ABSTRACT
Exidavnemab is a monoclonal antibody (mAb) with a high affinity and selectivity for pathological aggregated forms of α-synuclein and a low affinity for physiological monomers, which is in clinical development as a disease-modifying treatment for patients with synucleinopathies such as Parkinson's disease. Safety, tolerability, pharmacokinetics, immunogenicity, and exploratory biomarkers were assessed in two separate Phase 1 single ascending dose studies, including single intravenous (IV) (100 to 6000 mg) or subcutaneous (SC) (300 mg) administration of exidavnemab in healthy volunteers (HVs). Across the two studies, a total of 98 Western, Caucasian, Japanese, and Han Chinese HVs were enrolled, of which 95 completed the study. Exidavnemab was generally well tolerated. There were no serious adverse events or safety issues identified in laboratory analyses. Headache, asymptomatic COVID-19, back pain, and post lumbar puncture syndrome were the most frequently reported treatment-emergent adverse events. Following IV infusion, the pharmacokinetics of exidavnemab was approximately dose linear in the range 100-6000 mg. The terminal half-life was approximately 30 days, and the exposure was comparable across Western, Caucasian, Japanese, and Han Chinese volunteers. The absolute SC bioavailability was â¼71%. Cerebrospinal fluid exposure relative to serum after single dose was within the range expected for mAbs (approximately 0.2%). The anti-drug antibody rates were low and there was no effect of immunogenicity on the pharmacokinetics or safety. Dose-dependent reduction of free α-synuclein in plasma was observed. In summary, exidavnemab was found to have an excellent pharmacokinetic profile and was well tolerated in HVs, supporting the continued clinical development.
ABSTRACT
A growing body of evidence suggests that aggregated α-synuclein, the major constituent of Lewy bodies, plays a key role in the pathogenesis of Parkinson's disease and related α-synucleinopathies. Immunotherapies, both active and passive, against α-synuclein have been developed and are promising novel treatment strategies for such disorders. Here, we report on the humanization and pharmacological characteristics of ABBV-0805, a monoclonal antibody that exhibits a high selectivity for human aggregated α-synuclein and very low affinity for monomers. ABBV-0805 binds to a broad spectrum of soluble aggregated α-synuclein, including small and large aggregates of different conformations. Binding of ABBV-0805 to pathological α-synuclein was demonstrated in Lewy body-positive post mortem brains of Parkinson's disease patients. The functional potency of ABBV-0805 was demonstrated in several cellular assays, including Fcγ-receptor mediated uptake of soluble aggregated α-synuclein in microglia and inhibition of neurotoxicity in primary neurons. In vivo, the murine version of ABBV-0805 (mAb47) displayed significant dose-dependent decrease of α-synuclein aggregates in brain in several mouse models, both in prophylactic and therapeutic settings. In addition, mAb47 treatment of α-synuclein transgenic mice resulted in a significantly prolonged survival. ABBV-0805 selectively targets soluble toxic α-synuclein aggregates with a picomolar affinity and demonstrates excellent in vivo efficacy. Based on the strong preclinical findings described herein, ABBV-0805 has been progressed into clinical development as a potential disease-modifying treatment for Parkinson's disease.
Subject(s)
Antibodies, Monoclonal , Parkinson Disease , Synucleinopathies , Animals , Antibodies, Monoclonal/therapeutic use , Humans , Longevity , Mice , Mice, Transgenic , Parkinson Disease/metabolism , Parkinson Disease/therapy , Synucleinopathies/therapy , alpha-Synuclein/metabolismABSTRACT
Down syndrome (DS) is caused by trisomy of chromosome 21, which leads to a propensity to develop amyloid ß (Aß) brain pathology in early adulthood followed later by cognitive and behavioral deterioration. Characterization of the Aß 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 Aß species. Immunohistochemical staining using antibodies towards Aß was also performed. Elevated levels of soluble Aß protofibrils and insoluble Aßx-40 and Aßx-42 in formic acid brain extracts, and elevated immunohistochemical staining of Aß 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.
