RESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Morus alba L. are widely used as ethnomedicine and functional food in China, Japan, Korea and other Asian countries. Morus alba L. have a variety of pharmacological activity such as antiviral, antioxidation, anti-cholesterol, anticancer, hypoglycemia, and neuroprotection. Morus alba L. has demonstrated antiviral efficacy against influenza viruses, SARS-CoV-2 and so on, but its potential activity against pseudorabies virus (PRV) remains uncertain. AIM OF THE STUDY: This study endeavors to delve into the anti-pseudorabies virus (PRV) potential of the ethanol extract of Morus alba L. leaves (MLE), while simultaneously elucidating its underlying mechanism of action. MATERIALS AND METHODS: The anti-PRV activities of Morus alba L. extracts at different concentrations were evaluated by qPCR and immunoblotting. The inhibitory effects of MLE on PRV replication in three distinct treatment modes (pretreatment, co-treatment, and post-treatment) were detected by qPCR and indirect immunofluorescence assays. qPCR was used to investigate the effects of MLE on PRV attachment, entrance, and cytokine expression in PRV-infected cells. The chemical components in MLE were analyzed by UPLC-MS/MS. RESULTS: MLE significantly inhibits PRV replication and protein expression in a dose-dependent manner. MLE displays inhibitory effects against PRV at three different modes of treatment. The most significant inhibitory effect of MLE was observed when used in co-treatment mode, resulting in an inhibition rate of 99.42%. MLE inhibits PRV infection in the early stage. MLE inhibits PRV infection by affecting viral attachment and viral entry. Furthermore, MLE exerts its inhibition on PRV replication by mitigating the heightened expression of cytokines (TNF-α and IFN-α) triggered by PRV. Analysis of its chemical composition highlights phenolic acids and flavonoids as the principal constituents of MLE. CONCLUSION: The results illustrate that MLE effectively impedes PRV infection by suppressing viral adsorption and entry, while also curbing the expression of antiviral cytokines. Therefore, MLE may be a potential resource for creating new medications to treat human and animal PRV infections.
Assuntos
Antivirais , Herpesvirus Suídeo 1 , Morus , Extratos Vegetais , Folhas de Planta , Replicação Viral , Herpesvirus Suídeo 1/efeitos dos fármacos , Morus/química , Antivirais/farmacologia , Antivirais/isolamento & purificação , Extratos Vegetais/farmacologia , Animais , Replicação Viral/efeitos dos fármacos , Folhas de Planta/química , Citocinas/metabolismo , Cães , Células Madin Darby de Rim Canino , Internalização do Vírus/efeitos dos fármacos , Ligação Viral/efeitos dos fármacosRESUMO
BACKGROUND: Dengue virus (DENV) causes the most significant mosquito-borne viral disease with a wide spectrum of clinical manifestation, including neurological symptoms associated with lethal dengue diseases. Dopamine receptors are expressed in central nervous system, and dopamine antagonists have been reported to exhibit antiviral activity against DENV infection in vivo and in vitro. Although identification of host-cell receptor is critical to understand dengue neuropathogenesis and neurotropism, the involvement of dopamine receptors in DENV infection remains unclear. RESULTS: We exploited the sensitivity and precision of force spectroscopy to address whether dopamine type-2 receptors (D2R) directly interact with DENV particles at the first step of infection. Using optical tweezers, we quantified and characterized DENV binding to D2R expressed on Chinese hamster ovary (CHO) cells. Our finding suggested that the binding was D2R- and DENV-dependent, and that the binding force was in the range of 50-60 pN. We showed that dopamine antagonists prochlorperazine (PCZ) and trifluoperazine (TFP), previously reported to inhibit dengue infection, interrupt the DENV-D2R specific binding. CONCLUSIONS: This study demonstrates that D2R could specifically recognize DENV particles and function as an attachment factor on cell surfaces for DENV. We propose D2R as a host receptor for DENV and as a potential therapeutic target for anti-DENV drugs.
