RESUMEN
With continued expansion of the coronavirus disease (COVID-19) pandemic, caused by severe acute respiratory syndrome 2 (SARS-CoV-2), both antiviral drugs as well as effective vaccines are desperately needed to treat patients at high risk of life-threatening disease. Here, we present in vitro evidence for significant inhibition of SARS-CoV-2 by oleandrin and a defined extract of N. oleander (designated as PBI-06150). Using Vero cells, we found that prophylactic (pre-infection) oleandrin (as either the pure compound or as the active principal ingredient in PBI-06150) administration at concentrations as low as 0.05 µg/ml exhibited potent antiviral activity against SARS-CoV-2, with an 800-fold reduction in virus production, and a 0.1 µg/ml concentration resulted in a greater than 3000-fold reduction in infectious virus production. The half maximal effective concentration (EC50) values were 11.98 ng/ml when virus output was measured at 24 h post-infection, and 7.07 ng/ml measured at 48 h post-infection. Therapeutic (post-infection) treatment up to 24 h after SARS-CoV-2 infection of Vero cells also reduced viral titers, with 0.1 µg/ml and 0.05 µg/ml concentrations causing greater than 100-fold reduction as measured at 48 h, and the 0.05 µg/ml concentration resulting in a 78-fold reduction. Concentrations of oleandrin up to 10 µg/ml were well tolerated in Vero cells. We also present in vivo evidence of the safety and efficacy of defined N. oleander extract (PBI-06150), which was administered to golden Syrian hamsters in a preparation containing as high as 130 µg/ml of oleandrin. In comparison to administration of control vehicle, PBI-06150 provided a statistically significant reduction of the viral titer in the nasal turbinates (nasal conchae). The potent prophylactic and therapeutic antiviral activities demonstrated here, together with initial evidence of its safety and efficacy in a relevant hamster model of COVID-19, support the further development of oleandrin and/or defined extracts containing this molecule for the treatment of SARS-CoV-2 and associated COVID-19 disease and potentially also for reduction of virus spread by persons diagnosed early after infection.
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Antivirales/uso terapéutico , Tratamiento Farmacológico de COVID-19 , Cardenólidos/uso terapéutico , Nerium , Extractos Vegetales/uso terapéutico , SARS-CoV-2 , Animales , Antivirales/farmacología , COVID-19/prevención & control , Cardenólidos/farmacología , Chlorocebus aethiops , Cricetinae , Femenino , Genoma Viral , Fitoterapia , Extractos Vegetales/farmacología , SARS-CoV-2/genética , Células VeroRESUMEN
The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 2019 has triggered an ongoing global pandemic of the severe pneumonia-like disease coronavirus disease 2019 (COVID-19)1. The development of a vaccine is likely to take at least 12-18 months, and the typical timeline for approval of a new antiviral therapeutic agent can exceed 10 years. Thus, repurposing of known drugs could substantially accelerate the deployment of new therapies for COVID-19. Here we profiled a library of drugs encompassing approximately 12,000 clinical-stage or Food and Drug Administration (FDA)-approved small molecules to identify candidate therapeutic drugs for COVID-19. We report the identification of 100 molecules that inhibit viral replication of SARS-CoV-2, including 21 drugs that exhibit dose-response relationships. Of these, thirteen were found to harbour effective concentrations commensurate with probable achievable therapeutic doses in patients, including the PIKfyve kinase inhibitor apilimod2-4 and the cysteine protease inhibitors MDL-28170, Z LVG CHN2, VBY-825 and ONO 5334. Notably, MDL-28170, ONO 5334 and apilimod were found to antagonize viral replication in human pneumocyte-like cells derived from induced pluripotent stem cells, and apilimod also demonstrated antiviral efficacy in a primary human lung explant model. Since most of the molecules identified in this study have already advanced into the clinic, their known pharmacological and human safety profiles will enable accelerated preclinical and clinical evaluation of these drugs for the treatment of COVID-19.
