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1.
Proc Natl Acad Sci U S A ; 117(32): 19190-19200, 2020 08 11.
Artículo en Inglés | MEDLINE | ID: mdl-32723828

RESUMEN

The 26S proteasome, a self-compartmentalized protease complex, plays a crucial role in protein quality control. Multiple levels of regulatory systems modulate proteasomal activity for substrate hydrolysis. However, the destruction mechanism of mammalian proteasomes is poorly understood. We found that inhibited proteasomes are sequestered into the insoluble aggresome via HDAC6- and dynein-mediated transport. These proteasomes colocalized with the autophagic receptor SQSTM1 and cleared through selective macroautophagy, linking aggresomal segregation to autophagic degradation. This proteaphagic pathway was counterbalanced with the recovery of proteasomal activity and was critical for reducing cellular proteasomal stress. Changes in associated proteins and polyubiquitylation on inhibited 26S proteasomes participated in the targeting mechanism to the aggresome and autophagosome. The STUB1 E3 Ub ligase specifically ubiquitylated purified human proteasomes in vitro, mainly via Lys63-linked chains. Genetic and chemical inhibition of STUB1 activity significantly impaired proteasome processing and reduced resistance to proteasomal stress. These data demonstrate that aggresomal sequestration is the crucial upstream event for proteasome quality control and overall protein homeostasis in mammals.


Asunto(s)
Macroautofagia , Orgánulos/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Células A549 , Humanos , Orgánulos/genética , Complejo de la Endopetidasa Proteasomal/genética , Proteína Sequestosoma-1/genética , Proteína Sequestosoma-1/metabolismo , Ubiquitina-Proteína Ligasas/genética , Ubiquitinación
2.
Molecules ; 26(11)2021 May 31.
Artículo en Inglés | MEDLINE | ID: mdl-34073016

RESUMEN

The pathogenesis of several neurodegenerative diseases such as Alzheimer's or Huntington's disease has been associated with metabolic dysfunctions caused by imbalances in the brain and cerebral spinal fluid levels of neuroactive metabolites. Kynurenine monooxygenase (KMO) is considered an ideal therapeutic target for the regulation of neuroactive tryptophan metabolites. Despite significant efforts, the known KMO inhibitors lack blood-brain barrier (BBB) permeability and upon the mimicking of the substrate binding mode, are subject to produce reactive oxygen species as a side reaction. The computational drug design is further complicated by the absence of complete crystal structure information for human KMO (hKMO). In the current work, we performed virtual screening of readily available compounds using several protein-ligand complex pharmacophores. Each of the pharmacophores accounts for one of three distinct reported KMO protein-inhibitor binding conformations. As a result, six novel KMO inhibitors were discovered based on an in vitro fluorescence assay. Compounds VS1 and VS6 were predicted to be BBB permeable and avoid the hydrogen peroxide production dilemma, making them valuable, novel hit compounds for further drug property optimization and advancement in the drug design pipeline.


Asunto(s)
Quinurenina 3-Monooxigenasa/antagonistas & inhibidores , Quinurenina 3-Monooxigenasa/metabolismo , Barrera Hematoencefálica/metabolismo , Encéfalo/metabolismo , Biología Computacional/métodos , Diseño de Fármacos , Evaluación Preclínica de Medicamentos/métodos , Inhibidores Enzimáticos/farmacología , Humanos , Quinurenina/metabolismo , Quinurenina 3-Monooxigenasa/química , Simulación del Acoplamiento Molecular/métodos , Enfermedades Neurodegenerativas/tratamiento farmacológico , Conformación Proteica
3.
Metab Eng ; 57: 193-202, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31786244

RESUMEN

Monophosphoryl lipid A (MPLA) species, including MPL (a trade name of GlaxoSmithKline) and GLA (a trade name of Immune Design, a subsidiary of Merck), are widely used as an adjuvant in vaccines, allergy drugs, and immunotherapy to boost the immune response. Even though MPLA is a derivative of lipopolysaccharide (LPS), a component of the outer membrane of Gram-negative bacteria, bacterial strains producing MPLA have not been found in nature nor engineered. In fact, MPLA generation involves expensive and laborious procedures based on synthetic routes or chemical transformation of precursors isolated from Gram-negative bacteria. Here, we report the engineering of an Escherichia coli strain for in situ production and accumulation of MPLA. Furthermore, we establish a succinct method for purifying MPLA from the engineered E. coli strain. We show that the purified MPLA (named EcML) stimulates the mouse immune system to generate antigen-specific IgG antibodies similarly to commercially available MPLA, but with a dramatically reduced manufacturing time and cost. Our system, employing the first engineered E. coli strain that directly produces the adjuvant EcML, could transform the current standard of industrial MPLA production.


