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Synapse loss correlates with cognitive decline in Alzheimer's disease, and soluble oligomeric amyloid beta (Aß) is implicated in synaptic dysfunction and loss. An important knowledge gap is the lack of understanding of how Aß leads to synapse degeneration. In particular, there has been difficulty in determining whether there is a synaptic receptor that binds Aß and mediates toxicity. While many candidates have been observed in model systems, their relevance to human AD brain remains unknown. This is in part due to methodological limitations preventing visualization of Aß binding at individual synapses. To overcome this limitation, we combined two high resolution microscopy techniques: array tomography and Förster resonance energy transfer (FRET) to image over 1 million individual synaptic terminals in temporal cortex from AD (n = 11) and control cases (n = 9). Within presynapses and post-synaptic densities, oligomeric Aß generates a FRET signal with transmembrane protein 97. Further, Aß generates a FRET signal with cellular prion protein, and post-synaptic density 95 within post synapses. Transmembrane protein 97 is also present in a higher proportion of post synapses in Alzheimer's brain compared to controls. We inhibited Aß/transmembrane protein 97 interaction in a mouse model of amyloidopathy by treating with the allosteric modulator CT1812. CT1812 drug concentration correlated negatively with synaptic FRET signal between transmembrane protein 97 and Aß. In human-induced pluripotent stem cell derived neurons, transmembrane protein 97 is present in synapses and colocalizes with Aß when neurons are challenged with human Alzheimer's brain homogenate. Transcriptional changes are induced by Aß including changes in genes involved in neurodegeneration and neuroinflammation. CT1812 treatment of these neurons caused changes in gene sets involved in synaptic function. These data support a role for transmembrane protein 97 in the synaptic binding of Aß in human Alzheimer's disease brain where it may mediate synaptotoxicity.
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Enfermedad de Alzheimer , Disfunción Cognitiva , Proteínas de la Membrana , Animales , Humanos , Ratones , Péptidos beta-Amiloides , Encéfalo , Sinapsis , Proteínas de la Membrana/metabolismoRESUMEN
There is a large unmet medical need to develop disease-modifying treatment options for individuals with age-related degenerative diseases of the central nervous system. The sigma-2 receptor (S2R), encoded by TMEM97, is expressed in brain and retinal cells, and regulates cell functions via its co-receptor progesterone receptor membrane component 1 (PGRMC1), and through other protein-protein interactions. Studies describing functions of S2R involve the manipulation of expression or pharmacological modulation using exogenous small-molecule ligands. These studies demonstrate that S2R modulates key pathways involved in age-related diseases including autophagy, trafficking, oxidative stress, and amyloid-ß and α-synuclein toxicity. Furthermore, S2R modulation can ameliorate functional deficits in cell-based and animal models of disease. This review summarizes the current evidence-based understanding of S2R biology and function, and its potential as a therapeutic target for age-related degenerative diseases of the central nervous system, including Alzheimer's disease, α-synucleinopathies, and dry age-related macular degeneration.