Subject(s)
Alzheimer Disease/metabolism , Amyloid/metabolism , Brain/metabolism , Down Syndrome/metabolism , Aged , Aged, 80 and over , Alzheimer Disease/pathology , Brain/pathology , Down Syndrome/pathology , Female , Humans , Male , Middle AgedABSTRACT
The major characteristics of Alzheimer's disease (AD) are amyloid plaques, consisting of aggregated beta amyloid (Aß) 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 Aß 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 Aß species including oligomeric Aß. Mass spectrometry was then used to evaluate the presence of Aß species in the different patient groups. A large number of Aß peptides were identified including Aß1-X, 2-X, 3-X, 4-X, 5-X, 11-X, and Aß peptides extended N terminally of the BACE1 cleavage site and ending at amino 15 in the Aß sequence APP/Aß(-X to 15), as well as peptides post-translationally modified by pyroglutamate formation. Most Aß peptides had higher abundance in AD and DS compared to controls, except the APP/Aß(-X to 15) peptides which were most abundant in DS followed by controls and AD. Furthermore, the abundancies of AßX-40 and AßX-34 were increased in DS compared with AD. Aß1-40, Aß1-42, and Aß4-42 were identified as the main constitutes of protofibrils (IP'd using mAb158) and higher relative Aß1-42 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 Aß peptides found in AD were also present in DS indicating similar pathways of Aß peptide production, degradation and accumulation, except for APP/Aß(-X to 15). Likewise, the Aß 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.
Subject(s)
Alzheimer Disease/pathology , Amyloid beta-Peptides/metabolism , Brain/pathology , Down Syndrome/pathology , Peptide Fragments/metabolism , Aged , Aged, 80 and over , Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/analysis , Aspartic Acid Endopeptidases/metabolism , Case-Control Studies , Female , Humans , Male , Mass Spectrometry , Middle Aged , Peptide Fragments/analysis , Protein AggregatesABSTRACT
BACKGROUND: Currently, there is no established biomarker for Parkinson's disease (PD) and easily accessible biomarkers are crucial for developing disease-modifying treatments. OBJECTIVE: To develop a novel method to quantify cerebrospinal fluid (CSF) levels of α-synuclein protofibrils (α-syn PF) and apply it to clinical cohorts of patients with PD and atypical parkinsonian disorders. METHODS: A cohort composed of 49 patients with PD, 12 with corticobasal degeneration (CBD), 22 with progressive supranuclear palsy, and 33 controls, that visited the memory clinic but had no biomarker signs of Alzheimer's disease (AD, tau<350âpg/mL, amyloid-beta 42 (Aß42)>530âpg/mL, and phosphorylated tau (p-tau)<60âpg/mL) was used in this study. The CSF samples were analyzed with the Single molecule array (Simoa) technology. Total α-synuclein (α-syn) levels were analyzed with a commercial ELISA-kit. RESULTS: The assay is specific to α-syn PF, with no cross-reactivity to monomeric α-syn, or the ß- and γ-synuclein variants. CSF α-syn PF levels were increased in PD compared with controls (62.1 and 40.4âpg/mL, respectively, pâ=â0.03), and CBD (62.1 and 34.2âpg/mL, respectively, pâ=â0.02). The accuracy of predicting PD using α-syn PF is significantly different from controls (area under the curve 0.68, pâ=â0.0097) with a sensitivity of 62.8% and specificity of 67.7%. Levels of total α-syn were significantly different between the PD and CBD groups (pâ=â0.04). CONCLUSION: The developed method specifically quantifies α-syn PF in human CSF with increased concentrations in PD, but with an overlap with asymptomatic elderly controls.
Subject(s)
Parkinson Disease/cerebrospinal fluid , Parkinsonian Disorders/cerebrospinal fluid , alpha-Synuclein/cerebrospinal fluid , Aged , Biomarkers/cerebrospinal fluid , Female , Humans , Immunoassay , Male , Middle Aged , Supranuclear Palsy, Progressive/cerebrospinal fluidABSTRACT
Scientists increasingly find themselves working in bilateral drug development alliances. Alliances are conceptually simple, but operationally challenging, resulting in the value-eroding misalignment and delays that alliances often experience. This case study of an exemplary collaboration between a small biotech and a global biopharmaceutical company is based on 15 interviews and a lessons-learned workshop conducted with the principal alliance team members. We outline five repeatable practices identified as contributing to their success that other alliance teams can follow.
