D Lorne Tyrrell - Academia.edu (original) (raw)
Papers by D Lorne Tyrrell
Analytical Chemistry, May 8, 2023
Journal of the Canadian Association of Gastroenterology, Mar 1, 2019
FEBS Journal, Jul 23, 2014
Cell-death-inducing DFF45-like effector B (CIDEB) is an apoptotic host factor, which was recently... more Cell-death-inducing DFF45-like effector B (CIDEB) is an apoptotic host factor, which was recently found to also regulate hepatic lipid homeostasis. Herein we delineate the relevance of these dual roles of CIDEB in apoptosis and lipid metabolism in the context of hepatitis C virus (HCV) replication. We demonstrate that HCV upregulates CIDEB expression in human serum differentiated hepatoma cells. CIDEB overexpression inhibits HCV replication in HCV replicon expressing Huh7.5 cells, while small interfering RNA knockdown of CIDEB expression in human serum differentiated hepatoma cells promotes HCV replication and secretion of viral proteins. Furthermore, we characterize a CIDEB mutant, KRRA, which is deficient in lipid droplet clustering and fusion and demonstrate that CIDEB-mediated inhibition of HCV is independent of the protein's lipid droplet fusogenic role. Our results suggest that higher levels of CIDEB expression, which favour an apoptotic role for the host factor, inhibit HCV. Collectively, our data demonstrate that CIDEB can act as an anti-HCV host factor and contribute to altered triglyceride homeostasis.
Circulation, Nov 8, 2022
Introduction: Angiostatin - plasmin(ogen) break-down product - is generated by platelets in an ur... more Introduction: Angiostatin - plasmin(ogen) break-down product - is generated by platelets in an urokinase(uPA)-dependent manner. Angiostatin is anti-inflammatory during normoxia, and pro-apoptotic during hypoxia/acidosis. In SARS-CoV-infected mice, the uPA-plasminogen pathway was transcriptionally enriched regulating sublethal vs. lethal infection. Similarly, uPA was transcriptionally upregulated in SARS-CoV-2; however, role of angiostatin has not been investigated. Angiostatin level progressively increases in the blood of severe COVID-19 patients and this increases risk of lethal infection. We aimed to assess role of angiostatin in COVID-19. Methods: VeroE6 cells were infected with wild-type SARS-CoV-2 and treated with angiostatin (140 μg/ml) at pH 7.5 or 6.9. Cell death was quantified by the percentage of detached cells. Immunofluorescent staining against spike protein was used to confirm cellular infection. Additionally, HEK293-ACE2 cells were infected with replication-incompetent SARS-CoV-2 Reporter Virus Particles (RVP; GFP-expressing; Wuhan-Hu-1 D614G) with/without angiostatin and analyzed by flow cytometry. Co-immunoprecipitation of angiostatin and spike protein (Alpha variant) was carried out, followed by SDS-PAGE. Results: At pH 7.5 angiostatin significantly decreased the percentage of detached (24±7 vs 58±6%, p=0.002) VeroE6 following infection, and did not have a significant effect on uninfected cells. Conversely, at pH 6.9 angiostatin alone increased the percentage of detached cells (25% detachment, p=0.014), and failed to reduce the death of infected cells (detachment:53.5±4 vs 50±5%). Angiostatin lowered the percentage of spike protein-positive VeroE6 (59±3 vs 89±3%, p<0.001) and GFP-expressing HEK293-ACE2 (7±0.5 vs 29±3, p=0.001) at both pH 7.4 and 6.9. Spike protein and angiostatin were co-immunoprecipitated. Conclusions: Angiostatin likely has a complex role in COVID-19 pathophysiology. High angiostatin concentrations such as those observed in COVID-19 promote cell death in acidotic microenvironments. Conversely, at physiological pH, angiostatin may have protective effects. Irrespective of pH, angiostatin reduced SARS-CoV-2 cellular entry, possibly by interacting with spike protein.
