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Model-based pharmacokinetic and pharmacodynamic analysis for acute effects of a small molecule inhibitor of diacylglycerol acyltransferase-1 in the TallyHo/JngJ polygenic mouse
Abstract
• The purpose of this study was to evaluate the acute effect of a small molecule inhibitor of DGAT-1 on triglycerides and cholesterol in polygenic type 2 diabetic TallyHo/JngJ (TH) mice. PF-04620110, a potent and selective DGAT-1 inhibitor, was used as a model compound in this study and which was administered to TH and ICR mice.
• The concentration of the model compound that produced 50% of maximum lowering of triglyceride level (IC50) in TH mice was not significantly different from that in ICR mice, when estimated using the model-based pharmacokinetic and pharmacodynamic assay, a 2-compartmental model and an indirect response model.
• The clearance of the inhibitor in TH mice was 5-fold higher than that in ICR mice, suggesting significantly altered pharmacokinetics. Moreover, the in vitro metabolic elimination kinetic parameters (ke,met), determined using liver microsomes from TH and ICR mice were 1.24 ± 0.14 and 0.174 ± 0.116 min⁻¹, respectively.
• Thus, we report that the differences in the acute effects of the small molecule DAGT-1 inhibitor between TH mice and ICR mice can be attributed to altered pharmacokinetics caused by an altered metabolic rate for the compound in TH mice.
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Triacylglycerols are quantitatively the most important storage form of energy for eukaryotic cells. Acyl CoA:diacylglycerol
acyltransferase (DGAT, EC 2.3.1.20) catalyzes the terminal and only committed step in triacylglycerol synthesis, by using diacylglycerol and fatty acyl CoA
as substrates. DGAT plays a fundamental role in the metabolism of cellular diacylglycerol and is important in higher eukaryotes
for physiologic processes involving triacylglycerol metabolism such as intestinal fat absorption, lipoprotein assembly, adipose
tissue formation, and lactation. DGAT is an integral membrane protein that has never been purified to homogeneity, nor has
its gene been cloned. We identified an expressed sequence tag clone that shared regions of similarity with acyl CoA:cholesterol
acyltransferase, an enzyme that also uses fatty acyl CoA as a substrate. Expression of a mouse cDNA for this expressed sequence
tag in insect cells resulted in high levels of DGAT activity in cell membranes. No other acyltransferase activity was detected
when a variety of substrates, including cholesterol, were used as acyl acceptors. The gene was expressed in all tissues examined;
during differentiation of NIH 3T3-L1 cells into adipocytes, its expression increased markedly in parallel with increases in
DGAT activity. The identification of this cDNA encoding a DGAT will greatly facilitate studies of cellular glycerolipid metabolism
and its regulation.
Studies involving the cloning and disruption of the gene for acyl-CoA:diacylglycerol acyltransferase (DGAT) have shown that
alternative mechanisms exist for triglyceride synthesis. In this study, we cloned and characterized a second mammalian DGAT,
DGAT2, which was identified by its homology to a DGAT in the fungus Mortierella rammaniana. DGAT2 is a member of a gene family that has no homology with DGAT1 and includes several mouse and human homologues that
are candidates for additional DGAT genes. The expression of DGAT2 in insect cells stimulated triglyceride synthesis 6-fold
in assays with cellular membranes, and DGAT2 activity was dependent on the presence of fatty acyl-CoA and diacylglycerol,
indicating that this protein is a DGAT. Activity was not observed for acyl acceptors other than diacylglycerol. DGAT2 activity
was inhibited by a high concentration (100 mm) of MgCl2 in anin vitro assay, a characteristic that distinguishes DGAT2 from DGAT1. DGAT2 is expressed in many tissues with high expression levels
in the liver and white adipose tissue, suggesting that it may play a significant role in mammalian triglyceride metabolism.