Asunto(s)
Adyuvantes Inmunológicos , Escherichia coli , Lípido A/análogos & derivados , Ingeniería Metabólica , Adyuvantes Inmunológicos/biosíntesis , Adyuvantes Inmunológicos/genética , Adyuvantes Inmunológicos/aislamiento & purificación , Adyuvantes Inmunológicos/farmacología , Animales , Formación de Anticuerpos/efectos de los fármacos , Escherichia coli/genética , Escherichia coli/metabolismo , Inmunoglobulina G/biosíntesis , Lípido A/biosíntesis , Lípido A/genética , Lípido A/aislamiento & purificación , Lípido A/farmacología , Ratones , Ratones Endogámicos BALB C
4.
Bioorg Med Chem Lett ; 30(4): 126882, 2020 02 15.
Artículo en Inglés | MEDLINE | ID: mdl-31889666

RESUMEN

Development of highly effective, safe, and fast-acting anti-depressants is urgently required for the treatment of major depressive disorder. It has been suggested that targeting 5-HT2A and 5-HT2C in addition to inhibition of serotonin reuptake may be beneficial in generating anti-depressant agents with better pharmacology and less adverse effects. We have developed phthalazinone-based compounds that potently bind to 5-HT2A, 5-HT2C, and the serotonin transporter. The representative compounds 11j and 11l displayed strong binding affinities against these targets, and showed favorable toxicity profiles as determined by hERG binding and CYP inhibition assays. Furthermore, these compounds presented promising anti-depressant effects comparable to fluoxetine and also synergistic effects with fluoxetine in forced swimming test, which implicates these compounds can be developed to help the treatment of major depressive disorder.


Asunto(s)
Antidepresivos/química , Azoles/química , Receptor de Serotonina 5-HT2A/química , Receptor de Serotonina 5-HT2C/química , Proteínas de Transporte de Serotonina en la Membrana Plasmática/química , Animales , Antidepresivos/metabolismo , Antidepresivos/farmacología , Conducta Animal/efectos de los fármacos , Diseño de Fármacos , Fluoxetina/química , Fluoxetina/farmacología , Humanos , Concentración 50 Inhibidora , Ratones , Receptor de Serotonina 5-HT2A/metabolismo , Receptor de Serotonina 5-HT2C/metabolismo , Antagonistas del Receptor de Serotonina 5-HT2/química , Antagonistas del Receptor de Serotonina 5-HT2/metabolismo , Antagonistas del Receptor de Serotonina 5-HT2/farmacología , Proteínas de Transporte de Serotonina en la Membrana Plasmática/metabolismo , Inhibidores Selectivos de la Recaptación de Serotonina/química , Inhibidores Selectivos de la Recaptación de Serotonina/metabolismo , Inhibidores Selectivos de la Recaptación de Serotonina/farmacología , Relación Estructura-Actividad
5.
Proc Natl Acad Sci U S A ; 114(13): 3497-3502, 2017 03 28.
Artículo en Inglés | MEDLINE | ID: mdl-28292893

RESUMEN

Stauprimide is a staurosporine analog that promotes embryonic stem cell (ESC) differentiation by inhibiting nuclear localization of the MYC transcription factor NME2, which in turn results in down-regulation of MYC transcription. Given the critical role the oncogene MYC plays in tumor initiation and maintenance, we explored the potential of stauprimide as an anticancer agent. Here we report that stauprimide suppresses MYC transcription in cancer cell lines derived from distinct tissues. Using renal cancer cells, we confirmed that stauprimide inhibits NME2 nuclear localization. Gene expression analysis also confirmed the selective down-regulation of MYC target genes by stauprimide. Consistent with this activity, administration of stauprimide inhibited tumor growth in rodent xenograft models. Our study provides a unique strategy for selectively targeting MYC transcription by pharmacological means as a potential treatment for MYC-dependent tumors.