Assuntos
Cricetulus , Vírus da Dengue , Pinças Ópticas , Receptores de Dopamina D2 , Receptores de Dopamina D2/metabolismo , Vírus da Dengue/fisiologia , Vírus da Dengue/efeitos dos fármacos , Animais , Células CHO , Dengue/virologia , Ligação Proteica , Humanos , Ligação Viral/efeitos dos fármacos , Cricetinae , Antagonistas de Dopamina/farmacologiaRESUMO
The ongoing COVID-19 pandemic, caused by SARS-CoV-2, continues to pose significant global health challenges. The results demonstrated that GB-2 at 200 µg/mL effectively increased the population of 293T-ACE2 cells with low RBD binding for both SARS-CoV-2 Omicron EG.5.1 and HV.1 variants by dual-color flow cytometry, indicating its ability to inhibit virus attachment. Further investigation revealed that (+)-catechin at 25 and 50 µg/mL did not significantly alter the ACE2-RBD interaction for the EG.5.1 variant. In contrast, theaflavin showed inhibitory effects at both 25 and 50 µg/mL for EG.5.1, while only the higher concentration was effective for HV.1. Notably, theaflavin 3-gallate exhibited a potent inhibition of ACE2-RBD binding for both variants at both concentrations tested. Molecular docking studies provided insight into the binding mechanisms of theaflavin and theaflavin 3-gallate with the RBD of EG.5.1 and HV.1 variants. Both compounds showed favorable docking scores, with theaflavin 3-gallate demonstrating slightly lower scores (-8 kcal/mol) compared to theaflavin (-7 kcal/mol) for both variants. These results suggest stable interactions between the compounds and key residues in the RBD, potentially explaining their inhibitory effects on virus attachment. In conclusion, GB-2, theaflavin, and theaflavin 3-gallate demonstrate significant potential as inhibitors of the ACE2-RBD interaction in Omicron variants, highlighting their therapeutic promise against COVID-19. However, these findings are primarily based on computational and in vitro studies, necessitating further in vivo research and clinical trials to confirm their efficacy and safety in humans.
Assuntos
Enzima de Conversão de Angiotensina 2 , Antivirais , Biflavonoides , Catequina , Ligação Proteica , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Catequina/análogos & derivados , Catequina/farmacologia , Catequina/química , Enzima de Conversão de Angiotensina 2/metabolismo , Enzima de Conversão de Angiotensina 2/química , Humanos , Biflavonoides/farmacologia , Biflavonoides/química , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Antivirais/farmacologia , Antivirais/química , Simulação de Acoplamento Molecular , Células HEK293 , COVID-19/virologia , Tratamento Farmacológico da COVID-19 , Ligação Viral/efeitos dos fármacos , Enterovirus Humano B/efeitos dos fármacos , Ácido Gálico/análogos & derivadosRESUMO
BACKGROUND: Pseudorabies virus (PRV), a member of the family Herpesviridae, is responsible for significant economic losses in the pig industry and has recently been associated with human viral encephalitis, leading to severe neurological symptoms post-recovery. Despite the widespread impact of PRV, there are currently no approved effective drugs for treating PRV-related diseases in humans or pigs. Therefore, the exploration and discovery of safe and effective drugs for the prevention and treatment of PRV infection is of paramount importance. PURPOSE: The objective of this study is to screen and identify natural compounds with antiviral activity against PRV. METHODS: First, we used a strain of PRV with green fluorescent protein (PRV-GFP) to screen a natural product chemical library to identify potential antiviral drugs. Next, we assessed the antiviral abilities of salvianolic acid A (SAA) in vitro using virus titer assay, qPCR, and IFA. We investigated the mechanisms of SAA's antiviral activity through viral attachment, internalization, inactivation, and nuclease digestion assay. Finally, we evaluated the efficacy of SAA in inactivating PRV using mice as the experimental subjects. RESULTS: This study screened 206 natural compounds for anti-PRV activity in vitro, resulting in the identification of seven potential antiviral agents. Notably, SAA emerged as a promising candidate with significant anti-PRV activity. The mechanism of action may be that SAA can directly inactivate the virus by disrupting viral envelope. In vivo experiments have shown that pre-incubation of SAA and PRV can effectively inhibit the infectivity and pathogenicity of PRV in mice. CONCLUSION: This study offers valuable insights into the antiviral properties of SAA, potentially informing strategies for controlling PRV epidemics and treating related diseases in both humans and animals.
Assuntos
Antivirais , Herpesvirus Suídeo 1 , Pseudorraiva , Herpesvirus Suídeo 1/efeitos dos fármacos , Animais , Antivirais/farmacologia , Pseudorraiva/tratamento farmacológico , Pseudorraiva/virologia , Camundongos , Ácidos Cafeicos/farmacologia , Lactatos/farmacologia , Internalização do Vírus/efeitos dos fármacos , Vírion/efeitos dos fármacos , Suínos , Ligação Viral/efeitos dos fármacos , Linhagem Celular , FemininoRESUMO
Plant polyphenols possess diverse bioactivities, including antiviral activity against a broad spectrum of viruses. Here, we investigated the virucidal properties of an Kalanchoe daigremontiana extract using an in vitro model of human herpesvirus type 1 (HHV-1) infection. Chromatographic analysis indicated that the extract of Kalanchoe daigremontiana is rich in various compounds, among which are polyphenols with virucidal activity confirmed in the literature. We found that Kalanchoe daigremontiana extract shows an ability to prevent HHV-1 infection by direct inhibition of the virus attachment, penetration, and blocking of infection when used in pretreatment or post-entry treatment. Our results indicate that Kalanchoe daigremontiana extract may be a good candidate drug against HHV-1, both as a substance to prevent infection and to treat an already ongoing infection. Our findings illustrate that Kalanchoe daigremontiana could be a potential new candidate for clinical consideration in the treatment of HHV-1 infection alone or in combination with other therapeutics.
