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(A) Seven TLR agonists were compared. (B) Representative structures of the TLR7/8 agonists used in this study. HEK-293 cells transfected with (C) human TLR7 and (D) TLR8 and an NF-κB-driven reporter SEAP gene were stimulated for 18–24 h with TLR agonists. The y-axis shows the level of SEAP activity in the Quanti-blue™ assay optical density (OD). The x-axis shows the concentration of each compound in µM. Each data point represents the mean ± SD of OD at 650 nm of triplicate culture wells. <t>VTX-217</t> (black) is a structurally matched negative control for VTX-744 (gray), VTX-087 (purple) and VTX-294 (red). The IMQ benchmarks R848 and CL075 are denoted in blue and green, respectively.
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(A) Seven TLR agonists were compared. (B) Representative structures of the TLR7/8 agonists used in this study. HEK-293 cells transfected with (C) human TLR7 and (D) TLR8 and an NF-κB-driven reporter SEAP gene were stimulated for 18–24 h with TLR agonists. The y-axis shows the level of SEAP activity in the Quanti-blue™ assay optical density (OD). The x-axis shows the concentration of each compound in µM. Each data point represents the mean ± SD of OD at 650 nm of triplicate culture wells. <t>VTX-217</t> (black) is a structurally matched negative control for VTX-744 (gray), VTX-087 (purple) and VTX-294 (red). The IMQ benchmarks R848 and CL075 are denoted in blue and green, respectively.
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(A) Seven TLR agonists were compared. (B) Representative structures of the TLR7/8 agonists used in this study. HEK-293 cells transfected with (C) human TLR7 and (D) TLR8 and an NF-κB-driven reporter SEAP gene were stimulated for 18–24 h with TLR agonists. The y-axis shows the level of SEAP activity in the Quanti-blue™ assay optical density (OD). The x-axis shows the concentration of each compound in µM. Each data point represents the mean ± SD of OD at 650 nm of triplicate culture wells. <t>VTX-217</t> (black) is a structurally matched negative control for VTX-744 (gray), VTX-087 (purple) and VTX-294 (red). The IMQ benchmarks R848 and CL075 are denoted in blue and green, respectively.
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(A) Seven TLR agonists were compared. (B) Representative structures of the TLR7/8 agonists used in this study. HEK-293 cells transfected with (C) human TLR7 and (D) TLR8 and an NF-κB-driven reporter SEAP gene were stimulated for 18–24 h with TLR agonists. The y-axis shows the level of SEAP activity in the Quanti-blue™ assay optical density (OD). The x-axis shows the concentration of each compound in µM. Each data point represents the mean ± SD of OD at 650 nm of triplicate culture wells. <t>VTX-217</t> (black) is a structurally matched negative control for VTX-744 (gray), VTX-087 (purple) and VTX-294 (red). The IMQ benchmarks R848 and CL075 are denoted in blue and green, respectively.
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(A) Seven TLR agonists were compared. (B) Representative structures of the TLR7/8 agonists used in this study. HEK-293 cells transfected with (C) human TLR7 and (D) TLR8 and an NF-κB-driven reporter SEAP gene were stimulated for 18–24 h with TLR agonists. The y-axis shows the level of SEAP activity in the Quanti-blue™ assay optical density (OD). The x-axis shows the concentration of each compound in µM. Each data point represents the mean ± SD of OD at 650 nm of triplicate culture wells. <t>VTX-217</t> (black) is a structurally matched negative control for VTX-744 (gray), VTX-087 (purple) and VTX-294 (red). The IMQ benchmarks R848 and CL075 are denoted in blue and green, respectively.
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(A) Seven TLR agonists were compared. (B) Representative structures of the TLR7/8 agonists used in this study. HEK-293 cells transfected with (C) human TLR7 and (D) TLR8 and an NF-κB-driven reporter SEAP gene were stimulated for 18–24 h with TLR agonists. The y-axis shows the level of SEAP activity in the Quanti-blue™ assay optical density (OD). The x-axis shows the concentration of each compound in µM. Each data point represents the mean ± SD of OD at 650 nm of triplicate culture wells. <t>VTX-217</t> (black) is a structurally matched negative control for VTX-744 (gray), VTX-087 (purple) and VTX-294 (red). The IMQ benchmarks R848 and CL075 are denoted in blue and green, respectively.
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(A) Seven TLR agonists were compared. (B) Representative structures of the TLR7/8 agonists used in this study. HEK-293 cells transfected with (C) human TLR7 and (D) TLR8 and an NF-κB-driven reporter SEAP gene were stimulated for 18–24 h with TLR agonists. The y-axis shows the level of SEAP activity in the Quanti-blue™ assay optical density (OD). The x-axis shows the concentration of each compound in µM. Each data point represents the mean ± SD of OD at 650 nm of triplicate culture wells. VTX-217 (black) is a structurally matched negative control for VTX-744 (gray), VTX-087 (purple) and VTX-294 (red). The IMQ benchmarks R848 and CL075 are denoted in blue and green, respectively.

Journal: PLoS ONE

Article Title: The Ultra-Potent and Selective TLR8 Agonist VTX-294 Activates Human Newborn and Adult Leukocytes

doi: 10.1371/journal.pone.0058164

Figure Lengend Snippet: (A) Seven TLR agonists were compared. (B) Representative structures of the TLR7/8 agonists used in this study. HEK-293 cells transfected with (C) human TLR7 and (D) TLR8 and an NF-κB-driven reporter SEAP gene were stimulated for 18–24 h with TLR agonists. The y-axis shows the level of SEAP activity in the Quanti-blue™ assay optical density (OD). The x-axis shows the concentration of each compound in µM. Each data point represents the mean ± SD of OD at 650 nm of triplicate culture wells. VTX-217 (black) is a structurally matched negative control for VTX-744 (gray), VTX-087 (purple) and VTX-294 (red). The IMQ benchmarks R848 and CL075 are denoted in blue and green, respectively.

Article Snippet: VTX-217 (a negative control with a similar core structure to other VTX compounds but with modifications to make inactive towards TLRs 7/8), VTX-744 (TLR8), VTX-087 (TLR8) and VTX-294 (TLR8) (VentiRx Pharmaceuticals, Inc., Seattle, WA, USA) were produced using a method of synthesis of substituted benzazepine derivatives .

Techniques: Transfection, Activity Assay, Concentration Assay, Negative Control