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1.
J Toxicol Environ Health A ; 84(20): 846-857, 2021 10 18.
Artículo en Inglés | MEDLINE | ID: mdl-34196262

RESUMEN

The aim of this study was to examine the potential effects of long-term evolution (LTE) radiofrequency electromagnetic fields (RF-EMF) on cell proliferation using SH-SY5Y neuronal cells. The growth rate and proliferation of SH-SY5Y cells were significantly decreased upon exposure to 1760 MHz RF-EMF at 4 W/kg specific absorption rate (SAR) for 4 hr/day for 4 days. Cell cycle analysis indicated that the cell cycle was delayed in the G0/G1 phase after RF-EMF exposure. However, DNA damage or apoptosis was not involved in the reduced cellular proliferation following RF-EMF exposure because the expression levels of histone H2A.X at Ser139 (γH2AX) were not markedly altered and the apoptotic pathway was not activated. However, SH-SY5Y cells exposed to RF-EMF exhibited a significant elevation in Akt and mTOR phosphorylation levels. In addition, the total amount of p53 and phosphorylated-p53 was significantly increased. Data suggested that Akt/mTOR-mediated cellular senescence led to p53 activation via stimulation of the mTOR pathway in SH-SY5Y cells. The transcriptional activation of p53 led to a rise in expression of cyclin-dependent kinase (CDK) inhibitors p21 and p27. Further, subsequent inhibition of CDK2 and CDK4 produced a fall in phosphorylated retinoblastoma (pRb at Ser807/811), which decreased cell proliferation. Taken together, these data suggest that exposure to RF-EMF might induce Akt/mTOR-mediated cellular senescence, which may delay the cell cycle without triggering DNA damage in SH-SY5Y neuroblastoma cells.


Asunto(s)
Proliferación Celular/efectos de la radiación , Senescencia Celular/efectos de la radiación , Campos Electromagnéticos/efectos adversos , Neuroblastoma/fisiopatología , Ondas de Radio/efectos adversos , Senescencia Celular/genética , Humanos , Neuroblastoma/etiología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismo
2.
Front Immunol ; 12: 686462, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34276672

RESUMEN

Immune homeostasis is disturbed during severe viral infections, which can lead to loss of tolerance to self-peptides and result in short- or long-term autoimmunity. Using publicly available transcriptomic datasets, we conducted an in-silico analyses to evaluate the expression levels of 52 autoantigens, known to be associated with 24 autoimmune diseases, during SAR-CoV-2 infection. Seven autoantigens (MPO, PRTN3, PADI4, IFIH1, TRIM21, PTPRN2, and TSHR) were upregulated in whole blood samples. MPO and TSHR were overexpressed in both lung autopsies and whole blood tissue and were associated with more severe COVID-19. Neutrophil activation derived autoantigens (MPO, PRTN3, and PADI4) were prominently increased in blood of both SARS-CoV-1 and SARS-CoV-2 viral infections, while TSHR and PTPRN2 autoantigens were specifically increased in SARS-CoV-2. Using single-cell dataset from peripheral blood mononuclear cells (PBMCs), we observed an upregulation of MPO, PRTN3, and PADI4 autoantigens within the low-density neutrophil subset. To validate our in-silico analysis, we measured plasma protein levels of two autoantigens, MPO and PRTN3, in severe and asymptomatic COVID-19. The protein levels of these two autoantigens were significantly upregulated in more severe COVID-19 infections. In conclusion, the immunopathology and severity of COVID-19 could result in transient autoimmune activation. Longitudinal follow-up studies of confirmed cases of COVID-19 could determine the enduring effects of viral infection including development of autoimmune disease.


Asunto(s)
Autoantígenos/genética , Autoinmunidad/genética , COVID-19/inmunología , SARS-CoV-2/inmunología , Transcriptoma , Enfermedades Asintomáticas , Autoantígenos/sangre , Enfermedades Autoinmunes/sangre , Enfermedades Autoinmunes/inmunología , COVID-19/sangre , COVID-19/patología , COVID-19/virología , Simulación por Computador , Bases de Datos Genéticas , Humanos , Pulmón/patología , Mieloblastina/sangre , Mieloblastina/genética , Activación Neutrófila , Neutrófilos/inmunología , Peroxidasa/sangre , Peroxidasa/genética , RNA-Seq , Índice de Severidad de la Enfermedad , Regulación hacia Arriba/genética
3.
Nature ; 596(7871): 281-284, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-34290409

