ABSTRACT
In neurodegenerative diseases like Alzheimer's disease (AD), endogenous proteins or peptides aggregate with themselves. These proteins may lose their function or aggregates and/or oligomers can obtain toxicity, causing injury or death to cells. Aggregation of two major proteins characterizes AD. Amyloid-ß peptide (Aß) is deposited in amyloid plaques within the extracellular space of the brain and Tau in so-called neurofibrillary tangles in neurons. Finding peptide ligands to halt protein aggregation is a promising therapeutical approach. Using mirror-image phage display with a commercially available, randomized 12-mer peptide library, we have selected D-amino acid peptides, which bind to the Tau protein and modulate its aggregation in vitro. Peptides can bind specifically and selectively to a target molecule, but natural L-amino acid peptides may have crucial disadvantages for in vivo applications, as they are sensitive to protease degradation and may elicit immune responses. One strategy to circumvent these disadvantages is the use of non-naturally occurring D-amino acid peptides as they exhibit increased protease resistance and generally do not activate the immune system. To perform mirror-image phage display, the target protein needs to be synthesized as D-amino acid version. If the target protein sequence is too long to be synthesized properly, smaller peptides derived from the full length protein can be used for the selection process. This also offers the possibility to influence the binding region of the selected D-peptides in the full-length target protein. Here we provide the protocols for mirror-image phage display selection on the PHF6* peptide of Tau, based on the commercially available Ph.D.™-12 Phage Display Peptide Library Kit, leading to D-peptides that also bind the full length Tau protein (Tau441), next to PHF6*. In addition, we provide protocols and data for the first characterization of those D-peptides that inhibit Tau aggregation in vitro. © 2024 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Mirror image phage display selection against D-PHF6* fibrils Support Protocol 1: Single phage ELISA Basic Protocol 2: Sequencing and D-peptide generation Basic Protocol 3: Thioflavin-T (ThT) test to control inhibition of Tau aggregation Support Protocol 2: Purification of full-length Tau protein Basic Protocol 4: ELISA to demonstrate the binding of the generated D-peptides to PHF6* and full-length Tau fibrils.
Subject(s)
Alzheimer Disease , Bacteriophages , Humans , tau Proteins/genetics , tau Proteins/chemistry , tau Proteins/metabolism , Amino Acids , Peptide Library , Amyloid beta-Peptides/chemistry , Amyloid beta-Peptides/genetics , Amyloid beta-Peptides/metabolism , Alzheimer Disease/drug therapy , Alzheimer Disease/metabolism , Peptide Hydrolases/metabolism , Bacteriophages/metabolismABSTRACT
Alzheimer disease (AD) is the most common progressive neurodegenerative disorder. AD causes enormous personal and economic burden to society as currently only limited palliative therapeutic options are available. The pathological hallmarks of the disease are extracellular plaques, composed of fibrillar amyloid-ß (Aß), and neurofibrillary tangles inside neurons, composed of Tau protein. Until recently, the search for AD therapeutics was focussed more on the Aß peptide and its pathology, but the results were unsatisfying. As an alternative, Tau might be a promising therapeutic target as its pathology is closely correlated to clinical symptoms. In addition, pathological Tau aggregation occurs in a large group of diseases, called Tauopathies, and in most of them Aß aggregation does not play a role in disease pathogenesis. The formation of Tau aggregates is triggered by two hexapeptide motifs within Tau; PHF6* and PHF6. Both fragments are interesting targets for the development of Tau aggregation inhibitors (TAI). Peptides represent a unique class of pharmaceutical compounds and are reasonable alternatives to chemical substances or antibodies. They are attributed with high biological activity, valuable specificity and low toxicity, and often are developed as drug candidates to interrupt protein-protein interactions. The preparation of peptides is simple, controllable and the peptides can be easily modified. However, their application may also have disadvantages. Currently, a few peptide compounds acting as TAI are described in the literature, most of them developed by structure-based design or phage display. Here, we review the current state of research in this promising field of AD therapy development.
