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To discover a novel series of potent inhibitors of enteropeptidase, a membrane-bound serine protease localized to the duodenal brush border, 4-guanidinobenzoate derivatives were evaluated with minimal systemic exposure. The 1cdocking model enabled the installation of an additional carboxylic acid moiety to obtain an extra interaction with enteropeptidase, yielding 2a. The oral administration of 2asignificantly elevated the fecal protein output, a pharmacodynamic marker, in diet-induced obese (DIO) mice, whereas subcutaneous administration did not change this parameter. Thus, systemic exposure of 2awas not required for its pharmacological effects. Further optimization focusing on the in vitro IC50value and T1/2, an indicator of dissociation time, followed by enhanced in vivo pharmacological activity based on the ester stability of the compounds, revealed two series of potent enteropeptidase inhibitors, a dihydrobenzofuran analogue ((S)-5b, SCO-792) and phenylisoxazoline (6b), which exhibited potent anti-obesity effects despite their low systemic exposure following their oral administration to DIO rats.

Magnetic resonance imaging (MRI) and MRI‐based elastography (MRE) are the most promising noninvasive techniques in assessing liver diseases. The purpose of this study was to evaluate an advanced multiparametric imaging method for staging disease and assessing treatment response in realistic preclinical alcohol‐associated liver disease (ALD). We utilized four different preclinical mouse models in our study: Model 1—mice were fed a fast‐food diet and fructose water for 48 weeks to induce nonalcoholic fatty liver disease; Model 2—mice were fed chronic‐binge ethanol (EtOH) for 10 days or 8 weeks to induce liver steatosis/inflammation. Two groups of mice were treated with interleukin‐22 at different time points to induce disease regression; Model 3—mice were administered CCl4for 2 to 4 weeks to establish liver fibrosis followed by 2 or 4 weeks of recovery; and Model 4—mice were administered EtOH plus CCl4for 12 weeks. Mouse liver imaging biomarkers including proton density fat fraction (PDFF), liver stiffness (LS), loss modulus (LM), and damping ratio (DR) were assessed. Liver and serum samples were obtained for histologic and biochemical analyses. Ordinal logistic regression and generalized linear regression analyses were used to model the severity of steatosis, inflammation, and fibrosis, and to assess the regression of these conditions. Multiparametric models with combinations of biomarkers (LS, LM, DR, and PDFF) used noninvasively to predict the histologic severity and regression of steatosis, inflammation, and fibrosis were highly accurate (area under the curve > 0.84 for all). A three‐parameter model that incorporates LS, DR, and ALT predicted histologic fibrosis progression (r= 0.84, p< 0.0001) and regression (r= 0.79, p< 0.0001) as measured by collagen content in livers. This preclinical study provides evidence that multiparametric MRI/MRE can be used noninvasively to assess disease severity and monitor treatment response in ALD. Non‐contrast hepatic magnetic resonance imaging (MRI) and magnetic resonance elastography (MRE) can provide a composite metric to discriminate different steatosis grades, inflammation grades, and fibrosis stages in alcohol‐related liver disease (ALD) with high accuracies (AUCs all > 0.80), respectively. Combined with serum biomarkers, multiparametric hepatic MRE can assess the subtle changes of liver collagen content in fibrosis progression and regression. In summary, multiparametric MRI/MRE has the potentials to assess the severity of ALD and monitor response to treatment.