RESUMEN

The mTOR complex 1 (mTORC1) controls cell growth in response to amino acid levels1. Here we report SAR1B as a leucine sensor that regulates mTORC1 signalling in response to intracellular levels of leucine. Under conditions of leucine deficiency, SAR1B inhibits mTORC1 by physically targeting its activator GATOR2. In conditions of leucine sufficiency, SAR1B binds to leucine, undergoes a conformational change and dissociates from GATOR2, which results in mTORC1 activation. SAR1B-GATOR2-mTORC1 signalling is conserved in nematodes and has a role in the regulation of lifespan. Bioinformatic analysis reveals that SAR1B deficiency correlates with the development of lung cancer. The silencing of SAR1B and its paralogue SAR1A promotes mTORC1-dependent growth of lung tumours in mice. Our results reveal that SAR1B is a conserved leucine sensor that has a potential role in the development of lung cancer.


Asunto(s)
Leucina/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Proteínas de Unión al GTP Monoméricas/metabolismo , Transducción de Señal , Animales , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiología , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Secuencia Conservada , GTP Fosfohidrolasas/genética , GTP Fosfohidrolasas/metabolismo , Células HEK293 , Humanos , Leucina/deficiencia , Longevidad/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/agonistas , Ratones , Proteínas de Unión al GTP Monoméricas/química , Proteínas de Unión al GTP Monoméricas/deficiencia , Proteínas de Unión al GTP Monoméricas/genética , Complejos Multiproteicos/metabolismo , Proteínas Nucleares/metabolismo , Unión Proteica , Proteínas Supresoras de Tumor/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto
4.
ACS Infect Dis ; 7(8): 2522-2535, 2021 08 13.
Artículo en Inglés | MEDLINE | ID: mdl-34260210

RESUMEN

Invasive and superficial infections caused by the Candida species result in significant global morbidity and mortality. As the pathogenicity of these organisms is intimately intertwined with host immune response, therapies to target both the fungus and host inflammation may be warranted. Structural similarities exist between established inhibitors of the NLRP3 inflammasome and those of fungal acetohydroxyacid synthase (AHAS). Therefore, we leveraged this information to conduct an in silico molecular docking screen to find novel polypharmacologic inhibitors of these targets that resulted in the identification of 12 candidate molecules. Of these, compound 10 significantly attenuated activation of the NLPR3 inflammasome by LPS + ATP, while also demonstrating growth inhibitory activity against C. albicans that was alleviated in the presence of exogenous branched chain amino acids, consistent with targeting of fungal AHAS. SAR studies delineated an essential molecular scaffold required for dual activity. Ultimately, 10 and its analog 10a resulted in IC50 (IL-1ß release) and MIC50 (fungal growth) values with low µM potency against several Candida species. Collectively, this work demonstrates promising potential of dual-target approaches for improved management of fungal infections.


Asunto(s)
Acetolactato Sintasa , Inflamasomas , Antifúngicos/farmacología , Candida albicans , Simulación del Acoplamiento Molecular
5.
Neurobiol Dis ; 156: 105410, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-34087381

RESUMEN

We have recently demonstrated the role of the Fyn-PKCδ signaling pathway in status epilepticus (SE)-induced neuroinflammation and epileptogenesis in experimental models of temporal lobe epilepsy (TLE). In this study, we show a significant disease-modifying effect and the mechanisms of a Fyn/Src tyrosine kinase inhibitor, saracatinib (SAR, also known as AZD0530), in the rat kainate (KA) model of TLE. SAR treatment for a week, starting the first dose (25 mg/kg, oral) 4 h after the onset of SE, significantly reduced spontaneously recurring seizures and epileptiform spikes during the four months of continuous video-EEG monitoring. Immunohistochemistry of brain sections and Western blot analyses of hippocampal lysates at 8-day (8d) and 4-month post-SE revealed a significant reduction of SE-induced astrogliosis, microgliosis, neurodegeneration, phosphorylated Fyn/Src-419 and PKCδ-tyr311, in SAR-treated group when compared with the vehicle control. We also found the suppression of nitroxidative stress markers such as iNOS, 3-NT, 4-HNE, and gp91phox in the hippocampus, and nitrite and ROS levels in the serum of the SAR-treated group at 8d post-SE. The qRT-PCR (hippocampus) and ELISA (serum) revealed a significant reduction of key proinflammatory cytokines TNFα and IL-1ß mRNA in the hippocampus and their protein levels in serum, in addition to IL-6 and IL-12, in the SAR-treated group at 8d in contrast to the vehicle-treated group. These findings suggest that SAR targets some of the key biomarkers of epileptogenesis and modulates neuroinflammatory and nitroxidative pathways that mediate the development of epilepsy. Therefore, SAR can be developed as a potential disease-modifying agent to prevent the development and progression of TLE.