Subject(s)
Alzheimer Disease , Tauopathies , Humans , Alzheimer Disease/metabolism , tau Proteins/metabolism , Tauopathies/metabolism , Neurofibrillary Tangles/metabolism , Amyloid beta-Peptides/metabolismABSTRACT
BACKGROUND: Alzheimer's disease (AD), the most common form of dementia, is a progressive neurodegenerative disorder that mainly affects older adults. One of the pathological hallmarks of AD is abnormally aggregated Tau protein that forms fibrillar deposits in the brain. In AD, Tau pathology correlates strongly with clinical symptoms, cognitive dysfunction, and neuronal death. METHODS: We aimed to develop novel therapeutic D-amino acid peptides as Tau fibrillization inhibitors. It has been previously demonstrated that D-amino acid peptides are protease stable and less immunogenic than L-peptides, and these characteristics may render them suitable for in vivo applications. Using a phage display procedure against wild type full-length Tau (TauFL), we selected a novel Tau binding L-peptide and synthesized its D-amino acid version ISAD1 and its retro inversed form, ISAD1rev, respectively. RESULTS: While ISAD1rev inhibited Tau aggregation only moderately, ISAD1 bound to Tau in the aggregation-prone PHF6 region and inhibited fibrillization of TauFL, disease-associated mutant full-length Tau (TauFLΔK, TauFL-A152T, TauFL-P301L), and pro-aggregant repeat domain Tau mutant (TauRDΔK). ISAD1 and ISAD1rev induced the formation of large high molecular weight TauFL and TauRDΔK oligomers that lack proper Thioflavin-positive ß-sheet conformation even at lower concentrations. In silico modeling of ISAD1 Tau interaction at the PHF6 site revealed a binding mode similar to those known for other PHF6 binding peptides. Cell culture experiments demonstrated that ISAD1 and its inverse form are taken up by N2a-TauRDΔK cells efficiently and prevent cytotoxicity of externally added Tau fibrils as well as of internally expressed TauRDΔK. CONCLUSIONS: ISAD1 and related peptides may be suitable for therapy development of AD by promoting off-pathway assembly of Tau, thus preventing its toxicity.
Subject(s)
Alzheimer Disease , Peptides , tau Proteins , Aged , Alzheimer Disease/drug therapy , Alzheimer Disease/pathology , Amino Acids/therapeutic use , Cells, Cultured , Humans , Peptides/therapeutic use , Protein Conformation, beta-Strand , tau Proteins/metabolism , tau Proteins/toxicityABSTRACT
Alzheimer's disease and other Tauopathies are associated with neurofibrillary tangles composed of Tau protein, as well as toxic Tau oligomers. Therefore, inhibitors of pathological Tau aggregation are potentially useful candidates for future therapies targeting Tauopathies. Two hexapeptides within Tau, designated PHF6* (275-VQIINK-280) and PHF6 (306-VQIVYK-311), are known to promote Tau aggregation. Recently, the PHF6* segment has been described as the more potent driver of Tau aggregation. We therefore employed mirror-image phage display with a large peptide library to identify PHF6* fibril binding peptides consisting of D-enantiomeric amino acids. The suitability of D-enantiomeric peptides for inâ vivo applications, which are protease stable and less immunogenic than L-peptides, has already been demonstrated. The identified D-enantiomeric peptide MMD3 and its retro-inverso form, designated MMD3rev, inhibited inâ vitro fibrillization of the PHF6* peptide, the repeat domain of Tau as well as full-length Tau. Dynamic light scattering, pelleting assays and atomic force microscopy demonstrated that MMD3 prevents the formation of tau ß-sheet-rich fibrils by diverting Tau into large amorphous aggregates. NMR data suggest that the D-enantiomeric peptides bound to Tau monomers with rather low affinity, but ELISA (enzyme-linked immunosorbent assay) data demonstrated binding to PHF6* and full length Tau fibrils. In addition, molecular insight into the binding mode of MMD3 to PHF6* fibrils were gained by in silico modelling. The identified PHF6*-targeting peptides were able to penetrate cells. The study establishes PHF6* fibril binding peptides consisting of D-enantiomeric amino acids as potential molecules for therapeutic and diagnostic applications in AD research.
