1,722,730 research outputs found
Molecular Determinants of GS-9620-Dependent TLR7 Activation.
No full textGS-9620 is an orally administered agonist of Toll-like receptor (TLR)7 currently being evaluated in clinical studies for the treatment of chronic HBV and HIV patients. GS-9620 has shown antiviral efficacy in preclinical models of chronic hepadnavirus infection in woodchuck as well as chimpanzee. However, the molecular determinants of GS-9620-dependent activation of TLR7 are not well defined. The studies presented here elucidate GS-9620 subcellular distribution and characterize its molecular interactions with human TLR7 using structure-guided mutational analysis. Based on our results we present a molecular model of TLR7 bound to GS-9620. We also determine that several coding SNPs had no effect on GS-9620-dependent TLR7 activation. In addition, our studies provide evidence that TLR7 exists in a ligand-independent oligomeric state and that, TLR7 activation by GS-9620 is likely associated with compound-induced conformational changes. Finally, we demonstrate that activation of NF-κB and Akt pathways in primary plasmacytoid dendritic cells occur as immediate downstream cellular responses to GS-9620 stimulation. The data presented here further our understanding of the molecular parameters governing TLR7 activation by GS-9620, and more generally by nucleos/tide-related ligands
Biological sex and age influence GS-9620 activity ex vivo
No full textToll-like receptors (TLRs) are being explored to enhance immunity in HIV cure strategies. The TLR7 agonist GS-9620 promotes immune activation, reactivates latent HIV, and delays viral rebound in some people with HIV. Previous work has shown that biological sex influences TLR7 signaling. This study examined the interplay between biological sex, age, and the sex hormones 17β-estradiol, progesterone, and testosterone on GS-9620\u27s ability to promote cytokine secretion and activate CD4, CD8, and NK cells ex vivo. Interestingly, sex hormones had no effect on GS-9620-mediated immune activation or cytokine induction. However, we found that GS-9620 activity was influenced by age only in female donors. Further, we found that GS-9620-mediated CD4 T cell activation was positively correlated with the induction of IFN-γ and IL-12, while CD4 T cell activation and IL-12 production were negatively correlated with age. Additionally, CD8 T cell activation was positively correlated with IFN-γ production. Mechanistically, IFN-γ was sufficient to promote higher immune activation of both CD4 and CD8 T cells in female versus male donors. In conclusion, biological sex and age, but not sex hormones, influence GS-9620-mediated immune activation. Understanding these factors will help design and evaluate future clinical trials using GS-9620 for an HIV cure
GS-9620 rapidly distributes to and signals through the endo-lysosomal compartments.
No full text<p>(a-b) Chemical structure of (a) GS-9620 and (b) Resiquimod. (c) Kinetics of intracellular accumulation of <sup>3</sup>H-GS-9620 in Daudi cells. (d) Sub-cellular distribution of <sup>3</sup>H-GS-9620 in relation to major organelle markers obtained by isopycnic density gradient centrifugation. (e) IFN-α2 secretion by enriched pDCs upon stimulation with GS-9620 or resiquimod (control) in the presence of bafilomycin A1 (BAF) or PBS (mock). Data shown in (c), (d), (e) are representative of 3 independent experiments. (e) Error bars represent mean of triplicate assay conditions and SEM. Abbreviations: Early endosome (Early Endo), endoplasmic reticulum (ER), mitochondria (Mt), lysosome (Lyso).</p
GS-9620 induces phosphorylation of NF-κB and Akt in pDCs.
