Journal: BMC Biology

Article Title: Identification of Epinastine as CD96/PVR inhibitor for cancer immunotherapy

doi: 10.1186/s12915-025-02132-y

Figure Lengend Snippet: Virtual screening of small molecules from the FDA-approved drug library. A Crystal structure of CD96 in complex with PVR (PDB Code: 6ARQ), with the binding interface highlighted in blue. B Detailed depiction of amino acids involved in the CD96/PVR interaction, with residues shown in stick representation. C Relationship between molecular weight and S-value for selected small molecules. Red indicates molecules with molecular weights between 250 and 700 and S-values of -5 or less. D-F Results of further screening of small molecules interacting with the CD96/PVR binding interface. G Distribution of S-values for the final 15 selected small molecules

Article Snippet: 1729 small molecules were obtained from FDA-approved drug library (Topscience, Shanghai, China).

Techniques: Drug discovery, Binding Assay, Molecular Weight

Journal: Pharmaceutics

Article Title: Evaluation of In Vitro and In Vivo Antiviral Activities of Vitamin D for SARS-CoV-2 and Variants

doi: 10.3390/pharmaceutics15030925

Figure Lengend Snippet: Schematic overview of the anti-SARS-CoV-2 drug screening procedure. ( a ) ACE2 targeted inhibitors and compounds from the natural product library were applied prior to infection with SARS-CoV-2 (pre-infection treatment) while compounds from the FDA-approved library and the flavonoid library were applied post infection (post- infection treatment). Out of 2191 compounds tested, 121 displayed viral CPE reductions and 7 were selected for further validations using Vero E6 and HuH7 cell lines and hNEC. ( b ) Uninfected cells, infected cells treated with 0.1% DMSO, and infected cells treated with remdesivir were included as controls in the screen. Following treatment with citicoline, pravastatin sodium, tenofovir alafenamide, imatinib mesylate, calcitriol, dexlansoprazole, and prochlorperazine dimaleate, reduced CPE was observed when compared to the DMSO control.

Article Snippet: A primary screen to identify novel compounds with potential antiviral effects against SARS-CoV-2 infection was performed on four different libraries, namely a 462-compound ACE2-targeted compound library (CADD) (TargetMol, Boston, MA, USA), a 57-compound natural product library, a 500-compound flavonoids library (TimTec, Tampa, FL, USA), and a 1172-compound FDA-approved drug library (Selleckchem, Houston, TX, USA).

Techniques: Drug discovery, Infection, Control

Journal: Pharmaceutics

Article Title: Evaluation of In Vitro and In Vivo Antiviral Activities of Vitamin D for SARS-CoV-2 and Variants

doi: 10.3390/pharmaceutics15030925

Figure Lengend Snippet: Summary of compounds identified from the primary screen that exhibit potential antiviral effects against SARS-CoV-2 infection.

Article Snippet: A primary screen to identify novel compounds with potential antiviral effects against SARS-CoV-2 infection was performed on four different libraries, namely a 462-compound ACE2-targeted compound library (CADD) (TargetMol, Boston, MA, USA), a 57-compound natural product library, a 500-compound flavonoids library (TimTec, Tampa, FL, USA), and a 1172-compound FDA-approved drug library (Selleckchem, Houston, TX, USA).

Techniques: Infection, Reverse Transcription, Reflux

Journal: Pharmaceutics

Article Title: Evaluation of In Vitro and In Vivo Antiviral Activities of Vitamin D for SARS-CoV-2 and Variants

doi: 10.3390/pharmaceutics15030925

Figure Lengend Snippet: Validation of primary hits. For validating compounds with activity pre-infection, Vero E6 cells were first pre-treated with increasing concentrations of ( a ) citicoline, ( b ) pravastatin sodium and ( c ) tenofovir alafenamide prior to infection with SARS-CoV-2. Similarly, Huh7 cells were pre-treated with ( d ) citicoline, ( e ) pravastatin sodium and ( f ) tenofovir alafenamide and subsequently infected with SARS-CoV-2. For post-infection treatment validation, Vero E6 cells were infected with SARS-CoV-2 and treated with increasing concentrations of ( g ) imatinib mesylate, ( h ) calcitriol, ( i ) dexlansoprazole and ( j ) prochlorperazine dimaleate. Similarly, HuH7 cells were also infected then treated with a range of concentrations of ( k ) imatinib mesylate, ( l ) calcitriol, ( m ) dexlansoprazole and ( n ) prochlorperazine dimaleate. The primary and secondary exes correspond to the viral titre and relative cell viability, and the dashed line represents the CC 50 cut-off for cell viability. One-way ANOVA and Dunnett’s post-test were used to determine statistical differences, with * denoting p < 0.05, ** denoting p < 0.01 and *** denoting p < 0.001. Error bars represent the standard deviation observed from the means of triplicates performed for both cell viability and dose-dependent inhibition studies.

