RESUMEN
Dengue is a global public health threat, with about half of the world's population at risk of contracting this mosquito-borne viral disease. Climate change, urbanization, and global travel accelerate the spread of dengue virus (DENV) to new areas, including southern parts of Europe and the US. Currently, no dengue-specific small-molecule antiviral for prophylaxis or treatment is available. Here, we report the discovery of JNJ-1802 as a potent, pan-serotype DENV inhibitor (EC50's ranging from 0.057 to 11 nM against the four DENV serotypes). The observed oral bioavailability of JNJ-1802 across preclinical species, its low clearance in human hepatocytes, the absence of major in vitro pharmacology safety alerts, and a dose-proportional increase in efficacy against DENV-2 infection in mice were all supportive of its selection as a development candidate against dengue. JNJ-1802 is being progressed in clinical studies for the prevention or treatment of dengue.
Asunto(s)
Virus del Dengue , Dengue , Hidrocarburos Halogenados , Indoles , Ratones , Humanos , Animales , Serogrupo , Dengue/tratamiento farmacológicoRESUMEN
In the absence of any approved dengue-specific treatment, the discovery and development of a novel small-molecule antiviral for the prevention or treatment of dengue are critical. We previously reported the identification of a novel series of 3-acyl-indole derivatives as potent and pan-serotype dengue virus inhibitors. We herein describe our optimization efforts toward preclinical candidates 24a and 28a with improved pan-serotype coverage (EC50's against the four DENV serotypes ranging from 0.0011 to 0.24 µM for 24a and from 0.00060 to 0.084 µM for 28a), chiral stability, and oral bioavailability in preclinical species, as well as showing a dose-proportional increase in efficacy against DENV-2 infection in vivo in mice.
Asunto(s)
Virus del Dengue , Dengue , Ratones , Animales , Serogrupo , Antivirales/farmacología , Antivirales/uso terapéutico , Dengue/tratamiento farmacológico , Indoles/farmacología , Indoles/uso terapéuticoRESUMEN
Dengue is a major health threat and the number of symptomatic infections caused by the four dengue serotypes is estimated to be 96 million1 with annually around 10,000 deaths2. However, no antiviral drugs are available for the treatment or prophylaxis of dengue. We recently described the interaction between non-structural proteins NS3 and NS4B as a promising target for the development of pan-serotype dengue virus (DENV) inhibitors3. Here we present JNJ-1802-a highly potent DENV inhibitor that blocks the NS3-NS4B interaction within the viral replication complex. JNJ-1802 exerts picomolar to low nanomolar in vitro antiviral activity, a high barrier to resistance and potent in vivo efficacy in mice against infection with any of the four DENV serotypes. Finally, we demonstrate that the small-molecule inhibitor JNJ-1802 is highly effective against viral infection with DENV-1 or DENV-2 in non-human primates. JNJ-1802 has successfully completed a phase I first-in-human clinical study in healthy volunteers and was found to be safe and well tolerated4. These findings support the further clinical development of JNJ-1802, a first-in-class antiviral agent against dengue, which is now progressing in clinical studies for the prevention and treatment of dengue.
