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1.
Front Immunol ; 13: 834988, 2022.
Article in English | MEDLINE | ID: covidwho-1753370

ABSTRACT

Patients with COVID-19 present with a wide variety of clinical manifestations. Thromboembolic events constitute a significant cause of morbidity and mortality in patients infected with SARS-CoV-2. Severe COVID-19 has been associated with hyperinflammation and pre-existing cardiovascular disease. Platelets are important mediators and sensors of inflammation and are directly affected by cardiovascular stressors. In this report, we found that platelets from severely ill, hospitalized COVID-19 patients exhibited higher basal levels of activation measured by P-selectin surface expression and had poor functional reserve upon in vitro stimulation. To investigate this question in more detail, we developed an assay to assess the capacity of plasma from COVID-19 patients to activate platelets from healthy donors. Platelet activation was a common feature of plasma from COVID-19 patients and correlated with key measures of clinical outcome including kidney and liver injury, and APACHEIII scores. Further, we identified ferritin as a pivotal clinical marker associated with platelet hyperactivation. The COVID-19 plasma-mediated effect on control platelets was highest for patients that subsequently developed inpatient thrombotic events. Proteomic analysis of plasma from COVID-19 patients identified key mediators of inflammation and cardiovascular disease that positively correlated with in vitro platelet activation. Mechanistically, blocking the signaling of the FcγRIIa-Syk and C5a-C5aR pathways on platelets, using antibody-mediated neutralization, IgG depletion or the Syk inhibitor fostamatinib, reversed this hyperactivity driven by COVID-19 plasma and prevented platelet aggregation in endothelial microfluidic chamber conditions. These data identified these potentially actionable pathways as central for platelet activation and/or vascular complications and clinical outcomes in COVID-19 patients. In conclusion, we reveal a key role of platelet-mediated immunothrombosis in COVID-19 and identify distinct, clinically relevant, targetable signaling pathways that mediate this effect.


Subject(s)
Blood Platelets/immunology , COVID-19/immunology , Complement C5a/metabolism , Receptor, Anaphylatoxin C5a/metabolism , Receptors, IgG/metabolism , SARS-CoV-2/physiology , Thromboembolism/immunology , Adult , Aminopyridines/pharmacology , Cells, Cultured , Female , Hospitalization , Humans , Male , Morpholines/pharmacology , Platelet Activation , Pyrimidines/pharmacology , Severity of Illness Index , Signal Transduction , Syk Kinase/antagonists & inhibitors
3.
Molecules ; 27(3)2022 Jan 26.
Article in English | MEDLINE | ID: covidwho-1648677

ABSTRACT

The human population is still facing appalling conditions due to several outbreaks of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) virus. The absence of specific drugs, appropriate vaccines for mutants, and knowledge of potential therapeutic agents makes this situation more difficult. Several 1, 2, 4-triazolo [1, 5-a] pyrimidine (TP)-derivative compounds were comprehensively studied for antiviral activities against RNA polymerase of HIV, HCV, and influenza viruses, and showed immense pharmacological interest. Therefore, TP-derivative compounds can be repurposed against the RNA-dependent RNA polymerase (RdRp) protein of SARS-CoV-2. In this study, a meta-analysis was performed to ensure the genomic variability and stability of the SARS-CoV-2 RdRp protein. The molecular docking of natural and synthetic TP compounds to RdRp and molecular dynamic (MD) simulations were performed to analyse the dynamic behaviour of TP compounds at the active site of the RdRp protein. TP compounds were also docked against other non-structural proteins (NSP1, NSP2, NSP3, NSP5, NSP8, NSP13, and NSP15) of SARS-CoV-2. Furthermore, the inhibition potential of TP compounds was compared with Remdesivir and Favipiravir drugs as a positive control. Additionally, TP compounds were analysed for inhibitory activity against SARS-CoV RdRp protein. This study demonstrates that TP analogues (monomethylated triazolopyrimidine and essramycin) represent potential lead molecules for designing an effective inhibitor to control viral replication. Furthermore, in vitro and in vivo studies will strengthen the use of these inhibitors as suitable drug candidates against SARS-CoV-2.


Subject(s)
Coronavirus RNA-Dependent RNA Polymerase/drug effects , Coronavirus RNA-Dependent RNA Polymerase/metabolism , Pyrimidines/pharmacology , Triazoles/pharmacology , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , Amides/pharmacology , COVID-19/drug therapy , COVID-19/metabolism , Catalytic Domain/drug effects , Computational Biology/methods , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Pyrazines/pharmacology , Pyrimidines/chemistry , RNA, Viral/drug effects , RNA-Dependent RNA Polymerase/drug effects , RNA-Dependent RNA Polymerase/metabolism , SARS-CoV-2/drug effects , SARS-CoV-2/metabolism , Triazoles/chemistry , Virus Replication/drug effects
4.
Brief Bioinform ; 23(1)2022 01 17.
Article in English | MEDLINE | ID: covidwho-1597924