Triglyceride accumulation is associated with obesity and type 2 diabetes. Genetic disruption of diacylglycerol acyltransferase 1 (DGAT1), which catalyzes the final reaction of triglyceride synthesis, confers dramatic resistance to high-fat diet induced obesity. Hence, DGAT1 is considered a potential therapeutic target for treating obesity and related metabolic disorders. However, the molecular events shaping the mechanism of action of DGAT1 pharmacological inhibition have not been fully explored yet. Here, we investigate the metabolic molecular mechanisms induced in response to pharmacological inhibition of DGAT1 using a recently developed computational systems biology approach, the Causal Reasoning Engine (CRE). The CRE algorithm utilizes microarray transcriptomic data and causal statements derived from the biomedical literature to infer upstream molecular events driving these transcriptional changes. The inferred upstream events (also called hypotheses) are aggregated into biological models using a set of analytical tools that allow for evaluation and integration of the hypotheses in context of their supporting evidence. In comparison to gene ontology enrichment analysis which pointed to high-level changes in metabolic processes, the CRE results provide detailed molecular hypotheses to explain the measured transcriptional changes. CRE analysis of gene expression changes in high fat habituated rats treated with a potent and selective DGAT1 inhibitor demonstrate that the majority of transcriptomic changes support a metabolic network indicative of reversal of high fat diet effects that includes a number of molecular hypotheses such as PPARG, HNF4A and SREBPs. Finally, the CRE-generated molecular hypotheses from DGAT1 inhibitor treated rats were found to capture the major molecular characteristics of DGAT1 deficient mice, supporting a phenotype of decreased lipid and increased insulin sensitivity.
Acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1) is one of two known DGAT enzymes that catalyze the final step in triglyceride
synthesis. Findings from genetically modified mice as well as pharmacological studies suggest that inhibition of DGAT1 is
a promising strategy for the treatment of obesity and type 2 diabetes. Here we characterize a tool DGAT1 inhibitor compound,
T863. We found that T863 is a potent inhibitor for both human and mouse DGAT1 in vitro, which acts on the acyl-CoA binding
site of DGAT1, and inhibits DGAT1-mediated triacylglycerol formation in cells. In an acute lipid challenge model, oral administration
of T863 significantly delayed fat absorption and resulted in lipid accumulation in the distal small intestine of mice, mimicking
the effects of genetic ablation of DGAT1. In diet-induced obese mice, oral administration of T863 for two weeks caused weight
loss, reduction in serum and liver triglycerides, and improved insulin sensitivity. In addition to the expected triglyceride-lowering
activity, T863 also lowered serum cholesterol. Hepatic IRS2 protein was dramatically upregulated in mice treated with T863,
possibly contributing to improved insulin sensitivity. In differentiated 3T3-L1 adipocytes, T863 enhanced insulin-stimulated
glucose uptake, suggesting a possible role for adipocytes to improve insulin sensitivity upon DGAT1 inhibition. These results
reveal novel mechanistic insights into the insulin-sensitizing effects of DGAT1 inhibition in mouse models. Taken together,
our study provides a comprehensive evaluation of a small molecule inhibitor for DGAT1 and suggests that pharmacological inhibition
of DGAT1 holds promise in treating diverse metabolic disorders.
Dietary triacylglycerols are a major source of energy for animals. The absorption of dietary triacylglycerols involves their hydrolysis to free fatty acids and monoacylglycerols in the intestinal lumen, the uptake of these products into enterocytes, the resynthesis of triacylgylcerols, and the incorporation of newly synthesized triacylglycerols into nascent chylomicrons for secretion. In enterocytes, the final step in triacylglycerol synthesis is believed to be catalyzed primarily through the actions of acyl-CoA:diacylglycerol acyltransferase (DGAT) enzymes. In this study, we analyzed intestinal triacylglycerol absorption and chylomicron synthesis and secretion in DGAT1-deficient (Dgat1(-/-)) mice. Surprisingly, DGAT1 was not essential for quantitative dietary triacylglycerol absorption, even in mice fed a high fat diet, or for the synthesis of chylomicrons. However, Dgat1(-/-) mice had reduced postabsorptive chylomicronemia (1 h after a high fat challenge) and accumulated neutral-lipid droplets in the cytoplasm of enterocytes when chronically fed a high fat diet. These results suggest a reduced rate of triacylglycerol absorption in Dgat1(-/-) mice. Analysis of intestine from Dgat1(-/-) mice revealed activity for two other enzymes, DGAT2 and diacylglycerol transacylase, that catalyze triacylglycerol synthesis and apparently help to compensate for the absence of DGAT1. Our findings indicate that multiple mechanisms for triacylglycerol synthesis in the intestine facilitate triacylglycerol absorption.
