FIGURE 2 from Dual Blockade of EP2 and EP4 Signaling is Required for Optimal Immune Activation and Antitumor Activity Against Prostaglandin-Expressing Tumors
Year: 2023
Type: other
Abstract: TPST-1495 is more effective than a single EP4 or EP2 antagonists in overcoming PGE2-mediated immune suppression of recovery of cytokine production from murine monocytes and T cells. A, Schematic of PGE2 blockade assay. Murine whole blood (B–F) or splenic and lymph node T cells (G, H) were collected and incubated with inhibitors for 30 minutes at 37°C, followed by incubation with PGE2 at labeled concentrations for 30 minutes at 37°C, followed by stimulation with LPS (0.5 μg/mL, overnight) or SIINFEKL peptide (2 μmol/L, 5 hours). Detection of TNFα production by ELISA (B–D) and maximum percent recovery (E, F) after blockade assay was performed on murine whole blood. Representative data shown from one of three experiments performed with N = 3 technical replicates per experiment. Some technical replicates failed to meet threshold TNFα values in low Stim groups. Percent recovery as calculated Percent recovery = 100*(“Experimental group” − “LPS+PGE2”)/(“LPS” − “LPS+PGE2”). G and H, IFNγ production detected by Luminex in culture supernatants after treatment with TPST-1495 across a PGE2 concentration gradient (F), or with 333 nmol/L PGE2 comparing multiple PGE2 receptor inhibitors (G) in the PGE2 blockade assay performed with a single-cell suspension of OT-1 splenic and lymph node isolates stimulated with SIINFEKL peptide. Data are representative of three experiments with N = 3 technical replicates per experiment. H, IFNγ production levels in OT-1 T cell cultures containing 333 nmol/L PGE2 and incubated with TPST-1495 EP2/EP4 dual antagonist, or EP2 (PF04418948) or EP4 (E7046) single antagonists. Mean and SD are depicted in all graphs and the ordinary one-way ANOVA with Tukey multiple comparisons test was used to assess significance between individual groups. ****, P < 0.0001; ***, P < 0.001; **, P < 0.01; *, P < 0.05.
Language: English
Subfield: Pharmacology
Field: Medicine
Domain: Health Sciences
Open Access status: gold
OpenAlex ID: W4385658237