7 research outputs found
Comprehensive Fragment Screening of the SARS-CoV-2 Proteome Explores Novel Chemical Space for Drug Development
12 pags., 4 figs., 3 tabs.SARS-CoV-2 (SCoV2) and its variants of concern pose serious challenges to the public health. The variants increased challenges to vaccines, thus necessitating for development of new intervention strategies including anti-virals. Within the international Covid19-NMR consortium, we have identified binders targeting the RNA genome of SCoV2. We established protocols for the production and NMR characterization of more than 80 % of all SCoV2 proteins. Here, we performed an NMR screening using a fragment library for binding to 25 SCoV2 proteins and identified hits also against previously unexplored SCoV2 proteins. Computational mapping was used to predict binding sites and identify functional moieties (chemotypes) of the ligands occupying these pockets. Striking consensus was observed between NMR-detected binding sites of the main protease and the computational procedure. Our investigation provides novel structural and chemical space for structure-based drug design against the SCoV2 proteome.Work at BMRZ is supported by the state of Hesse. Work in Covid19-NMR
was supported by the Goethe Corona Funds, by the IWBEFRE-program 20007375 of state of Hesse, the DFG
through CRC902: “Molecular Principles of RNA-based regulation.” and through infrastructure funds (project
numbers: 277478796, 277479031, 392682309, 452632086, 70653611) and by European Union’s Horizon 2020 research and innovation program iNEXT-discovery under grant agreement No 871037. BY-COVID receives funding from the European Union’s Horizon Europe Research and Innovation Programme under grant agreement number 101046203. “INSPIRED” (MIS 5002550) project, implemented under the Action “Reinforcement of the Research and Innovation Infrastructure,” funded by the Operational
Program “Competitiveness, Entrepreneurship and Innovation” (NSRF 2014–2020) and co-financed by Greece and the EU (European Regional Development Fund) and the FP7 REGPOT CT-2011-285950—“SEE-DRUG” project (purchase of UPAT’s 700 MHz NMR equipment). The support of the CERM/CIRMMP center of Instruct-ERIC is gratefully acknowledged. This work has been funded in part by a grant of the Italian Ministry of University and Research (FISR2020IP_02112, ID-COVID) and by Fondazione CR
Firenze. A.S. is supported by the Deutsche Forschungsgemeinschaft [SFB902/B16, SCHL2062/2-1] and the Johanna Quandt Young Academy at Goethe [2019/AS01]. M.H. and C.F. thank SFB902 and the Stiftung Polytechnische Gesellschaft for the Scholarship. L.L. work was supported by the French National Research Agency (ANR, NMR-SCoV2-ORF8), the Fondation de la Recherche Médicale (FRM, NMR-SCoV2-ORF8), FINOVI and the IR-RMN-THC Fr3050 CNRS. Work at UConn Health was supported by grants from the US National Institutes of Health (R01 GM135592 to B.H., P41 GM111135 and R01 GM123249 to J.C.H.) and the US National Science Foundation (DBI 2030601 to J.C.H.). Latvian Council of Science Grant No. VPP-COVID-2020/1-0014. National Science Foundation EAGER MCB-2031269. This work was supported by the grant Krebsliga KFS-4903-08-2019 and SNF-311030_192646 to J.O. P.G. (ITMP) The EOSC Future project is co-funded by the European Union Horizon Programme call INFRAEOSC-03-2020—Grant Agreement
Number 101017536. Open Access funding enabled and organized by Projekt DEALPeer reviewe
Large-Scale Recombinant Production of the SARS-CoV-2 Proteome for High-Throughput and Structural Biology Applications
The highly infectious disease COVID-19 caused by the Betacoronavirus SARS-CoV-2 poses a severe threat to humanity and demands the redirection of scientific efforts and criteria to organized research projects. The international COVID19-NMR consortium seeks to provide such new approaches by gathering scientific expertise worldwide. In particular, making available viral proteins and RNAs will pave the way to understanding the SARS-CoV-2 molecular components in detail. The research in COVID19-NMR and the resources provided through the consortium are fully disclosed to accelerate access and exploitation. NMR investigations of the viral molecular components are designated to provide the essential basis for further work, including macromolecular interaction studies and high-throughput drug screening. Here, we present the extensive catalog of a holistic SARS-CoV-2 protein preparation approach based on the consortium’s collective efforts. We provide protocols for the large-scale production of more than 80% of all SARS-CoV-2 proteins or essential parts of them. Several of the proteins were produced in more than one laboratory, demonstrating the high interoperability between NMR groups worldwide. For the majority of proteins, we can produce isotope-labeled samples of HSQC-grade. Together with several NMR chemical shift assignments made publicly available on covid19-nmr.com, we here provide highly valuable resources for the production of SARS-CoV-2 proteins in isotope-labeled form
Identification of Natural Products Inhibiting SARS-CoV-2 by Targeting Viral Proteases: A Combined in Silico and in Vitro Approach