Subject(s)
Peptides/pharmacology , tau Proteins/antagonists & inhibitors , Humans , Peptide Library , Peptides/chemistry , Protein Aggregates/drug effects , tau Proteins/metabolismABSTRACT
A variety of neurodegenerative disorders, including Alzheimer disease (AD), are associated with neurofibrillary tangles composed of the tau protein, as well as toxic tau oligomers. Inhibitors of pathological tau aggregation, interrupting tau self-assembly, might be useful for the development of therapeutics. Employing mirror image phage display with a large peptide library (over 109 different peptides), we have identified tau fibril binding peptides consisting of d-enantiomeric amino acids. d-enantiomeric peptides are extremely protease stable and not or less immunogenic than l-peptides, and the suitability of d-peptides for in vivo applications have already been demonstrated. Phage display selections were performed using fibrils of the d-enantiomeric hexapeptide VQIVYK, representing residues 306 to 311 of the tau protein, as a target. VQIVYK has been demonstrated to be important for fibril formation of the full lengths protein and forms fibrils by itself. Here, we report on d-enantiomeric peptides, which bind to VQIVYK, tau isoforms like tau3RD (K19) as well as to full lengths tau fibrils, and modulate the aggregation of the respective tau form. The peptides are able to penetrate cells and might be interesting for therapeutic and diagnostic applications in AD research.
Subject(s)
Oligopeptides/metabolism , tau Proteins/metabolism , Alzheimer Disease/drug therapy , Alzheimer Disease/metabolism , Alzheimer Disease/pathology , Amino Acid Sequence , Binding Sites , Dynamic Light Scattering , Fluoresceins/chemistry , Humans , Molecular Dynamics Simulation , Oligopeptides/chemistry , Oligopeptides/therapeutic use , Peptide Library , Protein Aggregates , Protein Binding , Protein Isoforms/chemistry , Protein Isoforms/metabolism , Protein Structure, Secondary , Recombinant Proteins/biosynthesis , Recombinant Proteins/chemistry , Recombinant Proteins/isolation & purification , Stereoisomerism , tau Proteins/chemistry , tau Proteins/geneticsABSTRACT
[This corrects the article DOI: 10.1371/journal.pone.0147470.].
ABSTRACT
Protein misfolding and aggregation are fundamental features of the majority of neurodegenerative diseases, like Alzheimer's disease (AD), Parkinson's disease, frontotemporal dementia, and prion diseases. Proteinaceous deposits in the brain of the patient, e.g., amyloid plaques consisting of the amyloid-ß (Aß) peptide and tangles composed of tau protein, are the hallmarks of AD. Soluble oligomers of Aß and tau play a fundamental role in disease progression, and specific detection and quantification of the respective oligomeric proteins in cerebrospinal fluid may provide presymptomatically detectable biomarkers, paving the way for early diagnosis or even prognosis. Several studies on the development of techniques for the specific detection of Aß oligomers were published, but some of the existing tools do not yet seem to be satisfactory, and the study results are contradicting. The detection of oligomers is challenging due to their polymorphous and unstable nature, their low concentration, and the presence of competing proteins and Aß monomers in body fluids. Here, we present an overview of the current state of the development of methods for Aß oligomer specific detection and quantitation. The methods are divided in the three subgroups: (i) enzyme linked immunosorbent assays (ELISA), (ii) methods for single oligomer detection, and (iii) others, which are mainly biosensor based methods.
Subject(s)
Alzheimer Disease/cerebrospinal fluid , Amyloid beta-Peptides/analysis , Amyloid beta-Peptides/cerebrospinal fluid , Biosensing Techniques , Peptide Fragments/cerebrospinal fluid , Enzyme-Linked Immunosorbent Assay , Humans , tau Proteins/analysis , tau Proteins/cerebrospinal fluidABSTRACT
The aggregation of amyloid-ß (Aß) is postulated to be the crucial event in Alzheimer's disease (AD). In particular, small neurotoxic Aß oligomers are considered to be responsible for the development and progression of AD. Therefore, elimination of thesis oligomers represents a potential causal therapy of AD. Starting from the well-characterized d-enantiomeric peptide D3, we identified D3 derivatives that bind monomeric Aß. The underlying hypothesis is that ligands bind monomeric Aß and stabilize these species within the various equilibria with Aß assemblies, leading ultimately to the elimination of Aß oligomers. One of the hereby identified d-peptides, DB3, and a head-to-tail tandem of DB3, DB3DB3, were studied in detail. Both peptides were found to: (i) inhibit the formation of Thioflavin T-positive fibrils; (ii) bind to Aß monomers with micromolar affinities; (iii) eliminate Aß oligomers; (iv) reduce Aß-induced cytotoxicity; and (v) disassemble preformed Aß aggregates. The beneficial effects of DB3 were improved by DB3DB3, which showed highly enhanced efficacy. Our approach yielded Aß monomer-stabilizing ligands that can be investigated as a suitable therapeutic strategy against AD.