Asunto(s)
Neoplasias/tratamiento farmacológico , Neoplasias/genética , Proteínas Proto-Oncogénicas c-myc/genética , Bibliotecas de Moléculas Pequeñas/administración & dosificación , Animales , Línea Celular Tumoral , Regulación hacia Abajo/efectos de los fármacos , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Ratones Endogámicos NOD , Ratones SCID , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido Difosfato Quinasas NM23/metabolismo , Neoplasias/metabolismo , Proteínas Proto-Oncogénicas c-myc/metabolismo
6.
Bioorg Med Chem Lett ; 29(10): 1168-1172, 2019 05 15.
Artículo en Inglés | MEDLINE | ID: mdl-30928197

RESUMEN

Highly effective and safe drugs for the treatment of neuropathic pain are urgently required and it was shown that blocking T-type calcium channels can be a promising strategy for drug development for neuropathic pain. We have developed pyrrolidine-based T-type calcium channel inhibitors by structural hybridization and subsequent assessment of in vitro activities against Cav3.1 and Cav3.2 channels. Profiling of in vitro ADME properties of compounds was also carried out. The representative compound 17h showed comparable in vivo efficacy to gabapentin in the SNL model, which indicates T-type calcium channel inhibitors can be developed as effective therapeutics for neuropathic pain.


Asunto(s)
Analgésicos/química , Bloqueadores de los Canales de Calcio/química , Canales de Calcio Tipo T/metabolismo , Neuralgia/tratamiento farmacológico , Pirrolidinas/química , Analgésicos/farmacología , Animales , Bloqueadores de los Canales de Calcio/farmacología , Señalización del Calcio/efectos de los fármacos , Modelos Animales de Enfermedad , Gabapentina/metabolismo , Ganglios Espinales/efectos de los fármacos , Humanos , Ligadura , Microsomas Hepáticos/efectos de los fármacos , Estructura Molecular , Pirrolidinas/farmacología , Ratas , Relación Estructura-Actividad
7.
Molecules ; 24(22)2019 Nov 12.
Artículo en Inglés | MEDLINE | ID: mdl-31726716

RESUMEN

In this study, we investigate the atomistic details of Keap1-Nrf2 inhibitors by in-depth modeling techniques, including molecular dynamics (MD) simulations, and the path-based free energy method of umbrella sampling (US). The protein-protein interaction (PPI) of Keap1-Nrf2 is implicated in several neurodegenerative diseases like cancer, diabetes, and cardiomyopathy. A better understanding of the five sub-pocket binding sites for Nrf2 (ETGE and DLG motifs) inside the Kelch domain would expedite the inhibitor design process. We selected four protein-ligand complexes with distinct co-crystal ligands and binding occupancies inside the Nrf2 binding site. We performed 100 ns of MD simulation for each complex and analyzed the trajectories. From the results, it is evident that one ligand (1VV) has flipped inside the binding pocket, whereas the remaining three were stable. We found that Coulombic (Arg483, Arg415, Ser363, Ser508, and Ser602) and Lennard-Jones (Tyr525, Tyr334, and Tyr572) interactions played a significant role in complex stability. The obtained binding free energy values from US simulations were consistent with the potencies of simulated ligands. US simulation highlight the importance of basic and aromatic residues in the binding pocket. A detailed description of the dissociation process brings valuable insight into the interaction of the four selected protein-ligand complexes, which could help in the future to design more potent PPI inhibitors.