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Enfermedad de Alzheimer , Enfermedad por Cuerpos de Lewy , Receptores sigma , Animales , Enfermedad de Alzheimer/tratamiento farmacológico , Receptores sigma/metabolismo , alfa-Sinucleína/metabolismo , Péptidos beta-Amiloides , BiologíaRESUMEN
Several mutations conferring protection against Alzheimer's disease (AD) have been described, none as profound as the A673T mutation, where carriers are four times less likely to get AD compared to noncarriers. This mutation results in reduced amyloid beta (Aß) protein production in vitro and lower lifetime Aß concentration in carriers. Better understanding of the protective mechanisms of the mutation may provide important insights into AD pathophysiology and identify productive therapeutic intervention strategies for disease modification. Aß(1-42) protein forms oligomers that bind saturably to a single receptor site on neuronal synapses, initiating the downstream toxicities observed in AD. Decreased formation, toxicity, or stability of soluble Aß oligomers, or reduction of synaptic binding of these oligomers, may combine with overall lower Aß concentration to underlie A673T's disease protecting mechanism. To investigate these possibilities, we compared the formation rate of soluble oligomers made from Icelandic A673T mutant and wild type (wt) Aß(1-42) synthetic protein, the amount and intensity of oligomer bound to mature primary rat hippocampal/cortical neuronal synapses, and the potency of bound oligomers to impact trafficking rate in neurons in vitro using a physiologically relevant oligomer preparation method. At equal protein concentrations, mutant protein forms approximately 50% or fewer oligomers of high molecular weight (>50 kDa) compared to wt protein. Mutant oligomers are twice as potent at altering the cellular vesicle trafficking rate as wt at equivalent concentrations, however, mutant oligomers have a >4-fold lower binding affinity to synaptic receptors (Kd = 1,950 vs. 442 nM). The net effect of these differences is a lower overall toxicity at a given concentration. This study demonstrates for the first time that mutant A673T Aß oligomers prepared with this method have fundamentally different assembly characteristics and biological impact from wt protein and indicates that its disease protecting mechanism may result primarily from the mutant protein's much lower binding affinity to synaptic receptors. This suggests that therapeutics that effectively reduce oligomer binding to synapses in the brain may be beneficial in AD.
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Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/genética , Péptidos beta-Amiloides/metabolismo , Neuronas/metabolismo , Animales , Humanos , Unión Proteica , Transporte de Proteínas/fisiología , Ratas , Ratas Sprague-DawleyRESUMEN
α-Synuclein oligomers are thought to have a pivotal role in sporadic and familial Parkinson's disease (PD) and related α-synucleinopathies, causing dysregulation of protein trafficking, autophagy/lysosomal function, and protein clearance, as well as synaptic function impairment underlying motor and cognitive symptoms of PD. Moreover, trans-synaptic spread of α-synuclein oligomers is hypothesized to mediate disease progression. Therapeutic approaches that effectively block α-synuclein oligomer-induced pathogenesis are urgently needed. Here, we show for the first time that α-synuclein species isolated from human PD patient brain and recombinant α-synuclein oligomers caused similar deficits in lipid vesicle trafficking rates in cultured rat neurons and glia, while α-synuclein species isolated from non-PD human control brain samples did not. Recombinant α-synuclein oligomers also increased neuronal expression of lysosomal-associated membrane protein-2A (LAMP-2A), the lysosomal receptor that has a critical role in chaperone-mediated autophagy. Unbiased screening of several small molecule libraries (including the NIH Clinical Collection) identified sigma-2 receptor antagonists as the most effective at blocking α-synuclein oligomer-induced trafficking deficits and LAMP-2A upregulation in a dose-dependent manner. These results indicate that antagonists of the sigma-2 receptor complex may alleviate α-synuclein oligomer-induced neurotoxicity and are a novel therapeutic approach for disease modification in PD and related α-synucleinopathies.
