Biology subjects

Ma, C.

Publications and source records attributed to Ma, C..

At least 163 records · Page 9Linked to original sources

bioRxiv2021

Discovery of SARS-CoV-2 papain-like protease inhibitors through a combination of high-throughput screening and FlipGFP-based reporter assay

The papain-like protease (PLpro) of SARS-CoV-2 is a validated antiviral drug target. PLpro is involved in the cleavage of viral polyproteins and antagonizing host innate immune response through its deubiquitinating and deISG15ylating activities, rendering it a high profile antiviral drug target. Through a FRET-based high-throughput screening, several hits were identified as PLpro inhibitors with IC50 values at the single-digit micromolar range. Subsequent lead optimization led to potent inhibitors with IC50 values ranging from 0.56 to 0.90 {micro}M. To help prioritize lead compounds for the cellular antiviral assay against SARS-CoV-2, we developed the cell-based FlipGFP assay that is suitable for quantifying the intracellular enzymatic inhibition potency of PLpro inhibitors in the BSL-2 setting. Two compounds selected from the FlipGFP-PLpro assay, Jun9-53-2 and Jun9-72-2, inhibited SARS-CoV-2 replication in Caco-2 hACE2 cells with EC50 values of 8.89 and 8.32 {micro}M, respectively, which were 3-fold more potent than GRL0617 (EC50 = 25.1 {micro}M). The X-ray crystal structures of PLpro in complex with GRL0617 showed that binding of GRL0617 to SARS-CoV-2 induced a conformational change in the BL2 loop to the more closed conformation. Overall, the PLpro inhibitors identified in this study represent promising starting points for further development as SARS-CoV-2 antivirals, and FlipGFP-PLpro assay might be a suitable surrogate for screening PLpro inhibitors in the BSL-2 setting.

pharmacology and toxicology↗

bioRxiv2021

A global multiregional proteomic map of the human cerebral cortex

The Brodmann area (BA)-based map is one of the most widely used cortical maps for studies of human brain functions and in clinical practice; however, the molecular architecture of BAs remain unknown. The present study provided a global multiregional proteomic map of the human cerebral cortex by analyzing 29 BAs. These 29 BAs were grouped into 6 clusters based on similarities in proteomic patterns: the motor and sensory cluster, vision cluster, auditory cluster and Brocas area, Wernickes area cluster, cingulate cortex cluster, and heterogeneous function cluster. We identified 474 cluster-specific and 134 BA-specific signature proteins whose functions are closely associated with specialized functions and disease vulnerability of the corresponding cluster or BA. The findings of the present study could provide explanations for the functional connections between the anterior cingulate cortex and sensorimotor cortex and for anxiety-related function in the sensorimotor cortex. The brain transcriptomic and proteomic comparison indicated that they both could reflect the function of cerebral cortex, but showed different characteristics. These proteomic data are publicly available at the Human Brain Proteome Atlas (www.brain-omics.com). Our results may enhance our understanding of the molecular basis of brain functions and provide an important resource to support human brain research.

neuroscience↗

bioRxiv2021

Site-specific phosphorylation of PSD-95 dynamically regulates the post-synaptic density as observed by phase separation