Subject(s)
Amyloid beta-Peptides/chemistry , Oligopeptides/chemistry , Amino Acid Sequence , Animals , Cell Line, Tumor , Enzyme-Linked Immunosorbent Assay , RatsABSTRACT
Alzheimer´s disease is the most prominent type of dementia and currently no causative treatment is available. According to recent studies, oligomeric species of the amyloid beta (Aß) peptide appear to be the most toxic Aß assemblies. Aß monomers, however, may be not toxic per se and may even have a neuroprotective role. Here we describe a competitive mirror image phage display procedure that allowed us to identify preferentially Aß1-42 monomer binding and thereby stabilizing peptides, which destabilize and thereby eliminate toxic oligomer species. One of the peptides, called Mosd1 (monomer specific d-peptide 1), was characterized in more detail. Mosd1 abolished oligomers from a mixture of Aß1-42 species, reduced Aß1-42 toxicity in cell culture, and restored the physiological phenotype in neuronal cells stably transfected with the gene coding for human amyloid precursor protein.
Subject(s)
Amyloid beta-Peptides/metabolism , Amyloid beta-Peptides/toxicity , Cell Surface Display Techniques , Peptides/pharmacology , Protein Aggregates/drug effects , Protein Aggregation, Pathological , Amyloid Precursor Protein Secretases/metabolism , Cell Line , Cell Survival/drug effects , HumansABSTRACT
Recent evidence suggests Alzheimer-Disease (AD) to be driven by aggregated Aß. Capitalizing on the mechanism of molecular mimicry and applying several selection layers, we screened peptide libraries for moieties inducing antibodies selectively reacting with Aß-aggregates. The technology identified a pool of peptide candidates; two, AFFITOPES AD01 and AD02, were assessed as vaccination antigens and compared to Aß1-6, the targeted epitope. When conjugated to Keyhole Limpet Hemocyanin (KLH) and adjuvanted with aluminum, all three peptides induced Aß-targeting antibodies (Abs). In contrast to Aß1-6, AD01- or AD02-induced Abs were characterized by selectivity for aggregated forms of Aß and absence of reactivity with related molecules such as Amyloid Precursor Protein (APP)/ secreted APP-alpha (sAPPa). Administration of AFFITOPE-vaccines to APP-transgenic mice was found to reduce their cerebral amyloid burden, the associated neuropathological alterations and to improve their cognitive functions. Thus, the AFFITOME-technology delivers vaccines capable of inducing a distinct Ab response. Their features may be beneficial to AD-patients, a hypothesis currently tested within a phase-II-study.
Subject(s)
Alzheimer Disease/therapy , Alzheimer Vaccines/therapeutic use , Amyloid beta-Peptides/immunology , Antibodies/immunology , Alzheimer Disease/immunology , Alzheimer Vaccines/immunology , Animals , Female , Mice , Mice, Inbred C57BL , Peptide LibraryABSTRACT
BACKGROUND: Amyloid fibrils such as Semen-Derived Enhancer of Viral Infection (SEVI) or amyloid-ß-peptide (Aß) enhance HIV-1 attachment and entry. Inhibitors destroying or converting those fibrils into non-amyloidogenic aggregates effectively reduce viral infectivity. Thus, they seem to be suitable as therapeutic drugs expanding the current HIV-intervening repertoire of antiretroviral compounds. FINDINGS: In this study, we demonstrate that the small D-amino acid peptide D3, which was investigated for therapeutic studies on Alzheimer's disease (AD), significantly reduces both SEVI and Aß fibril boosted infectivity of HIV-1. CONCLUSIONS: Since amyloids could play an important role in the progression of AIDS dementia complex (ADC), the treatment of HIV-1 infected individuals with D3, that inhibits Aß fibril formation and converts preformed Aß fibrils into non-amyloidogenic and non-fibrillar aggregates, may reduce the vulnerability of the central nervous system of HIV patients for HIV associated neurological disorders.