Asunto(s)
Antivirales , Virus del Dengue , Dengue , Primates , Proteínas no Estructurales Virales , Animales , Humanos , Ratones , Antivirales/efectos adversos , Antivirales/farmacología , Antivirales/uso terapéutico , Ensayos Clínicos Fase I como Asunto , Dengue/tratamiento farmacológico , Dengue/prevención & control , Dengue/virología , Virus del Dengue/clasificación , Virus del Dengue/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Farmacorresistencia Viral , Técnicas In Vitro , Terapia Molecular Dirigida , Primates/virología , Unión Proteica/efectos de los fármacos , Proteínas no Estructurales Virales/antagonistas & inhibidores , Proteínas no Estructurales Virales/metabolismo , Replicación ViralRESUMEN
The SARS-CoV-2 main protease (3CLpro) has an indispensable role in the viral life cycle and is a therapeutic target for the treatment of COVID-19. The potential of 3CLpro-inhibitors to select for drug-resistant variants needs to be established. Therefore, SARS-CoV-2 was passaged in vitro in the presence of increasing concentrations of ALG-097161, a probe compound designed in the context of a 3CLpro drug discovery program. We identified a combination of amino acid substitutions in 3CLpro (L50F E166A L167F) that is associated with a >20× increase in 50% effective concentration (EC50) values for ALG-097161, nirmatrelvir (PF-07321332), PF-00835231, and ensitrelvir. While two of the single substitutions (E166A and L167F) provide low-level resistance to the inhibitors in a biochemical assay, the triple mutant results in the highest levels of resistance (6× to 72×). All substitutions are associated with a significant loss of enzymatic 3CLpro activity, suggesting a reduction in viral fitness. Structural biology analysis indicates that the different substitutions reduce the number of inhibitor/enzyme interactions while the binding of the substrate is maintained. These observations will be important for the interpretation of resistance development to 3CLpro inhibitors in the clinical setting. IMPORTANCE Paxlovid is the first oral antiviral approved for treatment of SARS-CoV-2 infection. Antiviral treatments are often associated with the development of drug-resistant viruses. In order to guide the use of novel antivirals, it is essential to understand the risk of resistance development and to characterize the associated changes in the viral genes and proteins. In this work, we describe for the first time a pathway that allows SARS-CoV-2 to develop resistance against Paxlovid in vitro. The characteristics of in vitro antiviral resistance development may be predictive for the clinical situation. Therefore, our work will be important for the management of COVID-19 with Paxlovid and next-generation SARS-CoV-2 3CLpro inhibitors.
Asunto(s)
COVID-19 , SARS-CoV-2 , Humanos , Antivirales/farmacología , Antivirales/química , Inhibidores Enzimáticos , Inhibidores de Proteasas/farmacología , Inhibidores de Proteasas/química , SARS-CoV-2/genéticaRESUMEN
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, has led to a pandemic, that continues to be a huge public health burden. Despite the availability of vaccines, there is still a need for small-molecule antiviral drugs. In an effort to identify novel and drug-like hit matter that can be used for subsequent hit-to-lead optimization campaigns, we conducted a high-throughput screening of a 160 K compound library against SARS-CoV-2, yielding a 1-heteroaryl-2-alkoxyphenyl analog as a promising hit. Antiviral profiling revealed this compound was active against various beta-coronaviruses and preliminary mode-of-action experiments demonstrated that it interfered with viral entry. A systematic structure-activity relationship (SAR) study demonstrated that a 3- or 4-pyridyl moiety on the oxadiazole moiety is optimal, whereas the oxadiazole can be replaced by various other heteroaromatic cycles. In addition, the alkoxy group tolerates some structural diversity.
Asunto(s)
Antivirales/química , Antivirales/farmacología , Compuestos Heterocíclicos/farmacología , SARS-CoV-2/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Animales , Chlorocebus aethiops , Ensayos Analíticos de Alto Rendimiento , Pruebas de Sensibilidad Microbiana , Relación Estructura-Actividad , Células VeroRESUMEN
Dengue virus causes approximately 96 million symptomatic infections annually, manifesting as dengue fever or occasionally as severe dengue1,2. There are no antiviral agents available to prevent or treat dengue. Here, we describe a highly potent dengue virus inhibitor (JNJ-A07) that exerts nanomolar to picomolar activity against a panel of 21 clinical isolates that represent the natural genetic diversity of known genotypes and serotypes. The molecule has a high barrier to resistance and prevents the formation of the viral replication complex by blocking the interaction between two viral proteins (NS3 and NS4B), thus revealing a previously undescribed mechanism of antiviral action. JNJ-A07 has a favourable pharmacokinetic profile that results in outstanding efficacy against dengue virus infection in mouse infection models. Delaying start of treatment until peak viraemia results in a rapid and significant reduction in viral load. An analogue is currently in further development.
