RESUMEN
Outbreaks of acute respiratory viral diseases, such as influenza and COVID-19 caused by influenza A virus (IAV) and SARS-CoV-2, pose a serious threat to global public health, economic security, and social stability. This calls for the development of broad-spectrum antivirals to prevent or treat infection or co-infection of IAV and SARS-CoV-2. Hemagglutinin (HA) on IAV and spike (S) protein on SARS-CoV-2, which contain various types of glycans, play crucial roles in mediating viral entry into host cells. Therefore, they are key targets for the development of carbohydrate-binding protein-based antivirals. This study demonstrated that griffithsin (GRFT) and the GRFT-based bivalent entry inhibitor GL25E (GRFT-L25-EK1) showed broad-spectrum antiviral effects against IAV infection in vitro by binding to HA in a carbohydrate-dependent manner and effectively protected mice from lethal IAV infection. Although both GRFT and GL25E could inhibit infection of SARS-CoV-2 Omicron variants, GL25E proved to be significantly more effective than GRFT and EK1 alone. Furthermore, GL25E effectively inhibited in vitro co-infection of IAV and SARS-CoV-2 and demonstrated good druggability, including favorable safety and stability profiles. These findings suggest that GL25E is a promising candidate for further development as a broad-spectrum antiviral drug for the prevention and treatment of infection or co-infection from IAV and SARS-CoV-2.IMPORTANCEInfluenza and COVID-19 are highly contagious respiratory illnesses caused by the influenza A virus (IAV) and SARS-CoV-2, respectively. IAV and SARS-CoV-2 co-infection exacerbates damage to lung tissue and leads to more severe clinical symptoms, thus calling for the development of broad-spectrum antivirals for combating IAV and SARS-CoV-2 infection or co-infection. Here we found that griffithsin (GRFT), a carbohydrate-binding protein, and GL25E, a recombinant protein consisting of GRFT, a 25 amino acid linker, and EK1, a broad-spectrum coronavirus inhibitor, could effectively inhibit IAV and SARS-CoV-2 infection and co-infection by targeting glycans on HA of IAV and spike (S) protein of SARS-CoV-2. GL25E is more effective than GRFT because GL25E can also interact with the HR1 domain in SARS-CoV-2 S protein. Furthermore, GL25E possesses favorable safety and stability profiles, suggesting that it is a promising candidate for development as a drug to prevent and treat IAV and SARS-CoV-2 infection or co-infection.
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Antivirales , COVID-19 , Coinfección , Virus de la Influenza A , Lectinas de Plantas , SARS-CoV-2 , Internalización del Virus , Animales , Antivirales/farmacología , Virus de la Influenza A/efectos de los fármacos , Ratones , SARS-CoV-2/efectos de los fármacos , Humanos , Internalización del Virus/efectos de los fármacos , Coinfección/tratamiento farmacológico , Coinfección/virología , Lectinas de Plantas/farmacología , COVID-19/virología , Glicoproteína de la Espiga del Coronavirus/metabolismo , Glicoproteína de la Espiga del Coronavirus/antagonistas & inhibidores , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Infecciones por Orthomyxoviridae/virología , Tratamiento Farmacológico de COVID-19 , Perros , Ratones Endogámicos BALB C , Femenino , Glicoproteínas Hemaglutininas del Virus de la Influenza/metabolismo , Gripe Humana/tratamiento farmacológico , Gripe Humana/virología , Células de Riñón Canino Madin DarbyRESUMEN
Hantaan virus (HTNV) is the etiological pathogen of hemorrhagic fever with renal syndrome in East Asia. There are currently no effective therapeutics approved for HTNV and other hantavirus infections. We found that griffithsin (GRFT), an algae-derived lectin with broad-spectrum antiviral activity against various enveloped viruses, can inhibit the growth and spread of HTNV. In vitro experiments using recombinant vesicular stomatitis virus (rVSV) with HTNV glycoproteins as a model revealed that the GRFT inhibited the entry of rVSV-HTNV-G into host cells. In addition, we demonstrated that GRFT prevented authentic HTNV infection in vitro by binding to the viral N-glycans. In vivo experiments showed that GRFT partially protected the suckling mice from death induced by intracranial exposure to HTNV. These results demonstrated that GRFT can be a promising agent for inhibiting HTNV infection.
