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2.
J Pharmacol Exp Ther ; 377(1): 11-19, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33509901

RESUMO

Gain-of-function mutations in leucine-rich kinase 2 (LRRK2) are associated with increased incidence of Parkinson disease (PD); thus, pharmacological inhibition of LRRK2 kinase activity is postulated as a disease-modifying treatment of PD. Histomorphological changes in lungs of nonhuman primates (NHPs) treated with small-molecule LRRK2 kinase inhibitors have brought the safety of this treatment approach into question. Although it remains unclear how LRRK2 kinase inhibition affects the lung, continued studies in NHPs prove to be both cost- and resource-prohibitive. To develop a tractable alternative animal model platform, we dosed male mice in-diet with the potent, highly selective LRRK2 kinase inhibitor MLi-2 and induced histomorphological changes in lung within 1 week. Oral bolus dosing of MLi-2 at a frequency modeled to provide steady-state exposure equivalent to that achieved with in-diet dosing induced type II pneumocyte vacuolation, suggesting pulmonary changes require sustained LRRK2 kinase inhibition. Treating mice with MLi-2 in-diet for up to 6 months resulted in type II pneumocyte vacuolation that progressed only modestly over time and was fully reversible after withdrawal of MLi-2. Immunohistochemical analysis of lung revealed a significant increase in prosurfactant protein C staining within type II pneumocytes. In the present study, we demonstrated the kinetics for onset, progression, and rapid reversibility of chronic LRRK2 kinase inhibitor effects on lung histomorphology in rodents and provide further evidence for the derisking of safety and tolerability concerns for chronic LRRK2 kinase inhibition in PD. SIGNIFICANCE STATEMENT: We have defined a mouse model by which the on-target lung effects of leucine-rich kinase 2 (LRRK2) kinase inhibition can be monitored, whereas previous in vivo testing relied solely on nonhuman primates. Data serve to derisk long-term treatment with LRRK2 kinase inhibitors, as all lung changes were mild and readily reversible.


Assuntos
Células Epiteliais Alveolares/efeitos dos fármacos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/antagonistas & inibidores , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/metabolismo , Animais , Indazóis/administração & dosagem , Indazóis/farmacologia , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Morfolinas/administração & dosagem , Morfolinas/farmacologia , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacologia , Proteína C Associada a Surfactante Pulmonar/genética , Proteína C Associada a Surfactante Pulmonar/metabolismo , Pirimidinas/administração & dosagem , Pirimidinas/farmacologia
3.
Brain ; 143(12): 3816-3826, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-33253354