Mice lacking acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1), an enzyme that catalyzes the terminal step in triacylglycerol synthesis, have enhanced insulin sensitivity and are protected from obesity, a result of increased energy expenditure. In these mice, factors derived from white adipose tissue (WAT) contribute to the systemic changes in metabolism. One such factor, adiponectin, increases fatty acid oxidation and enhances insulin sensitivity. To test the hypothesis that adiponectin is required for the altered energy and glucose metabolism in DGAT1-deficient mice, we generated adiponectin-deficient mice and introduced adiponectin deficiency into DGAT1-deficient mice by genetic crosses. Although adiponectin-deficient mice fed a high-fat diet were heavier, exhibited worse glucose tolerance, and had more hepatic triacylglycerol accumulation than wild-type controls, mice lacking both DGAT1 and adiponectin, like DGAT1-deficient mice, were protected from diet-induced obesity, glucose intolerance, and hepatic steatosis. These findings indicate that adiponectin is required for normal energy, glucose, and lipid metabolism but that the metabolic changes induced by DGAT1-deficient WAT are independent of adiponectin and are likely due to other WAT-derived factors. Our findings also suggest that the pharmacological inhibition of DGAT1 may be useful for treating human obesity and insulin resistance associated with low circulating adiponectin levels.
The TALLYHO/JngJ (TH) strain is a newly established, polygenic mouse model for type 2 diabetes (T2D) and obesity, and we have previously reported some key physiological features of this model after the overt onset of diabetes. In the present work, we conducted a comprehensive phenotypic characterization of TH in order to completely characterize this new and relevant model for human T2D and obesity. We monitored the development of obesity and diabetes starting at 4 weeks of age by measuring body weight, glucose tolerance, and plasma levels of insulin, glucose, and triglyceride. Additionally, histological alterations in the pancreas and glucose uptake and glucose transporter 4 (GLUT4) content in soleus muscle were also examined. Compared with age- and sex-matched C57BL/6J (B6) mice, both male and female TH mice were significantly heavier, hyperleptinemic, and hyperinsulinemic at 4 weeks of age, without glucose intolerance or hyperglycemia. TH mice maintained higher body weights throughout the study period of 16 weeks. The hyperinsulinemia in TH mice worsened with age, but to a lesser degree in females than in males. Both the male and the female TH mice had enlarged pancreatic islets. Male TH mice showed impaired glucose tolerance at 8 weeks that became more prominent at 16 weeks. Plasma glucose levels continuously increased with age in male TH mice resulting in frank diabetes, while female TH mice remained normoglycemic throughout the study. Impaired glucose tolerance and hyperglycemia in male TH mice were accompanied by impaired 2-deoxyglucose uptake in the soleus muscle at basal and insulin-stimulated states, but without any reduction in GLUT4 content. Interestingly, male TH mice exhibited a drastic elevation in plasma triglyceride levels in the pre-diabetic stage that was maintained throughout the study. These findings suggest that obesity and insulin resistance are an inherent part of the TH phenotype and glucose intolerance is evident preceding progression to overt diabetes in male TH mice.
Cytochrome P450 (CYP) epoxygenase-derived epoxyeicosatrienoic acids (EETs) elicit cell proliferation and promote angiogenesis. The aim of this study was to determine the expression of CYP epoxygenases in the bovine retina and the potential role of EETs in hypoxia-induced angiogenesis in bovine retinal endothelial cells.
