scholarly journals Enisamium is an inhibitor of the SARS-CoV-2 RNA polymerase and shows improvement of recovery in COVID-19 patients in an interim analysis of a clinical trial

2021 ◽  
Author(s):  
Olha Holubovska ◽  
Denisa Bojkova ◽  
Stefano Elli ◽  
Marco Bechtel ◽  
David Boltz ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Medical Care ◽  
Interim Analysis ◽  
Influenza A ◽  
Controlled Trial ◽  
Rank Test ◽  
Supplementary Oxygen ◽  
Active Inhibitor ◽  
Significant Difference

AbstractPandemic SARS-CoV-2 causes a mild to severe respiratory disease called Coronavirus Disease 2019 (COVID-19). Control of SARS-CoV-2 spread will depend on vaccine-induced or naturally acquired protective herd immunity. Until then, antiviral strategies are needed to manage COVID-19, but approved antiviral treatments, such as remdesivir, can only be delivered intravenously. Enisamium (laboratory code FAV00A, trade name Amizon®) is an orally active inhibitor of influenza A and B viruses in cell culture and clinically approved in countries of the Commonwealth of Independent States. Here we show that enisamium can inhibit SARS-CoV-2 infections in NHBE and Caco-2 cells. In vitro, the previously identified enisamium metabolite VR17-04 directly inhibits the activity of the SARS-CoV-2 RNA polymerase. Docking and molecular dynamics simulations suggest that VR17-04 prevents GTP and UTP incorporation. To confirm enisamium’s antiviral properties, we conducted a double-blind, randomized, placebo-controlled trial in adult, hospitalized COVID-19 patients, which needed medical care either with or without supplementary oxygen. Patients received either enisamium (500 mg per dose) or placebo for 7 days. A pre-planned interim analysis showed in the subgroup of patients needing supplementary oxygen (n = 77) in the enisamium group a mean recovery time of 11.1 days, compared to 13.9 days for the placebo group (log-rank test; p=0.0259). No significant difference was found for all patients (n = 373) or those only needing medical care (n = 296). These results thus suggest that enisamium is an inhibitor of SARS-CoV-2 RNA synthesis and that enisamium treatment shortens the time to recovery for COVID-19 patients needing oxygen.Significance statementSARS-CoV-2 is the causative agent of COVID-19. Although vaccines are now becoming available to prevent SARS-CoV-2 spread, the development of antivirals remains necessary for treating current COVID-19 patients and combating future coronavirus outbreaks. Here, we report that enisamium, which can be administered orally, can prevent SARS-CoV-2 replication and that its metabolite VR17-04 can inhibit the SARS-CoV-2 RNA polymerase in vitro. Moreover, we find that COVID-19 patients requiring supplementary oxygen, recover more quickly than patients treated with a placebo. Enisamium may therefore be an accessible treatment for COVID-19 patients.

scholarly journals The human H5N1 influenza A virus polymerase complex is active in vitro over a broad range of temperatures, in contrast to the WSN complex, and this property can be attributed to the PB2 subunit

2008 ◽  
Vol 89 (12) ◽  
pp. 2923-2932 ◽  
Author(s):  
Birgit G. Bradel-Tretheway ◽  
Z. Kelley ◽  
Shikha Chakraborty-Sett ◽  
Toru Takimoto ◽  
Baek Kim ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Influenza A Virus ◽  
Influenza A ◽  
Elevated Temperatures ◽  
Expression System ◽  
Lung Epithelial Cells ◽  
Upper Respiratory Tract ◽  
Polymerase Complex ◽  
H5n1 Influenza