RESUMO

In the phase 3 EPOCH trial (Clinicaltrials.gov; NCT01739348), treatment with the BACE inhibitor verubecestat failed to improve cognition in patients with mild-to-moderate Alzheimer's disease, but was associated with reduced hippocampal volume after 78 weeks as assessed by MRI. The aims of the present exploratory analyses were to: (i) characterize the effect of verubecestat on brain volume by evaluating the time course of volumetric MRI changes for a variety of brain regions; and (ii) understand the mechanism through which verubecestat might cause hippocampal (and other brain region) volume loss by assessing its relationship to measures of amyloid, neurodegeneration, and cognition. Participants were aged 55-85 years with probable Alzheimer's disease dementia and a Mini Mental State Examination score ≥15 and ≤26. MRIs were obtained at baseline and at Weeks 13, 26, 52 and 78 of treatment. MRIs were segmented using Freesurfer and analysed using a tensor-based morphometry method. PET amyloid data were obtained with 18F-flutemetamol (Vizamyl®) at baseline and Week 78. Standardized uptake value ratios were generated with subcortical white matter as a reference region. Neurofilament light chain in the CSF was assessed as a biomarker of neurodegeneration. Compared with placebo, verubecestat showed increased MRI brain volume loss at Week 13 with no evidence of additional loss through Week 78. The verubecestat-related volumetric MRI loss occurred predominantly in amyloid-rich brain regions. Correlations between amyloid burden at baseline and verubecestat-related volumetric MRI reductions were not significant (r = 0.05 to 0.26, P-values > 0.27). There were no significant differences between verubecestat and placebo in changes from baseline in CSF levels of neurofilament light chain at Week 78 (increases of 7.2 and 14.6 pg/ml for verubecestat versus 19.7 pg/ml for placebo, P-values ≥ 0.1). There was a moderate correlation between volumetric MRI changes and cognitive decline in all groups including placebo at Week 78 (e.g. r = -0.45 to -0.55, P < 0.001 for whole brain), but the correlations were smaller at Week 13 and significant only for the verubecestat groups (e.g. r = -0.15 and -0.11, P < 0.04 for whole brain). Our results suggest that the verubecestat-associated MRI brain volume loss is not due to generalized, progressive neurodegeneration, but may be mediated by specific effects on BACE-related amyloid processes.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Encéfalo/diagnóstico por imagem , Óxidos S-Cíclicos/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Tiadiazinas/uso terapêutico , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/diagnóstico por imagem , Doença de Alzheimer/psicologia , Peptídeos beta-Amiloides/metabolismo , Encéfalo/efeitos dos fármacos , Imagem de Tensor de Difusão , Método Duplo-Cego , Feminino , Hipocampo/diagnóstico por imagem , Hipocampo/efeitos dos fármacos , Humanos , Imageamento por Ressonância Magnética , Masculino , Testes de Estado Mental e Demência , Pessoa de Meia-Idade , Proteínas de Neurofilamentos/líquido cefalorraquidiano , Tomografia por Emissão de Pósitrons , Resultado do Tratamento , Substância Branca/diagnóstico por imagem , Substância Branca/metabolismo
4.
Neuroimage ; 213: 116725, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32173412

RESUMO

Functional magnetic resonance imaging (fMRI) is a valuable tool for studying neural activations in the central nervous system of animals due to its wide spatial coverage and non-invasive nature. However, the advantages of fMRI have not been fully realized in functional studies in mice, especially in the olfactory system, possibly due to the lack of suitable anesthesia protocols with spontaneous breathing. Since mice are widely used in biomedical research, it is desirable to evaluate different anesthesia protocols for olfactory fMRI studies in mice. Dexmedetomidine (DEX) as a sedative/anesthetic has been introduced to fMRI studies in mice, but it has a limited anesthesia duration. To extend the anesthesia duration, DEX has been combined with a low dose of isoflurane (ISO) or ketamine (KET) in previous functional studies in mice. In this report, olfactory fMRI studies were performed under three anesthesia protocols (DEX alone, DEX/ISO, and DEX/KET) in three different groups of mice. Isoamyl-acetate was used as an odorant, and the odorant-induced neural activations were measured by blood oxygenation-level dependent (BOLD) fMRI. BOLD fMRI responses were observed in the olfactory bulb (OB), anterior olfactory nuclei (AON), and piriform cortex (Pir). Interestingly, BOLD fMRI activations were also observed in the prefrontal cortical region (PFC), which are most likely caused by the draining vein effect. The response in the OB showed no adaptation to either repeated odor stimulations or continuous odor exposure, but the response in the Pir showed adaptation during the continuous odor exposure. The data also shows that ISO suppresses the olfactory response in the OB and AON, while KET enhances the olfactory response in the Pir. Thus, DEX/KET should be an attractive anesthesia for olfactory fMRI in mice.


Assuntos
Dexmedetomidina/farmacologia , Isoflurano/farmacologia , Ketamina/farmacologia , Bulbo Olfatório/efeitos dos fármacos , Percepção Olfatória/efeitos dos fármacos , Anestésicos/farmacologia , Animais , Hipnóticos e Sedativos/farmacologia , Imageamento por Ressonância Magnética/métodos , Camundongos , Modelos Animais
5.
J Pharmacol Exp Ther ; 365(3): 507-518, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29563326