Bovine retinal endothelial cells were cultured under normoxic (21% O(2)) or hypoxic (1% O(2)) conditions, and CYP2C expression was determined by Western blot analysis. The effect of hypoxia on EET levels was determined by LC-MS/MS. Cell migration (Transwell filter assays) and endothelial cell tube formation (on basement membrane matrix) were assessed in vitro in the absence and presence of pharmacologic inhibitors and CYP2C antisense oligonucleotides.
Bovine retinal endothelial cells expressed CYP2C protein in culture and generated detectable levels of EETs under basal conditions. Hypoxia (6-48 hours) enhanced CYP2C protein expression (2-fold) and EET formation (1.5-fold). Moreover, endothelial cells preexposed to hypoxia demonstrated an increase in serum-induced cell migration that was sensitive to the CYP2C inhibitors sulfaphenazole and MS-PPOH and the EET antagonist 14,15-epoxyeicosa-5(Z)-enoic acid. Furthermore, preventing the hypoxia-induced expression of CYP2C (antisense oligonucleotides) suppressed hypoxia-induced cell migration. In an in vitro angiogenesis model, the preexposure of endothelial cells to hypoxia increased CYP2C expression and enhanced endothelial tube formation, which was blocked by the EET antagonist and by the CYP2C antisense oligonucleotides.
Taken together, these data indicate that CYP2C-derived EETs are implicated in angiogenesis by retinal endothelial cells, especially under hypoxic conditions.
Six studies of primary prevention in patients with diabetes were identified. The Air Force Coronary Atherosclerosis Prevention Study/Texas Coronary Atherosclerosis Prevention Study (AFCAPS/TexCAPS) randomly assigned patients with average cholesterol levels and lower than average high-density lipoprotein (HDL) cholesterol levels to lovastatin, 20 to 40 mg/d, or placebo (in addition to a low-fat and low-cholesterol diet) for an average follow-up of 5.2 years (11). Based on data from the Third National Health and Nutrition Examination Survey, mean total cholesterol level was 5.72 mmol/L (221 mg/dL), mean LDL cholesterol level was 3.88 mmol/L (150 mg/dL), and mean HDL cholesterol level was 0.93 mmol/L (36 mg/dL) for men and 1.03 mmol/L (40 mg/dL) for women. One hundred fifty-five patients had diabetes. Lovastatin therapy led to a relative risk of 0.56 (95% CI, 0.17 to 1.92) for any atherosclerotic cardiovascular event (first fatal or nonfatal myocardial infarction, unstable angina, or sudden cardiac death) and an absolute risk reduction of 0.04 (CI, −0.04 to 0.12), neither of which was statistically significant. The mean LDL cholesterol level at the end of the study was 2.97 mmol/L (115 mg/dL), and the mean HDL cholesterol level was 1.00 mmol/L (39 mg/dL).
The current literature suggests that statins are extremely safe. Although rates of discontinuation and nonadherence in clinical trials are approximately 15% or more in many cases, rates of discontinuation typically are not different from those of placebo. Rates of elevated levels of liver or muscle enzymes did not differ between the statin and placebo groups in recent large-scale studies. For example, in the HPS, rates of elevated alanine aminotransferase levels above twice the upper limit of normal were 1.8% in the simvastatin group and 1.6% in the placebo group, and rates of elevated creatine kinase levels were 0.3% in the simvastatin group and 0.2% in the placebo group (15). Neither of these differences was statistically significant. Similarly, among the 5804 patients in PROSPER, only 1 person in each group had an alanine aminotransferase or aspartate aminotransferase level more than 3 times the upper limit of normal (16). In addition, no patients had rhabdomyolysis and 36 patients in the pravastatin group had myalgia compared with 32 in the placebo group. Based on the safety of these drugs, routine monitoring of liver or muscle enzymes is probably not warranted except in patients with symptoms, patients who have liver enzyme abnormalities at baseline, or patients taking drugs that interact with statins to increase the risk for adverse events.