6.
J Med Chem ; 64(9): 5384-5403, 2021 05 13.
Artículo en Inglés | MEDLINE | ID: mdl-33942619

RESUMEN

Ventricular cardiac arrhythmia (VA) arises in acquired or congenital heart disease. Long QT syndrome type-3 (LQT3) is a congenital form of VA caused by cardiac sodium channel (INaL) SCN5A mutations that prolongs cardiac action potential (AP) and enhances INaL current. Mexiletine inhibits INaL and shortens the QT interval in LQT3 patients. Above therapeutic doses, mexiletine prolongs the cardiac AP. We explored structure-activity relationships (SAR) for AP shortening and prolongation using dynamic medicinal chemistry and AP kinetics in human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Using patient-derived LQT3 and healthy hiPSC-CMs, we resolved distinct SAR for AP shortening and prolongation effects in mexiletine analogues and synthesized new analogues with enhanced potency and selectivity for INaL. This resulted in compounds with decreased AP prolongation effects, increased metabolic stability, increased INaL selectivity, and decreased avidity for the potassium channel. This study highlights using hiPSC-CMs to guide medicinal chemistry and "drug development in a dish".


Asunto(s)
Antiarrítmicos/química , Trastorno del Sistema de Conducción Cardíaco/patología , Síndrome de QT Prolongado/patología , Mexiletine/análogos & derivados , Potenciales de Acción/efectos de los fármacos , Animales , Antiarrítmicos/farmacología , Conducta Animal/efectos de los fármacos , Trastorno del Sistema de Conducción Cardíaco/metabolismo , Células Cultivadas , Diseño de Fármacos , Estabilidad de Medicamentos , Semivida , Humanos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/metabolismo , Síndrome de QT Prolongado/metabolismo , Masculino , Mexiletine/farmacología , Ratones , Ratones Endogámicos BALB C , Miocitos Cardíacos/citología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/fisiología , Canal de Sodio Activado por Voltaje NAV1.5/genética , Canal de Sodio Activado por Voltaje NAV1.5/metabolismo , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad
7.
Bioorg Med Chem Lett ; 43: 128089, 2021 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-33964438

RESUMEN

Several boron-containing small molecules have been approved by the US FDA to treat human diseases. We explored potential applications of boron-containing compounds in modern agriculture by pursuing multiple research and development programs. Here, we report a novel series of multi-substitution benzoxaboroles (1-36), a compound class that we recently reported as targeting geranylgeranyl transferase I (GGTase I) and thereby inhibiting protein prenylation (Kim et al., 2020). These compounds were designed, synthesized, and tested against the agriculturally important fungal pathogens Mycosphaerella fijiensis and Colletotrichum sublineolum in a structure-activity relationship (SAR) study. Compounds 13, 28, 30, 34 and 36 were identified as active leads with excellent antifungal MIC95 values in the range of 1.56-3.13 ppm against M. fijiensis and 0.78-3.13 ppm against C. sublineolum.

8.
Bioorg Chem ; 112: 104989, 2021 07.
Artículo en Inglés | MEDLINE | ID: mdl-34022709

RESUMEN

Eight new jatrophane diterpenoids, Euphosorophane F-M (1-8), as well as fourteen known jatrophane diterpenoids (9-22) were separated and purified from the fructus of Euphorbia sororia, and the chemical structures were determined based on extensive spectroscopic analysis, 1D, 2D NMR and HRESIMS data included. Their absolute configurations of compounds 1, 2, 9, and 22 were elucidated by X-ray crystallographic analysis. These jatrophane diterpenoids showed lower cytotoxicity and compounds 3, 4, 11, 12, 13, 14, and 20 revealed promising multidrug resistance (MDR) reversal ability as modulators compared to verapamil (VRP) by MTT assay. The structure-activity relationship (SAR) exhibited that the absence of keto-carbonyl at C-9 and C-14 was essential to MDR reversal activity and the acyloxies substitution at C-5, C-7, C-8, and C-14 also made the activity difference. Euphosorophane I (4) particularly unfold greater potency (EC50 = 1.82 µM) in reversing P-gp-mediated resistance to doxorubicin (DOX). As shown by fluorescence microscopy, 4 promoted intracellular accumulation of rhodamine 123 (Rh123) and DOX in a dose-dependentmanner than VRP. Flow cytometry indicated that 4 inhibitedP-glycoprotein (P-gp) -dependentRh123 efflux in drug-resistant MCF-7/ADR cells. 4 stimulated P-gp-ATPase activity in a concentration-dependent way and inhibited DOX transport activity. Western blot and real-time qPCR results further illustrated that 4 exhibited superior MDR reversal effect in MCF-7/ADR cells attributed to the activation of ATPase rather than the upregulation of P-gp expression and mRNA levels. In addition, 4 bond to the drug-binding site of P-gp predicted by the molecular docking analysis. Collectively, these results indicated that 4 efficiently reversed P-gp-mediated MDR via inhibiting the ABCB1 drug efflux function. 4 with the advantage of low toxicity and efficient could be used as an adjuvanttherapy drug for breast cancer.

