A Systems Model for Ursodeoxycholic Acid Metabolism in Healthy and Patients With Primary Biliary Cirrhosis.

A systems model was developed to describe the metabolism and disposition of ursodeoxycholic acid (UDCA) and its conjugates in healthy subjects based on pharmacokinetic (PK) data from published studies in order to study the distribution of oral UDCA and potential interactions influencing therapeutic effects upon interruption of its enterohepatic recirculation. The base model was empirically adapted to patients with primary biliary cirrhosis (PBC) based on current understanding of disease pathophysiology and clinical measurements. Simulations were performed for patients with PBC under two competing hypotheses: one for inhibition of ileal absorption of both UDCA and conjugates and the other only of conjugates. The simulations predicted distinctly different bile acid distribution patterns in plasma and bile. The UDCA model adapted to patients with PBC provides a platform to investigate a complex therapeutic drug interaction among UDCA, UDCA conjugates, and inhibition of ileal bile acid transport in this rare disease population.

GSK256073, a selective agonist of G-protein coupled receptor 109A (GPR109A) reduces serum glucose in subjects with type 2 diabetes mellitus.

AIMS: This clinical trial assessed whether a potent, selective GPR109A agonist, GSK256073, could, through inhibition of lipolysis, acutely improve glucose homeostasis in subjects with type 2 diabetes mellitus. METHODS: Thirty-nine diabetic subjects were enrolled in the randomized, single-blind, placebo-controlled, three-period crossover trial. Each subject received placebo and two of four regimens of GSK256073 for 2 days. GSK256073 was dosed 5 mg every 12 h before breakfast and supper (BID), 10 mg every 24 h before breakfast (QD), 25 mg BID and 50 mg QD. RESULTS: The change from baseline weighted mean glucose concentration for an interval from 24 to 48 h after the initial drug dose was significantly reduced for all GSK256073 regimens, reaching a maximum of -0.87 mmol/l (-1.20, -0.52) with the 25 mg BID dose. Sustained suppression of non-esterified fatty acid (NEFA) and glycerol concentrations was observed with all GSK256073 doses throughout the 48-h dosing period. Serum insulin and C-peptide concentrations fell in concert with glucose concentrations and calculated HOMA-IR scores decreased 27-47%, consistent with insulin sensitization. No marked differences were evident between either 10 and 50 mg total daily doses or QD versus BID dosing. CONCLUSIONS: Administration of a GPR109A agonist for 2 days significantly decreased serum NEFA and glucose concentrations in diabetic subjects. Glucose improvements were associated with decreased insulin concentrations and measures of enhanced insulin sensitivity. Improved glucose control occurred with GSK256073 doses that were generally safe and not associated with events of flushing or gastrointestinal disturbances.

Remogliflozin etabonate, a selective inhibitor of the sodium-dependent transporter 2 reduces serum glucose in type 2 diabetes mellitus patients.

AIMS: Remogliflozin etabonate (RE) is the pro-drug of remogliflozin (R), a selective inhibitor of renal sodium-dependent glucose transporter 2 (SGLT2) that improves glucose control via enhanced urinary glucose excretion (UGE). This study evaluated the safety, tolerability, pharmacokinetics and pharmacodynamics of repeated doses of RE in subjects with type 2 diabetes mellitus (T2DM). METHODS: In a double-blinded, randomized, placebo-controlled trial, subjects who were drug-naïve or had metformin discontinued received RE [100 mg BID (n = 9), 1000 mg QD (n = 9), 1000 mg BID (n = 9)], or placebo (n = 8) for 12 days. Safety parameters were assessed, including urine studies to evaluate renal function. Plasma concentrations of RE and metabolites were measured with the first dose and at steady state. RE effects on glucose levels were assessed with fasting glucose concentrations, frequently sampled 24-h glucose profiles and oral glucose tolerance tests. RESULTS: No significant laboratory abnormalities or safety events were reported; the most frequent adverse events were headache and flatulence. Plasma exposure to RE and R were proportional to administered dose with negligible accumulation. Mean 24-h UGE increased in RE treatment groups. Compared with the placebo group, 24-h mean (95% CI) changes in plasma glucose were -1.2 (-2.2 to -0.3) (100 mg BID), -0.8 (-1.7 to 0.2) (1000 mg QD) and -1.7 (-2.7 to -0.8) mmol/l (1000 mg BID). CONCLUSIONS: Administration of RE for 12 days is well-tolerated and results in clinically meaningful improvements in plasma glucose, accompanied by changes in body weight and blood pressure in subjects with T2DM.

