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1.
Nat Methods ; 21(7): 1288-1297, 2024 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-38877316

RESUMEN

Precision pharmacology aims to manipulate specific cellular interactions within complex tissues. In this pursuit, we introduce DART.2 (drug acutely restricted by tethering), a second-generation cell-specific pharmacology technology. The core advance is optimized cellular specificity-up to 3,000-fold in 15 min-enabling the targeted delivery of even epileptogenic drugs without off-target effects. Additionally, we introduce brain-wide dosing methods as an alternative to local cannulation and tracer reagents for brain-wide dose quantification. We describe four pharmaceuticals-two that antagonize excitatory and inhibitory postsynaptic receptors, and two that allosterically potentiate these receptors. Their versatility is showcased across multiple mouse-brain regions, including cerebellum, striatum, visual cortex and retina. Finally, in the ventral tegmental area, we find that blocking inhibitory inputs to dopamine neurons accelerates locomotion, contrasting with previous optogenetic and pharmacological findings. Beyond enabling the bidirectional perturbation of chemical synapses, these reagents offer intersectional precision-between genetically defined postsynaptic cells and neurotransmitter-defined presynaptic partners.


Asunto(s)
Sinapsis , Animales , Ratones , Sinapsis/efectos de los fármacos , Sinapsis/fisiología , Sinapsis/metabolismo , Encéfalo/metabolismo , Masculino , Ratones Endogámicos C57BL , Humanos , Femenino , Neuronas Dopaminérgicas/efectos de los fármacos , Neuronas Dopaminérgicas/metabolismo
2.
Tetrahedron Lett ; 1172023 Mar 06.
Artículo en Inglés | MEDLINE | ID: mdl-36936473

RESUMEN

KLS-13019 is a structural analogue of cannabidiol, that shows improved bioavailability and potency in both preventing and reversing paclitaxel-induced neurotoxicity in vitro and in vivo. KLS-13019 was selected as a development candidate and attention was turned to development of a scalable synthesis. The original synthesis of KLS-13019 was not efficient, regioselective, or high yielding. Two new syntheses are reported that make use of the palladium catalyzed cross couplings to a chemically advanced intermediate 5, dramatically shortening (3-4 steps) and improving the overall yield. In addition, a convenient one pot Boc-cleavage and acetylation procedure is described to avoid impurities generated from a step-wise process.

3.
Bioorg Med Chem Lett ; 30(14): 127243, 2020 07 15.
Artículo en Inglés | MEDLINE | ID: mdl-32527545

RESUMEN

Monoacylglycerol lipase (MAGL) is the enzyme that is primarily responsible for hydrolyzing the endocannabinoid 2-arachidononylglycerol (2-AG) to arachidonic acid (AA). It has emerged in recent years as a potential drug target for a number of diseases. Herein, we report the discovery of compound 6g from a series of azetidine-piperazine di-amide compounds as a potent, selective, and reversible inhibitor of MAGL. Oral administration of compound 6g increased 2-AG levels in rat brain and produced full efficacy in the rat complete Freund's adjuvant (CFA) model of inflammatory pain.


Asunto(s)
Amidas/farmacología , Azetidinas/farmacología , Descubrimiento de Drogas , Inhibidores Enzimáticos/farmacología , Monoacilglicerol Lipasas/antagonistas & inhibidores , Piperazinas/farmacología , Amidas/química , Azetidinas/química , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Humanos , Modelos Moleculares , Estructura Molecular , Monoacilglicerol Lipasas/metabolismo , Piperazinas/química , Relación Estructura-Actividad
4.
Neurobiol Dis ; 119: 13-25, 2018 11.
Artículo en Inglés | MEDLINE | ID: mdl-30031156