Influenza A virus (IAV) replicates in the upper respiratory tract of humans at 33 °C and in the intestinal tract of birds at close to 41 °C. The viral RNA polymerase complex comprises three subunits (PA, PB1 and PB2) and plays an important role in host adaptation. We therefore developed an in vitro system to examine the temperature sensitivity of IAV RNA polymerase complexes from different origins. Complexes were prepared from human lung epithelial cells (A549) using a novel adenoviral expression system. Affinity-purified complexes were generated that contained either all three subunits (PA/PB1/PB2) from the A/Viet/1203/04 H5N1 virus (H/H/H) or the A/WSN/33 H1N1 strain (W/W/W). We also prepared chimeric complexes in which the PB2 subunit was exchanged (H/H/W, W/W/H) or substituted with an avian PB2 from the A/chicken/Nanchang/3-120/01 H3N2 strain (W/W/N). All complexes were functional in transcription, cap-binding and endonucleolytic activity. Complexes containing the H5N1 or Nanchang PB2 protein retained transcriptional activity over a broad temperature range (30–42 °C). In contrast, complexes containing the WSN PB2 protein lost activity at elevated temperatures (39 °C or higher). The E627K mutation in the avian PB2 was not required for this effect. Finally, the avian PB2 subunit was shown to confer enhanced stability to the WSN 3P complex. These results show that PB2 plays an important role in regulating the temperature optimum for IAV RNA polymerase activity, possibly due to effects on the functional stability of the 3P complex.

scholarly journals Recovery rate of children with moderate acute malnutrition treated with ready-to-use supplementary food (RUSF) or improved corn–soya blend (CSB+): a randomized controlled trial

2015 ◽  
Vol 19 (2) ◽  
pp. 363-370 ◽  
Author(s):  
Gabriel Nama Medoua ◽  
Patricia M Ntsama ◽  
Anne Christine A Ndzana ◽  
Véronique J Essa’a ◽  
Julie Judith T Tsafack ◽  
...  
Keyword(s):  
Nutrition Education ◽  
Recovery Rate ◽  
Energy Requirement ◽  
Controlled Trial ◽  
Acute Malnutrition ◽  
Rank Test ◽  
Supplementary Food ◽  
Significant Difference ◽  
Moderate Acute Malnutrition

AbstractObjectiveTo compare an improved corn–soya blend (CSB+) with a ready-to-use supplementary food (RUSF) to test the hypothesis that satisfactory recovery rate will be achieved with CSB+ or RUSF when these foods provide 50 % of the child’s energy requirement, the 50 % remaining coming from usual diet.DesignA comparative efficacy trial study was conducted with moderately wasted children, using a controlled randomized design, with parallel assignment for RUSF or CSB+. Every child received a daily ration of 167 kJ (40 kcal)/kg body weight during 56 d with a follow-up performed every 14 d. Every caregiver received nutrition counselling at enrolment and at each follow-up visit.SettingHealth districts of Mvog-Beti and Evodoula in the Centre region of Cameroon.SubjectsEight hundred and thirty-three children aged 6–59 months were screened and eighty-one malnourished children (weight-for-height Z-score between −3 and −2) aged 25–59 months were selected.ResultsOf children treated with CSB+ and RUSF, 73 % (95 % CI 59 %, 87 %) and 85 % (95 % CI 73 %, 97 %), respectively, recovered from moderate acute malnutrition, with no significant difference between groups. The mean duration of treatment required to achieve recovery was 44 d in the RUSF group and 51 d in the CSB+ group (log-rank test, P=0·0048).ConclusionsThere was no significant difference in recovery rate between the groups. Both CSB+ and RUSF were relatively successful for the treatment of moderate acute malnutrition in children. Despite the relatively low ration size provided, the recovery rates observed for both groups were comparable to or higher than those reported in previous studies, a probable effect of nutrition education.

scholarly journals Cheilectomy With or Without Cryopreserved Umbilical Cord for Hallux Rigidus: A Prospective Randomized Controlled Trial

2019 ◽  
Vol 4 (4) ◽  
pp. 2473011419S0020
Author(s):  
Sara Heintzman ◽  
Chad Ferguson ◽  
W. Hodges Davis ◽  
Robert B. Anderson ◽  
Bruce E. Cohen ◽  
...  
Keyword(s):  
Range Of Motion ◽  
Umbilical Cord ◽  
Articular Surface ◽  
Controlled Trial ◽  
Treatment Algorithm ◽  
Hallux Rigidus ◽  
Least Square ◽  
Rank Test ◽  
Motion Data ◽  
Significant Difference