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Antivirales/análisis , Antivirales/farmacología , Betacoronavirus/efectos de los fármacos , Infecciones por Coronavirus/tratamiento farmacológico , Infecciones por Coronavirus/virología , Evaluación Preclínica de Medicamentos , Reposicionamiento de Medicamentos , Neumonía Viral/tratamiento farmacológico , Neumonía Viral/virología , Adenosina Monofosfato/análogos & derivados , Adenosina Monofosfato/farmacología , Alanina/análogos & derivados , Alanina/farmacología , Células Epiteliales Alveolares/citología , Células Epiteliales Alveolares/efectos de los fármacos , Betacoronavirus/crecimiento & desarrollo , COVID-19 , Línea Celular , Inhibidores de Cisteína Proteinasa/análisis , Inhibidores de Cisteína Proteinasa/farmacología , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Hidrazonas , Células Madre Pluripotentes Inducidas/citología , Modelos Biológicos , Morfolinas/análisis , Morfolinas/farmacología , Pandemias , Pirimidinas , Reproducibilidad de los Resultados , SARS-CoV-2 , Bibliotecas de Moléculas Pequeñas/análisis , Bibliotecas de Moléculas Pequeñas/farmacología , Triazinas/análisis , Triazinas/farmacología , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Tratamiento Farmacológico de COVID-19RESUMEN
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in the city of Wuhan, Hubei Province, China, in late 2019. Since then, the virus has spread globally and caused a pandemic. Assays that can measure the antiviral activity of antibodies or antiviral compounds are needed for SARS-CoV-2 vaccine and drug development. Here, we describe in detail a microneutralization assay, which can be used to assess in a quantitative manner if antibodies or drugs can block entry and/or replication of SARS-CoV-2 in vitro. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Microneutralization assay to test inhibition of virus by antibodies (purified antibodies or serum/plasma) Basic Protocol 2: Screening of anti-SARS-CoV-2 compounds in vitro Support Protocol: SARS-CoV-2 propagation.
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Anticuerpos Antivirales/inmunología , Betacoronavirus/inmunología , Evaluación Preclínica de Medicamentos/métodos , Pruebas de Neutralización/métodos , Animales , COVID-19 , Chlorocebus aethiops , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/virología , Humanos , Ratones , Pandemias , Neumonía Viral/inmunología , Neumonía Viral/virología , SARS-CoV-2 , Células Vero , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
Influenza A (IAV) and influenza B (IBV) viruses are highly contagious pathogens that cause fatal respiratory disease every year, with high economic impact. In addition, IAV can cause pandemic infections with great consequences when new viruses are introduced into humans. In this study, we evaluated 10 previously described compounds with antiviral activity against mammarenaviruses for their ability to inhibit IAV infection using our recently described bireporter influenza A/Puerto Rico/8/34 (PR8) H1N1 (BIRFLU). Among the 10 tested compounds, eight (antimycin A [AmA], brequinar [BRQ], 6-azauridine, azaribine, pyrazofurin [PF], AVN-944, mycophenolate mofetil [MMF], and mycophenolic acid [MPA]), but not obatoclax or Osu-03012, showed potent anti-influenza virus activity under posttreatment conditions [median 50% effective concentration (EC50) = 3.80 nM to 1.73 µM; selective index SI for 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, >28.90 to 13,157.89]. AmA, 6-azauridine, azaribine, and PF also showed potent inhibitory effect in pretreatment (EC50 = 0.14 µM to 0.55 µM; SI-MTT = 70.12 to >357.14) or cotreatment (EC50 = 34.69 nM to 7.52 µM; SI-MTT = 5.24 to > 1,441.33) settings. All of the compounds tested inhibited viral genome replication and gene transcription, and none of them affected host cellular RNA polymerase II activities. The antiviral activity of the eight identified compounds against BIRFLU was further confirmed with seasonal IAVs (A/California/04/2009 H1N1 and A/Wyoming/3/2003 H3N2) and an IBV (B/Brisbane/60/2008, Victoria lineage), demonstrating their broad-spectrum prophylactic and therapeutic activity against currently circulating influenza viruses in humans. Together, our results identified a new set of antiviral compounds for the potential treatment of influenza viral infections.IMPORTANCE Influenza viruses are highly contagious pathogens and are a major threat to human health. Vaccination remains the most effective tool to protect humans against influenza infection. However, vaccination does not always guarantee complete protection against drifted or, more noticeably, shifted influenza viruses. Although U.S. Food and Drug Administration (FDA) drugs are approved for the treatment of influenza infections, influenza viruses resistant to current FDA antivirals have been reported and continue to emerge. Therefore, there is an urgent need to find novel antivirals for the treatment of influenza viral infections in humans, a search that could be expedited by repurposing currently approved drugs. In this study, we assessed the influenza antiviral activity of 10 compounds previously shown to inhibit mammarenavirus infection. Among them, eight drugs showed antiviral activities, providing a new battery of drugs that could be used for the treatment of influenza infections.