Assuntos
Antivirais , Herpesvirus Humano 1 , Kalanchoe , Extratos Vegetais , Extratos Vegetais/farmacologia , Extratos Vegetais/química , Antivirais/farmacologia , Antivirais/química , Kalanchoe/química , Herpesvirus Humano 1/efeitos dos fármacos , Herpesvirus Humano 1/fisiologia , Humanos , Polifenóis/farmacologia , Polifenóis/química , Internalização do Vírus/efeitos dos fármacos , Ligação Viral/efeitos dos fármacosRESUMO
SARS-CoV-2 infection starts from the association of its spike 1 (S1) subunit with sensitive cells. Vesicular endothelial cells and platelets are among the cell types that bind SARS-CoV-2, but the effectors that mediate viral attachment on the cell membrane have not been fully elucidated. Herein, we show that P-selectin (SELP), a biomarker for endothelial dysfunction and platelet activation, can facilitate the attachment of SARS-CoV-2 S1. Since we observe colocalization of SELP with S1 in the lung tissues of COVID-19 patients, we perform molecular biology experiments on human umbilical vein endothelial cells (HUVECs) to confirm the intermolecular interaction between SELP and S1. SELP overexpression increases S1 recruitment to HUVECs and enhances SARS-CoV-2 spike pseudovirion infection. The opposite results are determined after SELP downregulation. As S1 causes endothelial inflammatory responses in a dose-dependent manner, by activating the interleukin (IL)-17 signaling pathway, SELP-induced S1 recruitment may contribute to the development of a "cytokine storm" after viral infection. Furthermore, SELP also promotes the attachment of S1 to the platelet membrane. Employment of PSI-697, a small inhibitor of SELP, markedly decreases S1 adhesion to both HUVECs and platelets. In addition to the role of membrane SELP in facilitating S1 attachment, we also discover that soluble SELP is a prognostic factor for severe COVID-19 through a meta-analysis. In this study, we identify SELP as an adhesive site for the SARS-CoV-2 S1, thus providing a potential drug target for COVID-19 treatment.
Assuntos
Plaquetas , COVID-19 , Células Endoteliais da Veia Umbilical Humana , Selectina-P , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Humanos , Glicoproteína da Espícula de Coronavírus/metabolismo , Selectina-P/metabolismo , SARS-CoV-2/fisiologia , COVID-19/virologia , Plaquetas/metabolismo , Ligação Viral/efeitos dos fármacosRESUMO
Porcine epidemic diarrhea virus (PEDV) remains one of the major causative microorganisms of viral diarrhea in piglets worldwide, with no approved drugs for treatment. We identified a natural molecule, flavonol, which is widely found in tea, vegetables and herbs. Subsequently, the antiviral activity of compound flavonol was evaluated in Vero cells and IPEC-J2 cells, and its anti-PEDV mechanism was analyzed by molecular docking and molecular dynamics. The results showed that flavonol could effectively inhibit viral progeny production, RNA synthesis and protein expression of PEDV strains in a dose-dependent manner. When flavonol was added simultaneously with viral infection in Vero cells, it demonstrated potent anti-PEDV activity by affecting the viral attachment and internalization phases. Similarly, in IPEC-J2 cells, flavonol effectively inhibited PEDV infection at different stages of infection, except for the release phase. Moreover, flavonol mainly interacts with PEDV Mpro through hydrogen bonds and hydrophobic forces, and the complex formed by it has high stability. Importantly, flavonol also showed broad-spectrum activity against other porcine enteric coronaviruses such as TGEV and PDCoV in vitro. These findings suggest that flavonol may exert antiviral effects by interacting with viral Mpro, thereby affecting viral replication. This means that flavonol is expected to become a potential drug to prevent or treat porcine enteric coronavirus.