ABSTRACT

The pharmacological arsenal against the COVID-19 pandemic is largely based on generic anti-inflammatory strategies or poorly scalable solutions. Moreover, as the ongoing vaccination campaign is rolling slower than wished, affordable and effective therapeutics are needed. To this end, there is increasing attention toward computational methods for drug repositioning and de novo drug design. Here, multiple data-driven computational approaches are systematically integrated to perform a virtual screening and prioritize candidate drugs for the treatment of COVID-19. From the list of prioritized drugs, a subset of representative candidates to test in human cells is selected. Two compounds, 7-hydroxystaurosporine and bafetinib, show synergistic antiviral effects in vitro and strongly inhibit viral-induced syncytia formation. Moreover, since existing drug repositioning methods provide limited usable information for de novo drug design, the relevant chemical substructures of the identified drugs are extracted to provide a chemical vocabulary that may help to design new effective drugs.


Subject(s)
Antiviral Agents/pharmacology , COVID-19 , Giant Cells , Pyrimidines/pharmacology , SARS-CoV-2/metabolism , Staurosporine/analogs & derivatives , A549 Cells , COVID-19/drug therapy , COVID-19/metabolism , Computational Biology , Drug Evaluation, Preclinical , Drug Repositioning , Giant Cells/metabolism , Giant Cells/virology , Humans , Staurosporine/pharmacology
5.
Int Immunopharmacol ; 103: 108463, 2022 Feb.
Article in English | MEDLINE | ID: covidwho-1587490

ABSTRACT

Therapeutics that impair the innate immune responses of the liver during the inflammatory cytokine storm like that occurring in COVID-19 are greatly needed. Much interest is currently directed toward Janus kinase (JAK) inhibitors as potential candidates to mitigate this life-threatening complication. Accordingly, this study investigated the influence of the novel JAK inhibitor ruxolitinib (RXB) on concanavalin A (Con A)-induced hepatitis and systemic hyperinflammation in mice to simulate the context occurring in COVID-19 patients. Mice were orally treated with RXB (75 and 150 mg/kg) 2 h prior to the intravenous administration of Con A (20 mg/kg) for a period of 12 h. The results showed that RXB pretreatments were efficient in abrogating Con A-instigated hepatocellular injury (ALT, AST, LDH), necrosis (histopathology), apoptosis (cleaved caspase-3) and nuclear proliferation due to damage (PCNA). The protective mechanism of RXB were attributed to i) prevention of Con A-enhanced hepatic production and systemic release of the proinflammatory cytokines TNF-α, IFN-γ and IL-17A, which coincided with decreasing infiltration of immune cells (monocytes, neutrophils), ii) reducing Con A-induced hepatic overexpression of IL-1ß and CD98 alongside NF-κB activation, and iii) lessening Con A-induced consumption of GSH and GSH peroxidase and generation of oxidative stress products (MDA, 4-HNE, NOx) in the liver. In summary, JAK inhibition by RXB led to eminent protection of the liver against Con A-deleterious manifestations primarily via curbing the inflammatory cytokine storm driven by TNF-α, IFN-γ and IL-17A.


Subject(s)
Concanavalin A/toxicity , Cytokine Release Syndrome/chemically induced , Cytokine Release Syndrome/drug therapy , Nitriles/pharmacology , Pyrazoles/pharmacology , Pyrimidines/pharmacology , Aldehydes/metabolism , Animals , Chemical and Drug Induced Liver Injury , Dose-Response Relationship, Drug , Inflammation/chemically induced , Liver/drug effects , Liver/metabolism , Male , Malondialdehyde/metabolism , Mice , Mice, Inbred BALB C , Nitrates/metabolism , Nitriles/administration & dosage , Nitrites/metabolism , Oxidative Stress , Peroxidase/metabolism , Pyrazoles/administration & dosage , Pyrimidines/administration & dosage
6.
Molecules ; 26(22)2021 Nov 09.
Article in English | MEDLINE | ID: covidwho-1538425

ABSTRACT

Novel complexes of type [Cu(N-N)(dmtp)2(OH2)](ClO4)2·dmtp ((1) N-N: 2,2'-bipyridine; (2) L: 1,10-phenantroline and dmtp: 5,7-dimethyl-1,2,4-triazolo[1,5-a]pyrimidine) were designed in order to obtain biologically active compounds. Complexes were characterized as mononuclear species that crystallized in the space group P-1 of the triclinic system with a square pyramidal geometry around the copper (II). In addition to the antiproliferative effect on murine melanoma B16 cells, complex (1) exhibited low toxicity on normal BJ cells and did not affect membrane integrity. Complex (2) proved to be a more potent antimicrobial in comparison with (1), but both compounds were more active in comparison with dmtp-both against planktonic cells and biofilms. A stronger antimicrobial and antibiofilm effect was noticed against the Gram-positive strains, including methicillin-resistant Staphylococcus aureus (MRSA). Both electron paramagnetic resonance (EPR) and Saccharomyces cerevisiae studies indicated that the complexes were scavengers rather than reactive oxygen species promoters. Their DNA intercalating capacity was evidenced by modifications in both absorption and fluorescence spectra. Furthermore, both complexes exhibited nuclease-like activity, which increased in the presence of hydrogen peroxide.


