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1.
Bioorg Med Chem Lett ; 28(8): 1381-1385, 2018 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-29555153

RESUMEN

The approach of tethering together two known receptor ligands, to be used as molecular probes for the study of G protein-coupled receptor (GPCR) systems, has proven to be a valuable approach. Selective ligands that possess functionality that can be used to link to other ligands, are useful in the development of novel antagonists and agonists. Such molecules can also be attached to reporter molecules, such as fluorophores, for the study of GPCR dimerization and its role in signaling. The highly selective serotonin (5-HT) 5-HT2A receptor (5-HT2AR) antagonist M100907 (volinanserin) is of clinical interest in the treatment of neurological and mental health disorders. Here, we synthesized the most active (+)-M100907 enantiomer as well as a series of derivatives that possessed either an alkyne or an azide. The triazole resulting from the dipolar cycloaddition of these groups did not interfere with the ability of the bivalent ligand to act as an antagonist. Thus, we have synthesized a number of compounds which will prove useful in elucidating the role of the 5-HT2AR in the central nervous system.


Asunto(s)
Fluorobencenos/farmacología , Piperidinas/farmacología , Receptor de Serotonina 5-HT2A/metabolismo , Antagonistas del Receptor de Serotonina 5-HT2/farmacología , Alquinos/síntesis química , Alquinos/química , Alquinos/farmacología , Animales , Azidas/síntesis química , Azidas/química , Azidas/farmacología , Células CHO , Calcio/metabolismo , Cricetulus , Fluorobencenos/síntesis química , Fluorobencenos/química , Piperidinas/síntesis química , Piperidinas/química , Antagonistas del Receptor de Serotonina 5-HT2/síntesis química , Antagonistas del Receptor de Serotonina 5-HT2/química , Estereoisomerismo
2.
Bioorg Med Chem Lett ; 26(18): 4446-4450, 2016 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-27530289

RESUMEN

Due to the oncogenic activity of cohesin protease, separase in human cancer cells, modulation of separase enzymatic activity could constitute a new therapeutic strategy for targeting resistant, separase-overexpressing aneuploid tumors. Herein, we report the synthesis, structural information, and structure-activity relationship (SAR) of separase inhibitors based on modification of the lead molecule 2,2-dimethyl-5-nitro-2H-benzimidazole-1,3-dioxide, named Sepin-1, (1) identified from a high-throughput-screen. Replacement of -NO2 at C5 with other functional groups reduce the inhibitory activity in separase enzymatic assay. Substitution of the two methyl groups with other alkyl chains at the C2 moderately improves the effects on the inhibitory activity of those compounds. Modifications on 2H-benzimidazole-1,3-dioxide or the skeleton have variable effect on inhibition of separase enzymatic activity. Density-functional theory (DFT) calculations suggest there may be a correlation between the charges on the oxide moieties on these compounds and their activity in inhibiting separase enzyme.


Asunto(s)
Bencimidazoles/síntesis química , Bencimidazoles/farmacología , Inhibidores Enzimáticos/farmacología , Separasa/antagonistas & inhibidores , Bencimidazoles/química , Inhibidores Enzimáticos/química , Relación Estructura-Actividad
3.
J Neurosci ; 33(4): 1615-30, 2013 Jan 23.
Artículo en Inglés | MEDLINE | ID: mdl-23345234

RESUMEN

Serotonin (5-hydroxytryptamine; 5-HT) signaling through the 5-HT(2C) receptor (5-HT(2C)R) is essential in normal physiology, whereas aberrant 5-HT(2C)R function is thought to contribute to the pathogenesis of multiple neural disorders. The 5-HT(2C)R interacts with specific protein partners, but the impact of such interactions on 5-HT(2C)R function is poorly understood. Here, we report convergent cellular and behavioral data that the interaction between the 5-HT(2C)R and protein phosphatase and tensin homolog (PTEN) serves as a regulatory mechanism to control 5-HT(2C)R-mediated biology but not that of the closely homologous 5-HT(2A)R. A peptide derived from the third intracellular loop of the human 5-HT(2C)R [3L4F (third loop, fourth fragment)] disrupted the association, allosterically augmented 5-HT(2C)R-mediated signaling in live cells, and acted as a positive allosteric modulator in rats in vivo. We identified the critical residues within an 8 aa fragment of the 3L4F peptide that maintained efficacy (within the picomolar range) in live cells similar to that of the 3L4F peptide. Last, molecular modeling identified key structural features and potential interaction sites of the active 3L4F peptides against PTEN. These compelling data demonstrate the specificity and importance of this protein assembly in cellular events and behaviors mediated by 5-HT(2C)R signaling and provide a chemical guidepost to the future development of drug-like peptide or small-molecule inhibitors as neuroprobes to study 5-HT(2C)R allostery and therapeutics for 5-HT(2C)R-mediated disorders.