Subject(s)
Drug Development/methods , Drug Industry/methods , Humans , Intersectoral Collaboration , Partnership PracticeABSTRACT
A growing body of pathological, biomarker, genetic, and mechanistic data suggests that amyloid accumulation, as a result of changes in production, processing, and/or clearance of brain amyloid-ß peptide (Aß) concentrations, plays a key role in the pathogenesis of Alzheimer's disease (AD). Beta-secretase 1 (BACE1) mediates the first step in the processing of amyloid-ß protein precursor (AßPP) to Aß peptides, with the soluble N terminal fragment of AßPP (sAßPPß) as a direct product, and BACE1 inhibition is an attractive target for therapeutic intervention to reduce the production of Aß. Here, we report the in vitro and in vivo pharmacological profile of AZD3293, a potent, highly permeable, orally active, blood-brain barrier (BBB) penetrating, BACE1 inhibitor with unique slow off-rate kinetics. The in vitro potency of AZD3293 was demonstrated in several cellular models, including primary cortical neurons. In vivo in mice, guinea pigs, and dogs, AZD3293 displayed significant dose- and time-dependent reductions in plasma, cerebrospinal fluid, and brain concentrations of Aß40, Aß42, and sAßPPß. The in vitro potency of AZD3293 in mouse and guinea pig primary cortical neuronal cells was correlated to the in vivo potency expressed as free AZD3293 concentrations in mouse and guinea pig brains. In mice and dogs, the slow off-rate from BACE1 may have translated into a prolongation of the observed effect beyond the turnover rate of Aß. The preclinical data strongly support the clinical development of AZD3293, and patients with AD are currently being recruited into a combined Phase 2/3 study to test the disease-modifying properties of AZD3293.
Subject(s)
Amyloid Precursor Protein Secretases/antagonists & inhibitors , Enzyme Inhibitors/administration & dosage , Enzyme Inhibitors/pharmacokinetics , Imidazoles/administration & dosage , Imidazoles/pharmacokinetics , Spiro Compounds/administration & dosage , Spiro Compounds/pharmacokinetics , Administration, Oral , Amyloid beta-Peptides/metabolism , Animals , Blood Chemical Analysis , Blood-Brain Barrier/drug effects , Blood-Brain Barrier/enzymology , Brain/drug effects , Brain/enzymology , Dogs , Dose-Response Relationship, Drug , Drug Evaluation, Preclinical , Female , Guinea Pigs , Humans , Kinetics , Male , Mice, Inbred C57BL , Peptide Fragments/metabolismABSTRACT
A major hallmark of Alzheimer's disease (AD) is the deposition of amyloid-ß (Aß) peptides in amyloid plaques. Aß peptides are produced by sequential cleavage of the amyloid precursor protein by the ß amyloid cleaving enzyme (BACE) and the γ-secretase (γ-sec) complex. Pharmacological treatments that decrease brain levels of in particular the toxic Aß42 peptide are thought to be promising approaches for AD disease modification. Potent and selective BACE1 inhibitors as well as γ-sec modulators (GSMs) have been designed. Pharmacological intervention of secretase function is not without risks of either on- or off-target adverse effects. One way of improving the therapeutic window could be to combine treatment on multiple targets, using smaller individual doses and thereby minimizing adverse effect liability. We show that combined treatment of primary cortical neurons with a BACE1 inhibitor and a GSM gives an additive effect on Aß42 level change compared with the individual treatments. We extend this finding to C57BL/6 mice, where the combined treatment results in reduction of brain Aß42 levels reflecting the sum of the individual treatment efficacies. These results show that pharmacological targeting of two amyloid precursor protein processing steps is feasible without negatively interfering with the mechanism of action on individual targets. We conclude that targeting Aß production by combining a BACE inhibitor and a GSM could be a viable approach for therapeutic intervention in AD modification.
Subject(s)
Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/metabolism , Alzheimer Disease/drug therapy , Alzheimer Disease/metabolism , Amyloid beta-Peptides/biosynthesis , Animals , Aspartic Acid Endopeptidases/antagonists & inhibitors , Aspartic Acid Endopeptidases/metabolism , Brain/drug effects , Brain/metabolism , Drug Synergism , Female , HEK293 Cells , Humans , Mice , Mice, Inbred C57BL , Peptide Fragments/metabolism , Protease Inhibitors/administration & dosage , Protease Inhibitors/pharmacology , Pyrans/administration & dosage , Pyrans/pharmacology , Pyrimidines/administration & dosage , Pyrimidines/pharmacologyABSTRACT
Aggregation of amyloid beta (Aß) peptides and the subsequent neural plaque formation is a central aspect of Alzheimer's disease. Various strategies to reduce Aß load in the brain are therefore intensely pursued. It has been hypothesized that reducing Aß peptides in the periphery, that is in organs outside the brain, would be a way to diminish Aß levels and plaque load in the brain. In this report, we put this peripheral sink hypothesis to test by investigating how selective inhibition of Aß production in the periphery using a ß-secretase (BACE)1 inhibitor or reduced BACE1 gene dosage affects Aß load in the brain. Selective inhibition of peripheral BACE1 activity in wild-type mice or mice over-expressing amyloid precursor protein (APPswe transgenic mice; Tg2576) reduced Aß levels in the periphery but not in the brain, not even after chronic treatment over several months. In contrast, a BACE1 inhibitor with improved brain disposition reduced Aß levels in both brain and periphery already after acute dosing. Mice heterozygous for BACE1, displayed a 62% reduction in plasma Aß40, whereas brain Aß40 was only lowered by 11%. These data suggest that reduction of Aß in the periphery is not sufficient to reduce brain Aß levels and that BACE1 is not the rate-limiting enzyme for Aß processing in the brain. This provides evidence against the peripheral sink hypothesis and suggests that a decrease in Aß via BACE1 inhibition would need to be carried out in the brain. Aggregation of amyloid beta (Aß) peptides in the brain is a central aspect of Alzheimer's disease. In this study, we demonstrate that inhibition of Aß formation by BACE1 inhibitors needs to be carried out in the brain and that reduction of Aß in the periphery is not sufficient to reduce brain Aß levels. This information is useful for developing future Aß-targeting therapies for Alzheimer's disease.