Category: Midfoot/Forefoot Introduction/Purpose: Arthritis of the first MTP joint (hallux rigidus) is the most common form of osteoarthritis affecting the foot. Despite advances in interpositional techniques and devices, dorsal cheilectomy remains part of the treatment algorithm after failed conservative treatment of hallux rigidus. Dorsal cheilectomy aims to surgically remove dorsal impingement and improve pain and function as well as range of motion. However, prospective data on outcomes following this procedure is lacking. Cryopreserved umbilical cord (UC) allografts have been shown to mitigate inflammation and decrease scar formation. This has theoretical benefit for recovery and disease progression following dorsal cheilectomy. In the first prospective randomized and blinded cheilectomy trial reported, we aimed to compare outcomes of patients undergoing dorsal cheilectomy alone and dorsal cheilectomy with cryopreserved umbilical cord. Methods: After obtaining institutional board review approval, patients were randomized to cheilectomy alone(CA) or cheilectomy with cryopreserved UC. Surgeries were performed by fellowship trained surgeons. Dorsal cheilectomy was performed utilizing fluoroscopy to remove ˜25% articular surface. UC was applied to cheilectomy site and secured inside capsule with absorbable “stay-stitch.” Patients were followed for 1 year with AOFAS MTP-IP, Foot Function Index (FFI), and VAS-pain (walking, waking, and end of day) outcomes collected preoperatively and at 6 months and 1 year. In addition, radiographic range of motion data was collected (maximal dorsiflexion and plantarflexion). Power analysis determined 27 patients per group was needed to detect a significant difference between AOFAS scores of 95(UC) and 85(control). Data was analyzed utilizing statistical analysis software(SAS v9.4). AOFAS MTP-IP, FFI, and VAS scores were analyzed using Wilcoxon signed-rank test. Range of motion data was analyzed using two-way ANOVA with Tukey adjusted least square means test. Results: 51 patients (26 UC, 25 CA) completed the study. There were 5 bilateral surgeries in UC group and 2 in CA group, totaling 31 and 27 feet respectively. Post-operatively, UC group had significantly improved AOFAS and FFI scores at 1 year compared to CA group. There was no difference between groups for VAS-pain scores (walking, waking, or end of day at any time point), but overall VAS-pain improved in both groups from preoperative values. There was no difference seen in range of motion between groups. However, there was an overall improvement in maximal plantarflexion at 6 months and 1 year and maximal dorsiflexion at 6 months in both groups. Conclusion: We present the results of the first randomized and blinded prospective study of cheilectomy surgery patients. There was improvement in range of motion, pain, AOFAS, and FFI scores in all patients with statistically significant improvement at 1 year in AOFAS and FFI scores in the UC group compared to CA group. When appropriately selected, cheilectomy remains a good option for patients with symptomatic hallux rigidus. Cryopreserved umbilical cord is a potential adjuvant to cheilectomy to modulate inflammation and scarring with early 1 year results showing improvements in functional outcome scores.

Depth of invasion in early oral cancers: Is it an independent prognostic factor?

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. 6058-6058
Author(s):  
Harsh Dhar ◽  
Anil D'Cruz ◽  
Richa Vaish ◽  
Rohini W Hawaldar ◽  
Sudeep Gupta ◽  
...  
Keyword(s):  
Multivariate Analysis ◽  
Prognostic Factors ◽  
Controlled Trial ◽  
Tnm Staging ◽  
Rank Test ◽  
Depth Of Invasion ◽  
Entire Cohort ◽  
Oral Cancers ◽  
T Stage ◽  
Significant Difference