Asunto(s)
Descubrimiento de Drogas , Proteína 1 Asociada A ECH Tipo Kelch/química , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Factor 2 Relacionado con NF-E2/química , Unión Proteica/efectos de los fármacos , Sitios de Unión , Descubrimiento de Drogas/métodos , Enlace de Hidrógeno , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Ligandos , Estructura Molecular , Factor 2 Relacionado con NF-E2/metabolismo , Relación Estructura-Actividad Cuantitativa , Relación Estructura-Actividad
8.
J Enzyme Inhib Med Chem ; 33(1): 1460-1471, 2018 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-30231778

RESUMEN

The treatment of neuropathic pain is one of the urgent unmet medical needs and T-type calcium channels are promising therapeutic targets for neuropathic pain. Several potent T-type channel inhibitors showed promising in vivo efficacy in neuropathic pain animal models and are being investigated in clinical trials. Herein we report development of novel pyrrolidine-based T-type calcium channel inhibitors by pharmacophore mapping and structural hybridisation followed by evaluation of their Cav3.1 and Cav3.2 channel inhibitory activities. Among potent inhibitors against both Cav3.1 and Cav3.2 channels, a promising compound 20n based on in vitro ADME properties displayed satisfactory plasma and brain exposure in rats according to in vivo pharmacokinetic studies. We further demonstrated that 20n effectively improved the symptoms of neuropathic pain in both SNL and STZ neuropathic pain animal models, suggesting modulation of T-type calcium channels can be a promising therapeutic strategy for the treatment of neuropathic pain.


Asunto(s)
Bloqueadores de los Canales de Calcio/farmacología , Canales de Calcio Tipo T/metabolismo , Neuralgia/tratamiento farmacológico , Pirrolidinas/farmacología , Animales , Bloqueadores de los Canales de Calcio/síntesis química , Bloqueadores de los Canales de Calcio/química , Modelos Animales de Enfermedad , Células HEK293 , Humanos , Ligadura , Masculino , Ratones , Ratones Noqueados , Estructura Molecular , Neuralgia/inducido químicamente , Neuralgia/metabolismo , Pirrolidinas/síntesis química , Pirrolidinas/química , Ratas , Ratas Sprague-Dawley , Nervios Espinales/cirugía , Estreptozocina
9.
J Comput Aided Mol Des ; 31(10): 929-941, 2017 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-28913661

RESUMEN

Cyclophilin D (CypD) is a mitochondria-specific cyclophilin that is known to play a pivotal role in the formation of the mitochondrial permeability transition pore (mPTP).The formation and opening of the mPTP disrupt mitochondrial homeostasis, cause mitochondrial dysfunction and eventually lead to cell death. Several recent studies have found that CypD promotes the formation of the mPTP upon binding to ß amyloid (Aß) peptides inside brain mitochondria, suggesting that neuronal CypD has a potential to be a promising therapeutic target for Alzheimer's disease (AD). In this study, we generated an energy-based pharmacophore model by using the crystal structure of CypD-cyclosporine A (CsA) complex and performed virtual screening of ChemDiv database, which yielded forty-five potential hit compounds with novel scaffolds. We further tested those compounds using mitochondrial functional assays in neuronal cells and identified fifteen compounds with excellent protective effects against Aß-induced mitochondrial dysfunction. To validate whether these effects derived from binding to CypD, we performed surface plasmon resonance (SPR)-based direct binding assays with selected compounds and discovered compound 29 was found to have the equilibrium dissociation constants (KD) value of 88.2 nM. This binding affinity value and biological activity correspond well with our predicted binding mode. We believe that this study offers new insights into the rational design of small molecule CypD inhibitors, and provides a promising lead for future therapeutic development.


Asunto(s)
Ciclofilinas/antagonistas & inhibidores , Ciclosporina/química , Mitocondrias/efectos de los fármacos , Fármacos Neuroprotectores/química , Péptidos beta-Amiloides/química , Animales , Sitios de Unión , Supervivencia Celular , Peptidil-Prolil Isomerasa F , Ciclofilinas/metabolismo , Ciclosporina/farmacología , Bases de Datos Farmacéuticas , Células HT29 , Humanos , Mitocondrias/metabolismo , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Poro de Transición de la Permeabilidad Mitocondrial , Simulación del Acoplamiento Molecular , Fármacos Neuroprotectores/farmacología , Unión Proteica , Conformación Proteica , Relación Estructura-Actividad
10.
Proc Natl Acad Sci U S A ; 111(24): 8895-900, 2014 Jun 17.
Artículo en Inglés | MEDLINE | ID: mdl-24889603