Acyl-CoA:diacylglycerol acyltransferase-1 (DGAT-1) catalyzes the final committed step in the biosynthesis of triglycerides. DGAT-1 knockout mice have been shown to be resistant to diet-induced obesity and have increased insulin sensitivity. Thus, inhibition of DGAT-1 may represent an attractive target for the treatment of obesity or type II diabetes. Herein, we report the discovery and characterization of a potent and selective DGAT-1 inhibitor PF-04620110 (3). Compound 3 inhibits DGAT-1 with an IC50 of 19 nM and shows high selectivity versus a broad panel of off-target pharmacologic end points. In vivo DGAT-1 inhibition has been demonstrated through reduction of plasma triglyceride levels in rodents at doses of ≥0.1 mg/kg following a lipid challenge. On the basis of this pharmacologic and pharmacokinetic profile, compound 3 has been advanced to human clinical studies.Keywords: DGAT-1; diabetes; obesity; triglyceride; phototoxicity; acyl glucuronide
Purpose. The effect of hyperlipidemia on nifedipine pharmacokinetics was studied. The mechanisms by which hyperlipidemia affects pharmacokinetics of drugs are mainly undetermined. Hyperlipidemia may decrease the fraction of unbound drug in plasma and/or decrease intrinsic ability of the cytochrome P-450 systems due to excess membrane cholesterol. Hyperlipidemia is a primary risk factor for coronary artery disease leading to hypertension and ischemic heart disease, for which nifedipine, a calcium channel blocker, is used.
Methods. Poloxamer 407 (P407)-induced hyperlipidemic rat model was used to study the effects of hyperlipidemia on the pharmacokinetics of nifedipine (6 mg kg–1 given iv, ip and po). Total plasma cholesterol levels increased from 0.82–2.02 to 5.27–11.05 mmol L–1 48 h post P407 administration (Ig kg–1, ip). Protein binding studies were conducted by an ultrafiltration method.
Results. Hyperlipidemia significantly decreased CLTB by 38% and CLTB/F by 45 and 42% following po and ip doses, respectively, thereby increasing AUC0–, Cmax and half-life. Absolute bioavailability and Vdss remained unchanged. AUC0– was affected to the same extent in each route of administration, therefore, the effect was mainly systemic rather than presystemic. Hyperlipidemia significantly lowered the fraction unbound in plasma by approximately 31%.
Conclusions. The altered pharmacokinetics of nifedipine by P407-induced HYPERLIPIDEMIA may be, at least in part, due to the decrease in fraction unbound in plasma. A decrease in intrinsic clearance, however, cannot be ruled out.
See related article, pp 216–222
The pandemic of obesity is a devastating health problem and contributes to premature morbidity and mortality. Results from clinical and experimental studies have identified a variety of unfavorable consequences of obesity including cardiovascular diseases, pulmonary diseases, cancer, and sleep disorders. An obesity-triggered parallel increase in the prevalence of type 2 diabetes mellitus is also expected to add to the overall cardiovascular burden of obesity. Components of metabolic syndrome such as dyslipidemia, hyperglycemia, insulin resistance, and hypertension are thought to play pivotal roles in obesity-associated sequelae responsible for atherosclerosis, cardiac hypertrophy, and ventricular dysfunction. The presence of 1 or more of these metabolic syndrome components can adversely affect multiple metabolic pathways resulting in alterations in glucose and lipid metabolism, fatty acid (FA) transport/storage/oxidation, oxygen consumption, redox status, and high-energy phosphate metabolism. Although the precise mechanism(s) of action responsible for metabolic derangement-induced cardiac abnormalities in obesity remains poorly understood, 1 theory that has received increasing attention focuses on lipid transport and storage, excessive FA oxidation (FAO), and lipotoxic injury to the heart.1,2 When energy intake exceeds expenditure, fat is stored as triacylglycerol (TG) in adipose tissue. In turn, once fat levels exceed the storage capacity of adipocytes, a variety of neutral lipids are released and accumulated in other cells and tissues including the heart. The presence of lipid inclusions within cardiomyocytes, a condition referred to as cardiac steatosis, has been confirmed in obesity and diabetes.3,4 Although recent evidence indicates that cardiac steatosis, increased availability of FA and excess FAO contribute to cardiac anomalies associated with obesity and type 2 diabetes, it has also been suggested that cardiac steatosis may be a compensatory mechanism used to neutralize FAs and their metabolites through esterification to neutral lipids.