Subject(s)
Anti-Infective Agents , Chelating Agents , Coordination Complexes , Methicillin-Resistant Staphylococcus aureus/growth & development , Pyrimidines , Saccharomyces cerevisiae/growth & development , Animals , Anti-Infective Agents/chemical synthesis , Anti-Infective Agents/chemistry , Anti-Infective Agents/pharmacology , Cell Line, Tumor , Chelating Agents/chemical synthesis , Chelating Agents/chemistry , Chelating Agents/pharmacology , Coordination Complexes/chemical synthesis , Coordination Complexes/chemistry , Coordination Complexes/pharmacology , Humans , Mice , Pyrimidines/chemical synthesis , Pyrimidines/chemistry , Pyrimidines/pharmacology
7.
Molecules ; 26(19)2021 Oct 04.
Article in English | MEDLINE | ID: covidwho-1463767

ABSTRACT

Antimicrobial resistance was one of the top priorities for global public health before the start of the 2019 coronavirus pandemic (COVID-19). Moreover, in this changing medical landscape due to COVID-19, finding new organic structures with antimicrobial and antiviral properties is a priority in current research. The Biginelli synthesis that mediates the production of pyrimidine compounds has been intensively studied in recent decades, especially due to the therapeutic properties of the resulting compounds, such as calcium channel blockers, anticancer, antiviral, antimicrobial, anti-inflammatory or antioxidant compounds. In this review we aim to review the Biginelli syntheses reported recently in the literature that mediates the synthesis of antimicrobial compounds, the spectrum of their medicinal properties, and the structure-activity relationship in the studied compounds.


Subject(s)
Anti-Infective Agents/chemical synthesis , Pyrimidines/chemical synthesis , Animals , Anti-Infective Agents/chemistry , Anti-Infective Agents/pharmacology , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , COVID-19/drug therapy , Chemistry Techniques, Synthetic/methods , Drug Discovery , Drug Resistance, Microbial , Humans , Models, Molecular , Pyrimidines/chemistry , Pyrimidines/pharmacology , SARS-CoV-2/drug effects
8.
Cancer Res ; 80(17): 3530-3541, 2020 09 01.
Article in English | MEDLINE | ID: covidwho-1455780

ABSTRACT

Reactive oxygen species (ROS) oxidize nucleotide triphosphate pools (e.g., 8-oxodGTP), which may kill cells if incorporated into DNA. Whether cancers avoid poisoning from oxidized nucleotides by preventing incorporation via the oxidized purine diphosphatase MTH1 remains under debate. Also, little is known about DNA polymerases incorporating oxidized nucleotides in cells or how oxidized nucleotides in DNA become toxic. Here we show that replacement of one of the main DNA replicases in human cells, DNA polymerase delta (Pol δ), with an error-prone variant allows increased 8-oxodG accumulation into DNA following treatment with TH588, a dual MTH1 inhibitor and microtubule targeting agent. The resulting elevated genomic 8-oxodG correlated with increased cytotoxicity of TH588. Interestingly, no substantial perturbation of replication fork progression was observed, but rather mitotic progression was impaired and mitotic DNA synthesis triggered. Reducing mitotic arrest by reversin treatment prevented accumulation of genomic 8-oxodG and reduced cytotoxicity of TH588, in line with the notion that mitotic arrest is required for ROS buildup and oxidation of the nucleotide pool. Furthermore, delayed mitosis and increased mitotic cell death was observed following TH588 treatment in cells expressing the error-prone but not wild-type Pol δ variant, which is not observed following treatments with antimitotic agents. Collectively, these results link accumulation of genomic oxidized nucleotides with disturbed mitotic progression. SIGNIFICANCE: These findings uncover a novel link between accumulation of genomic 8-oxodG and perturbed mitotic progression in cancer cells, which can be exploited therapeutically using MTH1 inhibitors.See related commentary by Alnajjar and Sweasy, p. 3459.


Subject(s)
8-Hydroxy-2'-Deoxyguanosine , Phosphoric Monoester Hydrolases , DNA Repair Enzymes/genetics , Genomics , Humans , Mitosis/genetics , Phosphoric Monoester Hydrolases/genetics , Pyrimidines/pharmacology
9.
Eur J Med Chem ; 221: 113494, 2021 Oct 05.
Article in English | MEDLINE | ID: covidwho-1446590