Frontiers in Immunology, Sep 29, 2017
ACS Measurement Au, Feb 9, 2022
Samples of nasopharyngeal swabs (NPS) are commonly used for the detection of SARS-CoV-2 and diagn... more Samples of nasopharyngeal swabs (NPS) are commonly used for the detection of SARS-CoV-2 and diagnosis of COVID-19. As an alternative, self-collection of saliva and gargle samples minimizes transmission to healthcare workers and relieves the pressure of resources and healthcare personnel during the pandemic. This study aimed to develop an enhanced method enabling simultaneous viral inactivation and RNA preservation during on-site self-collection of saliva and gargle samples. Our method involves the addition of saliva or gargle samples to a newly formulated viral inactivation and RNA preservation (VIP) buffer, concentration of the viral RNA on magnetic beads, and detection of SARS-CoV-2 using reverse transcription quantitative polymerase chain reaction directly from the magnetic beads. This method has a limit of detection of 25 RNA copies per 200 μL of gargle or saliva sample and 9−111 times higher sensitivity than the viral RNA preparation kit recommended by the United States Centers for Disease Control and Prevention. The integrated method was successfully used to analyze more than 200 gargle and saliva samples, including the detection of SARS-CoV-2 in 123 gargle and saliva samples collected daily from two NPS-confirmed positive SARS-CoV-2 patients throughout the course of their infection and recovery. The VIP buffer is stable at room temperature for at least 6 months. SARS-CoV-2 RNA (65 copies/200 μL sample) is stable in the VIP buffer at room temperature for at least 3 weeks. The on-site inactivation of SARS-CoV-2 and preservation of the viral RNA enables self-collection of samples, reduces risks associated with SARS-CoV-2 transmission, and maintains the stability of the target analyte.
![Research paper thumbnail of Erratum to “Saikosaponin b2 is a naturally occurring terpenoid that efficiently inhibits hepatitis C virus entry” [J Hepatol 2015;62:541–548]](https://attachments.academia-assets.com/116210873/thumbnails/1.jpg)
](Journal of Hepatology, Jul 1, 2015
bioRxiv (Cold Spring Harbor Laboratory), Jul 20, 2020
and is also made available for use under a CC0 license.
Journal of the Canadian Association of Gastroenterology, Mar 1, 2023
Frontiers in Immunology
Interferon regulatory factors (IRFs) are key elements of antiviral innate responses that regulate... more Interferon regulatory factors (IRFs) are key elements of antiviral innate responses that regulate the transcription of interferons (IFNs) and IFN-stimulated genes (ISGs). While the sensitivity of human coronaviruses to IFNs has been characterized, antiviral roles of IRFs during human coronavirus infection are not fully understood. Type I or II IFN treatment protected MRC5 cells from human coronavirus 229E infection, but not OC43. Cells infected with 229E or OC43 upregulated ISGs, indicating that antiviral transcription is not suppressed. Antiviral IRFs, IRF1, IRF3 and IRF7, were activated in cells infected with 229E, OC43 or severe acute respiratory syndrome-associated coronavirus 2 (SARS-CoV-2). RNAi knockdown and overexpression of IRFs demonstrated that IRF1 and IRF3 have antiviral properties against OC43, while IRF3 and IRF7 are effective in restricting 229E infection. IRF3 activation effectively promotes transcription of antiviral genes during OC43 or 229E infection. Our study s...
Liver Transplantation, 2010
Human hepatocyte transplantation is an alternative treatment for acute liver failure and liver di... more Human hepatocyte transplantation is an alternative treatment for acute liver failure and liver diseases involving enzyme deficiencies. Although it has been successfully applied in selected recipients, both isolation and transplantation outcomes have the potential to be improved by better donor selection. This study assessed the impact of various donor variables on isolation outcomes (yield and viability) and posttransplant engraftment, using the SCID/Alb-uPA (severe combined immunodeficient/urokinase type plasminogen activator under the control of an albumin promoter) human liver chimeric mouse model. Human hepatocytes were obtained from 90 human liver donor specimens and were transplanted into 3942 mice. Multivariate analysis revealed improved viability with younger donors (P ¼ 0.038) as well as with shorter warm ischemic time (P ¼ 0.012). Hepatocyte engraftment, assessed by the posttransplant level of serum human a1-antitrypsin, was improved with shorter warm ischemia time. Hepatocytes isolated from older donors (!60 years) had lower viability and posttransplant engraftment (P 0.01). In conclusion, the selection of young donors (<60 years) and rapid liver specimen retrieval, allowing for shorter warm ischemia time, are key determinants for the success of both the isolation of high viability human hepatocytes and their subsequent posttransplantation capacity for engraftment and expansion.
Antimicrobial Agents and Chemotherapy, 2002
N -Nonyl-deoxy-galactonojirimycin ( N -nonyl-DGJ) has been shown to reduce the amount of hepatiti... more N -Nonyl-deoxy-galactonojirimycin ( N -nonyl-DGJ) has been shown to reduce the amount of hepatitis B virus (HBV) produced by tissue cultures under conditions where cell viability is not affected. We show here that the compound N -nonyl-DGJ was effective against lamivudine-resistant HBV mutants bearing the YMDD motif in the polymerase gene, consistent with the compound's activity being distinct from those of nucleoside inhibitors. To better understand the chemical structures that influence its antiviral activity, a series of imino sugar derivatives were made and tested for their antiviral activity against HBV. This work suggests that the antiviral activity of the alkovirs requires an alkyl chain length of at least eight carbons but that the galactose-based head group can be modified with little or no loss in activity.
