nsc95397  (MedChemExpress)

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: NSC95397 identified as a HIV-1 latency-reversal agent (LRA) from a medium-throughput screen. ( A ) Schematic detailing the screen methodology. ( B , C ) Z-scores of total cell count or GFP intensity vs. percent of cells GFP positive. The grey square and purple diamond are two known latency-reversal agents, PMA and IBET151, respectively, and the blue triangle is a previously unknown LRA, NSC95397. ( D ) In total, 83 compounds were rescreened in triplicate at 24 and 48 h. Each row of the heatmap represents a different compound, with the top and second rows being the controls: untreated (UT) and PMA. ( E ) A circle graph representing the pathways/targets of the 35 compounds that were revalidated in ( D ). ( F , G ) The EC 50 titration of NSC95397 and IBET151 in J-Lat 10.6 and 5A8 cells. PMA was used as a positive control at 5.0 ng/mL.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques: Cell Counting, Titration, Positive Control

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: NSC9597 reactivates GFP and p24 expression in J-Lat 10.6 and 5A8 cells. J-Lat 10.6 ( A – E ) or 5A8 ( F ) cells were untreated or treated with 5.0 ng/mL PMA (positive control) or a titration of NSC95397 (NSC) from 12.5 μM to 1.0 μM. ( A , B ) Bar graphs showing GFP expression ( A ) or intracellular HIV-1 p24 ( B ) measured via flow cytometry. ( C , D ) Bar graphs showing GFP ( C ) or unspliced HIV-1 RNA transcripts ( D ) measured via qRT-PCR. ( E , F ) Western blot of whole-cell lysates of either J-Lat 10.6 ( E ) or 5A8 cells ( F ) probed for GFP and actin (loading control); 25.0 μM SAHA was used as a positive control. L = ladder; UT = untreated. Results were analyzed with a one-way ANOVA with Tukey’s multiple comparison tests in ( A , B ) or an unpaired t -test in ( C , D ); ns = not significant, * = p ≤ 0.0332, ** = p ≤ 0.0021, *** = p ≤ 0.0002, **** = p ≤ 0.0001.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques: Expressing, Positive Control, Titration, Flow Cytometry, Quantitative RT-PCR, Western Blot, Control, Comparison

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: NSC95397 increases HIV-1 reactivation in combination with SAHA. ( A , B ) J-Lat 10.6 ( A ) or 5A8 ( B ) cells were untreated or treated with 5.0 ng/mL PMA, 1.5 μM NSC95397 (NSC), a titration of SAHA from 12.5 to 1.0 μM, or a titration of SAHA with NSC held constant at 1.5 μM. ( C , D ) ACH-2 ( C ) or U1 ( D ) cells were untreated or treated with 5.0 ng/mL ( C ) or 20 ng/mL PMA ( D ), 2.5 μM NSC, a titration of SAHA from 10 to 0.5 μM, or a titration of SAHA with NSC held constant at 2.5 μM. Bar graphs show GFP mean fluorescence intensity (MFI) measured via flow cytometry. Results were analyzed with a one-way ANOVA with Tukey’s multiple comparison tests; ns = not significant, * = p ≤ 0.0332, ** = p ≤ 0.0021, *** = p ≤ 0.0002, **** = p ≤ 0.0001.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques: Titration, Fluorescence, Flow Cytometry, Comparison

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: Summary of NSC95397 and SAHA combination index synergy calculations. CI (combination index) values for drug–drug effect were calculated using CompuSyn based on average percent reactivation (see Methods). Concentrations with CI ≤ 0.90 are considered to be synergistic (green)- or greater than additive (with lower values indicating stronger synergy). Concentrations close to 1.0 are considered to have an additive effect (white), in which the two drugs exhibit the same effect from an independent dosage. Concentrations with CI ≥ 1.10 are considered to be antagonistic (purple) or less than additive (with higher values indicating stronger antagonism). Unless otherwise stated, experiments were done in normoxic conditions.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques:

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: NSC95397 increases HIV-1 reactivation in combination with prostratin. ( A , B ) J-Lat 10.6 ( A ) or 5A8 ( B ) cells were untreated or treated with 5.0 ng/mL PMA, NSC95397 (NSC) at 1.5 μM, a titration of prostratin (PS) from 0.66 to 0.1 μM ( A ) or 5.0 to 0.5 μM ( B ), or a titration of PS with NSC held constant at 1.5 μM. ( C , D ) ACH-2 ( C ) or U1 ( D ) cells were untreated or treated with 5.0 ng/mL ( C ) or 20.0 ng/mL PMA ( D ), 2.5 μM NSC, a titration of PS from 0.5 to 0.125 μM ( C ) or 5.0 to 0.5 μM ( D ), or a titration of PS with NSC held constant at 2.5 μM. Bar graphs show GFP MFI measured via flow cytometry. Results were analyzed with a one-way ANOVA with Tukey’s multiple comparison tests; **** = p ≤ 0.0001.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques: Titration, Flow Cytometry, Comparison

