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The prostaglandin D2 receptor 1 (DP1), a member of the prostanoid G protein-coupled receptor (GPCR) family, plays critical roles in allergic responses, sleep regulation, immune modulation, and vasodilation. Here, we present five high-resolution cryo-electron microscopy (cryo-EM) structures of the human DP1 receptor, including an apo structure, two inactive state structures bound to two different inverse agonists developed by ONO Pharmaceutical, and two active state structures in complex with the G s protein and bound to the endogenous agonist PGD2 and its selective derivative BW245C. Structural analysis, complemented by molecular dynamics simulations and site-directed mutagenesis, reveals key residues involved in ligand recognition and suggests a distinct activation mechanism for DP1, which lacks most of the conserved class A GPCR motifs. Notably, the unique residue K76 within the conserved sodium pocket acts as a major activation switch, while amphiphilic helix 8 adopts an unconventional orientation essential for receptor function. These findings offer valuable insights into the structure and function of prostanoid receptors and may facilitate the development of therapeutics targeting DP1. The prostaglandin DP1 receptor plays critical roles in allergic responses, sleep regulation, immune modulation, and vasodilation. Here, authors present atomic structures of DP1 in different states revealing molecular mechanisms of receptor inhibition and activation.

Group 2 innate lymphoid cells (ILC2s) are involved in human diseases, such as allergy, atopic dermatitis and nasal polyposis, but their function in human cancer remains unclear. Here we show that, in acute promyelocytic leukaemia (APL), ILC2s are increased and hyper-activated through the interaction of CRTH2 and NKp30 with elevated tumour-derived PGD2 and B7H6, respectively. ILC2s, in turn, activate monocytic myeloid-derived suppressor cells (M-MDSCs) via IL-13 secretion. Upon treating APL with all-trans retinoic acid and achieving complete remission, the levels of PGD2, NKp30, ILC2s, IL-13 and M-MDSCs are restored. Similarly, disruption of this tumour immunosuppressive axis by specifically blocking PGD2, IL-13 and NKp30 partially restores ILC2 and M-MDSC levels and results in increased survival. Thus, using APL as a model, we uncover a tolerogenic pathway that may represent a relevant immunosuppressive, therapeutic targetable, mechanism operating in various human tumour types, as supported by our observations in prostate cancer. Group 2 innate lymphoid cells (ILC2s) modulate inflammatory and allergic responses, but their function in cancer immunity is still unclear. Here the authors show that, in acute promyelocytic leukaemia, tumour-activated ILC2s secrete IL-13 to induce myeloid-derived suppressor cells and support tumour growth.

Exosomes are bioactive vesicles released from multivesicular bodies by intact cells and participate in intercellular signalling. We investigated the presence of lipid-related proteins and bioactive lipids in RBL-2H3 exosomes. Besides a phospholipid scramblase and a Fatty Acid Binding Protein, the exosomes contained the whole set of phospholipases (A2, C and D) together with interacting proteins such as aldolase A and Hsp 70. They also contained the PLD / PAP1 pathway leading to the formation of diglycerides. RBL-2H3 exosomes also carried members of the three phospholipase A2 classes, i.e. the calcium-dependent cPLA2-IVA, the calcium-independent iPLA2-VIA and the secreted sPLA2-IIA and V. Remarkably, almost all members of the Ras GTPase superfamily were present, and incubation of exosomes with GTPgammaS triggered activation of PLA2s and PLD2. A large panel of free fatty acids, including arachidonic acid, and derivatives such as prostaglandin E2 and 15-deoxy-Delta12,14-prostaglandinJ2 were detected. We observed that the exosomes were internalized by resting and activated RBLcells, and that they accumulated in an endosomal compartment. Endosomal concentrations were in the micromolar range for prostaglandins, i.e. concentrations able to trigger prostaglandin-dependent biological responses. Therefore exosomes are carriers of GTP-activatable phospholipases and lipid mediators from cell to cell.

Proinflammatory macrophages are key mediators in several pathologies; thus, controlling their activation is necessary. The endocannabinoid system is implicated in various inflammatory processes. Here we show that in macrophages, the newly characterized enzyme α/β-hydrolase domain 6 (ABHD6) controls 2-arachidonoylglycerol (2-AG) levels and thus its pharmacological effects. Furthermore, we characterize a unique pathway mediating the effects of 2-AG through its oxygenation by cyclooxygenase-2 to give rise to the anti-inflammatory prostaglandin D ₂-glycerol ester (PGD ₂-G). Pharmacological blockade of cyclooxygenase-2 or of prostaglandin D synthase prevented the effects of increasing 2-AG levels by ABHD6 inhibition in vitro, as well as the 2-AG–induced increase in PGD ₂-G levels. Together, our data demonstrate the physiological relevance of the interaction between the endocannabinoid and prostanoid systems. Moreover, we show that ABHD6 inhibition in vivo allows for fine-tuning of 2-AG levels in mice, therefore reducing lipopolysaccharide-induced inflammation, without the characteristic central side effects of strong increases in 2-AG levels obtained following monoacylglycerol lipase inhibition. In addition, administration of PGD ₂-G reduces lipopolysaccharide-induced inflammation in mice, thus confirming the biological relevance of this 2-AG metabolite. This points to ABHD6 as an interesting therapeutic target that should be relevant in treating inflammation-related conditions, and proposes PGD ₂-G as a bioactive lipid with potential anti-inflammatory properties in vivo.