6058 Background: Depth of invasion (DOI) has been incorporated in the new AJCC TNM staging (8th edition) for oral cancers. We hypothesized that the negative effect of increasing DOI on outcomes was a result of an increased propensity to node metastasis and appropriate neck treatment would negate its detrimental effect on outcomes. Methods: Patients with T1/ T2 oral squamous cell carcinoma, clinically node negative, from a previously reported Randomized Controlled Trial (NCT 00193765) formed the cohort for this study. Patients were restaged according to the new staging system . Overall survival(OS) was estimated by the revised T stage for the entire cohort and separately for those who underwent END and those who did not (TND arm) using Kaplan Meier and log rank test . Multivariate analysis was performed using Cox proportional hazard model making adjustment for neck treatment, T stage, site, prognostic factors and the interaction between revised T stage and neck treatment. Results: Of the 596 patients 577 were evaluable, with a median follow up of 77.57 months. Initial pT staging was pT1, 389(67.4%); pT2, 181(31.4%); pT3, 7(1.2%) and was modified to pT1, 195(33.8%); pT2, 280(48.5%); pT3, 102(17.7%) on restaging . 288 patients underwent END and 289 did not (TND arm). For the entire cohort 5-year OS rates were 79.0% [95 %CI, 73.12-84.88] for pT1, 69.4% [95% CI, 63.91-74.89] for pT2 and 53.0% [95% CI, 43.2 -62.8] for pT3 with significant difference between the 3 groups (p < 0.001). In those without upfront neck treatment( TND ), OS difference was maintained between the pT1 and pT2 groups [81.1% (95%CI, 73.26-88.94) vs 65.0% (95%CI, 56.77-73.23)], p = 0.004. This difference was not apparent in the END arm ,pT1 -76.9% (95 %CI, 68.47-85.33) vs pT2 -73.7% (95%CI, 66.25-81.15), p = 0.73. T3 tumours had uniformly poor survival irrespective of neck treatment. On multivariate analysis of the revised pT1/T2 cohort (n = 475), pT stage, neck treatment and grade were independent prognostic factors impacting OS. There was a significant interaction between the T stage and neck treatment (p = 0.03). Conclusions: When DOI < 10 mm, END supplants the prognostic implication of depth with similar outcomes for T1 and T2 tumours (new AJCC staging). The exact role of DOI on outcomes warrants further research. Clinical trial information: NCT00193765.

scholarly journals Pulmonary Collectins Modulate Strain-Specific Influenza A Virus Infection and Host Responses

2004 ◽  
Vol 78 (16) ◽  
pp. 8565-8572 ◽  
Author(s):  
Samuel Hawgood ◽  
Cynthia Brown ◽  
Jess Edmondson ◽  
Amber Stumbaugh ◽  
Lennell Allen ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Host Response ◽  
Influenza A ◽  
Small Difference ◽  
Alveolar Epithelium ◽  
Host Responses ◽  
Cytokine Release ◽  
Significant Difference

ABSTRACT Collectins are secreted collagen-like lectins that bind, agglutinate, and neutralize influenza A virus (IAV) in vitro. Surfactant proteins A and D (SP-A and SP-D) are collectins expressed in the airway and alveolar epithelium and could have a role in the regulation of IAV infection in vivo. Previous studies have shown that binding of SP-D to IAV is dependent on the glycosylation of specific sites on the HA1 domain of hemagglutinin on the surface of IAV, while the binding of SP-A to the HA1 domain is dependent on the glycosylation of the carbohydrate recognition domain of SP-A. Here, using SP-A and SP-D gene-targeted mice on a common C57BL6 background, we report that viral replication and the host response as measured by weight loss, neutrophil influx into the lung, and local cytokine release are regulated by SP-D but not SP-A when the IAV is glycosylated at a specific site (N165) on the HA1 domain. SP-D does not protect against IAV infection with a strain lacking glycosylation at N165. With the exception of a small difference on day 2 after infection with X-79, we did not find any significant difference in viral load in SP-A−/− mice with either IAV strain, although small differences in the cytokine responses to IAV were detected in SP-A−/− mice. Mice deficient in both SP-A and SP-D responded to IAV similarly to mice deficient in SP-D alone. Since most strains of IAV currently circulating are glycosylated at N165, SP-D may play a role in protection from IAV infection.

scholarly journals Codon Deletions in the Influenza A Virus PA Gene Generate Temperature-Sensitive Viruses

2016 ◽  
Vol 90 (7) ◽  
pp. 3684-3693 ◽  
Author(s):  
Léa Meyer ◽  
Alix Sausset ◽  
Laura Sedano ◽  
Bruno Da Costa ◽  
Ronan Le Goffic ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Rna Polymerase ◽  
Influenza A Virus ◽  
Influenza A ◽  
Wild Type Virus ◽  
Deletion Mutants ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Wild Type