RESUMEN

Cytosolic PSD-95 interactor (cypin), the primary guanine deaminase in the brain, plays key roles in shaping neuronal circuits and regulating neuronal survival. Despite this pervasive role in neuronal function, the ability for cypin activity to affect recovery from acute brain injury is unknown. A key barrier in identifying the role of cypin in neurological recovery is the absence of pharmacological tools to manipulate cypin activity in vivo. Here, we use a small molecule screen to identify two activators and one inhibitor of cypin's guanine deaminase activity. The primary screen identified compounds that change the initial rate of guanine deamination using a colorimetric assay, and secondary screens included the ability of the compounds to protect neurons from NMDA-induced injury and NMDA-induced decreases in frequency and amplitude of miniature excitatory postsynaptic currents. Hippocampal neurons pretreated with activators preserved electrophysiological function and survival after NMDA-induced injury in vitro, while pretreatment with the inhibitor did not. The effects of the activators were abolished when cypin was knocked down. Administering either cypin activator directly into the brain one hour after traumatic brain injury significantly reduced fear conditioning deficits 5 days after injury, while delivering the cypin inhibitor did not improve outcome after TBI. Together, these data demonstrate that cypin activation is a novel approach for improving outcome after TBI and may provide a new pathway for reducing the deficits associated with TBI in patients.


Asunto(s)
Lesiones Traumáticas del Encéfalo/metabolismo , Lesiones Traumáticas del Encéfalo/prevención & control , Guanina Desaminasa/metabolismo , Animales , Lesiones Traumáticas del Encéfalo/fisiopatología , Células COS , Células Cultivadas , Chlorocebus aethiops , Dimetilsulfóxido/farmacología , Miedo/efectos de los fármacos , Miedo/fisiología , Guanina Desaminasa/antagonistas & inhibidores , Compuestos Heterocíclicos con 3 Anillos/farmacología , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Hipocampo/fisiopatología , Masculino , Ratones , Ratones Endogámicos C57BL , N-Metilaspartato/farmacología , Técnicas de Cultivo de Órganos , Ratas
5.
Bioorg Med Chem Lett ; 24(4): 1116-21, 2014 Feb 15.
Artículo en Inglés | MEDLINE | ID: mdl-24461299

RESUMEN

We designed a series of anilino-indoylmaleimides based on structural elements from literature JAK3 inhibitors 3 and 4, and our lead 5. These new compounds were tested as inhibitors of JAKs 1, 2 and 3 and TYK2 for therapeutic intervention in rheumatoid arthritis (RA). Our requirements, based on current scientific rationale for optimum efficacy against RA with reduced side effects, was for potent, mixed JAK1 and 3 inhibition, and selectivity over JAK2. Our efforts yielded a potent JAK3 inhibitor 11d and its eutomer 11e. These compounds were highly selective for inhibition of JAK3 over JAK2 and TYK. The compounds displayed only modest JAK1 inhibition.


Asunto(s)
Compuestos de Anilina/farmacología , Janus Quinasa 3/antagonistas & inhibidores , Maleimidas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Compuestos de Anilina/síntesis química , Compuestos de Anilina/química , Relación Dosis-Respuesta a Droga , Humanos , Janus Quinasa 3/metabolismo , Maleimidas/síntesis química , Maleimidas/química , Modelos Moleculares , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Relación Estructura-Actividad
6.
Tetrahedron Lett ; 55(30): 4193-4195, 2014 Jul 23.
Artículo en Inglés | MEDLINE | ID: mdl-25018567

RESUMEN

We have found that α-amino acid amide derivatives of 2-aminobenzothiazoles undergo a time-dependent, thermal rearrangement in which the amine group attacks the 2-position carbon of the thiazole ring to form a 5,5-spiro ring system. This is followed by sulfur leaving and air oxidation to the corresponding symmetrical disulfide. The isolated yields of such products are quite high (>70%) if there is conformational bias to further promote the intramolecular reaction such as for the 2-aminobenzothiazole amides derived from proline or 4-aminopiperidine-4-carboxylic acid. This rearrangement has not been described previously for α-amino acid amide derivatives of 2-aminobenzothiazoles. However, a related reaction involving 2-semicarbazido benzothiazoles has been recently reported.