ABSTRACT

In the search for new anti-influenza virus (IV) compounds, we have identified the 1,2,4-triazolo[1,5-a]pyrimidine (TZP) as a very suitable scaffold to obtain compounds able to disrupt IV RNA-dependent RNA polymerase (RdRP) PA-PB1 subunits heterodimerization. In this work, in order to acquire further SAR insights for this class of compounds and identify more potent derivatives, we designed and synthesized additional series of analogues to investigate the role of the substituents around the TZP core. To this aim, we developed four facile and efficient one-step procedures for the synthesis of 5-phenyl-, 6-phenyl- and 7-phenyl-2-amino-[1,2,4]triazolo[1,5-a]pyrimidines, and 2-amino-5-phenyl-[1,2,4]triazolo[1,5-a]pyrimidin-7-ol. Two analogues having the ethyl carboxylate moiety at the C-2 position of the TZP were also prepared in good yields. Then, the scaffolds herein synthesized and two previous scaffolds were functionalized and evaluated for their anti-IAV activity, leading to the identification of compound 22 that showed both anti-PA-PB1 (IC50 = 19.5 µM) and anti-IAV activity (EC50 = 16 µM) at non-toxic concentrations, thus resulting among the most active TZP derivatives reported to date by us. A selection of the synthesized compounds, along with a set of in-house available analogues, was also tested against SARS-CoV-2. The most promising compound 49 from this series displayed an EC50 value of 34.47 µM, highlighting the potential of the TPZ scaffold in the search for anti-CoV agents.


Subject(s)
Antiviral Agents/pharmacology , Protein Multimerization/drug effects , Pyrimidines/pharmacology , RNA-Dependent RNA Polymerase/antagonists & inhibitors , Triazoles/pharmacology , Viral Proteins/antagonists & inhibitors , Animals , Antiviral Agents/chemical synthesis , Chlorocebus aethiops , Dogs , Drug Design , HEK293 Cells , Humans , Influenza A virus/drug effects , Madin Darby Canine Kidney Cells , Microbial Sensitivity Tests , Pyrimidines/chemical synthesis , SARS-CoV-2/drug effects , Triazoles/chemical synthesis , Vero Cells
10.
J Biomol Struct Dyn ; 39(15): 5768-5778, 2021 Sep.
Article in English | MEDLINE | ID: covidwho-1390294

ABSTRACT

The entire human population over the globe is currently facing appalling conditions due to the spread of infection from coronavirus disease-2019 (COVID-19). The spike glycoprotein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) present on the surface of the virion mediates the virus entry into the host cells and therefore is targeted by several scientific groups as a novel drug target site. The spike glycoprotein binds to the human angiotensin-converting enzyme-2 (hACE2) cell surface receptor abundantly expressed in lung tissues, and this binding phenomenon is a primary determinant of cell tropism and pathogenesis. The binding and internalization of the virus is the primary and most crucial step in the process of infection, and therefore the molecules targeting the inhibition of this process certainly hold a significant therapeutic value. Thus, we systematically applied the computational techniques to identify the plausible inhibitor from a chosen set of well characterized diaryl pyrimidine analogues which may disrupt interfacial interaction of spike glycoprotein (S) at the surface of hACE2. Using molecular docking, molecular dynamics (MD) simulation and binding free energy calculation, we have identified AP-NP (2-(2-amino-5-(naphthalen-2-yl)pyrimidin-4-yl)phenol), AP-3-OMe-Ph (2-(2-amino-5-(3-methoxyphenyl)pyrimidin-4-yl)phenol) and AP-4-Me-Ph (2-(2-amino-5-(p-tolyl) pyrimidin-4-yl)phenol) from a group of diaryl pyrimidine derivatives which appears to bind at the interface of the hACE2-S complex with low binding free energy. Thus, pyrimidine derivative AP-NP may be explored as an effective inhibitor for hACE2-S complex. Furthermore, in vitro and in vivo studies will strengthen the use of these inhibitors as suitable drug candidates against SARS-COV-2. Communicated by Ramaswamy H. Sarma.


Subject(s)
COVID-19 , Pharmaceutical Preparations , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Pyrimidines/pharmacology , SARS-CoV-2 , Spike Glycoprotein, Coronavirus
11.
J Virol ; 95(21): e0097521, 2021 10 13.
Article in English | MEDLINE | ID: covidwho-1361966

ABSTRACT

Repurposing FDA-approved inhibitors able to prevent infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) could provide a rapid path to establish new therapeutic options to mitigate the effects of coronavirus disease 2019 (COVID-19). Proteolytic cleavages of the spike (S) protein of SARS-CoV-2, mediated by the host cell proteases cathepsin and TMPRSS2, alone or in combination, are key early activation steps required for efficient infection. The PIKfyve kinase inhibitor apilimod interferes with late endosomal viral traffic and through an ill-defined mechanism prevents in vitro infection through late endosomes mediated by cathepsin. Similarly, inhibition of TMPRSS2 protease activity by camostat mesylate or nafamostat mesylate prevents infection mediated by the TMPRSS2-dependent and cathepsin-independent pathway. Here, we combined the use of apilimod with camostat mesylate or nafamostat mesylate and found an unexpected ∼5- to 10-fold increase in their effectiveness to prevent SARS-CoV-2 infection in different cell types. Comparable synergism was observed using both a chimeric vesicular stomatitis virus (VSV) containing S of SARS-CoV-2 (VSV-SARS-CoV-2) and SARS-CoV-2. The substantial ∼5-fold or higher decrease of the half-maximal effective concentrations (EC50s) suggests a plausible treatment strategy based on the combined use of these inhibitors. IMPORTANCE Infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is causing the coronavirus disease 2019 (COVID-2019) global pandemic. There are ongoing efforts to uncover effective antiviral agents that could mitigate the severity of the disease by controlling the ensuing viral replication. Promising candidates include small molecules that inhibit the enzymatic activities of host proteins, thus preventing SARS-CoV-2 entry and infection. They include apilimod, an inhibitor of PIKfyve kinase, and camostat mesylate and nafamostat mesylate, inhibitors of TMPRSS2 protease. Our research is significant for having uncovered an unexpected synergism in the effective inhibitory activity of apilimod used together with camostat mesylate or nafamostat mesylate.