ABSTRACTThe influenza virus RNA-dependent RNA polymerase, which is composed of three subunits, PB1, PB2, and PA, catalyzes genome replication and transcription within the cell nucleus. The PA linker (residues 197 to 256) can be altered by nucleotide substitutions to engineer temperature-sensitive (ts), attenuated mutants that display a defect in the transport of the PA–PB1 complex to the nucleus at a restrictive temperature. In this study, we investigated the ability of the PA linker to tolerate deletion mutations for furtherin vitroandin vivocharacterization. Four viable mutants with single-codon deletions were generated; all of them exhibited atsphenotype that was associated with the reduced efficiency of replication/transcription of a pseudoviral reporter RNA in a minireplicon assay. Using fluorescently tagged PB1, we observed that the deletion mutants did not efficiently recruit PB1 to reach the nucleus at a restrictive temperature (39.5°C). Mouse infections showed that the four mutants were attenuated and induced antibodies that were able to protect mice from challenge with a lethal homologous wild-type virus. Serialin vitropassages of two deletion mutants at 39.5°C and 37°C did not allow the restoration of a wild-type phenotype among virus progeny. Thus, our results identify codons that can be deleted in the PA gene to engineer genetically stabletsmutants that could be used to design novel attenuated vaccines.IMPORTANCEIn order to generate genetically stable live influenza A virus vaccines, we constructed viruses with single-codon deletions in a discrete domain of the RNA polymerase PA gene. The four rescued viruses exhibited a temperature-sensitive phenotype that we found was associated with a defect in the transport of the PA–PB1 dimer to the nucleus, where viral replication occurs. Thesetsdeletion mutants were shown to be attenuated and to be able to produce antibodies in mice and to protect them from a lethal challenge. Assays to select revertants that were able to grow efficiently at a restrictive temperature failed, showing that these deletion mutants are genetically more stable than conventional substitution mutants. These results are of interest for the design of genetically stable live influenza virus vaccines.

scholarly journals Abstract P-18: Phosphorylation of RNA Polymerase II C-Terminal Domain Affects Transcript Elongation Through Chromatin

2021 ◽  
Vol 11 (Suppl_1) ◽  
pp. S19-S19
Author(s):  
Sohail Akhtar ◽  
Elena Kotova ◽  
Nadezhda Gerasimova ◽  
Vasily Studitsky
Keyword(s):  
Rna Polymerase ◽  
Chromatin Structure ◽  
Full Length ◽  
Pol Ii ◽  
Terminal Domain ◽  
Significant Difference ◽  
Transcript Elongation ◽  
Hyperphosphorylated Form ◽  
Ctd Phosphorylation

Background: Transcription is the central point of gene regulation where the efficient maintenance of chromatin structure during the passage of RNA polymerase (Pol II) is critical for cell survival and functioning. The phosphorylation of carboxy-terminal domain (CTD) of the large subunit (Rpb1) of Pol II plays a key role in transcription through chromatin providing the binding and dissociation of factors essential for the mRNA biogenesis. Although the regulatory effect of chromatin structure on multiple stages of transcription has been well established, the role of CTD phosphorylation itself has not been systematically addressed. Methods: The effect of differentially phosphorylated Pol II-CTD on transcript elongation through chromatin was studied using in vitro transcription system based on mononucleosomes precisely positioned on DNA. The unphosphorylated and hyperphosphorylated Pol II-CTD were obtained using yeast genetics as well as in vitro kinase or phosphatases. Transcription rate and positions of pausing were measured using authentic elongation complexes comprising Pol II having different CTD phosphorylation states. The quantitative analysis of the transcripts was conducted using denaturing PAGE. Results: We observed a significant difference in the transcription through chromatin depending on CTD phosphorylation level. Thus, experiments on transcription of nucleosomes with Pol II isoforms have shown that the hyperphosphorylated form more efficiently transcribes the nucleosome and leads to a faster accumulation of the full-length RNA product than the non-phosphorylated isoform of Pol II. The non-phosphorylated isoform of the enzyme is characterized by a stronger pause in the early nucleosomal region and a slower accumulation of the full-length RNA product. Conclusion: Hyperphosphorylated form more efficiently transcribes the nucleosome and leads to a faster accumulation of the full-length RNA product as compared with the non-phosphorylated isoform of Pol II. A preliminary model of the effect of Pol II hyperphosphorylation on nucleosomal DNA transcription is proposed.