9.
Bioorg Chem ; 113: 104994, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-34052738

RESUMEN

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secreted protein and its deficiency markedly enhanced the survival rate of patient with cardiovascular diseases (CVDs). Forty berberine (BBR) derivatives were synthesized and evaluated for their activities on down-regulating the transcription of PCSK9 in HepG2 cells, taking BBR as the lead. Structure-activity relationship (SAR) analysis revealed that 2,3-dimethoxy moiety might be beneficial for activity. Among them, 9k displayed the most potent activity with IC50 value of 9.5 ± 0.5 µM, better than that of BBR. Also, it significantly decreased PCSK9 protein level at cellular level, as well as in the liver and serum of mice in vivo. Furthermore, 9k markedly increased LDLR expression and LDL-C clearance via down-regulating PCSK9 protein. The mechanism of action of 9k is targeting HNF1α and/or Sp1 cluster modulation upstream of PCSK9, a different one from BBR. Therefore, 9k might have the potential to be a novel PCSK9 transcriptional inhibitor for the treatment of atherosclerosis, worthy for further investigation.

10.
Antimicrob Agents Chemother ; 65(8): e0234920, 2021 07 16.
Artículo en Inglés | MEDLINE | ID: mdl-34001508

RESUMEN

Here, we identified a novel class of compounds which demonstrated good antiviral activity against dengue and Zika virus infection. These derivatives constitute intermediates in the synthesis of indole (ervatamine-silicine) alkaloids and share a tetracyclic structure, with an indole and a piperidine fused to a seven-membered carbocyclic ring. Structure-activity relationship studies indicated the importance of substituent at position C-6 and especially the presence of a benzyl ester for the activity and cytotoxicity of the molecules. In addition, the stereochemistry at C-7 and C-8, as well as the presence of an oxazolidine ring, influenced the potency of the compounds. Mechanism of action studies with two analogues of this family (compounds 22 and trans-14) showed that this class of molecules can suppress viral infection during the later stages of the replication cycle (RNA replication/assembly). Moreover, a cell-dependent antiviral profile of the compounds against several Zika strains was observed, possibly implying the involvement of a cellular factor(s) in the activity of the molecules. Sequencing of compound-resistant Zika mutants revealed a single nonsynonymous amino acid mutation (aspartic acid to histidine) at the beginning of the predicted transmembrane domain 1 of NS4B protein, which plays a vital role in the formation of the viral replication complex. To conclude, our study provides detailed information on a new class of NS4B-associated inhibitors and strengthens the importance of identifying host-virus interactions in order to tackle flavivirus infections.


Asunto(s)
Dengue , Infección por el Virus Zika , Virus Zika , Humanos , Alcaloides Indólicos , Proteínas no Estructurales Virales , Replicación Viral , Infección por el Virus Zika/tratamiento farmacológico
11.
NMR Biomed ; 34(7): e4525, 2021 07.
Artículo en Inglés | MEDLINE | ID: mdl-33955061

RESUMEN

PURPOSE: To investigate inter-subject variability of B1 + , SAR and temperature rise in a database of human models using a local transmit array for 7 T cardiac imaging. METHODS: Dixon images were acquired of 14 subjects and segmented in dielectric models with an eight-channel local transmit array positioned around the torso for cardiac imaging. EM simulations were done to calculate SAR distributions. Based on the SAR distributions, temperature simulations were performed for exposure times of 6 min and 30 min. Peak local SAR and temperature rise levels were calculated for different RF shim settings. A statistical analysis of the resulting peak local SAR and temperature rise levels was performed to arrive at safe power limits. RESULTS: For RF shim vectors with random phase and uniformly distributed power, a safe average power limit of 35.7 W was determined (first level controlled mode). When RF amplitude and phase shimming was performed on the heart, a safe average power limit of 35.0 W was found. According to Pennes' model, our numerical study suggests a very low probability of exceeding the absolute local temperature limit of 40 °C for a total exposure time of 6 min and a peak local SAR of 20 W/kg. For a 30 min exposure time at 20 W/kg, it was shown that the absolute temperature limit can be exceeded in the case where perfusion does not change with temperature. CONCLUSION: Safe power constraints were found for 7 T cardiac imaging with an eight-channel local transmit array, while considering the inter-subject variability of B1 + , SAR and temperature rise.