Postsynaptic density protein 95 is a key scaffolding protein in the postsynaptic density of excitatory glutamatergic neurons, organizing signaling complexes primarily via its three PSD-95/Discs-large/Zona occludens domains. PSD-95 is regulated by phosphorylation, but technical challenges have limited studies of the molecular details. Here, we genetically introduced site-specific phosphorylations in single, tandem and full-length PSD-95 and generated a total of 11 phosphorylated protein variants. We examined how these phosphorylations affected binding to known interaction partners and the impact on phase separation of PSD-95 complexes, and identified two new phosphorylation sites with opposing effects. Phosphorylation of Ser78 inhibited phase separation with the glutamate receptor subunit GluN2B and the auxiliary protein stargazin, whereas phosphorylation of Ser116 induced phase separation with stargazin only. Thus, by genetically introducing phosphoserine site-specifically and exploring the impact on phase separation, we have provided new insights into the regulation of PSD-95 by phosphorylation and the dynamics of the PSD. Graphical Abstract: Phosphorylation of PSD-95. O_LIIllustration of the interaction between the PDZ domains of PSD-95 and the NMDA receptor (NMDAR) (PDB: 4PE5) at the plasma membrane. PSD-95 interacts with the AMPA receptor (AMPAR) (PDB: 3KG2) through Stargazin (PDB: 5KBU). Using the software Ranch, a model of FL PSD-95 was generated using known structural domains of PSD-95 connected via a fully-flexible linker (PDZ1-2, PDB: 3GSL; PDZ3, PDB: 5JXB; SH3-GK, PDB: 1KJW). C_LIO_LIIllustration of the effect of phosphorylation of PSD-95 on the formation of condensates at the PSD. PSD-95 can phase separate with receptors and intracellular proteins, and phosphorylation is can affect this interaction and lead to alterations, both negative and positive, in the formation of condensates. C_LI

biochemistry↗

bioRxiv2021

A D-2-hydroxyglutarate biosensor based on specific transcriptional regulator DhdR

D-2-Hydroxyglutarate (D-2-HG) is a metabolite in many physiological metabolic processes. When D-2-HG is aberrantly accumulated due to mutations in isocitrate dehydrogenases or D-2-HG dehydrogenase, it functions in a pro-oncogenic manner and is thus considered a therapeutic target and biomarker in many cancers. In this study, DhdR from Achromobacter denitrificans NBRC 15125 was identified as an allosteric transcription factor that negatively regulates D-2-HG dehydrogenase expression and responds to presence of D-2-HG. It is the first known transcription regulator specifically responding to D-2-HG across all domains of life. Based on the allosteric effect of DhdR, a D-2-HG biosensor was developed by combining DhdR with amplified luminescent proximity homogeneous assay technology. The biosensor was able to detect D-2-HG in serum, urine, and cell culture with high specificity and sensitivity. Additionally, this biosensor was also successfully used to identify the role of D-2-HG metabolism in lipopolysaccharide biosynthesis of Pseudomonas aeruginosa, demonstrating its broad usages.

microbiology↗

bioRxiv2021

Brain transcriptional regulatory architecture and schizophrenia etiology converge between East Asian and European ancestral populations

Understanding the genetic architecture of gene expression and splicing in human brain is critical to unlocking the mechanisms of complex neuropsychiatric disorders like schizophrenia (SCZ). Large-scale brain transcriptomic studies are based primarily on populations of European (EUR) ancestry. The uniformity of mono-racial resources may limit important insights into the disease etiology. Here, we characterized brain transcriptional regulatory architecture of East Asians (EAS; n=151), identifying 3,278 expression quantitative trait loci (eQTL) and 4,726 spliceQTL (sQTL). Comparing these to PsychENCODE/BrainGVEX confirmed our hypothesis that the transcriptional regulatory architecture in EAS and EUR brains align. Furthermore, distinctive allelic frequency and linkage disequilibrium impede QTL translation and gene-expression prediction accuracy. Integration of eQTL/sQTL with genome-wide association studies reveals common and novel SCZ risk genes. Pathway-based analyses showing shared SCZ biology point to synaptic and GTPase dysfunction as a prospective pathogenesis. This study elucidates the transcriptional landscape of the EAS brain and emphasizes an essential convergence between EAS and EUR populations.

genetics↗

bioRxiv2021

BnaA03.WRKY28, interacting with BnaA09.VQ12, acts as a brake factor of activated BnWRKY33-mediated resistance outburst against Sclerotinia sclerotiorum in Brassica napus