No full text<p>PBMC isolated from healthy donors were cultured with GS-9620, resiquimod, or DMSO control. Phosphorylation of NF-κB and Akt was assessed in gated pDC and mDC subsets. (a) Flow cytometric histogram of p-NF-κB (using a phosho NF-κB (S529) specific antibody) following 30min stimulation (left panel) or p-Akt (using a phospho Akt (S473) specific antibody) following 60min stimulation (right panel) with 1μM GS-9620 (blue histogram) or DMSO control (grey histogram). (b) Fold increase in mean fluorescence intensity (MFI) of pDCs for pNF-κB (left panel) or p-Akt (right panel) after stimulation with 1μM GS-9620 normalized to DMSO control treatment. Statistically significant differences relative to DMSO control (p<0.05) are observed with GS-9620 stimulation for all assessed time points (p-NF-κB), and for 10, 15, 30, and 60min time points (p-Akt). (c) Fold increase in MFI upon stimulation with 1μM GS-9620 or resiquimod (R848) normalized to DMSO control treatment in mDCs or pDCs. As expected, phospho-responses to GS-9620 and R848 for both readouts, p-NF-κB and p-Akt, were significantly stronger in pDC compared to donor-matched mDC (p<0.01 for all comparisons). Graphs show data obtained from 6 (p-NF-κB) and 4 (p-Akt) independent healthy donors with mean ±SEM (bars).</p
Toll-like receptor 7 agonist GS-9620 induces prolonged inhibition of HBV via a type I interferon-dependent mechanism
No full textInternational audienceBackground & Aims: GS-9620, an oral agonist of toll-like receptor 7 (TLR7), is in clinical development for the treatment of chronic hepatitis B (CHB). GS-9620 was previously shown to induce prolonged suppression of serum viral DNA and antigens in the woodchuck and chimpanzee models of CHB. Herein, we investigated the molecular mechanisms that contribute to the antiviral response to GS-9620 using in vitro models of hepatitis B virus (HBV) infection.Methods: Cryopreserved primary human hepatocytes (PHH) and differentiated HepaRG (dHepaRG) cells were infected with HBV and treated with GS-9620, conditioned media from human peripheral blood mononuclear cells treated with GS-9620 (GS-9620 conditioned media [GS-9620-CM]), or other innate immune stimuli. The antiviral and transcriptional response to these agents was determined.Results: GS-9620 had no antiviral activity in HBV-infected PHH, consistent with low level TLR7 mRNA expression in human hepatocytes. In contrast, GS-9620-CM induced prolonged reduction of HBV DNA, RNA, and antigen levels in PHH and dHepaRG cells via a type I interferon (IFN)-dependent mechanism. GS-9620-CM did not reduce covalently closed circular DNA (cccDNA) levels in either cell type. Transcriptional profiling demonstrated that GS-9620-CM strongly induced various HBV restriction factors – although not APOBEC3A or the Smc5/6 complex – and indicated that established HBV infection does not modulate innate immune sensing or signaling in cryopreserved PHH. GS-9620-CM also induced expression of immunoproteasome subunits and enhanced presentation of an immunodominant viral peptide in HBV-infected PHH.Conclusions: Type I IFN induced by GS-9620 durably suppressed HBV in human hepatocytes without reducing cccDNA levels. Moreover, HBV antigen presentation was enhanced, suggesting additional components of the TLR7-induced immune response played a role in the antiviral response to GS-9620 in animal models of CHB
Linked collectors and determiners for: Description of a new species of the fish genus Acanthoplesiops Regan (Teleostei: Plesiopidae: Acanthoclininae) from Tonga..
No full textNatural history specimen data linked to collectors and determiners held within, "Description of a new species of the fish genus Acanthoplesiops Regan (Teleostei: Plesiopidae: Acanthoclininae) from Tonga.". Claims or attributions were made on Bionomia by volunteer Scribes, <a href="http://bionomia.net/dataset/a7d8b32e-84ec-457d-9620-ce9faddd44e3">https://bionomia.net/dataset/a7d8b32e-84ec-457d-9620-ce9faddd44e3</a> using specimen data from the dataset aggregated by the Global Biodiversity Information Facility, <a href="https://gbif.org/dataset/a7d8b32e-84ec-457d-9620-ce9faddd44e3">https://gbif.org/dataset/a7d8b32e-84ec-457d-9620-ce9faddd44e3</a>. Formatted as a Frictionless Data package
Optimization of Cellulase (E.C. 3.2.1: 4) Production Using Penicillium citrinum MTCC 9620 in Solid State Fermentation
No full textAim: The objective of the present work was to optimize the environmental parameters for Cellulase (1,4 β-endoglucanase, E.C.3.2.1:4) production using Penicillium citrinum MTCC 9620 in Solid State Fermentation.
Study Design: One unit of Cellulase (1,4 β-endoglucanase, E.C.3.2.1:4) activity is defined as the amount of enzyme producing 1μmole of glucose equivalent/min measured using UV visible spectrophotometer at 540 nm.
Place and Duration of Study: Food Technology laboratory of Dr. S. S. Bhatnagar University Institute of Chemical Engineering & Technology, Panjab University, Chandigarh between January and June 2011.
