High throughput screening of single atomic catalysts with optimized local structures for the electrochemical oxygen reduction by machine learning

Single atomic catalysts(SACs),especially metal-nitrogen doped carbon(M-NC)catalysts,have been extensively explored for the electrochemical oxygen reduction reaction(ORR),owing to their high activity and atomic utilization efficiency.However,there is still a lack of systematic screening and optimization of local structures surrounding active centers of SACs for ORR as the local coordination has an essential impact on their electronic structures and catalytic performance.Herein,we systematic study the ORR catalytic performance of M-NC SACs with different central metals and environmental atoms in the first and second coordination sphere by using density functional theory(DFT)calculation and machine learning(ML).The geometric and electronic informed overpotential model(GEIOM)based on random forest algorithm showed the highest accuracy,and its R^(2) and root mean square errors(RMSE)were 0.96 and 0.21,respectively.30 potential high-performance catalysts were screened out by GEIOM,and the RMSE of the predicted result was only 0.12 V.This work not only helps us fast screen high-performance catalysts,but also provides a low-cost way to improve the accuracy of ML models.

Discovery of natural products capable of inducing porcine host defense peptide gene expression using cell-based high throughput screening

Background:In-feed antibiotics are being phased out in livestock production worldwide.Alternatives to antibiotics are urgently needed to maintain animal health and production performance.Host defense peptides(HDPs)are known for their broad-spectrum antimicrobial and immunomodulatory capabilities.Enhancing the synthesis of endogenous HDPs represents a promising antibiotic alternative strategy to disease control and prevention.Methods:To identify natural products with an ability to stimulate the synthesis of endogenous HDPs,we performed a high-throughput screening of 1261 natural products using a newly-established stable luciferase reporter cell line known as IPEC-J2/pBD3-luc.The ability of the hit compounds to induce HDP genes in porcine IPEC-J2 intestinal epithelial cells,3D4/31 macrophages,and jejunal explants were verified using RT-qPCR.Augmentation of the antibacterial activity of porcine 3D4/31 macrophages against a Gram-negative bacterium(enterotoxigenic E.coli)and a Gram-positive bacterium(Staphylococcus aureus)were further confirmed with four selected HDP-inducing compounds.Results:A total of 48 natural products with a minimum Z-score of 2.0 were identified after high-throughput screening,with 21 compounds giving at least 2-fold increase in luciferase activity in a follow-up dose-response experiment.Xanthohumol and deoxyshikonin were further found to be the most potent in inducing pBD3 mRNA expression,showing a minimum 10-fold increase in IPEC-J2,3D4/31 cells,and jejunal explants.Other compounds such as isorhapontigenin and calycosin also enhanced pBD3 mRNA expression by at least 10-fold in both IPEC-J2 cells and jejunal explants,but not 3D4/31 cells.In addition to pBD3,other porcine HDP genes such as pBD2,PG1-5,and pEP2C were induced to different magnitudes by xanthohumol,deoxyshikonin,isorhapontigenin,and calycosin,although clear gene-and cell type-specific patterns of regulation were observed.Desirably,these four compounds had a minimum effect on the expression of several representative inflammatory cytokine genes.Furthermore,when used at HDP-inducing concentrations,these compounds showed no obvious direct antibacterial activity,but significantly augmented the antibacterial activity of 3D4/31 macrophages(P<0.05)against both Gram-negative and Gram-positive bacteria.Conclusions:Our results indicate that these newly-identified natural HDP-inducing compounds have the potential to be developed as novel alternatives to antibiotics for prophylactic and therapeutic treatment of infectious diseases in livestock production.

A High-affinity Activator of G551D-CFTR Chloride Channel Identified By High Throughput Screening

A stably transfected CHO cell line coexpressing G551D-CFTR and iodide-sensitive yellow fluorescent protein mutant EYFP-H148Q-I152L was successfully established and used as assay model to identify small-molecule activators of G551D-CFTR chloride channel from 100000 diverse combinatorial compounds by high throughput screening on a customized Beckman robotic system. A bicyclooctane compound was identified to activate G551D-CFTR chloride channel with high-affinity(K d=1.8 μmol/L). The activity of the bicyclooctane compound is G551D-CFTR-specific, reversible and non-toxic. The G551D-CFTR activator may be useful as a tool to study the mutant G551D-CFTR chloride channel structure and transport properties and as a candidate drug to cure cystic fibrosis caused by G551D-CFTR mutation.