7.
Res Sq ; 2024 Feb 28.
Artículo en Inglés | MEDLINE | ID: mdl-38464007

RESUMEN

KLS-13019 was reported previously to reverse paclitaxel-induced mechanical allodynia in a mouse model of chemotherapy-induced peripheral neuropathy (CIPN). Recent studies demonstrated that paclitaxel-induced increases in inflammatory markers (GPR55, NLRP3 and IL-1b) of dorsal root ganglion (DRG) cultures were shown to be reversed by KLS-13019 treatment. The mechanism of action for KLS-13019-mediated reversal of paclitaxel-induced neuroinflammation now has been explored using GPR55 siRNA. Pretreatment of DRG cultures with GPR55 siRNA produced a 21% decrease of immunoreactive (IR) area for GPR55 in cell bodies and a 59% decrease in neuritic IR area, as determined by high content imaging. Using a 24-hour reversal treatment paradigm, paclitaxel-induced increases in the inflammatory markers were reversed back to control levels after KLS-3019 treatment. Decreases in these inflammatory markers produced by KLS-13019 were significantly attenuated by GPR55 siRNA co-treatment, with mean IR area responses being attenuated by 56% in neurites and 53% in cell bodies. These data indicate that the percentage decreases in siRNA-mediated attenuation of KLS-13019-related efficacy on the inflammatory markers were similar to the percentage knockdown observed for neuritic GPR55 IR area. Similar studies conducted with cannabidiol (CBD), the parent compound of KLS-13019, produced low efficacy (25%) reversal of all inflammatory markers that were poorly attenuated (29%) by GPR55 siRNA. CBD was shown previously to be ineffective in reversing paclitaxel-induced mechanical allodynia. The present studies indicated significant differences between the anti-inflammatory properties of KLS-13019 and CBD which may play a role in their observed differences in the reversibility of mechanical allodynia in a mouse model of CIPN.

8.
J Mol Neurosci ; 74(2): 41, 2024 Apr 11.
Artículo en Inglés | MEDLINE | ID: mdl-38602576

RESUMEN

KLS-13019 was reported previously to reverse paclitaxel-induced mechanical allodynia in a mouse model of chemotherapy-induced peripheral neuropathy (CIPN). Recent studies demonstrated that paclitaxel-induced increases in inflammatory markers (GPR55, NLRP3, and IL-1ß) of dorsal root ganglion (DRG) cultures were shown to be reversed by KLS-13019 treatment. The mechanism of action for KLS-13019-mediated reversal of paclitaxel-induced neuroinflammation now has been explored using GPR55 siRNA. Pre-treatment of DRG cultures with GPR55 siRNA produced a 21% decrease of immunoreactive (IR) area for GPR55 in cell bodies and a 59% decrease in neuritic IR area, as determined by high-content imaging. Using a 24-h reversal treatment paradigm, paclitaxel-induced increases in the inflammatory markers were reversed back to control levels after KLS-3019 treatment. Decreases in these inflammatory markers produced by KLS-13019 were significantly attenuated by GPR55 siRNA co-treatment, with mean IR area responses being attenuated by 56% in neurites and 53% in cell bodies. These data indicate that the percentage decreases in siRNA-mediated attenuation of KLS-13019-related efficacy on the inflammatory markers were similar to the percentage knockdown observed for neuritic GPR55 IR area. Similar studies conducted with cannabidiol (CBD), the parent compound of KLS-13019, produced low efficacy (25%) reversal of all inflammatory markers that were poorly attenuated (29%) by GPR55 siRNA. CBD was shown previously to be ineffective in reversing paclitaxel-induced mechanical allodynia. The present studies indicated significant differences between the anti-inflammatory properties of KLS-13019 and CBD which may play a role in their observed differences in the reversibility of mechanical allodynia in a mouse model of CIPN.