Article Snippet: A primary screen to identify novel compounds with potential antiviral effects against SARS-CoV-2 infection was performed on four different libraries, namely a 462-compound ACE2-targeted compound library (CADD) (TargetMol, Boston, MA, USA), a 57-compound natural product library, a 500-compound flavonoids library (TimTec, Tampa, FL, USA), and a 1172-compound FDA-approved drug library (Selleckchem, Houston, TX, USA).

Techniques: Biomarker Discovery, Activity Assay, Infection, Standard Deviation, Inhibition

Journal: Pharmaceutics

Article Title: Evaluation of In Vitro and In Vivo Antiviral Activities of Vitamin D for SARS-CoV-2 and Variants

doi: 10.3390/pharmaceutics15030925

Figure Lengend Snippet: Validation of primary hits in hNECs. Selected primary hits were further validated in hNECs. Cells were either pre-treated with 10 μM citicoline prior to SARS-CoV-2 infection or treated following SARS-CoV-2 infection with 10 μM imatinib mesylate and 10 μM calcitriol. One-way ANOVA and Dunnett’s post-test were used to determine statistical differences, with ** denoting that p < 0.01. Error bars represent the standard deviation observed from the means of triplicates performed for dose-dependent inhibition studies.

Article Snippet: A primary screen to identify novel compounds with potential antiviral effects against SARS-CoV-2 infection was performed on four different libraries, namely a 462-compound ACE2-targeted compound library (CADD) (TargetMol, Boston, MA, USA), a 57-compound natural product library, a 500-compound flavonoids library (TimTec, Tampa, FL, USA), and a 1172-compound FDA-approved drug library (Selleckchem, Houston, TX, USA).

Techniques: Biomarker Discovery, Infection, Standard Deviation, Inhibition

Journal: Pharmaceutics

Article Title: Evaluation of In Vitro and In Vivo Antiviral Activities of Vitamin D for SARS-CoV-2 and Variants

doi: 10.3390/pharmaceutics15030925

Figure Lengend Snippet: SARS-CoV-2 and calcitriol modulate vitamin D Receptor (VDR), 24-hydroxylase (24(OH)ase), and cathelicidin (LL-37) in Vero E6 and Huh7 cells. In the first part of the experiment, Vero E6 and Huh7 cells were either mock-infected with media, or SARS-CoV-2 infected and harvested at indicated timepoints in the absence of calcitriol for baseline VDR, 24(OH)ase, and LL-37 mRNA expression. Vero E6 expression of ( a ) VDR, ( b ) 24(OH)ase and ( c ) LL-37 and Huh7 expression of ( d ) VDR, ( e ) 24(OH)ase and ( f ) LL-37 were as shown. In the second part of the experiment, Vero E6 and Huh7 cells were either mock-infected with media or SARS-CoV-2 for 1 h as above, then treated with 0.01, 0.1, 1, 10 or 20 µM calcitriol following infection. At 48 h post infection, the cells were harvested to check for VDR, 24(OH)ase, and LL-37 mRNA expression. Vero E6 mRNA expression of ( g ) VDR, ( i ) 24(OH)ase and ( k ) LL-37 and Huh7 expression of ( h ) VDR, ( j ) 24(OH)ase and ( l ) LL-37 were as shown. Data are represented as relative quantity to 0.1% DMSO control, or as relative quantity to mock-infected control. One-way ANOVA and Dunnett’s post-test and paired t -test were used to determine statistical differences, with * denoting p < 0.05, ** denoting p < 0.01 and *** denoting p < 0.001. Error bars represent the standard deviation observed from the means of triplicates performed for mRNA expression studies.

Article Snippet: A primary screen to identify novel compounds with potential antiviral effects against SARS-CoV-2 infection was performed on four different libraries, namely a 462-compound ACE2-targeted compound library (CADD) (TargetMol, Boston, MA, USA), a 57-compound natural product library, a 500-compound flavonoids library (TimTec, Tampa, FL, USA), and a 1172-compound FDA-approved drug library (Selleckchem, Houston, TX, USA).