Biochemical and Biophysical Research Communications, Nov 1, 1988
... The DNA washybridized with the 32p-nick translated DHBV probe.Extraction of DNA from hepatocy... more ... The DNA washybridized with the 32p-nick translated DHBV probe.Extraction of DNA from hepatocyte cultures: The methoddescribed by Tuttleman et al (9) was used. ... 58, 17-25.ii. Lein, JM, Petcu, DG, Aldrich, CE, and Mason, WS(1987) J. Virol.61, 3832-3840.12. ...
ChemInform, Sep 26, 1989
The deoxyuridines (I) react with the succinimides (II) to give the compounds (III).
PubMed, Aug 19, 2000
Purpose: The objective of this study was to evaluate a dual action prodrug concept wherein an unn... more Purpose: The objective of this study was to evaluate a dual action prodrug concept wherein an unnatural myristic acid analogue is coupled via an ester moiety to the 5'-position of FLT or AZT. Subsequent intracellular cleavage of the prodrug ester would simultaneously release FLT or AZT that could inhibit reverse transcriptase (RT), and the myristic acid analogue that could inhibit myristoyl-CoA:protein N-myristoyltransferase (NMT). Methods: Cytotoxicity (2.2.15 cell culture), and anti-hepatitis B activity of 5'-O-myristoyl analogue prodrug derivatives of FLT and AZT (2-8) were evaluated in vitro using human liver hepatitis B virus (HBV) producing 2.2.15 cell lines. Results: The 5'-O-(12-methoxydodecanoyl) ester derivatives of AZT (2, EC(50) = 2. 7 +/- 0.3 microM; CC(50)= 727 +/- 19 microM) and FLT (4, EC(50)= 2.8 +/- 0.3 microM; CC(50)= 186 +/- 20 microM) were the most effective anti-hepatitis B virus (anti-HBV) compounds of this series in a replication assay. In the series of 5'-O-myristic acid analogue ester prodrug derivatives of FLT, the relative anti-HBV potency order was MeO(CH(2))(11)CO(2)- > N(3)(CH(2))(11)CO(2)- and Br(CH(2))(11)CO(2)- > EtS(CH(2))(n)CO(2)- (n = 10 or 11) > Me(CH(2))(12)CO(2)- (myristoyl). Conclusions: The in vitro data suggest that the 5'-O-myristoyl analogue prodrug concept offers a potential drug design approach to design dual acting antiviral agents, with superior pharmacokinetic, biodistribution, reduced cytotoxicity and/or increased efficacy. In this regard, the 5'-O-(12-methoxydodecanoyl) prodrug ester of 3'-thia-3'-deoxythymidine (3TC) may offer the greatest potential for the treatment of HBV infection.
Biochemical Pharmacology, May 1, 1995
The intracellular fate of the potent duck hepatitis B virus (DHBV) inhibitor 2,6_diaminopurine 2'... more The intracellular fate of the potent duck hepatitis B virus (DHBV) inhibitor 2,6_diaminopurine 2',3'-dideoxyriboside (ddDAPR), its deamination product 2',3'-dideoxyguanosine (ddG), and the less effective DHBV-inhibitor 2',3'-dideoxycytidine (ddC) was investigated in duck hepatocyte primary cultures. After a l-mitt exposure of [3H]ddDAPR to duck blood, 95% of the compound was converted to ddG. Similarly, [3H]ddDAPR was converted rapidly to ddG in duck hepatocyte primary cultures, with ddG exhibiting resistance to further catabolism. The major pathway of ddG utilization in these cells was phosphorylation, yielding a concentration of 2.1 and 1.9 ,wM total ddG nucleotides after 5 and 26 hr, respectively, of exposure to 4 PM ddG. Removal of exogenous ddG led to a rapid (Ti,r = 1.6 hr) decrease in the total intracellular ddG nucleotide pools. Duck hepatocytes treated with 4pMddC exhibited a time-dependent accumulation of ddC nucleotides, culminating in a maximum intracellular total ddC nucleotide concentration of 1.4 PM after 2426 hr. The intracellular total ddC nucleotide level decreased with a T,,* of 4.4 hr following the removal of exogenous ddC. The formation of ddC nucleotides was reduced in the presence of excess 2'-dideoxycytidine implicating deoxycytidine kinase in the initial step of ddC phosphorylation. A 25fold excess of 2'-deoxycytidine had no effect on ddG phosphorylation in duck hepatocytes. However, a 92% inhibition of ddG nucleotide formation occurred in duck hepal:ocytes treated for 5 hr with 4 PM ['H]ddG + 100 PM adenosine in the presence of the adenosine deaminase inhibitor 2'-deoxycoformycin, suggesting that, in these cells, adenosine kinase is involved in the ddG phosphorylation process.
