The Experts below are selected from a list of 3867 Experts worldwide ranked by ideXlab platform
Craig E. Cameron - One of the best experts on this subject based on the ideXlab platform.
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More than efficacy revealed by single-cell analysis of Antiviral Therapeutics
Science Advances, 2019Co-Authors: Wu Liu, Mehmet Umut Caglar, Zhangming Mao, Andrew Woodman, Jamie J. Arnold, Claus O. Wilke, Craig E. CameronAbstract:Because many aspects of viral infection dynamics and inhibition are governed by stochastic processes, single-cell analysis should provide more information than approaches using population averaging. We have developed a microfluidic device composed of ~6000 wells, with each well containing a microstructure to capture single, infected cells replicating an enterovirus expressing a fluorescent reporter protein. We have used this system to characterize enterovirus inhibitors with distinct mechanisms of action. Single-cell analysis reveals that each class of inhibitor interferes with the viral infection cycle in a manner that can be distinguished by principal component analysis. Single-cell analysis of Antiviral candidates not only reveals efficacy but also facilitates clustering of drugs with the same mechanism of action and provides some indication of the ease with which resistance will develop.
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More than efficacy revealed by single-cell analysis of Antiviral Therapeutics
2019Co-Authors: Wu Liu, Mehmet Umut Caglar, Zhangming Mao, Andrew Woodman, Jamie J. Arnold, Claus O. Wilke, Craig E. CameronAbstract:SUMMARY Development of Antiviral Therapeutics emphasizes minimization of the effective dose and maximization of the toxic dose, first in cell culture and later in animal models. Long-term success of an Antiviral therapeutic is determined not only by its efficacy but also by the duration of time required for drug-resistance to evolve. We have developed a microfluidic device comprised of ~6000 wells, with each well containing a microstructure to capture single cells. We have used this device to characterize enterovirus inhibitors with distinct mechanisms of action. In contrast to population methods, single-cell analysis reveals that each class of inhibitor interferes with the viral infection cycle in a manner that can be distinguished by principal component analysis. Single-cell analysis of Antiviral candidates reveals not only efficacy but also properties of the members of the viral population most sensitive to the drug, the stage of the lifecycle most affected by the drug, and perhaps even if the drug targets an interaction of the virus with its host.
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Hepatitis C Virus Non-structural Protein 3 (HCV NS3): A Multifunctional Antiviral Target
The Journal of biological chemistry, 2010Co-Authors: Kevin D. Raney, Suresh D. Sharma, Ibrahim M. Moustafa, Craig E. CameronAbstract:Hepatitis C virus non-structural protein 3 contains a serine protease and an RNA helicase. Protease cleaves the genome-encoded polyprotein and inactivates cellular proteins required for innate immunity. Protease has emerged as an important target for the development of Antiviral Therapeutics, but drug resistance has turned out to be an obstacle in the clinic. Helicase is required for both genome replication and virus assembly. Mechanistic and structural studies of helicase have hurled this enzyme into a prominent position in the field of helicase enzymology. Nevertheless, studies of helicase as an Antiviral target remain in their infancy.
Boon Chin Heng - One of the best experts on this subject based on the ideXlab platform.