Subject(s)
Antiviral Agents/pharmacology , Benzamidines/pharmacology , Esters/pharmacology , Guanidines/pharmacology , Hydrazones/pharmacology , Morpholines/pharmacology , Phosphatidylinositol 3-Kinases/metabolism , Pyrimidines/pharmacology , SARS-CoV-2/drug effects , Serine Endopeptidases/metabolism , Animals , COVID-19/drug therapy , Cell Line, Tumor , Chlorocebus aethiops , Drug Synergism , Humans , Phosphoinositide-3 Kinase Inhibitors/pharmacology , Protease Inhibitors/pharmacology , SARS-CoV-2/physiology , Vero Cells , Virus Internalization
12.
Molecules ; 26(15)2021 Jul 29.
Article in English | MEDLINE | ID: covidwho-1346516

ABSTRACT

We recently developed a molecule (GT-73) that blocked leukocyte transendothelial migration from blood to the peripheral tissues, supposedly by affecting the platelet endothelial cell adhesion molecule (PECAM-1) function. GT-73 was tested in an LPS-induced acute respiratory distress syndrome (ARDS) mouse model. The rationale for this is based on the finding that the mortality of COVID-19 patients is partly caused by ARDS induced by a massive migration of leukocytes to the lungs. In addition, the role of tert-butyl and methyl ester moieties in the biological effect of GT-73 was investigated. A human leukocyte, transendothelial migration assay was applied to validate the blocking effect of GT-73 derivatives. Finally, a mouse model of LPS-induced ARDS was used to evaluate the histological and biochemical effects of GT-73. The obtained results showed that GT-73 has a unique structure that is responsible for its biological activity; two of its chemical moieties (tert-butyl and a methyl ester) are critical for this effect. GT-73 is a prodrug, and its lipophilic tail covalently binds to PECAM-1 via Lys536. GT-73 significantly decreased the number of infiltrating leukocytes in the lungs and reduced the inflammation level. Finally, GT-73 reduced the levels of IL-1ß, IL-6, and MCP-1 in bronchoalveolar lavage fluid (BALF). In summary, we concluded that GT-73, a blocker of white blood cell transendothelial migration, has a favorable profile as a drug candidate for the treatment of ARDS in COVID-19 patients.


Subject(s)
COVID-19/drug therapy , Leukocytes/drug effects , Platelet Endothelial Cell Adhesion Molecule-1/antagonists & inhibitors , Pyrimidines/pharmacology , Respiratory Distress Syndrome/drug therapy , Transendothelial and Transepithelial Migration/drug effects , Animals , COVID-19/pathology , Cell Adhesion/drug effects , Cell Adhesion/immunology , Cell Movement/drug effects , Cytokine Release Syndrome/drug therapy , Cytokines/metabolism , Disease Models, Animal , Female , Humans , Leukocytes/immunology , Lipopolysaccharides/adverse effects , Mice , Mice, Inbred BALB C , Platelet Endothelial Cell Adhesion Molecule-1/immunology , Pyrimidines/chemistry , Respiratory Distress Syndrome/chemically induced , SARS-CoV-2
13.
Cell Rep ; 36(5): 109479, 2021 08 03.
Article in English | MEDLINE | ID: covidwho-1328702

ABSTRACT

Coronaviruses rely on host membranes for entry, establishment of replication centers, and egress. Compounds targeting cellular membrane biology and lipid biosynthetic pathways have previously shown promise as antivirals and are actively being pursued as treatments for other conditions. Here, we test small molecule inhibitors that target the PI3 kinase VPS34 or fatty acid metabolism for anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) activity. Our studies determine that compounds targeting VPS34 are potent SARS-CoV-2 inhibitors. Mechanistic studies with compounds targeting multiple steps up- and downstream of fatty acid synthase (FASN) identify the importance of triacylglycerol production and protein palmitoylation as requirements for efficient viral RNA synthesis and infectious virus production. Further, FASN knockout results in significantly impaired SARS-CoV-2 replication that can be rescued with fatty acid supplementation. Together, these studies clarify roles for VPS34 and fatty acid metabolism in SARS-CoV-2 replication and identify promising avenues for the development of countermeasures against SARS-CoV-2.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/virology , Class III Phosphatidylinositol 3-Kinases/antagonists & inhibitors , Lipid Metabolism/drug effects , SARS-CoV-2/drug effects , SARS-CoV-2/physiology , Virus Replication/drug effects , Aminopyridines/pharmacology , Animals , Caco-2 Cells , Cell Line , Chlorocebus aethiops , Class III Phosphatidylinositol 3-Kinases/metabolism , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/genetics , Gene Knockout Techniques , Humans , Lipoylation/drug effects , Pyrimidines/pharmacology , RNA, Viral/metabolism , Triglycerides/metabolism , Vero Cells
14.
Molecules ; 26(13)2021 Jun 22.
Article in English | MEDLINE | ID: covidwho-1304687