RESUMO

The abnormal accumulation of amyloid-ß (Aß) in the brain parenchyma has been posited as a central event in the pathophysiology of Alzheimer's disease. Recently, we have proposed a systems pharmacology model of the amyloid precursor protein (APP) pathway, describing the Aß APP metabolite responses (Aß40, Aß42, sAPPα, and sAPPß) to ß-secretase 1 (BACE1) inhibition. In this investigation this model was challenged to describe Aß dynamics following γ-secretase (GS) inhibition. This led an extended systems pharmacology model, with separate descriptions to characterize the sequential cleavage steps of APP by BACE1 and GS, to describe the differences in Aß response to their respective inhibition. Following GS inhibition, a lower Aß40 formation rate constant was observed, compared with BACE1 inhibition. Both BACE1 and GS inhibition were predicted to lower Aß oligomer levels. Further model refinement and new data may be helpful to fully understand the difference in Aß dynamics following BACE1 versus GS inhibition.


Assuntos
Secretases da Proteína Precursora do Amiloide/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Modelos Biológicos , Proteólise , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Animais , Inibidores Enzimáticos/farmacologia , Macaca mulatta , Proteólise/efeitos dos fármacos
7.
J Pharmacol Exp Ther ; 357(1): 205-16, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26826190

RESUMO

The deposition of amyloid-ß (Aß) oligomers in brain parenchyma has been implicated in the pathophysiology of Alzheimer's disease. Here we present a systems pharmacology model describing the changes in the amyloid precursor protein (APP) pathway after administration of three different doses (10, 30, and 125 mg/kg) of the ß-secretase 1 (BACE1) inhibitor MBi-5 in cisterna magna ported rhesus monkeys. The time course of the MBi-5 concentration in plasma and cerebrospinal fluid (CSF) was analyzed in conjunction with the effect on the concentrations of the APP metabolites Aß42, Aß40, soluble ß-amyloid precursor protein (sAPP) α, and sAPPß in CSF. The systems pharmacology model contained expressions to describe the production, elimination, and brain-to-CSF transport for the APP metabolites. Upon administration of MBi-5, a dose-dependent increase of the metabolite sAPPα and dose-dependent decreases of sAPPß and Aß were observed. Maximal inhibition of BACE1 was close to 100% and the IC50 value was 0.0256 µM (95% confidence interval, 0.0137-0.0375). A differential effect of BACE1 inhibition on Aß40 and Aß42 was observed, with the Aß40 response being larger than the Aß42 response. This enabled the identification of an Aß42 oligomer pool in the systems pharmacology model. These findings indicate that decreases in monomeric Aß responses resulting from BACE1 inhibition are partially compensated by dissociation of Aß oligomers and suggest that BACE1 inhibition may also reduce the putatively neurotoxic oligomer pool.


Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/efeitos dos fármacos , Precursor de Proteína beta-Amiloide/metabolismo , Fragmentos de Peptídeos/metabolismo , Algoritmos , Peptídeos beta-Amiloides/efeitos dos fármacos , Animais , Biotransformação , Química Encefálica/efeitos dos fármacos , Cisterna Magna , Relação Dose-Resposta a Droga , Injeções , Macaca mulatta , Masculino , Modelos Estatísticos , Fragmentos de Peptídeos/efeitos dos fármacos
8.
J Neurosci ; 34(24): 8336-46, 2014 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-24920637