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Pluripotent Human embryonic stem cell derived neural lineages for in vitro modelling of enterovirus 71 infection and therapy
Virology Journal, 2016Co-Authors: May Shin Yap, Yin Quan Tang, Yin Yeo, Wei Ling Lim, Lee Wei Lim, Kuan Onn Tan, Mark Richards, Iekhsan Othman, Chit Laa Poh, Boon Chin HengAbstract:Background The incidence of neurological complications and fatalities associated with Hand, Foot & Mouth disease has increased over recent years, due to emergence of newly-evolved strains of Enterovirus 71 (EV71). In the search for new Antiviral Therapeutics against EV71, accurate and sensitive in vitro cellular models for preliminary studies of EV71 pathogenesis is an essential prerequisite, before progressing to expensive and time-consuming live animal studies and clinical trials. Methods This study thus investigated whether neural lineages derived from pluripotent human embryonic stem cells (hESC) can fulfil this purpose. EV71 infection of hESC-derived neural stem cells (NSC) and mature neurons (MN) was carried out in vitro, in comparison with RD and SH-SY5Y cell lines. Results Upon assessment of post-infection survivability and EV71 production by the various types, it was observed that NSC were significantly more susceptible to EV71 infection compared to MN, RD (rhabdomyosarcoma) and SH-SY5Y cells, which was consistent with previous studies on mice. The SP81 peptide had significantly greater inhibitory effect on EV71 production by NSC and MN compared to the cancer-derived RD and SH-SY5Y cell lines. Conclusions Hence, this study demonstrates that hESC-derived neural lineages can be utilized as in vitro models for studying EV71 pathogenesis and for screening of Antiviral Therapeutics.
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Pluripotent Human embryonic stem cell derived neural lineages for in vitro modelling of enterovirus 71 infection and therapy.
Virology journal, 2016Co-Authors: May Shin Yap, Yin Quan Tang, Yin Yeo, Wei Ling Lim, Lee Wei Lim, Kuan Onn Tan, Mark Richards, Iekhsan Othman, Chit Laa Poh, Boon Chin HengAbstract:The incidence of neurological complications and fatalities associated with Hand, Foot & Mouth disease has increased over recent years, due to emergence of newly-evolved strains of Enterovirus 71 (EV71). In the search for new Antiviral Therapeutics against EV71, accurate and sensitive in vitro cellular models for preliminary studies of EV71 pathogenesis is an essential prerequisite, before progressing to expensive and time-consuming live animal studies and clinical trials. This study thus investigated whether neural lineages derived from pluripotent human embryonic stem cells (hESC) can fulfil this purpose. EV71 infection of hESC-derived neural stem cells (NSC) and mature neurons (MN) was carried out in vitro, in comparison with RD and SH-SY5Y cell lines. Upon assessment of post-infection survivability and EV71 production by the various types, it was observed that NSC were significantly more susceptible to EV71 infection compared to MN, RD (rhabdomyosarcoma) and SH-SY5Y cells, which was consistent with previous studies on mice. The SP81 peptide had significantly greater inhibitory effect on EV71 production by NSC and MN compared to the cancer-derived RD and SH-SY5Y cell lines. Hence, this study demonstrates that hESC-derived neural lineages can be utilized as in vitro models for studying EV71 pathogenesis and for screening of Antiviral Therapeutics.
Nicholas S. Heaton - One of the best experts on this subject based on the ideXlab platform.
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Development of a fluorescence based, high-throughput SARS-CoV-2 3CLpro reporter assay
2020Co-Authors: Heather M. Froggatt, Brook E. Heaton, Nicholas S. HeatonAbstract:In late 2019 a human coronavirus, now known as SARS-CoV-2, emerged, likely from a zoonotic reservoir. This virus causes COVID-19 disease, has infected millions of people, and has led to hundreds of thousands of deaths across the globe. While the best interventions to control and ultimately stop the pandemic are prophylactic vaccines, Antiviral Therapeutics are important to limit morbidity and mortality in those already infected. At this time, only one FDA approved anti-SARS-CoV-2 Antiviral drug, remdesivir, is available and unfortunately, its efficacy appears to be limited. Thus, the identification of new and efficacious Antivirals is of highest importance. In order to facilitate rapid drug discovery, flexible, sensitive, and high-throughput screening methods are required. With respect to drug targets, most attention is focused on either the viral RNA-dependent RNA polymerase or the main viral protease, 3CLpro. 3CLpro is an attractive target for Antiviral Therapeutics as it is essential for processing newly translated viral proteins, and the viral lifecycle cannot be completed without protease activity. In this work, we present a new assay to identify inhibitors of the SARS-CoV-2 main protease, 3CLpro. Our reporter is based on a GFP-derived protein that only fluoresces after cleavage by 3CLpro. This experimentally optimized reporter assay allows for Antiviral drug screening in human cell culture at biosafety level-2 (BSL2) with high-throughput compatible protocols. Using this screening approach in combination with existing drug libraries may lead to the rapid identification of novel Antivirals to suppress SARS-CoV-2 replication and spread.