Asunto(s)
Antivirales/farmacología , Virus del Dengue/clasificación , Virus del Dengue/efectos de los fármacos , Dengue/virología , Proteínas de la Membrana/metabolismo , Proteínas no Estructurales Virales/metabolismo , Animales , Antivirales/farmacocinética , Antivirales/uso terapéutico , Dengue/tratamiento farmacológico , Virus del Dengue/genética , Virus del Dengue/metabolismo , Modelos Animales de Enfermedad , Femenino , Masculino , Proteínas de la Membrana/antagonistas & inhibidores , Ratones , ARN Helicasas/antagonistas & inhibidores , ARN Helicasas/metabolismo , Serina Endopeptidasas/metabolismo , Carga Viral/efectos de los fármacos , Proteínas no Estructurales Virales/antagonistas & inhibidores , Viremia/tratamiento farmacológico , Viremia/virología , Replicación Viral/efectos de los fármacosRESUMEN
There is an urgent need for antivirals targeting the SARS-CoV-2 virus to fight the current COVID-19 pandemic. The SARS-CoV-2 main protease (3CLpro) represents a promising target for antiviral therapy. The lack of selectivity for some of the reported 3CLpro inhibitors, specifically versus cathepsin L, raises potential safety and efficacy concerns. ALG-097111 potently inhibited SARS-CoV-2 3CLpro (IC50 = 7 nM) without affecting the activity of human cathepsin L (IC50 > 10 µM). When ALG-097111 was dosed in hamsters challenged with SARS-CoV-2, a robust and significant 3.5 log10 (RNA copies/mg) reduction of the viral RNA copies and 3.7 log10 (TCID50/mg) reduction in the infectious virus titers in the lungs was observed. These results provide the first in vivo validation for the SARS-CoV-2 3CLpro as a promising therapeutic target for selective small molecule inhibitors.
Asunto(s)
Amidas/farmacología , Tratamiento Farmacológico de COVID-19 , Proteasas 3C de Coronavirus/antagonistas & inhibidores , Inhibidores de Cisteína Proteinasa/farmacología , Modelos Animales de Enfermedad , SARS-CoV-2/efectos de los fármacos , SARS-CoV-2/enzimología , Amidas/farmacocinética , Animales , COVID-19/virología , Catepsina L/antagonistas & inhibidores , Línea Celular , Cricetinae , Inhibidores de Cisteína Proteinasa/farmacocinética , Femenino , Humanos , Concentración 50 Inhibidora , Masculino , Mesocricetus/virología , Reproducibilidad de los Resultados , SARS-CoV-2/crecimiento & desarrollo , Serina Endopeptidasas , Especificidad por Sustrato , Replicación Viral/efectos de los fármacosRESUMEN
The 3-chymotrypsin-like cysteine protease (3CLpro) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is considered a major target for the discovery of direct antiviral agents. We previously reported the evaluation of SARS-CoV-2 3CLpro inhibitors in a novel self-assembled monolayer desorption ionization mass spectrometry (SAMDI-MS) enzymatic assay (Gurard-Levin et al., 2020). The assay was further improved by adding the rhinovirus HRV3C protease to the same well as the SARS-CoV-2 3CLpro enzyme. High substrate specificity for each enzyme allowed the proteases to be combined in a single assay reaction without interfering with their individual activities. This novel duplex assay was used to profile a diverse set of reference protease inhibitors. The protease inhibitors were grouped into three categories based on their relative potency against 3CLpro and HRV3C including those that are: equipotent against 3CLpro and HRV3C (GC376 and calpain inhibitor II), selective for 3CLpro (PF-00835231, calpain inhibitor XII, boceprevir), and selective for HRV3C (rupintrivir). Structural analysis showed that the combination of minimal interactions, conformational flexibility, and limited bulk allows GC376 and calpain inhibitor II to potently inhibit both enzymes. In contrast, bulkier compounds interacting more tightly with pockets P2, P3, and P4 due to optimization for a specific target display a more selective inhibition profile. Consistently, the most selective viral protease inhibitors were relatively weak inhibitors of human cathepsin L. Taken together, these results can guide the design of cysteine protease inhibitors that are either virus-specific or retain a broad antiviral spectrum against coronaviruses and rhinoviruses.
Asunto(s)
Antivirales/farmacología , Proteasas 3C de Coronavirus/antagonistas & inhibidores , Inhibidores de Proteasas/farmacología , Rhinovirus/efectos de los fármacos , SARS-CoV-2/efectos de los fármacos , Antivirales/química , Sitios de Unión , Catepsina L/metabolismo , Descubrimiento de Drogas , Glicoproteínas/farmacología , Humanos , Cinética , Modelos Moleculares , Inhibidores de Proteasas/química , Pirrolidinas/farmacología , Ácidos SulfónicosRESUMEN
3-Acyl-indole derivative 1 was identified as a novel dengue virus (DENV) inhibitor from a DENV serotype 2 (DENV-2) phenotypic antiviral screen. Extensive SAR studies led to the discovery of new derivatives with improved DENV-2 potency as well as activity in nanomolar to micromolar range against the other DENV serotypes. In addition to the potency, physicochemical properties and metabolic stability in rat and human microsomes were improved during the optimization process. Chiral separation of the racemic mixtures showed a clear preference for one of the two enantiomers. Furthermore, rat pharmacokinetics of two compounds will be discussed in more detail, demonstrating the potential of this new series of pan-serotype-DENV inhibitors.
