Journal: bioRxiv

Article Title: OKN4395, a first-in-class EP2/EP4/DP1 triple antagonist reprograms prostanoid-driven immunosuppression to restore antitumor immunity

doi: 10.64898/2026.02.08.704632

Figure Lengend Snippet: ( A ) Compound 1: lead compound from [29] with moderate dual inhibition; Compound 2. lead compound from [30] with potent EP2 inhibition; Compound 3. lead compound from [31] with potent dual EP2/EP4 inhibition. The OKN4395 series is derived from [31]. Most of the diversity in the series comes from the exploration of the right hand aryl system bearing the carboxylic acid. Compound 3 is an example of that series. ( B ) cAMP production after stimulation with PGE2 (EC80, 2nM) measured on HEK293 stable clones expressing human EP2 (hEP2). Values represent mean of n=10 technical replicates ± SEM. ( C ) cAMP production after stimulation with PGE2 (EC80, 6nM) measured on HEK293 stable clones expressing human EP4 (hEP4). Values represent mean of n=10 technical replicates ± SEM. ( D ) cAMP production after stimulation with PGD2 (EC80, 300pM) measured on CHO-K1 cells stable clone transfected for human DP1 (hDP1). Values represent mean of n=4 technical replicates ± SEM. ( E ) Visualization of OKN4395 antagonist activity at 1µM screened with gpcrMAX GPCR LeadHunter Panel (Eurofins) obtained with the plotrender spatial data mapper tool developed by gpcrdb. ( F ) Summary table of OKN4395 IC50 on human prostanoid receptors. ( G ) Plausible binding mode of a representative of OKN4395 series in DP1 (DP1: orange, ligand: green). Hydrogen bonds are yellow dashed lines.

Article Snippet: RNA was reverse transcribed using the High-Capacity cDNA Reverse Transcription kit (4374967, Fisher Scientific). qPCR was performed in a QuantStudio 7 Pro using the TaqMan Fast Advanced Master Mix (4444557, Fisher Scientific) and the following TaqMan assay ID (Thermo Fisher Scientific) specific for the respective human genes of interest: PTGER1 (Hs00168752_m1), PTGER2 (Hs00168754_m1), PTGER3 (Hs00168755_m1), PTGER4 (Hs00168761_m1), PTGDR (Hs00235003_m1), PTGDR2 (Hs00173717_m1).

Techniques: Inhibition, Derivative Assay, Clone Assay, Expressing, Stable Transfection, Transfection, Activity Assay, Binding Assay

Journal: bioRxiv

Article Title: OKN4395, a first-in-class EP2/EP4/DP1 triple antagonist reprograms prostanoid-driven immunosuppression to restore antitumor immunity