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Antivirales/farmacología , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H3N2 del Virus de la Influenza A/efectos de los fármacos , Virus de la Influenza B/efectos de los fármacos , Células A549 , Animales , Proliferación Celular , Perros , Evaluación Preclínica de Medicamentos , Genoma Viral , Células HEK293 , Interacciones Huésped-Patógeno , Humanos , Subtipo H1N1 del Virus de la Influenza A/efectos de la radiación , Subtipo H3N2 del Virus de la Influenza A/fisiología , Virus de la Influenza B/fisiología , Células de Riñón Canino Madin Darby , Replicación Viral/efectos de los fármacosRESUMEN
The mammarenavirus Lassa (LASV) is highly prevalent in West Africa where it infects several hundred thousand individuals annually resulting in a high number of Lassa fever (LF) cases, a febrile disease associated with high morbidity and significant mortality. Mounting evidence indicates that the worldwide-distributed prototypic mammarenavirus lymphocytic choriomeningitis virus (LCMV) is a neglected human pathogen of clinical significance. There are not Food and Drug Administration (FDA) licensed vaccines and current anti-mammarenavirus therapy is limited to an off-label use of ribavirin that is only partially effective and can cause significant side effects. Therefore, there is an unmet need for novel antiviral drugs to combat LASV. This task would be facilitated by the implementation of high throughput screens (HTS) to identify inhibitors of the activity of the virus ribonucleoprotein (vRNP) responsible for directing virus RNA genome replication and gene transcription. The use of live LASV for this purpose is jeopardized by the requirement of biosafety level 4 (BSL4) containment. We have developed a virus-free cell platform, where expression levels of reporter genes serve as accurate surrogates of vRNP activity, to develop cell-based assays compatible with HTS to identify inhibitors of LASV and LCMV mammarenavirus vRNP activities.
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Antivirales/farmacología , Evaluación Preclínica de Medicamentos/métodos , Virus Lassa/efectos de los fármacos , Ribonucleoproteínas/antagonistas & inhibidores , Proteínas Virales/antagonistas & inhibidores , Animales , Chlorocebus aethiops , Relación Dosis-Respuesta a Droga , Expresión Génica , Ingeniería Genética , Células HEK293 , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Interferencia de ARN , Reproducibilidad de los Resultados , Ribonucleoproteínas/genética , Ribonucleoproteínas/metabolismo , Bibliotecas de Moléculas Pequeñas , Células VeroRESUMEN
Influenza A viruses (IAVs) cause seasonal epidemics and occasional pandemics, representing a serious public health concern. It has been described that one mechanism used by some IAV strains to escape the host innate immune responses and modulate virus pathogenicity involves the ability of the PA-X and NS1 proteins to inhibit the host protein synthesis in infected cells. It was reported that for the 2009 pandemic H1N1 IAV (pH1N1) only the PA-X protein had this inhibiting capability, while the NS1 protein did not. In this work, we have evaluated, for the first time, the combined effect of PA-X- and NS1-mediated inhibition of general gene expression on virus pathogenesis, using a temperature-sensitive, live-attenuated 2009 pandemic H1N1 IAV (pH1N1 LAIV). We found that viruses containing PA-X and NS1 proteins that simultaneously have (PAWT+/NS1MUT+) or do not have (PAMUT-/NS1WT-) the ability to block host gene expression showed reduced pathogenicity in vivo However, a virus where the ability to inhibit host protein expression was switched between PA-X and NS1 (PAMUT-/NS1MUT+) presented pathogenicity similar to that of a virus containing both wild-type proteins (PAWT+/NS1WT-). Our findings suggest that inhibition of host protein expression is subject to a strict balance, which can determine the successful progression of IAV infection. Importantly, knowledge obtained from our studies could be used for the development of new and more effective vaccine approaches against IAV.IMPORTANCE Influenza A viruses (IAVs) are one of the most common causes of respiratory infections in humans, resulting in thousands of deaths annually. Furthermore, IAVs can cause unpredictable pandemics of great consequence when viruses not previously circulating in humans are introduced into humans. The defense machinery provided by the host innate immune system limits IAV replication; however, to counteract host antiviral activities, IAVs have developed different inhibition mechanisms, including prevention of host gene expression mediated by the viral PA-X and NS1 proteins. Here, we provide evidence demonstrating that optimal control of host protein synthesis by IAV PA-X and/or NS1 proteins is required for efficient IAV replication in the host. Moreover, we demonstrate the feasibility of genetically controlling the ability of IAV PA-X and NS1 proteins to inhibit host immune responses, providing an approach to develop more effective vaccines to combat disease caused by this important respiratory pathogen.
