scholarly journals Virtual screening identifies novel high-affinity σ1 receptor ligands

2019 ◽  
Author(s):  
Daniel A. Greenfield ◽  
Hayden R. Schmidt ◽  
Piotr Sliz ◽  
Andrew C. Kruse
Keyword(s):  
Virtual Screening ◽  
High Throughput ◽  
Transmembrane Protein ◽  
Cross Reactivity ◽  
Receptor Ligands ◽  
Computational Docking ◽  
High Affinity ◽  
Σ1 Receptor ◽  
Ligand Discovery

AbstractThe σ1 receptor is a transmembrane protein implicated in several pathophysiological conditions, including neurodegenerative disease1, drug addiction2, cancer3, and pain4. However, there are no high-throughput functional assays for σ1 receptor drug discovery. Here, we assessed high-throughput structure-based computational docking for discovery of novel ligands of the σ1 receptor. We screened a library of over 6 million compounds using the Schrödinger Glide package, followed by experimental characterization of top-scoring candidates. 77% of tested candidates bound σ1 with high affinity (10-550 nM). These include compounds with high selectivity for the σ1 receptor compared to the genetically unrelated but pharmacologically similar σ2 receptor, as well as compounds with substantial cross-reactivity between the two receptors. These results establish structure-based virtual screening as a highly effective platform for σ1 receptor ligand discovery.

scholarly journals Characterization of protease activity of Nsp3 from SARS-CoV-2 and its in vitro inhibition by nanobodies

2020 ◽  
Author(s):  
Lee A. Armstrong ◽  
Sven M. Lange ◽  
Virginia de Cesare ◽  
Stephen P. Matthews ◽  
Raja Sekar Nirujogi ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Transmembrane Protein ◽  
Cross Reactivity ◽  
Host Cells ◽  
Alternative Methods ◽  
Structural Proteins ◽  
Active Protease ◽  
Micromolar Range

AbstractOf the 16 non-structural proteins (Nsps) encoded by SARS CoV-2, Nsp3 is the largest and plays important roles in the viral life cycle. Being a large, multidomain, transmembrane protein, Nsp3 has been the most challenging Nsp to characterize. Encoded within Nsp3 is the papain-like protease PLpro domain that cleaves not only the viral protein but also polyubiquitin and the ubiquitin-like modifier ISG15 from host cells. We here compare the interactors of PLpro and Nsp3 and find a largely overlapping interactome. Intriguingly, we find that near full length Nsp3 is a more active protease compared to the minimal catalytic domain of PLpro. Using a MALDI-TOF based assay, we screen 1971 approved clinical compounds and identify five compounds that inhibit PLpro with IC50s in the low micromolar range but showed cross reactivity with other human deubiquitinases and had no significant antiviral activity in cellular SARS-CoV-2 infection assays. We therefore looked for alternative methods to block PLpro activity and engineered competitive nanobodies that bind to PLpro at the substrate binding site with nanomolar affinity thus inhibiting the enzyme. Our work highlights the importance of studying Nsp3 and provides tools and valuable insights to investigate Nsp3 biology during the viral infection cycle.

Development of novel phenoxyalkylpiperidines as high-affinity Sigma-1 (σ1) receptor ligands with potent anti-amnesic effect

2021 ◽  
pp. 114038
Author(s):  
Francesca S. Abatematteo ◽  
Philip D. Mosier ◽  
Mauro Niso ◽  
Leonardo Brunetti ◽  
Francesco Berardi ◽  
...  
Keyword(s):  
Receptor Ligands ◽  
Amnesic Effect ◽  
High Affinity ◽  
Σ1 Receptor ◽  
Sigma 1

scholarly journals Deep phenotypic characterization of immunization-induced antibacterial IgG repertoires in mice using a single-antibody bioassay

2020 ◽  
Vol 3 (1) ◽  
Author(s):  
Millie Heo ◽  
Guilhem Chenon ◽  
Carlos Castrillon ◽  
Jérôme Bibette ◽  
Pierre Bruhns ◽  
...  
Keyword(s):  
Humoral Response ◽  
Cross Reactivity ◽  
Phenotypic Characterization ◽  
Antibody Repertoire ◽  
Apparent Dissociation Constant ◽  
Density Profiles ◽  
Bacterial Surface ◽  
High Affinity ◽  
Efficient Elimination

Abstract Antibodies with antibacterial activity need to bind to the bacterial surface with affinity, specificity, and sufficient density to induce efficient elimination. To characterize the anti-bacterial antibody repertoire, we developed an in-droplet bioassay with single-antibody resolution. The assay not only allowed us to identify whether the secreted antibodies recognized a bacterial surface antigen, but also to estimate the apparent dissociation constant (KD app) of the interaction and the density of the recognized epitope on the bacteria. Herein, we found substantial differences within the KD app/epitope density profiles in mice immunized with various species of heat-killed bacteria. The experiments further revealed a high cross-reactivity of the secreted IgG repertoires, binding to even unrelated bacteria with high affinity. This application confirmed the ability to quantify the anti-bacterial antibody repertoire and the utility of the developed bioassay to study the interplay between bacteria and the humoral response.

scholarly journals COVID-19 Repurposed Therapeutics Targeting the Viral Protease and Spike-protein:ACE2 Interface using MD-based Pharmacophore and Consensus Virtual Screening