Generation of ATP for normal cardiac …
Acyl CoA/diacylglycerol acyltransferase (DGAT) 1 is one of two known DGAT enzymes that catalyze the final and only committed step in triglyceride biosynthesis. The purpose of this study was to test the hypothesis that chronic inhibition of DGAT-1 with a small-molecule inhibitor will reduce serum triglyceride concentrations in both genetic and diet-induced models of hypertriglyceridemia. Zucker fatty rats and diet-induced dyslipidemic hamsters were dosed orally with A-922500 (0.03, 0.3, and 3-mg/kg), a potent and selective DGAT-1 inhibitor, for 14 days. Serum triglycerides were significantly reduced by the 3 mg/kg dose of the DGAT-1 inhibitor in both the Zucker fatty rat (39%) and hyperlipidemic hamster (53%). These serum triglyceride changes were accompanied by significant reductions in free fatty acid levels by 32% in the Zucker fatty rat and 55% in the hyperlipidemic hamster. In addition, high-density lipoprotein-cholesterol was significantly increased (25%) in the Zucker fatty rat by A-922500 administered at 3 mg/kg. This study provides the first report that inhibition of DGAT-1, the final and only committed step of triglyceride synthesis, with a selective small-molecule inhibitor, significantly reduces serum triglyceride levels in both genetic and diet-induced animal models of hypertriglyceridemia. The results of this study support further investigation of DGAT-1 inhibition as a novel therapeutic approach to the treatment of hypertriglyceridemia in humans, and they suggest that inhibition of triglyceride synthesis may have more diverse beneficial effects on serum lipid profiles beyond triglyceride lowering.
These studies provide new insight into the complex mechanisms wherby hyperlipidemia causes progressive atherosclerosis. It has been shown that physical injury to the endothelial lining of arteries sets off a process which probably is an attempt at healing the injury but which can lead to atherosclerosis. It has also been found that chemical agents such as homocystine can produce a similar series of events leading to atherosclerosis. These events include focal loss of endothelium, exposure of subendothelial connective tissue, and adherence of platelets followed by release of factors that stimulate intimal smooth muscle proliferation. The present studies indicate that the effects of chronic hyperlipidemia are complex in that the condition results not only in the deposition of lipids in the atheromatous lesions but that it may produce the primary endothelial injury that initiates the process of atherosclerosis as well.
Four basic models for characterizing indirect pharmacodynamic responses after drug administration have been developed and compared. The models are based on drug effects (inhibition or stimulation) on the factors controlling either the input or the dissipation of drug response. Pharmacokinetic parameters of methylprednisolone were used to generate plasma concentration and response-time profiles using computer simulations. It was found that the responses produced showed a slow onset and a slow return to baseline. The time of maximal response was dependent on the model and dose. In each case, hysteresis plots showed that drug concentrations preceded the response. When the responses were fitted with pharmacodynamic models based on distribution to a hypothetical effect compartment, the resulting parameters were dose-dependent and inferred biological implausibility. Indirect response models must be treated as distinct from conventional pharmacodynamic models which assume direct action of drugs. The assumptions, equations, and data patterns for the four basic indirect response models provide a starting point for evaluation of pharmacologic effects where the site of action precedes or follows the measured response variable.
Unlabelled:
The study was carried out on male rabbits divided into two groups: a control and an experimental one, fed on a high-fat diet. Humans were also ascribed into two groups: control and those affected with primary, mixed form of hyperlipidemia. The animals and humans were given theophylline intravenously as a single dose. Blood was sampled after 5, 10, 15, 30 and 45 min and 1, 2, 4, 6, 8, 12 and 24 h following theophylline administration. FPIA method was used to determine blood serum concentrations of theophylline. Considerable alterations of theophylline pharmacokinetics in humans suffering from mixed form of hyperlipidemia were observed. Marked decrease in area under the concentration-time curve (AUC), diminished volume of distribution, increased total body clearance, and shortened elimination half-life were observed. On the contrary, in rabbits with alimentary induced lipid metabolism disturbances t1/2 of theophylline was practically unchanged and AUC only slightly increased.