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Interacciones Huésped-Patógeno , Subtipo H1N1 del Virus de la Influenza A/fisiología , Infecciones por Orthomyxoviridae/virología , Proteínas Represoras/fisiología , Proteínas no Estructurales Virales/fisiología , Replicación Viral , Animales , Replicación del ADN , Evaluación Preclínica de Medicamentos , Femenino , Células HEK293 , Humanos , Inmunidad Innata , Subtipo H1N1 del Virus de la Influenza A/genética , Ratones , Ratones Endogámicos C57BL , Infecciones por Orthomyxoviridae/inmunología , Biosíntesis de Proteínas , Proteínas Represoras/genética , Temperatura , Proteínas no Estructurales Virales/genéticaRESUMEN
INTRODUCTION: Several arenaviruses, chiefly Lassa virus (LASV), cause hemorrhagic fever (HF) disease in humans and pose significant public health problems in their endemic regions. Moreover, HF arenaviruses represent credible biodefense threats. There are not FDA-approved arenavirus vaccines and current anti-arenaviral therapy is limited to an off-label use of ribavirin that is only partially effective. AREAS COVERED: Live-attenuated vaccines (LAV) represent the most feasible approach to control HF arenaviruses within their endemic regions. Different platforms, including recombinant viral vectors expressing LASV antigens, and the use of attenuated reassortant arenaviruses, have been used to develop LAV candidates against LASV with promising results in animal models of LASV infection, but none of them has entered a clinical trial. These vaccine efforts have been the subject of recent reviews and will not be examined in this review, which is focused on new avenues for the development of safe and effective LAV to combat HF arenaviruses. Expert commentary: The development of arenavirus reverse genetics has provided investigators with a novel powerful approach to manipulate the genomes of HF arenaviruses, which has opened new avenues for the rapid development of safe and effective LAV to combat these human pathogens.
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Infecciones por Arenaviridae/prevención & control , Arenavirus/inmunología , Fiebres Hemorrágicas Virales/prevención & control , Vacunas Virales/inmunología , Vacunas Virales/aislamiento & purificación , Animales , Infecciones por Arenaviridae/virología , Descubrimiento de Drogas/tendencias , Evaluación Preclínica de Medicamentos , Fiebres Hemorrágicas Virales/virología , Humanos , Vacunas Atenuadas/inmunología , Vacunas Atenuadas/aislamiento & purificaciónRESUMEN
Influenza viruses remain a critical global health concern. More efficacious vaccines are needed to protect against influenza virus, yet few adjuvants are approved for routine use. Specialized proresolving mediators (SPMs) are powerful endogenous bioactive regulators of inflammation, with great clinical translational properties. In this study, we investigated the ability of the SPM 17-HDHA to enhance the adaptive immune response using an OVA immunization model and a preclinical influenza vaccination mouse model. Our findings revealed that mice immunized with OVA plus 17-HDHA or with H1N1-derived HA protein plus 17-HDHA increased Ag-specific Ab titers. 17-HDHA increased the number of Ab-secreting cells in vitro and the number of HA-specific Ab-secreting cells present in the bone marrow. Importantly, the 17-HDHA-mediated increased Ab production was more protective against live pH1N1 influenza infection in mice. To our knowledge, this is the first report on the biological effects of ω-3-derived SPMs on the humoral immune response. These findings illustrate a previously unknown biological link between proresolution signals and the adaptive immune system. Furthermore, this work has important implications for the understanding of B cell biology, as well as the development of new potential vaccine adjuvants.