RESUMEN

Directly targeting oncogenic V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (K-Ras) with small-molecule inhibitors has historically been considered prohibitively challenging. Recent reports of compounds that bind directly to the K-Ras G12C mutant suggest avenues to overcome key obstacles that stand in the way of developing such compounds. We aim to target the guanine nucleotide (GN)-binding pocket because the natural contents of this pocket dictate the signaling state of K-Ras. Here, we characterize the irreversible inhibitor SML-8-73-1 (SML), which targets the GN-binding pocket of K-Ras G12C. We report a high-resolution X-ray crystal structure of G12C K-Ras bound to SML, revealing that the compound binds in a manner similar to GDP, forming a covalent linkage with Cys-12. The resulting conformation renders K-Ras in the open, inactive conformation, which is not predicted to associate productively with or activate downstream effectors. Conservation analysis of the Ras family GN-binding pocket reveals variability in the side chains surrounding the active site and adjacent regions, especially in the switch I region. This variability may enable building specificity into new iterations of Ras and other GTPase inhibitors. High-resolution in situ chemical proteomic profiling of SML confirms that SML effectively discriminates between K-Ras G12C and other cellular GTP-binding proteins. A biochemical assay provides additional evidence that SML is able to compete with millimolar concentrations of GTP and GDP for the GN-binding site.


Asunto(s)
Acetamidas/química , Genes ras , Guanosina Difosfato/análogos & derivados , Proteínas ras/antagonistas & inhibidores , Proteínas ras/química , Sitios de Unión , Biotina/química , Dominio Catalítico , Secuencia Conservada , Cristalografía por Rayos X , GTP Fosfohidrolasas/química , Proteínas de Unión al GTP/química , Guanosina Difosfato/química , Guanosina Trifosfato/química , Humanos , Ligandos , Modelos Moleculares , Mutación , Fosfatidilinositol 3-Quinasas/química , Unión Proteica , Conformación Proteica , Proteómica , Transducción de Señal
11.
Angew Chem Int Ed Engl ; 56(51): 16262-16266, 2017 12 18.
Artículo en Inglés | MEDLINE | ID: mdl-29105973

RESUMEN

Regioselective C4-, C5-, and di-alkenylations of pyrazoles were achieved. An electrophilic Pd catalyst generated by trifluoroacetic acid (TFA) and 4,5-diazafluoren-9-one (DAF) leads to C4-alkenylation, whereas KOAc and mono-protected amino acid (MPAA) ligand Ac-Val-OH give C5-alkenylation. A combination of palladium acetate, silver carbonate, and pivalic acid affords dialkenylation products. Annulation through sequential alkenylation, thermal 6π-electrocyclization, and oxidation gives functionalized indazoles. This comprehensive strategy greatly expands the range of readily accessible pyrazole and indazole derivatives, enabling useful regiodivergent C-H functionalization of pyrazoles and other heteroaromatic systems.

12.
Nat Chem Biol ; 10(12): 1006-12, 2014 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-25326665

RESUMEN

Her3 (also known as ErbB3) belongs to the epidermal growth factor receptor tyrosine kinases and is well credentialed as an anti-cancer target but is thought to be 'undruggable' using ATP-competitive small molecules because it lacks appreciable kinase activity. Here we report what is to our knowledge the first selective Her3 ligand, TX1-85-1, that forms a covalent bond with Cys721 located in the ATP-binding site of Her3. We demonstrate that covalent modification of Her3 inhibits Her3 signaling but not proliferation in some Her3-dependent cancer cell lines. Subsequent derivatization with a hydrophobic adamantane moiety demonstrates that the resultant bivalent ligand (TX2-121-1) enhances inhibition of Her3-dependent signaling. Treatment of cells with TX2-121-1 results in partial degradation of Her3 and serendipitously interferes with productive heterodimerization between Her3 with either Her2 or c-Met. These results suggest that small molecules will be capable of perturbing the biological function of Her3 and ∼60 other pseudokinases found in human cells.


