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1.
Viruses ; 14(2)2022 02 17.
Article in English | MEDLINE | ID: covidwho-1703374

ABSTRACT

Coronaviruses (CoVs) are common among humans and many animals, causing respiratory or gastrointestinal diseases. Currently, only a few antiviral drugs against CoVs are available. Especially for SARS-CoV-2, new compounds for treatment of COVID-19 are urgently needed. In this study, we characterize the antiviral effects of two high-sulfated glycosaminoglycan (GAG) derivatives against SARS-CoV-2 and bovine coronaviruses (BCoV), which are both members of the Betacoronavirus genus. The investigated compounds are based on hyaluronan (HA) and chondroitin sulfate (CS) and exhibit a strong inhibitory effect against both CoVs. Yield assays were performed using BCoV-infected PT cells in the presence and absence of the compounds. While the high-sulfated HA (sHA3) led to an inhibition of viral growth early after infection, high-sulfated CS (sCS3) had a slightly smaller effect. Time of addition assays, where sHA3 and sCS3 were added to PT cells before, during or after infection, demonstrated an inhibitory effect during all phases of infection, whereas sHA3 showed a stronger effect even after virus absorbance. Furthermore, attachment analyses with prechilled PT cells revealed that virus attachment is not blocked. In addition, sHA3 and sCS3 inactivated BCoV by stable binding. Analysis by quantitative real-time RT PCR underlines the high potency of the inhibitors against BCoV, as well as B.1-lineage, Alpha and Beta SARS-CoV-2 viruses. Taken together, these results demonstrated that the two high-sulfated GAG derivatives exhibit low cytotoxicity and represent promising candidates for an anti-CoV therapy.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus Infections/veterinary , Coronavirus, Bovine/drug effects , Glycosaminoglycans/pharmacology , SARS-CoV-2/drug effects , Animals , COVID-19/drug therapy , Cattle , Cell Line , Chondroitin Sulfates/chemistry , Chondroitin Sulfates/pharmacology , Coronavirus Infections/drug therapy , Glycosaminoglycans/chemistry , Glycosaminoglycans/metabolism , Humans , Hyaluronic Acid/chemistry , Hyaluronic Acid/pharmacology , Sulfates/chemistry , Sulfates/pharmacology , Virus Attachment/drug effects
2.
Cells ; 10(6)2021 06 07.
Article in English | MEDLINE | ID: covidwho-1259431

ABSTRACT

Coronaviruses such as SARS-CoV-2, which is responsible for COVID-19, depend on virus spike protein binding to host cell receptors to cause infection. The SARS-CoV-2 spike protein binds primarily to ACE2 on target cells and is then processed by membrane proteases, including TMPRSS2, leading to viral internalisation or fusion with the plasma membrane. It has been suggested, however, that receptors other than ACE2 may be involved in virus binding. We have investigated the interactions of recombinant versions of the spike protein with human epithelial cell lines that express low/very low levels of ACE2 and TMPRSS2 in a proxy assay for interaction with host cells. A tagged form of the spike protein containing the S1 and S2 regions bound in a temperature-dependent manner to all cell lines, whereas the S1 region alone and the receptor-binding domain (RBD) interacted only weakly. Spike protein associated with cells independently of ACE2 and TMPRSS2, while RBD required the presence of high levels of ACE2 for interaction. As the spike protein has previously been shown to bind heparin, a soluble glycosaminoglycan, we tested the effects of various heparins on ACE2-independent spike protein interaction with cells. Unfractionated heparin inhibited spike protein interaction with an IC50 value of <0.05 U/mL, whereas two low-molecular-weight heparins were less effective. A mutant form of the spike protein, lacking the arginine-rich putative furin cleavage site, interacted only weakly with cells and had a lower affinity for unfractionated and low-molecular-weight heparin than the wild-type spike protein. This suggests that the furin cleavage site might also be a heparin-binding site and potentially important for interactions with host cells. The glycosaminoglycans heparan sulphate and dermatan sulphate, but not chondroitin sulphate, also inhibited the binding of spike protein, indicating that it might bind to one or both of these glycosaminoglycans on the surface of target cells.