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Enfermedad de Parkinson/metabolismo , Receptores sigma/antagonistas & inhibidores , Receptores sigma/metabolismo , alfa-Sinucleína/metabolismo , Anciano , Anciano de 80 o más Años , Animales , Autofagia/efectos de los fármacos , Encéfalo/metabolismo , Femenino , Ensayos Analíticos de Alto Rendimiento , Humanos , Metabolismo de los Lípidos , Proteína 2 de la Membrana Asociada a los Lisosomas/metabolismo , Masculino , Enfermedad de Parkinson/patología , Cultivo Primario de Células , Ratas , Ratas Sprague-Dawley , Proteínas de Transporte Vesicular/metabolismo , alfa-Sinucleína/farmacologíaRESUMEN
INTRODUCTION: Amyloid beta (Aß) oligomers are one of the most toxic structural forms of the Aß protein and are hypothesized to cause synaptotoxicity and memory failure as they build up in Alzheimer's disease (AD) patients' brain tissue. We previously demonstrated that antagonists of the sigma-2 receptor complex effectively block Aß oligomer toxicity. CT1812 is an orally bioavailable, brain penetrant small molecule antagonist of the sigma-2 receptor complex that appears safe and well tolerated in healthy elderly volunteers. We tested CT1812's effect on Aß oligomer pathobiology in preclinical AD models and evaluated CT1812's impact on cerebrospinal fluid (CSF) protein biomarkers in mild to moderate AD patients in a clinical trial (ClinicalTrials.gov NCT02907567). METHODS: Experiments were performed to measure the impact of CT1812 versus vehicle on Aß oligomer binding to synapses in vitro, to human AD patient post mortem brain tissue ex vivo, and in living APPSwe /PS1dE9 transgenic mice in vivo. Additional experiments were performed to measure the impact of CT1812 versus vehicle on Aß oligomer-induced deficits in membrane trafficking rate, synapse number, and protein expression in mature hippocampal/cortical neurons in vitro. The impact of CT1812 on cognitive function was measured in transgenic Thy1 huAPPSwe/Lnd+ and wild-type littermates. A multicenter, double-blind, placebo-controlled parallel group trial was performed to evaluate the safety, tolerability, and impact on protein biomarker expression of CT1812 or placebo given once daily for 28 days to AD patients (Mini-Mental State Examination 18-26). CSF protein expression was measured by liquid chromatography with tandem mass spectrometry or enzyme-linked immunosorbent assay in samples drawn prior to dosing (Day 0) and at end of dosing (Day 28) and compared within each patient and between pooled treated versus placebo-treated dosing groups. RESULTS: CT1812 significantly and dose-dependently displaced Aß oligomers bound to synaptic receptors in three independent preclinical models of AD, facilitated oligomer clearance into the CSF, increased synaptic number and protein expression in neurons, and improved cognitive performance in transgenic mice. CT1812 significantly increased CSF concentrations of Aß oligomers in AD patient CSF, reduced concentrations of synaptic proteins and phosphorylated tau fragments, and reversed expression of many AD-related proteins dysregulated in CSF. DISCUSSION: These preclinical studies demonstrate the novel disease-modifying mechanism of action of CT1812 against AD and Aß oligomers. The clinical results are consistent with preclinical data and provide evidence of target engagement and impact on fundamental disease-related signaling pathways in AD patients, supporting further development of CT1812.
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Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/líquido cefalorraquídeo , Biomarcadores/líquido cefalorraquídeo , Cognición/efectos de los fármacos , Ratones Transgénicos , Receptores sigma/antagonistas & inhibidores , Anciano , Animales , Encéfalo/metabolismo , Método Doble Ciego , Ensayo de Inmunoadsorción Enzimática , Hipocampo/metabolismo , Humanos , Masculino , Ratones , Neuronas/metabolismo , Sinapsis/metabolismoRESUMEN
Progesterone receptor membrane component 1 (PGRMC1) is a multi-functional protein with a heme-binding moiety related to that of cytochrome b5, which is a putative progesterone receptor. The recently solved PGRMC1 structure revealed that heme-binding involves coordination by a tyrosinate ion at Y113, and induces dimerization which is stabilized by hydrophobic stacking of heme on adjacent monomers. Dimerization is required for association with cytochrome P450 (cyP450) enzymes, which mediates chemoresistance to doxorubicin and may be responsible for PGRMC1's anti-apoptotic activity. Here we review the multiple attested involvement of PGRMC1 in diverse functions, including regulation of cytochrome P450, steroidogenesis, vesicle trafficking, progesterone signaling and mitotic spindle and cell cycle regulation. Its wide range of biological functions is attested to particularly by its emerging association with cancer and progesterone-responsive female reproductive tissues. PGRMC1 exhibits all the hallmarks of a higher order nexus signal integration hub protein. It appears capable of acting as a detector that integrates information from kinase/phosphatase pathways with heme and CO levels and probably redox status.