G protein-coupled receptor 40 (GPR40)/free fatty acid 1 (FFA1) is a G protein-coupled receptor involved in free fatty acid-induced insulin secretion. To analyze the effect of our novel GPR40/FFA1-selective agonist, [(3S)-6-({2′,6′-dimethyl-4′-[3-(methylsulfonyl)propoxy]biphenyl-3-yl}methoxy)-2,3-dihydro-1-benzofuran-3-yl]acetic acid hemi-hydrate (TAK-875), on insulin and glucagon secretion, we performed hormone secretion assays and measured intracellular Ca2+concentration ([Ca2+]i) in both human and rat islets. Insulin and glucagon secretion were measured in static and dynamic conditions by using groups of isolated rat and human pancreatic islets. [Ca2+]iwas recorded by using confocal microscopy. GPR40/FFA1 expression was measured by quantitative polymerase chain reaction. In both human and rat islets, TAK-875 enhanced glucose-induced insulin secretion in a glucose-dependent manner. The stimulatory effect of TAK-875 was similar to that produced by glucagon-like peptide-1 and correlated with the elevation of β-cell [Ca2+]i. TAK-875 was without effect on glucagon secretion at both 1 and 16 mM glucose in human islets. These data indicate that GPR40/FFA1 influences mainly insulin secretion in a glucose-dependent manner. The β-cell-specific action of TAK-875 in human islets may represent a therapeutically useful feature that allows plasma glucose control without compromising counter-regulation of glucagon secretion, thus minimizing the risk of hypoglycemia.

G protein-coupled receptor 40 (GPR40)/free fatty acid 1 (FFA1) is a G protein-coupled receptor involved in free fatty acid-induced insulin secretion. To analyze the effect of our novel GPR40/FFA1-selective agonist, [(3S)-6-({2',6'-dimethyl-4'-[3-(methylsulfonyl)propoxy]biphenyl-3-yl}methoxy)-2,3-dihydro-1-benzofuran-3-yl]acetic acid hemi-hydrate (TAK-875), on insulin and glucagon secretion, we performed hormone secretion assays and measured intracellular Ca²⁺ concentration ([Ca²⁺](i)) in both human and rat islets. Insulin and glucagon secretion were measured in static and dynamic conditions by using groups of isolated rat and human pancreatic islets. [Ca²⁺](i) was recorded by using confocal microscopy. GPR40/FFA1 expression was measured by quantitative polymerase chain reaction. In both human and rat islets, TAK-875 enhanced glucose-induced insulin secretion in a glucose-dependent manner. The stimulatory effect of TAK-875 was similar to that produced by glucagon-like peptide-1 and correlated with the elevation of β-cell [Ca²⁺](i). TAK-875 was without effect on glucagon secretion at both 1 and 16 mM glucose in human islets. These data indicate that GPR40/FFA1 influences mainly insulin secretion in a glucose-dependent manner. The β-cell-specific action of TAK-875 in human islets may represent a therapeutically useful feature that allows plasma glucose control without compromising counter-regulation of glucagon secretion, thus minimizing the risk of hypoglycemia.

Cibenzoline, (±)‐2‐(2,2‐diphenylcyclopropyl‐2‐imidazoline succinate, has been clinically used as one of the Class I type antiarrhythmic agents and also reported to block ATP‐sensitive K+channels in excised membranes from heart and pancreatic β cells. In the present study, we investigated if this drug inhibited gastric H+,K+‐ATPase activity in vitro.Cibenzoline inhibited H+,K+‐ATPase activity of permeabilized leaky hog gastric vesicles in a concentration‐dependent manner (IC50: 201 μM), whereas no effect was shown on Na+,K+‐ATPase activity of dog kidney (IC50: >1000 μM). Similarly, cibenzoline inhibited H+,K+‐ATPase activity of HEK‐293 cells (human embryonic kidney cell line) co‐transfected with rabbit gastric H+,K+‐ATPase α‐ and β‐subunit cDNAs (IC50: 183 μM).In leaky gastric vesicles, inhibition of H+,K+‐ATPase activity by cibenzoline was attenuated by the addition of K+(0.5 – 5 mM) in a concentration‐dependent manner. The Lineweaver‐Burk plot of the H+,K+‐ATPase activity shows that cibenzoline increases Kmvalue for K+without affecting Vmax, indicating that this drug inhibits H+,K+‐ATPase activity competitively with respect to K+.The inhibitory effect of H+,K+‐ATPase activity by cibenzoline with normal tight gastric vesicles did not significantly differ from that with permeabilized leaky gastric vesicles, indicating that this drug reacted to the ATPase from the cytoplasmic side of the membrane.These findings suggest that cibenzoline is an inhibitor of gastric H+,K+‐ATPase with a novel inhibition mechanism, which inhibits gastric H+,K+‐ATPase by binding its K+‐recognition site from the cytoplasmic side.