scholarly journals Enisamium Inhibits SARS-CoV-2 RNA Synthesis

Biomedicines ◽  
2021 ◽  
Vol 9 (9) ◽  
pp. 1254
Author(s):  
Stefano Elli ◽  
Denisa Bojkova ◽  
Marco Bechtel ◽  
Thomas Vial ◽  
David Boltz ◽  
...  
Keyword(s):  
Cell Culture ◽  
Rna Polymerase ◽  
Influenza A ◽  
Rna Synthesis ◽  
Herd Immunity ◽  
Virus Rna ◽  
Independent States ◽  
Dynamics Simulations ◽  
Insight Into

Pandemic SARS-CoV-2 causes a mild to severe respiratory disease called coronavirus disease 2019 (COVID-19). While control of the SARS-CoV-2 spread partly depends on vaccine-induced or naturally acquired protective herd immunity, antiviral strategies are still needed to manage COVID-19. Enisamium is an inhibitor of influenza A and B viruses in cell culture and clinically approved in countries of the Commonwealth of Independent States. In vitro, enisamium acts through metabolite VR17-04 and inhibits the activity of the influenza A virus RNA polymerase. Here we show that enisamium can inhibit coronavirus infections in NHBE and Caco-2 cells, and the activity of the SARS-CoV-2 RNA polymerase in vitro. Docking and molecular dynamics simulations provide insight into the mechanism of action and indicate that enisamium metabolite VR17-04 prevents GTP and UTP incorporation. Overall, these results suggest that enisamium is an inhibitor of SARS-CoV-2 RNA synthesis in vitro.

scholarly journals Effect of dual tocolysis with fenoterol and atosiban in human myometrium

2019 ◽  
Vol 47 (2) ◽  
pp. 190-194 ◽  
Author(s):  
Bernhard Stoiber ◽  
Christian Haslinger ◽  
Marie Kristin Schäffer ◽  
Roland Zimmermann ◽  
Leonhard Schäffer
Keyword(s):  
Area Under The Curve ◽  
Antagonistic Effect ◽  
Additive Effect ◽  
University Hospital ◽  
Rank Test ◽  
Human Myometrium ◽  
Tissue Samples ◽  
Significant Difference ◽  
Signed Rank Test

Abstract Objectives To measure the tocolytic effect of the combination of the oxytocin receptor antagonist atosiban with the β-mimetic agent fenoterol on human myometrium of pregnant women. Methods An in vitro study of contractility in human myometrium at the Laboratory of the Department of Obstetrics, University Hospital of Zürich, Switzerland, was performed. Thirty-six human myometrial biopsies were obtained during elective caesarean sections of singleton pregnancies at term. Tissue samples were exposed to atosiban, fenoterol and the combination of atosiban with fenoterol. Contractility was measured as area under the curve during 30 min of spontaneous contractions. The effect of treatment was expressed as the percentage of change from basal activity during 30 min of exposure. Differences were calculated using a paired Wilcoxon signed-rank test. An additive effect of dual tocolysis was assumed when no significant difference was detected between the observed and expected inhibition of dual tocolysis. When inhibition was greater or lower than expected, the dual combination was characterised as “synergistic” or “antagonistic”, respectively. Results Atosiban and fenoterol alone suppressed contractions by a median of 43.2% and 29.8%, respectively. The combination of atosiban plus fenoterol was measured at a level of 67.3% inhibition. There was no significant difference in the expected (63.2%) and observed inhibition effect of dual tocolysis (P=0.945). Conclusion This study demonstrated an additive effect of dual tocolysis of atosiban and fenoterol on human myometrium in vitro, but no synergistic or antagonistic effect.

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