scholarly journals LY-CoV555, a rapidly isolated potent neutralizing antibody, provides protection in a non-human primate model of SARS-CoV-2 infection

2020 ◽  
Author(s):  
Bryan E. Jones ◽  
Patricia L. Brown-Augsburger ◽  
Kizzmekia S. Corbett ◽  
Kathryn Westendorf ◽  
Julian Davies ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Functional Characterization ◽  
Neutralizing Antibody ◽  
Therapeutic Agents ◽  
Receptor Binding Domain ◽  
Primate Model ◽  
Binding Inhibition ◽  
Antiviral Antibody ◽  
Human Primate

AbstractSARS-CoV-2 poses a public health threat for which therapeutic agents are urgently needed. Herein, we report that high-throughput microfluidic screening of antigen-specific B-cells led to the identification of LY-CoV555, a potent anti-spike neutralizing antibody from a convalescent COVID-19 patient. Biochemical, structural, and functional characterization revealed high-affinity binding to the receptor-binding domain, ACE2 binding inhibition, and potent neutralizing activity. In a rhesus macaque challenge model, prophylaxis doses as low as 2.5 mg/kg reduced viral replication in the upper and lower respiratory tract. These data demonstrate that high-throughput screening can lead to the identification of a potent antiviral antibody that protects against SARS-CoV-2 infection.One Sentence SummaryLY-CoV555, an anti-spike antibody derived from a convalescent COVID-19 patient, potently neutralizes SARS-CoV-2 and protects the upper and lower airways of non-human primates against SARS-CoV-2 infection.

scholarly journals The neutralizing antibody, LY-CoV555, protects against SARS-CoV-2 infection in non-human primates

2021 ◽  
pp. eabf1906
Author(s):  
Bryan E. Jones ◽  
Patricia L. Brown-Augsburger ◽  
Kizzmekia S. Corbett ◽  
Kathryn Westendorf ◽  
Julian Davies ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Functional Characterization ◽  
Neutralizing Antibody ◽  
Therapeutic Agents ◽  
Receptor Binding Domain ◽  
Angiotensin Converting Enzyme 2 ◽  
Cynomolgus Monkeys ◽  
Binding Inhibition ◽  
Vaccination Campaigns

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) poses a public health threat for which preventive and therapeutic agents are urgently needed. Neutralizing antibodies are a key class of therapeutics which may bridge widespread vaccination campaigns and offer a treatment solution in populations less responsive to vaccination. Herein, we report that high-throughput microfluidic screening of antigen-specific B-cells led to the identification of LY-CoV555 (also known as bamlanivimab), a potent anti-spike neutralizing antibody from a hospitalized, convalescent patient with coronavirus disease 2019 (COVID-19). Biochemical, structural, and functional characterization of LY-CoV555 revealed high-affinity binding to the receptor-binding domain, angiotensin converting enzyme 2 binding inhibition, and potent neutralizing activity. A pharmacokinetic study of LY-CoV555 conducted in cynomolgus monkeys demonstrated a mean half-life of 13 days, and clearance of 0.22 mL/hr/kg, consistent with a typical human therapeutic antibody. In a rhesus macaque challenge model, prophylactic doses as low as 2.5 mg/kg reduced viral replication in the upper and lower respiratory tract in samples collected through study Day 6 following viral inoculation. This antibody has entered clinical testing and is being evaluated across a spectrum of COVID-19 indications, including prevention and treatment.

scholarly journals High-Throughput Generation of Product Profiles for Arabinoxylan-Active Enzymes from Metagenomes

2020 ◽  
Vol 86 (23) ◽  
Author(s):  
Maria João Maurício da Fonseca ◽  
Zachary Armstrong ◽  
Stephen G. Withers ◽  
Yves Briers
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Functional Characterization ◽  
Enzymatic Reaction ◽  
Added Value ◽  
Screening Methods ◽  
Reaction Products ◽  
High Throughput Analysis ◽  
Substrate Preferences ◽  
Promising Source