Asunto(s)
Antivirales/química , Antivirales/farmacología , Virus del Dengue/efectos de los fármacos , Dengue/tratamiento farmacológico , Descubrimiento de Drogas , Indoles/química , Microsomas Hepáticos/metabolismo , Animales , Chlorocebus aethiops , Dengue/virología , Virus del Dengue/clasificación , Diseño de Fármacos , Humanos , Microsomas Hepáticos/efectos de los fármacos , Modelos Moleculares , Estructura Molecular , Conformación Proteica , Ratas , Relación Estructura-Actividad , Células VeroRESUMEN
Objectives: We report the use of reconstituted 3D human airway epithelium cells (HuAECs) of bronchial origin in an air-liquid interface to study respiratory syncytial virus (RSV) infection and to assess the efficacy of RSV inhibitors in (pre-)clinical development. Methods: HuAECs were infected with RSV-A Long strain (0.01 CCID50/cell, where CCID50 represents 50% cell culture infectious dose in HEp2 cells) on the apical compartment of the culture. At the time of infection or at 1 or 3 days post-infection, selected inhibitors were added and refreshed daily on the basal compartment of the culture. Viral shedding was followed up by apical washes collected daily and quantifying viral RNA by RT-qPCR. Results: RSV-A replicates efficiently in HuAECs and viral RNA is shed for weeks after infection. RSV infection reduces the ciliary beat frequency of the ciliated cells as of 4 days post-infection, with complete ciliary dyskinesia observed by day 10. Treatment with RSV fusion inhibitors resulted in an antiviral effect only when added at the time of infection. In contrast, the use of replication inhibitors (both nucleoside and non-nucleoside) elicited a marked antiviral effect even when the start of treatment was delayed until 1 day or even 3 days after infection. Levels of the inflammation marker RANTES (mRNA) increased â¼200-fold in infected, untreated cultures (at 3 weeks post-infection), but levels were comparable to those of uninfected cultures in the presence of PC786, an RSV replication inhibitor, suggesting that an efficient antiviral treatment might inhibit virus-induced inflammation in this model. Conclusions: Overall, HuAECs offer a robust and physiologically relevant model to study RSV replication and to assess the efficacy of antiviral compounds.
Asunto(s)
Antivirales/farmacología , Mucosa Respiratoria/virología , Virus Sincitial Respiratorio Humano/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Benzamidas , Benzazepinas , Técnicas de Cultivo de Célula , Evaluación Preclínica de Medicamentos , Células Epiteliales/virología , Humanos , Técnicas de Cultivo de Órganos , ARN Viral/genética , Infecciones por Virus Sincitial Respiratorio/tratamiento farmacológico , Virus Sincitial Respiratorio Humano/genética , Compuestos de Espiro/farmacologíaRESUMEN
From a fungicidal screen, we identified 2-(2-oxo-morpholin-3-yl)-acetamide derivatives as fungicidal agents against Candida species, additionally characterized by antifungal activity against Aspergillus species. However, development of this series was hampered by low plasmatic stability. Introduction of a gem-dimethyl on the 6-position of the morpholin-2-one core led to considerable improvement in plasmatic stability while maintaining in vitro antifungal activity. Further optimization of the series resulted in the discovery of N-(biphenyl-3-ylmethyl)-2-(4-ethyl-6,6-dimethyl-2-oxomorpholin-3-yl)acetamide (87), which, in addition to fungicidal activity against Candida species, shows promising and broad antifungal in vitro activity against various fungi species, such as molds and dermatophytes. In vivo efficacy was also demonstrated in a murine model of systemic Candida albicans infection with a significant fungal load reduction in kidneys.