Asunto(s)
Modelos Moleculares , Fosfohidrolasa PTEN/química , Fosfohidrolasa PTEN/metabolismo , Receptor de Serotonina 5-HT2C/química , Receptor de Serotonina 5-HT2C/metabolismo , Transducción de Señal/fisiología , Secuencia de Aminoácidos , Animales , Humanos , Immunoblotting , Inmunoprecipitación , Masculino , Datos de Secuencia Molecular , Actividad Motora/fisiología , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Ratas , Ratas Sprague-Dawley , Serotonina/metabolismo , Transfección
4.
Bioorg Med Chem ; 20(1): 368-76, 2012 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-22154558

RESUMEN

Edema factor (EF) toxin of Bacillus anthracis (NIAID category A), and several other toxins from NIAID category B Biodefense target bacteria are adenylyl cyclases or adenylyl cyclase agonists that catalyze the conversion of ATP to 3',5'-cyclic adenosine monophosphate (cAMP). We previously identified compound 1 (3-[(9-oxo-9H-fluorene-1-carbonyl)-amino]-benzoic acid), that inhibits EF activity in cultured mammalian cells, and reduces diarrhea caused by enterotoxigenic Escherichia coli (ETEC) at an oral dosage of 15µg/mouse. Here, molecular docking was used to predict improvements in potency and solubility of new derivatives of compound 1 in inhibiting edema toxin (ET)-catalyzed stimulation of cyclic AMP production in murine monocyte-macrophage cells (RAW 264.7). Structure-activity relationship (SAR) analysis of the bioassay results for 22 compounds indicated positions important for activity. Several derivatives demonstrated superior pharmacological properties compared to our initial lead compound, and are promising candidates to treat anthrax infections and diarrheal diseases induced by toxin-producing bacteria.


Asunto(s)
Bacillus anthracis/metabolismo , Toxinas Bacterianas/antagonistas & inhibidores , Diseño de Fármacos , Inhibidores de Adenilato Ciclasa , Adenilil Ciclasas/metabolismo , Administración Oral , Animales , Antígenos Bacterianos/metabolismo , Toxinas Bacterianas/metabolismo , Benzoatos/síntesis química , Benzoatos/química , Sitios de Unión , Línea Celular , Simulación por Computador , AMP Cíclico/metabolismo , Fluorenos/química , Ratones , Estructura Terciaria de Proteína , Relación Estructura-Actividad
5.
ChemMedChem ; 17(9): e202100653, 2022 05 04.
Artículo en Inglés | MEDLINE | ID: mdl-35018729

RESUMEN

STAG2 (SA2) is a critical component of the cohesin complex that regulates gene expression and the separation of sister chromatids in cells. Mutations in STAG2 have been identified in over thirty different types of cancers including myeloid leukaemia, non-small cell lung, bladder and Ewing sarcoma. Selectively inhibiting cancer cells lacking of STAG2 is an attractive approach for the cancer therapy. Here we report that a small molecule, StagX1, identified through a high-throughput screening, inhibits the growth of Ewing sarcoma cells possessing mutant STAG2. A new synthetic route to the StagX1 scaffold and new versions of the molecule along with their activity in a cell viability assay are reported.


Asunto(s)
Sarcoma de Ewing , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Humanos , Mutación , Sarcoma de Ewing/tratamiento farmacológico , Sarcoma de Ewing/genética
6.
Proc Natl Acad Sci U S A ; 105(24): 8440-5, 2008 Jun 17.
Artículo en Inglés | MEDLINE | ID: mdl-18559851