ACS Applied Materials & Interfaces, Jun 14, 2023
bioRxiv (Cold Spring Harbor Laboratory), May 10, 2021
SARS-CoV-2 is the etiological agent of COVID19. There are currently several licensed vaccines app... more SARS-CoV-2 is the etiological agent of COVID19. There are currently several licensed vaccines approved for human use and most of them are targeting the spike protein (or virion) in the virion envelope to induce protective immunity. Recently, variants that spread more quickly have emerged. There is evidence that some of these variants are less sensitive to neutralization in vitro, but it is not clear whether they can evade vaccine induced protection. In this study, we tested the utility of SARS-CoV-2 spike RBD as a vaccine antigen and explore the effect of formulation with Alum/MPLA or AddaS03 adjuvants. Our results indicate RBD induces high titers of neutralizing antibodies and activates strong cellular immune responses. There is also significant cross-neutralisation of variants B1.1.7 and B.1.351 and to a lesser extent, SARS-CoV-1. These results indicate that recombinant RBD can be a viable candidate as a stand-alone vaccine or as a booster shot to diversify our strategy for COVID19 protection.
PubMed, Nov 1, 1988
Liposomal encapsulation of radioiodinated anti-herpes nucleosides was undertaken to reduce metabo... more Liposomal encapsulation of radioiodinated anti-herpes nucleosides was undertaken to reduce metabolic inactivation and increase blood-brain barrier penetration of the nucleosides, and so provide formulations suitable for use in the non-invasive scintigraphic diagnosis of herpes simplex encephalitis (HSE). The nucleosides investigated were [125I]-IVdU and its more lipophilic 3-methyl derivative, and they were encapsulated in phosphatidylcholine:cholesterol:sulfatide liposomes with an efficiency of 4.4% and 1.7% respectively. The encapsulation of IVdU reduced in vitro phosphorolysis (by 31% in serum at 25 degrees C over 3 hr as compared with non-encapsulated IVdU), and markedly increased in vivo stability (fifty-fold greater than that of free drug). In normal mice, higher radioactivity levels were observed in most tissues and there was prolonged, constant blood and brain uptake. Biodistribution of liposomal IVdU in herpes simplex virus (HSV) infected animals was similar, but somewhat lower concentrations were attained. Liposomal encapsulation of [125I]-3-CH3-IVdU produced less dramatic changes in tissue distribution in either healthy or HSV infected mice, as compared with the non-encapsulated drug. Whilst, in HSV infected mice, liposomal encapsulation of both drugs caused increased uptake by spleen, liver and lung, the uptake by brain was still too low for detection by whole-body scintigraphy.
PubMed, Mar 1, 1987
The biological activities of 1-(2'-deoxy-2'-fluoro-beta-D-arabinofuranosyl)-5-fluorouracil (2'F-a... more The biological activities of 1-(2'-deoxy-2'-fluoro-beta-D-arabinofuranosyl)-5-fluorouracil (2'F-ara-FU), 1-(3'-deoxy-3'-fluoro-beta-D-arabinofuranosyl)-5-fluorouracil (3'F-ara-FU) and 1-beta-D-arabinofuranosylthymine (ara-T) were compared in human cytomegalovirus (HCMV)-infected and noninfected human fibroblasts. 2'F-ara-FU inhibited HCMV plaque formation (ED50, 16 microM for AD 169 strain) at lower concentrations than 3'F-ara-FU (ED50, 100 microM for AD 169). These nucleoside analogues are expected to be phosphorylated to their 5'-phosphate forms by cellular thymidine kinase in HCMV-infected cells. The thymidine kinase activities in the virus-infected and noninfected cells were compared. Cellular thymidine kinase was increased in the virus-infected cells and showed better phosphorylation of 2'F-ara-FU than did 3'F-ara-FU. HCMV DNA polymerase was purified using affinity column chromatography, and the inhibitory effect of the 5'-triphosphate derivatives of 2'F-ara-FU (2'F-ara-FUTP) and 3'F-ara-FU (3'F-ara-FUTP) against viral and host DNA polymerase alpha was examined. No significant difference in the effectiveness of inhibition was observed between viral DNA polymerase and host polymerase alpha. However, viral polymerase incorporated 2'F-ara-FUTP into newly synthesized DNA, whereas polymerase alpha did not utilize 2'F-ara-FUTP as a substrate. Thus, viral polymerase differs from host polymerase alpha in its recognition and utilization of 2'F-ara-FUTP. This difference may be important to the design of selective antiviral agents for HCMV.