In this study, an integrated in silico-in vitro approach was employed to discover natural products (NPs) active against SARS-CoV-2. The two SARS-CoV-2 viral proteases, i.e., main protease (Mpro) and papain-like protease (PLpro), were selected as targets for the in silico study. Virtual hits were obtained by docking more than 140,000 NPs and NP derivatives available in-house and from commercial sources, and 38 virtual hits were experimentally validated in vitro using two enzyme-based assays. Five inhibited the enzyme activity of SARS-CoV-2 Mpro by more than 60% at a concentration of 20 mu M, and four of them with high potency (IC50 < 10 mu M). These hit compounds were further evaluated for their antiviral activity against SARS-CoV-2 in Calu-3 cells. The results from the cell-based assay revealed three mulberry Diels-Alder-type adducts (MDAAs) from Morus alba with pronounced anti-SARS-CoV-2 activities. Sanggenons C (12), O (13), and G (15) showed IC50 values of 4.6, 8.0, and 7.6 mu M and selectivity index values of 5.1, 3.1 and 6.5, respectively. The docking poses of MDAAs in SARS-CoV-2 Mpro proposed a butterfly-shaped binding conformation, which was supported by the results of saturation transfer difference NMR experiments and competitive 1H relaxation dispersion NMR spectroscopy.ISSN:0163-3864ISSN:1520-602
Targeting the main protease (Mpro, nsp5) by growth of fragment scaffolds exploiting structure-based methodologies
The main protease Mpro, nsp5, of SARS-CoV-2 (SCoV2) is one of its most attractive drug targets. Here, we report primary screening data using nuclear magnetic resonance spectroscopy (NMR) of four different libraries and detailed follow-up synthesis on the promising uracil-containing fragment Z604 derived from these libraries. Z604 shows time-dependent binding. Its inhibitory effect is sensitive to reducing conditions. Starting with Z604, we synthesized and characterized 13 compounds designed by fragment growth strategies. Each compound was characterized by NMR and/or activity assays to investigate their interaction with Mpro. These investigations resulted in the four-armed compound 35b that binds directly to Mpro. 35b could be cocrystallized with Mpro revealing its noncovalent binding mode, which fills all four active site subpockets. Herein, we describe the NMR-derived fragment-to-hit pipeline and its application for the development of promising starting points for inhibitors of the main protease of SCoV2
Comprehensive Fragment Screening of the SARS‐CoV‐2 Proteome Explores Novel Chemical Space for Drug Development
SARS‐CoV‐2 (SCoV2) and its variants of concern pose serious challenges to the public health. The variants increased challenges to vaccines, thus necessitating for development of new intervention strategies including anti‐virals. Within the international Covid19‐NMR consortium, we have identified binders targeting the RNA genome of SCoV2. We established protocols for the production and NMR characterization of more than 80 % of all SCoV2 proteins. Here, we performed an NMR screening using a fragment library for binding to 25 SCoV2 proteins and identified hits also against previously unexplored SCoV2 proteins. Computational mapping was used to predict binding sites and identify functional moieties (chemotypes) of the ligands occupying these pockets. Striking consensus was observed between NMR‐detected binding sites of the main protease and the computational procedure. Our investigation provides novel structural and chemical space for structure‐based drug design against the SCoV2 proteome
Large-Scale Recombinant Production of the SARS-CoV-2 Proteome for High-Throughput and Structural Biology Applications
The highly infectious disease COVID-19 caused by the Betacoronavirus SARS-CoV-2 poses a severe threat to humanity and demands the redirection of scientific efforts and criteria to organized research projects. The international COVID19-NMR consortium seeks to provide such new approaches by gathering scientific expertise worldwide. In particular, making available viral proteins and RNAs will pave the way to understanding the SARS-CoV-2 molecular components in detail. The research in COVID19-NMR and the resources provided through the consortium are fully disclosed to accelerate access and exploitation. NMR investigations of the viral molecular components are designated to provide the essential basis for further work, including macromolecular interaction studies and high-throughput drug screening. Here, we present the extensive catalog of a holistic SARS-CoV-2 protein preparation approach based on the consortium’s collective efforts. We provide protocols for the large-scale production of more than 80% of all SARS-CoV-2 proteins or essential parts of them. Several of the proteins were produced in more than one laboratory, demonstrating the high interoperability between NMR groups worldwide. For the majority of proteins, we can produce isotope-labeled samples of HSQC-grade. Together with several NMR chemical shift assignments made publicly available on covid19-nmr.com, we here provide highly valuable resources for the production of SARS-CoV-2 proteins in isotope-labeled form.This work was supported by Goethe University (Corona funds),
the DFG-funded CRC: “Molecular Principles of RNA-Based
Regulation,” DFG infrastructure funds (project numbers:
277478796, 277479031, 392682309, 452632086, 70653611), the
state of Hesse (BMRZ), the Fondazione CR Firenze (CERM),
and the IWB-EFRE-program 20007375. This project has
received funding from the European Union’s Horizon 2020
research and innovation program under Grant Agreement No.