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Ácidos Docosahexaenoicos/farmacología , Inmunidad Humoral/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/inmunología , Infecciones por Orthomyxoviridae/inmunología , Adyuvantes Inmunológicos/farmacología , Animales , Anticuerpos Antivirales/inmunología , Formación de Anticuerpos/efectos de los fármacos , Formación de Anticuerpos/inmunología , Subgrupos de Linfocitos B/citología , Subgrupos de Linfocitos B/efectos de los fármacos , Subgrupos de Linfocitos B/inmunología , Subgrupos de Linfocitos B/metabolismo , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/inmunología , Glicoproteínas Hemaglutininas del Virus de la Influenza/inmunología , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunología , Masculino , Ratones , Infecciones por Orthomyxoviridae/virología , Células Plasmáticas/citología , Células Plasmáticas/efectos de los fármacos , Células Plasmáticas/inmunología , Células Plasmáticas/metabolismoRESUMEN
Arenaviruses merit significant interest as important human pathogens, since several of them cause severe hemorrhagic fever disease that is associated with high morbidity and significant mortality. Currently, there are no FDA-licensed arenavirus vaccines available, and current antiarenaviral therapy is limited to an off-labeled use of the nucleoside analog ribavirin, which has limited prophylactic efficacy. The pyrimidine biosynthesis inhibitor A3, which was identified in a high-throughput screen for compounds that blocked influenza virus replication, exhibits a broad-spectrum antiviral activity against negative- and positive-sense RNA viruses, retroviruses, and DNA viruses. In this study, we evaluated the antiviral activity of A3 against representative Old World (lymphocytic choriomeningitis virus) and New World (Junin virus) arenaviruses in rodent, monkey, and human cell lines. We show that A3 is significantly more efficient than ribavirin in controlling arenavirus multiplication and that the A3 inhibitory effect is in part due to its ability to interfere with viral RNA replication and transcription. We document an additive antiarenavirus effect of A3 and ribavirin, supporting the potential combination therapy of ribavirin and pyrimidine biosynthesis inhibitors for the treatment of arenavirus infections.
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Antivirales/farmacología , Infecciones por Arenaviridae/virología , Arenavirus/efectos de los fármacos , Pirimidinas/antagonistas & inhibidores , Animales , Infecciones por Arenaviridae/metabolismo , Arenavirus/genética , Arenavirus/fisiología , Línea Celular , Evaluación Preclínica de Medicamentos , Humanos , Pirimidinas/biosíntesis , Replicación Viral/efectos de los fármacosRESUMEN
Seasonal and pandemic influenza viruses continue to be a leading global health concern. Emerging resistance to the current drugs and the variable efficacy of vaccines underscore the need for developing new flu drugs that will be broadly effective against wild-type and drug-resistant influenza strains. Here, we report the discovery and development of a class of inhibitors targeting the cap-snatching endonuclease activity of the viral polymerase. A high-resolution crystal form of pandemic 2009 H1N1 influenza polymerase acidic protein N-terminal endonuclease domain (PAN) was engineered and used for fragment screening leading to the identification of new chemical scaffolds binding to the PAN active site cleft. During the course of screening, binding of a third metal ion that is potentially relevant to endonuclease activity was detected in the active site cleft of PAN in the presence of a fragment. Using structure-based optimization, we developed a highly potent hydroxypyridinone series of compounds from a fragment hit that defines a new mode of chelation to the active site metal ions. A compound from the series demonstrating promising enzymatic inhibition in a fluorescence-based enzyme assay with an IC50 value of 11 nM was found to have an antiviral activity (EC50) of 11 µM against PR8 H1N1 influenza A in MDCK cells.