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: Summary of NSC95397 and prostratin combination index synergy calculations. CI (combination index) values for drug–drug effect were calculated using CompuSyn based on average percent reactivation (see methods). Concentrations with CI ≤ 0.90 are considered to be synergistic (green), or greater than additive (with lower values indicating stronger synergy). Concentrations close to 1.0 are considered to have an additive effect (white), in which the two drugs exhibit the same effect from an independent dosage. Concentrations with CI ≥ 1.10 are considered to be antagonistic (purple) or less than additive (with higher values indicating stronger antagonism). Unless otherwise stated, experiments were done in normoxic conditions.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques:

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: NSC95397 does not increase global histone markers of open chromatin. ( A , C ) J-Lat 10.6 cells were untreated or treated with 5.0 ng/mL PMA, 25.0 μM SAHA (positive control), or NSC95397 (NSC) at 2.5 μM. Bar graphs show H3K9ac ( A ), H3K4me3 ( B ), or H3K27ac ( C ) MFI measured by flow cytometry. Bar graphs show GFP MFI measured via flow cytometry. Results were analyzed with a one-way ANOVA with Tukey’s multiple comparison tests; ns = not significant, **** = p ≤ 0.0001.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques: Positive Control, Flow Cytometry, Comparison

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: NSC95397 does not globally activate HIV-1 latency reversal-associated pathways. J-Lat 10.6 cells were untreated or treated with 5.0 ng/mL PMA, 25.0 μM SAHA, or 5.0 μM NSC95397 (NSC) and analyzed by bulk RNA sequencing in four biological replicates. ( A ) Principal component analysis (PCA) of all normalized transcripts in all samples. ( B ) Clustered heatmap of all normalized transcripts in all samples. ( C – F ) Volcano plot labeling HIV-associated reads ( C ), selected HIV-1 Tat-modulated genes ( D ), key heat shock response genes ( E ), and HIV-associated transcription factors ( F ). Cut-offs are drawn for log2 fold change above 1 and p -value greater than 0.05 where reads are separated to non-significant and non-enriched (gray), non-significant with log2 fold change above 1.0 (green), significant and log2 fold change below 1.0 (blue), or significant and log2 fold change above 1.0 (red). ( G ) Gene ontology using Gene Set Enrichment Analysis (GSEA) identifying top negatively enriched hallmarks.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques: RNA Sequencing, Labeling

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: NSC95397 causes accumulation of DNA damage markers. ( A ) Representative images of U2OS cells untreated or treated with 10.0 μL DMSO (negative control), 50.0 μM Etoposide (ETOP; DNA damage positive control), 5.0 ng/mL PMA, or 5.0 μM NSC95397 (NSC) for 24 h and assessed for presence of DNA damage markers by confocal microscopy. Blue (Hoechst 33342) shows nuclei, while damage markers (γH2AX, RPA32, or p53BP1) are in red. ( B ) Mean fluorescence intensity (MFI) of at least 100 cells per condition was quantified for γH2AX. Foci per cell of at least 100 cells per condition were quantified for RPA32 and 53BP1. Cells with greater than 60 (RPA32) or 15 (53BP1) foci per cell were binned at the maximum limit of 60 and 15 foci, respectively. Results were analyzed with a one-way ANOVA with Dunnett’s multiple comparison tests; ns = not significant, * = p ≤ 0.0332, ** = p ≤ 0.0021, **** = p ≤ 0.0001.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques: Negative Control, Positive Control, Confocal Microscopy, Fluorescence, Comparison