Asunto(s)
Cannabidiol , Animales , Ratones , ARN Interferente Pequeño/genética , Cannabidiol/farmacología , Cannabidiol/uso terapéutico , Hiperalgesia/tratamiento farmacológico , Antiinflamatorios , Modelos Animales de Enfermedad , Paclitaxel/toxicidad , Receptores de Cannabinoides/genética
9.
Bioorg Med Chem ; 20(11): 3609-14, 2012 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-22546203

RESUMEN

Inhibitors of both heat shock proteins Hsp90 and Hsp70 have been identified in assays measuring luciferase refolding containing rabbit reticulocyte lysate or purified chaperone components. Here, we report the discovery of a series of phenoxy-N-arylacetamides that disrupt Hsp70-mediated luciferase refolding by binding to DnaJ, the bacterial homolog of human Hsp40. Inhibitor characterization experiments demonstrated negative cooperativity with respect to DnaJ and luciferase concentration, but varying the concentration of ATP had no effect on potency. Thermal shift analysis suggested a direct interaction with DnaJ, but not with Hsp70. These compounds may be useful tools for studying DnaJ/Hsp40 in various cellular processes.


Asunto(s)
Acetamidas/química , Acetamidas/farmacología , Proteínas de Escherichia coli/metabolismo , Proteínas del Choque Térmico HSP40/metabolismo , Proteínas HSP70 de Choque Térmico/antagonistas & inhibidores , Luciferasas/química , Luciferasas/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Evaluación Preclínica de Medicamentos , Proteínas HSP70 de Choque Térmico/metabolismo , Humanos , Concentración 50 Inhibidora , Pliegue de Proteína , Conejos
10.
Bioorg Med Chem ; 20(18): 5642-8, 2012 Sep 15.
Artículo en Inglés | MEDLINE | ID: mdl-22892214

RESUMEN

Riluzole (1) is an approved therapeutic for the treatment of ALS and has also demonstrated anti-melanoma activity in metabotropic glutamate GRM1 positive cell lines, a mouse xenograft assay and human clinical trials. Highly variable drug exposure following oral administration among patients, likely due to variable first pass effects from heterogeneous CYP1A2 expression, hinders its clinical use. In an effort to mitigate effects of this clearance pathway and uniformly administer riluzole at efficacious exposure levels, several classes of prodrugs of riluzole were designed, synthesized, and evaluated in multiple in vitro stability assays to predict in vivo drug levels. The optimal prodrug would possess the following profile: stability while transiting the digestive system, stability towards first pass metabolism, and metabolic lability in the plasma releasing riluzole. (S)-O-Benzyl serine derivative 9 was identified as the most promising therapeutically acceptable prodrug.


Asunto(s)
Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Diseño de Fármacos , Melanoma/tratamiento farmacológico , Profármacos/síntesis química , Profármacos/farmacología , Riluzol/metabolismo , Riluzol/farmacología , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Citocromo P-450 CYP1A2/biosíntesis , Citocromo P-450 CYP1A2/metabolismo , Estabilidad de Medicamentos , Humanos , Melanoma/metabolismo , Ratones , Microsomas Hepáticos/metabolismo , Estructura Molecular , Profármacos/química , Profármacos/metabolismo , Riluzol/sangre , Riluzol/síntesis química
11.
Tetrahedron Lett ; 53(25): 3144-3146, 2012 Jun 20.
Artículo en Inglés | MEDLINE | ID: mdl-22822274

RESUMEN

3-Chloro-4-(dichloromethyl)-5-hydroxy-5H-furan-2-one (Mutagen X, MX) was synthesized in six steps from commercially-available and inexpensive starting materials (27% overall yield). This synthesis enables the preparation of MX analogs and does not require the use of chlorine gas, as do previously reported methods.