Analytical Chemistry, May 8, 2023
Journal of the Canadian Association of Gastroenterology, Mar 1, 2019
FEBS Journal, Jul 23, 2014
Cell-death-inducing DFF45-like effector B (CIDEB) is an apoptotic host factor, which was recently... more Cell-death-inducing DFF45-like effector B (CIDEB) is an apoptotic host factor, which was recently found to also regulate hepatic lipid homeostasis. Herein we delineate the relevance of these dual roles of CIDEB in apoptosis and lipid metabolism in the context of hepatitis C virus (HCV) replication. We demonstrate that HCV upregulates CIDEB expression in human serum differentiated hepatoma cells. CIDEB overexpression inhibits HCV replication in HCV replicon expressing Huh7.5 cells, while small interfering RNA knockdown of CIDEB expression in human serum differentiated hepatoma cells promotes HCV replication and secretion of viral proteins. Furthermore, we characterize a CIDEB mutant, KRRA, which is deficient in lipid droplet clustering and fusion and demonstrate that CIDEB-mediated inhibition of HCV is independent of the protein&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s lipid droplet fusogenic role. Our results suggest that higher levels of CIDEB expression, which favour an apoptotic role for the host factor, inhibit HCV. Collectively, our data demonstrate that CIDEB can act as an anti-HCV host factor and contribute to altered triglyceride homeostasis.
Circulation, Nov 8, 2022
Introduction: Angiostatin - plasmin(ogen) break-down product - is generated by platelets in an ur... more Introduction: Angiostatin - plasmin(ogen) break-down product - is generated by platelets in an urokinase(uPA)-dependent manner. Angiostatin is anti-inflammatory during normoxia, and pro-apoptotic during hypoxia/acidosis. In SARS-CoV-infected mice, the uPA-plasminogen pathway was transcriptionally enriched regulating sublethal vs. lethal infection. Similarly, uPA was transcriptionally upregulated in SARS-CoV-2; however, role of angiostatin has not been investigated. Angiostatin level progressively increases in the blood of severe COVID-19 patients and this increases risk of lethal infection. We aimed to assess role of angiostatin in COVID-19. Methods: VeroE6 cells were infected with wild-type SARS-CoV-2 and treated with angiostatin (140 μg/ml) at pH 7.5 or 6.9. Cell death was quantified by the percentage of detached cells. Immunofluorescent staining against spike protein was used to confirm cellular infection. Additionally, HEK293-ACE2 cells were infected with replication-incompetent SARS-CoV-2 Reporter Virus Particles (RVP; GFP-expressing; Wuhan-Hu-1 D614G) with/without angiostatin and analyzed by flow cytometry. Co-immunoprecipitation of angiostatin and spike protein (Alpha variant) was carried out, followed by SDS-PAGE. Results: At pH 7.5 angiostatin significantly decreased the percentage of detached (24±7 vs 58±6%, p=0.002) VeroE6 following infection, and did not have a significant effect on uninfected cells. Conversely, at pH 6.9 angiostatin alone increased the percentage of detached cells (25% detachment, p=0.014), and failed to reduce the death of infected cells (detachment:53.5±4 vs 50±5%). Angiostatin lowered the percentage of spike protein-positive VeroE6 (59±3 vs 89±3%, p<0.001) and GFP-expressing HEK293-ACE2 (7±0.5 vs 29±3, p=0.001) at both pH 7.4 and 6.9. Spike protein and angiostatin were co-immunoprecipitated. Conclusions: Angiostatin likely has a complex role in COVID-19 pathophysiology. High angiostatin concentrations such as those observed in COVID-19 promote cell death in acidotic microenvironments. Conversely, at physiological pH, angiostatin may have protective effects. Irrespective of pH, angiostatin reduced SARS-CoV-2 cellular entry, possibly by interacting with spike protein.