Sclerotinia sclerotiorum causes substantial damage to the growth of Brassica napus (rapeseed) and makes a significant loss of crop yield. The plant innate immune system may be the primary solution to defense against S. sclerotiorum for rapeseed. Here, we identify that BnWRKY33, a transcription factor in the innate immune pathway, can be rapidly phosphorylated and activated by the MAPK cascade after rapeseed is infected with S. sclerotiorum. In the MAPK cascade, activated BnaA03.MKK4 phosphorylates and activates BnaA06.MPK3 and BnaC03.MPK3. The activated BnMPK3 acts on the substrate BnWRKY33 to enhance its transcriptional activity and trigger a transcriptional burst of BnWRKY33, which helps plants effectively resist the pathogenic fungi by enhancing the expression of phytoalexin synthesis-related genes. With constant infection, BnaA03.WRKY28 and BnaA09.VQ12 are induced, and BnaA03.WRKY28 physically interacts with BnaA09.VQ12 to form a protein complex. BnaA03.WRKY28 preferentially binds to the promoter of BnWRKY33 with the help of BnaA09.VQ12. Compared with activated BnWRKY33, BnaA03.WRKY28 has a lower transcriptional activity on downstream BnWRKY33, which leads to weaker resistance against S. sclerotiorum for rapeseed in the later stage of infection. Furthermore, the induced BnaA03.WRKY28 may promote axillary bud activity and axillary meristem initiation by regulating the expression of branching-related genes (such as BnBRC1), thus promoting the formation of branches in the leaf axils. One-sentence summaryUnder constant infection by Sclerotinia sclerotiorum, BnaA03.WRKY28 interacts with BnaA09.VQ12 and takes precedence over phosphorylated BnWRKY33 to bind to the BnWRKY33 promoter, thereby weakening resistance but promoting branching.

plant biology↗

bioRxiv2021

TAK1: a potential therapeutic target for retinal neovascularization

Retinal neovascularization, or pathological angiogenesis in the retina, is a leading cause of blindness in developed countries. Transforming growth factor-{beta}-activated kinase 1 (TAK1) is a mitogen-activated protein kinase kinase kinase (MAPKKK) activated by TGF-{beta}1 and other pro-inflammatory cytokines. TAK1 is also a key mediator of inflammation, innate immune responses, apoptosis and tissue homeostasis and plays an important role in physiological angiogenesis. Its role in pathological angiogenesis, particularly in retinal neovascularization, remains unclear. We investigated the regulatory role of TAK1 in pathological angiogenesis in the retina. Transcriptome analysis of human retina featuring retinal neovascularization revealed enrichment of known TAK1-mediated signaling pathways. Selective inhibition of TAK1 activation by 5Z-7-oxozeaenol attenuated aberrant retinal angiogenesis in rats following oxygen-induced retinopathy. Transcriptome profiling revealed that TAK1 activation in human microvascular endothelial cells under TNF stimulation led to increase the gene expression related to cytokines and leukocyte-endothelial interaction, mainly through nuclear factor kappa B (NF{kappa}B) signaling pathways. These results reveal that inhibition of TAK1 signaling may have therapeutic value for the treatment of pathological angiogenesis in the retina.

biochemistry↗

bioRxiv2021

Convergent evolution of resistance pathways during early stage breast cancer treatment with combination cell cycle (CDK) and endocrine inhibitors