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Proteínas de la Membrana/fisiología , Neoplasias/metabolismo , Receptores de Progesterona/fisiología , Ciclo Celular , Proliferación Celular , Humanos , Proteínas de la Membrana/química , Neoplasias/patología , Multimerización de Proteína , Receptores de Progesterona/química , Receptores sigma/fisiologíaRESUMEN
BACKGROUND: Effective, disease-modifying therapeutics for the treatment of Alzheimer's disease (AD) remain a large unmet need. Extensive evidence suggests that amyloid beta (Aß) is central to AD pathophysiology, and Aß oligomers are among the most toxic forms of Aß. CT1812 is a novel brain penetrant sigma-2 receptor ligand that interferes with the binding of Aß oligomers to neurons. Preclinical studies of CT1812 have demonstrated its ability to displace Aß oligomers from neurons, restore synapses in cell cultures, and improve cognitive measures in mouse models of AD. CT1812 was found to be generally safe and well tolerated in a placebo-controlled phase 1 clinical trial in healthy volunteers and phase 1a/2 clinical trials in patients with mild to moderate dementia due to AD. The unique objective of this study was to incorporate synaptic positron emission tomography (PET) imaging as an outcome measure for CT1812 in AD patients. METHODS: The present phase 1/2 study was a randomized, double-blind, placebo-controlled, parallel-group trial conducted in 23 participants with mild to moderate dementia due to AD to primarily evaluate the safety of CT1812 and secondarily its pharmacodynamic effects. Participants received either placebo or 100 mg or 300 mg per day of oral CT1812 for 24 weeks. Pharmacodynamic effects were assessed using the exploratory efficacy endpoints synaptic vesicle glycoprotein 2A (SV2A) PET, fluorodeoxyglucose (FDG) PET, volumetric MRI, cognitive clinical measures, as well as cerebrospinal fluid (CSF) biomarkers of AD pathology and synaptic degeneration. RESULTS: No treatment differences relative to placebo were observed in the change from baseline at 24 weeks in either SV2A or FDG PET signal, the cognitive clinical rating scales, or in CSF biomarkers. Composite region volumetric MRI revealed a trend towards tissue preservation in participants treated with either dose of CT1812, and nominally significant differences with both doses of CT1812 compared to placebo were found in the pericentral, prefrontal, and hippocampal cortices. CT1812 was safe and well tolerated. CONCLUSIONS: The safety findings of this 24-week study and the observed changes on volumetric MRI with CT1812 support its further clinical development. TRIAL REGISTRATION: The clinical trial described in this manuscript is registered at clinicaltrials.gov (NCT03493282).
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Enfermedad de Alzheimer , Ratones , Animales , Humanos , Enfermedad de Alzheimer/diagnóstico por imagen , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/líquido cefalorraquídeo , Proyectos Piloto , Fluorodesoxiglucosa F18 , Tomografía de Emisión de Positrones , Biomarcadores/líquido cefalorraquídeoRESUMEN
In this study, a panel of 46 compounds containing five different scaffolds known to have high σ2 receptor affinity were screened. 6,7-Dimethoxy-2-[4-(4-methoxyphenyl)butan-2-yl]-1,2,3,4-tetrahydroisoquinoline [(±)-7] (Ki for σ1 = 48.4 ± 7.7 nM, and Ki for σ2 = 0.59 ± 0.02 nM) and its desmethyl analogue, (±)-8 (Ki for σ1 = 108 ± 35 nM, and Ki for σ2 = 4.92 ± 0.59 nM), showed excellent binding affinity and subtype selectivity for σ2 receptors. In vitro cell binding indicated that σ2 receptor binding of [11C]-(±)-7 and [11C]-(±)-8 was dependent on TMEM97 protein expression. In PET studies, the peak brain uptake of [11C]-(±)-7 (8.28 ± 2.52%ID/cc) was higher than that of [11C]-(±)-8 (4.25 ± 0.97%ID/cc) with specific distribution in the cortex and hypothalamus. Brain uptake or tissue binding was selectively inhibited by ligands with different σ2 receptor binding affinities. The results suggest [11C]-(±)-7 can be used as a PET radiotracer for imaging the function of σ2 receptors in central nervous system disorders.