871037. AS is supported by DFG Grant SCHL 2062/2-1 and by the
JQYA at Goethe through project number 2019/AS01. Work in the
lab of KV was supported by a CoRE grant from the University of
New Hampshire. The FLI is a member of the Leibniz Association
(WGL) and financially supported by the Federal Government of
Germany and the State of Thuringia. Work in the lab of RM was
supported by NIH (2R01EY021514) and NSF (DMR-2002837).
BN-B was supported by theNSF GRFP.MCwas supported byNIH
(R25 GM055246 MBRS IMSD), and MS-P was supported by the
HHMI Gilliam Fellowship. Work in the labs of KJ and KT was
supported by Latvian Council of Science Grant No. VPP-COVID
2020/1-0014. Work in the UPAT’s lab was supported by the
INSPIRED (MIS 5002550) project, which is implemented under
the Action “Reinforcement of the Research and Innovation
Infrastructure,” funded by the Operational Program
“Competitiveness, Entrepreneurship and Innovation” (NSRF
2014–2020) and cofinanced by Greece and the EU (European
Regional Development Fund) and the FP7 REGPOT CT-2011-
285950–“SEE-DRUG” project (purchase of UPAT’s 700MHz
NMR equipment). Work in the CM-G lab was supported by
the Helmholtz society. Work in the lab of ABö was supported
by the CNRS, the French National Research Agency (ANR, NMRSCoV2-
ORF8), the Fondation de la Recherche Médicale (FRM,
NMR-SCoV2-ORF8), and the IR-RMN-THC Fr3050 CNRS.
Work in the lab of BM was supported by the Swiss National
Science Foundation (Grant number 200020_188711), the
Günthard Stiftung für Physikalische Chemie, and the ETH
Zurich. Work in the labs of ABö and BM was supported by a
common grant from SNF (grant 31CA30_196256). This work was
supported by the ETHZurich, the grant ETH40 18 1, and the grant
Krebsliga KFS 4903 08 2019. Work in the lab of the IBS Grenoble
was supported by the Agence Nationale de Recherche (France)
RA-COVID SARS2NUCLEOPROTEIN and European Research
Council Advanced Grant DynamicAssemblies. Work in the
CA lab was supported by Patto per il Sud della Regione
Siciliana–CheMISt grant (CUP G77B17000110001). Part of
this work used the platforms of the Grenoble Instruct-ERIC
center (ISBG; UMS 3518 CNRS-CEA-UGA-EMBL) within the
Grenoble Partnership for Structural Biology (PSB), supported
by FRISBI (ANR-10-INBS-05-02) and GRAL, financed within
the University Grenoble Alpes graduate school (Ecoles
Universitaires de Recherche) CBH-EUR-GS (ANR-17-EURE-
0003). Work at the UW-Madison was supported by grant
numbers NSF MCB2031269 and NIH/NIAID AI123498. MM
is a Ramón y Cajal Fellow of the Spanish AEI-Ministry of
Science and Innovation (RYC2019-026574-I), and a “La
Caixa” Foundation (ID 100010434) Junior Leader Fellow
(LCR/BQ/PR19/11700003). Funded by project COV20/00764
fromthe Carlos III Institute of Health and the SpanishMinistry
of Science and Innovation to MMand DVL. VDJ was supported
by the Boehringer Ingelheim Fonds. Part of this work used the
resources of the Italian Center of Instruct-ERIC at the CERM/
CIRMMP infrastructure, supported by the Italian Ministry for
University and Research (FOE funding). CF was supported by
the Stiftung Polytechnische Gesellschaft. Work in the lab of
JH was supported by NSF (RAPID 2030601) and NIH
(R01GM123249).Peer reviewe
ISARIC-COVID-19 dataset: A Prospective, Standardized, Global Dataset of Patients Hospitalized with COVID-19
The International Severe Acute Respiratory and Emerging Infection Consortium (ISARIC) COVID-19 dataset is one of the largest international databases of prospectively collected clinical data on people hospitalized with COVID-19. This dataset was compiled during the COVID-19 pandemic by a network of hospitals that collect data using the ISARIC-World Health Organization Clinical Characterization Protocol and data tools. The database includes data from more than 705,000 patients, collected in more than 60 countries and 1,500 centres worldwide. Patient data are available from acute hospital admissions with COVID-19 and outpatient follow-ups. The data include signs and symptoms, pre-existing comorbidities, vital signs, chronic and acute treatments, complications, dates of hospitalization and discharge, mortality, viral strains, vaccination status, and other data. Here, we present the dataset characteristics, explain its architecture and how to gain access, and provide tools to facilitate its use