Effect of Neutralizing Monoclonal Antibody Treatment on Early Trajectories of Virologic and Immunologic Biomarkers in Patients Hospitalized With COVID-19

BACKGROUND: Neutralizing monoclonal antibodies (nmAbs) failed to show clear benefit for hospitalized patients with coronavirus disease 2019 (COVID-19). Dynamics of virologic and immunologic biomarkers remain poorly understood. METHODS: Participants enrolled in the Therapeutics for Inpatients with COVID-19 trials were randomized to nmAb versus placebo. Longitudinal differences between treatment and placebo groups in levels of plasma nucleocapsid antigen (N-Ag), anti-nucleocapsid antibody, C-reactive protein, interleukin-6, and D-dimer at enrollment, day 1, 3, and 5 were estimated using linear mixed models. A 7-point pulmonary ordinal scale assessed at day 5 was compared using proportional odds models. RESULTS: Analysis included 2149 participants enrolled between August 2020 and September 2021. Treatment resulted in 20% lower levels of plasma N-Ag compared with placebo (95% confidence interval, 12%-27%; P \u3c .001), and a steeper rate of decline through the first 5 days (P \u3c .001). The treatment difference did not vary between subgroups, and no difference was observed in trajectories of other biomarkers or the day 5 pulmonary ordinal scale. CONCLUSIONS: Our study suggests that nmAb has an antiviral effect assessed by plasma N-Ag among hospitalized patients with COVID-19, with no blunting of the endogenous anti-nucleocapsid antibody response. No effect on systemic inflammation or day 5 clinical status was observed. CLINICAL TRIALS REGISTRATION: NCT04501978

Structural Insights into the Inhibition of Cytosolic 5′-Nucleotidase II (cN-II) by Ribonucleoside 5′-Monophosphate Analogues

Cytosolic 5′-nucleotidase II (cN-II) regulates the intracellular nucleotide pools within the cell by catalyzing the dephosphorylation of 6-hydroxypurine nucleoside 5′-monophosphates. Beside this physiological function, high level of cN-II expression is correlated with abnormal patient outcome when treated with cytotoxic nucleoside analogues. To identify its specific role in the resistance phenomenon observed during cancer therapy, we screened a particular class of chemical compounds, namely ribonucleoside phosphonates to predict them as potential cN-II inhibitors. These compounds incorporate a chemically and enzymatically stable phosphorus-carbon linkage instead of a regular phosphoester bond. Amongst them, six compounds were predicted as better ligands than the natural substrate of cN-II, inosine 5′-monophosphate (IMP). The study of purine and pyrimidine containing analogues and the introduction of chemical modifications within the phosphonate chain has allowed us to define general rules governing the theoretical affinity of such ligands. The binding strength of these compounds was scrutinized in silico and explained by an impressive number of van der Waals contacts, highlighting the decisive role of three cN-II residues that are Phe 157, His 209 and Tyr 210. Docking predictions were confirmed by experimental measurements of the nucleotidase activity in the presence of the three best available phosphonate analogues. These compounds were shown to induce a total inhibition of the cN-II activity at 2 mM. Altogether, this study emphasizes the importance of the non-hydrolysable phosphonate bond in the design of new competitive cN-II inhibitors and the crucial hydrophobic stacking promoted by three protein residues

Fine Tuning of Globin Gene Expression by DNA Methylation

Expression patterns in the globin gene cluster are subject to developmental regulation in vivo. While the γ(A) and γ(G) genes are expressed in fetal liver, both are silenced in adult erythrocytes. In order to decipher the role of DNA methylation in this process, we generated a YAC transgenic mouse system that allowed us to control γ(A) methylation during development. DNA methylation causes a 20-fold repression of γ(A) both in non-erythroid and adult erythroid cells. In erythroid cells this modification works as a dominant mechanism to repress γ gene expression, probably through changes in histone acetylation that prevent the binding of erythroid transcription factors to the promoter. These studies demonstrate that DNA methylation serves as an elegant in vivo fine-tuning device for selecting appropriate genes in the globin locus. In addition, our findings provide a mechanism for understanding the high levels of γ-globin transcription seen in patients with Hereditary Persistence of Fetal Hemoglobin, and help explain why 5azaC and butyrate compounds stimulate γ-globin expression in patients with β-hemoglobinopathies