Techniques: Infection, Expressing, Control, Standard Deviation

Journal: Pharmaceutics

Article Title: Evaluation of In Vitro and In Vivo Antiviral Activities of Vitamin D for SARS-CoV-2 and Variants

doi: 10.3390/pharmaceutics15030925

Figure Lengend Snippet: Western blot analysis of SARS-CoV-2 spike and VDR proteins in Vero E6 cells. ( a ) Vero E6 cells are either mock infected with media (−) or with SARS-CoV-2 (+) for 1 h. After 1 h post-infection, the cells are then either untreated (−) or treated (+) with 20 µM calcitriol for 6-, 24- and 48- hours before harvesting of total cell lysate. ( b ) Densitometry quantitation of protein expression levels is shown as fold changes to beta-actin. Full-length blots are shown in .

Article Snippet: A primary screen to identify novel compounds with potential antiviral effects against SARS-CoV-2 infection was performed on four different libraries, namely a 462-compound ACE2-targeted compound library (CADD) (TargetMol, Boston, MA, USA), a 57-compound natural product library, a 500-compound flavonoids library (TimTec, Tampa, FL, USA), and a 1172-compound FDA-approved drug library (Selleckchem, Houston, TX, USA).

Techniques: Western Blot, Infection, Quantitation Assay, Expressing

Journal: Pharmaceutics

Article Title: Evaluation of In Vitro and In Vivo Antiviral Activities of Vitamin D for SARS-CoV-2 and Variants

doi: 10.3390/pharmaceutics15030925

Figure Lengend Snippet: In vivo study of calcitriol treatment. 8-week old K18-hACE2 male and female mice were separated into treatment and non-treatment groups. Treatment study group has been given 3 days pre-treatment and 5 days post-treatment of calcitriol (5 μg/kg) via intraperitoneal injection. The non-treatment group has been given a PBS as a treatment control. Next, 10 3 PFU of SARS-CoV-2 (L, Alpha, and Beta variants) were inoculated through intranasal delivery to all groups after pre-treatment. Physiological parameters: body weight ( a – c ), survival rate ( d – f ), and physiological conditions were monitored after infections. For comparison between individual physiological conditions, 5 dpi infected mice were compared between treatment groups of all three variant infected mice ( g – i ). Scoring of physiological conditions was based on five criteria: appearance of the mouse coat, level of consciousness, activity level, eye condition, and respiratory quality. To compare the severity of damage in mouse tissues after infection, left lung lobes from 4 dpi mice were processed for histological analyses and scored based on six criteria to determine severity ( j – l ). The six criteria are inflammatory cell infiltration, haemorrhage, oedema, bronchial epithelial cell damage, degeneration of alveolar epithelial cells, and parenchymal wall expansion. For viral load determination, right lung, brain, liver, and spleen tissues from the same 4 dpi mice were harvested and homogenised for virus titration ( m – o ). Data is not shown for liver and spleen tissues. Statistical significance is determined with a two-tailed unpaired t -test, with * denoting p < 0.05. Survival group: variant L: n = 6 (treatment and no treatment); Alpha: n = 6 (treatment), n = 4 (no treatment); Beta: n = 6 (treatment), n = 5 (no treatment). 4 dpi group: variant L: n = 7 (treatment and no treatment); Alpha and Beta: n = 5 each (treatment and no treatment). Mock infection, n = 3.

Article Snippet: A primary screen to identify novel compounds with potential antiviral effects against SARS-CoV-2 infection was performed on four different libraries, namely a 462-compound ACE2-targeted compound library (CADD) (TargetMol, Boston, MA, USA), a 57-compound natural product library, a 500-compound flavonoids library (TimTec, Tampa, FL, USA), and a 1172-compound FDA-approved drug library (Selleckchem, Houston, TX, USA).

Techniques: In Vivo, Injection, Control, Comparison, Infection, Variant Assay, Activity Assay, Virus, Titration, Two Tailed Test

Journal: Blood

Article Title: MDH1-mediated malate-aspartate NADH shuttle maintains the activity levels of fetal liver hematopoietic stem cells