Combining cyclin-dependent kinase (CDK) inhibitors with endocrine therapy improves outcomes for metastatic estrogen receptor positive (ER+), HER2 negative, breast cancer patients. However, the value of this combination in potentially curable earlier stage patients is not clear. Using single cell transcriptomic profiling, we examined the evolutionary trajectories of early stage breast cancer tumors using serial tumor biopsies from a clinical trial of preoperative endocrine therapy (letrozole) alone or in combination with the cell cycle inhibitor ribociclib. Applying hierarchical regression and Gaussian process mathematical modelling, we classified each tumor by whether it shrinks or persists with therapy and determined cancer phenotypes related to evolution of resistance and cell cycle transcriptional rewiring. We found that all patients tumors undergo subclonal evolution during therapy, irrespective of the clinical response. However, tumors subjected to endocrine therapy alone showed reduced diversity over time, while those facing combination therapy exhibited increased diversity. Despite different subclonal diversity, single nuclei RNA sequencing uncovered common phenotypic changes in tumor cells that persist following treatment. In these tumors, cancer cells with accelerated loss of estrogen signaling have convergent up-regulation of the JNK pathway, while cells that maintain estrogen signaling during therapy show potentiation of CDK4/6 activation consistent with ERBB4 and ERK signaling up-regulation. These convergent phenotypes were associated with growing tumors resistant to combination therapy. Cell cycle reconstruction identified that these tumors can rebound during combination therapy treatment, indicating stronger selection and promotion of a proliferative state. These results indicate that combination therapy in early stage ER+ breast cancers with ER and CDK inhibition drives rapid evolution of resistance via a shift from estrogen signaling to alternative growth factor receptor mediated proliferation and JNK signaling activation, concordant with a bypass in the G1 checkpoint.

cancer biology↗

bioRxiv2021

Neuromodulation of Behavioral Specialization: Tachykinin Signaling Inhibits Task-Specific Behavioral Responsiveness in Honeybee Workers

Behavioral specialization is key to the success of social insects and often compartmentalized among colony members leading to division of labor. Response thresholds to task-specific stimuli proximally regulate behavioral specialization but their neurobiological regulation is not understood. Here, we show that response thresholds to task-relevant stimuli correspond to the specialization of three behavioral phenotypes of honeybee workers. Quantitative neuropeptidome comparisons suggest two tachykinin-related peptides (TRP2 and TRP3) as candidates for the modification of these response thresholds. Based on our characterization of their receptor binding and downstream signaling, we then confirm the functional role of tachykinins: TRP2 injection and RNAi cause consistent, opposite effects on responsiveness to task-specific stimuli of each behaviorally specialized phenotype but not to stimuli that are unrelated to their tasks. Thus, our study demonstrates that TRP-signaling regulates the degree of task-specific responsiveness of specialized honeybee workers and may control the context-specificity of behavior in animals more generally.

evolutionary biology↗

bioRxiv2020

easyMF: A Web Platform for Matrix Factorization-based Biological Discovery from Large-scale Transcriptome Data

With the development of high-throughput experimental technologies, large-scale RNA sequencing (RNA-Seq) data have been and continue to be produced, but have led to challenges in extracting relevant biological knowledge hidden in the produced high-dimensional gene expression matrices. Here, we present easyMF, a user-friendly web platform that aims to facilitate biological discovery from large-scale transcriptome data through matrix factorization (MF). The easyMF platform enables users with little bioinformatics experience to streamline transcriptome analysis from raw reads to gene expression and to decompose expression matrix from thousands of genes to a handful of metagenes. easyMF also offers a series of functional modules for metagene-based exploratory analysis with an emphasis on functional gene discovery. As a modular, containerized and open-source platform, easyMF can be customized to satisfy users specific demands and deployed as a web server for broad applications. easyMF is freely available at https://github.com/cma2015/easyMF. We demonstrated the application of easyMF with four case studies using 940 RNA sequencing datasets from maize (Zea mays L.).

bioinformatics↗

bioRxiv2020

Dissecting human skeletal stem cell ontogeny by single-cell transcriptomic and functional analyses

Human skeletal stem cells (SSCs) have been discovered in fetal and adult bones. However, the spatiotemporal ontogeny of human SSCs during embryogenesis has been elusive. Here we map the transcriptional landscape of human embryonic skeletogenesis at single-cell resolution to address this fundamental question. We found remarkable heterogeneity within human limb bud mesenchyme and epithelium, as well as the earliest osteo-chondrogenic progenitors. Importantly, embryonic SSCs (eSSCs) were found in the perichondrium of human long bones, which self-renew and generate osteochondral lineage cells, but not adipocytes or hematopoietic stroma. eSSCs are marked by the adhesion molecule CADM1 and highly enrich FOXP1/2 transcriptional network. Interestingly, neural crest-derived cells with similar phenotypic markers and transcriptional network were also found in the sagittal suture of human embryonic calvaria. Taken together, this study revealed the cellular heterogeneity and lineage hierarchy during human embryonic skeletogenesis, and identified distinct skeletal stem/progenitor cells that orchestrate endochondral and intramembranous ossification.