Hyperimmune immunoglobulin for hospitalised patients with COVID-19 (ITAC): a double-blind, placebo-controlled, phase 3, randomised trial

BACKGROUND: Passive immunotherapy using hyperimmune intravenous immunoglobulin (hIVIG) to SARS-CoV-2, derived from recovered donors, is a potential rapidly available, specific therapy for an outbreak infection such as SARS-CoV-2. Findings from randomised clinical trials of hIVIG for the treatment of COVID-19 are limited. METHODS: In this international randomised, double-blind, placebo-controlled trial, hospitalised patients with COVID-19 who had been symptomatic for up to 12 days and did not have acute end-organ failure were randomly assigned (1:1) to receive either hIVIG or an equivalent volume of saline as placebo, in addition to remdesivir, when not contraindicated, and other standard clinical care. Randomisation was stratified by site pharmacy; schedules were prepared using a mass-weighted urn design. Infusions were prepared and masked by trial pharmacists; all other investigators, research staff, and trial participants were masked to group allocation. Follow-up was for 28 days. The primary outcome was measured at day 7 by a seven-category ordinal endpoint that considered pulmonary status and extrapulmonary complications and ranged from no limiting symptoms to death. Deaths and adverse events, including organ failure and serious infections, were used to define composite safety outcomes at days 7 and 28. Prespecified subgroup analyses were carried out for efficacy and safety outcomes by duration of symptoms, the presence of anti-spike neutralising antibodies, and other baseline factors. Analyses were done on a modified intention-to-treat (mITT) population, which included all randomly assigned participants who met eligibility criteria and received all or part of the assigned study product infusion. This study is registered with ClinicalTrials.gov, NCT04546581. FINDINGS: From Oct 8, 2020, to Feb 10, 2021, 593 participants (n=301 hIVIG, n=292 placebo) were enrolled at 63 sites in 11 countries; 579 patients were included in the mITT analysis. Compared with placebo, the hIVIG group did not have significantly greater odds of a more favourable outcome at day 7; the adjusted OR was 1·06 (95% CI 0·77–1·45; p=0·72). Infusions were well tolerated, although infusion reactions were more common in the hIVIG group (18·6% vs 9·5% for placebo; p=0·002). The percentage with the composite safety outcome at day 7 was similar for the hIVIG (24%) and placebo groups (25%; OR 0·98, 95% CI 0·66–1·46; p=0·91). The ORs for the day 7 ordinal outcome did not vary for subgroups considered, but there was evidence of heterogeneity of the treatment effect for the day 7 composite safety outcome: risk was greater for hIVIG compared with placebo for patients who were antibody positive (OR 2·21, 95% CI 1·14–4·29); for patients who were antibody negative, the OR was 0·51 (0·29–0·90; pinteraction=0·001). INTERPRETATION: When administered with standard of care including remdesivir, SARS-CoV-2 hIVIG did not demonstrate efficacy among patients hospitalised with COVID-19 without end-organ failure. The safety of hIVIG might vary by the presence of endogenous neutralising antibodies at entry. FUNDING: US National Institutes of Health

Interaction of human 3-phosphoglycerate kinase with Its two substrates : Is substrate antagonism a kinetic advantage ?

International audienceSubstrate antagonism has been described for a variety of enzymes with more than one substrate and is characterized by a lowering of the affinity of one substrate in the presence of the other(s). 3-Phosphoglycerate kinase (PGK) catalyzes phosphotransfer from 1,3-bisphosphoglycerate (bPG) to ADP to give 3-phosphoglycerate (PG) and ATP, and is subject to substrate antagonism. Because of the instability of bPG, antagonism has only been described between PG and ATP or ADP. Here, we show that antagonism also occurs between bPG and ADP. Using the stopped-flow method, we show that the dissociation constant for one substrate increases in the presence of the other, and that this decrease in affinity is mainly due to an increase in the dissociation rate constant. As a consequence, there is an increase in the overall interaction kinetics. Interestingly, in the presence of the mirror image of natural D-ADP, L-ADP (a good substrate for PGK), antagonism is absent. Using rapid-quench-flow, we studied the kinetics of ATP formation. The time courses present the following: (1) a lag with L-ADP, but not with D-ADP, the kinetics of which were similar to the interaction kinetics measured by stopped-flow; (2) a burst that is directed by the phosphotransfer; and (3) a steady-state that is rate limited by the release of product kinetics. Structural explanations for these results are proposed by analyzing the crystallographic structure of the fully closed conformation of PGK in complex with L-ADP, PG, and the transition-state analogue AlF4 − compared to previously determined structures