Journal: Viruses

Article Title: NSC95397 Is a Novel HIV-1 Latency-Reversing Agent

doi: 10.3390/v16111783

Figure Lengend Snippet: NSC95397 increases HIV-1 reactivation in hypoxic conditions. ( A , B ) Bar graphs showing GFP mean fluorescence intensity (MFI) for J-Lat 10.6 ( A ) or 5A8 ( B ) cells. Cells were untreated or treated with 5.0 ng/mL PMA (positive control) or a titration of NSC95397 (NSC) from 5.0 μM to 1.0 μM. Solid bars are cells maintained in normoxia, while patterned bars are cells maintained in hypoxia. ( C , D ) J-Lat 10.6 ( C ) or 5A8 ( D ) cells were grown in hypoxic conditions and untreated or treated with 5.0 ng/mL PMA (positive control), 1.5 μM NSC, or NSC held constant at 1.5 μM with a titration of SAHA from 12.5 to 1.0 μM. Bar graphs show GFP mean fluorescence intensity (MFI) measured via flow cytometry. ( E , F ) J-Lat 10.6 ( E ) or 5A8 ( F ) cells were untreated or treated with 5.0 ng/mL PMA, NSC at 1.5 μM, or NSC held constant at 1.5 μM with a titration of prostratin (PS) from 0.66 to 0.1 μM ( E ) or 5.0 to 0.5 μM ( F ). Bar graphs show GFP MFI measured via flow cytometry. Results were analyzed with a two-way ANOVA with Šídák’s multiple comparison test; ns = not significant, ** = p ≤ 0.0021, *** = p ≤ 0.0002, **** = p ≤ 0.0001.

Article Snippet: Follow-up studies were performed manually with IBET151 (Cayman Chemistry, Ann Arbor, MI, USA, Cat #11181), NSC95397 (Cayman Chemistry, Cat# 21431), PMA (VWR, Radnor, PA, USA, Cat# 102515-692), prostratin (Cayman Chemistry, Cat# 10272), SAHA (Cayman Chemistry, Cat# 10009929), and sunitinib (Cayman Chemistry, Cat# 13159).

Techniques: Fluorescence, Positive Control, Titration, Flow Cytometry, Comparison

Journal: Biology Direct

Article Title: PNSC928, a plant-derived compound, specifically disrupts CtBP2-p300 interaction and reduces inflammation in mice with acute respiratory distress syndrome

doi: 10.1186/s13062-024-00491-0

Figure Lengend Snippet: Proinflammatory cytokine genes were predominately upregulated in ARDS mice. (A) Experimental design: RNA-Seq analysis to identify differentially expressed genes (DGEs) in ARDS mice. (B) Volcano plot displaying RNA-Seq results. (C) Heatmap of top 20 upregulated genes and top 10 downregulated genes in ARDS lung tissues. (D) Biological processes of DGEs revealed by gene ontology (GO) analysis. (E and F) Verification of 12 DGEs by RT-qPCR analysis ( n = 3). (E) IL-1B, IL-6, IL-15, IL-18, TNFA, and IFNG. (F) S100A8, CtBP2, ICAM1, SPP1, FBN1, and SPSB1. ** P < 0.01; *** P < 0.001

Article Snippet: To assess the impact of CtBP2, p300, NF-κB inhibitors, and PNSC928 on ARDS progression, mice were further categorized post-4 h LPS administration into different groups: ARDS, ARDS + MTOB (700 mg/kg; MedChemExpress; Monmouth Junction, NJ; #HY-135,046), ARDS + NSC95397 (1.5 mg/kg; MedChemExpress; #HY-108,543), ARDS + C646 (10 mg/kg; MedChemExpress; #HY-13,823), ARDS + A-485 (50 mg/kg; MedChemExpress; #HY-107,455), ARDS + TPCA1 (10 mg/kg; MedChemExpress; #HY-10,074), ARDS + BOT64 (50 mg/kg; MedChemExpress; #HY-136,741), and ARDS + PNSC928 (20 mg/kg).

Techniques: RNA Sequencing Assay, Quantitative RT-PCR

Journal: Biology Direct

Article Title: PNSC928, a plant-derived compound, specifically disrupts CtBP2-p300 interaction and reduces inflammation in mice with acute respiratory distress syndrome

doi: 10.1186/s13062-024-00491-0

Figure Lengend Snippet: Inhibitors of CtBP2, p300, and NF-κB exhibited cytotoxicity in vivo. C57BL/6 mice were administrated with MTOB (400 and 800 mg/kg), NSC95397 (2 and 4 mg/kg), C646 (5 and 10 mg/kg), A-485 (40 and 80 mg/kg), TPCA1 (10 and 20 mg/kg), BOT64 (30 and 60 mg/kg) for a duration of 6 days ( n = 10 for each group). (A-F) Body weights were measured every two days. (G-K) Serum concentrations of different groups of mice. (G) IL-1β, (H) IL-6, (I) IL-15, (J) IL-18, and (K) TNF-α. * P < 0.05

Article Snippet: To assess the impact of CtBP2, p300, NF-κB inhibitors, and PNSC928 on ARDS progression, mice were further categorized post-4 h LPS administration into different groups: ARDS, ARDS + MTOB (700 mg/kg; MedChemExpress; Monmouth Junction, NJ; #HY-135,046), ARDS + NSC95397 (1.5 mg/kg; MedChemExpress; #HY-108,543), ARDS + C646 (10 mg/kg; MedChemExpress; #HY-13,823), ARDS + A-485 (50 mg/kg; MedChemExpress; #HY-107,455), ARDS + TPCA1 (10 mg/kg; MedChemExpress; #HY-10,074), ARDS + BOT64 (50 mg/kg; MedChemExpress; #HY-136,741), and ARDS + PNSC928 (20 mg/kg).