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Influenza viruses that require 10 genomic segments as Antiviral Therapeutics.
PLoS pathogens, 2019Co-Authors: Alfred T. Harding, Griffin D. Haas, Benjamin S. Chambers, Nicholas S. HeatonAbstract:Influenza A viruses (IAVs) encode their genome across eight, negative sense RNA segments. During viral assembly, the failure to package all eight segments, or packaging a mutated segment, renders the resulting virion incompletely infectious. It is known that the accumulation of these defective particles can limit viral disease by interfering with the spread of fully infectious particles. In order to harness this phenomenon therapeutically, we defined which viral packaging signals were amenable to duplication and developed a viral genetic platform which produced replication competent IAVs that require up to two additional artificial genome segments for full infectivity. The modified and artificial genome segments propagated by this approach are capable of acting as “decoy” segments that, when packaged by coinfecting wild-type viruses, lead to the production of non-infectious viral particles. Although IAVs which require 10 genomic segments for full infectivity are able to replicate themselves and spread in vivo, their genomic modifications render them avirulent in mice. Administration of these viruses, both prophylactically and therapeutically, was able to rescue animals from a lethal influenza virus challenge. Together, our results show that replicating IAVs designed to propagate and spread defective genomic segments represent a potent anti-influenza biological therapy that can target the conserved process of particle assembly to limit viral disease.
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Replication competent, 10-segmented influenza viruses as Antiviral Therapeutics
2019Co-Authors: Griffin D. Haas, Alfred T. Harding, Nicholas S. HeatonAbstract:AbstractInfluenza A viruses (IAVs) encode their genome as eight negative sense RNA segments. During viral assembly, the failure to package all eight segments, or packaging of a mutated segment, renders the resultant virion incompletely infectious. It is known that the accumulation of these defective particles can limit viral disease by interfering with the spread of fully infectious particles. In order to harness this phenomenon therapeutically, we defined which viral packaging signals were amenable to duplication and developed a viral genetic platform which allowed the production of replication competent IAVs that package up to two additional artificial genome segments for a total of 10 segments. These artificial genome segments are capable of acting as “decoy” segments that, when packaged by wild-type (WT) viruses, lead to the production of non-infectious viral particles. Despite 10-segmented viruses being able to replicate and spreadin vivo, these genomic modifications render the viruses avirulent. Excitingly, administration of 10-segmented viruses, both prophylactically and therapeutically, was able to rescue animals from normally lethally influenza virus infections. Thus, 10-segmented influenza viruses represent a potent anti-influenza biological therapy that targets the strain-independent process of viral assembly to slow the kinetics of productive viral spread and therefore limit viral disease.Author SummarySeasonal influenza infections are best prevented using vaccination. Vaccination, however, is not capable of completely preventing influenza infection, necessitating the use of anti-influenza Therapeutics. To date, several different classes of anti-influenza Therapeutics have been developed and used in order to combat these infections. Unfortunately, the incidence of influenza resistance to many of these Therapeutics has begun to rise, necessitating the development of new strategies. One such strategy is to mimic the activity of naturally occurring viral particles that harbor defective genomes. These defective interfering particles have the ability to interfere with productive viral assembly, preventing the spread of influenza viruses across the respiratory tract. Furthermore, given the manner in which they target influenza segment packaging, a conserved feature of all influenza A viruses, resistance to this therapeutic strategy is unlikely. Here, we report the development of a genetic platform that allows the production of replication competent, 10-segmented influenza viruses. These viruses are capable of amplifying themselves in isolation, but co-infection with a wild-type virus leads to segment exchange and compromises the spread of both viruses. This interference, while mechanistically distinct from naturally occurring defective particles, was able to target the same viral process and rescue animals exposed to an otherwise lethal viral infection. This viral-based approach may represent a cost effective and scalable method to generate effective anti-influenza Therapeutics when vaccines or anti-viral drugs become ineffective due to acquisition of viral resistance mutations.