Assuntos
Antivirais , Flavonóis , Vírus da Diarreia Epidêmica Suína , Replicação Viral , Vírus da Diarreia Epidêmica Suína/efeitos dos fármacos , Animais , Antivirais/farmacologia , Antivirais/química , Flavonóis/farmacologia , Chlorocebus aethiops , Suínos , Células Vero , Replicação Viral/efeitos dos fármacos , Simulação de Acoplamento Molecular , Infecções por Coronavirus/virologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/tratamento farmacológico , Internalização do Vírus/efeitos dos fármacos , Doenças dos Suínos/virologia , Doenças dos Suínos/tratamento farmacológico , Linhagem Celular , Simulação de Dinâmica Molecular , Ligação Viral/efeitos dos fármacosRESUMO
Gu-Sui-Bu, the dried rhizome of Davallia mariesii, is a traditional Chinese herbal remedy with a significant history of treating osteoporosis and inflammatory conditions. However, its potential as an anti-influenza agent and its underlying mechanisms of action remain unexplored. To obtain a more potent extract from D. mariesii and gain insights into its mechanism of action against influenza A virus (IAV), we utilized a partitioning process involving organic solvents and water, resulting in the isolation of butanolic subfractions of the D. mariesii extract (DMBE). DMBE exhibited a broad anti-viral spectrum, effectively inhibiting IAV, with an EC50 of 24.32 ± 6.19 µg/mL and a selectivity index of 6.05. We subsequently conducted a series of in vitro assays to evaluate the antiviral effects of DMBE and to uncover its mechanisms of action. DMBE was found to inhibit IAV during the early stages of infection by hindering the attachment of the virus onto and its penetration into host cells. Importantly, DMBE was observed to hinder IAV-mediated cell-cell fusion. It also inhibited neuraminidase activity, plaque size, and the expression levels of phospho-AKT. In summary, this study provides evidence for the effectiveness of D. mariesii as a complementary and alternative herbal remedy against IAV. Specifically, our data highlight DMBE's capabilities in inhibiting viral entry and the release of virions.
Assuntos
Antivirais , Vírus da Influenza A , Extratos Vegetais , Antivirais/farmacologia , Antivirais/química , Vírus da Influenza A/efeitos dos fármacos , Vírus da Influenza A/fisiologia , Humanos , Extratos Vegetais/farmacologia , Extratos Vegetais/química , Animais , Células Madin Darby de Rim Canino , Cães , Internalização do Vírus/efeitos dos fármacos , Sapindaceae/química , Replicação Viral/efeitos dos fármacos , Ligação Viral/efeitos dos fármacos , Influenza Humana/tratamento farmacológico , Influenza Humana/virologia , Medicamentos de Ervas Chinesas/farmacologia , Medicamentos de Ervas Chinesas/química , Neuraminidase/metabolismo , Células A549 , Linhagem CelularRESUMO
Rotavirus infections in suckling and weaning piglets cause severe dehydration and death, resulting in significant economic losses in the pig breeding industry. With the continuous emergence of porcine rotavirus (PoRV) variants and poor vaccine cross-protection among various genotypes, there is an urgent need to develop alternative strategies such as seeking effective antiviral products from nature, microbial metabolites and virus-host protein interaction. Sialidases play a crucial role in various physiopathological processes and offer a promising target for developing antivirus drugs. However, the effect of bacterial-derived sialidases on the infection of PoRVs remains largely unknown. Herein, we investigated the impact of bacterial-derived sialidases (sialidase Cp and Vc) on PoRV strain OSU(Group A) infection, using differentiated epithelial monkey kidney cells (MA104) as a model. Our results indicated that the pretreatment of MA104 with exogenous sialidases effectively suppressed PoRV OSU in a concentration-dependent manner. Notably, even at a concentration of 0.01 µU/mL, sialidases significantly inhibited the virus (MOI = 0.01). Meanwhile, we found that sialidase Vc pretreatment sharply reduced the binding rate of PoRV OSU. Last, we demonstrated that PoRV OSU might recognize α-2,3-linked sialic acid as the primary attachment factor in MA104. Our findings provide new insights into the underlying mechanism of PoRV OSU infections, shedding lights on the development of alternative antivirus approaches based on bacteria-virus interaction.