12.
Int J Radiat Biol ; 97(9): 1316-1323, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34047676

RESUMEN

PURPOSE: To assess the effects of 1800 MHz radiofrequency electromagnetic field (RF-EMF) exposure on the expression of signal transduction and antioxidant proteins in a human-derived A172 glioblastoma cell line. MATERIALS AND METHODS: Adherent human-derived A172 glioblastoma cells (1.0 × 105 cells per 35 mm culture dish, containing 2 mL DMEM media) were exposed to 1800 MHz continuous-wave (CW) or GSM-modulated RF fields, in the presence or absence of serum for 5, 30 or 240 min at a specific absorption rate (SAR) of 0 (sham) or 2.0 W/kg. Concurrent negative (vehicle) and positive controls (1 µg/mL anisomycin) were included in each experiment. Cell lysates were collected immediately after exposure, stabilized by protease and phosphatase inhibitors in lysis buffer, then frozen and maintained at -80 °C until analysis. The relative expression levels of phosphorylated- and total-signal transduction proteins (CREB, JNK, NF-κB, ERK1/2, Akt, p70S6K, STAT3 and STAT5) and antioxidant proteins (SOD1, SOD2, CAT, TRX1, PRX2) were assessed using Milliplex magnetic bead array panels and a MagPix Multiplex imaging system. RESULTS: In cells exposed to 1800 MHz continuous-wave RF-EMF with the presence of serum in the culture medium, CAT expression was statistically significantly decreased after a 30 min exposure, total JNK was decreased at both 30 and 240 min of exposure, STAT3 was decreased after 240 min of exposure and phosphorylated-CREB expression was decreased after 30 min of exposure. In cells exposed to 1800 MHz GSM-modulated RF-EMF in serum-free cultures, the expression level of total STAT5 was decreased after 30 and 240 min of exposure. These observed changes were detected sporadically across time-points, culture conditions and RF-EMF exposure conditions indicating the likelihood of false positive events. When cells were treated with anisomycin for 15 min as a positive control, dramatic increases in the expression of phosphorylated signaling proteins were observed in both serum-starved and serum-fed A172 cells, with larger fold change increases in the serum-free cultures. No statistically significant differences in the expression levels of SOD1, SOD2 or TRX1 were observed under any tested conditions after exposure to RF-EMF. CONCLUSIONS: The current study found no consistent evidence of changes in the expression of antioxidant proteins (SOD1, SOD2, CAT or TRX2) or a variety of signal transductions proteins (CREB, JNK, NF-κB, ERK1/2, Akt, p70S6K, STAT3, STAT5) in a human-derived glioblastoma A172 cell line in response to exposure to 1800 MHz continuous-wave or GSM-modulated RF-EMF for 5, 30 or 240 min in either serum-free or serum-containing cultures.


Asunto(s)
Antioxidantes/metabolismo , Glioblastoma/patología , Fosfoproteínas/metabolismo , Ondas de Radio , Transducción de Señal/efectos de la radiación , Línea Celular Tumoral , Humanos , Fosforilación/efectos de la radiación
13.
J Med Chem ; 64(11): 7275-7295, 2021 06 10.
Artículo en Inglés | MEDLINE | ID: mdl-33881312

RESUMEN

Pathogenic bacteria demonstrate incredible abilities to evade conventional antibiotics through the development of resistance and formation of dormant, surface-attached biofilms. Therefore, agents that target and eradicate planktonic and biofilm bacteria are of significant interest. We explored a new series of halogenated phenazines (HP) through the use of N-aryl-2-nitrosoaniline synthetic intermediates that enabled functionalization of the 3-position of this scaffold. Several HPs demonstrated potent antibacterial and biofilm-killing activities (e.g., HP 29, against methicillin-resistant Staphylococcus aureus: MIC = 0.075 µM; MBEC = 2.35 µM), and transcriptional analysis revealed that HPs 3, 28, and 29 induce rapid iron starvation in MRSA biofilms. Several HPs demonstrated excellent activities against Mycobacterium tuberculosis (HP 34, MIC = 0.80 µM against CDC1551). This work established new SAR insights, and HP 29 demonstrated efficacy in dorsal wound infection models in mice. Encouraged by these findings, we believe that HPs could lead to significant advances in the treatment of challenging infections.


