RESUMEN
BACKGROUND: The elevation of endocannabinoid levels through inhibiting their degradation afforded neuroprotection in CaMKIIα-TDP-43 mice, a conditional transgenic model of frontotemporal dementia. However, which cannabinoid receptors are mediating these benefits is still pending to be elucidated. METHODS: We have investigated the involvement of the CB1 and the CB2 receptor using chronic treatments with selective ligands in CaMKIIα-TDP-43 mice, analysis of their cognitive deterioration with the Novel Object Recognition test, and immunostaining for neuronal and glial markers in two areas of interest in frontotemporal dementia. RESULTS: Our results confirmed the therapeutic value of activating either the CB1 or the CB2 receptor, with improvements in the animal performance in the Novel Object Recognition test, preservation of pyramidal neurons, in particular in the medial prefrontal cortex, and attenuation of glial reactivity, in particular in the hippocampus. In addition, the activation of both CB1 and CB2 receptors reduced the elevated levels of TDP-43 in the medial prefrontal cortex of CaMKIIα-TDP-43 mice, an effect exerted by mechanisms that are currently under investigation. CONCLUSIONS: These data reinforce the notion that the activation of CB1 and CB2 receptors may represent a promising therapy against TDP-43-induced neuropathology in frontotemporal dementia. Future studies will have to confirm these benefits, in particular with one of the selective CB2 agonists used here, which has been thoroughly characterized for clinical development.
Asunto(s)
Cannabinoides , Modelos Animales de Enfermedad , Demencia Frontotemporal , Ratones Transgénicos , Fármacos Neuroprotectores , Receptor Cannabinoide CB1 , Receptor Cannabinoide CB2 , Animales , Receptor Cannabinoide CB2/agonistas , Receptor Cannabinoide CB2/metabolismo , Masculino , Fármacos Neuroprotectores/farmacología , Receptor Cannabinoide CB1/metabolismo , Receptor Cannabinoide CB1/agonistas , Demencia Frontotemporal/tratamiento farmacológico , Demencia Frontotemporal/metabolismo , Demencia Frontotemporal/patología , Ratones , Cannabinoides/farmacología , Corteza Prefrontal/efectos de los fármacos , Corteza Prefrontal/metabolismo , Corteza Prefrontal/patología , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Proteínas de Unión al ADN/metabolismo , Ratones Endogámicos C57BL , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Hipocampo/patologíaRESUMEN
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease without any effective treatment. Protein TDP-43 is a pathological hallmark of ALS in both sporadic and familiar patients. Post-translational modifications of TDP-43 promote its aggregation in the cytoplasm. Tau-Tubulin kinase (TTBK1) phosphorylates TDP-43 in cellular and animal models; thus, TTBK1 inhibitors emerge as a promising therapeutic strategy for ALS. The design, synthesis, biological evaluation, kinase-ligand complex structure determination, and molecular modeling studies confirmed novel pyrrolopyrimidine derivatives as valuable inhibitors for further development. Moreover, compound 29 revealed good brain penetration in vivo and was able to reduce TDP-43 phosphorylation not only in cell cultures but also in the spinal cord of transgenic TDP-43 mice. A shift to M2 anti-inflammatory microglia was also demonstrated in vivo. Both these activities led to motor neuron preservation in mice, proposing pyrrolopyrimidine 29 as a valuable lead compound for future ALS therapy.