Asunto(s)
Acetamidas/farmacología , Antifúngicos/farmacología , Morfolinas/farmacología , Acetamidas/síntesis química , Acetamidas/química , Animales , Antifúngicos/síntesis química , Antifúngicos/química , Candida/efectos de los fármacos , Candidiasis/tratamiento farmacológico , Candidiasis/microbiología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Masculino , Ratones , Ratones Endogámicos BALB C , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Morfolinas/síntesis química , Morfolinas/química , Especificidad de la Especie , Relación Estructura-Actividad , Especificidad por SustratoRESUMEN
We have identified two subseries of 2,6-disubstituted quinolines, consisting of 6-amide and 6-urea derivatives, which are characterized by fungicidal activity against Candida albicans with minimal fungicidal concentration (MFC) values < 15 µM. The 6-amide derivatives displayed the highest fungicidal activity against C. albicans, in particular compounds 1, 5 and 6 characterized by MFC values of 6.25-12.5 µM. Compounds 1 and 5 of this series displayed fungicidal activity against the emerging pathogen Candida glabrata (MFC < 50 µM). The 6-amide derivatives 1, 2, 5, and 6 and the 6-urea derivatives 10, 12, 13 and 15 could also eradicate C. albicans biofilms. We found that the 6-urea derivatives 10, 13, and 15 induced accumulation of endogenous reactive oxygen species in Candida albicans biofilms.
Asunto(s)
Antifúngicos/química , Antifúngicos/farmacología , Biopelículas/efectos de los fármacos , Candida/efectos de los fármacos , Quinolinas/química , Quinolinas/farmacología , Candida/metabolismo , Pruebas de Sensibilidad Microbiana , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Lens epithelium-derived growth factor (LEDGF/p75) is a cellular cofactor of HIV-1 integrase that promotes viral integration by tethering the preintegration complex to the chromatin. By virtue of its crucial role in the early steps of HIV replication, the interaction between LEDGF/p75 and integrase represents an attractive target for antiviral therapy. We have rationally designed a series of 2-(quinolin-3-yl)acetic acid derivatives (LEDGINs) that act as potent inhibitors of the LEDGF/p75-integrase interaction and HIV-1 replication at submicromolar concentration by blocking the integration step. A 1.84-A resolution crystal structure corroborates the binding of the inhibitor in the LEDGF/p75-binding pocket of integrase. Together with the lack of cross-resistance with two clinical integrase inhibitors, these findings define the 2-(quinolin-3-yl)acetic acid derivatives as the first genuine allosteric HIV-1 integrase inhibitors. Our work demonstrates the feasibility of rational design of small molecules inhibiting the protein-protein interaction between a viral protein and a cellular host factor.
Asunto(s)
Antivirales/farmacología , Inhibidores Enzimáticos/farmacología , Integrasa de VIH/metabolismo , VIH/fisiología , Integración Viral/fisiología , Replicación Viral/fisiología , Antivirales/síntesis química , Antivirales/química , Técnicas de Cultivo de Célula , Farmacorresistencia Viral , Inhibidores Enzimáticos/síntesis química , VIH/efectos de los fármacos , VIH/enzimología , VIH/patogenicidad , Integrasa de VIH/química , VIH-1/efectos de los fármacos , VIH-1/enzimología , VIH-1/fisiología , Inmunidad Innata , Modelos Moleculares , Fragmentos de Péptidos/metabolismo , Relación Estructura-Actividad Cuantitativa , Tetrahidroisoquinolinas/farmacología , Interfaz Usuario-Computador , Integración Viral/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
RNA viruses are the agents of numerous widespread and often severe diseases. Their unique RNA-dependent RNA polymerase (RDRP) is essential for replication and, thus, constitutes a valid target for the development of selective chemotherapeutic agents. In this regard, we have investigated sugar-modified ribonucleoside analogues as potential inhibitors of the RDRP. Title compounds retain 'natural' pyrimidine bases, but possess a beta-methyl substituent at the 2'-position of the D- or L-ribose moiety. Evaluation against a broad range of RNA viruses, either single-stranded positive (ssRNA+), single-stranded negative (ssRNA-) or double-stranded (dsRNA), revealed potent activities for D-2'-C-methyl-cytidine and -uridine against ssRNA+, and dsRNA viruses. None of the L-enantiomers were active. Moreover, the 5'-triphosphates of the active D-enantiomers were found to inhibit the bovine virus diarrhoea virus polymerase. Thus, the 2'-methyl branching of natural pyrimidine ribonucleosides transforms physiological molecules into potent, broad-spectrum antiviral agents that merit further development.