ABSTRACT
Alzheimer's disease (AD) is a progressive neurodegenerative disorder with devastating effects. Currently, therapeutic options are limited to symptomatic treatment. For more than a decade, research focused on immunotherapy for the causal treatment of AD. However, clinical trials with active immunization using Aß encountered severe complications, for example meningoencephalitis. Consequently, attention focused on passive immunization using antibodies. As an alternative to large immunoglobulins (IgGs), Aß binding single-chain variable fragments (scFvs) were used for diagnostic and therapeutic research approaches. scFvs can be expressed in E. coli and may provide improved pharmacokinetic properties like increased blood-brain barrier permeability or reduced side-effects in vivo. In this study, we constructed an scFv from an Aß binding IgG, designated IC16, which binds the N-terminal region of Aß (Aß(1-8)). scFv-IC16 was expressed in E. coli, purified and characterized with respect to its interaction with different Aß species and its influence on Aß fibril formation. We were able to show that scFv-IC16 strongly influenced the aggregation behavior of Aß and could be applied as an Aß detection probe for plaque staining in the brains of transgenic AD model mice. The results indicate potential for therapy and diagnosis of AD.
Subject(s)
Alzheimer Disease/immunology , Amyloid beta-Peptides/chemistry , Epitopes/chemistry , Single-Chain Antibodies/chemistry , Alzheimer Disease/metabolism , Amino Acid Sequence , Animals , Benzothiazoles , Brain/pathology , Enzyme-Linked Immunosorbent Assay , Humans , Immunoglobulin G/chemistry , Immunohistochemistry , Ligands , Mice , Mice, Transgenic , Models, Molecular , Molecular Sequence Data , Neurodegenerative Diseases/immunology , Neurodegenerative Diseases/metabolism , Permeability , Plasmids/metabolism , Protein Binding , Sequence Alignment , Thiazoles/chemistryABSTRACT
Recent studies indicate that small amyloid-ß peptide (Aß) oligomers are the major toxic species responsible for development and progression of Alzheimer's disease (AD). Therefore, we suggest that the number of Aß oligomers in body fluids is the most direct and relevant biomarker for AD. Determination of the Aß oligomer content of cerebrospinal fluid (CSF) samples from 14 AD patients and 12 age-matched controls revealed a clear distinction between both groups. All samples of the control group showed homogenously low numbers of Aß oligomers, while the samples of the AD group exhibited significantly higher levels of Aß oligomers. The Aß oligomer numbers correlated with the patients' Mini-Mental State Examination scores. This indicates that the quantity of Aß oligomers in CSF reflects the severity of the disease and that Aß oligomers play a crucial role in AD pathology and in turn can be used as a diagnostic biomarker.
Subject(s)
Alzheimer Disease/cerebrospinal fluid , Amyloid beta-Peptides/cerebrospinal fluid , Biomarkers/cerebrospinal fluid , Aged , Aged, 80 and over , Alzheimer Disease/diagnosis , Case-Control Studies , Enzyme-Linked Immunosorbent Assay , Female , Humans , Male , Mental Status Schedule , Microscopy, FluorescenceABSTRACT
One of the characteristic pathological hallmarks of Alzheimer's disease (AD) is neuritic plaques. The sequence of events leading to deposition of amyloid-ß (Aß) peptides in plaques is not clear. Here we investigate the effects of D3, an Aß oligomer directed D-enantiomeric peptide that was obtained from a mirror image phage display selection against monomeric or small oligomeric forms of Aß42, on Aß deposition in aged AßPP/PS1 double transgenic AD-model mice. Using Alzet minipumps, we infused the brains of these AD model mice for 8 weeks with FITC-labeled D3, and examined the subsequent changes in pathology and cognitive deficits. Initial cognitive deficits are similar comparing control and D3-FITC-treated mice, but the treated mice show a significant improvement on the last day of testing. Further, we show that there is a substantial reduction in the amount of amyloid deposits in the animals treated with D3-FITC, compared to the control mice. Finally, the amount of activated microglia and astrocytes surrounding Aß deposits is dramatically reduced in the D3-FITC-treated mice. Our findings demonstrate that treatments with the high affinity Aß42 oligomer binding D-enantiomeric peptide D3 significantly decrease Aß deposits and the associated inflammatory response, and improve cognition even when applied only at late stages and high age. Together, this suggests that the treatment reduces the level of Aß peptide in the brains of AßPP/PS1 mice, possibly by increasing Aß outflow from the brain. In conclusion, treatments with this D-peptide have great potential to be successful in AD patients.