RESUMEN

Acute secretory diarrhea induced by infection with enterotoxigenic strains of Escherichia coli involves binding of stable toxin (STa) to its receptor on the intestinal brush border, guanylyl cyclase type C (GC-C). Intracellular cGMP is elevated, inducing increase in chloride efflux and subsequent accumulation of fluid in the intestinal lumen. We have screened a library of compounds and identified a pyridopyrimidine derivatives {5-(3-bromophenyl)-1,3-dimethyl-5,11-dihydro-1H-indeno[2',1':5,6]pyrido[2,3-d]pyrimidine-2,4,6-trione; BPIPP} as an inhibitor of GC-C that can suppress STa-stimulated cGMP accumulation by decreasing GC-C activation in intact T84 human colorectal carcinoma cells. BPIPP inhibited stimulation of guanylyl cyclases, including types A and B and soluble isoform in various cells. BPIPP suppressed stimulation of adenylyl cyclase and significantly decreased the activities of adenylyl cyclase toxin of Bordetella pertussis and edema toxin of Bacillus anthracis. The effects of BPIPP on cyclic nucleotide synthesis were observed only in intact cells. The mechanism of BPIPP-dependent inhibition appears to be complex and indirect, possibly associated with phospholipase C and tyrosine-specific phosphorylation. BPIPP inhibited chloride-ion transport stimulated by activation of guanylyl or adenylyl cyclases and suppressed STa-induced fluid accumulation in an in vivo rabbit intestinal loop model. Thus, BPIPP may be a promising lead compound for treatment of diarrhea and other diseases.


Asunto(s)
Inhibidores de Adenilato Ciclasa , Antidiarreicos/química , Antidiarreicos/farmacología , Diarrea/tratamiento farmacológico , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Guanilato Ciclasa/antagonistas & inhibidores , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Animales , Antidiarreicos/uso terapéutico , Toxinas Bacterianas/farmacología , Línea Celular , Cricetinae , AMP Cíclico/antagonistas & inhibidores , AMP Cíclico/biosíntesis , GMP Cíclico/antagonistas & inhibidores , GMP Cíclico/biosíntesis , Diarrea/enzimología , Enterotoxinas/farmacología , Inhibidores Enzimáticos/uso terapéutico , Proteínas de Escherichia coli , Compuestos Heterocíclicos de 4 o más Anillos/uso terapéutico , Humanos , Ratas , Bibliotecas de Moléculas Pequeñas
7.
Org Lett ; 23(8): 2911-2914, 2021 Apr 16.
Artículo en Inglés | MEDLINE | ID: mdl-33783221

RESUMEN

A diastereoselective [4 + 2] cycloisomerization of asymmetric allenyl dienes is reported. The asymmetric dienyl allenes are synthesized using the method reported by Ma. These substrates readily undergo diastereoselective intramolecular rhodium catalyzed [4 + 2] cycloisomerization analogous to thermal intramolecular Diels-Alder reactions. Overall, 29 examples are presented with tethers possessing nitrogen, oxygen, and carbon. Diastereoselectivities range from 99:1 to 90:10 in most examples.

8.
Infect Immun ; 78(4): 1740-9, 2010 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-20123712

RESUMEN

Enterotoxigenic Escherichia coli (ETEC) produces the ADP-ribosyltransferase toxin known as heat-labile enterotoxin (LT). In addition to the toxic effect of LT resulting in increases of cyclic AMP (cAMP) and disturbance of cellular metabolic processes, this toxin promotes bacterial adherence to intestinal epithelial cells (A. M. Johnson, R. S. Kaushik, D. H. Francis, J. M. Fleckenstein, and P. R. Hardwidge, J. Bacteriol. 191:178-186, 2009). Therefore, we hypothesized that the identification of a compound that inhibits the activity of the toxin would have a suppressive effect on the ETEC colonization capabilities. Using in vivo and in vitro approaches, we present evidence demonstrating that a fluorenone-based compound, DC5, which inhibits the accumulation of cAMP in intoxicated cultured cells, significantly decreases the colonization abilities of adenylyl cyclase toxin-producing bacteria, such as ETEC. These findings established that DC5 is a potent inhibitor both of toxin-induced cAMP accumulation and of ETEC adherence to epithelial cells. Thus, DC5 may be a promising compound for treatment of diarrhea caused by ETEC and other adenylyl cyclase toxin-producing bacteria.