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replication competent 10 segmented influenza viruses as Antiviral Therapeutics
bioRxiv, 2019Co-Authors: Griffin D. Haas, Alfred T. Harding, Nicholas S. HeatonAbstract:Influenza A viruses (IAVs) encode their genome as eight negative sense RNA segments. During viral assembly, the failure to package all eight segments, or packaging of a mutated segment, renders the resultant virion incompletely infectious. It is known that the accumulation of these defective particles can limit viral disease by interfering with the spread of fully infectious particles. In order to harness this phenomenon therapeutically, we defined which viral packaging signals were amenable to duplication and developed a viral genetic platform which allowed the production of replication competent IAVs that package up to two additional artificial genome segments for a total of 10 segments. These artificial genome segments are capable of acting as decoy segments that, when packaged by wild-type (WT) viruses, lead to the production of non-infectious viral particles. Despite 10-segmented viruses being able to replicate and spread in vivo, these genomic modifications render the viruses avirulent. Excitingly, administration of 10-segmented viruses, both prophylactically and therapeutically, was able to rescue animals from normally lethally influenza virus infections. Thus, 10-segmented influenza viruses represent a potent anti-influenza biological therapy that targets the strain-independent process of viral assembly to slow the kinetics of productive viral spread and therefore limit viral disease.
Wu Liu - One of the best experts on this subject based on the ideXlab platform.
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Microfluidics in Single-Cell Virology: Technologies and Applications
Trends in biotechnology, 2020Co-Authors: Wu Liu, Si-yang ZhengAbstract:Microfluidics has proven to be a powerful tool for probing biology at the single-cell level. However, it is only in the past 5 years that single-cell microfluidics has been used in the field of virology. An array of strategies based on microwells, microvalves, and droplets is now available for tracking viral infection dynamics, identifying cell subpopulations with particular phenotypes, as well as high-throughput screening. The insights into the virus-host interactions gained at the single-cell level are unprecedented and usually inaccessible by population-based experiments. Therefore, single-cell microfluidics, which opens new avenues for mechanism elucidation and development of Antiviral Therapeutics, would be a valuable tool for the study of viral pathogenesis.
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More than efficacy revealed by single-cell analysis of Antiviral Therapeutics
Science Advances, 2019Co-Authors: Wu Liu, Mehmet Umut Caglar, Zhangming Mao, Andrew Woodman, Jamie J. Arnold, Claus O. Wilke, Craig E. CameronAbstract:Because many aspects of viral infection dynamics and inhibition are governed by stochastic processes, single-cell analysis should provide more information than approaches using population averaging. We have developed a microfluidic device composed of ~6000 wells, with each well containing a microstructure to capture single, infected cells replicating an enterovirus expressing a fluorescent reporter protein. We have used this system to characterize enterovirus inhibitors with distinct mechanisms of action. Single-cell analysis reveals that each class of inhibitor interferes with the viral infection cycle in a manner that can be distinguished by principal component analysis. Single-cell analysis of Antiviral candidates not only reveals efficacy but also facilitates clustering of drugs with the same mechanism of action and provides some indication of the ease with which resistance will develop.