Safety, tolerability, pharmacodynamics and pharmacokinetics of albiglutide, a long-acting glucagon-like peptide-1 mimetic, in healthy subjects.

AIMS: Albiglutide is a glucagon-like peptide-1 (GLP-1) mimetic generated by genetic fusion of a dipeptidyl peptidase-IV-resistant GLP-1 dimer to human albumin. Albiglutide was designed to retain the therapeutic effects of native GLP-1 while extending its duration of action. This study was conducted to determine the pharmacokinetics and initial safety/tolerability profile of albiglutide in non-diabetic volunteers. METHODS: In this single-blind, randomized, placebo-controlled trial, 39 subjects (18-60 years, body mass index 19.9-35.0 kg/m(2)) received placebo (n = 10) or escalating doses of albiglutide (n = 29) on days 1 and 8 in the following sequential cohorts: cohort 1: 0.25 + 1 mg; cohort 2: 3 + 6 mg; cohort 3: 16 + 24 mg; cohort 4: 48 + 60 mg; and cohort 5: 80 + 104 mg. Dose proportionality was evaluated based on area under the plasma drug concentration versus time curve [area under the curve (AUC((0-7 days)))] and maximum plasma drug concentration (C(max)) for cohorts 2-5 during week 1. RESULTS: Albiglutide had a terminal elimination half-life (T(1/2)) of 6-8 days and time to maximum observed plasma drug concentration (T(max)) of 3-4 days. A greater-than-dose proportional increase in albiglutide exposure was observed. Albiglutide demonstrated a dose-dependent trend in reductions of glucose weighted mean AUC and fructosamine levels in healthy subjects. The incidence and severity of adverse events (AEs) was similar between placebo and albiglutide groups. Headache was the most frequent drug-related AE, followed by constipation, flatulence and nausea. CONCLUSIONS: Albiglutide has a half-life that favours once weekly or less frequent dosing with an acceptable safety/tolerability profile in non-diabetic subjects.

Prolonged inhibition of muscle carnitine palmitoyltransferase-1 promotes intramyocellular lipid accumulation and insulin resistance in rats.

Cross-sectional studies in human subjects have used 1H magnetic resonance spectroscopy (HMRS) to demonstrate that insulin resistance correlates more tightly with the intramyocellular lipid (IMCL) concentration than with any other identified risk factor. To further explore the interaction between these two elements in the rat, we used two strategies to promote the storage of lipids in skeletal muscle and then evaluated subsequent changes in insulin-mediated glucose disposal. Normal rats received either a low-fat or a high-fat diet (20% lard oil) for 4 weeks. Two additional groups (lowfat + etoxomir and lard + etoxomir) consumed diets containing 0.01% of the carnitine palmitoyltransferase-1 inhibitor, R-etomoxir, which produced chronic blockade of enzyme activity in liver and skeletal muscle. Both the high-fat diet and drug treatment significantly impaired insulin sensitivity, as measured with the hyperinsulinemic-euglycemic clamp. Insulin-mediated glucose disposal (IMGD) fell from 12.57 +/- 0.72 in the low-fat group to 9.79 +/- 0.59, 8.96 +/- 0.38, and 7.32 +/- 0.28 micromol x min(-1) x 100 g(-1) in the low-fat + etoxomir, lard, and lard + etoxomir groups, respectively. We used HMRS, which distinguishes between fat within the myocytes and fat associated with contaminating adipocytes located in the muscle bed, to assess the IMCL content of isolated soleus muscle. A tight inverse relationship was found between IMGD and IMCL, the correlation (R = 0.96) being much stronger than that seen between IMGD and either fat mass or weight. In conclusion, either a diet rich in saturated fat or prolonged inhibition of fatty acid oxidation impairs IMGD in rats via a mechanism related to the accumulation of IMCL.

Measurement of intracellular triglyceride stores by H spectroscopy: validation in vivo.

