable.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12575-017-0060-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5603040PMC
September 2017
-->