ABSTRACT

Discovery of compound 1 as a Zika virus (ZIKV) inhibitor has prompted us to investigate its 7H-pyrrolo[2,3-d]pyrimidine scaffold, revealing structural features that elicit antiviral activity. Furthermore, we have demonstrated that 9H-purine or 1H-pyrazolo[3,4-d]pyrimidine can serve as an alternative core structure. Overall, we have identified 4,7-disubstituted 7H-pyrrolo[2,3-d]pyrimidines and their analogs including compounds 1, 8 and 11 as promising antiviral agents against flaviviruses ZIKV and dengue virus (DENV). While the molecular target of these compounds is yet to be elucidated, 4,7-disubstituted 7H-pyrrolo[2,3-d]pyrimidines and their analogs are new chemotypes in the design of small molecules against flaviviruses, an important group of human pathogens.


Subject(s)
Antiviral Agents , Pyrimidines , Virus Replication/drug effects , Zika Virus Infection/drug therapy , Zika Virus/physiology , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Cell Line, Tumor , Humans , Pyrimidines/chemical synthesis , Pyrimidines/chemistry , Pyrimidines/pharmacology , Zika Virus Infection/metabolism , Zika Virus Infection/pathology
15.
Eur J Med Chem ; 221: 113494, 2021 Oct 05.
Article in English | MEDLINE | ID: covidwho-1291451

ABSTRACT

In the search for new anti-influenza virus (IV) compounds, we have identified the 1,2,4-triazolo[1,5-a]pyrimidine (TZP) as a very suitable scaffold to obtain compounds able to disrupt IV RNA-dependent RNA polymerase (RdRP) PA-PB1 subunits heterodimerization. In this work, in order to acquire further SAR insights for this class of compounds and identify more potent derivatives, we designed and synthesized additional series of analogues to investigate the role of the substituents around the TZP core. To this aim, we developed four facile and efficient one-step procedures for the synthesis of 5-phenyl-, 6-phenyl- and 7-phenyl-2-amino-[1,2,4]triazolo[1,5-a]pyrimidines, and 2-amino-5-phenyl-[1,2,4]triazolo[1,5-a]pyrimidin-7-ol. Two analogues having the ethyl carboxylate moiety at the C-2 position of the TZP were also prepared in good yields. Then, the scaffolds herein synthesized and two previous scaffolds were functionalized and evaluated for their anti-IAV activity, leading to the identification of compound 22 that showed both anti-PA-PB1 (IC50 = 19.5 µM) and anti-IAV activity (EC50 = 16 µM) at non-toxic concentrations, thus resulting among the most active TZP derivatives reported to date by us. A selection of the synthesized compounds, along with a set of in-house available analogues, was also tested against SARS-CoV-2. The most promising compound 49 from this series displayed an EC50 value of 34.47 µM, highlighting the potential of the TPZ scaffold in the search for anti-CoV agents.


Subject(s)
Antiviral Agents/pharmacology , Protein Multimerization/drug effects , Pyrimidines/pharmacology , RNA-Dependent RNA Polymerase/antagonists & inhibitors , Triazoles/pharmacology , Viral Proteins/antagonists & inhibitors , Animals , Antiviral Agents/chemical synthesis , Chlorocebus aethiops , Dogs , Drug Design , HEK293 Cells , Humans , Influenza A virus/drug effects , Madin Darby Canine Kidney Cells , Microbial Sensitivity Tests , Pyrimidines/chemical synthesis , SARS-CoV-2/drug effects , Triazoles/chemical synthesis , Vero Cells
16.
Int J Biol Sci ; 17(9): 2348-2355, 2021.
Article in English | MEDLINE | ID: covidwho-1285527

ABSTRACT

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has led to more than 150 million infections and about 3.1 million deaths up to date. Currently, drugs screened are urgently aiming to block the infection of SARS-CoV-2. Here, we explored the interaction networks of kinase and COVID-19 crosstalk, and identified phosphoinositide 3-kinase (PI3K)/AKT pathway as the most important kinase signal pathway involving COVID-19. Further, we found a PI3K/AKT signal pathway inhibitor capivasertib restricted the entry of SARS-CoV-2 into cells under non-cytotoxic concentrations. Lastly, the signal axis PI3K/AKT/FYVE finger-containing phosphoinositide kinase (PIKfyve)/PtdIns(3,5)P2 was revealed to play a key role during the cellular entry of viruses including SARS-CoV-2, possibly providing potential antiviral targets. Altogether, our study suggests that the PI3K/AKT kinase inhibitor drugs may be a promising anti-SARS-CoV-2 strategy for clinical application, especially for managing cancer patients with COVID-19 in the pandemic era.