We validate the use of 1H magnetic resonance spectroscopy (MRS) to quantitatively differentiate between adipocyte and intracellular triglyceride (TG) stores by monitoring the TG methylene proton signals at 1.6 and 1.4 ppm, respectively. In two animal models of intracellular TG accumulation, intrahepatic and intramyocellular TG accumulation was confirmed histologically. Consistent with the histological changes, the methylene signal intensity at 1.4 ppm increased in both liver and muscle, whereas the signal at 1.6 ppm was unchanged. In response to induced fat accumulation, the TG concentration in liver derived from 1H MRS increased from 0 to 44.9 +/- 13.2 micromol/g, and this was matched by increases measured biochemically (2.1 +/- 1.1 to 46.1 +/- 10.9 micromol/g). Supportive evidence that the methylene signal at 1.6 ppm in muscle is derived from investing interfascial adipose tissue was the finding that, in four subjects with generalized lipodystrophy, a disease characterized by absence of interfacial fat, no signal was detected at 1.6 ppm; however, a strong signal was seen at 1.4 ppm. An identical methylene chemical shift at 1.4 ppm was obtained in human subjects with fatty liver where the fat is located exclusively within hepatocytes. In experimental animals, there was a close correlation between hepatic TG content measured in vivo by 1H MRS and chemically by liver biopsy [R = 0.934; P <.0001; slope 0.98, confidence interval (CI) 0.70-1.17; y-intercept 0.26, CI -0.28 to 0. 70]. When applied to human calf muscle, the coefficient of variation of the technique in measuring intramyocellular TG content was 11.8% in nonobese subjects and 7.9% in obese subjects and of extramyocellular (adipocyte) fat was 22.6 and 52.5%, respectively. This study demonstrates for the first time that noninvasive in vivo 1H MRS measurement of intracellular TG, including that within myocytes, is feasible at 1.5-T field strengths and is comparable in accuracy to biochemical measurement. In addition, in mixed tissue such as muscle, the method is clearly advantageous in differentiating between TG from contaminating adipose tissue compared with intramyocellular lipids.

Circulating fatty acids are essential for efficient glucose-stimulated insulin secretion after prolonged fasting in humans.

In the fasted rat, efficient glucose-stimulated insulin secretion (GSIS) is absolutely dependent on an elevated level of circulating free fatty acids (FFAs). To determine if this is also true in humans, nonobese volunteers were fasted for 24 h (n = 5) or 48 h (n = 5), after which they received an infusion of either saline or nicotinic acid (NA) to deplete their plasma FFA pool, followed by an intravenous bolus of glucose. NA treatment resulted in a fall in basal insulin concentrations of 35 and 45% and in the area under the insulin response curve (area under the curve [AUC]) to glucose of 47 and 42% in the 24- and 48-h fasted individuals, respectively. The 48-h fasted subjects underwent the same procedure with the addition of a coinfusion of Intralipid plus heparin (together with NA) to maintain a high concentration of plasma FFAs throughout the study. The basal level and AUC for insulin were now completely normalized (C-peptide profiles paralleled those for insulin). To assess the effect of an overnight fast, nonobese (n = 6) and obese (n = 6) subjects received an infusion of either saline or NA, followed by a hyperglycemic clamp (200 mg/dl). The insulin AUC in response to glucose was unaffected by lowering of the FFA level in nonobese subjects, but fell by 29% in the obese group. The data clearly demonstrate that in humans, the rise in circulating FFA levels after 24 and 48 h of food deprivation is critically important for pancreatic beta-cell function both basally and during subsequent glucose loading. They also suggest that the enhancement of GSIS by FFAs in obese individuals is more prominent than that seen in their nonobese counterparts.

A fatty acid- dependent step is critically important for both glucose- and non-glucose-stimulated insulin secretion.