Subject(s)
Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/biosynthesis , Aspartic Acid Endopeptidases/antagonists & inhibitors , Aspartic Acid Endopeptidases/metabolism , Brain/enzymology , Animals , Brain/drug effects , Caco-2 Cells , Cricetinae , Dose-Response Relationship, Drug , Enzyme Inhibitors/pharmacology , Female , Humans , Mice , Mice, Inbred C57BL , Mice, TransgenicABSTRACT
Corrected QT interval (QTc) prolongation in humans is usually predictable based on results from preclinical findings. This study confirms the signal from preclinical cardiac repolarization models (human ether-a-go-go-related gene, guinea pig monophasic action potential, and dog telemetry) on the clinical effects on the QTc interval. A thorough QT/QTc study is generally required for bioavailable pharmaceutical compounds to determine whether or not a drug shows a QTc effect above a threshold of regulatory interest. However, as demonstrated in this AZD3839 [(S)-1-(2-(difluoromethyl)pyridin-4-yl)-4-fluoro-1-(3-(pyrimidin-5-yl)phenyl)-1H-isoindol-3-amine hemifumarate] single-ascending-dose (SAD) study, high-resolution digital electrocardiogram data, in combination with adequate efficacy biomarker and pharmacokinetic data and nonlinear mixed effects modeling, can provide the basis to safely explore the margins to allow for robust modeling of clinical effect versus the electrophysiological risk marker. We also conclude that a carefully conducted SAD study may provide reliable data for effective early strategic decision making ahead of the thorough QT/QTc study.
Subject(s)
Amyloid Precursor Protein Secretases/antagonists & inhibitors , Electrocardiography/drug effects , Indoles/pharmacology , Pyrimidines/pharmacology , Animals , Arterial Pressure/drug effects , Dogs , Dose-Response Relationship, Drug , Double-Blind Method , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Ether-A-Go-Go Potassium Channels/genetics , Guinea Pigs , Humans , Indoles/adverse effects , Male , Models, Biological , Pyrimidines/adverse effectsABSTRACT
PURPOSE: The aims were to quantify the in vivo time-course between the oral dose, the plasma and brain exposure and the inhibitory effect on Amyloid ß (Aß) in brain and cerebrospinal fluid, and to establish the correlation between in vitro and in vivo potency of novel ß-secretase (BACE1) inhibitors. METHODS: BACE1-mediated inhibition of Aß was quantified in in vivo dose- and/or time-response studies and in vitro in SH-SY5Y cells, N2A cells, and primary cortical neurons (PCN). An indirect response model with inhibition on Aß production rate was used to estimate unbound in vivo IC 50 in a population pharmacokinetic-pharmacodynamic modeling approach. RESULTS: Estimated in vivo inhibitory potencies varied between 1 and 1,000 nM. The turnover half-life of Aß40 in brain was predicted to be 0.5 h in mouse and 1 h in guinea pig. An excellent correlation between PCN and in vivo potency was observed. Moreover, a strong correlation in potency was found between human SH-SY5Y cells and mouse PCN, being 4.5-fold larger in SH-SY5Y cells. CONCLUSION: The strong in vivo-in vitro correlation increased the confidence in using human cell lines for screening and optimization of BACE1 inhibitors. This can optimize the design and reduce the number of preclinical in vivo effect studies.
