Journal: Science translational medicine

Article Title: Metabolic adaptations of ovarian tumors in patients treated with an IDO1 inhibitor constrains antitumor immune responses

doi: 10.1126/scitranslmed.abg8402

Figure Lengend Snippet: (A) Cumulative data of the fold change in human healthy donor CD8+ T cell proliferation stimulated with anti-CD3/CD28 beads in the presence of the indicated combinations of NAD and antagonists to A2BR (PSB 1115), A2AR (SCH 58261), P2XR (oATP), P2X1R (NF 449), P2X3R (NF 110), A1R (PSB 36), P2X4R (5-BDBD) and P2Y1R (MRS 2279) respectively. In all box charts, the median and min-max of the values are presented; *P < 0.05, **P < 0.01, ***P < 0.001,****P < 0.0001. Wilcoxon and one-way ANOVA tests. (B) Survival data of IE9mp1-mIDO1 tumor-bearing mice treated with EPA, A2a, A2b receptor antagonist, or combination of EPA, A2a, and A2b receptor antagonist (n= 5 animals per group). Mice were treated by oral gavage with 300mg/kg EPA in 200μl 0.5% methylcellulose, i.p. injection 5 days on and two days off from d7-d21, with or without (i) NAMPT inhibitor FK866 (500 μg per mouse i.p.) once every week for 3 weeks (on day 13, 20, 27); and (ii) with or without A2a, A2b or A2a/A2b antagonists (1mg/kg, i.p. 5 days on two days off from d7-d21). *, P<0.05 compared to vehicle. Statistical analysis was calculated using a log-rank analysis. (C). Heatmap representing the Log2FC of core set genes of immune pathways that are significantly differentially expressed (FDR < 0.05) in the treatment groups compared with the vehicle group of mice. Blue and red indicate down- and up-regulation, respectively.

Article Snippet: C57BL/6 mice were challenged intraperitoneally (i.p.) with 1×10 7 IE9mp1-mIDO1 or IE9mp1-Empty-Vector tumor cells in a final volume of 500μl Dulbecco’s PBS (Corning Cellgro®) and treated by oral gavage with 300mg/kg IDO1 inhibitor INCB023843 (Incyte Corp.) in 200ul 0.5% methylcellulose, i.p. injection 5 days on and two days off from d7-d21, with or without (i) NAMPT inhibitor FK866 (Sigma-Aldrich, 500 μg per mouse i.p.) once every week for 3 weeks (on day 13, 20, 27); (ii) purinergic receptor A2a or A2b antagonists (Sch 58261 from Sigma-Aldrich or PSB 1115 from Tocris Bioscience, 1mg/kg, i.p. 5 days on two days off from d7-d21).

Techniques: Injection

Journal: JCI Insight

Article Title: Coordination of ENT2-dependent adenosine transport and signaling dampens mucosal inflammation

doi: 10.1172/jci.insight.121521

Figure Lengend Snippet: (A) Sex-, age-, and weight-matched Ent2-deficient mice (Ent2–/–) or WT controls (Ent2+/+, mice on a B6/129 background) were exposed to DSS for 6 days, followed by water for 24 hours. Colonic lavage was performed on terminally anesthetized mice with nucleoside preserving cocktail. Adenosine concentration in the lavage fluid was determined by HPLC and normalized to protein content. Results are shown as mean ± SEM, n = 6 mice/group, and are representative of 2 independent experiments. (B–D) Sex-, age-, and weight-matched Ent2-deficient mice (Ent2–/–, mice on a B6/129 background) were treated with an A2B receptor–specific antagonist (PSB 1115, 1 mg/kg/mouse, oral gavage) on days –1, 0, 2, 4, and 6 of DSS colitis. (B) Mice were weighed daily. Results are presented as percentage of bodyweight on day 0. (C) Histological analysis of whole colon harvested on day 7 after DSS. Scores were provided by a pathologist blinded to the groups and the study. (D) Representative histological sections from whole colon harvested on day 7 after DSS (Bar represents 100 μm; images acquired at ×10). Results are presented as mean ± SEM, n = 7–8 mice/treatment group, from 1 independent experiment. Two-way ANOVA with post hoc Bonferroni’s t test was used to determine statistical weight change; in all other cases, paired Student’s t test was used. *P < 0.05.

Article Snippet: A2B receptor antagonist (PSB 1115: 1 mg/kg, Tocris Bioscience) or vehicle (sterile water) was administered by oral gavage 1 day prior to DSS and once on days 0, 2, 4, and 6 of DSS.

Techniques: Preserving, Concentration Assay

Journal: JCI Insight

Article Title: Coordination of ENT2-dependent adenosine transport and signaling dampens mucosal inflammation

doi: 10.1172/jci.insight.121521

Figure Lengend Snippet: Matched mice with (A) Ent2 deletion on the intestinal epithelium (Ent2fl/flVillinCre+) or (B) A2B receptor deletion on the intestinal epithelium (Adora2bfl/flVillinCre+) were treated with soluflazine (Ent2 inhibitor, 7.7 μg/kg, Alzet pump) or vehicle 1 day prior to exposure to DSS or water. Mice were weighed, and results are presented as percentage of body weight on day 0. Two-way ANOVA with post hoc Bonferroni’s t test was used to determine statistical weight change. n = 6–8 mice/group from 1 independent experiment. (C) Schematic of functional consequence of Ent2 inhibition in colitis. Intestinal inflammation as observed in IBD is associated with decreased intestinal epithelial Ent2 expression, which leads to increased extracellular adenosine that can signal through the epithelial A2B adenosine receptor to protect the mucosal barrier.

Article Snippet: A2B receptor antagonist (PSB 1115: 1 mg/kg, Tocris Bioscience) or vehicle (sterile water) was administered by oral gavage 1 day prior to DSS and once on days 0, 2, 4, and 6 of DSS.

Techniques: Functional Assay, Inhibition, Expressing