RESUMO

BACE, a ß-secretase, is an attractive potential disease-modifying therapeutic strategy for Alzheimer's disease (AD) as it results directly in the decrease of amyloid precursor protein (APP) processing through the ß-secretase pathway and a lowering of CNS amyloid-ß (Aß) levels. The interaction of the ß-secretase and α-secretase pathway-mediated processing of APP in the rhesus monkey (nonhuman primate; NHP) CNS is not understood. We hypothesized that CNS inhibition of BACE would result in decreased newly generated Aß and soluble APPß (sAPPß), with increased newly generated sAPPα. A stable isotope labeling kinetics experiment in NHPs was performed with a (13)C6-leucine infusion protocol to evaluate effects of BACE inhibition on CNS APP processing by measuring the kinetics of sAPPα, sAPPß, and Aß in CSF. Each NHP received a low, medium, or high dose of MBI-5 (BACE inhibitor) or vehicle in a four-way crossover design. CSF sAPPα, sAPPß, and Aß were measured by ELISA and newly incorporated label following immunoprecipitation and liquid chromatography-mass spectrometry. Concentrations, kinetics, and amount of newly generated APP fragments were calculated. sAPPß and sAPPα kinetics were similar, but both significantly slower than Aß. BACE inhibition resulted in decreased labeled sAPPß and Aß in CSF, without observable changes in labeled CSF sAPPα. ELISA concentrations of sAPPß and Aß both decreased and sAPPα increased. sAPPα increased by ELISA, with no difference by labeled sAPPα kinetics indicating increases in product may be due to APP shunting from the ß-secretase to the α-secretase pathway. These results provide a quantitative understanding of pharmacodynamic effects of BACE inhibition on NHP CNS, which can inform about target development.


Assuntos
Secretases da Proteína Precursora do Amiloide/líquido cefalorraquidiano , Peptídeos beta-Amiloides/líquido cefalorraquidiano , Precursor de Proteína beta-Amiloide/líquido cefalorraquidiano , Sistema Nervoso Central/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Isótopos de Carbono/metabolismo , Linhagem Celular Tumoral , Sistema Nervoso Central/efeitos dos fármacos , Estudos Cross-Over , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Humanos , Imunoprecipitação , Leucina/metabolismo , Macaca mulatta , Espectrometria de Massas , Neuroblastoma , Fragmentos de Peptídeos , Transfecção
9.
J Pharmacol Exp Ther ; 355(3): 397-409, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26407721

RESUMO

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most common genetic cause of familial and sporadic Parkinson's disease (PD). That the most prevalent mutation, G2019S, leads to increased kinase activity has led to a concerted effort to identify LRRK2 kinase inhibitors as a potential disease-modifying therapy for PD. An internal medicinal chemistry effort identified several potent and highly selective compounds with favorable drug-like properties. Here, we characterize the pharmacological properties of cis-2,6-dimethyl-4-(6-(5-(1-methylcyclopropoxy)-1H-indazol-3-yl)pyrimidin-4-yl)morpholine (MLi-2), a structurally novel, highly potent, and selective LRRK2 kinase inhibitor with central nervous system activity. MLi-2 exhibits exceptional potency in a purified LRRK2 kinase assay in vitro (IC50 = 0.76 nM), a cellular assay monitoring dephosphorylation of LRRK2 pSer935 LRRK2 (IC50 = 1.4 nM), and a radioligand competition binding assay (IC50 = 3.4 nM). MLi-2 has greater than 295-fold selectivity for over 300 kinases in addition to a diverse panel of receptors and ion channels. Acute oral and subchronic dosing in MLi-2 mice resulted in dose-dependent central and peripheral target inhibition over a 24-hour period as measured by dephosphorylation of pSer935 LRRK2. Treatment of MitoPark mice with MLi-2 was well tolerated over a 15-week period at brain and plasma exposures >100× the in vivo plasma IC50 for LRRK2 kinase inhibition as measured by pSer935 dephosphorylation. Morphologic changes in the lung, consistent with enlarged type II pneumocytes, were observed in MLi-2-treated MitoPark mice. These data demonstrate the suitability of MLi-2 as a compound to explore LRRK2 biology in cellular and animal models.


Assuntos
Antiparkinsonianos/efeitos adversos , Antiparkinsonianos/uso terapêutico , Indazóis/farmacologia , Inibidores de Proteínas Quinases/efeitos adversos , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Pirimidinas/farmacologia , Células Epiteliais Alveolares/efeitos dos fármacos , Células Epiteliais Alveolares/patologia , Animais , Comportamento Animal/efeitos dos fármacos , Ligação Competitiva , Encéfalo/metabolismo , Química Encefálica/efeitos dos fármacos , Linhagem Celular , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Relação Dose-Resposta a Droga , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mutação/genética , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/patologia , Doença de Parkinson/psicologia , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo
10.
J Neurochem ; 130(1): 4-28, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24646365