Subject(s)
Alzheimer Disease/drug therapy , Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid beta-Peptides/antagonists & inhibitors , Amyloid beta-Peptides/metabolism , Aspartic Acid Endopeptidases/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Neurons/drug effects , Alzheimer Disease/blood , Alzheimer Disease/cerebrospinal fluid , Alzheimer Disease/metabolism , Amyloid beta-Peptides/blood , Amyloid beta-Peptides/cerebrospinal fluid , Animals , Brain/cytology , Brain/drug effects , Brain/metabolism , Cell Line , Cells, Cultured , Female , Guinea Pigs , Humans , Male , Mice , Mice, Inbred C57BL , Models, Biological , Neurons/metabolismABSTRACT
Modulating deposition of Aß-containing plaques in the brain may be beneficial in treating Alzheimer's disease. ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibitors have been shown to reduce Aß in plasma and CSF in healthy volunteers. In this study safety, pharmacokinetics and pharmacodynamics that is reduction of the plasma biomarkers Aß40 and Aß42 , of the BACE1 inhibitor AZD3839 were evaluated. Single oral ascending doses (1-300 mg) of AZD3839 were administered to 54 young healthy volunteers in a randomized, double-blind, placebo-controlled study. The data was analyzed using non-linear mixed effects modeling. AZD3839 reduced Aß40 and Aß42 in plasma with estimated potencies (EC50 ) of 46 and 59 nM, respectively, and a maximum effect of approximately 55%. This was in excellent agreement with the concentration-response relationships obtained in mouse and guinea pig. AZD3839 exposure displayed non-linear kinetics, described by a three-compartment model with a saturated binding compartment and an increase in bioavailability with dose. AZD3839 was safe, although, a dose-dependent QTcF prolongation was observed (mean 20 milliseconds at 300 mg). In conclusion, AZD3839 reduced plasma Aß40 and Aß42 , demonstrating clinical peripheral proof of mechanism. Pre-clinical models were predictive for the effect of AZD3839 on the human plasma biomarker in a strictly quantitative manner.
Subject(s)
Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid beta-Peptides/blood , Aspartic Acid Endopeptidases/antagonists & inhibitors , Indoles/administration & dosage , Indoles/pharmacokinetics , Peptide Fragments/blood , Protease Inhibitors/administration & dosage , Protease Inhibitors/pharmacokinetics , Pyrimidines/administration & dosage , Pyrimidines/pharmacokinetics , Administration, Oral , Adolescent , Adult , Amyloid Precursor Protein Secretases/metabolism , Aspartic Acid Endopeptidases/metabolism , Biomarkers/blood , Computer Simulation , Double-Blind Method , Down-Regulation , Drug Administration Schedule , Female , Healthy Volunteers , Humans , Indoles/adverse effects , Indoles/blood , London , Male , Middle Aged , Models, Biological , Nonlinear Dynamics , Protease Inhibitors/adverse effects , Protease Inhibitors/blood , Pyrimidines/adverse effects , Pyrimidines/blood , Young AdultABSTRACT
Brown adipose tissue is unusual in that the neurotransmitter norepinephrine influences cell destiny in ways generally associated with effects of classical growth factors: regulation of cell proliferation, of apoptosis, and progression of differentiation. The norepinephrine effects are mediated through G-protein-coupled receptors; further mediation of such stimulation to e.g. Erk1/2 activation is in cell biology in general accepted to occur through transactivation of the EGF receptor (by external or internal pathways). We have examined here the significance of such transactivation in brown adipocytes. Stimulation of mature brown adipocytes with cirazoline (α1-adrenoceptor coupled via Gq), clonidine (α2 via Gi) or CL316243 (ß3 via Gs) or via ß1-receptors significantly activated Erk1/2. Pretreatment with the EGF receptor kinase inhibitor AG1478 had, remarkably, no significant effect on Erk1/2 activation induced by any of these adrenergic agonists (although it fully abolished EGF-induced Erk1/2 activation), demonstrating absence of EGF receptor-mediated transactivation. Results with brown preadipocytes (cells in more proliferative states) were not qualitatively different. Joint stimulation of all adrenoceptors with norepinephrine did not result in synergism on Erk1/2 activation. AG1478 action on EGF-stimulated Erk1/2 phosphorylation showed a sharp concentration-response relationship (IC50 0.3µM); a minor apparent effect of AG1478 on norepinephrine-stimulated Erk1/2 phosphorylation showed nonspecific kinetics, implying caution in interpretation of partial effects of AG1478 as reported in other systems. Transactivation of the EGF receptor is clearly not a universal prerequisite for coupling of G-protein coupled receptors to Erk1/2 signalling cascades.