Self-encoding Functional Resin Applying for Combinatorial Chemistry and High Throughput Screening

A novel solid phase organic synthesis resin was synthesized for combinatorial high-throughput screening,which based on FTIR spectra self-encoding functional resin technology. A new deconvolution strategy termed position encoding deconvolution had illustrated and was compared with some popular combinatorial deconvolution strategies in efficiency and information content. The mimic high throughput screening of hexapeptide library successfully proved the applying of the self-encoding functional resin technology and the position encoding deconvolution strategy.

A Class of High-affinity Bicyclooctane G551D-CFTR Activators Identified by High Throughput Screening

The glycine-to-aspartic acid missense mutation at the codon 551(G551D) of the cystic fibrosis transmembrane conductance regulator(CFTR) is one of the five most frequent cystic fibrosis(CF) mutations associated with a severe CF phenotype. To explore the feasibility of pharmacological correction of disrupted activation of CFTR chloride channel caused by G551D mutation, we developed a halide-sensitive fluorescence miniassay for G551D-CFTR in Fisher rat thyroid(FRT) epithelial cells for the discovery of novel activators of G551D-CFTR. A class of bicyclooctane small molecule compounds that efficiently stimulate G551D-CFTR chloride channel activity was identified by high throughput screening via the FRT cell-based assay. This class of compounds selectively activates G551D-CFTR with a high affinity, whereas little effect of the compounds on wildtype CFTR can be seen. The discovery of a class of bicyclooctane G551D-CFTR activators will permit the analysis of structure-activity relationship of the compounds to identify ideal leads for in vivo therapeutic studies.

INTRODUCTION OF THE HIGH THROUGHPUT SCREENING SYSTEM

In this article, we introduce the system of high throughput screening (HTS). Its role in new drug study and current development is described. The relationship between research achievements of genome study and new type screening model of new drugs is emphasized. The personal opinions of current problems about HTS study in China are raised.

High throughput screening of mesenchymal stromal cell morphological response to inflammatory signals for bioreactor-based manufacturing of extracellular vesicles that modulate microglia

Due to their immunomodulatory function,mesenchymal stromal cells(MSCs)are a promising therapeutic with the potential to treat neuroinflammation associated with neurodegenerative diseases.This function is mediated by secreted extracellular vesicles(MSC-EVs).Despite established safety,MSC clinical translation has been unsuccessful due to inconsistent clinical outcomes resulting from functional heterogeneity.Current approaches to mitigate functional heterogeneity include‘priming’MSCs with inflammatory signals to enhance function.However,comprehensive evaluation of priming and its effects on MSC-EV function has not been performed.Furthermore,clinical translation of MSC-EV therapies requires significant manufacturing scale-up,yet few studies have investigated the effects of priming in bioreactors.As MSC morphology has been shown to predict their immunomodulatory function,we screened MSC morphological response to an array of priming signals and evaluated MSC-EV identity and potency in response to priming in flasks and bioreactors.We identified unique priming conditions corresponding to distinct morphologies.These conditions demonstrated a range of MSC-EV preparation quality and lipidome,allowing us to discover a novel MSC-EV manufacturing condition,as well as gain insight into potential mechanisms of MSC-EV microglia modulation.Our novel screening approach and application of priming to MSC-EV bioreactor manufacturing informs refinement of larger-scale manufacturing and enhancement of MSC-EV function.

The development of a sensitive fluorescent protein-based transcript reporter for high throughput screening of negative modulators of lncRNAs

While the human genome is pervasively transcribed,<2%of the human genome is transcribed into protein-coding mRNAs,leaving most of the transcripts as noncoding RNAs,such as microRNAs and long-noncoding RNAs(lncRNAs),which are critical components of epigenetic regulation.lncRNAs are emerging as critical regulators of gene expression and genomic stability.However,it remains largely unknown about how lncRNAs are regulated.Here,we develop a highly sensitive and dynamic reporter that allows us to identify and/or monitor negative modulators of lncRNA transcript levels in a high throughput fashion.Specifically,we engineer a fluorescent fusion protein by fusing three copies of the PEST destruction domain of mouse ornithine decarboxylase(MODC)to the C-terminal end of the codon-optimized bilirubin-inducible fluorescent protein,designated as dBiFP,and show that the dBiFP protein is highly destabilized,compared with the commonly-used eGFP protein.We further demonstrate that the dBiFP signal is effectively down-regulated when the dBiFP and mouse lncRNA H19 chimeric transcript is silenced by mouse H19-specific siRNAs.Therefore,our results strongly suggest that the dBiFP fusion protein may serve as a sensitive and dynamic transcript reporter to monitor the inhibition of lncRNAs by microRNAs,synthetic regulatory RNA molecules,RNA binding proteins,and/or small molecule inhibitors so that novel and efficacious inhibitors targeting the epigenetic circuit can be discovered to treat human diseases such as cancer and other chronic disorders.