RESUMO

The ß-site APP cleaving enzymes 1 and 2 (BACE1 and BACE2) were initially identified as transmembrane aspartyl proteases cleaving the amyloid precursor protein (APP). BACE1 is a major drug target for Alzheimer's disease because BACE1-mediated cleavage of APP is the first step in the generation of the pathogenic amyloid-ß peptides. BACE1, which is highly expressed in the nervous system, is also required for myelination by cleaving neuregulin 1. Several recent proteomic and in vivo studies using BACE1- and BACE2-deficient mice demonstrate a much wider range of physiological substrates and functions for both proteases within and outside of the nervous system. For BACE1 this includes axon guidance, neurogenesis, muscle spindle formation, and neuronal network functions, whereas BACE2 was shown to be involved in pigmentation and pancreatic ß-cell function. This review highlights the recent progress in understanding cell biology, substrates, and functions of BACE proteases and discusses the therapeutic options and potential mechanism-based liabilities, in particular for BACE inhibitors in Alzheimer's disease. The protease BACE1 is a major drug target in Alzheimer disease. Together with its homolog BACE2, both proteases have an increasing number of functions within and outside of the nervous system. This review highlights recent progress in understanding cell biology, substrates, and functions of BACE proteases and discusses the therapeutic options and potential mechanism-based liabilities, in particular for BACE inhibitors in Alzheimer disease.


Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Secretases da Proteína Precursora do Amiloide/fisiologia , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Ácido Aspártico Endopeptidases/fisiologia , Líquido Intracelular/enzimologia , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/enzimologia , Precursor de Proteína beta-Amiloide/fisiologia , Animais , Previsões , Humanos , Líquido Intracelular/efeitos dos fármacos , Transporte Proteico/fisiologia
11.
CPT Pharmacometrics Syst Pharmacol ; 13(3): 449-463, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38078626

RESUMO

Alzheimer's disease (AD) is characterized by beta-amyloid (Aß) plaques in the brain and widespread neuronal damage. Because of the high drug attrition rates in AD, there is increased interest in characterizing neuroimmune responses to Aß plaques. In response to AD pathology, microglia are innate phagocytotic immune cells that transition into a neuroprotective state and form barriers around plaques. We seek to understand the role of microglia in modifying Aß dynamics and barrier formation. To quantify the influence of individual microglia behaviors (activation, chemotaxis, phagocytosis, and proliferation) on plaque size and barrier coverage, we developed an agent-based model to characterize the spatiotemporal interactions between microglia and Aß. Our model qualitatively reproduces mouse data trends where the fraction of microglia coverage decreases as plaques become larger. In our model, the time to microglial arrival at the plaque boundary is significantly negatively correlated (p < 0.0001) with plaque size, indicating the importance of the time to microglial activation for regulating plaque size. In addition, in silico behavioral knockout simulations show that phagocytosis knockouts have the strongest impact on plaque size, but modest impacts on microglial coverage and activation. In contrast, the chemotaxis knockouts had a strong impact on microglial coverage with a more modest impact on plaque volume and microglial activation. These simulations suggest that phagocytosis, chemotaxis, and replication of activated microglia have complex impacts on plaque volume and coverage, whereas microglial activation remains fairly robust to perturbations of these functions. Thus, our work provides insights into the potential and limitations of targeting microglial activation as a pharmacological strategy for the treatment of AD.


Assuntos
Doença de Alzheimer , Camundongos , Animais , Doença de Alzheimer/tratamento farmacológico , Microglia/metabolismo , Microglia/patologia , Camundongos Transgênicos , Peptídeos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Placa Amiloide
12.
Mol Neurodegener ; 18(1): 13, 2023 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-36810097