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Virus Hantaan , Infecciones por Hantavirus , Fiebre Hemorrágica con Síndrome Renal , Animales , Ratones , Lectinas/farmacología , Fiebre Hemorrágica con Síndrome Renal/tratamiento farmacológicoRESUMEN
Candida albicans is a fungal pathogen that can form biofilms on medical devices and host tissue, resulting in serious, life-threatening infections. These fungal biofilms are inherently resistant to traditional antifungal therapies and the host immune system; therefore, biofilm-associated infections are a huge clinical challenge. This review summarizes the most important insights into C. albicans biofilm-associated antifungal drug resistance mechanisms and immune evasion strategies. In addtion, this review also discusses the strategies for antifungal drug use to combat these processes, providing further evidence for novel drugs research and clinical therapies.
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Candida albicans , Evasión Inmune , Farmacorresistencia Fúngica , Antifúngicos/farmacología , Antifúngicos/uso terapéutico , BiopelículasRESUMEN
The development of broad-spectrum antivirals against human coronaviruses (HCoVs) is critical to combat the current coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its variants, as well as future outbreaks of emerging CoVs. We have previously identified a polyethylene glycol-conjugated (PEGylated) lipopeptide, EK1C4, with potent pan-CoV fusion inhibitory activity. However, PEG linkers in peptide or protein drugs may reduce stability or induce anti-PEG antibodies in vivo. Therefore, we herein report the design and synthesis of a series of dePEGylated lipopeptide-based pan-CoV fusion inhibitors featuring the replacement of the PEG linker with amino acids in the heptad repeat 2 C-terminal fragment (HR2-CF) of HCoV-OC43. Among these lipopeptides, EKL1C showed the most potent inhibitory activity against infection by SARS-CoV-2 and its spike (S) mutants, as well as other HCoVs and some bat SARS-related coronaviruses (SARSr-CoVs) tested. The dePEGylated lipopeptide EKL1C exhibited significantly stronger resistance to proteolytic enzymes, better metabolic stability in mouse serum, higher thermostability than the PEGylated lipopeptide EK1C4, suggesting that EKL1C could be further developed as a candidate prophylactic and therapeutic for COVID-19 and other coronavirus diseases.
RESUMEN
The COVID-19 pandemic caused by SARS-CoV-2 infection poses a serious threat to global public health and the economy. The enzymatic product of cholesterol 25-hydroxylase (CH25H), 25-Hydroxycholesterol (25-HC), was reported to have potent anti-SARS-CoV-2 activity. Here, we found that the combination of 25-HC with EK1 peptide, a pan-coronavirus (CoV) fusion inhibitor, showed a synergistic antiviral activity. We then used the method of 25-HC modification to design and synthesize a series of 25-HC-modified peptides and found that a 25-HC-modified EK1 peptide (EK1P4HC) was highly effective against infections caused by SARS-CoV-2, its variants of concern (VOCs), and other human CoVs, such as HCoV-OC43 and HCoV-229E. EK1P4HC could protect newborn mice from lethal HCoV-OC43 infection, suggesting that conjugation of 25-HC with a peptide-based viral inhibitor was a feasible and universal strategy to improve its antiviral activity.