Techniques: In Vivo

Journal: Biology Direct

Article Title: PNSC928, a plant-derived compound, specifically disrupts CtBP2-p300 interaction and reduces inflammation in mice with acute respiratory distress syndrome

doi: 10.1186/s13062-024-00491-0

Figure Lengend Snippet: Administration of PNSC928 significantly improve the inflammatory outcomes of ARDS mice. (A) A schematic representation illustrating PNSC928 administration. (B-G) Serum concentrations of proinflammatory cytokines by ELISA assays. (B) IL-1β, (C) IL-6, (D) IL-15, (E) IL-18, (F) TNF-α, and (G) IFN-γ. (H ) Body weights of mice measured at 0, 1, and 2 days. (I) pO 2 levels in mice measured at 0, 1, and 2 days. (J) Effects of PNSC928 on the expression levels of IL-1B, IL-6, IL-15, IL-18, TNFA, and IFNG. (K) Effects of PNSC928 on the expression levels of S100A8, CtBP2, ICAM1, SPP1, FBN1, and SPSB1. (L) Representative H&E staining images of lung from control, ARDS, PNSC928 groups of mice. Bars = 100 μm. (M) Quantification of histological scores. Images in (L) were quantified. n = 3 for each experiment. ns: no significant difference. * P < 0.05; ** P < 0.01; *** P < 0.001

Article Snippet: To assess the impact of CtBP2, p300, NF-κB inhibitors, and PNSC928 on ARDS progression, mice were further categorized post-4 h LPS administration into different groups: ARDS, ARDS + MTOB (700 mg/kg; MedChemExpress; Monmouth Junction, NJ; #HY-135,046), ARDS + NSC95397 (1.5 mg/kg; MedChemExpress; #HY-108,543), ARDS + C646 (10 mg/kg; MedChemExpress; #HY-13,823), ARDS + A-485 (50 mg/kg; MedChemExpress; #HY-107,455), ARDS + TPCA1 (10 mg/kg; MedChemExpress; #HY-10,074), ARDS + BOT64 (50 mg/kg; MedChemExpress; #HY-136,741), and ARDS + PNSC928 (20 mg/kg).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Staining, Control

Journal: Biology Direct

Article Title: PNSC928, a plant-derived compound, specifically disrupts CtBP2-p300 interaction and reduces inflammation in mice with acute respiratory distress syndrome

doi: 10.1186/s13062-024-00491-0

Figure Lengend Snippet: A schematic model of targeting CtBP2-p300 by PNSC928 to suppress the expression of proinflammatory cytokine genes and improve ARDS outcomes. (A) This schematic model illustrates the role of the CtBP2-p300-NF-κB complex in the activation of proinflammatory cytokine genes. CtBP2 forms a transcriptional complex with p300 and NF-κB subunits, leading to the activation of proinflammatory cytokine genes (IL-1B, IL-6, IL-15, IL-18, TNFA, and IFNG). The induction of these proinflammatory cytokines promotes the inflammatory response, contributing to the pathogenesis of ARDS. (B) This schematic model demonstrates the mechanism of action of PNSC928 in targeting the CtBP2-p300 complex. PNSC928 specifically disrupts the interaction between CtBP2 and p300, effectively suppressing the expression of proinflammatory cytokine genes. This intervention ultimately leads to improved outcomes in ARDS by mitigating the inflammatory response

Article Snippet: To assess the impact of CtBP2, p300, NF-κB inhibitors, and PNSC928 on ARDS progression, mice were further categorized post-4 h LPS administration into different groups: ARDS, ARDS + MTOB (700 mg/kg; MedChemExpress; Monmouth Junction, NJ; #HY-135,046), ARDS + NSC95397 (1.5 mg/kg; MedChemExpress; #HY-108,543), ARDS + C646 (10 mg/kg; MedChemExpress; #HY-13,823), ARDS + A-485 (50 mg/kg; MedChemExpress; #HY-107,455), ARDS + TPCA1 (10 mg/kg; MedChemExpress; #HY-10,074), ARDS + BOT64 (50 mg/kg; MedChemExpress; #HY-136,741), and ARDS + PNSC928 (20 mg/kg).

Techniques: Expressing, Activation Assay