12.
J Mol Neurosci ; 72(9): 1859-1874, 2022 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-35779192

RESUMEN

KLS-13019, a novel devised cannabinoid-like compound, was explored for anti-inflammatory actions in dorsal root ganglion cultures relevant to chemotherapy-induced peripheral neuropathy (CIPN). Time course studies with 3 µM paclitaxel indicated > 1.9-fold increases in immunoreactive (IR) area for cell body GPR55 after 30 min as determined by high content imaging. To test for reversibility of paclitaxel-induced increases in GPR55, cultures were treated for 8 h with paclitaxel alone and then a dose response to KLS-13019 added for another 16 h. This "reversal" paradigm indicated established increases in cell body GPR55 IR areas were decreased back to control levels. Because GPR55 had previously reported inflammatory actions, IL-1ß and NLRP3 (inflammasome-3 marker) were also measured in the "reversal" paradigm. Significant increases in all inflammatory markers were produced after 8 h of paclitaxel treatment alone that were reversed to control levels with KLS-13019 treatment. Accompanying studies using alamar blue indicated that decreased cellular viability produced by paclitaxel treatment was reverted back to control levels by KLS-13019. Similar studies conducted with lysophosphatidylinositol (GPR55 agonist) in DRG or hippocampal cultures demonstrated significant increases in neuritic GPR55, NLRP3 and IL-1ß areas that were reversed to control levels with KLS-13019 treatment. Studies with a human GPR55-ß-arrestin assay in Discover X cells indicated that KLS-13019 was an antagonist without agonist activity. These studies indicated that KLS-13019 has anti-inflammatory properties mediated through GPR55 antagonist actions. Together with previous studies, KLS-13019 is a potent neuroprotective, anti-inflammatory cannabinoid with therapeutic potential for high efficacy treatment of neuropathic pain.


Asunto(s)
Cannabinoides , Neuralgia , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Cannabinoides/uso terapéutico , Ganglios Espinales/metabolismo , Hipocampo/metabolismo , Humanos , Proteína con Dominio Pirina 3 de la Familia NLR , Neuralgia/tratamiento farmacológico , Paclitaxel/farmacología , Receptores de Cannabinoides/metabolismo
13.
Biochem Pharmacol ; 167: 149-162, 2019 09.
Artículo en Inglés | MEDLINE | ID: mdl-30880062

RESUMEN

In our previous studies of the molecular mechanisms of poly(ADP-ribose) polymerase 1 (PARP-1)-mediated transcriptional regulation we identified a novel class of PARP-1 inhibitors targeting the histone-dependent route of PARP-1 activation. Because histone-dependent activation is unique to PARP-1, non-NAD-like PARP-1 inhibitors have the potential to bypass the off-target effects of classical NAD-dependent PARP-1 inhibitors, such as olaparib, veliparib, and rucaparib. Furthermore, our recently published studies demonstrate that, compared to NAD-like PARP-1 inhibitors that are used clinically, the non-NAD-like PARP-1 inhibitor 5F02 exhibited superior antitumor activity in cell and animal models of human prostate cancer (PC). In this study, we further evaluated the antitumor activity of 5F02 and several of its novel analogues against PC cells. In contrast to NAD-like PARP-1 inhibitors, non-NAD-like PARP-1 inhibitors demonstrated efficacy against androgen-dependent and -independent routes of androgen receptor signaling activation. Our experiments reveal that methylation of the quaternary ammonium salt and the presence of esters were critical for the antitumor activity of 5F02 against PC cells. In addition, we examined the role of a related regulatory protein of PARP-1, called Poly(ADP-ribose) glycohydrolase (PARG), in prostate carcinogenesis. Our study reveals that PARG expression is severely disrupted in PC cells, which is associated with decreased integrity and localization of Cajal bodies (CB). Overall, the results of our study strengthen the justification for using non-NAD-like PARP-1 inhibitors as a novel therapeutic strategy for the treatment of advanced prostate cancer.


Asunto(s)
Antineoplásicos/farmacología , NAD , Poli(ADP-Ribosa) Polimerasa-1/antagonistas & inhibidores , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Neoplasias de la Próstata/enzimología , Animales , Antineoplásicos/uso terapéutico , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Relación Dosis-Respuesta a Droga , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/patología
14.
Sci Transl Med ; 11(482)2019 03 06.
Artículo en Inglés | MEDLINE | ID: mdl-30842315