Subject(s)
Adipocytes, Brown/metabolism , ErbB Receptors/metabolism , Mitogen-Activated Protein Kinase 1/metabolism , Mitogen-Activated Protein Kinase 3/metabolism , Receptors, Adrenergic/metabolism , Adipocytes, Brown/drug effects , Adrenergic Antagonists/pharmacology , Adrenergic alpha-1 Receptor Agonists/pharmacology , Adrenergic alpha-2 Receptor Agonists/pharmacology , Adrenergic beta-1 Receptor Agonists/pharmacology , Adrenergic beta-3 Receptor Agonists/pharmacology , Animals , Clonidine/pharmacology , Dioxoles/pharmacology , Dose-Response Relationship, Drug , Drug Synergism , ErbB Receptors/antagonists & inhibitors , Imidazoles/pharmacology , MAP Kinase Signaling System/drug effects , Male , Mice , Norepinephrine/pharmacology , Phosphorylation , Quinazolines/pharmacology , Tyrphostins/pharmacologyABSTRACT
Aß, the product of APP (amyloid precursor protein), has been implicated in the pathophysiology of Alzheimer's disease (AD). ß-Site APP cleaving enzyme1 (BACE1) is the enzyme initiating the processing of the APP to Aß peptides. Small molecule BACE1 inhibitors are expected to decrease Aß-peptide generation and thereby reduce amyloid plaque formation in the brain, a neuropathological hallmark of AD. BACE1 inhibition thus addresses a key mechanism in AD and its potential as a therapeutic target is currently being addressed in clinical studies. Here, we report the discovery and the pharmacokinetic and pharmacodynamic properties of BACE1 inhibitor AZ-4217, a high potency compound (IC50 160 pM in human SH-SY5Y cells) with an excellent in vivo efficacy. Central efficacy of BACE1 inhibition was observed after a single dose in C57BL/6 mice, guinea pigs, and in an APP transgenic mouse model of cerebral amyloidosis (Tg2576). Furthermore, we demonstrate that in a 1 month treatment paradigm BACE1 inhibition of Aß production does lower amyloid deposition in 12-month-old Tg2576 mice. These results strongly support BACE1 inhibition as concretely impacting amyloid deposition and therefore potentially an important approach for therapeutic intervention in AD.
Subject(s)
Alzheimer Disease/pathology , Amyloid/metabolism , Enzyme Inhibitors/pharmacology , Neurons/drug effects , Neurons/metabolism , Alzheimer Disease/drug therapy , Alzheimer Disease/genetics , Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Animals , Aspartic Acid Endopeptidases/antagonists & inhibitors , Aspartic Acid Endopeptidases/metabolism , Cells, Cultured , Cerebral Cortex/pathology , Disease Models, Animal , Dose-Response Relationship, Drug , Embryo, Mammalian , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/therapeutic use , Female , Guinea Pigs , Humans , Isoindoles/pharmacology , Isoindoles/therapeutic use , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Mutation/genetics , Peptide Fragments/metabolism , Pyridones/pharmacology , Pyridones/therapeutic use , Time FactorsABSTRACT
Abnormal tau phosphorylation resulting in detachment of tau from microtubules and aggregation are critical events in neuronal dysfunction, degeneration, and neurofibrillary pathology seen in Alzheimer's disease. Glycogen synthase kinase-3ß (GSK3ß) is a key target for drug discovery in the treatment of Alzheimer's disease and related tauopathies because of its potential to abnormally phosphorylate proteins and contribute to synaptic degeneration. We report the discovery of AZD1080, a potent and selective GSK3 inhibitor that demonstrates peripheral target engagement in Phase 1 clinical studies. AZD1080 inhibits tau phosphorylation in cells expressing human tau and in intact rat brain. Interestingly, subchronic but not acute administration with AZD1080 reverses MK-801-induced deficits, measured by long-term potentiation in hippocampal slices and in a cognitive test in mice, suggesting that reversal of synaptic plasticity deficits in dysfunctional systems requires longer term modifications of proteins downstream of GSK3ß signaling. The inhibitory pattern on tau phosphorylation reveals a prolonged pharmacodynamic effect predicting less frequent dosing in humans. Consistent with the preclinical data, in multiple ascending dose studies in healthy volunteers, a prolonged suppression of glycogen synthase activity was observed in blood mononuclear cells providing evidence of peripheral target engagement with a selective GSK3 inhibitor in humans.