Frontiers in Immunology, Sep 29, 2017
ACS Measurement Au, Feb 9, 2022
Samples of nasopharyngeal swabs (NPS) are commonly used for the detection of SARS-CoV-2 and diagn... more Samples of nasopharyngeal swabs (NPS) are commonly used for the detection of SARS-CoV-2 and diagnosis of COVID-19. As an alternative, self-collection of saliva and gargle samples minimizes transmission to healthcare workers and relieves the pressure of resources and healthcare personnel during the pandemic. This study aimed to develop an enhanced method enabling simultaneous viral inactivation and RNA preservation during on-site self-collection of saliva and gargle samples. Our method involves the addition of saliva or gargle samples to a newly formulated viral inactivation and RNA preservation (VIP) buffer, concentration of the viral RNA on magnetic beads, and detection of SARS-CoV-2 using reverse transcription quantitative polymerase chain reaction directly from the magnetic beads. This method has a limit of detection of 25 RNA copies per 200 μL of gargle or saliva sample and 9−111 times higher sensitivity than the viral RNA preparation kit recommended by the United States Centers for Disease Control and Prevention. The integrated method was successfully used to analyze more than 200 gargle and saliva samples, including the detection of SARS-CoV-2 in 123 gargle and saliva samples collected daily from two NPS-confirmed positive SARS-CoV-2 patients throughout the course of their infection and recovery. The VIP buffer is stable at room temperature for at least 6 months. SARS-CoV-2 RNA (65 copies/200 μL sample) is stable in the VIP buffer at room temperature for at least 3 weeks. The on-site inactivation of SARS-CoV-2 and preservation of the viral RNA enables self-collection of samples, reduces risks associated with SARS-CoV-2 transmission, and maintains the stability of the target analyte.
Journal of Hepatology, Jul 1, 2015
bioRxiv (Cold Spring Harbor Laboratory), Jul 20, 2020
and is also made available for use under a CC0 license.
Journal of the Canadian Association of Gastroenterology, Mar 1, 2023
Frontiers in Immunology
Interferon regulatory factors (IRFs) are key elements of antiviral innate responses that regulate... more Interferon regulatory factors (IRFs) are key elements of antiviral innate responses that regulate the transcription of interferons (IFNs) and IFN-stimulated genes (ISGs). While the sensitivity of human coronaviruses to IFNs has been characterized, antiviral roles of IRFs during human coronavirus infection are not fully understood. Type I or II IFN treatment protected MRC5 cells from human coronavirus 229E infection, but not OC43. Cells infected with 229E or OC43 upregulated ISGs, indicating that antiviral transcription is not suppressed. Antiviral IRFs, IRF1, IRF3 and IRF7, were activated in cells infected with 229E, OC43 or severe acute respiratory syndrome-associated coronavirus 2 (SARS-CoV-2). RNAi knockdown and overexpression of IRFs demonstrated that IRF1 and IRF3 have antiviral properties against OC43, while IRF3 and IRF7 are effective in restricting 229E infection. IRF3 activation effectively promotes transcription of antiviral genes during OC43 or 229E infection. Our study s...
Liver Transplantation, 2010
Human hepatocyte transplantation is an alternative treatment for acute liver failure and liver di... more Human hepatocyte transplantation is an alternative treatment for acute liver failure and liver diseases involving enzyme deficiencies. Although it has been successfully applied in selected recipients, both isolation and transplantation outcomes have the potential to be improved by better donor selection. This study assessed the impact of various donor variables on isolation outcomes (yield and viability) and posttransplant engraftment, using the SCID/Alb-uPA (severe combined immunodeficient/urokinase type plasminogen activator under the control of an albumin promoter) human liver chimeric mouse model. Human hepatocytes were obtained from 90 human liver donor specimens and were transplanted into 3942 mice. Multivariate analysis revealed improved viability with younger donors (P ¼ 0.038) as well as with shorter warm ischemic time (P ¼ 0.012). Hepatocyte engraftment, assessed by the posttransplant level of serum human a1-antitrypsin, was improved with shorter warm ischemia time. Hepatocytes isolated from older donors (!60 years) had lower viability and posttransplant engraftment (P 0.01). In conclusion, the selection of young donors (<60 years) and rapid liver specimen retrieval, allowing for shorter warm ischemia time, are key determinants for the success of both the isolation of high viability human hepatocytes and their subsequent posttransplantation capacity for engraftment and expansion.
Antimicrobial Agents and Chemotherapy, 2002
N -Nonyl-deoxy-galactonojirimycin ( N -nonyl-DGJ) has been shown to reduce the amount of hepatiti... more N -Nonyl-deoxy-galactonojirimycin ( N -nonyl-DGJ) has been shown to reduce the amount of hepatitis B virus (HBV) produced by tissue cultures under conditions where cell viability is not affected. We show here that the compound N -nonyl-DGJ was effective against lamivudine-resistant HBV mutants bearing the YMDD motif in the polymerase gene, consistent with the compound's activity being distinct from those of nucleoside inhibitors. To better understand the chemical structures that influence its antiviral activity, a series of imino sugar derivatives were made and tested for their antiviral activity against HBV. This work suggests that the antiviral activity of the alkovirs requires an alkyl chain length of at least eight carbons but that the galactose-based head group can be modified with little or no loss in activity.