Assuntos
Neuraminidase , Infecções por Rotavirus , Rotavirus , Replicação Viral , Animais , Neuraminidase/metabolismo , Neuraminidase/genética , Rotavirus/efeitos dos fármacos , Rotavirus/fisiologia , Suínos , Replicação Viral/efeitos dos fármacos , Linhagem Celular , Células Epiteliais/virologia , Células Epiteliais/microbiologia , Ligação Viral/efeitos dos fármacos , Ácido N-Acetilneuramínico/metabolismo , Ácido N-Acetilneuramínico/farmacologia , Antivirais/farmacologia , Haplorrinos , Doenças dos Suínos/virologia , Doenças dos Suínos/microbiologiaRESUMO
Entry of influenza A viruses (IAVs) into host cells is initiated by binding to sialic acids (Sias), their primary host cell receptor, followed by endocytosis and membrane fusion to release the viral genome into the cytoplasm of the host cell. Host tropism is affected by these entry processes, with a primary factor being receptor specificity. Sias exist in several different chemical forms, including the hydroxylated N-glycolylneuraminic acid (Neu5Gc), which is found in many hosts; however, it has not been clear how modified Sias affect viral binding and entry. Neu5Gc is commonly found in many natural influenza hosts, including pigs and horses, but not in humans or ferrets. Here, we engineered HEK293 cells to express the hydoxylase gene (CMAH) that converts Neu5Ac to Neu5Gc, or knocked out the Sia-CMP transport gene (SLC35A1), resulting in cells that express 95% Neu5Gc or minimal level of Sias, respectively. H3N2 (X-31) showed significantly reduced infectivity in Neu5Gc-rich cells compared to wild-type HEK293 (>95% Neu5Ac). To determine the effects on binding and fusion, we generated supported lipid bilayers (SLBs) derived from the plasma membranes of these cells and carried out single particle microscopy. H3N2 (X-31) exhibited decreased binding to Neu5Gc-containing SLBs, but no significant difference in H3N2 (X-31)'s fusion kinetics to either SLB type, suggesting that reduced receptor binding does not affect subsequent membrane fusion. This finding suggests that for this virus to adapt to host cells rich in Neu5Gc, only receptor affinity changes are required without further adaptation of virus fusion machinery. IMPORTANCE Influenza A virus (IAV) infections continue to threaten human health, causing over 300,000 deaths yearly. IAV infection is initiated by the binding of influenza glycoprotein hemagglutinin (HA) to host cell sialic acids (Sias) and the subsequent viral-host membrane fusion. Generally, human IAVs preferentially bind to the Sia N-acetylneuraminic acid (Neu5Ac). Yet, other mammalian hosts, including pigs, express diverse nonhuman Sias, including N-glycolylneuraminic acid (Neu5Gc). The role of Neu5Gc in human IAV infections in those hosts is not well-understood, and the variant form may play a role in incidents of cross-species transmission and emergence of new epidemic variants. Therefore, it is important to investigate how human IAVs interact with Neu5Ac and Neu5Gc. Here, we use membrane platforms that mimic the host cell surface to examine receptor binding and membrane fusion events of human IAV H3N2. Our findings improve the understanding of viral entry mechanisms that can affect host tropism and virus evolution.
Assuntos
Interações entre Hospedeiro e Microrganismos , Vírus da Influenza A Subtipo H3N2 , Ácidos Siálicos , Internalização do Vírus , Animais , Humanos , Células HEK293 , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza A Subtipo H3N2/metabolismo , Fusão de Membrana , Proteínas de Transporte de Nucleotídeos/genética , Proteínas de Transporte de Nucleotídeos/metabolismo , Ácidos Siálicos/química , Ácidos Siálicos/farmacologia , Imagem Individual de Molécula , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Interações entre Hospedeiro e Microrganismos/genética , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/virologiaRESUMO
Two years since the outbreak of the novel coronavirus SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) pandemic, there remain few clinically effective drugs to complement vaccines. One is the anticoagulant, heparin, which in 2004 was found able to inhibit invasion of SARS-CoV (CoV-1) and which has been employed during the current pandemic to prevent thromboembolic complications and moderate potentially damaging inflammation. Heparin has also been shown experimentally to inhibit SARS-CoV-2 attachment and infection in susceptible cells. At high therapeutic doses however, heparin increases the risk of bleeding and prolonged use can cause heparin-induced thrombocytopenia, a serious side effect. One alternative, with structural similarities to heparin, is the plant-derived, semi-synthetic polysaccharide, pentosan polysulfate (PPS). PPS is an established drug for the oral treatment of interstitial cystitis, is well-tolerated, and exhibits weaker anticoagulant effects than heparin. In an established Vero cell model, PPS and its fractions of varying molecular weights inhibited invasion by SARS-CoV-2. Intact PPS and its size-defined fractions were characterized by molecular weight distribution and chemical structure using nuclear magnetic resonance spectroscopy and liquid chromatography-mass spectrometry, then employed to explore the structural basis of interactions with SARS-CoV-2 spike protein receptor-binding domain (S1 RBD) and the inhibition of Vero cell invasion. PPS was as effective as unfractionated heparin, but more effective in inhibiting cell infection than low-molecular-weight heparin (on a weight/volume basis). Isothermal titration calorimetry and viral plaque-forming assays demonstrated size-dependent binding to S1 RBD and inhibition of Vero cell invasion, suggesting the potential application of PPS as a novel inhibitor of SARS-CoV-2 infection.
