G protein-coupled receptor (GPCR) signaling in the mouse and human gastrointestinal (GI) tract.
Lead Research Organisation:
King's College London
Department Name: Wolfson Centre for Age Related Diseases
Abstract
AIM: To establish the mechanisms of GPR35 signaling in the mouse and human GI tract.
REMIT: This proposal falls within the BBSRC strategy 'Bioscience for Health', and remit of 'Food, nutrition & health'.
BACKGROUND: GPR35 is an orphan receptor that lacks an endogenous ligand but is highly expressed in mammalian GI tract, particularly in the colon, and is implicated in the onset of inflammatory bowel disease (IBD). Without selective agonists GPR35's mechanisms of action remain obscure. Some studies suggest that aromatic acids derived from microbial metabolism activate GPR35, implicating a microbial sensor role for the GPCR. At high doses the clinically validated drug, sodium chromoglycate also activates GPR35 and appears to protect GI mucosa from inflammation. GPR35 is therefore a high priority for our partner Heptares Therapeutics Ltd, who have synthesised a range of structurally diverse, potent GPR35 agonists. We will evaluate their efficacies alongside chromoglycate and the aromatic acid, kynurenic acid (KA) a microbial metabolite of tryptophan. Elucidation of specific GPR35 signaling in healthy GI tissues will inform investigations of inflamed mucosae and the receptor's protective potential in models of IBD. To this end we will include a mild colitis model refined in mice during a recent BBSRC Sparking Impact project (to Cox) with alternate models that most closely recapitulate human IBD, collectively tackling an unmet need for improved translational biology.
PLAN
Yr 1 Cellular GPR35 signaling in epithelia, enteric nerves, enteroendocrine or mast cells will be investigated using proven strategies in murine and human colon. The student will compare the efficacies of novel GPR35 agonists, chromoglycate and KA in normal colonic mucosae using established methods (Cox lab). Determining whether mucosal responses are luminally or basally-directed will be important for predicting future formulation, delivery and exposure effects to support work in Yr 2-4. In vivo transit assays will establish if i.p. or oral drug alters GI transit in normal mice.
Yr 2 At Heptares the student will utilise mutated GPR35 (generated at Heptares) to establish a high confidence homology model to elucidate the molecular mechanisms of ligand binding and signaling. Autoradiographic analysis of ex vivo GI tissue will identify the cells targetted by oral or i.p. [3H]-GPR35 ligand, complementing Yr 1 functional studies. Inflamed GI tissues from IBD models e.g. dextran sulphate sodium, or oxazolone-induced colitis, will be assessed functionally (at KCL) and for GPR35 expression and other key genes implicated in mucosal changes that most closely recapitulate those in human IBD.
Yr 3-4 Using chosen IBD models the student (in KCL) will treat mice with GPR35 ligand, monitoring the onset of GI inflammation by measuring epithelial integrity functionally and histologically (scoring the degree of inflammation and stool consistency) and comparing murine mucosal erosion and dysplasia with that in human IBD colon. Optimised GPR35 ligand dosing (oral or i.p.) will test whether administered drug protects against a given inflammatory challenge and maintains mucosal health and normal rates of GI and colonic transit. Finally, the student will integrate their results into a predictive model of GPR35 signaling, i) common to healthy mouse and human tissue, ii) in inflamed colon and in-so-doing determine whether, and hopefully where, GPR35 is a therapeutic target for treating IBD.
REMIT: This proposal falls within the BBSRC strategy 'Bioscience for Health', and remit of 'Food, nutrition & health'.
BACKGROUND: GPR35 is an orphan receptor that lacks an endogenous ligand but is highly expressed in mammalian GI tract, particularly in the colon, and is implicated in the onset of inflammatory bowel disease (IBD). Without selective agonists GPR35's mechanisms of action remain obscure. Some studies suggest that aromatic acids derived from microbial metabolism activate GPR35, implicating a microbial sensor role for the GPCR. At high doses the clinically validated drug, sodium chromoglycate also activates GPR35 and appears to protect GI mucosa from inflammation. GPR35 is therefore a high priority for our partner Heptares Therapeutics Ltd, who have synthesised a range of structurally diverse, potent GPR35 agonists. We will evaluate their efficacies alongside chromoglycate and the aromatic acid, kynurenic acid (KA) a microbial metabolite of tryptophan. Elucidation of specific GPR35 signaling in healthy GI tissues will inform investigations of inflamed mucosae and the receptor's protective potential in models of IBD. To this end we will include a mild colitis model refined in mice during a recent BBSRC Sparking Impact project (to Cox) with alternate models that most closely recapitulate human IBD, collectively tackling an unmet need for improved translational biology.
PLAN
Yr 1 Cellular GPR35 signaling in epithelia, enteric nerves, enteroendocrine or mast cells will be investigated using proven strategies in murine and human colon. The student will compare the efficacies of novel GPR35 agonists, chromoglycate and KA in normal colonic mucosae using established methods (Cox lab). Determining whether mucosal responses are luminally or basally-directed will be important for predicting future formulation, delivery and exposure effects to support work in Yr 2-4. In vivo transit assays will establish if i.p. or oral drug alters GI transit in normal mice.
Yr 2 At Heptares the student will utilise mutated GPR35 (generated at Heptares) to establish a high confidence homology model to elucidate the molecular mechanisms of ligand binding and signaling. Autoradiographic analysis of ex vivo GI tissue will identify the cells targetted by oral or i.p. [3H]-GPR35 ligand, complementing Yr 1 functional studies. Inflamed GI tissues from IBD models e.g. dextran sulphate sodium, or oxazolone-induced colitis, will be assessed functionally (at KCL) and for GPR35 expression and other key genes implicated in mucosal changes that most closely recapitulate those in human IBD.
Yr 3-4 Using chosen IBD models the student (in KCL) will treat mice with GPR35 ligand, monitoring the onset of GI inflammation by measuring epithelial integrity functionally and histologically (scoring the degree of inflammation and stool consistency) and comparing murine mucosal erosion and dysplasia with that in human IBD colon. Optimised GPR35 ligand dosing (oral or i.p.) will test whether administered drug protects against a given inflammatory challenge and maintains mucosal health and normal rates of GI and colonic transit. Finally, the student will integrate their results into a predictive model of GPR35 signaling, i) common to healthy mouse and human tissue, ii) in inflamed colon and in-so-doing determine whether, and hopefully where, GPR35 is a therapeutic target for treating IBD.