RESUMEN

Epstein-Barr virus (EBV) is a DNA tumor virus responsible for 1 to 2% of human cancers including subtypes of Burkitt's lymphoma, Hodgkin's lymphoma, gastric carcinoma, and nasopharyngeal carcinoma (NPC). Persistent latent infection drives EBV-associated tumorigenesis. Epstein-Barr nuclear antigen 1 (EBNA1) is the only viral protein consistently expressed in all EBV-associated tumors and is therefore an attractive target for therapeutic intervention. It is a multifunctional DNA binding protein critical for viral replication, genome maintenance, viral gene expression, and host cell survival. Using a fragment-based approach and x-ray crystallography, we identify a 2,3-disubstituted benzoic acid series that selectively inhibits the DNA binding activity of EBNA1. We characterize these inhibitors biochemically and in cell-based assays, including chromatin immunoprecipitation and DNA replication assays. In addition, we demonstrate the potency of EBNA1 inhibitors to suppress tumor growth in several EBV-dependent xenograft models, including patient-derived xenografts for NPC. These inhibitors selectively block EBV gene transcription and alter the cellular transforming growth factor-ß (TGF-ß) signaling pathway in NPC tumor xenografts. These EBNA1-specific inhibitors show favorable pharmacological properties and have the potential to be further developed for the treatment of EBV-associated malignancies.


Asunto(s)
ADN Viral/metabolismo , Diseño de Fármacos , Antígenos Nucleares del Virus de Epstein-Barr/metabolismo , Herpesvirus Humano 4/fisiología , Neoplasias Nasofaríngeas/virología , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Latencia del Virus/efectos de los fármacos , Animales , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Regulación Viral de la Expresión Génica/efectos de los fármacos , Humanos , Ratones Desnudos , Neoplasias Nasofaríngeas/patología , Relación Estructura-Actividad
15.
16.
ACS Med Chem Lett ; 7(4): 424-8, 2016 Apr 14.
Artículo en Inglés | MEDLINE | ID: mdl-27096053

RESUMEN

Cannabidiol is the nonpsychoactive natural component of C. sativa that has been shown to be neuroprotective in multiple animal models. Our interest is to advance a therapeutic candidate for the orphan indication hepatic encephalopathy (HE). HE is a serious neurological disorder that occurs in patients with cirrhosis or liver failure. Although cannabidiol is effective in models of HE, it has limitations in terms of safety and oral bioavailability. Herein, we describe a series of side chain modified resorcinols that were designed for greater hydrophilicity and "drug likeness", while varying hydrogen bond donors, acceptors, architecture, basicity, neutrality, acidity, and polar surface area within the pendent group. Our primary screen evaluated the ability of the test agents to prevent damage to hippocampal neurons induced by ammonium acetate and ethanol at clinically relevant concentrations. Notably, KLS-13019 was 50-fold more potent and >400-fold safer than cannabidiol and exhibited an in vitro profile consistent with improved oral bioavailability.

17.
Biochimie ; 94(9): 1974-81, 2012 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-22659571

RESUMEN

Dysfunction of the heterogeneous ribonucleoprotein TAR DNA binding protein 43 (TDP-43) is associated with neurodegeneration in diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Here we examine the effects of a series of 4-aminoquinolines with affinity for TDP-43 upon caspase-7-induced cleavage of TDP-43 and TDP-43 cellular function. These compounds were mixed inhibitors of biotinylated TG6 binding to TDP-43, binding to both free and occupied TDP-43. Incubation of TDP-43 and caspase-7 in the presence of these compounds stimulated caspase-7 mediated cleavage of TDP-43. This effect was antagonized by the oligonucleotide TG12, prevented by denaturing TDP-43, and exhibited a similar relation of structure to function as for the displacement of bt-TG6 binding to TDP-43. In addition, the compounds did not affect caspase-7 enzyme activity. In human neuroglioma H4 cells, these compounds lowered levels of TDP-43 and increased TDP-43 C-terminal fragments via a caspase-dependent mechanism. Subsequent experiments demonstrated that this was due to induction of caspases 3 and 7 leading to increased PARP cleavage in H4 cells with similar rank order of the potency among the compounds tests for displacement of bt-TG6 binding. Exposure to these compounds also reduced HDAC-6, ATG-7, and increased LC3B, consistent with the effects of TDP-43 siRNA described by other investigators. These data suggest that such compounds may be useful biochemical probes to further understand both the normal and pathological functions of TDP-43, and its cleavage and metabolism promoted by caspases.