2020 ◽  
Author(s):  
brady garabato ◽  
Federico Falchi ◽  
Andrea Cavalli
Keyword(s):  
Molecular Dynamics ◽  
Virtual Screening ◽  
Crystal Structures ◽  
High Throughput ◽  
Binding Energies ◽  
Enhanced Sampling ◽  
Viral Protease ◽  
High Affinity ◽  
Receptor Interface

<p>Molecular dynamics (MD) and enhanced sampling MD was performed for 100 ns on the biological assembly of the COVID-19 protease (<a href="https://www.rcsb.org/structure/6lu7">6LU7</a>), and a template of the COVID-19 S-protein:ACE2 receptor interface (99.88% coverage of 6M0J; model03, <a href="https://swissmodel.expasy.org/interactive/HLkhkP/models/">swissmodel</a>). Apo-site pharmacophores of the resulting structural clusters were used to mine the FDA database (8700 compounds), and a multi-target library was developed from MD-based hits in high affinity sites across 100 ns. Consensus hits from high throughput docking in crystal structures 5R82, 6LU7 and 6Y2F (protease), and 6VW1 (S-protein:ACE2) were also added, and the resulting libraries were re-docked into MD sites to collect potential COVID-19 re-purposed therapeutics by estimated binding energies. </p>

scholarly journals Biochemical characterization of protease activity of Nsp3 from SARS-CoV-2 and its inhibition by nanobodies

PLoS ONE ◽  
2021 ◽  
Vol 16 (7) ◽  
pp. e0253364
Author(s):  
Lee A. Armstrong ◽  
Sven M. Lange ◽  
Virginia Dee Cesare ◽  
Stephen P. Matthews ◽  
Raja Sekhar Nirujogi ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Catalytic Domain ◽  
Transmembrane Protein ◽  
Cross Reactivity ◽  
Alternative Methods ◽  
Protease Domain ◽  
Host Cell Proteins ◽  
Active Protease ◽  
Micromolar Range

Of the 16 non-structural proteins (Nsps) encoded by SARS CoV-2, Nsp3 is the largest and plays important roles in the viral life cycle. Being a large, multidomain, transmembrane protein, Nsp3 has been the most challenging Nsp to characterize. Encoded within Nsp3 is the papain-like protease domain (PLpro) that cleaves not only the viral polypeptide but also K48-linked polyubiquitin and the ubiquitin-like modifier, ISG15, from host cell proteins. We here compare the interactors of PLpro and Nsp3 and find a largely overlapping interactome. Intriguingly, we find that near full length Nsp3 is a more active protease compared to the minimal catalytic domain of PLpro. Using a MALDI-TOF based assay, we screen 1971 approved clinical compounds and identify five compounds that inhibit PLpro with IC50s in the low micromolar range but showed cross reactivity with other human deubiquitinases and had no significant antiviral activity in cellular SARS-CoV-2 infection assays. We therefore looked for alternative methods to block PLpro activity and engineered competitive nanobodies that bind to PLpro at the substrate binding site with nanomolar affinity thus inhibiting the enzyme. Our work highlights the importance of studying Nsp3 and provides tools and valuable insights to investigate Nsp3 biology during the viral infection cycle.

scholarly journals COVID-19 Repurposed Therapeutics Targeting the Viral Protease and Spike-protein:ACE2 Interface using MD-based Pharmacophore and Consensus Virtual Screening

2020 ◽  
Author(s):  
brady garabato ◽  
Federico Falchi ◽  
Andrea Cavalli
Keyword(s):  
Molecular Dynamics ◽  
Virtual Screening ◽  
Crystal Structures ◽  
High Throughput ◽  
Binding Energies ◽  
Enhanced Sampling ◽  
Viral Protease ◽  
High Affinity ◽  
Receptor Interface

<p>Molecular dynamics (MD) and enhanced sampling MD was performed for 100 ns on the biological assembly of the COVID-19 protease (<a href="https://www.rcsb.org/structure/6lu7">6LU7</a>), and a template of the COVID-19 S-protein:ACE2 receptor interface (99.88% coverage of 6M0J; model03, <a href="https://swissmodel.expasy.org/interactive/HLkhkP/models/">swissmodel</a>). Apo-site pharmacophores of the resulting structural clusters were used to mine the FDA database (8700 compounds), and a multi-target library was developed from MD-based hits in high affinity sites across 100 ns. Consensus hits from high throughput docking in crystal structures 5R82, 6LU7 and 6Y2F (protease), and 6VW1 (S-protein:ACE2) were also added, and the resulting libraries were re-docked into MD sites to collect potential COVID-19 re-purposed therapeutics by estimated binding energies. </p>

scholarly journals Virtual Screening for Ligand Discovery at the σ1 Receptor

2020 ◽  
Vol 11 (8) ◽  
pp. 1555-1561
Author(s):  
Daniel A. Greenfield ◽  
Hayden R. Schmidt ◽  
Meredith A. Skiba ◽  
Michael D. Mandler ◽  
Jacob R. Anderson ◽  
...  
Keyword(s):  
Virtual Screening ◽  
Σ1 Receptor ◽  
Ligand Discovery

scholarly journals Pharmacological characterization of high‐affinity σ 1 receptor ligands with spirocyclic thienopyran and thienofuran scaffold

2019 ◽  
Vol 72 (2) ◽  
pp. 236-248
Author(s):  
Dirk Schepmann ◽  
Christina Neue ◽  
Stefanie Westphälinger ◽  
Christoph Müller ◽  
Franz Bracher ◽  
...  
Keyword(s):  
Receptor Ligands ◽  
Pharmacological Characterization ◽  
High Affinity
Sign in / Sign up

Export Citation Format

Share Document