Asunto(s)
Inhibidores de Adenilato Ciclasa , Adhesinas Bacterianas/metabolismo , Toxinas Bacterianas/antagonistas & inhibidores , Escherichia coli Enterotoxigénica/patogenicidad , Enterotoxinas/antagonistas & inhibidores , Inhibidores Enzimáticos/administración & dosificación , Infecciones por Escherichia coli/prevención & control , Proteínas de Escherichia coli/antagonistas & inhibidores , Animales , Adhesión Bacteriana/efectos de los fármacos , Línea Celular , Recuento de Colonia Microbiana , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/toxicidad , Células Epiteliales/microbiología , Femenino , Fluorenos/administración & dosificación , Fluorenos/farmacología , Fluorenos/toxicidad , Humanos , Concentración 50 Inhibidora , Intestino Delgado/microbiología , Intestino Delgado/patología , Macrófagos/microbiología , Ratones
9.
Chem Commun (Camb) ; 56(14): 2211-2213, 2020 Feb 18.
Artículo en Inglés | MEDLINE | ID: mdl-32039415

RESUMEN

Correction for 'Diastereoselective synthesis of 1,3-disubstituted isoindolines and sultams via bronsted acid catalysis' by Ye Tao et al., Chem. Commun., 2018, 54, 11292-11295.

10.
Biochem Pharmacol ; 174: 113808, 2020 04.
Artículo en Inglés | MEDLINE | ID: mdl-31930961

RESUMEN

Separase, a sister chromatid cohesion-resolving enzyme, is an oncogene and overexpressed in many human cancers. Sepin-1 (2,2-dimethyl-5-nitro-2H-benzimidazole-1,3-dioxide) is a potent separase inhibitor that impedes cancer cell growth, cell migration, and wound healing, suggesting that Sepin-1 possesses a great potential to target separase-overexpressing tumors. As a part of the IND-enabling studies to bring Sepin-1 to clinic, herein we report the results from a 28-day repeat-dose pharmacokinetic study of Sepin-1 in rats. Sepin-1 was intravenously administered to Sprague-Dawley rats once daily for 28 days at three different (5, 10, and 20 mg/kg) doses. Blood samples were collected after administration of doses on days 1 and 28. Sepin-1 is unstable and isomerizes in basic solutions, but it is stable in acidic buffer such as citrate-buffered saline (pH 4.0). UHPLC-MS analysis indicated Sepin-1 was rapidly metabolized in vivo. One of the major metabolites was an amine adduct of 2,2-dimethyl-5-nitro-2H-benzimidazole (named Sepin-1.55). The concentration of Sepin-1.55 in blood samples was Sepin-1 dose-dependent and used for pharmacokinetic analysis of Sepin-1. Tmax was approximately 5-15 min. The data suggest that no Sepin-1 accumulation occurred from daily repeat dosing and similar exposures on the first and final day of dosing. Data also suggest a gender difference, namely that female rats have more exposure and slower clearance than male rats. The data support that Sepin-1 is a potential drug candidate that can be further developed to treat Separase-overexpressing human tumors.


Asunto(s)
Bencimidazoles , Inhibidores de Cisteína Proteinasa , Separasa/antagonistas & inhibidores , Animales , Bencimidazoles/química , Bencimidazoles/farmacocinética , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Inhibidores de Cisteína Proteinasa/química , Inhibidores de Cisteína Proteinasa/farmacocinética , Estabilidad de Medicamentos , Femenino , Masculino , Ratas , Ratas Sprague-Dawley , Separasa/sangre
11.
ACS Chem Biol ; 15(11): 2916-2928, 2020 11 20.
Artículo en Inglés | MEDLINE | ID: mdl-33074669

RESUMEN

Pancreatic ductal adenocarcinoma (PDAC) is the predominant form of pancreatic cancer. PDACs harbor oncogenic mutations in the KRAS gene, and ongoing efforts to directly target its mutant protein product to inhibit tumor growth are a priority not only in pancreatic cancer but in other malignancies such as lung and colorectal cancers where KRAS is also commonly mutated. An alternative strategy to directly targeting KRAS is to identify and target druggable receptors involved in dysregulated cancer hallmarks downstream of KRAS dysregulation. Liver X receptors (LXRs) are members of the nuclear receptor family of ligand-modulated transcription factors and are involved in the regulation of genes which function in key cancer-related processes, including cholesterol transport, lipid and glucose metabolism, and inflammatory and immune responses. Modulation of LXRs via small molecule ligands has emerged as a promising approach for directly targeting tumor cells or the stromal and immune cells within the tumor microenvironment. We have previously shown that only one of the two LXR subtypes (LXRß) is expressed in pancreatic cancer cells, and targeting LXR with available synthetic ligands blocked the proliferation of PDAC cells and tumor formation. In a screen of a focused library of drug-like small molecules predicted to dock in the ligand-binding pocket of LXRß, we identified two novel LXR ligands with more potent antitumor activity than current LXR agonists used in our published studies. Characterization of the two lead compounds (GAC0001E5 and GAC0003A4) indicates that they function as LXR inverse agonists which inhibit their transcriptional activity. Prolonged treatments with novel ligands further revealed their function as LXR "degraders" which significantly reduced LXR protein levels in all three PDAC cell lines tested. These findings support the utility of these novel inhibitors in basic research on ligand design, allosteric mechanisms, and LXR functions and their potential application as treatments for advanced pancreatic cancer and other recalcitrant malignancies.