Phyto-extraction des terres rares à partir de résidus de bauxite modifiés

International audienc

Phyto-extraction des terres rares à partir de résidus de bauxite modifiés

International audienc

Phyto-extraction des terres rares à partir de résidus de bauxite modifiés

International audienc

Colocalization analysis to understand Yttrium uptake in Saxifraga paniculata using complementary imaging technics

International audienceOver the last decades, yttrium (Y) has gained importance inhigh tech applications. Due to its chemical similarities with thelanthanides, Y is often considered a rare earth element (REE).Despite their increased usage, the environmental behaviour ofREEs remains poorly understood. Especially regarding theirinteractions with plants many uncertainties exist as both, positiveand negative effects on plant development have been observed[1]. In order to understand these phenomena a precise knowledgeis necessary about how Y is absorbed by the plant and how it ishandled once inside the organism. Contradictory studies exist,stating that due to similar ionic radius, Y and the other REEsmight be absorbed through Ca2+-channels while others suspect ashared pathway with Al3+ [2].In this study, we used laser ablation coupled ICP-MS andsynchrotron-based micro-X-ray fluorescence spectroscopy(μXRF, beamline Nanoscopium, SOLEIL, France) to localise Ywithin the plant tissue and identify colocalized elements. Theplant used in this study is Saxifraga paniculata, a rugged alpineplant that has shown an affinity for Y in a previous study (inprep.). The results show that after growing on a Y-doped soil(500mg/kg), Y is mainly concentrated in the roots of Saxifragapaniculata and only a small amount is translocated to the aerialparts.μXRF analysis indicates that within the roots the majority of Yremains in the outer cortex and epidermis and hardly penetratesthe stele. Laser ablation coupled ICP-MS confirms this findingand shows a colocalization between Y, Fe and Al and to a lesserextent Ca. In the stem and the leaves Ca disappears from thisgroup of correlated elements while especially in Y-hotspots, Feand Al remain strongly associated. Accordingly, a relationbetween Ca and Y during root uptake remains possible whereasthe correlation to Fe and Al appears to be dominant in the aerialparts, indicating the formation of complexes or a shared pathwayduring translocation

Direct kinetic evidence that lysine 215 is involved in the phospho-transfer step of human 3-phosphoglycerate kinase.

International audience3-Phosphoglycerate kinase (PGK) is a promising candidate for the activation of nucleotide analogues used in antiviral and anticancer therapies. PGK is a key enzyme in glycolysis; it catalyzes the reversible reaction 1,3-bisphosphoglycerate + ADP 3-phosphoglycerate + ATP. Here we explored the catalytic role in human PGK of the highly conserved Lys 215 that has been proposed to be essential for PGK function by a transient and equilibrium kinetic study with the active site mutant K215A. By the stopped-flow method we show that the kinetics of substrate binding and the associated protein isomerization steps are fast and identical for the wild-type PGK and mutant K215A. By the use of a chemical sampling method (rapid quench flow) under multiple and single turnover conditions and in both directions of the reaction, we show that the rate-limiting step with wild-type PGK follows product formation (presumably product release), whereas with the mutant it is the phospho-transfer step itself that is rate-limiting. Mutant K215A has a low inherent phosphotransferase activity, and to explain this, we carried out a molecular modeling study. This suggests that with the mutant the conserved Arg 65 replaces the missing Lys 215 by helping to position the transferable phospho group during the reaction. Molecular dynamics simulations suggest that in the mutant the closed conformation of the enzyme is stabilized by a salt bridge between Asp 218 and Arg 170 rather than Arg 65 in the wild-type PGK