Asunto(s)
Acrilamidas/farmacología , Adenina/análogos & derivados , Antineoplásicos/farmacología , Regulación Neoplásica de la Expresión Génica , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-met/química , Receptor ErbB-2/química , Receptor ErbB-3/antagonistas & inhibidores , Acrilamidas/síntesis química , Adamantano/química , Adenina/síntesis química , Adenina/farmacología , Adenosina Trifosfato/química , Adenosina Trifosfato/metabolismo , Antineoplásicos/síntesis química , Dominio Catalítico , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cisteína/química , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Simulación del Acoplamiento Molecular , Terapia Molecular Dirigida , Inhibidores de Proteínas Quinasas/síntesis química , Multimerización de Proteína , Proteolisis , Proteínas Proto-Oncogénicas c-met/genética , Proteínas Proto-Oncogénicas c-met/metabolismo , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Receptor ErbB-3/química , Receptor ErbB-3/genética , Transducción de Señal
13.
Bioprocess Biosyst Eng ; 39(5): 713-23, 2016 May.
Artículo en Inglés | MEDLINE | ID: mdl-26857371

RESUMEN

Culturing microalgae in the ocean has potentials that may reduce the production cost and provide an option for an economic biofuel production from microalgae. The ocean holds great potentials for mass microalgal cultivation with its high specific heat, mixing energy from waves, and large cultivable area. Suitable photobioreactors (PBRs) that are capable of integrating marine energy into the culture systems need to be developed for the successful ocean cultivation. In this study, prototype floating PBRs were designed and constructed using transparent low-density polyethylene film for microalgal culture in the ocean. To improve the mixing efficiency, various types of internal partitions were introduced within PBRs. Three different types of internal partitions were evaluated for their effects on the mixing efficiency in terms of mass transfer (k(L)a) and mixing time in the PBRs. The partition type with the best mixing efficiency was selected, and the number of partitions was varied from one to three for investigation of its effect on mixing efficiency. When the number of partitions is increased, mass transfer increased in proportion to the number of partitions. However, mixing time was not directly related to the number of partitions. When a green microalga, Tetraselmis sp. was cultivated using PBRs with the selected partition under semi-continuous mode in the ocean, biomass and fatty acid productivities in the PBRs were increased by up to 50 % and 44% at high initial cell density, respectively, compared to non-partitioned ones. The results of internally partitioned PBRs demonstrated potentials for culturing microalgae by efficiently utilizing ocean wave energy into culture mixing in the ocean.


Asunto(s)
Microalgas/metabolismo , Océanos y Mares , Fotobiorreactores , Ácidos Grasos/metabolismo , Biología Marina , Microalgas/crecimiento & desarrollo
15.
Glycoconj J ; 32(3-4): 161-72, 2015 May.
Artículo en Inglés | MEDLINE | ID: mdl-25971702

RESUMEN

Human cytotoxic T-lymphocyte antigen 4-immunoglobulin (hCTLA4Ig) is an immunosuppressive therapeutic, and recently produced rice cell-derived hCTLA4Ig (hCTLA4Ig(P)) reportedly exhibits in vitro immunosuppressive activities equivalent to those of Chinese hamster ovary cell-derived hCTLA4Ig (hCTLA4Ig(M)). However, limitations of hCTLA4Ig(P) include shortened in vivo half-life as well as the presence of nonhuman N-glycans containing (ß1-2)-xylose and α1,3-fucose, which cause immunogenic reactions in humans. In the present study, human ß1,4-galactose-extended hCTLA4Ig(P) (hCTLA4Ig(P)-Gal) was expressed through the coexpression of human ß1,4-galactosyltransferase (hGalT) and hCTLA4Ig in an attempt to overcome these unfavorable effects. The results indicated that both encoding hGalT and hCTLA4Ig were successfully coexpressed, and the analysis of N-glycan and its relative abundance in purified hCTLA4Ig(P)-Gal indicated that not only were the two glycans containing (ß1-4)-galactose newly extended, but also glycans containing both ß1,2-xylose and α1,3-fucose were markedly reduced and high-mannose-type glycans were increased compared to those of hCTLA4Ig(P), respectively. Unlike hCTLA4Ig(P), hCTLA4Ig(P)-Gal was effective as an acceptor via (ß1-4)-galactose for in vitro sialylation. Additionally, the serum half-life of intravenously injected hCTLA4Ig(P)-Gal in Sprague-Dawley rats was 1.9 times longer than that of hCTLA4Ig(P), and the clearance pattern of hCTLA4Ig(P)-Gal was close to that for hCTLA4Ig(M). These results indicate that the coexpression with hGalT and hCTLA4Ig(P) is useful for both reducing glycan immunogens and increasing in vivo stability. This is the first report of hCTLA4Ig as an effective therapeutics candidate in glycoengineered rice cells.