Asunto(s)
Antineoplásicos/farmacología , Carcinoma Ductal Pancreático/tratamiento farmacológico , Receptores X del Hígado/metabolismo , Neoplasias Pancreáticas/tratamiento farmacológico , Bibliotecas de Moléculas Pequeñas/farmacología , Antineoplásicos/química , Carcinoma Ductal Pancreático/metabolismo , Línea Celular Tumoral , Agonismo Inverso de Drogas , Humanos , Ligandos , Receptores X del Hígado/agonistas , Neoplasias Pancreáticas/metabolismo , Proteolisis/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/química
12.
Bioorg Med Chem Lett ; 19(11): 3067-71, 2009 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-19409779

RESUMEN

A series of pyridopyrimidine derivatives were synthesized and evaluated for their ability to inhibit cyclic nucleotide synthesis in the presence of stable toxin a of Escherichia coli. The structure activity relationships around the basic core structure were examined and examples with better activity and potentially better pharmacological properties are presented.


Asunto(s)
Toxinas Bacterianas/metabolismo , GMP Cíclico/biosíntesis , Enterotoxinas/metabolismo , Pirimidinas/química , Línea Celular Tumoral , Escherichia coli/metabolismo , Proteínas de Escherichia coli , Guanilato Ciclasa/antagonistas & inhibidores , Guanilato Ciclasa/metabolismo , Humanos , Pirimidinas/síntesis química , Pirimidinas/farmacología , Receptores de Enterotoxina , Receptores Acoplados a la Guanilato-Ciclasa , Receptores de Péptidos/antagonistas & inhibidores , Receptores de Péptidos/metabolismo , Relación Estructura-Actividad
13.
Front Pharmacol ; 10: 907, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31507411

RESUMEN

Hypofunction of the serotonin (5-HT) 5-HT2C receptor (5-HT2CR) has been implicated in a variety of disorders including substance use disorders. As such, approaches to enhance 5-HT2CR signaling display therapeutic potential. In the present study, we show that disruption of the 5-HT2CR interaction with the protein phosphatase and tensin homolog (PTEN) via peptidomimetics enhances 5-HT2CR-mediating signaling in vitro and potentiates selective 5-HT2CR agonists in behavioral rodent models. Overall, the present study provides further evidence that 5-HT2CR activity can be modulated through an allosteric protein-protein interaction. This work provides the groundwork for the continued exploration of protein-protein interactions that can allosterically modulate this critical receptor and other important G protein-coupled receptors (GPCRs) for new therapeutic development through mechanisms that may display clinical utility.

14.
Bioorg Med Chem Lett ; 18(1): 427-31, 2008 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-17981463

RESUMEN

This paper reports a study to find small peptide substrates for the important virulence factor of Yersinia pestis, plasminogen activator, Pla. The method used to find small substrates for this protease is reported along with studies examining the ability of these peptides to inhibit activity of the enzyme. Through the use of parallel synthesis and positional scanning, small tripeptides were identified that are viable substrates for the protease.