RESUMO

BACKGROUND: The protease BACE1 is a major drug target for Alzheimer's disease, but chronic BACE1 inhibition is associated with non-progressive cognitive worsening that may be caused by modulation of unknown physiological BACE1 substrates. METHODS: To identify in vivo-relevant BACE1 substrates, we applied pharmacoproteomics to non-human-primate cerebrospinal fluid (CSF) after acute treatment with BACE inhibitors. RESULTS: Besides SEZ6, the strongest, dose-dependent reduction was observed for the pro-inflammatory cytokine receptor gp130/IL6ST, which we establish as an in vivo BACE1 substrate. Gp130 was also reduced in human CSF from a clinical trial with a BACE inhibitor and in plasma of BACE1-deficient mice. Mechanistically, we demonstrate that BACE1 directly cleaves gp130, thereby attenuating membrane-bound gp130 and increasing soluble gp130 abundance and controlling gp130 function in neuronal IL-6 signaling and neuronal survival upon growth-factor withdrawal. CONCLUSION: BACE1 is a new modulator of gp130 function. The BACE1-cleaved, soluble gp130 may serve as a pharmacodynamic BACE1 activity marker to reduce the occurrence of side effects of chronic BACE1 inhibition in humans.


Assuntos
Doença de Alzheimer , Camundongos , Humanos , Animais , Doença de Alzheimer/tratamento farmacológico , Secretases da Proteína Precursora do Amiloide , Receptor gp130 de Citocina/uso terapêutico , Ácido Aspártico Endopeptidases , Interleucina-6 , Proteínas do Tecido Nervoso
13.
NPJ Parkinsons Dis ; 9(1): 74, 2023 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-37169750

RESUMO

Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are progressive neurodegenerative diseases characterized by the accumulation of misfolded α-synuclein in the form of Lewy pathology. While most cases are sporadic, there are rare genetic mutations that cause disease and more common variants that increase incidence of disease. The most prominent genetic mutations for PD and DLB are in the GBA1 and LRRK2 genes. GBA1 mutations are associated with decreased glucocerebrosidase activity and lysosomal accumulation of its lipid substrates, glucosylceramide and glucosylsphingosine. Previous studies have shown a link between this enzyme and lipids even in sporadic PD. However, it is unclear how the protein pathologies of disease are related to enzyme activity and glycosphingolipid levels. To address this gap in knowledge, we examined quantitative protein pathology, glucocerebrosidase activity and lipid substrates in parallel from 4 regions of 91 brains with no neurological disease, idiopathic, GBA1-linked, or LRRK2-linked PD and DLB. We find that several biomarkers are altered with respect to mutation and progression to dementia. We found mild association of glucocerebrosidase activity with disease, but a strong association of glucosylsphingosine with α-synuclein pathology, irrespective of genetic mutation. This association suggests that Lewy pathology precipitates changes in lipid levels related to progression to dementia.

14.
J Med Chem ; 66(21): 14912-14927, 2023 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-37861679

RESUMO

Genetic mutation of the leucine-rich repeat kinase 2 (LRRK2) protein has been associated with Parkinson's disease (PD), a disabling and progressive neurodegenerative disorder that is devoid of efficacious disease-modifying therapies. Herein, we describe the invention of an amidoisoquinoline (IQ)-derived LRRK2 inhibitor lead chemical series. Knowledge-, structure-, and property-based drug design in concert with rigorous application of in silico calculations and presynthesis predictions enabled the prioritization of molecules with favorable CNS "drug-like" physicochemical properties. This resulted in the discovery of compound 8, which was profiled extensively before human ether-a-go-go (hERG) ion channel inhibition halted its progression. Strategic reduction of lipophilicity and basicity resulted in attenuation of hERG ion channel inhibition while maintaining a favorable CNS efflux transporter profile. Further structure- and property-based optimizations resulted in the discovery of preclinical candidate MK-1468. This exquisitely selective LRRK2 inhibitor has a projected human dose of 48 mg BID and a preclinical safety profile that supported advancement toward GLP toxicology studies.