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Antivirales/farmacología , Hidroxicolesteroles/química , Lipopéptidos/química , SARS-CoV-2/efectos de los fármacos , Secuencia de Aminoácidos , Animales , Antivirales/química , Antivirales/uso terapéutico , Peso Corporal/efectos de los fármacos , COVID-19/virología , Coronavirus Humano 229E/efectos de los fármacos , Coronavirus Humano 229E/patogenicidad , Infecciones por Coronavirus/tratamiento farmacológico , Infecciones por Coronavirus/mortalidad , Infecciones por Coronavirus/virología , Coronavirus Humano OC43/efectos de los fármacos , Coronavirus Humano OC43/patogenicidad , Modelos Animales de Enfermedad , Sinergismo Farmacológico , Humanos , Hidroxicolesteroles/farmacología , Hidroxicolesteroles/uso terapéutico , Lipopéptidos/farmacología , Lipopéptidos/uso terapéutico , Ratones , Ratones Endogámicos BALB C , Polietilenglicoles/química , SARS-CoV-2/aislamiento & purificación , SARS-CoV-2/fisiología , Tasa de Supervivencia , Internalización del Virus/efectos de los fármacos , Tratamiento Farmacológico de COVID-19RESUMEN
BACKGROUND: Our previous studies have shown that combining the antiviral lectin GRFT and the pan-CoV fusion inhibitory peptide EK1 results in highly potent inhibitory activity against SARS-CoV-2 infection. In this study, we aimed to design and construct a bivalent protein consisting of GRFT and EK1 components and evaluate its inhibitory activity and mechanism of action against infection by SARS-CoV-2 and its mutants, as well as other human coronaviruses (HCoVs). METHODS: The bivalent proteins were expressed in E. coli and purified with Ni-NTA column. HIV backbone-based pseudovirus (PsV) infection and HCoV S-mediated cell-cell fusion assays were performed to test their inhibitory activity. ELISA and Native-PAGE were conducted to illustrate the mechanism of action of these bivalent proteins. Five-day-old newborn mice were intranasally administrated with a selected bivalent protein before or after HCoV-OC43 challenge, and its protective effect was monitored for 14 days. RESULTS: Among the three bivalent proteins purified, GL25E exhibited the most potent inhibitory activity against infection of SARS-CoV-2 PsVs expressing wild-type and mutated S protein. GL25E was significantly more effective than GRFT and EK1 alone in inhibiting HCoV S-mediated cell-cell fusion, as well as infection by SARS-CoV-2 and other HCoVs, including SARS-CoV, MERS-CoV, HCoV-229E, HCoV-NL63 and HCoV-OC43. GL25E could inhibit authentic SASR-CoV-2, HCoV-OC43 and HCoV-229E infection in vitro and prevent newborn mice from authentic HCoV-OC43 infection in vivo. GL25E could bind to glycans in the S1 subunit and HR1 in the S2 subunit in S protein, showing a mechanism of action similar to that of GRFT and EK1 alone. CONCLUSIONS: Since GL25E showed highly potent and broad-spectrum inhibitory activity against infection of SARS-CoV-2 and its mutants, as well as other HCoVs, it is a promising candidate for further development as a broad-spectrum anti-HCoV therapeutic and prophylactic to treat and prevent COVID-19 and other emerging HCoV diseases.
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COVID-19 , Yema de Huevo , Animales , Pollos , Femenino , Humanos , Inmunoglobulinas , SARS-CoV-2Asunto(s)
Coronavirus , VIH-1/fisiología , Lipopéptidos/farmacología , Proteínas Virales de Fusión/antagonistas & inhibidores , Acoplamiento Viral/efectos de los fármacos , Internalización del Virus/efectos de los fármacos , Línea Celular , Coronavirus/efectos de los fármacos , Coronavirus/fisiología , Inhibidores de Fusión de VIH , Humanos , Lipopéptidos/químicaAsunto(s)
Antivirales/farmacología , Lectinas de Plantas/farmacología , Polisacáridos/metabolismo , SARS-CoV-2/efectos de los fármacos , Glicoproteína de la Espiga del Coronavirus/metabolismo , Línea Celular , Reposicionamiento de Medicamentos , Sinergismo Farmacológico , Humanos , Concentración 50 InhibidoraRESUMEN