Asunto(s)
Proteínas Bacterianas/antagonistas & inhibidores , Péptidos/síntesis química , Péptidos/farmacología , Activadores Plasminogénicos/antagonistas & inhibidores , Inhibidores de Proteasas/síntesis química , Inhibidores de Proteasas/farmacología , Secuencia de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Fluorometría , Cinética , Datos de Secuencia Molecular , Oligopéptidos/síntesis química , Oligopéptidos/química , Oligopéptidos/farmacología , Péptidos/química , Activadores Plasminogénicos/química , Activadores Plasminogénicos/metabolismo , Inhibidores de Proteasas/química , Conformación Proteica , Relación Estructura-Actividad , Especificidad por Sustrato , Yersinia pestis/enzimología
15.
Bioorg Med Chem Lett ; 18(14): 4215-8, 2008 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-18539457

RESUMEN

The synthesis and development of a novel class of molecules that inhibit anthrax edema factor, an adenylyl cyclase, is reported. These molecules are derived from the initial discovery that histidine and imidazole adducts of the prostaglandin PGE(2) reduce the net secretory response of cholera toxin-challenged mice and act directly on the action of anthrax edema factor, a calmodulin-dependent adenylyl cyclase. The simple enones examined in this letter were prepared by palladium-catalyzed Suzuki reaction.


Asunto(s)
Adenilil Ciclasas/química , Carbunco/metabolismo , Antiinfecciosos/síntesis química , Antiinfecciosos/farmacología , Adenilil Ciclasas/metabolismo , Animales , Antígenos Bacterianos , Bacillus anthracis/enzimología , Toxinas Bacterianas , Sitios de Unión , Calmodulina/metabolismo , Catálisis , Dinoprostona/metabolismo , Diseño de Fármacos , Cetonas , Ratones , Modelos Químicos , Paladio/química
16.
J Comb Chem ; 10(5): 655-7, 2008.
Artículo en Inglés | MEDLINE | ID: mdl-18611055

RESUMEN

This paper reports the preparation of a library of unsymmetrical ureas based on 8-azabicyclo[3.2.1]octane scaffold. The reported synthetic route uses nortropane-8-carbonyl chlorides as key intermediates that, when treated with a slight excess of amine, give the corresponding ureas in high yield (129 examples).


Asunto(s)
Técnicas Químicas Combinatorias/métodos , Tropanos/química , Urea/síntesis química , Aminas/química , Hidrocarburos Clorados/química , Modelos Químicos , Peso Molecular , Nortropanos/química , Solubilidad , Solventes/química
17.
Chem Commun (Camb) ; 54(80): 11292-11295, 2018 Oct 04.
Artículo en Inglés | MEDLINE | ID: mdl-30232479

RESUMEN

The bis(trifluoromethane)sulfonimide (Tf2NH) catalyzed intramolecular hydroamidation of terminal alkynes is reported. The combination of Et3SiH and Tf2NH provides cis-1,3-disubstituted isoindolines and sultams in high yield (up to 98%) and high diastereoselectivity (up to 99 : 1 d.r.).

18.
Front Pharmacol ; 9: 313, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-29867452

RESUMEN

Separase, a known oncogene, is widely overexpressed in numerous human tumors of breast, bone, brain, blood, and prostate. Separase is an emerging target for cancer therapy, and separase enzymatic inhibitors such as sepin-1 are currently being developed to treat separase-overexpressed tumors. Drug metabolism plays a critical role in the efficacy and safety of drug development, as well as possible drug-drug interactions. In this study, we investigated the in vitro metabolism of sepin-1 in human, mouse, and rat liver microsomes (RLM) using metabolomic approaches. In human liver microsomes (HLM), we identified seven metabolites including one cysteine-sepin-1 adduct and one glutathione-sepin-1 adduct. All the sepin-1 metabolites in HLM were also found in both mouse and RLM. Using recombinant CYP450 isoenzymes, we demonstrated that multiple enzymes contributed to the metabolism of sepin-1, including CYP2D6 and CYP3A4 as the major metabolizing enzymes. Inhibitory effects of sepin-1 on seven major CYP450s were also evaluated using the corresponding substrates recommended by the US Food and Drug Administration. Our studies indicated that sepin-1 moderately inhibits CYP1A2, CYP2C19, and CYP3A4 with IC50 < 10 µM but weakly inhibits CYP2B6, CYP2C8/9, and CYP2D6 with IC50 > 10 µM. This information can be used to optimize the structures of sepin-1 for more suitable pharmacological properties and to predict the possible sepin-1 interactions with other chemotherapeutic drugs.