Asunto(s)
Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Encéfalo/efectos de los fármacos , Proteínas de Unión al ADN/metabolismo , Inflamación/tratamiento farmacológico , Macrófagos/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Animales , Encéfalo/metabolismo , Estudios de Casos y Controles , Humanos , Inflamación/metabolismo , Inflamación/patología , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Transgénicos , Fosforilación , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacocinética , Ratas , Ratas Wistar , Médula Espinal/efectos de los fármacos , Médula Espinal/metabolismo , Distribución TisularRESUMEN
Cannabinoids act as pleiotropic compounds exerting, among others, a broad-spectrum of neuroprotective effects. These effects have been investigated in the last years in different preclinical models of neurodegeneration, with the cannabinoid type-1 (CB1) and type-2 (CB2) receptors concentrating an important part of this research. However, the issue has also been extended to additional targets that are also active for cannabinoids, such as the orphan G-protein receptor 55 (GPR55). In the present study, we investigated the neuroprotective potential of VCE-006.1, a chromenopyrazole derivative with biased orthosteric and positive allosteric modulator activity at GPR55, in murine models of two neurodegenerative diseases. First, we proved that VCE-006.1 alone could induce ERK1/2 activation and calcium mobilization, as well as increase cAMP response but only in the presence of lysophosphatidyl inositol. Next, we investigated this compound administered chronically in two neurotoxin-based models of Parkinson's disease (PD), as well as in some cell-based models. VCE-006.1 was active in reversing the motor defects caused by 6-hydroxydopamine (6-OHDA) in the pole and the cylinder rearing tests, as well as the losses in tyrosine hydroxylase-containing neurons and the elevated glial reactivity detected in the substantia nigra. Similar cytoprotective effects were found in vitro in SH-SY5Y cells exposed to 6-OHDA. We also investigated VCE-006.1 in LPS-lesioned mice with similar beneficial effects, except against glial reactivity and associated inflammatory events, which remained unaltered, a fact confirmed in BV2 cells treated with LPS and VCE-006.1. We also analyzed GPR55 in these in vivo models with no changes in its gene expression, although GPR55 was down-regulated in BV2 cells treated with LPS, which may explain the lack of efficacy of VCE-006.1 in such an assay. Furthermore, we investigated VCE-006.1 in two genetic models of amyotrophic lateral sclerosis (ALS), mutant SOD1, or TDP-43 transgenic mice. Neither the neurological decline nor the deteriorated rotarod performance were prevented with this compound, and the same happened with the elevated microglial and astroglial reactivities, albeit modest spinal motor neuron preservation was achieved in both models. We also analyzed GPR55 in these in vivo models and found no changes in both TDP-43 transgenic and mSOD1 mice. Therefore, our findings support the view that targeting the GPR55 may afford neuroprotection in experimental PD, but not in ALS, thus stressing the specificities for the development of cannabinoid-based therapies in the different neurodegenerative disorders.
Asunto(s)
Esclerosis Amiotrófica Lateral , Fármacos Neuroprotectores/farmacología , Enfermedad de Parkinson , Receptores de Cannabinoides/metabolismo , Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Modelos Animales de Enfermedad , Humanos , Ligandos , Masculino , Ratones , Ratones Transgénicos , Neuroglía/metabolismo , Fármacos Neuroprotectores/química , Enfermedad de Parkinson/tratamiento farmacológico , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/metabolismo , Sustancia Negra/metabolismo , Células U937RESUMEN
Amyotrophic Lateral Sclerosis (ALS) is the most common degenerative motor neuron disease in adults. About 97% of ALS patients present TDP-43 aggregates with post-translational modifications, such as hyperphosphorylation, in the cytoplasm of affected cells. GSK-3ß is one of the protein kinases involved in TDP-43 phosphorylation. Up-regulation of its expression and activity is reported on spinal cord and cortex tissues of ALS patients. Here, we propose the repurposing of Tideglusib, an in-house non-ATP competitive GSK-3ß inhibitor that is currently in clinical trials for autism and myotonic dystrophy, as a promising therapeutic strategy for ALS. With this aim we have evaluated the efficacy of Tideglusib in different experimental ALS models both in vitro and in vivo. Moreover, we observed that GSK-3ß activity is increased in lymphoblasts from sporadic ALS patients, with a simultaneous increase in TDP-43 phosphorylation and cytosolic TDP-43 accumulation. Treatment with Tideglusib decreased not only phospho-TDP-43 levels but also recovered its nuclear localization in ALS lymphoblasts and in a human TDP-43 neuroblastoma model. Additionally, we found that chronic oral treatment with Tideglusib is able to reduce the increased TDP-43 phosphorylation in the spinal cord of Prp-hTDP-43A315T mouse model. Therefore, we consider Tideglusib as a promising drug candidate for ALS, being proposed to start a clinical trial phase II by the end of the year.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Tiadiazoles/farmacología , Anciano , Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Proteínas de Unión al ADN/fisiología , Modelos Animales de Enfermedad , Reposicionamiento de Medicamentos , Glucógeno Sintasa Quinasa 3 beta/fisiología , Humanos , Masculino , Ratones , Ratones Transgénicos , Persona de Mediana Edad , Neuronas Motoras/metabolismo , Preparaciones Farmacéuticas/metabolismo , Fosforilación , Proteínas Quinasas/metabolismo , Médula Espinal/metabolismoRESUMEN
Cannabinoids form a singular group of plant-derived compounds, endogenous lipids and synthetic derivatives with multiple therapeutic effects exerted by targeting different elements of the endocannabinoid system. One of their therapeutic applications is the preservation of neuronal integrity exerted by attenuating the multiple neurotoxic events that kill neurons in neurodegenerative disorders. In this review, we will address the potential of cannabinoids as neuroprotective agents in amyotrophic lateral sclerosis (ALS), a devastating neurodegenerative disorder characterized by muscle denervation, atrophy and paralysis, and progressive deterioration in upper and/or lower motor neurons. The emphasis will be paid on the cannabinoid type 2 (CB2 ) receptor, whose activation limits glial reactivity, but the potential of additional endocannabinoid-related targets will be also addressed. The evidence accumulated so far at the preclinical level supports the need to soon move towards the patients and initiate clinical trials to confirm the potential of cannabinoid-based medicines as disease modifiers in ALS. LINKED ARTICLES: This article is part of a themed issue on Neurochemistry in Japan. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.6/issuetoc.