Assuntos
Poliéster Sulfúrico de Pentosana , SARS-CoV-2 , Ligação Viral , Animais , Anticoagulantes/farmacologia , Chlorocebus aethiops , Heparina/uso terapêutico , Poliéster Sulfúrico de Pentosana/farmacologia , Ligação Proteica , SARS-CoV-2/efeitos dos fármacos , Glicoproteína da Espícula de Coronavírus , Células Vero , Ligação Viral/efeitos dos fármacosRESUMO
Porcine pseudorabies (PR) is an important infectious disease caused by pseudorabies virus (PRV), which poses a major threat to food safety and security. Vaccine immunization has become the main means to prevent and control the disease. However, since 2011, a new PRV variant has caused huge economic losses to the Chinese pig industry. Panax notoginseng polysaccharides have immunomodulatory activity and other functions, but the antiviral effect has not been reported. We studied the anti-PRV activity of Panax notoginseng polysaccharides in vitro. A less cytopathic effect was observed by increasing the concentration of Panax notoginseng polysaccharides. Western blot, TCID50, plaque assay, and IFA revealed that Panax notoginseng polysaccharides could significantly inhibit the infectivity of PRV XJ5 on PK15 cells. In addition, we also found that Panax notoginseng polysaccharides blocked the adsorption and replication of PRV to PK15 cells in a dose-dependent manner. These results show that Panax notoginseng polysaccharides play an antiviral effect mainly by inhibiting virus adsorption and replication in vitro. Therefore, Panax notoginseng polysaccharides may be a potential anti-PRV agent.
Assuntos
Herpesvirus Suídeo 1/fisiologia , Fatores Imunológicos/farmacologia , Panax notoginseng/química , Polissacarídeos/farmacologia , Pseudorraiva/metabolismo , Doenças dos Suínos/metabolismo , Ligação Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Animais , Linhagem Celular , Fatores Imunológicos/química , Polissacarídeos/química , Pseudorraiva/tratamento farmacológico , Pseudorraiva/patologia , Suínos , Doenças dos Suínos/patologia , Doenças dos Suínos/virologiaRESUMO
The continuous emergence of severe acute respiratory coronavirus 2 (SARS-CoV-2) variants and the increasing number of breakthrough infection cases among vaccinated people support the urgent need for research and development of antiviral drugs. Viral entry is an intriguing target for antiviral drug development. We found that diltiazem, a blocker of the L-type calcium channel Cav1.2 pore-forming subunit (Cav1.2 α1c) and an FDA-approved drug, inhibits the binding and internalization of SARS-CoV-2, and decreases SARS-CoV-2 infection in cells and mouse lung. Cav1.2 α1c interacts with SARS-CoV-2 spike protein and ACE2, and affects the attachment and internalization of SARS-CoV-2. Our finding suggests that diltiazem has potential as a drug against SARS-CoV-2 infection and that Cav1.2 α1c is a promising target for antiviral drug development for COVID-19.
Assuntos
Tratamento Farmacológico da COVID-19 , COVID-19 , Diltiazem/farmacologia , Pulmão/efeitos dos fármacos , SARS-CoV-2/efeitos dos fármacos , Células A549 , Animais , COVID-19/patologia , COVID-19/virologia , Células Cultivadas , Chlorocebus aethiops , Diltiazem/uso terapêutico , Modelos Animais de Doenças , Feminino , Células HEK293 , Células HeLa , Humanos , Pulmão/patologia , Pulmão/virologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , SARS-CoV-2/fisiologia , Células Vero , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacosRESUMO
Since the beginning of the HIV epidemic, lasting more than 30 years, the main goal of scientists was to develop effective methods for the prevention and treatment of HIV infection. Modern medicines have reduced the death rate from AIDS by 80%. However, they still have side effects and are very expensive, dictating the need to search for new drugs. Earlier, it was shown that phospholipases A2 (PLA2s) from bee and snake venoms block HIV replication, the effect being independent on catalytic PLA2 activity. However, the antiviral activity of human PLA2s against Lentiviruses depended on catalytic function and was mediated through the destruction of the viral membrane. To clarify the role of phospholipolytic activity in antiviral effects, we analyzed the anti-HIV activity of several snake PLA2s and found that the mechanisms of their antiviral activity were similar to that of mammalian PLA2. Our results indicate that snake PLA2s are capable of inhibiting syncytium formation between chronically HIV-infected cells and healthy CD4-positive cells and block HIV binding to cells. However, only dimeric PLA2s had pronounced virucidal and anti-HIV activity, which depended on their catalytic activity. The ability of snake PLA2s to inactivate the virus may provide an additional barrier to HIV infection. Thus, snake PLA2s might be considered as candidates for lead molecules in anti-HIV drug development.
Assuntos
Fármacos Anti-HIV/farmacologia , Linfócitos T CD4-Positivos/citologia , Células Gigantes/citologia , HIV-1/fisiologia , Fosfolipases A2/farmacologia , Venenos de Serpentes/enzimologia , Serpentes/metabolismo , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/virologia , Linhagem Celular , Células Cultivadas , Células Gigantes/efeitos dos fármacos , Células Gigantes/virologia , HIV-1/efeitos dos fármacos , Humanos , Concentração Inibidora 50 , Proteínas de Répteis/farmacologia , Serpentes/classificação , Ativação Viral/efeitos dos fármacos , Ligação Viral/efeitos dos fármacosRESUMO
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.