Assuntos
Doença de Parkinson , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Inibidores de Proteínas Quinases/química , Encéfalo/metabolismo , Mutação , Canais Iônicos/metabolismo
15.
Bioorg Med Chem Lett ; 22(7): 2444-9, 2012 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-22390835
16.
Sci Rep ; 12(1): 14985, 2022 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-36056033

RESUMO

Evidence suggests that ß-secretase (BACE1), which cleaves Amyloid Precursor Protein (APP) to form sAPPß and amyloid-ß, is elevated in Alzheimer's disease (AD) brains and biofluids and, thus, BACE1 is a therapeutic target for this devastating disease. The direct product of BACE1 cleavage of APP, sAPPß, serves as a surrogate marker of BACE1 activity in the central nervous system. This biomarker could be utilized to better understand normal APP processing, aberrant processing in the disease setting, and modulations to processing during therapeutic intervention. In this paper, we present a method for measuring the metabolism of sAPPß and another APP proteolytic product, sAPPα, in vivo in humans using stable isotope labeling kinetics, paired with immunoprecipitation and liquid chromatography/tandem mass spectrometry. The method presented herein is robust, reproducible, and precise, and allows for the study of these analytes by taking into account their full dynamic potential as opposed to the traditional methods of absolute concentration quantitation that only provide a static view of a dynamic system. A study of in vivo cerebrospinal fluid sAPPß and sAPPα kinetics using these methods could reveal novel insights into pathophysiological mechanisms of AD, such as increased BACE1 processing of APP.


Assuntos
Doença de Alzheimer , Precursor de Proteína beta-Amiloide , Doença de Alzheimer/líquido cefalorraquidiano , Secretases da Proteína Precursora do Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Ácido Aspártico Endopeptidases/metabolismo , Biomarcadores , Humanos , Marcação por Isótopo , Isoformas de Proteínas , Espectrometria de Massas em Tandem
17.
J Med Chem ; 65(1): 838-856, 2022 01 13.
Artigo em Inglês | MEDLINE | ID: mdl-34967623

RESUMO

The leucine-rich repeat kinase 2 (LRRK2) protein has been genetically and functionally linked to Parkinson's disease (PD), a disabling and progressive neurodegenerative disorder whose current therapies are limited in scope and efficacy. In this report, we describe a rigorous hit-to-lead optimization campaign supported by structural enablement, which culminated in the discovery of brain-penetrant, candidate-quality molecules as represented by compounds 22 and 24. These compounds exhibit remarkable selectivity against the kinome and offer good oral bioavailability and low projected human doses. Furthermore, they showcase the implementation of stereochemical design elements that serve to enable a potency- and selectivity-enhancing increase in polarity and hydrogen bond donor (HBD) count while maintaining a central nervous system-friendly profile typified by low levels of transporter-mediated efflux and encouraging brain penetration in preclinical models.


Assuntos
Antiparkinsonianos/síntese química , Antiparkinsonianos/farmacologia , Encéfalo/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/antagonistas & inibidores , Quinazolinas/síntese química , Quinazolinas/farmacologia , Antiparkinsonianos/farmacocinética , Disponibilidade Biológica , Desenho de Fármacos , Humanos , Modelos Moleculares , Inibidores de Proteínas Quinases/farmacologia , Quinazolinas/farmacocinética , Relação Estrutura-Atividade
18.
J Med Chem ; 65(24): 16801-16817, 2022 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-36475697

RESUMO

Inhibition of leucine-rich repeat kinase 2 (LRRK2) kinase activity represents a genetically supported, chemically tractable, and potentially disease-modifying mechanism to treat Parkinson's disease. Herein, we describe the optimization of a novel series of potent, selective, central nervous system (CNS)-penetrant 1-heteroaryl-1H-indazole type I (ATP competitive) LRRK2 inhibitors. Type I ATP-competitive kinase physicochemical properties were integrated with CNS drug-like properties through a combination of structure-based drug design and parallel medicinal chemistry enabled by sp3-sp2 cross-coupling technologies. This resulted in the discovery of a unique sp3-rich spirocarbonitrile motif that imparted extraordinary potency, pharmacokinetics, and favorable CNS drug-like properties. The lead compound, 25, demonstrated exceptional on-target potency in human peripheral blood mononuclear cells, excellent off-target kinase selectivity, and good brain exposure in rat, culminating in a low projected human dose and a pre-clinical safety profile that warranted advancement toward pre-clinical candidate enabling studies.