Group 2 innate lymphoid cells (ILC2s) promote type 2 cytokine-dependent immunity, inflammation, and tissue repair. Although epithelial cell-derived cytokines regulate ILC2 effector functions, the pathways that control the in vivo migration of ILC2s into inflamed tissues remain poorly understood. Here, we provide the first demonstration that expression of the prostaglandin D2 (PGD2) receptor CRTH2 (chemoattractant receptor-homologous molecule expressed on Th2 cells) regulates the in vivo accumulation of ILC2s in the lung. Although a significant proportion of ILC2s isolated from healthy human peripheral blood expressed CRTH2, a smaller proportion of ILC2s isolated from nondiseased human lung expressed CRTH2, suggesting that dynamic regulation of CRTH2 expression might be associated with the migration of ILC2s into tissues. Consistent with this, murine ILC2s expressed CRTH2, migrated toward PGD2in vitro, and accumulated in the lung in response to PGD2in vivo. Furthermore, mice deficient in CRTH2 exhibited reduced ILC2 responses and inflammation in a murine model of helminth-induced pulmonary type 2 inflammation. Critically, adoptive transfer of CRTH2-sufficient ILC2s restored pulmonary inflammation in CRTH2-deficient mice. Together, these data identify a role for the PGD2–CRTH2 pathway in regulating the in vivo accumulation of ILC2s and the development of type 2 inflammation in the lung.

Increased production of Prostaglandin D 2 (PGD 2 ) is linked to development and progression of asthma and allergy. PGD 2 is rapidly degraded to its metabolites, which initiate type 2 innate lymphoid cells (ILC2) migration and IL-5/IL-13 cytokine secretion in a PGD 2 receptor 2 (DP 2 )-dependent manner. Blockade of DP 2 has shown therapeutic benefit in subsets of asthma patients. Cellular mechanisms of ILC2 activity in response to PGD 2 and its metabolites are still unclear. We hypothesized that ILC2 respond non-uniformly to PGD 2 metabolites. ILC2s were isolated from peripheral blood of patients with atopic asthma. ILC2s were stimulated with PGD 2 and four PGD 2 metabolites (Δ 12 -PGJ 2 , Δ 12 -PGD 2 , 15-deoxyΔ 12,14 -PGD 2 , 9α,11β-PGF 2 ) with or without the selective DP 2 antagonist fevipiprant. Total RNA was sequenced, and differentially expressed genes (DEG) were identified by DeSeq2. Differential gene expression analysis revealed an upregulation of pro-inflammatory DEGs in ILC2s stimulated with PGD 2 (14 DEGs), Δ 12 -PGD 2 (27 DEGs), 15-deoxyΔ 12,14 -PGD 2 (56 DEGs) and Δ 12 -PGJ 2 (136 DEGs), but not with 9α,11β-PGF 2 . Common upregulated DEGs were i.e. ARG2, SLC43A2, LAYN, IGFLR1, or EPHX2. Inhibition of DP 2 via fevipiprant mainly resulted in downregulation of pro-inflammatory genes such as DUSP4, SPRED2, DUSP6, ETV1, ASB2, CD38, ADGRG1, DDIT4, TRPM2, or CD69. DEGs were related to migration and various immune response-relevant pathways such as “ chemokine (C-C motif) ligand 4 production ”, “ cell migration” , “interleukin-13 production” , “regulation of receptor signaling pathway via JAK-STAT” , or “lymphocyte apoptotic process” , underlining the pro-inflammatory effects of PGD 2 metabolite-induced immune responses in ILC2s as well as the anti-inflammatory effects of DP 2 inhibition via fevipiprant. Furthermore, PGD 2 and metabolites showed distinct profiles in ILC2 activation. Overall, these results expand our understanding of DP 2 initiated ILC2 activity.