Asunto(s)
Compuestos de Anilina/química , Antibacterianos/síntesis química , Fenazinas/química , Animales , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Biopelículas/efectos de los fármacos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Diseño de Fármacos , Femenino , Halogenación , Humanos , Hierro/química , Hierro/deficiencia , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Staphylococcus aureus Resistente a Meticilina/fisiología , Ratones , Ratones Endogámicos BALB C , Mycobacterium tuberculosis/efectos de los fármacos , Fenazinas/farmacología , Fenazinas/uso terapéutico , Infecciones Estafilocócicas/tratamiento farmacológico , Relación Estructura-Actividad , Cicatrización de Heridas/efectos de los fármacos
14.
J Med Chem ; 64(9): 5404-5428, 2021 05 13.
Artículo en Inglés | MEDLINE | ID: mdl-33844533

RESUMEN

The connection with acute myelogenous leukemia (AML) of dihydroorotate dehydrogenase (hDHODH), a key enzyme in pyrimidine biosynthesis, has attracted significant interest from pharma as a possible AML therapeutic target. We recently discovered compound 1, a potent hDHODH inhibitor (IC50 = 1.2 nM), able to induce myeloid differentiation in AML cell lines (THP1) in the low nM range (EC50 = 32.8 nM) superior to brequinar's phase I/II clinical trial (EC50 = 265 nM). Herein, we investigate the 1 drug-like properties observing good metabolic stability and no toxic profile when administered at doses of 10 and 25 mg/kg every 3 days for 5 weeks (Balb/c mice). Moreover, in order to identify a backup compound, we investigate the SAR of this class of compounds. Inside the series, 17 is characterized by higher potency in inducing myeloid differentiation (EC50 = 17.3 nM), strong proapoptotic properties (EC50 = 20.2 nM), and low cytotoxicity toward non-AML cells (EC30(Jurkat) > 100 µM).


Asunto(s)
Compuestos de Bifenilo/química , Inhibidores Enzimáticos/química , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/antagonistas & inhibidores , Pirazoles/química , Piridinas/química , Animales , Apoptosis/efectos de los fármacos , Sitios de Unión , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Diseño de Fármacos , Inhibidores Enzimáticos/metabolismo , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Femenino , Semivida , Humanos , Leucemia Mieloide Aguda/tratamiento farmacológico , Masculino , Ratones , Ratones Endogámicos BALB C , Microsomas Hepáticos/metabolismo , Simulación del Acoplamiento Molecular , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/metabolismo , Pirazoles/metabolismo , Pirazoles/farmacología , Pirazoles/uso terapéutico , Piridinas/metabolismo , Piridinas/farmacología , Piridinas/uso terapéutico , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad
15.
J Biol Chem ; 296: 100695, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33894200

RESUMEN

Upon pathogen infection, receptors in plants will activate a localized immune response, the effector-triggered immunity (ETI), and a systemic immune response, the systemic acquired response (SAR). Infection also induces oscillations in the redox environment of plant cells, triggering response mechanisms involving sensitive cysteine residues that subsequently alter protein function. Arabidopsis thaliana thimet oligopeptidases TOP1 and TOP2 are required for plant defense against pathogens and the oxidative stress response. Herein, we evaluated the biochemical attributes of TOP isoforms to determine their redox sensitivity using ex vivo Escherichia coli cultures and recombinant proteins. Moreover, we explored the link between their redox regulation and plant immunity in wild-type and mutant Arabidopsis lines. These analyses revealed that redox regulation of TOPs occurs through two mechanisms: (1) oxidative dimerization of full-length TOP1 via intermolecular disulfides engaging cysteines in the N-terminal signal peptide, and (2) oxidative activation of all TOPs via cysteines that are unique and conserved. Further, we detected increased TOP activity in wild-type plants undergoing ETI or SAR following inoculation with Pseudomonas syringae strains. Mutants unable to express the chloroplast NADPH-dependent thioredoxin reductase C (NTRC) showed elevated TOP activity under unstressed conditions and were SAR-incompetent. A top1top2 knockout mutant challenged with P. syringae exhibited misregulation of ROS-induced gene expression in pathogen-inoculated and distal tissues. Furthermore, TOP1 and TOP2 could cleave a peptide derived from the immune component ROC1 with distinct efficiencies at common and specific sites. We propose that Arabidopsis TOPs are thiol-regulated peptidases active in redox-mediated signaling of local and systemic immunity.