Statistical considerations for high throughput screening data

High throughput screening(HTS)is a widely used effective approach in genome-wide association and large scale protein expression studies,drug discovery,and biomedical imaging research.How to accurately identify candidate‘targets’or biologically meaningful features with a high degree of confidence has led to extensive statistical research in an effort to minimize both false-positive and false-negative rates.A large body of literature on this topic with in-depth statistical contents is available.We examine currently available statistical methods on HTS and aim to summarize some selected methods into a concise,easy-tofollow introduction for experimental biologists.

A high throughput antiviral screening platform for alphaviruses based on Semliki Forest virus expressing eGFP reporter gene

Alphaviruses,which contain a variety of mosquito-borne pathogens,are important pathogens of emerging/reemerging infectious diseases and potential biological weapons.Currently,no specific antiviral drugs are available for the treatment of alphaviruses infection.For most highly pathogenic alphaviruses are classified as risk group-3 agents,the requirement of biosafety level 3(BSL-3)facilities limits the live virus-based antiviral study.To facilitate the antiviral development of alphaviruses,we developed a high throughput screening(HTS)platform based on a recombinant Semliki Forest virus(SFV)which can be manipulated in BSL-2 laboratory.Using the reverse genetics approach,the recombinant SFV and SFV reporter virus expressing eGFP(SFV-eGFP)were successfully rescued.The SFV-eGFP reporter virus exhibited robust eGFP expression and remained relatively stable after four passages in BHK-21 cells.Using a broad-spectrum alphavirus inhibitor ribavirin,we demonstrated that the SFV-eGFP can be used as an effective tool for antiviral study.The SFV-eGFP reporter virus-based HTS assay in a 96-well format was then established and optimized with a robust Z0 score.A section of reference compounds that inhibit highly pathogenic alphaviruses were used to validate that the SFV-eGFP reporter virus-based HTS assay enables rapid screening of potent broad-spectrum inhibitors of alphaviruses.This assay provides a safe and convenient platform for antiviral study of alphaviruses.

Drug discovery from traditional Chinese herbal medicine using high content imaging technology

Traditional Chinese medicine(TCM) has been widely used in China and other Asia countries for thousands of years to treat or prevent human diseases. Chinese herbal medicine, one of the most important components of TCM, has unique diversities in chemical components, and thus results in a wide range of biological activities. However, pharmaceutical industry is facing a major challenge to develop a large population of novel natural products and drugs, and considerable efforts have not resulted in highvolume of novel drug discovery and productivity. At present, increasing attention has been paid to Chinese herb medicine modernization in combination with the cutting-age technologies of drug discovery, especially the high throughput selection. High content imaging is an image-based high throughput screening method by using automated microscopy and image analysis software to capture and analyze phenotypes at a large scale to investigate multiple biological features simultaneously in the biological complex. Here, we described the pipeline of the state-of-the-art high content imaging technology, summarized the applications of the high content imaging technology in drug discovery from traditional Chinese herbal medicine, and finally discussed the current challenges and future perspectives for development of high throughput image-based screening technology in novel drug research and discovery.