Asunto(s)
Abatacept/química , Galactosiltransferasas/genética , Inmunosupresores/farmacocinética , Oryza/genética , Polisacáridos/química , Abatacept/sangre , Animales , Células CHO , Secuencia de Carbohidratos , Técnicas de Cultivo de Célula/métodos , Cricetulus , Galactosiltransferasas/metabolismo , Semivida , Humanos , Inmunosupresores/sangre , Masculino , Datos de Secuencia Molecular , Oryza/citología , Plantas Modificadas Genéticamente , Ratas Sprague-Dawley , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo
16.
Bioorg Med Chem Lett ; 25(22): 5186-9, 2015 Nov 15.
Artículo en Inglés | MEDLINE | ID: mdl-26442778

RESUMEN

Enzymatic kinase assays and docking simulation studies have shown that the natural product wrightiadione displays inhibitory activity toward TrkA and PLK3. In this study, the template of wrightiadione served as a starting point for Trk inhibitor development campaigns. Molecular simulation provided structural insights for the design of derivatives that were efficiently generated by our recently developed 3-step tandem synthetic approach, resulting in the discovery of compound 2h with biochemical potency at the single-digit micromolar level.


Asunto(s)
Antineoplásicos Fitogénicos/química , Indenos/química , Isoflavonas/química , Inhibidores de Proteínas Quinasas/química , Quinolinas/química , Receptor trkA/antagonistas & inhibidores , Humanos , Simulación del Acoplamiento Molecular , Receptor trkA/química
17.
Bioorg Med Chem Lett ; 25(16): 3382-9, 2015 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-26094118

RESUMEN

Her3 is a member of the human epidermal growth factor receptor (EGFR) tyrosine kinase family, and it is often either overexpressed or deregulated in many types of human cancer. Her3 has not been the subject of small-molecule inhibitor development because it is a pseudokinase and does not possess appreciable kinase activity. We recently reported on the development of the first selective irreversible Her3 ligand (TX1-85-1) that forms a covalent bond with cysteine 721 which is unique to Her3 among all kinases. We also developed a bi-functional compound (TX2-121-1) containing a hydrophobic adamantane moiety and the same warhead of TX1-85-1 that is capable of inhibiting Her3-dependent signaling and growth. Here we report on the structure-based medicinal chemistry effort that resulted in the discovery of these two compounds.


Asunto(s)
Acrilamidas/síntesis química , Acrilamidas/farmacología , Adenina/análogos & derivados , Sistemas de Liberación de Medicamentos , Pirazoles/síntesis química , Pirazoles/farmacología , Pirimidinas/síntesis química , Pirimidinas/farmacología , Receptor ErbB-3/antagonistas & inhibidores , Acrilamidas/química , Adenina/síntesis química , Adenina/química , Adenina/farmacología , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Concentración 50 Inhibidora , Estructura Molecular , Pirazoles/química , Pirimidinas/química , Bibliotecas de Moléculas Pequeñas/síntesis química , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología
18.
Angew Chem Int Ed Engl ; 53(1): 199-204, 2014 Jan 03.
Artículo en Inglés | MEDLINE | ID: mdl-24259466

RESUMEN

We report the synthesis of a GDP analogue, SML-8-73-1, and a prodrug derivative, SML-10-70-1, which are selective, direct-acting covalent inhibitors of the K-Ras G12C mutant relative to wild-type Ras. Biochemical and biophysical measurements suggest that modification of K-Ras with SML-8-73-1 renders the protein in an inactive state. These first-in-class covalent K-Ras inhibitors demonstrate that irreversible targeting of the K-Ras guanine-nucleotide binding site is potentially a viable therapeutic strategy for inhibition of Ras signaling.