Asunto(s)
Esclerosis Amiotrófica Lateral , Cannabinoides , Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Cannabinoides/uso terapéutico , Progresión de la Enfermedad , Endocannabinoides , Humanos , Neuronas Motoras , Receptor Cannabinoide CB1 , Receptor Cannabinoide CB2RESUMEN
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with no known cure. Aggregates of the nuclear protein TDP-43 have been recognized as a hallmark of proteinopathy in both familial and sporadic cases of ALS. Post-translational modifications of this protein, include hyperphosphorylation, cause disruption of TDP-43 homeostasis and as a consequence, promotion of its neurotoxicity. Among the kinases involved in these changes, cell division cycle kinase 7 (CDC7) plays an important role by directly phosphorylating TDP-43. In the present manuscript the discovery, synthesis, and optimization of a new family of selective and ATP-competitive CDC7 inhibitors based on 6-mercaptopurine scaffold are described. Moreover, we demonstrate the ability of these inhibitors to reduce TDP-43 phosphorylation in both cell cultures and transgenic animal models such as C. elegans and Prp-hTDP43 (A315T) mice. Altogether, the compounds described here may be useful as versatile tools to explore the role of CDC7 in TDP-43 phosphorylation and also as new drug candidates for the future development of ALS therapies.
Asunto(s)
Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Proteínas de Ciclo Celular/antagonistas & inhibidores , Proteínas de Unión al ADN/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Conducta Animal/efectos de los fármacos , Proteínas de Ciclo Celular/metabolismo , Proteínas de Unión al ADN/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Ratones , Ratones Transgénicos , Simulación del Acoplamiento Molecular , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Recombinantes/metabolismo , Relación Estructura-Actividad , Células Tumorales CultivadasRESUMEN
The quinone derivative of the non-psychotropic cannabinoid cannabigerol (CBG), so-called VCE-003.2, has been recently investigated for its neuroprotective properties in inflammatory models of Parkinson's disease (PD) in mice. Such potential derives from its activity at the peroxisome proliferator-activated receptor-γ (PPAR-γ). In the present study, we investigated the neuroprotective properties of VCE-003.2 against the parkinsonian neurotoxin 6-hydroxydopamine (6-OHDA), in comparison with two new CBG-related derivatives, the cannabigerolic acid quinone (CBGA-Q) and its sodium salt CBGA-Q-Salt, which, similarly to VCE-003.2, were found to be active at the PPAR-γ receptor, but not at the cannabinoid CB1 and CB2 receptors. First, we investigated their cytoprotective properties in vitro by analyzing cell survival in cultured SH-SY5Y cells exposed to 6-OHDA. We found an important cytoprotective effect of VCE-003.2 at a concentration of 20 µM, which was not reversed by the blockade of PPAR-γ receptors with GW9662, supporting its activity at an alternative site (non-sensitive to classic antagonists) in this receptor. We also found CBGA-Q and CBGA-Q-Salt being cytoprotective in this cell assay, but their effects were completely eliminated by GW9662, thus indicating that they are active at the canonical site in the PPAR-γ receptor. Then, we moved to in vivo testing using mice unilaterally lesioned with 6-OHDA. Our data confirmed that VCE-003.2 administered orally (20 mg/kg) preserved tyrosine hydroxylase (TH)-positive nigral neurons against 6-OHDA-induced damage, whereas it completely attenuated the astroglial (GFAP) and microglial (CD68) reactivity found in the substantia nigra of lesioned mice. Such neuroprotective effects caused an important recovery in the motor deficiencies displayed by 6-OHDA-lesioned mice