Lowering of the plasma FFA level in intact fasted rats by infusion of nicotinic acid (NA) caused essentially complete ablation of insulin secretion (IS) in response to a subsequent intravenous bolus of arginine, leucine, or glibenclamide (as previously found using glucose as the beta-cell stimulus). However, in all cases, IS became supranormal when a high FFA level was maintained by co-infusion of lard oil plus heparin. Each of these secretagogues elicited little, if any, IS from the isolated, perfused "fasted" pancreas when tested simply on the background of 3 mM glucose, but all became extremely potent when 0.5 mM palmitate was also included in the medium. Similarly, IS from the perfused pancreas, in response to depolarizing concentrations of KCl, was markedly potentiated by palmitate. As was the case with intravenous glucose administration, fed animals produced an equally robust insulin response to glibenclamide regardless of whether their low basal FFA concentration was further reduced by NA. In the fasted state, arginine-induced glucagon secretion appeared to be independent of the prevailing FFA concentration. The findings establish that the essential role of circulating FFA for glucose-stimulated IS after food deprivation also applies in the case of nonglucose secretagogues. In addition, they imply that (i) a fatty acid-derived lipid moiety, which plays a pivotal role in IS, is lost from the pancreatic beta-cell during fasting; (ii) in the fasted state, the elevated level of plasma FFA compensates for this deficit; and (iii) the lipid factor acts at a late step in the insulin secretory pathway that is common to the action of a wide variety of secretagogues.

Rates of glucagon activation and deactivation of hepatic glucose production in conscious dogs.

To determine the time course of glucagon activation and deactivation of hepatic glucose production (HGP), studies were conducted in 18-hour fasted, conscious dogs. Somatostatin was infused with insulin replaced intraportally at 1.8 pmol x kg(-1) x min(-1) and glucagon replaced peripherally at 1.0 ng x kg(-1) x min(-1). After a 2-hour control period, glucagon infusion was either (1) increased fourfold for 4 hours (GGN 4X), (2) increased fourfold for 30 minutes and returned to a basal rate for 3.5 hours (GGN 4X/1X), or (3) fixed at the basal rate for 4 hours (GGN 1X). In the latter two protocols, glucose was infused peripherally to match glucose concentrations observed during GGN 4X. Glucose turnover was determined by deconvolution with the impulse response of the glucose system described by a two-compartment, time-varying model identified from high-performance liquid chromatography (HPLC)-purified [3-3H]glucose tracer data. In GGN 4X, HGP was stimulated from 15.2 +/- 0.9 micromol x kg(-1) x min(-1) to 52.7 +/- 6.5 micromol x kg(-1) x min(-1) after just 15 minutes, but it decreased over the subsequent 3 hours to a rate 25% above basal. In GGN 4X/1X, the increase in HGP during the first 30 minutes equaled that observed in GGN 4X, but when glucagon infusion was returned to basal, HGP decreased in 15 minutes to rates equal to those observed in GGN 1X. The times for half-maximal activation and deactivation of glucagon action were equal (4.5 +/- 1.0 and 4.0 +/- 1.1 minutes, respectively). The very rapid and sensitive hepatic response to glucagon makes pancreatic glucagon release a key component of minute-to-minute glucose homeostasis.

Counterregulation by epinephrine and glucagon during insulin-induced hypoglycemia in the conscious dog.

We assessed the combined role of epinephrine and glucagon in regulating gluconeogenic precursor metabolism during insulin-induced hypoglycemia in the overnight-fasted, adrenalectomized, conscious dog. In paired studies (n = 5), insulin was infused intraportally at 5 mU.kg-1.min-1 for 3 h. Epinephrine was infused at a basal rate (B-EPI) or variable rate to simulate the normal epinephrine response to hypoglycemia (H-EPI), whereas in both groups the hypoglycemia-induced rise in cortisol was simulated by cortisol infusion. Plasma glucose fell to approximately 42 mg/dl in both groups. Glucagon failed to rise in B-EPI, but increased normally in H-EPI. Hepatic glucose release fell in B-EPI but increased in H-EPI. In B-EPI, the normal rise in lactate levels and net hepatic lactate uptake was prevented. Alanine and glycerol metabolism were similar in both groups. Since glucagon plays little role in regulating gluconeogenic precursor metabolism during 3 h of insulin-induced hypoglycemia, epinephrine must be responsible for increasing lactate release from muscle, but is minimally involved in the lipolytic response. In conclusion, a normal rise in epinephrine appears to be required to elicit an increase in glucagon during insulin-induced hypoglycemia in the dog. During insulin-induced hypoglycemia, epinephrine plays a major role in maintaining an elevated rate of glucose production, probably via muscle lactate release and hepatic lactate uptake.

Compartmental modeling of glucagon kinetics in the conscious dog.