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Receptores sigma , Tetrahidroisoquinolinas , Encéfalo/diagnóstico por imagen , Encéfalo/metabolismo , Ligandos , Tomografía de Emisión de Positrones , Radiofármacos/química , Tetrahidroisoquinolinas/químicaRESUMEN
BACKGROUND: Mature primary neuronal cultures are an important model of the nervous system, but limited scalability has been a major challenge in their use for drug discovery of neurodegenerative diseases. This work describes a method for improving scalability through the use of larger format microtiter plates while preserving culture quality. NEW METHOD: Here we describe a method and quality control procedures for growing embryonic day 18 rat hippocampal/cortical neuronal cultures in 384-well microtiter plates for three weeks in vitro. RESULTS: We use these cultures in two assays measuring intracellular lipid vesicle trafficking and synapse density for routine screening of small molecule libraries. Together this culture system and screening platform have successfully identified therapeutics capable of improving cognitive function in transgenic models of Alzheimer's disease that have advanced to clinical trials, validating their translational applicability. COMPARISON WITH EXISTING METHODS: Our method enables the growth of healthy, mature neurons in larger format microtiter plates than in traditional primary neuronal culturing protocols, making it ideal for drug screening and mechanism of action studies. CONCLUSION: The predictive capacity of this culture system and screening platform provides a method for rapidly identifying novel disease-modifying neurodegenerative therapeutics.
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Enfermedad de Alzheimer , Enfermedades Neurodegenerativas , Enfermedad de Alzheimer/tratamiento farmacológico , Péptidos beta-Amiloides , Animales , Descubrimiento de Drogas , Enfermedades Neurodegenerativas/tratamiento farmacológico , Neuronas , RatasRESUMEN
An unbiased phenotypic neuronal assay was developed to measure the synaptotoxic effects of soluble Aß oligomers. A collection of CNS druglike small molecules prepared by conditioned extraction was screened. Compounds that prevented and reversed synaptotoxic effects of Aß oligomers in neurons were discovered to bind to the sigma-2 receptor complex. Select development compounds displaced receptor-bound Aß oligomers, rescued synapses, and restored cognitive function in transgenic hAPP Swe/Ldn mice. Our first-in-class orally administered small molecule investigational drug 7 (CT1812) has been advanced to Phase II clinical studies for Alzheimer's disease.
RESUMEN
The σ-2 receptor (S2R) complex has been implicated in CNS disorders ranging from anxiety and depression to neurodegenerative disorders such as Alzheimer's disease (AD). The proteins comprising the S2R complex impact processes including autophagy, cholesterol synthesis, progesterone signaling, lipid membrane-bound protein trafficking, and receptor stabilization at the cell surface. While there has been much progress in understanding the role of S2R in cellular processes and its potential therapeutic value, a great deal remains unknown. The International Symposium on Sigma-2 Receptors is held in conjunction with the annual Society for Neuroscience (SfN) conference to promote collaboration and advance the field of S2R research. This review summarizes updates presented at the Fourth International Symposium on Sigma-2 Receptors: Role in Health and Disease, a Satellite Symposium held at the 2019 SfN conference. Interdisciplinary members of the S2R research community presented both previously published and preliminary results from ongoing studies of the role of S2R in cellular metabolism, the anatomic and cellular expression patterns of S2R, the relationship between S2R and amyloid ß (Aß) in AD, the role of S2R complex protein PGRMC1 in health and disease, and the efforts to design new S2R ligands for the purposes of research and drug development. The proceedings from this symposium are reported here as an update on the field of S2R research, as well as to highlight the value of the symposia that occur yearly in conjunction with the SfN conference.