Asunto(s)
Aminoquinolinas/farmacología , Caspasa 7/metabolismo , Proteínas de Unión al ADN/antagonistas & inhibidores , Proteínas de Unión al ADN/metabolismo , Proteolisis/efectos de los fármacos , Aminoquinolinas/metabolismo , Proteína 7 Relacionada con la Autofagia , Biotinilación/efectos de los fármacos , Caspasa 3/metabolismo , Línea Celular Tumoral , Proteínas de Unión al ADN/deficiencia , Proteínas de Unión al ADN/genética , Activación Enzimática/efectos de los fármacos , Ensayos Analíticos de Alto Rendimiento , Histona Desacetilasa 6 , Histona Desacetilasas/metabolismo , Humanos , Poli(ADP-Ribosa) Polimerasas/metabolismo , ARN Interferente Pequeño/genética , Enzimas Activadoras de Ubiquitina/metabolismo
18.
Protein Sci ; 20(4): 670-83, 2011 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-21308848

RESUMEN

A high-resolution structure of a ligand-bound, soluble form of human monoglyceride lipase (MGL) is presented. The structure highlights a novel conformation of the regulatory lid-domain present in the lipase family as well as the binding mode of a pharmaceutically relevant reversible inhibitor. Analysis of the structure lacking the inhibitor indicates that the closed conformation can accommodate the native substrate 2-arachidonoyl glycerol. A model is proposed in which MGL undergoes conformational and electrostatic changes during the catalytic cycle ultimately resulting in its dissociation from the membrane upon completion of the cycle. In addition, the study outlines a successful approach to transform membrane associated proteins, which tend to aggregate upon purification, into a monomeric and soluble form.


Asunto(s)
Monoacilglicerol Lipasas/antagonistas & inhibidores , Monoacilglicerol Lipasas/química , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Ácidos Araquidónicos/química , Ácidos Araquidónicos/metabolismo , Moduladores de Receptores de Cannabinoides/química , Moduladores de Receptores de Cannabinoides/metabolismo , Dominio Catalítico , Cristalografía por Rayos X , Endocannabinoides , Glicéridos/química , Glicéridos/metabolismo , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Estructura Molecular , Monoacilglicerol Lipasas/genética , Monoacilglicerol Lipasas/metabolismo , Mutagénesis Sitio-Dirigida , Unión Proteica , Electricidad Estática
19.
Curr Med Res Opin ; 26(10): 2375-84, 2010 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-20804444

RESUMEN

OBJECTIVE: Diclofenac potassium liquid-filled soft gelatin capsule (DPSGC) is a rapidly absorbed formulation of diclofenac approved for the treatment of mild to moderate acute pain in adults (≥18 years of age). The objective of this study was to investigate the efficacy and safety of DPSGC 25 mg in a multicenter, randomized, double-blind, placebo-controlled study in patients experiencing pain following first metatarsal bunionectomy. RESEARCH DESIGN AND METHODS: Patients experiencing a requisite level of pain (≥4 based on an 11-point numeric pain rating scale [NPRS]; 0 = no pain, 10 = worst pain possible) on the day following surgery were randomized to receive DPSGC 25 mg or placebo. Patients received a second dose (remedication) on request or at 8 hours postdose followed by additional doses every 6 hours through the end of postsurgery Day 4. Rescue medication (hydrocodone/acetaminophen) was available as needed after the second dose. CLINICAL TRIAL REGISTRATION: NCT00375934. MAIN OUTCOME MEASURE: The primary efficacy endpoint was the average NPRS score over the 48 hour inpatient multiple-dose period. RESULTS: DPSGC provided a significant improvement in mean 48 hour NPRS scores over placebo (3.29 vs 5.74, respectively; p < 0.0001), as well as for summed pain intensity difference (203.1 vs 86.6; p < 0.0001). Patients treated with DPSGC experienced a faster onset of meaningful pain relief compared with placebo (p = 0.0034). Rescue medication use on Day 1 and Day 2 was reduced in the DPSGC group compared with placebo (53.5% vs 92.1% on Day 1; 30.3% vs 67.3% on Day 2; p < 0.0001). DPSGC was well tolerated and no patients treated with DPSGC reported serious adverse events. As with any study, there are potential limitations including study design and patient population. CONCLUSION: These results indicate that DPSGC reduced pain in patients who underwent bunionectomy and this novel formulation of diclofenac potassium may be a practical option for treating mild to moderate acute pain.