Asunto(s)
Arabidopsis/enzimología , Arabidopsis/inmunología , Metaloendopeptidasas/metabolismo , Arabidopsis/citología , Arabidopsis/microbiología , Metaloendopeptidasas/química , Metaloendopeptidasas/genética , Modelos Moleculares , Mutación , Oxidación-Reducción , Conformación Proteica , Señales de Clasificación de Proteína , Pseudomonas syringae/fisiología , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal
16.
Viruses ; 13(3)2021 03 15.
Artículo en Inglés | MEDLINE | ID: mdl-33804121

RESUMEN

Small molecules targeting the PF74 binding site of the HIV-1 capsid protein (CA) confer potent and mechanistically unique antiviral activities. Structural modifications of PF74 could further the understanding of ligand binding modes, diversify ligand chemical classes, and allow identification of new variants with balanced antiviral activity and metabolic stability. In the current work, we designed and synthesized three series of PF74-like analogs featuring conformational constraints at the aniline terminus or the phenylalanine carboxamide moiety, and characterized them using a biophysical thermal shift assay (TSA), cell-based antiviral and cytotoxicity assays, and in vitro metabolic stability assays in human and mouse liver microsomes. These studies showed that the two series with the phenylalanine carboxamide moiety replaced by a pyridine or imidazole ring can provide viable hits. Subsequent SAR identified an improved analog 15 which effectively inhibited HIV-1 (EC50 = 0.31 µM), strongly stabilized CA hexamer (ΔTm = 8.7 °C), and exhibited substantially enhanced metabolic stability (t1/2 = 27 min for 15 vs. 0.7 min for PF74). Metabolic profiles from the microsomal stability assay also indicate that blocking the C5 position of the indole ring could lead to increased resistance to oxidative metabolism.


Asunto(s)
Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/farmacología , Proteínas de la Cápside/metabolismo , VIH-1/efectos de los fármacos , Indoles/metabolismo , Fenilalanina/análogos & derivados , Bibliotecas de Moléculas Pequeñas/metabolismo , Bibliotecas de Moléculas Pequeñas/farmacología , Animales , Fármacos Anti-VIH/aislamiento & purificación , Sitios de Unión , Proteínas de la Cápside/química , Proteínas de la Cápside/genética , Línea Celular , Diseño de Fármacos , Células HEK293 , Humanos , Indoles/farmacología , Hígado/efectos de los fármacos , Ratones , Microsomas/efectos de los fármacos , Modelos Moleculares , Conformación Molecular , Fenilalanina/metabolismo , Fenilalanina/farmacología , Replicación Viral/efectos de los fármacos
17.
IUBMB Life ; 73(6): 855-865, 2021 06.
Artículo en Inglés | MEDLINE | ID: mdl-33724683

RESUMEN

Despite sharing conserved substrate-binding residues, members of 3-hydroxyisobutyrate dehydrogenase (HIBADH) superfamily show remarkable differences in substrate preference. Cysteine residues were identified within a radius of 6 Å surrounding both the active site and the substrate entry site of HIBADH enzyme from Mycobacterium tuberculosis (MtHIBADH). Chemical modification with thiol-modifying reagents, pCMB and DTNB, abrogated the dehydrogenase activity of the enzyme. The loss in activity followed pseudo-first-order kinetics as a function of the concentration of pCMB. S-HIBA (substrate) binding provided partial protection, while NAD (cofactor) binding provided ~70% protection from thiol-modifying reagent. Site-directed mutagenesis of cysteine residues present in the MtHIBADH enzyme identified the indispensable role of Cys-210 residue, located at C-terminal domain, for its dehydrogenase activity. Cys-210 mutation to serine reduced the dehydrogenase activity by ~2-fold while mutation to alanine strikingly reduced the activity by ~140-fold. C210A mutation did not perturb the state of oligomerization of the enzyme but perturbed the secondary structure content. Structural analysis revealed the involvement of Cys-210 residue in inter-chain interaction with Gln-178, which acts as hydrogen bond donor and coordinates with Cys-210 and Gly-208 of the adjacent subunit. The data demonstrate a critical role of Cys-210 residue in maintaining the conformation and rigidity of loop composed of substrate-interacting residues involved in the entry of S-HIBA substrate in MtHIBADH.

18.
J Med Chem ; 64(4): 2077-2109, 2021 02 25.
Artículo en Inglés | MEDLINE | ID: mdl-33538581

RESUMEN

In the present work, 103 novel acyclic nucleosides were designed, synthesized, and evaluated for their anticancer activities in vitro and in vivo. The structure-activity relationship (SAR) studies revealed that most target compounds inhibited the growth of colon cancer cells in vitro, of which 3-(6-chloro-9H-purin-9-yl)dodecan-1-ol (9b) exhibited the most potent effect against the HCT-116 and SW480 cells with IC50 values of 0.89 and 1.15 µM, respectively. Furthermore, all of the (R)-configured acyclic nucleoside derivatives displayed more potent anticancer activity compared to their (S)-counterparts. Mechanistic studies revealed that compound 9b triggered apoptosis in the cancer cell lines via depolarization of the mitochondrial membrane and effectively inhibited colony formation. Importantly, compound 9b inhibited the growth of the SW480 xenograft in a mouse model with low systemic toxicity. These results indicated that acyclic nucleoside compounds are viable as potent and effective anticancer agents, and compound 9b may serve as a promising lead compound that merits further attention in future anticancer drug discovery.