BACKGROUND: Macrophage M2b polarization conferred by self-DNA immunization initiates and propagates lupus nephritis. RESULTS: Knockdown of DNA-dependent activator of interferon-regulatory factors (DAI) ameliorates SLE syndrome via blunting macrophage M2b polarization. CONCLUSION: DAI functions as a DNA sensor in self-DNA-induced macrophage M2b polarization and lupus nephritis. SIGNIFICANCE: We disclose the mechanism by which self-DNA induces macrophage M2b polarization and lupus nephritis DNA-dependent activator of interferon-regulatory factors (DAI) functions as a cytoplasmic DNA sensor that activates the innate immune system. We previously found that activated lymphocyte-derived self-apoptotic DNA (ALD-DNA) immunization led to pathological macrophage activation and M2b polarization, which could initiate and propagate murine lupus nephritis. However, the specific DNA sensor(s) as well as underlying molecular mechanisms involved in ALD-DNA-induced macrophage M2b polarization in systemic lupus erythematosus (SLE) disease remains unknown. In this study, we reported that DAI expression was significantly increased in SLE patients as well as in lupus mice. Gain- and loss-of-function studies revealed that DAI was involved in ALD-DNA-induced macrophage activation and M2b polarization. Moreover, ALD-DNA notably induced dimerization/oligomerization of DAI and consequently activation of nuclear factor κB (NF-κB) and interferon regulatory factor 3 (IRF3) signaling pathways via calcium signaling, resulting in macrophage activation and M2b polarization. More importantly, blockade of DAI in vivo or selective knockdown of DAI in macrophages could ameliorate SLE syndrome via blunting macrophage M2b polarization and inhibiting inflammatory response in lupus mice. Our results suggest that DAI could function as a DNA sensor and a regulator in ALD-DNA-induced macrophage M2b polarization and lupus nephritis, providing the possible molecular mechanisms involved in ALD-DNA-induced macrophage M2b polarization in SLE disease and making DAI as a potential therapeutic target for the treatment of SLE.
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Señalización del Calcio , Proteínas de Unión al ADN/metabolismo , Lupus Eritematoso Sistémico/metabolismo , Nefritis Lúpica/metabolismo , Adolescente , Traslado Adoptivo , Adulto , Animales , Estudios de Casos y Controles , Línea Celular , Polaridad Celular , ADN/inmunología , Proteínas de Unión al ADN/genética , Femenino , Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Leucocitos Mononucleares/metabolismo , Lupus Eritematoso Sistémico/inmunología , Nefritis Lúpica/inmunología , Linfocitos/inmunología , Activación de Macrófagos , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/trasplante , Masculino , Ratones , Ratones Endogámicos BALB C , Persona de Mediana Edad , Multimerización de Proteína , ARN Interferente Pequeño/genética , Proteínas de Unión al ARN , Regulación hacia Arriba , Adulto JovenRESUMEN
BACKGROUND: Deficiency in clearance of self nuclear antigens, including DNA, is the hallmark of systemic lupus erythematosus (SLE), a chronic autoimmnue disease characterized by the production of various autoantibodies, immune complex deposition and severe organ damage. Our previous studies revealed that administration of syngeneic BALB/c mice with activated lymphocyte-derived DNA (ALD-DNA) could induce SLE disease. Mannose-binding lectin (MBL), a secreted pattern recognition receptor with binding activity to DNA, has been proved to be a modulator of inflammation, but whether MBL takes responsibility for DNA clearance, modulates the DNA-mediated immune responses, and is involved in the development of DNA-induced SLE disease remain poorly understood. METHODOLOGY/PRINCIPAL FINDINGS: The levels of serum MBL significantly decreased in lupus mice induced by ALD-DNA and were negatively correlated with SLE disease. MBL blunted macrophage M2b polarization by inhibiting the MAPK and NF-κB signaling while enhancing the activation of CREB. Furthermore, MBL suppressed the ability of ALD-DNA-stimulated macrophages to polarize T cells toward Th1 cells and Th17 cells. Importantly, MBL supplement in vivo could ameliorate lupus nephritis. CONCLUSION/SIGNIFICANCE: These results suggest MBL supplement could alleviate SLE disease and might imply a potential therapeutic strategy for DNA-induced SLE, which would further our understanding of the protective role of MBL in SLE disease.