Subject(s)
Angiotensin-Converting Enzyme 2/physiology , Epithelial Cells/metabolism , Heparin/pharmacology , Spike Glycoprotein, Coronavirus/metabolism , A549 Cells , Angiotensin-Converting Enzyme 2/genetics , Animals , Binding Sites/drug effects , Binding Sites/genetics , Caco-2 Cells , Cell Line , Chlorocebus aethiops , Dermatan Sulfate/pharmacology , Down-Regulation/drug effects , Epithelial Cells/drug effects , Epithelial Cells/virology , Glycosaminoglycans/pharmacology , HEK293 Cells , HaCaT Cells , Heparitin Sulfate/pharmacology , Humans , Protein Binding/drug effects , Protein Binding/genetics , SARS-CoV-2/drug effects , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/chemistry , Vero Cells , Virus Internalization/drug effects
3.
Blood Coagul Fibrinolysis ; 32(3): 167-171, 2021 Apr 01.
Article in English | MEDLINE | ID: covidwho-1171412

ABSTRACT

Coronavirus disease 2019 infection produce a prothrombotic state. This is initiated through multiple pathways and is finally aggravated by cross talks with cytokine storm and neutrophil, platelet, complement activation. All these combine towards the second week of illness to produce thrombosis in the lung capillaries surrounding the alveolus producing characteristic pulmonary dysfunction (PaO2/FiO2 > 300, normal or minimally increased lung compliance and very high d-dimer levels) and a high rate of peripheral venous thrombosis. International and many national guidelines have approached this state in different ways but all emphasized the need for management and prevention of widespread thrombosis. It is felt more aggressive and graded thrombosis prevention and management should be initiated early in the treatment. d-Dimer, neutrophil count, SaO2, fibrinogen levels should be used to control the hypercoagulability. Drugs like statins which have anti-inflammatory action as well as ability to reduce fibrinogen and other clotting factors should be used in the beginning along with antiplatelet drugs and progressively complement activation and neutrophil extracellular traps inhibitors, oral mucopolysaccharides, full-scale anticoagulation along with judicial use of fibrinolysis supporting drugs should be added. In the present review, we have evaluated the various studies and argued the rationality that the anticoagulation in this condition should be initiated early during the infection and should be increased in a graded manner depending on clinical and laboratory progression of the condition until a strong specific antiviral drug for coronavirus disease 2019 infection is available.


Subject(s)
Blood Coagulation/drug effects , COVID-19/drug therapy , COVID-19/physiopathology , Thrombophilia/drug therapy , Thrombosis/drug therapy , Anticoagulants/therapeutic use , Antiviral Agents/therapeutic use , Blood Platelets/drug effects , Extracellular Traps/drug effects , Fibrinolytic Agents/therapeutic use , Glycosaminoglycans/pharmacology , Glycosaminoglycans/therapeutic use , Humans , Hydroxymethylglutaryl-CoA Reductase Inhibitors/pharmacology , Hydroxymethylglutaryl-CoA Reductase Inhibitors/therapeutic use , Inflammation/drug therapy , Lung/drug effects , Lung/physiopathology , Lung/virology , Platelet Aggregation Inhibitors/therapeutic use
4.
Clin Appl Thromb Hemost ; 26: 1076029620954913, 2020.
Article in English | MEDLINE | ID: covidwho-841480

ABSTRACT

INTRODUCTION: Sulodexide represents a mixture of fast-moving heparin (FMH) and dermatan sulfate (DS) and has been used for the management of venous diseases such as DVT and related disorders. The purpose of this study is to compare sulodexide and its components with unfractionated heparin (UFH) to determine its suitability for the indications in which UFH is used. MATERIALS AND METHOD: Active pharmaceutical ingredients (API) versions of sulodexide, FMH and DS were obtained from Alfasigma. API versions of UFH were obtained from Medefil Inc. Normal human citrated plasma was obtained from blood bank of the Loyola University Medical Center. Each of the individual agents were supplemented in plasma at a graded concentration of 0.0-10 µg/mL. Clotting assays (PiCT, aPTT, PT and TT), anti-Xa and anti-IIa and thrombin generation studies were carried out. Results were compiled as mean ± SD of 3 individual determination. RESULT: In the clot based (PiCT, aPTT and TT), anti-Xa and IIa assays, both the UFH and FMH produced stronger activities in these assays followed by sulodexide. DS did not show any anticoagulant activity. In the thrombin generation assay, FMH and UFH produced comparable inhibition of thrombin generation as measured by various parameters. Sulodexide was slightly weaker in this assay, whereas DS produced relatively weaker effects. CONCLUSION: In comparison to sulodexide, both UFH and FMH exhibit comparable anticoagulant activity despite differences in their molecular weight. These results suggest that sulodexide can be developed as a parenteral anticoagulant for indications in which UFH is used.


Subject(s)
Anticoagulants/pharmacology , Blood Coagulation/drug effects , Glycosaminoglycans/pharmacology , Thrombin/pharmacology , Anticoagulants/administration & dosage , Antithrombins/administration & dosage , Antithrombins/pharmacology , Glycosaminoglycans/administration & dosage , Heparin/administration & dosage , Heparin/pharmacology , Humans , Italy , Sensitivity and Specificity , Thrombin/administration & dosage
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