Asunto(s)
Dominio Catalítico/genética , Proteínas ras/química , Proteínas ras/genética , Diseño de Fármacos , Transducción de Señal , Proteínas ras/metabolismo
19.
J Med Chem ; 66(15): 10381-10412, 2023 08 10.
Artículo en Inglés | MEDLINE | ID: mdl-37489798

RESUMEN

Because of the wide use of Fingolimod for the treatment of multiple sclerosis (MS) and its cardiovascular side effects such as bradycardia, second-generation sphingosine 1-phosphate receptor 1 (S1P1) agonist drugs for MS have been developed and approved by FDA. The issue of bradycardia is still present with the new drugs, however, which necessitates further exploration of S1P1 agonists with improved safety profiles for next-generation MS drugs. Herein, we report a tetrahydroisoquinoline or a benzo[c]azepine core-based S1P1 agonists such as 32 and 60 after systematic examination of hydrophilic groups and cores. We investigated the binding modes of our representative compounds and their molecular interactions with S1P1 employing recent S1P1 cryo-EM structures. Also, favorable ADME properties of our compounds were shown. Furthermore, in vivo efficacy of our compounds was clearly demonstrated with PLC and EAE studies. Also, the preliminary in vitro cardiovascular safety of our compound was verified with human iPSC-derived cardiomyocytes.


Asunto(s)
Esclerosis Múltiple , Tetrahidroisoquinolinas , Humanos , Esclerosis Múltiple/tratamiento farmacológico , Esclerosis Múltiple/metabolismo , Receptores de Esfingosina-1-Fosfato , Bradicardia/inducido químicamente , Receptores de Lisoesfingolípidos/agonistas , Receptores de Lisoesfingolípidos/metabolismo , Receptores de Lisoesfingolípidos/uso terapéutico , Clorhidrato de Fingolimod/farmacología , Clorhidrato de Fingolimod/uso terapéutico , Tetrahidroisoquinolinas/uso terapéutico , Esfingosina/metabolismo
20.
Exp Mol Med ; 55(3): 612-627, 2023 03.
Artículo en Inglés | MEDLINE | ID: mdl-36914856

RESUMEN

Tau oligomers play critical roles in tau pathology and are responsible for neuronal cell death and transmitting the disease in the brain. Accordingly, preventing tau oligomerization has become an important therapeutic strategy to treat tauopathies, including Alzheimer's disease. However, progress has been slow because detecting tau oligomers in the cellular context is difficult. Working toward tau-targeted drug discovery, our group has developed a tau-BiFC platform to monitor and quantify tau oligomerization. By using the tau-BiFC platform, we screened libraries with FDA-approved and passed phase I drugs and identified levosimendan as a potent anti-tau agent that inhibits tau oligomerization. 14C-isotope labeling of levosimendan revealed that levosimendan covalently bound to tau cysteines, directly inhibiting disulfide-linked tau oligomerization. In addition, levosimendan disassembles tau oligomers into monomers, rescuing neurons from aggregation states. In comparison, the well-known anti-tau agents methylene blue and LMTM failed to protect neurons from tau-mediated toxicity, generating high-molecular-weight tau oligomers. Levosimendan displayed robust potency against tau oligomerization and rescued cognitive declines induced by tauopathy in the TauP301L-BiFC mouse model. Our data present the potential of levosimendan as a disease-modifying drug for tauopathies.


Asunto(s)
Enfermedad de Alzheimer , Tauopatías , Ratones , Animales , Proteínas tau/metabolismo , Simendán/farmacología , Simendán/uso terapéutico , Simendán/metabolismo , Tauopatías/tratamiento farmacológico , Tauopatías/metabolismo , Tauopatías/patología , Enfermedad de Alzheimer/metabolismo , Neuronas/metabolismo , Modelos Animales de Enfermedad , Ratones Transgénicos
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