Subject(s)
Enzyme Inhibitors/pharmacology , Glycogen Synthase Kinase 3/antagonists & inhibitors , Long-Term Potentiation/drug effects , tau Proteins/metabolism , Animals , Cell Line, Transformed , Cognition Disorders/chemically induced , Cognition Disorders/drug therapy , Crystallography , Disease Models, Animal , Dizocilpine Maleate/toxicity , Dose-Response Relationship, Drug , Double-Blind Method , Electric Stimulation , Enzyme Inhibitors/chemistry , Excitatory Amino Acid Antagonists/toxicity , Excitatory Postsynaptic Potentials/drug effects , Glycogen Synthase/metabolism , Glycogen Synthase Kinase 3/metabolism , Hippocampus/cytology , Hippocampus/drug effects , Humans , In Vitro Techniques , Indoles/pharmacology , Indoles/therapeutic use , Leukocytes, Mononuclear/drug effects , Long-Term Potentiation/physiology , Male , Mice , Middle Aged , Phosphorylation/drug effects , Protein Binding/drug effects , Protein Kinases/metabolism , Pyridines/pharmacology , Pyridines/therapeutic use , Rats , Rats, Sprague-DawleyABSTRACT
By use of iterative design aided by predictive models for target affinity, brain permeability, and hERG activity, novel and diverse compounds based on cyclic amidine and guanidine cores were synthesized with the goal of finding BACE-1 inhibitors as a treatment for Alzheimer's disease. Since synthesis feasibility had low priority in the design of the cores, an extensive synthesis effort was needed to make the relevant compounds. Syntheses of these compounds are reported, together with physicochemical properties and structure-activity relationships based on in vitro data. Four crystal structures of diverse amidines binding in the active site are deposited and discussed. Inhibitors of BACE-1 with 3 µM to 32 nM potencies in cells are shown, together with data on in vivo brain exposure levels for four compounds. The results presented show the importance of the core structure for the profile of the final compounds.
Subject(s)
Amidines/chemical synthesis , Amyloid Precursor Protein Secretases/antagonists & inhibitors , Aspartic Acid Endopeptidases/antagonists & inhibitors , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Guanidines/chemical synthesis , Amidines/chemistry , Amidines/pharmacology , Amyloid Precursor Protein Secretases/chemistry , Animals , Aspartic Acid Endopeptidases/chemistry , Brain/metabolism , CHO Cells , Cell Line, Tumor , Cell Membrane Permeability , Computer Simulation , Cricetinae , Crystallography, X-Ray , Dogs , Drug Stability , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels/metabolism , Female , Guanidines/chemistry , Guanidines/pharmacology , Humans , Madin Darby Canine Kidney Cells , Mice , Mice, Inbred C57BL , Protein Conformation , Quantitative Structure-Activity Relationship , StereoisomerismABSTRACT
ß-Site amyloid precursor protein cleaving enzyme1 (BACE1) is one of the key enzymes involved in the processing of the amyloid precursor protein (APP) and formation of amyloid ß peptide (Aß) species. Because cerebral deposition of Aß species might be critical for the pathogenesis of Alzheimer disease, BACE1 has emerged as a key target for the treatment of this disease. Here, we report the discovery and comprehensive preclinical characterization of AZD3839, a potent and selective inhibitor of human BACE1. AZD3839 was identified using fragment-based screening and structure-based design. In a concentration-dependent manner, AZD3839 inhibited BACE1 activity in a biochemical fluorescence resonance energy transfer (FRET) assay, Aß and sAPPß release from modified and wild-type human SH-SY5Y cells and mouse N2A cells as well as from mouse and guinea pig primary cortical neurons. Selectivity against BACE2 and cathepsin D was 14 and >1000-fold, respectively. AZD3839 exhibited dose- and time-dependent lowering of plasma, brain, and cerebrospinal fluid Aß levels in mouse, guinea pig, and non-human primate. Pharmacokinetic/pharmacodynamic analyses of mouse and guinea pig data showed a good correlation between the potency of AZD3839 in primary cortical neurons and in vivo brain effects. These results suggest that AZD3839 effectively reduces the levels of Aß in brain, CSF, and plasma in several preclinical species. It might, therefore, have disease-modifying potential in the treatment of Alzheimer disease and related dementias. Based on the overall pharmacological profile and its drug like properties, AZD3839 has been progressed into Phase 1 clinical trials in man.