doi: 10.64898/2026.02.08.704632

Figure Lengend Snippet: ( A ) cAMP production after stimulation with PGE2 (EC80) measured on HEK293 stable clones expressing EP2 or EP4 from different species or stimulation with PGD2 (EC80) measured on transfected HEK293 cells transiently expressing DP1 from different species. EC80 PGE2 concentrations are mouse EP2 (mEP2) 300pM, mouse EP4 (mEP4) 200pM, rat EP2 (rEP2) 35-45pM, rat EP4 (rEP4) 5.8-7.8pM, monkey EP2 (mkEP2) 35-40pM, monkey EP4 (mkEP4) 4.2-7.8pM, dog EP2 (dEP2) 65-80pM and dog EP4 (dEP4) 17.8-22.7pM. EC80 PGD2 concentrations are mouse DP1 (mDP1) 2.5nM, rat DP1 (rDP1) 4.4nM and monkey DP1 (mkDP1) 0.75nM. Values represent mean of n=4 (mEP2), n=6 (mEP4), n=10 (rEP2, rEP4, mkEP2, mkEP4, dEP2, dEP4), n=2 (mDP1, rDP1, mkDP1) technical replicates ± SEM. ( B ) cAMP production after stimulation with different PGE2 concentrations measured on SF295 cells (left) or BT549 cells (right) naturally expressing EP2 and EP4 receptor respectively. Values represent mean of n=2 technical replicates ± SEM. ( C ) Activation of (left) hEP1 and (middle) hEP3 after stimulation with PGE2 (EC80, 45nM and 15nM respectively) was measurement on U2OS cells and CHO-K1 cells expressing human EP1 and human EP3 respectively. (Right) cAMP production after stimulation with iloprost (EC80, 60pM) measured on HEK293 stable clones expressing hIP. Values represent mean of n=8 (hEP1, hEP3) and n=4 (hIP) technical replicates ± SEM. ( D ) Selectivity of OKN4395 (top 1µM; bottom 10µM) across 166 G protein-coupled receptors. OKN4395 activity was quantified as the percentage of the natural ligand response, with reduced activity indicating inhibition. ( E ) Superimposition of EP4 (cyan), EP2 (purple) and DP1 (green) binding sites (left). Interactions made by a representative of OKN4395 series in DP1 (right). All the main residues essential for the interactions are conserved in the three proteins. ( F ) Evolution of the distance between the carboxylate of OKN4395 and the interacting arginine residue in EP3, EP4 and DP1, showing the gradual weakening of its strength through movement of the ligand in EP3 and the stability in EP4 and DP1 (solid line: averaged across the triplicates, shaded area: standard deviation).

Article Snippet: RNA was reverse transcribed using the High-Capacity cDNA Reverse Transcription kit (4374967, Fisher Scientific). qPCR was performed in a QuantStudio 7 Pro using the TaqMan Fast Advanced Master Mix (4444557, Fisher Scientific) and the following TaqMan assay ID (Thermo Fisher Scientific) specific for the respective human genes of interest: PTGER1 (Hs00168752_m1), PTGER2 (Hs00168754_m1), PTGER3 (Hs00168755_m1), PTGER4 (Hs00168761_m1), PTGDR (Hs00235003_m1), PTGDR2 (Hs00173717_m1).

Techniques: Clone Assay, Expressing, Transfection, Activation Assay, Activity Assay, Inhibition, Binding Assay, Residue, Standard Deviation

Journal: Animals : an Open Access Journal from MDPI

Article Title: FABP3 Mediates Lipid Droplet Accumulation and Adhesive Capacity in Bovine Endometrial Epithelial Cells via PGE 2 /PTGER4/PPAR Axis

doi: 10.3390/ani15233417

Figure Lengend Snippet: PGE 2 binds to PTGER4 to regulate FABP3 expression in bEECs: ( A ) The difference in PTGER ( PTGER1 , PTGER2 , PTGER3 and PTGER4 ) expression in bEECs after treatment with P 4 and/or PGE 2 . ( B – D ) The representative images and analysis of PTGER2 and PTGER4 expression in bEECs after treatment with P 4 and/or PGE 2 . ( E , F ) The representative images and analysis of PTGER2 expression in bEECs between shN and shPTGER2 groups after treatment with DMSO or P 4 + PGE 2 . ( G , H ) The representative images and analysis of PTGER4 expression in bEECs between shN and shPTGER4 groups after treatment with DMSO or P 4 + PGE 2 . ( I ) The analysis of FABP3 expression in bEECs after treatment with DMSO, PGE 2 or the inhibitor of PTGER4 ( GW627368 ). The data are means ± SEM of three independent experiments. Statistically significant differences are indicated by asterisks: ** p < 0.01, ns p > 0.05. Bars with different letters are significantly different ( p < 0.05).

Article Snippet: Then, PGE 2 (10 −8 –10 −6 M; 14010, Cayman, Ann Arbor, MI, USA) was added to the medium, and the cells were cultured for 24 h. For inhibitor treatment, 5 and 10 μM PTGER2 inhibitor (TG-155; MedChemExpress, Newark, NJ, USA) or 5 and 10 μM PTGER4 inhibitor ( GW627368 ; Selleck, Houston, TX, USA) were added to the medium 1 h prior to the PGE 2 treatment.

Techniques: Expressing