Biochemical and Biophysical Research Communications, Nov 1, 1988
... The DNA washybridized with the 32p-nick translated DHBV probe.Extraction of DNA from hepatocy... more ... The DNA washybridized with the 32p-nick translated DHBV probe.Extraction of DNA from hepatocyte cultures: The methoddescribed by Tuttleman et al (9) was used. ... 58, 17-25.ii. Lein, JM, Petcu, DG, Aldrich, CE, and Mason, WS(1987) J. Virol.61, 3832-3840.12. ...
ChemInform, Sep 26, 1989
The deoxyuridines (I) react with the succinimides (II) to give the compounds (III).
PubMed, Aug 19, 2000
Purpose: The objective of this study was to evaluate a dual action prodrug concept wherein an unn... more Purpose: The objective of this study was to evaluate a dual action prodrug concept wherein an unnatural myristic acid analogue is coupled via an ester moiety to the 5'-position of FLT or AZT. Subsequent intracellular cleavage of the prodrug ester would simultaneously release FLT or AZT that could inhibit reverse transcriptase (RT), and the myristic acid analogue that could inhibit myristoyl-CoA:protein N-myristoyltransferase (NMT). Methods: Cytotoxicity (2.2.15 cell culture), and anti-hepatitis B activity of 5'-O-myristoyl analogue prodrug derivatives of FLT and AZT (2-8) were evaluated in vitro using human liver hepatitis B virus (HBV) producing 2.2.15 cell lines. Results: The 5'-O-(12-methoxydodecanoyl) ester derivatives of AZT (2, EC(50) = 2. 7 +/- 0.3 microM; CC(50)= 727 +/- 19 microM) and FLT (4, EC(50)= 2.8 +/- 0.3 microM; CC(50)= 186 +/- 20 microM) were the most effective anti-hepatitis B virus (anti-HBV) compounds of this series in a replication assay. In the series of 5'-O-myristic acid analogue ester prodrug derivatives of FLT, the relative anti-HBV potency order was MeO(CH(2))(11)CO(2)- > N(3)(CH(2))(11)CO(2)- and Br(CH(2))(11)CO(2)- > EtS(CH(2))(n)CO(2)- (n = 10 or 11) > Me(CH(2))(12)CO(2)- (myristoyl). Conclusions: The in vitro data suggest that the 5'-O-myristoyl analogue prodrug concept offers a potential drug design approach to design dual acting antiviral agents, with superior pharmacokinetic, biodistribution, reduced cytotoxicity and/or increased efficacy. In this regard, the 5'-O-(12-methoxydodecanoyl) prodrug ester of 3'-thia-3'-deoxythymidine (3TC) may offer the greatest potential for the treatment of HBV infection.
Biochemical Pharmacology, May 1, 1995
The intracellular fate of the potent duck hepatitis B virus (DHBV) inhibitor 2,6_diaminopurine 2'... more The intracellular fate of the potent duck hepatitis B virus (DHBV) inhibitor 2,6_diaminopurine 2',3'-dideoxyriboside (ddDAPR), its deamination product 2',3'-dideoxyguanosine (ddG), and the less effective DHBV-inhibitor 2',3'-dideoxycytidine (ddC) was investigated in duck hepatocyte primary cultures. After a l-mitt exposure of [3H]ddDAPR to duck blood, 95% of the compound was converted to ddG. Similarly, [3H]ddDAPR was converted rapidly to ddG in duck hepatocyte primary cultures, with ddG exhibiting resistance to further catabolism. The major pathway of ddG utilization in these cells was phosphorylation, yielding a concentration of 2.1 and 1.9 ,wM total ddG nucleotides after 5 and 26 hr, respectively, of exposure to 4 PM ddG. Removal of exogenous ddG led to a rapid (Ti,r = 1.6 hr) decrease in the total intracellular ddG nucleotide pools. Duck hepatocytes treated with 4pMddC exhibited a time-dependent accumulation of ddC nucleotides, culminating in a maximum intracellular total ddC nucleotide concentration of 1.4 PM after 2426 hr. The intracellular total ddC nucleotide level decreased with a T,,* of 4.4 hr following the removal of exogenous ddC. The formation of ddC nucleotides was reduced in the presence of excess 2'-dideoxycytidine implicating deoxycytidine kinase in the initial step of ddC phosphorylation. A 25fold excess of 2'-deoxycytidine had no effect on ddG phosphorylation in duck hepatocytes. However, a 92% inhibition of ddG nucleotide formation occurred in duck hepal:ocytes treated for 5 hr with 4 PM ['H]ddG + 100 PM adenosine in the presence of the adenosine deaminase inhibitor 2'-deoxycoformycin, suggesting that, in these cells, adenosine kinase is involved in the ddG phosphorylation process.