Subject(s)
Alzheimer Disease/drug therapy , Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid beta-Peptides/antagonists & inhibitors , Amyloid beta-Peptides/cerebrospinal fluid , Aspartic Acid Endopeptidases/antagonists & inhibitors , Indoles/pharmacology , Pyrimidines/pharmacology , Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/metabolism , Animals , Aspartic Acid Endopeptidases/metabolism , Biomarkers/metabolism , Brain/metabolism , Cathepsin D/metabolism , Cell Line , Disease Progression , Dose-Response Relationship, Drug , Drug Design , Drug Evaluation, Preclinical , Female , Fluorescence Resonance Energy Transfer/methods , Guinea Pigs , Humans , Male , Mice , Mice, Inbred C57BL , Treatment OutcomeABSTRACT
Amino-2H-imidazoles are described as a new class of BACE-1 inhibitors for the treatment of Alzheimer's disease. Synthetic methods, crystal structures, and structure-activity relationships for target activity, permeability, and hERG activity are reported and discussed. Compound (S)-1m was one of the most promising compounds in this report, with high potency in the cellular assay and a good overall profile. When guinea pigs were treated with compound (S)-1m, a concentration and time dependent decrease in Aß40 and Aß42 levels in plasma, brain, and CSF was observed. The maximum reduction of brain Aß was 40-50%, 1.5 h after oral dosing (100 µmol/kg). The results presented highlight the potential of this new class of BACE-1 inhibitors with good target potency and with low effect on hERG, in combination with a fair CNS exposure in vivo.
Subject(s)
Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid beta-Peptides/metabolism , Aspartic Acid Endopeptidases/antagonists & inhibitors , Brain/drug effects , Imidazoles/chemical synthesis , Peptide Fragments/metabolism , Amyloid Precursor Protein Secretases/chemistry , Amyloid beta-Peptides/blood , Amyloid beta-Peptides/cerebrospinal fluid , Animals , Aspartic Acid Endopeptidases/chemistry , Brain/metabolism , Cell Line , Crystallography, X-Ray , Dogs , Female , Guinea Pigs , Humans , Imidazoles/chemistry , Imidazoles/pharmacology , Male , Mice , Mice, Inbred C57BL , Models, Molecular , Molecular Structure , Peptide Fragments/blood , Peptide Fragments/cerebrospinal fluid , Permeability , Stereoisomerism , Structure-Activity Relationship , Tissue DistributionABSTRACT
The evaluation of a series of aminoisoindoles as ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibitors and the discovery of a clinical candidate drug for Alzheimer's disease, (S)-32 (AZD3839), are described. The improvement in permeability properties by the introduction of fluorine adjacent to the amidine moiety, resulting in in vivo brain reduction of Aß40, is discussed. Due to the basic nature of these compounds, they displayed affinity for the human ether-a-go-go related gene (hERG) ion channel. Different ways to reduce hERG inhibition and increase hERG margins for this series are described, culminating in (S)-16 and (R)-41 showing large in vitro margins with BACE1 cell IC(50) values of 8.6 and 0.16 nM, respectively, and hERG IC(50) values of 16 and 2.8 µM, respectively. Several compounds were advanced into pharmacodynamic studies and demonstrated significant reduction of ß-amyloid peptides in mouse brain following oral dosing.
Subject(s)
Amyloid Precursor Protein Secretases/antagonists & inhibitors , Aspartic Acid Endopeptidases/antagonists & inhibitors , Indoles/chemical synthesis , Pyrimidines/chemical synthesis , Administration, Oral , Alkynes/chemical synthesis , Alkynes/pharmacokinetics , Alkynes/pharmacology , Amides/chemical synthesis , Amides/pharmacokinetics , Amides/pharmacology , Amyloid Precursor Protein Secretases/chemistry , Amyloid beta-Peptides/metabolism , Animals , Aspartic Acid Endopeptidases/chemistry , Biological Availability , Brain/drug effects , Brain/metabolism , Cell Line , Crystallography, X-Ray , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Female , Fluorescence Resonance Energy Transfer , Humans , Hydrogen Bonding , Indoles/pharmacokinetics , Indoles/pharmacology , Mice , Mice, Inbred C57BL , Models, Molecular , Molecular Structure , Peptide Fragments/metabolism , Permeability , Pyrimidines/pharmacokinetics , Pyrimidines/pharmacology , Stereoisomerism , Structure-Activity RelationshipABSTRACT
The evaluation of a series of bicyclic aminoimidazoles as potent BACE-1 inhibitors is described. The crystal structures of compounds 14 and 23 in complex with BACE-1 reveal hydrogen bond interactions with the protein important for achieving potent inhibition. The optimization of permeability and efflux properties of the compounds is discussed as well as the importance of these properties for attaining in vivo brain efficacy. Compound (R)-25 was selected for evaluation in vivo in wild type mice and 1.5h after oral co-administration of 300µmol/kg (R)-25 and efflux inhibitor GF120918 the brain Aß40 level was reduced by 17% and the plasma Aß40 level by 76%.