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Nefritis Lúpica/metabolismo , Macrófagos/metabolismo , Lectina de Unión a Manosa/metabolismo , Animales , Línea Celular , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , ADN/inmunología , Modelos Animales de Enfermedad , Femenino , Lupus Eritematoso Sistémico/complicaciones , Lupus Eritematoso Sistémico/inmunología , Lupus Eritematoso Sistémico/metabolismo , Nefritis Lúpica/genética , Nefritis Lúpica/inmunología , Activación de Linfocitos/genética , Activación de Linfocitos/inmunología , Linfocitos/inmunología , Linfocitos/metabolismo , Macrófagos/inmunología , Lectina de Unión a Manosa/sangre , Lectina de Unión a Manosa/genética , Ratones , Proteínas Quinasas Activadas por Mitógenos/metabolismo , FN-kappa B/metabolismo , Transducción de SeñalRESUMEN
PURPOSE: Lupus nephritis, a major cause of morbidity in patients with systemic lupus erythematosus (SLE), is generally thought to be induced by macrophage-mediated inflammation following deposition of various autoantibodies in kidneys. We previously reported that macrophage aberrant activation induced by activated lymphocyte-derived apoptotic DNA (apopDNA) have been found to play pathogenic roles in the immunodysregulation in lupus nephritis. However, DNA sensor(s) involved in apopDNA-induced macrophage activation and lupus nephritis remains largely undefined. Herein, we aimed to reveal the DNA sensor(s) involved in SLE disease. METHODS: Correlation between the level of absent in melanoma 2 (AIM2), a cytoplasmic DNA receptor in the inflammasome pathway, and the clinical severity of SLE disease were analyzed in SLE patients as well as in lupus mice. Activated macrophages induced by apopDNA were analyzed by real-time PCR and western blot for AIM2 expression. After silencing of AIM2 via siRNA-mediated knockdown in vitro and in vivo, macrophage activation, inflammatory response, and SLE syndrome were assessed. RESULTS: AIM2 expression was closely correlated with the severity of disease in SLE patients and in lupus mice. Importantly, AIM2 expression was significantly increased in apopDNA-induced macrophages and closely correlated with macrophage activation. Knockdown of AIM2 significantly blunted apopDNA-induced macrophage activation. Furthermore, blockade of AIM2 expression notably ameliorated SLE syndrome via impeding macrophage activation and dampening inflammatory response in apopDNA-induced lupus mice. CONCLUSIONS: Our results implied that AIM2 might act as an important DNA sensor and a potential biomarker for apopDNA-induced macrophage functional maturation and SLE disease.
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Apoptosis/genética , ADN/genética , Lupus Eritematoso Sistémico/genética , Activación de Macrófagos/genética , Macrófagos/metabolismo , Proteínas Nucleares/genética , Adolescente , Adulto , Animales , Anticuerpos Antinucleares/inmunología , Apoptosis/inmunología , Células Cultivadas , ADN/inmunología , Proteínas de Unión al ADN , Femenino , Humanos , Inflamasomas/genética , Inflamasomas/inmunología , Inflamasomas/metabolismo , Lupus Eritematoso Sistémico/inmunología , Lupus Eritematoso Sistémico/metabolismo , Nefritis Lúpica/genética , Activación de Macrófagos/inmunología , Macrófagos/inmunología , Masculino , Melanoma/inmunología , Melanoma/metabolismo , Ratones , Ratones Endogámicos BALB C , Persona de Mediana Edad , Negociación , Proteínas Nucleares/inmunología , Proteínas Nucleares/metabolismo , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/inmunología , Receptores de Superficie Celular/metabolismo , Adulto JovenRESUMEN
C-reactive protein (CRP), an acute-phase protein with an ability to bind to nuclear antigen, has been reported to regulate cytokine secretion and modulate immune responses. We previously reported that activated syngeneic lymphocyte-derived apoptotic DNA (apopDNA) could induce macrophage activation and contribute to the initiation and progression of lupus nephritis. It is reasonable to hypothesize that CRP might regulate apopDNA-induced macrophage activation. Herein, CRP was shown to promote macrophage-mediated apopDNA uptake by binding to apopDNA (CRP/apopDNA complex). Notably, CRP/apopDNA treatment inhibited the production of inflammatory cytokines and chemokines by macrophages which could be induced by apopDNA alone. Further coculture and transwell studies revealed that CRP/apopDNA-induced macrophages prohibited apopDNA-induced macrophage activation in an IL-10 dependent manner. These results provide insight into the potential mechanism of CRP regulatory activity in macrophage activation induced by apopDNA in the context of lupus nephritis and other autoimmune diseases.