RESUMO
INTRODUCTION: Prion diseases are a group of rare and lethal, rapidly progressive neurodegenerative diseases arising due to conversion of the physiological cellular prion protein into its pathological counterparts, denoted as 'prions.' These agents are resistant to inactivation by standard decontamination procedures and can be transmitted between individuals, consequently driving the irreversible brain damage typical of the diseases. AREAS COVERED: Since its infancy, prion research has mainly depended on animal models for untangling the pathogenesis of the disease as well as for the drug development studies. With the advent of prion-infected cell lines, relevant animal models have been complemented by a variety of cell-based models presenting a much faster, ethically acceptable alternative. EXPERT OPINION: To date, there are still either no effective prophylactic regimens or therapies for human prion diseases. Therefore, there is an urgent need for more relevant cellular models that best approximate in vivo models. Each cellular model presented and discussed in detail in this review has its own benefits and limitations. Once embarking in a drug screening campaign for the identification of molecules that could interfere with prion conversion and replication, one should carefully consider the ideal cellular model.
Assuntos
Doenças Priônicas , Príons , Animais , Desenvolvimento de Medicamentos , Avaliação Pré-Clínica de Medicamentos , Humanos , Doenças Priônicas/tratamento farmacológico , Proteínas Priônicas , Príons/metabolismoRESUMO
Prion diseases or transmissible spongiform encephalopathies (TSEs) are a group of rare neurodegenerative disorders. TSEs are characterized by the accumulation of prions (PrPSc) that represent pathological isoforms of the physiological cellular prion protein PrPC. Although the conversion of PrPC to PrPSc is still not completely understood, blocking this process may lead to develop new therapies. Here, we have generated a pharmacophore model, based on anti-prion molecules reported in literature to be effective in phenotypic assay. The model was used to conduct a virtual screen of commercial compound databases that selected a small library of ten compounds. These molecules were then screened in mouse neuroblastoma cell line chronically infected with prions (ScN2a) after excluding neurotoxicity. 1 has been identified as the therapeutic hit on the basis of the following evidence: chronic treatments of ScN2a cells using 1 eliminate PrPSc loaded in both Western blotting analysis and Real-Time Quaking-Induced Conversion (RT-QuIC) assay. We also proposed the mechanism of action of 1 by which it has the ability to bind PrPC and consequentially blocks prion conversion. Herein we describe the results of these efforts.
Assuntos
Fenotiazinas/farmacologia , Proteínas Priônicas/antagonistas & inibidores , Animais , Linhagem Celular , Avaliação Pré-Clínica de Medicamentos , Camundongos , Modelos Moleculares , Estrutura Molecular , Fenotiazinas/química , Proteínas Priônicas/isolamento & purificação , Proteínas Priônicas/metabolismo , Relação Quantitativa Estrutura-AtividadeRESUMO
α-Synuclein (AS) is an intrinsically disordered protein highly expressed in dopaminergic neurons. Its amyloid aggregates are the major component of Lewy bodies, a hallmark of Parkinson's disease (PD). AS is particularly exposed to oxidation of its methionine residues, both in vivo and in vitro Oxidative stress has been implicated in PD and oxidized α-synuclein has been shown to assemble into soluble, toxic oligomers, rather than amyloid fibrils. However, the structural effects of methionine oxidation are still poorly understood. In this work, oxidized AS was obtained by prolonged incubations with dopamine (DA) or epigallocatechin-3-gallate (EGCG), two inhibitors of AS aggregation, indicating that EGCG promotes the same final oxidation product as DA. The conformational transitions of the oxidized and non-oxidized protein were monitored by complementary biophysical techniques, including MS, ion mobility (IM), CD, and FTIR spectroscopy assays. Although the two variants displayed very similar structures under conditions that stabilize highly disordered or highly ordered states, differences emerged in the intermediate points of transitions induced by organic solvents, such as trifluoroethanol (TFE) and methanol (MeOH), indicating a lower propensity of the oxidized protein for forming either α- or ß-type secondary structures. Furthermore, oxidized AS displayed restricted secondary-structure transitions in response to dehydration and slightly amplified tertiary-structure transitions induced by ligand binding. This difference in susceptibility to induced folding could explain the loss of fibrillation potential observed for oxidized AS. Finally, site-specific oxidation kinetics point out a minor delay in Met-127 modification, likely due to the effects of AS intrinsic structure.
Assuntos
Catequina/análogos & derivados , Metionina/química , Agregados Proteicos , Dobramento de Proteína , alfa-Sinucleína/química , Catequina/química , Humanos , Corpos de Lewy/metabolismo , Corpos de Lewy/patologia , Metionina/metabolismo , Oxirredução , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , alfa-Sinucleína/metabolismoRESUMO
Several findings propose the altered tau protein network as an important target for Alzheimer's disease (AD). Particularly, two points of pharmacological intervention can be envisaged: inhibition of phosphorylating tau kinase GSK-3ß and tau aggregation process. On the basis of this consideration and on our interest in multitarget paradigms in AD, we report on the discovery of 2,4-thiazolidinedione derivatives endowed with such a profile. 28 and 30 displayed micromolar IC50 values toward GSK-3ß, together with the capacity of inhibiting AcPHF6 aggregation of 60% and 80% at 10 µM, respectively. In addition, they showed PAMPA-BBB permeability, together with a suitable cellular safety profile. 30 also displayed inhibition of both K18 and full-length tau aggregations. Finally, both compounds were able to improve cell viability in an okadaic acid-induced neurodegeneration cell model. To the best of our knowledge, 28 and 30 are the first balanced, nontoxic, dual-acting compounds hitting tau cascade at two different hubs.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Proteínas tau/metabolismo , Animais , Barreira Hematoencefálica/efeitos dos fármacos , Fármacos do Sistema Nervoso Central/efeitos adversos , Fármacos do Sistema Nervoso Central/química , Fármacos do Sistema Nervoso Central/farmacologia , Dicroísmo Circular , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos/métodos , Transferência Ressonante de Energia de Fluorescência , Quinase 3 da Glicogênio Sintase/genética , Quinase 3 da Glicogênio Sintase/metabolismo , Células Hep G2 , Humanos , Microscopia de Força Atômica , Terapia de Alvo Molecular/métodos , Ácido Okadáico/toxicidade , Fosforilação/efeitos dos fármacos , Ratos , Relação Estrutura-Atividade , Suínos , Tiazolidinedionas/química , Proteínas tau/antagonistas & inibidoresRESUMO
Prion diseases are fatal neurodegenerative and infectious disorders for which effective pharmacological tools are not yet available. This unmet challenge and the recently proposed interplay between prion diseases and Alzheimer's have led to a more urgent demand for new antiprion agents. Herein, we report the identification of a novel bifunctional diketopiperazine (DKP) derivative 1 d, which exhibits activity in the low micromolar range against prion replication in ScGT1 cells, while showing low cytotoxicity. Supported by properly addressed molecular modeling studies, we hypothesized that a planar conformation is the major determinant for activity in this class of compounds. Moreover, studies aimed at assessing the mechanism-of-action at the molecular level showed that 1 d might interact directly with recombinant prion protein (recPrP) to prevent its conversion to the pathogenic misfolded prion protein (PrP(Sc))-like form. This investigation suggests that DKP based antiprion compounds can serve as a promising lead scaffold in developing new drugs to combat prion diseases.