Asunto(s)
Diclofenaco/administración & dosificación , Hallux Valgus/cirugía , Dolor Postoperatorio/tratamiento farmacológico , Adolescente , Adulto , Anciano , Antiinflamatorios no Esteroideos/administración & dosificación , Antiinflamatorios no Esteroideos/efectos adversos , Cápsulas , Diclofenaco/efectos adversos , Relación Dosis-Respuesta a Droga , Método Doble Ciego , Femenino , Gelatina , Hallux Valgus/tratamiento farmacológico , Humanos , Masculino , Huesos Metatarsianos/cirugía , Persona de Mediana Edad , Ortopedia/métodos , Placebos , Adulto Joven
20.
Toxicol Sci ; 117(2): 493-504, 2010 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-20616206

RESUMEN

The unexpected observation of a hyperglycemic effect of some tricycle-based delta opioid receptor (DOR) agonists led to a series of studies to better understand the finding. Single administration of two novel tricyclic DOR agonists dose dependently elevated rat plasma glucose levels; 4-week toxicology studies confirmed the hyperglycemic finding and further revealed pancreatic ß-cell hypertrophy, including vacuole formation, as well as bone dysplasia and Harderian gland degeneration with regeneration. Similar diabetogenic effects were observed in dog. A review of the literature on the antiserotonergic and antihistaminergic drug cyproheptadine (CPH) and its metabolites revealed shared structural features as well as similar hyperglycemic effects to the present series of DOR agonists. To further evaluate these effects, we established an assay measuring insulin levels in the rat pancreatic ß-cell-derived RINm5F cell line, extensively used to study CPH and its metabolites. Like CPH, the initial DOR agonists studied reduced RINm5F cell insulin levels in a concentration-dependent manner. Importantly, compound DOR potency did not correlate with the insulin-reducing potency. Furthermore, the RINm5F cell insulin results correlated with the diabetogenic effect of the compounds in a 5-day mouse study. The RINm5F cell insulin assay enabled the identification of aryl-aryl-amine DOR agonists that lacked an insulin-reducing effect and did not elevate blood glucose in repeated dosing studies conducted over a suprapharmacologic dose range. Thus, not only did the RINm5F cell assay open a path for the further discovery of DOR agonists lacking diabetogenic potential but also it established a reliable, economical, and high-throughput screen for such potential, regardless of chemotype or target pharmacology. The present findings also suggest a mechanistic link between the toxicity observed here and that underlying Wolcott-Rallison Syndrome.


Asunto(s)
Ciproheptadina/toxicidad , Hiperglucemia/inducido químicamente , Células Secretoras de Insulina/efectos de los fármacos , Antagonistas de Narcóticos/toxicidad , Páncreas/efectos de los fármacos , Antagonistas de la Serotonina/toxicidad , Animales , Glucemia/análisis , Glucemia/efectos de los fármacos , Aumento de la Célula/efectos de los fármacos , Línea Celular Tumoral , Ciproheptadina/análogos & derivados , Diabetes Mellitus Tipo 1/metabolismo , Perros , Epífisis/anomalías , Epífisis/metabolismo , Femenino , Ensayos Analíticos de Alto Rendimiento , Hiperglucemia/metabolismo , Insulina/sangre , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patología , Insulinoma/tratamiento farmacológico , Insulinoma/metabolismo , Masculino , Ratones , Osteocondrodisplasias/metabolismo , Páncreas/metabolismo , Páncreas/patología , Ratas , Ratas Sprague-Dawley , Vacuolas/efectos de los fármacos , Vacuolas/ultraestructura
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