Assuntos
Antivirais/farmacologia , Infecções por Coronavirus/veterinária , Coronavirus Bovino/efeitos dos fármacos , Glicosaminoglicanos/farmacologia , SARS-CoV-2/efeitos dos fármacos , Animais , Bovinos , Linhagem Celular , Sulfatos de Condroitina/química , Sulfatos de Condroitina/farmacologia , Infecções por Coronavirus/tratamento farmacológico , Glicosaminoglicanos/química , Glicosaminoglicanos/metabolismo , Humanos , Ácido Hialurônico/química , Ácido Hialurônico/farmacologia , Sulfatos/química , Sulfatos/farmacologia , Ligação Viral/efeitos dos fármacos , Tratamento Farmacológico da COVID-19RESUMO
The severe acute respiratory syndrome coronavirus 2 responsible for COVID-19 remains a persistent threat to mankind, especially for the immunocompromised and elderly for which the vaccine may have limited effectiveness. Entry of SARS-CoV-2 requires a high affinity interaction of the viral spike protein with the cellular receptor angiotensin-converting enzyme 2. Novel mutations on the spike protein correlate with the high transmissibility of new variants of SARS-CoV-2, highlighting the need for small molecule inhibitors of virus entry into target cells. We report the identification of such inhibitors through a robust high-throughput screen testing 15,000 small molecules from unique libraries. Several leads were validated in a suite of mechanistic assays, including whole cell SARS-CoV-2 infectivity assays. The main lead compound, calpeptin, was further characterized using SARS-CoV-1 and the novel SARS-CoV-2 variant entry assays, SARS-CoV-2 protease assays and molecular docking. This study reveals calpeptin as a potent and specific inhibitor of SARS-CoV-2 and some variants.
Assuntos
Antivirais/farmacologia , Tratamento Farmacológico da COVID-19 , Inibidores de Cisteína Proteinase/farmacologia , Dipeptídeos/farmacologia , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Enzima de Conversão de Angiotensina 2/metabolismo , Animais , Catepsina L/antagonistas & inibidores , Linhagem Celular , Chlorocebus aethiops , Avaliação Pré-Clínica de Medicamentos , Reposicionamento de Medicamentos , Células HEK293 , Humanos , Simulação de Acoplamento Molecular , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/crescimento & desenvolvimento , Serina Endopeptidases/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Células VeroRESUMO
Increasing outbreaks of new pathogenic viruses have promoted the exploration of novel alternatives to time-consuming vaccines. Thus, it is necessary to develop a universal approach to halt the spread of new and unknown viruses as they are discovered. One such promising approach is to target lipid membranes, which are common to all viruses and bacteria. The ongoing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has reaffirmed the importance of interactions between the virus envelope and the host cell plasma membrane as a critical mechanism of infection. Metadichol®, a nanolipid emulsion of long-chain alcohols, has been demonstrated as a strong candidate that inhibits the proliferation of SARS-CoV-2. Naturally derived substances, such as long-chain saturated lipid alcohols, reduce viral infectivity, including that of coronaviruses (such as SARS-CoV-2) by modifying their lipid-dependent attachment mechanism to human host cells. The receptor ACE2 mediates the entry of SARS-CoV-2 into the host cells, whereas the serine protease TMPRSS2 primes the viral S protein. In this study, Metadichol® was found to be 270 times more potent an inhibitor of TMPRSS2 (EC50 = 96 ng/mL) than camostat mesylate (EC50 = 26000 ng/mL). Additionally, it inhibits ACE with an EC50 of 71 ng/mL, but it is a very weak inhibitor of ACE2 at an EC50 of 31 µg/mL. Furthermore, the live viral assay performed in Caco-2 cells revealed that Metadichol® inhibits SARS-CoV-2 replication at an EC90 of 0.16 µg/mL. Moreover, Metadichol® had an EC90 of 0.00037 µM, making it 2081 and 3371 times more potent than remdesivir (EC50 = 0.77 µM) and chloroquine (EC50 = 1.14 µM), respectively.