19.
PLoS One ; 13(8): e0203137, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30157263

RESUMEN

The serotonin (5-HT) 5-HT2A receptor (5-HT2AR) and 5-HT2C receptor (5-HT2CR) in the central nervous system are implicated in a range of normal behaviors (e.g., appetite, sleep) and physiological functions (e.g., endocrine secretion) while dysfunctional 5-HT2AR and/or 5-HT2CR are implicated in neuropsychiatric disorders (e.g., addiction, obesity, schizophrenia). Preclinical studies suggest that the 5-HT2AR and 5-HT2CR may act in concert to regulate the neural bases for behavior. Here, we utilize three distinct biophysical and immunocytochemistry-based approaches to identify and study this receptor complex in cultured cells. Employing a split luciferase complementation assay (LCA), we demonstrated that formation of the 5-HT2AR:5-HT2CR complex exists within 50 nm, increases proportionally to the 5-HT2CR:5-HT2AR protein expression ratio, and is specific to the receptor interaction and not due to random complementation of the luciferase fragments. Using a proximity ligation assay (PLA), we found that cells stably expressing both the 5-HT2AR and 5-HT2CR exhibit 5-HT2AR:5-HT2CR heteroreceptor complexes within 40 nm of each other. Lastly, bioluminescence resonance energy transfer (BRET) analyses indicates the formation of a specific and saturable 5-HT2AR:5-HT2CR interaction, suggesting that the 5-HT2AR and 5-HT2CR form a close interaction within 10 nm of each other in intact live cells. The bioengineered receptors generated for the LCA and the BRET exhibit 5-HT-mediated intracellular calcium signaling as seen for the native receptors. Taken together, this study validates a very close 5-HT2AR:5-HT2CR interaction in cultured cells.


Asunto(s)
Receptor de Serotonina 5-HT2A/metabolismo , Receptor de Serotonina 5-HT2C/metabolismo , Animales , Fenómenos Biofísicos , Células CHO , Calcio/metabolismo , Señalización del Calcio/fisiología , Cricetulus , Células HEK293 , Humanos , Receptor de Serotonina 5-HT2A/genética , Receptor de Serotonina 5-HT2C/genética , Serotonina/metabolismo , Transfección
20.
ACS Chem Neurosci ; 9(3): 514-521, 2018 03 21.
Artículo en Inglés | MEDLINE | ID: mdl-29111677

RESUMEN

The 5-HT2A receptor (5-HT2AR) plays an important role in various neuropsychiatric disorders, including substance use disorder and schizophrenia. Homodimerization of this receptor has been suggested, but tools that allow direct assessment of the relevance of the 5-HT2AR:5-HT2AR homodimer in these disorders are necessary. We chemically modified the selective 5-HT2AR antagonist M100907 to synthesize a series of homobivalent ligands connected by ethylene glycol linkers of varying lengths that may be useful tools for probing 5-HT2AR:5-HT2AR homodimer function. We tested these molecules for 5-HT2AR antagonist activity in a cell line stably expressing the functional 5-HT2AR and quantified a downstream signaling target, activation (phosphorylation) of extracellular regulated kinases 1/2 (ERK1/2), in comparison to in vivo efficacy of altering spontaneous or cocaine-evoked locomotor activity in rats. All of the synthetic compounds inhibited 5-HT-mediated phosphorylation of ERK1/2 in the cellular signaling assay; the potency of the bivalent ligands varied as a function of linker length, with the intermediate linker lengths being the most potent. The Ki values for the binding of bivalent ligands to 5-HT2AR were only slightly lower than the values for the parent (+)-M100907 compound, but significant selectivity for 5-HT2AR over 5-HT2BR or 5-HT2CR binding was retained. In addition, the 11-atom-linked bivalent 5-HT2AR antagonist (2 mg/kg, intraperitoneally) demonstrated efficacy on par with that of (+)-M100907 in inhibiting cocaine-evoked hyperactivity. As we develop further strategies for ligand-evoked receptor assembly and analyses of diverse signaling and functional roles, these novel homobivalent 5-HT2AR antagonist ligands will serve as useful in vitro and in vivo probes of 5-HT2AR structure and function.


Asunto(s)
Calcio/metabolismo , Cocaína/farmacología , Receptor de Serotonina 5-HT2A/efectos de los fármacos , Antagonistas del Receptor de Serotonina 5-HT2/farmacología , Fosforilación/efectos de los fármacos , Unión Proteica/efectos de los fármacos , Receptor de Serotonina 5-HT2A/metabolismo , Transducción de Señal/efectos de los fármacos
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