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More than efficacy revealed by single-cell analysis of Antiviral Therapeutics
2019Co-Authors: Wu Liu, Mehmet Umut Caglar, Zhangming Mao, Andrew Woodman, Jamie J. Arnold, Claus O. Wilke, Craig E. CameronAbstract:SUMMARY Development of Antiviral Therapeutics emphasizes minimization of the effective dose and maximization of the toxic dose, first in cell culture and later in animal models. Long-term success of an Antiviral therapeutic is determined not only by its efficacy but also by the duration of time required for drug-resistance to evolve. We have developed a microfluidic device comprised of ~6000 wells, with each well containing a microstructure to capture single cells. We have used this device to characterize enterovirus inhibitors with distinct mechanisms of action. In contrast to population methods, single-cell analysis reveals that each class of inhibitor interferes with the viral infection cycle in a manner that can be distinguished by principal component analysis. Single-cell analysis of Antiviral candidates reveals not only efficacy but also properties of the members of the viral population most sensitive to the drug, the stage of the lifecycle most affected by the drug, and perhaps even if the drug targets an interaction of the virus with its host.
May Shin Yap - One of the best experts on this subject based on the ideXlab platform.
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Pluripotent Human embryonic stem cell derived neural lineages for in vitro modelling of enterovirus 71 infection and therapy
Virology Journal, 2016Co-Authors: May Shin Yap, Yin Quan Tang, Yin Yeo, Wei Ling Lim, Lee Wei Lim, Kuan Onn Tan, Mark Richards, Iekhsan Othman, Chit Laa Poh, Boon Chin HengAbstract:Background The incidence of neurological complications and fatalities associated with Hand, Foot & Mouth disease has increased over recent years, due to emergence of newly-evolved strains of Enterovirus 71 (EV71). In the search for new Antiviral Therapeutics against EV71, accurate and sensitive in vitro cellular models for preliminary studies of EV71 pathogenesis is an essential prerequisite, before progressing to expensive and time-consuming live animal studies and clinical trials. Methods This study thus investigated whether neural lineages derived from pluripotent human embryonic stem cells (hESC) can fulfil this purpose. EV71 infection of hESC-derived neural stem cells (NSC) and mature neurons (MN) was carried out in vitro, in comparison with RD and SH-SY5Y cell lines. Results Upon assessment of post-infection survivability and EV71 production by the various types, it was observed that NSC were significantly more susceptible to EV71 infection compared to MN, RD (rhabdomyosarcoma) and SH-SY5Y cells, which was consistent with previous studies on mice. The SP81 peptide had significantly greater inhibitory effect on EV71 production by NSC and MN compared to the cancer-derived RD and SH-SY5Y cell lines. Conclusions Hence, this study demonstrates that hESC-derived neural lineages can be utilized as in vitro models for studying EV71 pathogenesis and for screening of Antiviral Therapeutics.
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Pluripotent Human embryonic stem cell derived neural lineages for in vitro modelling of enterovirus 71 infection and therapy.
Virology journal, 2016Co-Authors: May Shin Yap, Yin Quan Tang, Yin Yeo, Wei Ling Lim, Lee Wei Lim, Kuan Onn Tan, Mark Richards, Iekhsan Othman, Chit Laa Poh, Boon Chin HengAbstract:The incidence of neurological complications and fatalities associated with Hand, Foot & Mouth disease has increased over recent years, due to emergence of newly-evolved strains of Enterovirus 71 (EV71). In the search for new Antiviral Therapeutics against EV71, accurate and sensitive in vitro cellular models for preliminary studies of EV71 pathogenesis is an essential prerequisite, before progressing to expensive and time-consuming live animal studies and clinical trials. This study thus investigated whether neural lineages derived from pluripotent human embryonic stem cells (hESC) can fulfil this purpose. EV71 infection of hESC-derived neural stem cells (NSC) and mature neurons (MN) was carried out in vitro, in comparison with RD and SH-SY5Y cell lines. Upon assessment of post-infection survivability and EV71 production by the various types, it was observed that NSC were significantly more susceptible to EV71 infection compared to MN, RD (rhabdomyosarcoma) and SH-SY5Y cells, which was consistent with previous studies on mice. The SP81 peptide had significantly greater inhibitory effect on EV71 production by NSC and MN compared to the cancer-derived RD and SH-SY5Y cell lines. Hence, this study demonstrates that hESC-derived neural lineages can be utilized as in vitro models for studying EV71 pathogenesis and for screening of Antiviral Therapeutics.