There is a wealth of information available about endogenous fatty acid ligands for peroxisome proliferator-activated receptor α/δ/γ (PPARα/δ/γ); however, there are few comparative studies of PPARα/δ/γ activation using standardized experimental systems. This study investigated which of 14 major free long-chain fatty acids (LCFAs: C12:0-C22:6) and 15-deoxy-Δ -prostaglandin J2 (15d-PGJ2) activate PPARα/δ/γ using a coactivator recruitment assay. We recently discovered that eight different synthetic PPAR agonists recruit four different coactivator peptides (PGC1α, CBP, SRC1, TRAP220) with varying potency and efficacy, so we examined the ligand-concentration-dependent recruitment of these four coactivators. All 15 fatty acids (FAs) activated PPARα/δ at high concentrations, but only palmitic acid, stearic acid, oleic acid, and linoleic acid significantly activated PPARα/δ at physiologically relevant concentrations. Lauric acid, myristic acid, palmitic acid, and 15d-PGJ2 activated PPARγ at high concentrations, but only palmitic acid slightly activated PPARγ at physiologically relevant concentrations. FA ligands exhibited different coactivator preference compared to synthetic PPAR agonists, including approved drugs such as pemafibrate, seladelpar, and pioglitazone, suggesting that these agonists may regulate target gene transcription in a different manner than natural FA ligands. Such differences may be relevant to the pathogenesis of side effects of synthetic PPAR agonists occasionally observed in (pre)clinical studies.

20-carbon fatty acid-derived eicosanoids are versatile signaling oxylipins in mammals. In particular, a group of eicosanoids termed prostanoids are involved in multiple physiological processes, such as reproduction and immune responses. Although some eicosanoids such as prostaglandin E2 (PGE2) have been detected in some insect species, molecular mechanisms of eicosanoid synthesis and signal transduction in insects have not been thoroughly investigated. Our phylogenetic analysis indicated that, in clear contrast to the presence of numerous receptors for oxylipins and other lipid mediators in humans, the Drosophila genome only possesses a single ortholog of such receptors, which is homologous to human prostanoid receptors. This G protein-coupled receptor, named Prostaglandin Receptor or PGR, is activated by PGE2 and its isomer PGD2 in Drosophila S2 cells. PGR mutant flies die as pharate adults with insufficient tracheal development, which can be rescued by supplying high oxygen. Consistent with this, through a comprehensive mutagenesis approach, we identified a Drosophila PGE synthase whose mutants show similar pharate adult lethality with hypoxia responses. Drosophila thus has a highly simplified eicosanoid signaling pathway as compared to humans, and it may provide an ideal model system for investigating evolutionarily conserved aspects of eicosanoid signaling.

NSAIDs that inhibit cyclooxygenase-1 can provoke severe asthma and rhinosinusitis with nasal polyps and eosinophil infiltration. Cysteinyl leukotriene generation by the leukotriene C4 synthase pathway may cause the bronchoconstriction, vascular leak, and mucous secretion. The Clinical Implications of Basic Research series has focused on highlighting laboratory research that could lead to advances in clinical therapeutics. However, the path between the laboratory and the bedside runs both ways: clinical observations often pose new questions for laboratory investigations that then lead back to the clinic. One of a series of occasional articles drawing attention to the bedside-to-bench flow of information is presented here, under the Basic Implications of Clinical Observations rubric. We hope our readers will enjoy these stories of discovery, and we invite them to submit their own examples of clinical findings that have led . . .

Prostaglandin D2 (PGD2) signals via the DP1 and DP2 receptors. In Phase II trials, DP2 antagonism decreased airway inflammation and airway smooth muscle (ASM) area in moderate-to-severe asthma patients. However, in Phase III, DP2 antagonism failed to lower the rate of exacerbations, and DP2 as a target was shelved. Here, using a preclinical model of chronic experimental asthma, we demonstrate that rhinovirus-induced exacerbations increase PGD2 release, mucus production, transforming growth factor (TGF)-β1 and type-2 inflammation. DP2 antagonism or DP1 agonism ablates these phenotypes, increases epithelial EGF expression and decreases ASM area via increased IFN-γ. In contrast, dual DP1-DP2 antagonism or dual corticosteroid/DP2 antagonism, which attenuates endogenous PGD2, prevented ASM resolution. We demonstrate that DP2 antagonism resolves ASM remodelling via PGD2/DP1-mediated upregulation of IFN-γ expression, and that dual DP2 antagonism/corticosteroid therapy, as often occurred in the human trials, impairs the efficacy of DP2 antagonism by suppressing endogenous PGD2 and IFN-γ production. Antagonists of the DP2 receptor, activated by prostaglandin D2, showed clinical efficacy in Phase II trials but not in the reduction of asthma exacerbations in Phase III trials. Here the authors use a pre-clinical model of chronic experimental asthma to show that corticosteroids suppress endogenous PGD2 and IFN-γ production, necessary for the beneficial effects of DP2 antagonism, and hence dual therapy with corticosteroids was less effective than DP2 antagonism alone at resolving airway inflammation and airway remodelling.