Methodology: Penicillium citrinum MTCC 9620 was maintained on potato dextrose agar (PDA) at 4°C. For Cellulase production Czapek Yeast Extract medium was used as moistening medium. Incubation temperature, pH, incubation time and other parameters like suitable substrate, pre-treatment of the substrate on production of Cellulase in Solid State Fermentation (SSF) was optimized using agricultural residues by Penicillium citrinum MTCC 9620. Microscopic and Spectral properties of substrates were determined to detect the structural changes after pre-treatment.
Results: Production of extracellular Cellulase was greatly affected by variation in substrates, pre-treatments of substrate and variation in pH and incubation temperature. Cellulase activity was significantly (p < 0.05) higher when alkali treated wheat bran was used as substrate than untreated substrate. Among three substrates and their three pre-treatment conditions, It has been observed that alkali treated wheat bran was the most suitable substrate for maximum cellulase production (12.56± 0.097U/mL) at pH 5.5 and 30ºC without any extraneous nitrogen source by Penicillium citrinum MTCC 9620 after 120 h of fermentation time. SEM study revealed that during alkali treatment the solid surface become rough which results growth of fungus eventually maximum cellulase production.
Conclusion: P. citrinum MTCC 9620 is one of the potential cellulase producing fungal strain. Optimum condition of cellulase (1,4 β-endoglucanase, E.C.3.2.1:4) production by P. citrinum MTCC 9620 was 30°C temperature, 5.5 pH when alkali treated wheat bran was used as substrate. Growth kinetics of P. citrinum MTCC 9620 was studied and it showed adequacy of fit to Monod Model to describe the growth pattern of P. citrinum MTCC 9620 in SSF at 30°C for 120 h incubation period
TLR7 Agonist GS–9620 Combined with Nicotinamide Generate Viral Reactivation in Seronegative SHIVSF162P3-Infected Rhesus Monkeys
No full textAntiretroviral therapy is capable of inhibiting HIV replication, but it fails to completely achieve a cure due to HIV persistence. The commonly used HIV cure approach is the “shock and kill” strategy, which employs latency-reversing agents to trigger viral reactivation and boost cellular immunity. Finding the appropriate drug combination for the “shock and kill” strategy would greatly facilitate clinical trials. The toll-like receptor (TLR) 7 agonist GS–9620 and nicotinamide (NAM) are reported as potential latency-reversing agents. Herein, we found the absence of viral reactivation when SHIVSF162P3-aviremic rhesus macaques were treated with GS–9620 monotherapy. However, our findings demonstrate that viral blips emerged in half of the macaques treated with the combination therapy of GS–9620 and NAM. Notably, an increase in the reactivation of the replication-competent latent virus was measured in monkeys treated with the combination therapy. These findings suggest that the GS–9620 and NAM combination could be used as a multipronged HIV latency stimulation approach, with potential for optimizing antiviral therapy design
Sustained efficacy and seroconversion with the Toll-like receptor 7 agonist GS-9620 in the Woodchuck model of chronic hepatitis B
No full textBackground & AimsNew therapies for chronic hepatitis B (CHB) are urgently needed since current treatments rarely lead to cure. We evaluated whether the oral small molecule toll-like receptor (TLR7) agonist GS-9620 could induce durable antiviral efficacy in woodchucks chronically infected with woodchuck hepatitis virus (WHV), a hepadnavirus closely related to human hepatitis B virus (HBV).MethodsAfter evaluating the pharmacokinetics, pharmacodynamics and tolerability of oral GS-9620 in uninfected woodchucks, adult woodchucks chronically infected with WHV (n=7 per group) were dosed with GS-9620 or placebo for 4 or 8weeks with different treatment schedules.ResultsGS-9620 treatment induced rapid, marked and sustained reduction in serum viral DNA (mean maximal 6.2log10 reduction), and hepatic WHV DNA replicative intermediates, WHV cccDNA and WHV RNA, as well as loss of detectable serum WHV surface antigen (WHsAg). GS-9620 treatment also induced a sustained antibody response against WHsAg in a subset of animals. Strikingly, treatment reduced the incidence of hepatocellular carcinoma (HCC) from 71% in the placebo group to 8% in GS-9620-treated woodchucks with sustained viral load reduction. GS-9620 treatment was associated with reversible increases in serum liver enzymes and thrombocytopenia, and induced intrahepatic CD8+ T cell, NK cell, B cell and interferon response transcriptional signatures.ConclusionsThe data demonstrate that short duration, finite treatment with the oral TLR7 agonist GS-9620 can induce a sustained antiviral response in the woodchuck model of CHB, and support investigation of this compound as a therapeutic approach to attain a functional cure in CHB patients
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