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Enfermedad de Alzheimer , Receptores sigma , Péptidos beta-Amiloides , Humanos , Proteínas de la Membrana , Progesterona , Receptores de ProgesteronaRESUMEN
BACKGROUND: Synapse damage and loss are fundamental to the pathophysiology of Alzheimer's disease (AD) and lead to reduced cognitive function. The goal of this review is to address the challenges of forging new clinical development approaches for AD therapeutics that can demonstrate reduction of synapse damage or loss. The key points of this review include the following: Synapse loss is a downstream effect of amyloidosis, tauopathy, inflammation, and other mechanisms occurring in AD.Synapse loss correlates most strongly with cognitive decline in AD because synaptic function underlies cognitive performance.Compounds that halt or reduce synapse damage or loss have a strong rationale as treatments of AD.Biomarkers that measure synapse degeneration or loss in patients will facilitate clinical development of such drugs.The ability of methods to sensitively measure synapse density in the brain of a living patient through synaptic vesicle glycoprotein 2A (SV2A) positron emission tomography (PET) imaging, concentrations of synaptic proteins (e.g., neurogranin or synaptotagmin) in the cerebrospinal fluid (CSF), or functional imaging techniques such as quantitative electroencephalography (qEEG) provides a compelling case to use these types of measurements as biomarkers that quantify synapse damage or loss in clinical trials in AD. CONCLUSION: A number of emerging biomarkers are able to measure synapse injury and loss in the brain and may correlate with cognitive function in AD. These biomarkers hold promise both for use in diagnostics and in the measurement of therapeutic successes.
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Enfermedad de Alzheimer/líquido cefalorraquídeo , Enfermedad de Alzheimer/diagnóstico por imagen , Enfermedad de Alzheimer/patología , Biomarcadores/líquido cefalorraquídeo , Sinapsis/patología , Electroencefalografía/métodos , Neuroimagen Funcional/métodos , Humanos , Tomografía de Emisión de Positrones/métodosRESUMEN
Measurement of antiproliferative capacity of a compound is central to early oncology drug discovery, with information about the precise mechanism of compound action typically being acquired during later downstream assays. Here we describe the development and validation of an in vitro image-based assay that simultaneously measures tumor cell count, late apoptotic morphology, and nuclear DNA content (termed the proliferation, apoptosis, and DNA content [PAD] assay) by using a DNA binding fluorescent dye. The PAD assay determines whether a compound's antiproliferative effect occurs via cell cycle arrest or induction of apoptosis, replacing downstream assays for 1/50(th) the cost. We used this assay to screen a kinase inhibitor-biased library and discovered an Aurora kinase inhibitor, and we also used it to drive structure-activity relationship to clinical candidate Investigational New Drug filing within 2 years. The simplicity of the PAD assay was critical to the rapid time frame within which this candidate was identified and progressed. This inhibitor is currently beginning Phase II clinical trials.
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Apoptosis/efectos de los fármacos , ADN/análisis , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Aurora Quinasas , Bromodesoxiuridina/metabolismo , División Celular , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Fase G2 , HumanosRESUMEN
BACKGROUND: Elayta (CT1812) is a novel allosteric antagonist of the sigma-2 receptor complex that prevents and displaces binding of Aß oligomers to neurons. By stopping a key initiating event in Alzheimer's disease, this first-in-class drug candidate mitigates downstream synaptotoxicity and restores cognitive function in aged transgenic mouse models of Alzheimer's disease. METHODS: A phase 1, two-part single and multiple ascending dose study was conducted in 7 and 4 cohorts of healthy human subjects, respectively. In part A, healthy, young subjects (<65 years old) received CT1812 doses ranging from 10 to 1120 mg (6:2 active to placebo [A:P] per cohort). In part B, subjects were administered 280, 560, and 840 mg once daily for 14 days (8:2 A:P per cohort). An elderly cohort, aged 65-75 years, was dosed at 560 mg once daily for 14 days (7:2 A:P). Serum concentrations of CT1812 in part B were measured on day 3 and 14 and cerebrospinal fluid concentrations on day 7 or 9. Cognitive testing was performed in the healthy elderly cohort at baseline and at day 14 of treatment. RESULTS: Treatment with CT1812 was well tolerated in all cohorts. Adverse events were mild to moderate in severity and included headache and GI tract symptoms. Plasma concentrations of drug were dose proportional across two orders of magnitude with minimal accumulation over 14 days. Cognitive scores in the healthy elderly cohort were similar before and after treatment. CONCLUSIONS: CT1812 was well tolerated with single dose administration up to 1120 mg and with multiple dose administration up to 840 mg and 560 mg in healthy young and healthy elderly subjects, respectively. CT1812 is currently being studied in early phase 2 trials in patients with Alzheimer's disease.
