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compounds  (ATCC)
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compound 7  (MedChemExpress)
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Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with <t>Compound</t> <t>7</t> at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.
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Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with <t>Compound</t> <t>7</t> at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.
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Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with <t>Compound</t> <t>7</t> at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.
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compound 7 id5c  (Ambeed Inc)
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Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with <t>Compound</t> <t>7</t> at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.
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1 8 diazabicyclo  (Aladdin Scientific Corporation)
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Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with <t>Compound</t> <t>7</t> at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.
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compounds dalspinin 7 o β d galactopyranoside  (Pfizer Inc)
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Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with <t>Compound</t> <t>7</t> at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.
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Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with <t>Compound</t> <t>7</t> at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.
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Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with Compound 7 at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.

Journal: ACS Chemical Biology

Article Title: High-Throughput Phenotypic Screen to Identify FoxP3 Regulators in Primary T Cells

doi: 10.1021/acschembio.5c01019

Figure Lengend Snippet: Compounds validation in cell-based assay. ( A–C) CD3 + T cells were treated with the specified compounds with or without TCR stimulation (anti-CD2/CD3/CD28 beads) at the indicated concentrations for 24 h. Cells were then stained with viability dye Ax700, followed by flow cytometry analysis of the percentage of FoxP3 in live CD4 + T cells. D) CD3 + T cells were treated with Compound 7 at the indicated concentrations for 48 h under TCR stimulation. The expression of Treg markers was assessed through flow cytometry analysis by gating on CD4 + FoxP3 + T cells. For TCR stimulation, anti-CD2/CD3/CD28 beads were added to the cells in a 1:2 ratio. E ) Tregs were treated with Compound 7 for 48 h and then cocultured with CellTrace Far Red-stained Teffs for 4 days. The proliferation of CD4 + and CD8 + Teffs was determined by measuring the percentage of CellTrace Far Red + cells among live cells, which was normalized to the Teff-only controls. Data are represented as mean ± SEM ( n = 3 healthy donors). * p < 0.5, ** p < 0.05, *** p < 0.005, **** p < 0.0005, 2-way ANOVA.

Article Snippet: Compound 7 , Ethoxyquin , MedChemExpress , HY-B1425 , 98.76%.

Techniques: Biomarker Discovery, Cell Based Assay, Staining, Flow Cytometry, Expressing