Identification and optimization of a potent and highly selective D3 dopamine receptor agonist

OBJECTIVE Dopamine receptors(DRs) are involved in the development and treatment of many neuropsychiatric disorders.Currently available dopaminergic drugs modulate both DRD2 and DRD3,leading to side effects and uncertainty as to the roles each DR subtype plays physiologically.Our lab employed high throughput screening paradigms to discover highly selective modulators for the DRD3.METHODS The NIH Molecular Libraries Program 400,000 + small molecule library was screened using the Discove Rx Path Hunter?β-arrestin assay for compounds that activate the DRD3 without effects on the DRD2.Confirmation and counter-screens assessed selectivity and mechanisms of action.We identified 62 potential agonists,and chose the most promising to perform a structure-activity relationship(SAR) study to increase potency while maintaining selectivity.The lead compound identified through this process,ML417,was also characterized using bioluminescence resonance energy transfer(BRET)-based β-arrestin recruitment and G-protein activation assays as well as p-ERK assays.Potential neuroprotective properties of this compound were assessed using a SHSY5 Y neuronal cell model.RESULTS ML417 displays potent,DRD3-selective agonist activity in multiple functional assays.Binding and functional GPCR screens(>165 receptors) show ML417 has limited cross-reactivity with other GPCRs.ML417 also displays superior(compared to the reference compound pramipexole),dose-dependent protection against a decrease in neurite length induced by 10 μmol·L^(-1) of the neurotoxin,6-hydroxydopamine,in the SHSY5 Y cell model.CONCLUSION We have discovered and characterized ML417,a potent and highly selective DRD3 agonist.This compound will be useful as a research tool,and may prove useful as a therapeutic drug lead.

High-throughput screening of carbon-supported single metal atom catalysts for oxygen reduction reaction

Carbon-supported transition metal single atoms are promising oxygen reduction reaction(ORR)electrocatalyst.Since there are many types of carbon supports and transition metals,the accurate prediction of the components with high activity through theoretical calculations can greatly save experimental time and costs.In this work,the ORR catalytic properties of 180 types single-atom catalysts(SACs)composed of the eight representative carbon-based substrates(graphdiyne,C_(2)N,C_(3)N_(4),phthalocyanine,C-coordination graphene,N-coordination graphene,covalent organic frameworks and metal-organic frameworks)and 3d,4d,and 5d transition metal elements are investigated by density functional theory(DFT).The adsorption free energy of OH^(*) is proved a universal descriptor capable of accurately prediction of the ORR catalytic activity.It is found that the oxygen reduction reaction overpotentials of all the researched SACs follow one volcano shape very well with the adsorption free energy of OH^(*).Phthalocyanine,N-coordination graphene and metal-organic frameworks stand out as the promising supports for single metal atom due to the relatively lower overpotentials.Notably,the Co-doped metal-organic frameworks,Ir-doped phthalocyanine,Co-doped N-coordination graphene,Co-doped graphdiyne and Rh-doped phthalocyanine show extremely low overpotentials comparable to that of Pt(111).The study provides a guideline for design and selection of carbon-supported SACs toward oxygen reduction reaction.

A novel non-radioactive assay for HIV-RT(RdDp)based on pyrosequencing for high-throughput drug screening

Current in vitro assays for the activity of HIV-RT(reverse transcriptase)require radio-labeled or chemically modified nucleotides to detect reaction products.However,these assays are inherently end-point measurements and labor intensive.Here we describe a novel non-radioactive assay based on the principle of pyrosequencing coupledenzyme system to monitor the activity of HIV-RT by indirectly measuring the release of pyrophosphate(PPi),which is generated during nascent strand synthesis.The results show that our assay could monitor HIV-RT activity with high sensitivity and is suitable for rapid highthroughput drug screening targeting anti-HIV therapies due to its high speed and convenience.Moreover,this assay can be used to measure primase activity in an easy and sensitive manner,which suggests that this novel approach could be wildly used to analyze the activity of PPi-generated and ATP-free enzyme reactions.

Hepatitis C Virus Experimental Model Systems and Antiviral drug Research

An estimated 130 million people worldwide are chronically infected with hepatitis C virus (HCV) making it a leading cause of liver disease worldwide. Because the currently available therapy of pegylated interferon-alpha and ribavirin is only effective in a subset of patients, the development of new HCV antivirals is a healthcare imperative. This review discusses the experimental models available for HCV antiviral drug research, recent advances in HCV antiviral drug development, as well as active research being pursued to facilitate development of new HCV-specific therapeutics.

Advances of multiplex and high throughput biomolecular detection technologies based on encoding microparticles

The rapid developments of genomics and proteomics have driven the demand for multiplex and high throughput analysis of large numbers of biomolecules in the fields of medical diagnostics, drug discovery, and environmental monitoring. Encoding the biomolecular binding events is the key technique to fulfill this demand, in which microparticles play the most important roles. This review outlines the development of multiplex and high throughput biodetections, and highlights the most recent advances in the field of encoding microparticles, together with problems that need to be resolved.