developmental biology↗

bioRxiv2020

An expedited approach towards the rationale design of non-covalent SARS-CoV-2 main protease inhibitors with in vitro antiviral activity

The main protease (Mpro) of SARS-CoV-2 is a validated antiviral drug target. Several Mpro inhibitors have been reported with potent enzymatic inhibition and cellular antiviral activity, including GC376, boceprevir, calpain inhibitors II and XII, each containing a reactive warhead that covalently modifies the catalytic Cys145. In this study, we report an expedited drug discovery approach by coupling structure-based design and Ugi four-component (Ugi-4CR) reaction methodology to the design of non-covalent Mpro inhibitors. The most potent compound 23R had cellular antiviral activity similar to covalent inhibitors such as GC376. Our designs were guided by overlaying the structure of SARS-CoV Mpro + ML188 (R), a non-covalent inhibitor derived from Ug-4CR, with the X-ray crystal structures of SARS-CoV-2 Mpro + calpain inhibitor XII/GC376/UAWJ247. Binding site analysis suggests a strategy of extending the P2 and P3 substitutions in ML188 (R) to achieve optimal shape complementary with SARS-CoV-2 Mpro. Lead optimization led to the discovery of 23R, which inhibits SARS-CoV-2 Mpro and SARS-CoV-2 viral replication with an IC50 of 0.31 M and EC50 of 1.27 M, respectively. The binding and specificity of 23R to SARS-CoV-2 Mpro were confirmed in a thermal shift assay and native mass spectrometry assay. The co-crystal structure of SARS-CoV-2 Mpro with 23R revealed the P2 biphenyl fits snuggly into the S2 pocket and the benzyl group in the -methylbenzyl faces towards the core of the enzyme, occupying a previously unexplored binding site located in between the S2 and S4 pockets. Overall, this study revealed the most potent non-covalent SARS-CoV-2 Mpro inhibitors reported to date and a novel binding pocket that can be explored for Mpro inhibitor design.

pharmacology and toxicology↗

bioRxiv2020

Interactive Web-based Annotation of Plant MicroRNAs with iwa-miRNA

MicroRNAs (miRNAs) are important regulators of gene expression. The large-scale detection and profiling of miRNAs has accelerated with the development of high-throughput small RNA sequencing (sRNA-Seq) techniques and bioinformatics tools. However, generating high-quality comprehensive miRNA annotations remains challenging, due to the intrinsic complexity of sRNA-Seq data and inherent limitations of existing miRNA predictions. Here, we present iwa-miRNA, a Galaxy-based framework that can facilitate miRNA annotation in plant species by combining computational analysis and manual curation. iwa-miRNA is specifically designed to generate a comprehensive list of miRNA candidates, bridging the gap between already annotated miRNAs provided by public miRNA databases and new predictions from sRNA-Seq datasets. It can also assist users to select promising miRNA candidates in an interactive mode through the automated and manual steps, contributing to the accessibility and reproducibility of genome-wide miRNA annotation. iwa-miRNA is user-friendly and can be easily deployed as a web application for researchers without programming experience. With flexible, interactive, and easy-to-use features, iwa-miRNA is a valuable tool for annotation of miRNAs in plant species with reference genomes. We illustrated the application of iwa-miRNA for miRNA annotation of plant species with varying complexity. The sources codes and web server of iwa-miRNA is freely accessible at: http://iwa-miRNA.omicstudio.cloud/.

plant biology↗

bioRxiv2020

Analytical validation and performance characteristics of a 48-gene next-generation sequencing panel for detecting potentially actionable genomic alterations in myeloid neoplasms