Asunto(s)
Antineoplásicos/uso terapéutico , Neoplasias/tratamiento farmacológico , Nucleósidos de Purina/uso terapéutico , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Diseño de Fármacos , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones Endogámicos BALB C , Mitocondrias/efectos de los fármacos , Estructura Molecular , Nucleósidos de Purina/síntesis química , Nucleósidos de Purina/farmacología , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de Xenoinjerto
19.
Bioorg Chem ; 109: 104699, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33611138

RESUMEN

Aconitine linoleate (11) isolated from the Aconitum sinchiangense W. T. Wang exhibited significant anti-tumor activity. Based on this, a series of novel lipo-diterpenoid alkaloids were synthesized and evaluated for their anticancer activities against MCF-7 and MCF-7/ADR cell lines. Seventeen compounds, including 18-20, 22, 24-32, 36, 39, 41-42 possessed higher anti-proliferative activities (IC50 < 20 µM) against MCF-7 cell lines, which were better than the reference drug etoposide (IC50 = 18.01 ± 1.64 µM), among which compound 24 (IC50 = 4.00 ± 0.30 µM) was found to be the most potent derivative, being 4.5-fold more active than etoposide. Meanwhile, eighteen compounds, including 18-22, 24, 26-32, 36, 38-39, 41-42 presented excellent activities (IC50 < 20 µM) against MCF-7/ADR cell lines, better than etoposide (IC50 = 35.48 ± 0.29 µM) and doxorubicin (IC50 = 67.61 ± 6.5 µM). The most potent compound (19) was 13.5- and 25.7-fold more active than etoposide and doxorubicin against MCF-7/ADR cell lines, respectively. The structure-activity relationship (SAR) studies indicated that the 3-OH, 8-lipo, 14-benzene ring, and nitrogen atom with proper alkaline are crucial elements for anti-proliferative activity of target lipo-diterpenoid compounds. The proper length, the double bonds or di-fluoro-substituted at C-8 fatty acid chain, the para-donating electron group on 14-benzene group, and 13-OH are all favorable for the enhancement of anti-proliferative activities. In conclusion, the introduction of the 8-lipo group into aconitine leads to significant increase of anti-proliferative activity against MCF-7 and MCF-7/ADR cells, which suggests these kinds of lipo-alkaloids are powerful and promising antitumor compounds for breast cancer, especially for drug-resistant breast cancer.

20.
Int J Phytoremediation ; 23(9): 969-981, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33455421

RESUMEN

Salinity is a widespread soil and underground water contaminant threatening food security and economic stability. Phytoremediation is an efficient and environmental-friendly solution to mitigate salinity impacts. The present study was conducted to evaluate the phytoremediation potential of five multipurpose trees: Vachellia nilotica, Concorpus erectus, Syzygium cumini, Tamarix aphylla and Eucalyptus cammaldulensis under four salinity treatments: Control, 10, 20 and 30 dS m-1. Salinity negatively impacted all the tested species. However, E. cammaldulensis and T. aphylla exhibited the lowest reduction (28%) and (35%) in plant height respectively along with a minimal reduction in leaf gas exchange while V. nilotica, S. cumini and C. erectus showed severe dieback. Similarly, the antioxidant enzymes increased significantly in E. cammaldulensis and T. aphylla as Superoxide Dismutase (87% and 79%), Catalase (66% and 67%) and Peroxidase (89% and 81%), respectively. Furthermore, both of these species maintained optimum Na/K ratio reducing the highest levels of soil ECe and SAR, suggesting the best phytoremediation potential. The present study identifies that E. cammaldulensis and T. aphylla showed effective tolerance mechanisms and the highest salt sequestration; therefore, may be used for phyto-amelioration of salinity impacted lands. Novelty statement Although previous studies evaluated the tolerance potential of many tree species, comparative and physiochemical evaluation of multipurpose tree species has been remained unexplored. In this scenario, eco-physiological characterization of multipurpose tree species may inform tree species for phytoremediation of saline soils according to the level of salinity. Optimizing tree species selection also improves the success of wood for energy and revenue generation while restoring degraded soils.


Asunto(s)
Contaminantes del Suelo , Suelo , Biodegradación Ambiental , Salinidad , Árboles
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