Subject(s)
Alzheimer Disease/drug therapy , Amyloid beta-Protein Precursor/genetics , Cognition Disorders/drug therapy , Cognition Disorders/pathology , Oligopeptides/administration & dosage , Plaque, Amyloid/drug therapy , Plaque, Amyloid/pathology , Presenilin-1/genetics , Aging/genetics , Aging/pathology , Alzheimer Disease/genetics , Alzheimer Disease/pathology , Amyloid beta-Protein Precursor/antagonists & inhibitors , Animals , Cognition Disorders/genetics , Disease Models, Animal , Hippocampus/metabolism , Hippocampus/pathology , Inflammation/drug therapy , Inflammation/genetics , Inflammation/pathology , Infusions, Intraventricular , Male , Mice , Mice, Transgenic , Oligopeptides/genetics , Plaque, Amyloid/genetics , Treatment OutcomeABSTRACT
The aggregation of the Aß plays a fundamental role in the pathology of AD. Recently, N-terminally modified Aß species, pE-Aß, have been described as major constituents of Aß deposits in the brains of AD patients. pE-Aß has an increased aggregation propensity and shows increased toxicity compared with Aß1-40 and Aß1-42. In the present work, high-resolution NMR spectroscopy was performed to study pE-Aß3-40 in aqueous TFE-containing solution. Two-dimensional TOCSY and NOESY experiments were performed. On the basis of NOE and chemical shift data, pE-Aß3-40 was shown to contain two helical regions formed by residues 14-22 and 30-36. This is similar as previously described for Aß1-40. However, the secondary chemical shift data indicate decreased helical propensity in pE-Aß3-40 when compared with Aß1-40 under exactly the same conditions. This is in agreement with the observation that pE-Aß3-40 shows a drastically increased tendency to form ß-sheet-rich structures under more physiologic conditions. Structural studies of pE-Aß are crucial for better understanding the structural basis of amyloid fibril formation in the brain during development of AD, especially because an increasing number of reports indicate a decisive role of pE-Aß for the pathogenesis of AD.
Subject(s)
Amyloid beta-Peptides/chemistry , Magnetic Resonance Spectroscopy , Pyrrolidonecarboxylic Acid/chemistry , Alzheimer Disease/pathology , Humans , Protein Isoforms/chemistryABSTRACT
Alzheimer's disease (AD) is a devastating disease affecting predominantly the aging population. One of the characteristic pathological hallmarks of AD are neuritic plaques, consisting of amyloid-ß peptide (Aß). While there has been some advancement in diagnostic classification of AD patients according to their clinical severity, no fully reliable method for pre-symptomatic diagnosis of AD is available. To enable such early diagnosis, which will allow the initiation of treatments early in the disease progress, neuroimaging tools are under development, making use of Aß-binding ligands that can visualize amyloid plaques in the living brain. Here we investigate the properties of a newly designed series of D-enantiomeric peptides which are derivatives of ACI-80, formerly called D1, which was developed to specifically bind aggregated Aß1-42. We describe ACI-80 derivatives with increased stability and Aß binding properties, which were characterized using surface plasmon resonance and enzyme-linked immunosorbent assays. The specific interactions of the lead compounds with amyloid plaques were validated by ex vivo immunochemistry in transgenic mouse models of AD. The novel compounds showed increased binding affinity and are promising candidates for further development into in vivo imaging compounds.