Identification of genomic mutations by molecular testing plays an important role in diagnosis, prognosis, and treatment of myeloid neoplasms. Next-generation sequencing (NGS) is an efficient method for simultaneous detection of clinically significant genomic mutations with high sensitivity. However, due to lack of standard NGS protocols, the application of NGS for hematologic malignancies into clinical settings remains limited. We report development and validation of a 48-gene NGS panel for molecular profiling of myeloid neoplasms including acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), and myeloproliferative neoplasms (MPN). Target regions were captured by hybridization with complementary biotinylated DNA baits, and NGS was performed on an Illumina NextSeq500 instrument. A bioinformatics pipeline that was developed in-house was used to detect single nucleotide variations (SNVs), insertions/deletions (indels), and FLT3 internal tandem duplications (FLT3-ITD). An analytical validation study was performed on 184 unique specimens for variants with allele frequencies [≥]5%. Variants identified by the 48-gene panel were compared to those identified by a 35-gene hematologic neoplasms panel using an additional 137 unique specimens. The developed assay was applied to a large cohort (n=2,053) of patients with suspected myeloid neoplasms. Analytical validation yielded 99.6% sensitivity (95% CI: 98.9-99.9%) and 100% specificity (95% CI: 100%). Concordance of variants detected by the 2 tested panels was 100%. Among patients with suspected myeloid neoplasms (n=2,053), 54.5% patients harbored at least one clinically significant mutation: 77% in AML patients, 48% in MDS, and 45% in MPN. Together, these findings demonstrate that the assay can identify mutations associated with diagnosis, prognosis, and treatment options of myeloid neoplasms.

cancer biology↗

bioRxiv2020

Boceprevir, calpain inhibitors II and XII, and GC-376 have broad-spectrum antiviral activity against coronaviruses in cell culture

As the COVID-19 pandemic continues to fold out, the morbidity and mortality are increasing daily. Effective treatment for SARS-CoV-2 is urgently needed. We recently discovered four SARS-CoV-2 main protease (Mpro) inhibitors including boceprevir, calpain inhibitors II and XII and GC-376 with potent antiviral activity against infectious SARS-CoV-2 in cell culture. Despite the weaker enzymatic inhibition of calpain inhibitors II and XII against Mpro compared to GC-376, calpain inhibitors II and XII had more potent cellular antiviral activity. This observation promoted us to hypothesize that the cellular antiviral activity of calpain inhibitors II and XII might also involve the inhibition of cathepsin L in addition to Mpro. To test this hypothesis, we tested calpain inhibitors II and XII in the SARS-CoV-2 pseudovirus neutralization assay in Vero E6 cells and found that both compounds significantly decreased pseudoviral particle entry into cells, indicating their role in inhibiting cathepsin L. The involvement of cathepsin L was further confirmed in the drug time-of-addition experiment. In addition, we found that these four compounds not only inhibit SARS-CoV-2, but also SARS-CoV, MERS-CoV, as well as human coronaviruses (CoVs) 229E, OC43, and NL63. The mechanism of action is through targeting the viral Mpro, which was supported by the thermal shift binding assay and enzymatic FRET assay. We further showed that these four compounds have additive antiviral effect when combined with remdesivir. Altogether, these results suggest that boceprevir, calpain inhibitors II and XII, and GC-376 are not only promising antiviral drug candidates against existing human coronaviruses, but also might work against future emerging CoVs.