Assuntos
Álcoois Graxos/farmacologia , Sistemas de Liberação de Fármacos por Nanopartículas/farmacologia , SARS-CoV-2/efeitos dos fármacos , Vírus/efeitos dos fármacos , Animais , Antivirais/farmacologia , Linhagem Celular , Chlorocebus aethiops , Ésteres/farmacologia , Guanidinas/farmacologia , Humanos , Metabolismo dos Lipídeos/fisiologia , Lipídeos/química , SARS-CoV-2/metabolismo , SARS-CoV-2/patogenicidade , Serina Endopeptidases/efeitos dos fármacos , Serina Endopeptidases/metabolismo , Serina Proteases/metabolismo , Inibidores de Serina Proteinase/farmacologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Células Vero , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Tratamento Farmacológico da COVID-19RESUMO
GSK3732394 is a multi-specific biologic inhibitor of HIV entry currently under clinical evaluation. A key component of this molecule is an adnectin (6940_B01) that binds to CD4 and inhibits downstream actions of gp160. Studies were performed to determine the binding site of the adnectin on CD4 and to understand the mechanism of inhibition. Using hydrogen-deuterium exchange with mass spectrometry (HDX), CD4 peptides showed differential rates of deuteration (either enhanced or slowed) in the presence of the adnectin that mapped predominantly to the interface of domains 2 and 3 (D2-D3). In addition, an X-ray crystal structure of an ibalizumab Fab/CD4(D1-D4)/adnectin complex revealed an extensive interface between the adnectin and residues on CD4 domains D2-D4 that stabilize a novel T-shaped CD4 conformation. A cryo-EM map of the gp140/CD4/GSK3732394 complex clearly shows the bent conformation for CD4 while bound to gp140. Mutagenic analyses on CD4 confirmed that amino acid F202 forms a key interaction with the adnectin. In addition, amino acid L151 was shown to be a critical indirect determinant of the specificity for binding to the human CD4 protein over related primate CD4 molecules, as it appears to modulate CD4's flexibility to adopt the adnectin-bound conformation. The significant conformational change of CD4 upon adnectin binding brings the D1 domain of CD4 in proximity to the host cell membrane surface, thereby re-orienting the gp120 binding site in a direction that is inaccessible to incoming virus due to a steric clash between gp160 trimers on the virus surface and the target cell membrane.
Assuntos
Fármacos Anti-HIV/farmacologia , Antígenos CD4/química , Antígenos CD4/metabolismo , HIV-1/metabolismo , Ligação Viral/efeitos dos fármacos , Animais , Anticorpos Monoclonais , Sítios de Ligação , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Domínios Proteicos , Internalização do Vírus/efeitos dos fármacosRESUMO
Epimedium koreanum Nakai (EKN) is a popular plant in Korean and Chinese medicine for treating a variety of ailments. The aqueous extract of EKN has a significant inhibitory impact on influenza A virus (IAV) infection by directly blocking viral attachment and having a virucidal effect, according to this study. Using fluorescent microscopy and fluorescence-activated cell sorting (FACS) with a green fluorescent protein (GFP)-tagged Influenza A/PR/8/34 virus, we examined the effect of EKN on viral infection. By viral infection, EKN strongly suppresses GFP expression, and at a dosage of 100 µg/mL, EKN decreased GFP expression by up to 90% of the untreated infected control. Immunofluorescence and Western blot analyses against influenza viral proteins revealed that EKN decreased influenza viral protein expression in a dose-dependent manner. EKN inhibited the H1N1 influenza virus's hemagglutinin (HA) and neuraminidase (NA), preventing viral attachment to cells. Furthermore, EKN had a virucidal impact and inhibited the cytopathic effects of H1N1, H3N2 and influenza B virus infection. Finally, our findings show that EKN has the potential to be developed as a natural viral inhibitor against influenza virus infection.
Assuntos
Alphainfluenzavirus/efeitos dos fármacos , Antivirais/farmacologia , Epimedium , Extratos Vegetais/farmacologia , Animais , Hemaglutininas/efeitos dos fármacos , Humanos , Camundongos , Neuraminidase/efeitos dos fármacos , Proteínas Virais/efeitos dos fármacos , Ligação Viral/efeitos dos fármacosRESUMO
OBJECTIVE: With the recent direction in drug repurposing, many approved drugs have been evaluated to assess their effect on the coronavirus or SARS-CoV-2 infection (COVID-19). Driving this path, chloroquine (CQ) has been used in the treatment of malaria and hydroxychloroquine (HCQ) in immunomodulatory and anti-thrombotic action, playing a leading role in initial management of the viral infection. MATERIALS AND METHODS: Literature search was done using Google Scholar, PubMed and Scopus database using keywords "chloroquine" "SARS-CoV-2" "COVID-19" "mechanism of action" and articles of interest were selected providing evidence of the possible role of CQ in viral infection. RESULTS: In a bid to understand how and if CQ and HCQ would exert their anti-viral property, mechanistic exegesis was done to review various proposed mechanisms of action. This revealed the inhibition of viral attachment and entry, inhibition of enveloped glycoprotein, inhibition of the development and proliferation of new viral particles as the way they perform their action. There is an interplay between iron metabolism and homeostasis with COVID-19 infection and viral reproduction. CONCLUSIONS: This study aims to show the functional role of CQ and HCQ, as well as to provide possible mechanistic insight on the role of iron on viral infection, iron starvation and its downstream cellular pathways involving hepcidin and proinflammatory cytokines. The overall aim of providing possible mode of action of CQ and HCQ in the management of COVID-19 infection is exhibited via its anti-viral, anti-inflammatory and anti-thrombotic activities.