Assuntos
Doença de Parkinson , Ratos , Humanos , Animais , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina , Doença de Parkinson/tratamento farmacológico , Indazóis/farmacologia , Indazóis/uso terapêutico , Leucócitos Mononucleares/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Inibidores de Proteínas Quinases/química , Encéfalo/metabolismo , Trifosfato de Adenosina
19.
Neurotherapeutics ; 17(3): 973-988, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32236823

RESUMO

While the peripheral nervous system is able to repair itself following injury and disease, recovery is often slow and incomplete, with no available treatments to enhance the effectiveness of regeneration. Using knock-out and transgenic overexpressor mice, we previously reported that BACE1, an aspartyl protease, as reported by Hemming et al. (PLoS One 4:12, 2009), negatively regulates peripheral nerve regeneration. Here, we investigated whether pharmacological inhibition of BACE may enhance peripheral nerve repair following traumatic nerve injury or neurodegenerative disease. BACE inhibitor-treated mice had increased numbers of regenerating axons and enhanced functional recovery after a sciatic nerve crush while inhibition increased axonal sprouting following a partial nerve injury. In the SOD1G93A ALS mouse model, BACE inhibition increased axonal regeneration with improved muscle re-innervation. CHL1, a BACE1 substrate, was elevated in treated mice and may mediate enhanced regeneration. Our data demonstrates that pharmacological BACE inhibition accelerates peripheral axon regeneration after varied nerve injuries and could be used as a potential therapy.


Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Secretases da Proteína Precursora do Amiloide/metabolismo , Esclerose Lateral Amiotrófica/enzimologia , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Ácido Aspártico Endopeptidases/metabolismo , Axônios/fisiologia , Regeneração Nervosa/fisiologia , Traumatismos dos Nervos Periféricos/enzimologia , Esclerose Lateral Amiotrófica/tratamento farmacológico , Esclerose Lateral Amiotrófica/genética , Animais , Axônios/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Feminino , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Regeneração Nervosa/efeitos dos fármacos , Traumatismos dos Nervos Periféricos/tratamento farmacológico , Traumatismos dos Nervos Periféricos/genética , Superóxido Dismutase/genética
20.
Sci Transl Med ; 12(570)2020 11 18.
Artigo em Inglês | MEDLINE | ID: mdl-33208500

RESUMO

Axonal dystrophy, indicative of perturbed axonal transport, occurs early during Alzheimer's disease (AD) pathogenesis. Little is known about the mechanisms underlying this initial sign of the pathology. This study proves that Golgi-localized γ-ear-containing ARF binding protein 3 (GGA3) loss of function, due to Gga3 genetic deletion or a GGA3 rare variant that cosegregates with late-onset AD, disrupts the axonal trafficking of the ß-site APP-cleaving enzyme 1 (BACE1) resulting in its accumulation in axonal swellings in cultured neurons and in vivo. We show that BACE pharmacological inhibition ameliorates BACE1 axonal trafficking and diminishes axonal dystrophies in Gga3 null neurons in vitro and in vivo. These data indicate that axonal accumulation of BACE1 engendered by GGA3 loss of function results in local toxicity leading to axonopathy. Gga3 deletion exacerbates axonal dystrophies in a mouse model of AD before ß-amyloid (Aß) deposition. Our study strongly supports a role for GGA3 in AD pathogenesis, where GGA3 loss of function triggers BACE1 axonal accumulation independently of extracellular Aß, and initiates a cascade of events leading to the axonal damage distinctive of the early stage of AD.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Doença de Alzheimer , Secretases da Proteína Precursora do Amiloide/genética , Secretases da Proteína Precursora do Amiloide/metabolismo , Ácido Aspártico Endopeptidases/genética , Ácido Aspártico Endopeptidases/metabolismo , Doença de Alzheimer/genética , Peptídeos beta-Amiloides , Precursor de Proteína beta-Amiloide/genética , Animais , Axônios , Camundongos
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