Computational discovery of energy materials in the era of big data and machine learning:A critical review

The discovery of novel materials with desired properties is essential to the advancements of energy-related technologies.Despite the rapid development of computational infrastructures and theoretical approaches,progress so far has been limited by the empirical and serial nature of experimental work.Fortunately,the situation is changing thanks to the maturation of theoretical tools such as density functional theory,high-throughput screening,crystal structure prediction,and emerging approaches based on machine learning.Together these recent innovations in computational chemistry,data informatics,and machine learning have acted as catalysts for revolutionizing material design and hopefully will lead to faster kinetics in the development of energy-related industries.In this report,recent advances in material discovery methods are reviewed for energy devices.Three paradigms based on empiricism-driven experiments,database-driven high-throughput screening,and data informatics-driven machine learning are discussed critically.Key methodological advancements involved are reviewed including high-throughput screening,crystal structure prediction,and generative models for target material design.Their applications in energy-related devices such as batteries,catalysts,and photovoltaics are selectively showcased.

Effect of Guizhi Fuling capsule and its extracts on human breast cancer cells proliferation

OBJECTIVE To evaluate the effect of Guizhi Fuling Capsule active pharmaceutical ingredient(API)and its fractions on human breast cancer cells proliferation by high-throughput screening assay.METHODS The crude fractions were obtained from the extraction and elution of the API of Guizhi Fuling Capsule,and 929 standard fractions were obtained by the optimal separation conditions.Sulforhodamine B(SRB)method was used to evaluate the effects of the Guizhi Fuling capsule API and929 kinds of fractions on the proliferation of human breast cancer cells MCF-7 and MDA-MB-231.RESULTS The Guizhi Fuling capsule API had a strong ability to inhibit the proliferation of MCF-7 cells at high concentration and the ability to inhibit MDA-MB-231 cells' proliferate at low concentration following 72 h treatment;some samples of 929 fractions(5μg·mL^(-1))was found to have a breast cancer cell growth inhibition rate above 50%,without toxicity on HUVECs proliferation.CONCLUSION The API of Guizhi Fuling capsule had significant cytotoxicity effects on these two human breast cancer cells,with significant concentration-and time-dependent manner.

Current technologies to identify protein kinase substrates in high throughput

Since the discovery of protein phosphorylation as an important modulator of many cellular processes, the involvement of protein kinases in diseases, such as cancer, diabetes, cardiovascular diseases, and central nervous system pathologies, has been extensively documented. Our understanding of many disease pathologies at the molecular level, therefore, requires the comprehensive identification of substrates targeted by protein kinases. In this review, we focus on recent techniques for kinase substrate identification in high throughput, in particular on genetic and proteomic approaches. Each method with its inherent advantages and limitations is discussed.

Identification of novel small-molecule inhibitors of SARS-CoV-2 by chemical genetics

There are only eight approved small molecule antiviral drugs for treating COVID-19.Among them,four are nucleotide analogues(remdesivir,JT001,molnupiravir,and azvudine),while the other four are protease inhibitors(nirmatrelvir,ensitrelvir,leritrelvir,and simnotrelvir-ritonavir).Antiviral resistance,unfavourable drug‒drug interaction,and toxicity have been reported in previous studies.Thus there is a dearth of new treatment options for SARS-CoV-2.In this work,a three-tier cell-based screening was employed to identify novel compounds with anti-SARS-CoV-2 activity.One compound,designated 172,demonstrated broad-spectrum antiviral activity against multiple human pathogenic coronaviruses and different SARS-CoV-2 variants of concern.Mechanistic studies validated by reverse genetics showed that compound 172 inhibits the 3-chymotrypsin-like protease(3CLpro)by binding to an allosteric site and reduces 3CLpro dimerization.A drug synergistic checkerboard assay demonstrated that compound 172 can achieve drug synergy with nirmatrelvir in vitro.In vivo studies confirmed the antiviral activity of compound 172 in both Golden Syrian Hamsters and K18 humanized ACE2 mice.Overall,this study identified an alternative druggable site on the SARS-CoV-23CLpro,proposed a potential combination therapy with nirmatrelvir to reduce the risk of antiviral resistance and shed light on the development of allosteric protease inhibitors for treating a range of coronavirus diseases.