Subject(s)
COVID-19/drug therapy , Proto-Oncogene Proteins c-akt/antagonists & inhibitors , Pyrimidines/therapeutic use , Pyrroles/therapeutic use , SARS-CoV-2/drug effects , Virus Internalization/drug effects , Animals , COVID-19/enzymology , Chlorocebus aethiops , Computer Simulation , Humans , Neoplasms/enzymology , Neoplasms/mortality , Proto-Oncogene Proteins c-akt/metabolism , Pyrimidines/pharmacology , Pyrroles/pharmacology , Receptor Cross-Talk , Vero Cells
17.
Eur J Immunol ; 51(9): 2330-2340, 2021 09.
Article in English | MEDLINE | ID: covidwho-1261763

ABSTRACT

The molecular mechanism of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Spike protein was characterized to identify novel therapies. The impact of tofacitinib, IL-6R Ab, or TNFi therapy was determined on Spike protein or LPS/IFN-γ-induced signaling, inflammation, and metabolic reprogramming in MΦs and/or rheumatoid arthritis (RA) fibroblast-like synoviocyte (FLS). ACE2 frequency was markedly expanded in MΦs compared to T cells and RA FLS. Tofacitinib suppresses Spike protein potentiated STAT1 signaling, whereas this function was unchanged by TNFi. Tofacitinib impairs IL-6/IFN/LPS-induced STAT1 and STAT3 phosphorylation in RA MΦs and FLS. Interestingly, tofacitinib had a broader inhibitory effect on the monokines, glycolytic regulators, or oxidative metabolites compared to IL-6R Ab and TNFi in Spike-protein-activated MΦs. In contrast, all three therapies disrupted IFN-α and IFN-ß secretion in response to Spike protein; nonetheless, the IFN-γ was only curtailed by tofacitinib or IL-6R Ab. While tofacitinib counteracted MΦ metabolic rewiring instigated by Spike protein, it was inconsequential on the glycolysis expansion mediated via HK2 and/or LDHA in the activated RA MΦ and FLS. Nevertheless, the potentiated inflammatory response and the diminished oxidative phosphorylation modulated by Spike protein and/or LPS/IFN-γ stimulation in MΦs or RA FLS were reversed by tofacitinib. In conclusion, tofacitinib suppresses MΦ inflammation and immunometabolism triggered by Spike protein and may provide a promising strategy for COVID-19 patients.


Subject(s)
COVID-19/drug therapy , Macrophages/drug effects , Piperidines/pharmacology , Pyrimidines/pharmacology , SARS-CoV-2/drug effects , Spike Glycoprotein, Coronavirus/metabolism , Arthritis, Rheumatoid/metabolism , COVID-19/metabolism , Cells, Cultured , Fibroblasts/drug effects , Fibroblasts/metabolism , Humans , Interleukin-6/metabolism , Macrophages/metabolism , Receptors, Interleukin-6/metabolism , STAT1 Transcription Factor/metabolism , STAT3 Transcription Factor/metabolism , Signal Transduction/drug effects , Tumor Necrosis Factor-alpha/metabolism
18.
Front Immunol ; 12: 635018, 2021.
Article in English | MEDLINE | ID: covidwho-1211810

ABSTRACT

Objective: Bacterial and viral infectious triggers are linked to spondyloarthritis (SpA) including psoriatic arthritis (PsA) development, likely via dendritic cell activation. We investigated spinal entheseal plasmacytoid dendritic cells (pDCs) toll-like receptor (TLR)-7 and 9 activation and therapeutic modulation, including JAK inhibition. We also investigated if COVID-19 infection, a potent TLR-7 stimulator triggered PsA flares. Methods: Normal entheseal pDCs were characterized and stimulated with imiquimod and CpG oligodeoxynucleotides (ODN) to evaluate TNF and IFNα production. NanoString gene expression assay of total pDCs RNA was performed pre- and post- ODN stimulation. Pharmacological inhibition of induced IFNα protein was performed with Tofacitinib and PDE4 inhibition. The impact of SARS-CoV2 viral infection on PsA flares was evaluated. Results: CD45+HLA-DR+CD123+CD303+CD11c- entheseal pDCs were more numerous than blood pDCs (1.9 ± 0.8% vs 0.2 ± 0.07% of CD45+ cells, p=0.008) and showed inducible IFNα and TNF protein following ODN/imiquimod stimulation and were the sole entheseal IFNα producers. NanoString data identified 11 significantly upregulated differentially expressed genes (DEGs) including TNF in stimulated pDCs. Canonical pathway analysis revealed activation of dendritic cell maturation, NF-κB signaling, toll-like receptor signaling and JAK/STAT signaling pathways following ODN stimulation. Both tofacitinib and PDE4i strongly attenuated ODN induced IFNα. DAPSA scores elevations occurred in 18 PsA cases with SARS-CoV2 infection (9.7 ± 4 pre-infection and 35.3 ± 7.5 during infection). Conclusion: Entheseal pDCs link microbes to TNF/IFNα production. SARS-CoV-2 infection is associated with PsA Flares and JAK inhibition suppressed activated entheseal plasmacytoid dendritic Type-1 interferon responses as pointers towards a novel mechanism of PsA and SpA-related arthropathy.