ABSTRACT Metagenomics is an exciting alternative to seek carbohydrate-active enzymes from a range of sources. Typically, metagenomics reveals dozens of putative catalysts that require functional characterization for further application in industrial processes. High-throughput screening methods compatible with adequate natural substrates are crucial for an accurate functional elucidation of substrate preferences. Based on DNA sequencer-aided fluorophore-assisted carbohydrate electrophoresis (DSA-FACE) analysis of enzymatic-reaction products, we generated product profiles to consequently infer substrate cleavage positions, resulting in the generation of enzymatic-degradation maps. Product profiles were produced in high throughput for arabinoxylan (AX)-active enzymes belonging to the glycoside hydrolase families GH43 (subfamilies 2 [MG432], 7 [MG437], and 28 [MG4328]) and GH8 (MG8) starting from 12 (arabino)xylo-oligosaccharides. These enzymes were discovered through functional metagenomic studies of feces from the North American beaver (Castor canadensis). This work shows how enzyme loading alters the product profiles of all enzymes studied and gives insight into AX degradation patterns, revealing sequential substrate preferences of AX-active enzymes. IMPORTANCE Arabinoxylan is mainly found in the hemicellulosic fractions of rice straw, corn cobs, and rice husk. Converting arabinoxylan into (arabino)xylo-oligosaccharides as added-value products that can be applied in food, feed, and cosmetics presents a sustainable and economic alternative for the biorefinery industries. Efficient and profitable AX degradation requires a set of enzymes with particular characteristics. Therefore, enzyme discovery and the study of substrate preferences are of utmost importance. Beavers, as consumers of woody biomass, are a promising source of a repertoire of enzymes able to deconstruct hemicelluloses into soluble oligosaccharides. High-throughput analysis of the oligosaccharide profiles produced by these enzymes will assist in the selection of the most appropriate enzymes for the biorefinery.

High-Throughput Screening of Enzyme Inhibitors: Automatic Determination of Tight-Binding Inhibition Constants

2000 ◽  
Vol 281 (1) ◽  
pp. 62-67 ◽  
Author(s):  
Petr Kuzmič ◽  
Steve Sideris ◽  
Lynne M. Cregar ◽  
Kyle C. Elrod ◽  
Kenneth D. Rice ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Enzyme Inhibitors ◽  
Tight Binding ◽  
Automatic Determination ◽  
Binding Inhibition ◽  
Inhibition Constants

scholarly journals Broadening a SARS-CoV-1 neutralizing antibody for potent SARS-CoV-2 neutralization through directed evolution

2021 ◽  
Author(s):  
Fangzhu Zhao ◽  
Meng Yuan ◽  
Celina Keating ◽  
Namir Shabaani ◽  
Oliver Limbo ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Small Animal ◽  
Neutralizing Antibody ◽  
Therapeutic Agents ◽  
Crystallographic Analysis ◽  
Affinity Maturation ◽  
Receptor Binding Domain ◽  
Resistant Virus ◽  
Engineering Process ◽  
Small Animal Model

The emergence of SARS-CoV-2 underscores the need for strategies to rapidly develop neutralizing monoclonal antibodies that can function as prophylactic and therapeutic agents and to help guide vaccine design. Here, we demonstrate that engineering approaches can be used to refocus an existing neutralizing antibody to a related but resistant virus. Using a rapid affinity maturation strategy, we engineered CR3022, a SARS-CoV-1 neutralizing antibody, to bind SARS-CoV-2 receptor binding domain with >1000-fold improved affinity. The engineered CR3022 neutralized SARS-CoV-2 and provided prophylactic protection from viral challenge in a small animal model of SARS-CoV-2 infection. Deep sequencing throughout the engineering process paired with crystallographic analysis of an enhanced antibody elucidated the molecular mechanisms by which engineered CR3022 can accommodate sequence differences in the epitope between SARS-CoV-1 and SARS-CoV-2. The workflow described provides a blueprint for rapid broadening of neutralization of an antibody from one virus to closely related but resistant viruses.

scholarly journals Validation of FLIPR Membrane Potential Dye for High Throughput Screening of Potassium Channel Modulators

2001 ◽  
Vol 6 (5) ◽  
pp. 305-312 ◽  
Author(s):  
Kristi L. Whiteaker ◽  
Sujatha M. Gopalakrishnan ◽  
Duncan Groebe ◽  
Char-Chang Shieh ◽  
Usha Warrior ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Potassium Channel ◽  
High Throughput ◽  
High Throughput Screening ◽  
Time Course ◽  
Functional Characterization ◽  
K Channel ◽  
Potassium Channel Modulators ◽  
Alternative Choice ◽  
Fluorescent Compounds

A fluorescence-based assay using the FLIPR Membrane Potential Assay Kit (FMP) was evaluated for functional characterization and high throughput screening (HTS) of potassium channel (ATP-sensitive Ki channel; KATP) modulators. The FMP dye permits a more sensitive evaluation of changes in membrane potential with a more rapid response time relative to DiBAC4(3). The time course of responses is comparable to ligand-evoked activation of the channel measured by patch-clamp studies. The pharmacological profile of the K+ channel evaluated by using reference KATP channel openers is in good agreement with that derived previously by DiBAC4(3)-based FLIPR assays. Improved sensitivity of responses together with the diminished susceptibility to artifacts such as those evoked by fluorescent compounds or quenching agents makes the FMP dye an alternative choice for HTS screening of potassium channel modulators.

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