The aim of the present study was to examine glucagon metabolism and distribution using both compartmental-modeling approaches and steady-state organ-balance techniques in conscious, overnight-fasted dogs. Arterial plasma glucose concentrations were clamped at 14 mmol/L with a variable exogenous glucose infusion. Somatostatin was infused to block endogenous secretion of insulin and glucagon. Insulin was replaced intraportally at 2.4 pmol.kg-1.min-1 to maintain basal insulin concentrations in the range from 70 +/- 4 to 95 +/- 12 pmol/L. Glucagon was not given during the control period, but was subsequently infused peripherally in four 1-hour steps of 1.0, 3.0, 6.0, and 3.0 ng.kg-1.min-1. Glucagon levels increased from 0 to 68 +/- 6, 195 +/- 19, 378 +/- 47, and 181 +/- 20 ng/mL. Compartmental analysis of glucagon concentrations showed that glucagon was distributed in one compartment with a volume approximately equal to the plasma volume. The metabolic clearance rate of glucagon was 17.6 mL.kg-1.min-1. The liver cleared 24% of glucagon, and the kidneys, 17%.

Pulsatility does not alter the response to a physiological increment in glucagon in the conscious dog.

The present study was designed to investigate if pulsatile hyperglucagonemia of physiological magnitude has greater efficacy in stimulating hepatic glucose production than constant glucagon. Paired studies were performed in conscious dogs. After insulin and glucagon were clamped at basal concentrations for 2 h, glucagon was elevated for 4 h with either a continuous infusion or pulses having physiological frequency and amplitude. With continuous infusion, plasma glucagon concentrations increased from 56 +/- 7 to 194 +/- 27 ng/l. With pulsatile infusion, glucagon concentrations started at 53 +/- 6 ng/l and then oscillated between 157 +/- 15 and 253 +/- 28 ng/l. Plasma insulin concentrations remained constant at basal levels. Glucose production was determined using a time-varying two-compartment model for glucose kinetics and deconvolution. After 15 min, glucose production had risen from 13.6 +/- 1.1 to 53.8 +/- 3.9 mumol.kg-1.min-1 with continuous infusion and from 12.9 +/- 0.6 to 50.6 +/- 2.9 mumol.kg-1.min-1 with pulsatile infusion. After 4 h, the production had fallen to 16.1 +/- 1.2 and 17.1 +/- 0.7 mumol.kg-1.min-1. In the present animal model with insulin held constant, no difference was noted between the response to continuous or pulsatile glucagon infusion.

Role of glucagon in countering hypoglycemia induced by insulin infusion in dogs.

The aim of the present study was to characterize the role of glucagon in countering the prolonged hypoglycemia resulting from insulin infusion and to determine whether its effect is manifest through glycogenolysis and/or gluconeogenesis. Two groups of 18-h fasted somatostatin-treated dogs were given intraportal insulin at 5 mU.kg-1.min-1. In one group (SimGGN; n = 6), glucagon was infused intraportally so as to mimic the normal response to hypoglycemia. In a second group (BasGGN; n = 6), glucagon was infused at a basal rate. Glucose turnover and gluconeogenesis were assessed by combining tracer and hepatic balance techniques. Exogenous glucose was infused as needed to maintain equivalent hypoglycemia at approximately 45 mg/dl in the two groups. Although glucagon concentrations were significantly different, the levels of other counterregulatory hormones were equivalent in both experimental protocols. Endogenous glucose production (EGP) in SimGGN doubled from 2.4 +/- 0.2 to 5.4 +/- 0.8 mg.kg-1.min-1 by 1 h before dropping to 4.5 +/- 0.2 mg.kg-1.min-1 in the 3rd h of insulin infusion. EGP in BasGGN was initially 2.5 +/- 0.1 mg.kg-1.min-1, unchanged by 1 h, and increased to 3.9 +/- 0.2 mg.kg-1.min-1 by the 3rd h of insulin infusion. In the 1st h of insulin infusion, the rise in gluconeogenesis in both groups was equal and represented only a small part of total EGP. By the 3rd h, gluconeogenesis was the major contributor to total EGP, and gluconeogenic efficiency increased significantly more in SimGGN than BasGGN (261 vs. 140%, P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)