in the pole test and the cylinder rearing test. We also investigated CBGA-Q, given orally (20 mg/kg) or intraperitoneally (10 mg/kg, i.p.), having similar benefits compared to VCE-003.2 against the loss of TH-positive nigral neurons, glial reactivity and motor defects caused by 6-OHDA. Lastly, the sodium salt of CBGA-Q, given orally (40 mg/kg) to 6-OHDA-lesioned mice, also showed benefits at behavioral and histopathological levels, but to a lower extent compared to the other two compounds. In contrast, when given i.p., CBGA-Q-Salt (10 mg/kg) was poorly active. We also analyzed the concentrations of dopamine and its metabolite DOPAC in the striatum of 6-OHDA-lesioned mice after the treatment with the different compounds, but recovery in the contents of both dopamine and DOPAC was only found after the treatment with VCE-003.2. In summary, our data confirmed the neuroprotective potential of VCE-003.2 in 6-OHDA-lesioned mice, which adds to its previous activity found in an inflammatory model of PD (LPS-lesioned mice). Additional phytocannabinoid derivatives, CBGA-Q and CBGA-Q-Salt, also afforded neuroprotection in 6-OHDA-lesioned mice, but their effects were lower compared to VCE-003.2, in particular in the case of CBGA-Q-Salt. In vitro studies confirmed the relevance of PPAR-γ receptors for these effects.
Asunto(s)
Antiparkinsonianos/uso terapéutico , Cannabinoides/química , Fármacos Neuroprotectores/uso terapéutico , Enfermedad de Parkinson/tratamiento farmacológico , Quinonas/química , Animales , Antiparkinsonianos/síntesis química , Antiparkinsonianos/farmacología , Línea Celular Tumoral , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Fármacos Neuroprotectores/síntesis química , Fármacos Neuroprotectores/farmacología , Oxidopamina/toxicidad , PPAR gamma/antagonistas & inhibidores , PPAR gamma/metabolismo , Enfermedad de Parkinson/etiología , Sustancia Negra/citología , Sustancia Negra/efectos de los fármacos , Sustancia Negra/metabolismoRESUMEN
BACKGROUND AND PURPOSE: Δ9 -Tetrahydrocannabinolic acid (Δ9 -THCA-A), the precursor of Δ9 -THC, is a non-psychotropic phytocannabinoid that shows PPARγ agonist activity. Here, we investigated the ability of Δ9 -THCA-A to modulate the classic cannabinoid CB1 and CB2 receptors and evaluated its anti-arthritis activity in vitro and in vivo. EXPERIMENTAL APPROACH: Cannabinoid receptors binding and intrinsic activity, as well as their downstream signalling, were analysed in vitro and in silico. The anti-arthritis properties of Δ9 -THCA-A were studied in human chondrocytes and in the murine model of collagen-induced arthritis (CIA). Plasma disease biomarkers were identified by LC-MS/MS based on proteomic and elisa assays. KEY RESULTS: Functional and docking analyses showed that Δ9 -THCA-A can act as an orthosteric CB1 receptor agonist and also as a positive allosteric modulator in the presence of CP-55,940. Also, Δ9 -THCA-A seemed to be an inverse agonist for CB2 receptors. In vivo, Δ9 -THCA-A reduced arthritis in CIA mice, preventing the infiltration of inflammatory cells, synovium hyperplasia, and cartilage damage. Furthermore, Δ9 -THCA-A inhibited expression of inflammatory and catabolic genes on knee joints. The anti-arthritic effect of Δ9 -THCA-A was blocked by either SR141716 or T0070907. Analysis of plasma biomarkers, and determination of cytokines and anti-collagen antibodies confirmed that Δ9 -THCA-A mediated its activity mainly through PPARγ and CB1 receptor pathways. CONCLUSION AND IMPLICATIONS: Δ9 -THCA-A modulates CB1 receptors through the orthosteric and allosteric binding sites. In addition, Δ9 -THCA-A exerts anti-arthritis activity through CB1 receptors and PPARγ pathways, highlighting its potential for the treatment of chronic inflammatory diseases such as rheumatoid arthritis.