ACS Applied Materials & Interfaces, Jun 14, 2023
bioRxiv (Cold Spring Harbor Laboratory), May 10, 2021
SARS-CoV-2 is the etiological agent of COVID19. There are currently several licensed vaccines app... more SARS-CoV-2 is the etiological agent of COVID19. There are currently several licensed vaccines approved for human use and most of them are targeting the spike protein (or virion) in the virion envelope to induce protective immunity. Recently, variants that spread more quickly have emerged. There is evidence that some of these variants are less sensitive to neutralization in vitro, but it is not clear whether they can evade vaccine induced protection. In this study, we tested the utility of SARS-CoV-2 spike RBD as a vaccine antigen and explore the effect of formulation with Alum/MPLA or AddaS03 adjuvants. Our results indicate RBD induces high titers of neutralizing antibodies and activates strong cellular immune responses. There is also significant cross-neutralisation of variants B1.1.7 and B.1.351 and to a lesser extent, SARS-CoV-1. These results indicate that recombinant RBD can be a viable candidate as a stand-alone vaccine or as a booster shot to diversify our strategy for COVID19 protection.
PubMed, Nov 1, 1988
Liposomal encapsulation of radioiodinated anti-herpes nucleosides was undertaken to reduce metabo... more Liposomal encapsulation of radioiodinated anti-herpes nucleosides was undertaken to reduce metabolic inactivation and increase blood-brain barrier penetration of the nucleosides, and so provide formulations suitable for use in the non-invasive scintigraphic diagnosis of herpes simplex encephalitis (HSE). The nucleosides investigated were [125I]-IVdU and its more lipophilic 3-methyl derivative, and they were encapsulated in phosphatidylcholine:cholesterol:sulfatide liposomes with an efficiency of 4.4% and 1.7% respectively. The encapsulation of IVdU reduced in vitro phosphorolysis (by 31% in serum at 25 degrees C over 3 hr as compared with non-encapsulated IVdU), and markedly increased in vivo stability (fifty-fold greater than that of free drug). In normal mice, higher radioactivity levels were observed in most tissues and there was prolonged, constant blood and brain uptake. Biodistribution of liposomal IVdU in herpes simplex virus (HSV) infected animals was similar, but somewhat lower concentrations were attained. Liposomal encapsulation of [125I]-3-CH3-IVdU produced less dramatic changes in tissue distribution in either healthy or HSV infected mice, as compared with the non-encapsulated drug. Whilst, in HSV infected mice, liposomal encapsulation of both drugs caused increased uptake by spleen, liver and lung, the uptake by brain was still too low for detection by whole-body scintigraphy.
PubMed, Mar 1, 1987
The biological activities of 1-(2'-deoxy-2'-fluoro-beta-D-arabinofuranosyl)-5-fluorouracil (2'F-a... more The biological activities of 1-(2'-deoxy-2'-fluoro-beta-D-arabinofuranosyl)-5-fluorouracil (2'F-ara-FU), 1-(3'-deoxy-3'-fluoro-beta-D-arabinofuranosyl)-5-fluorouracil (3'F-ara-FU) and 1-beta-D-arabinofuranosylthymine (ara-T) were compared in human cytomegalovirus (HCMV)-infected and noninfected human fibroblasts. 2'F-ara-FU inhibited HCMV plaque formation (ED50, 16 microM for AD 169 strain) at lower concentrations than 3'F-ara-FU (ED50, 100 microM for AD 169). These nucleoside analogues are expected to be phosphorylated to their 5'-phosphate forms by cellular thymidine kinase in HCMV-infected cells. The thymidine kinase activities in the virus-infected and noninfected cells were compared. Cellular thymidine kinase was increased in the virus-infected cells and showed better phosphorylation of 2'F-ara-FU than did 3'F-ara-FU. HCMV DNA polymerase was purified using affinity column chromatography, and the inhibitory effect of the 5'-triphosphate derivatives of 2'F-ara-FU (2'F-ara-FUTP) and 3'F-ara-FU (3'F-ara-FUTP) against viral and host DNA polymerase alpha was examined. No significant difference in the effectiveness of inhibition was observed between viral DNA polymerase and host polymerase alpha. However, viral polymerase incorporated 2'F-ara-FUTP into newly synthesized DNA, whereas polymerase alpha did not utilize 2'F-ara-FUTP as a substrate. Thus, viral polymerase differs from host polymerase alpha in its recognition and utilization of 2'F-ara-FUTP. This difference may be important to the design of selective antiviral agents for HCMV.