RESUMEN
Amyloid beta-derived diffusible ligands (ADDLs) comprise the neurotoxic subset of soluble Abeta(1-42) oligomers, now widely considered to be the molecular cause of memory malfunction and neurodegeneration in Alzheimer's disease (AD). We have developed a screening cascade which identifies small molecule modulators of ADDL-mediated neurotoxicity. The primary screen involves a fluorescence resonance energy transfer (FRET)-based assay which selects inhibitors of Abeta1-42 oligomer assembly. The identified hits were further characterized by assessing their ability to inhibit the assembly and binding of ADDLs to cultures of primary hippocampal neurons. This approach has led to the identification of a number of small molecules which inhibit ADDL assembly and their subsequent binding to neurons. Here we describe our small molecule discovery efforts to identify ADDL assembly blocker and ADDL binding inhibitors, and to transform validated hits into pre-clinical lead compounds.
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Enfermedad de Alzheimer/tratamiento farmacológico , Péptidos beta-Amiloides/metabolismo , Antipsicóticos/uso terapéutico , Péptidos beta-Amiloides/antagonistas & inhibidores , Péptidos beta-Amiloides/química , Animales , Antipsicóticos/química , Diseño de Fármacos , Humanos , Bibliotecas de Moléculas PequeñasRESUMEN
The amyloid-beta (Abeta) cascade hypothesis of Alzheimer's disease (AD) has dominated research and subsequent therapeutic drug development for over two decades. Central to this hypothesis is the observation that Abeta is elevated in AD patients and that the disease is ultimately characterized by the central deposition of insoluble senile plaques. More recent evidence, however, suggests that the presence or absence of plaque is insufficient to fully account for the deleterious role of elevated Abeta in AD. Such studies support the basis for an alternate interpretation of the Abeta cascade hypothesis. Namely, that soluble oligomers of Abeta (i.e., ADDLs) accumulate and cause functional deficits prior to overt neuronal cell death or plaque deposition. Accordingly, the following review focuses on research describing the preparation and functional activity of ADDLs in vitro and in vivo. These studies provide the basis for an alternate, ADDL-based, view of the Abeta cascade hypothesis and accounts for the disconnect between plaque burden and cognitive deficits. Possible therapeutic approaches aimed at lowering ADDLs in AD patients are also considered.