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Endonucleasas/antagonistas & inhibidores , Inhibidores Enzimáticos/química , Orthomyxoviridae/enzimología , Antivirales/química , Antivirales/farmacología , Bioensayo , Dominio Catalítico , Células Cultivadas , Quelantes/química , Cristalografía por Rayos X , Evaluación Preclínica de Medicamentos , Inhibidores Enzimáticos/farmacología , Humanos , Concentración 50 InhibidoraRESUMEN
Epidemiological studies indicate that maternal influenza viral infection increases the risk for schizophrenia in the adult offspring. The serotonin and glutamate systems are suspected in the etiology of schizophrenia, as well as in the mechanism of action of antipsychotic drugs. The effects of hallucinogens, such as psilocybin and mescaline, require the serotonin 5-HT(2A) receptor, and induce schizophrenia-like psychosis in humans. In addition, metabotropic glutamate receptor mGlu(2/3) agonists show promise as a new treatment for schizophrenia. Here, we investigated the level of expression and behavioral function of 5-HT(2A) and mGlu(2) receptors in a mouse model of maternal influenza viral infection. We show that spontaneous locomotor activity is diminished by maternal infection with the mouse-adapted influenza A/WSN/33 (H1N1) virus. The behavioral responses to hallucinogens and glutamate antipsychotics are both affected by maternal exposure to influenza virus, with increased head-twitch response to hallucinogens and diminished antipsychotic-like effect of the glutamate agonist. In frontal cortex of mice born to influenza virus-infected mothers, the 5-HT(2A) receptor is upregulated and the mGlu(2) receptor is downregulated, an alteration that may be involved in the behavioral changes observed. Additionally, we find that the cortical 5-HT(2A) receptor-dependent signaling pathways are significantly altered in the offspring of infected mothers, showing higher c-fos, egr-1, and egr-2 expression in response to the hallucinogenic drug DOI. Identifying a biochemical alteration that parallels the behavioral changes observed in a mouse model of prenatal viral infection may facilitate targeting therapies for treatment and prevention of schizophrenia.
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Lóbulo Frontal/metabolismo , Infecciones por Orthomyxoviridae/metabolismo , Receptor de Serotonina 5-HT2A/metabolismo , Receptores de Glutamato Metabotrópico/metabolismo , Esquizofrenia , Hijos Adultos , Animales , Antipsicóticos/farmacología , Corteza Cerebral/metabolismo , Modelos Animales de Enfermedad , Regulación hacia Abajo/efectos de los fármacos , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Proteína 2 de la Respuesta de Crecimiento Precoz/metabolismo , Femenino , Ácido Glutámico/análogos & derivados , Alucinógenos/farmacología , Subtipo H1N1 del Virus de la Influenza A , Intercambio Materno-Fetal , Ratones , Vías Nerviosas/efectos de los fármacos , Infecciones por Orthomyxoviridae/fisiopatología , Infecciones por Orthomyxoviridae/virología , Embarazo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Receptor de Serotonina 5-HT2A/efectos de los fármacos , Esquizofrenia/metabolismo , Regulación hacia Arriba/efectos de los fármacosRESUMEN
A hallmark of immune activation by certain RNA sequences is the generation of interferon responses. However, the study of immunostimulatory RNA (isRNA) has been hindered by costly and slow methods, particularly in vitro. We have developed a cell-based assay to detect human type I interferon (IFN) that reliably senses both IFN-alpha and IFN-beta simultaneously. The human 293T cell line was stably transfected with a fusion gene of monomeric red fluorescent protein (mRFP) under the transcriptional control of an interferon-stimulated response element (ISRE). High levels of mRFP are expressed following activation of the type I IFN receptor (IFNAR). Using this method, detection limits for IFN similar to that of ELISA can be achieved but with a greater dynamic range and in a high-throughput manner. As a proof of concept, we utilized this method to screen a library of cationic lipid-like materials that form nanoparticle complexes with RNA for induction of innate immune responses in vitro. We expect the screening and detection methods described herein may provide a useful tool in elucidating mechanisms that govern the delivery of RNA molecules to effector cells and receptors of the innate immune system. We apply this tool to investigate isRNA drug delivery, but it may also find use in other applications for which high-throughput detection of type 1 IFN is desired.