doi: 10.1182/blood.2019003940

Figure Lengend Snippet: STAT3 transactivates Mdh1 expression to maintain FL-HSC activities. (A) Mdh1 luciferase reporter and different doses of Stat3 were cotransfected into 293T cells, followed by the determination of luciferase activities (n = 3). (B) ChIP assays were analyzed with 293T cells cotransfected with pGL4.27-mdh1 promoter vector and Stat3 plasmid or control plasmid. Input control and amplification of the Stat3-binding sequence of Mdh1 were determined by semiquantitative PCR. (C) Bisulfite sequencing analysis of the methylation patterns of the CpG islands in the Mdh1 promoter in CD45+ SoNar-high and -low FL hematopoietic cell fractions is shown. Each row represents a unique DNA clone; filled and open circles represent methylated and unmethylated CpGs, respectively. (D) Quantification data of methylated and unmethylated CpGs (n = 3). (E) Protein levels of pSTAT3, STAT3, and MDH1 were measured in CD45+ SoNar-high and -low FL cells. Ratios of pSTAT3/STAT3 and MDH1/actin were quantified and normalized against SoNar-high#1 cells. (F) Representative flow cytometric analysis of intracellular level of pSTAT3 of total CD45+ FL hematopoietic cells and FL-HSCs. Mean fluorescence intensity (MFI) of each group was shown (isotype control, yellow and green lines). (G) Quantification of MFI of intracellular level of pSTAT3 in panel F (n = 3). (H) Immunoblotting assay showed protein levels of pSTAT3, STAT3, and MDH1 in total CD45+ FL hematopoietic cells and FL-HSCs. Ratios of pSTAT3/STAT3 and MDH1/actin were quantified and normalized against total cells. (I) Representative flow cytometric analysis of intracellular level of pSTAT3 of CD45+ SoNar-high and -low FL-HSCs. MFI of each group is shown (isotype control, yellow and green lines). (J) Quantification of MFI of intracellular level of pSTAT3 in panel I (n = 3). (K) Protein level of MDH1 was measured in Stat3-overexpressed SoNar 32D cells and control cells by immunoblotting. Ratios of STAT3/actin and MDH1/actin were quantified and normalized against control cells. (L) Stat3-overexpressed SoNar 32D cells and control cells were evaluated for the ratios of SoNar fluorescence, and representative images are shown. Scale bar, 10 μm. (M) Quantification of the ratios of SoNar fluorescence in panel L. A total of 30 SoNar 32D cells were analyzed (n = 3). (N) Working model for the connections between FL-HSC activities and Mdh1-mediated malate-aspartate shuttle as indicated by SoNar. Data are represented as mean ± standard error of the mean. Student 2-tailed unpaired t test (G,J,M) and 2-way analysis of variance with Sidak’s multiple comparison test (D) were used for the comparison. *P < .05, ***P < .001.

Article Snippet: To test the effect of STAT3 inhibitor on MDH1 expression, purified Lin − Sca-1 + c-Kit + FL-HSCs and adult HSCs were treated with 10 or 20 μM of STAT3 inhibitor C188-9 (TargetMol) for 24 hours according to the previous study 37 and subjected to determination of pSTAT3, STAT3, or MDH1 level by western blot.

Techniques: Expressing, Luciferase, Plasmid Preparation, Control, Amplification, Binding Assay, Sequencing, Methylation Sequencing, Methylation, Fluorescence, Western Blot, Comparison

Journal: Science Advances

Article Title: Designer cellular spheroids with DNA origami for drug screening

doi: 10.1126/sciadv.ado9880

Figure Lengend Snippet: ( A ) Schematic of the high-throughput AI screening platform for identifying antifibrotic candidate molecules using fibrosis package NAC-Livers. ( B ) Representative high-content images of fibrotic pNAC-Livers treated with 37 candidate molecules, where PHHs were labeled with GFP (green) and HSCs were labeled with RFP (red). See fig. S21A for additional information on the locations and concentrations of 37 candidate molecules. n = 3 independent experiments. ( C and D ) Heatmap showing the fluorescence intensity of GFP (C) and RFP (D) in high-content imaging of each pNAC-Liver in (B). ( E ) The binding pose of axitinib (candidate 4) with PDK1. ( F ) H&E, Sirius Red, and α-SMA staining of mouse liver sections from three treatment groups: negative control (NC), DDC Ctrl, and axitinib. NC represents the group with normal diet, DDC Ctrl represents the group with 0.1% DDC diet modeling and intraperitoneal injection of solvent [saline with 2% dimethyl sulfoxide (DMSO) + 30% polyethylene glycol 300 + 2% Tween 80], and axitinib represents the group fed with 0.1% DDC diet and intraperitoneal injection of axitinib. See table S2 for the concentration for in vivo administration. ( G and H ) Quantification of Sirius Red–positive area (G) and the H-score in α-SMA immunohistochemical staining slices (H) of each group in (F). ( I and J ) The concentrations of serum aspartate aminotransferase (AST) (I) and alanine aminotransferase (ALT) (J) in mouse serum of each group in (F). Data in (G) to (J) are mean ± SD of n = 3 independent experiments.

Article Snippet: All small molecules used in hepatotoxicity tests and high-content drug screening were purchased from TopScience.

Techniques: High Throughput Screening Assay, Labeling, Fluorescence, Imaging, Binding Assay, Staining, Negative Control, Injection, Solvent, Saline, Concentration Assay, In Vivo, Immunohistochemical staining