pharmacology and toxicology↗

bioRxiv2020

Cooperation-based sperm clusters mediate sperm oviduct entry and fertilization

Sperm cooperation has been observed in multiple species, yet its existence and benefit for reproductive success in mammals remains underexplored. Here, combining tissue-clearing with deep three-dimensional imaging, we demonstrate that postcopulatory mouse sperm congregate into unidirectional sperm cooperative clusters at the utero-tubal junction (UTJ), a key physical barrier for passage into the oviduct. Reducing sperm number in male mice by unilateral vasoligation or busulfan-treatment impairs sperm cluster formation and oviduct entry. Interestingly, sperm derived from Tex101-/- mouse has normal number, motility and morphology, yet they cannot form sperm cluster and fail to pass through the UTJ, which is at least in part due to the altered tail beating pattern of the Tex101-/- sperm. Moreover, Tex101-/- sperms defect in oviduct entry cannot be rescued by the presence of wild-type (WT) sperm in the same uteri by sequential mating, suggesting sperm cooperative cluster as an essential behavior contributing to male fertility, which could be related to human infertility or subfertility.

developmental biology↗

bioRxiv2020

Ebselen, disulfiram, carmofur, PX-12, tideglusib, and shikonin are non-specific promiscuous SARS-CoV-2 main protease inhibitors

There is an urgent need for vaccines and antiviral drugs to combat the COVID-19 pandemic. Encouraging progress has been made in developing antivirals targeting SARS-CoV-2, the etiological agent of COVID-19. Among the drug targets being investigated, the viral main protease (Mpro) is one of the most extensively studied drug targets. Mpro is a cysteine protease that hydrolyzes the viral polyprotein at more than 11 sites and it is highly conserved among coronaviruses. In addition, Mpro has a unique substrate preference for glutamine in the P1 position. Taken together, it appears that Mpro inhibitors can achieve both broad-spectrum antiviral activity and a high selectivity index. Structurally diverse compounds have been reported as Mpro inhibitors, with several of which also showed antiviral activity in cell culture. In this study, we investigated the mechanism of action of six previously reported Mpro inhibitors, ebselen, disulfiram, tideglusib, carmofur, shikonin, and PX-12 using a consortium of techniques including FRET-based enzymatic assay, thermal shift assay, native mass spectrometry, cellular antiviral assays, and molecular dynamics simulations. Collectively, the results showed that the inhibition of Mpro by these six compounds is non-specific and the inhibition is abolished or greatly reduced with the addition of reducing reagent DTT. In the absence of DTT, these six compounds not only inhibit Mpro, but also a panel of viral cysteine proteases including SARS-CoV-2 papain-like protease, the 2Apro and 3Cpro from enterovirus A71 (EV-A71) and EV-D68. However, none of the compounds inhibits the viral replication of EV-A71 or EV-D68, suggesting that the enzymatic inhibition potency IC50 values obtained in the absence of DTT cannot be used to faithfully predict their cellular antiviral activity. Overall, we provide compelling evidence suggesting that ebselen, disulfiram, tideglusib, carmofur, shikonin, and PX-12 are non-specific SARS-CoV-2 Mpro inhibitors, and urge the scientific community to be stringent with hit validation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=92 SRC="FIGDIR/small/299164v1_ufig1.gif" ALT="Figure 1"> View larger version (13K): org.highwire.dtl.DTLVardef@1e68820org.highwire.dtl.DTLVardef@1f31c02org.highwire.dtl.DTLVardef@1b406e1org.highwire.dtl.DTLVardef@2fa8e8_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2020

Discovery of a Novel Inhibitor of Coronavirus 3CL Protease as a Clinical Candidate for the Potential Treatment of COVID-19

COVID-19 caused by the SARS-CoV-2 virus has become a global pandemic. 3CL protease is a virally encoded protein that is essential across a broad spectrum of coronaviruses with no close human analogs. The designed phosphate prodrug PF-07304814 is metabolized to PF-00835321 which is a potent inhibitor in vitro of the coronavirus family 3CL pro, with selectivity over human host protease targets. Furthermore, PF-00835231 exhibits potent in vitro antiviral activity against SARS-CoV-2 as a single agent and it is additive/synergistic in combination with remdesivir. We present the ADME, safety, in vitro, and in vivo antiviral activity data that supports the clinical evaluation of this compound as a potential COVID-19 treatment.

pharmacology and toxicology↗