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Enfermedad de Alzheimer/etiología , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/metabolismo , Enfermedad de Alzheimer/tratamiento farmacológico , Animales , Humanos , LigandosRESUMEN
Synaptic dysfunction and loss caused by age-dependent accumulation of synaptotoxic beta amyloid (Abeta) 1-42 oligomers is proposed to underlie cognitive decline in Alzheimer's disease (AD). Alterations in membrane trafficking induced by Abeta oligomers mediates reduction in neuronal surface receptor expression that is the basis for inhibition of electrophysiological measures of synaptic plasticity and thus learning and memory. We have utilized phenotypic screens in mature, in vitro cultures of rat brain cells to identify small molecules which block or prevent the binding and effects of Abeta oligomers. Synthetic Abeta oligomers bind saturably to a single site on neuronal synapses and induce deficits in membrane trafficking in neuronal cultures with an EC50 that corresponds to its binding affinity. The therapeutic lead compounds we have found are pharmacological antagonists of Abeta oligomers, reducing the binding of Abeta oligomers to neurons in vitro, preventing spine loss in neurons and preventing and treating oligomer-induced deficits in membrane trafficking. These molecules are highly brain penetrant and prevent and restore cognitive deficits in mouse models of Alzheimer's disease. Counter-screening these compounds against a broad panel of potential CNS targets revealed they are highly potent and specific ligands of the sigma-2/PGRMC1 receptor. Brain concentrations of the compounds corresponding to greater than 80% receptor occupancy at the sigma-2/PGRMC1 receptor restore cognitive function in transgenic hAPP Swe/Ldn mice. These studies demonstrate that synthetic and human-derived Abeta oligomers act as pharmacologically-behaved ligands at neuronal receptors--i.e. they exhibit saturable binding to a target, they exert a functional effect related to their binding and their displacement by small molecule antagonists blocks their functional effect. The first-in-class small molecule receptor antagonists described here restore memory to normal in multiple AD models and sustain improvement long-term, representing a novel mechanism of action for disease-modifying Alzheimer's therapeutics.
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Enfermedad de Alzheimer/tratamiento farmacológico , Péptidos beta-Amiloides/química , Neuronas/metabolismo , Fragmentos de Péptidos/química , Sinapsis/efectos de los fármacos , Enfermedad de Alzheimer/metabolismo , Animales , Encéfalo/efectos de los fármacos , Química Farmacéutica , Cognición/efectos de los fármacos , Trastornos del Conocimiento/tratamiento farmacológico , Diseño de Fármacos , Ensayo de Inmunoadsorción Enzimática , Humanos , Ratones , Ratones Transgénicos , Neuroglía/metabolismo , Unión Proteica , Transporte de Proteínas , Ratas , Ratas Sprague-Dawley , Sinapsis/metabolismoRESUMEN
Amyloid beta (Abeta) 1-42 oligomers accumulate in brains of patients with Mild Cognitive Impairment (MCI) and disrupt synaptic plasticity processes that underlie memory formation. Synaptic binding of Abeta oligomers to several putative receptor proteins is reported to inhibit long-term potentiation, affect membrane trafficking and induce reversible spine loss in neurons, leading to impaired cognitive performance and ultimately to anterograde amnesia in the early stages of Alzheimer's disease (AD). We have identified a receptor not previously associated with AD that mediates the binding of Abeta oligomers to neurons, and describe novel therapeutic antagonists of this receptor capable of blocking Abeta toxic effects on synapses in vitro and cognitive deficits in vivo. Knockdown of sigma-2/PGRMC1 (progesterone receptor membrane component 1) protein expression in vitro using siRNA results in a highly correlated reduction in binding of exogenous Abeta oligomers to neurons of more than 90%. Expression of sigma-2/PGRMC1 is upregulated in vitro by treatment with Abeta oligomers, and is dysregulated in Alzheimer's disease patients' brain compared to age-matched, normal individuals. Specific, high affinity small molecule receptor antagonists and antibodies raised against specific regions on this receptor can displace synthetic Abeta oligomer binding to synaptic puncta in vitro and displace endogenous human AD patient oligomers from brain tissue sections in a dose-dependent manner. These receptor antagonists prevent and reverse the effects of Abeta oligomers on membrane trafficking and synapse loss in vitro and cognitive deficits in AD mouse models. These findings suggest sigma-2/PGRMC1 receptors mediate saturable oligomer binding to synaptic puncta on neurons and that brain penetrant, small molecules can displace endogenous and synthetic oligomers and improve cognitive deficits in AD models. We propose that sigma-2/PGRMC1 is a key mediator of the pathological effects of Abeta oligomers in AD and is a tractable target for small molecule disease-modifying therapeutics.