Subject(s)
Alzheimer Disease/diagnosis , Alzheimer Disease/metabolism , Amyloid beta-Peptides/chemistry , Amyloid/chemistry , Diagnostic Imaging/methods , Oligopeptides/chemistry , Peptide Fragments/chemistry , Amyloid/metabolism , Amyloid beta-Peptides/metabolism , Animals , Ligands , Mice , Neurites/metabolism , Neurites/pathology , Oligopeptides/metabolism , Peptide Fragments/metabolism , Protein BindingABSTRACT
Peptides are attracting increasing attention as therapeutics. D-enantiomeric peptides are remarkably resistant to in vivo proteolysis and elicit low immunogenic responses when compared with the respective L-peptides. Therefore, D-peptides can serve as therapeutic and early diagnosis agents for drug development. Here we discuss the application of mirror image phage display in pharmaceutical biotechnology aiming to identify protease resistant D-peptides with biotechnological approaches.
Subject(s)
Biotechnology/methods , Cell Surface Display Techniques/methods , Drug Discovery , Peptide Hydrolases/chemistry , Peptide Hydrolases/metabolism , Peptides/genetics , Peptides/metabolism , StereoisomerismABSTRACT
A key feature of Alzheimer disease (AD) is the pathologic self-association of the amyloid-ß (Aß) peptide, leading to the formation of diffusible toxic Aß oligomers and extracellular amyloid plaques. Next to extracellular Aß, intraneuronal Aß has important pathological functions in AD. Agents that specifically interfere with the oligomerization processes either outside or inside of neurons are highly desired for the elucidation of the pathologic mechanisms of AD and might even pave the way for new AD gene therapeutic approaches. Here, we characterize the Aß binding peptide L3 and its influence on Aß oligomerization in vitro. Preliminary studies in cell culture demonstrate that stably expressed L3 reduces cell toxicity of externally added Aß in neuroblastoma cells.
Subject(s)
Alzheimer Disease/genetics , Alzheimer Disease/therapy , Amyloid beta-Peptides/chemistry , Genetic Therapy/methods , Animals , Benzothiazoles , Cell Line, Tumor , Centrifugation, Density Gradient , Disease Models, Animal , Humans , Mice , Nephelometry and Turbidimetry , Neurons/metabolism , Peptides/chemistry , Sequence Analysis, DNA , Thiazoles/chemistryABSTRACT
Alzheimer's disease (AD) is a progressive neurodegenerative disorder with devastating effects. The greatest risk factor to develop AD is age. Today, only symptomatic therapies are available. Additionally, AD can be diagnosed with certainty only post mortem, whereas the diagnosis "probable AD" can be established earliest when severe clinical symptoms appear. Specific neuropathological changes like neurofibrillary tangles and amyloid plaques define AD. Amyloid plaques are mainly composed of the amyloid-ßpeptide (Aß). Several lines of evidence suggest that the progressive concentration and subsequent aggregation and accumulation of Aß play a fundamental role in the disease progress. Therefore, substances which bind to Aß and influence aggregation thereof are of great interest. An enormous number of organic substances for therapeutic purposes are described. This review focuses on peptides developed for diagnosis and therapy of AD and discusses the pre- and disadvantages of peptide drugs.
Subject(s)
Alzheimer Disease/diagnosis , Alzheimer Disease/drug therapy , Peptides/therapeutic use , Amino Acid Sequence , Amyloid beta-Protein Precursor , Drug Discovery , Humans , Peptide LibraryABSTRACT
The diagnosis of probable Alzheimer's disease (AD) can be established premortem based on clinical criteria like neuropsychological tests. Post mortem, specific neuropathological changes like amyloid plaques define AD. However, the standard criteria based on medical history and mental status examinations do not take into account the long preclinical features of the disease, and a biomarker for improved diagnosis of AD is urgently needed. In a large number of studies, amyloid-ß (Aß) monomer concentrations in CSF of AD patients are consistently and significantly reduced when compared to healthy controls. Therefore, monomeric Aß in CSF was suggested to be a helpful biomarker for the diagnosis of preclinical AD. However, not the monomeric form, but Aß oligomers have been shown to be the toxic species in AD pathology, and their quantification and characterization could facilitate AD diagnosis and therapy monitoring. Here, we review the current status of assay development to reliably and routinely detect Aß oligomers and high-molecular-weight particles in CSF.