Subject(s)
Arthritis, Psoriatic/complications , COVID-19/complications , Dendritic Cells/metabolism , Interferon-alpha/metabolism , Janus Kinases/antagonists & inhibitors , Adjuvants, Immunologic/pharmacology , Adult , Aged , COVID-19/genetics , COVID-19/metabolism , Computational Biology , Cyclic Nucleotide Phosphodiesterases, Type 4/metabolism , Dendritic Cells/drug effects , Female , Gene Expression Regulation/drug effects , Gene Expression Regulation/genetics , Humans , Imiquimod/pharmacology , Janus Kinases/metabolism , Male , Middle Aged , NF-kappa B/metabolism , Oligonucleotides/pharmacology , Phosphodiesterase 4 Inhibitors/pharmacology , Piperidines/pharmacology , Protein Kinase Inhibitors/pharmacology , Pyrimidines/pharmacology , Signal Transduction/drug effects , Signal Transduction/genetics , Toll-Like Receptor 7/metabolism , Toll-Like Receptor 9/metabolism , Transcriptome , Tumor Necrosis Factor-alpha/metabolism
19.
Nat Commun ; 12(1): 1876, 2021 03 25.
Article in English | MEDLINE | ID: covidwho-1152854

ABSTRACT

Viruses hijack host cell metabolism to acquire the building blocks required for replication. Understanding how SARS-CoV-2 alters host cell metabolism may lead to potential treatments for COVID-19. Here we profile metabolic changes conferred by SARS-CoV-2 infection in kidney epithelial cells and lung air-liquid interface (ALI) cultures, and show that SARS-CoV-2 infection increases glucose carbon entry into the TCA cycle via increased pyruvate carboxylase expression. SARS-CoV-2 also reduces oxidative glutamine metabolism while maintaining reductive carboxylation. Consistent with these changes, SARS-CoV-2 infection increases the activity of mTORC1 in cell lines and lung ALI cultures. Lastly, we show evidence of mTORC1 activation in COVID-19 patient lung tissue, and that mTORC1 inhibitors reduce viral replication in kidney epithelial cells and lung ALI cultures. Our results suggest that targeting mTORC1 may be a feasible treatment strategy for COVID-19 patients, although further studies are required to determine the mechanism of inhibition and potential efficacy in patients.


Subject(s)
COVID-19/pathology , Citric Acid Cycle/physiology , Mechanistic Target of Rapamycin Complex 1/antagonists & inhibitors , Mechanistic Target of Rapamycin Complex 1/metabolism , Protein Kinase Inhibitors/pharmacology , Animals , Benzamides/pharmacology , Cell Line , Chlorocebus aethiops , Glucose/metabolism , Glutamine/metabolism , HEK293 Cells , Humans , Lung/metabolism , Lung/virology , Morpholines/pharmacology , Naphthyridines/pharmacology , Pyrimidines/pharmacology , Pyruvate Carboxylase/biosynthesis , SARS-CoV-2/metabolism , Vero Cells , Virus Replication/drug effects
20.
Viruses ; 13(2)2021 02 09.
Article in English | MEDLINE | ID: covidwho-1128060

ABSTRACT

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the newly emergent causative agent of coronavirus disease-19 (COVID-19), has resulted in more than two million deaths worldwide since it was first detected in 2019. There is a critical global need for therapeutic intervention strategies that can be deployed to safely treat COVID-19 disease and reduce associated morbidity and mortality. Increasing evidence shows that both natural and synthetic antimicrobial peptides (AMPs), also referred to as Host Defense Proteins/Peptides (HDPs), can inhibit SARS-CoV-2, paving the way for the potential clinical use of these molecules as therapeutic options. In this manuscript, we describe the potent antiviral activity exerted by brilacidin-a de novo designed synthetic small molecule that captures the biological properties of HDPs-on SARS-CoV-2 in a human lung cell line (Calu-3) and a monkey cell line (Vero). These data suggest that SARS-CoV-2 inhibition in these cell culture models is likely to be a result of the impact of brilacidin on viral entry and its disruption of viral integrity. Brilacidin demonstrated synergistic antiviral activity when combined with remdesivir. Collectively, our data demonstrate that brilacidin exerts potent inhibition of SARS-CoV-2 against different strains of the virus in cell culture.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Guanidines/pharmacology , Pyrimidines/pharmacology , SARS-CoV-2/drug effects , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , Animals , COVID-19/virology , Cell Culture Techniques , Cell Line , Chlorocebus aethiops , Defensins/pharmacology , Humans , Peptidomimetics/pharmacology , SARS-CoV-2/physiology , Vero Cells , Virus Internalization/drug effects , Virus Replication/drug effects
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