In conclusion:
(1) hyperlipidemia affects the pharmacokinetics of theophylline in human beings, (2) rabbit model with dietetary induced lipid metabolic disturbances is not a suitable subject for estimation of pharmacokinetics of xanthine derivatives.
The relationship between blood concentrations of the adenosine A
1-receptor agonist N6-(p-sulfophenyI)adenosine (SPA) and its effect on both plasma nonesterified fatty acid (NEFA) and glycerol release was described on the basis of an integrated pharmacokinetic–pharmacodynamic model. An indirect response model rather than a hypothetical “link” model was used to account for the delayed response. For that purpose an empirical solution to the differential equation describing the physiological indirect response model is presented. The model-estimated rate constant for the output of the glycerol response was compared to the elimination rate constant after exogenous administration of glycerol. In a crossover designed study, chronically cannulated male Wistar rats were subjected to either SPA administration (120 μg/kg for 15 min) for measurement of the effects on glycerol, or glycerol administration for determination of glycerol pharmacokinetics. Glycerol pharmacokinetics was determined in the presence of a stable level of SPA (171±6ng/ml) to suppress endogenous glycerol levels completely. The indirect response model adequately described the relationship between SPA concentrations and plasma glycerol levels. The PD parameter estimates for EC
50, Emax, and Hill factor were 23±2 ng/ml, 74±3% (change from baseline), and 3.3±0.5, respectively. These values were not different from those obtained when analyzing the data on basis of the differential equation directly. Furthermore the EC50 values for the reduction in glycerol or NEFA levels were identical (23±2 and 21±3 ng/ml, respectively) indicating that both PD end points reflect the same physiological process. The concentration–time profile after administration of glycerol could be described best on the basis of a biexponential function. The value for kout in the PK/PD model (0.19±0.03 min
−1) corresponded very well to the terminal elimination rate constant determined after iv administration of glycerol (0.25±0.03 min
−1). In conclusion, the antilipolytic effects of adenosine A
1-receptor agonists can be described by the indirect suppression model. The rate constant describing the delay between concentration and glycerol effect was shown to be a true reflection of the removal of glycerol.
Hyperlipidemia is recognized as one of the major risk factors for the development of coronary artery disease and progression of atherosclerotic lesions. Dietary therapy together with hypolipidemic drugs are central to the management of hyperlipidemia, which aims to prevent atherosclerotic plaque progression, induce regression, and so decrease the risk of acute coronary events in patients with pre-existing coronary or peripheral vascular disease. In patients at high risk of coronary artery disease but without evidence of atherosclerosis, treatment is designed to prevent the premature development of coronary artery disease, whereas in those with hypertriglyceridemia, treatment aims to prevent the development of hepatomegaly, splenomegaly, and pancreatitis. The 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, or statins, are the most potent lipid-lowering agents currently available, and their use in the treatment of hyperlipidemia provides the focus for this review. Particular emphasis is given to cerivastatin, a new HMG-CoA reductase inhibitor that combines potent cholesterol-lowering properties with significant triglyceride-reducing effects. Recently completed primary and secondary intervention trials have shown that the significant reductions in low-density lipoprotein (LDL) cholesterol achieved with statins result in significant reductions in morbidity and mortality associated with coronary artery disease as well as reductions in the incidence of stroke and total mortality. Such benefits occur early in the course of statin therapy and have led to suggestions that these drugs may possess antiatherogenic effects over and above their capacity to lower atherogenic lipids and lipoproteins. Experimental studies have also shown statin-induced improvements in endothelial function, decreased platelet thrombus formation, improvements in fibrinolytic activity, and reductions in the frequency of transient myocardial ischemia.
Statins (HMG-CoA reductase inhibitors) are used widely for the treatment of hypercholesterolemia. They inhibit HMG-CoA reductase competitively, reduce LDL levels more than other cholesterol-lowering drugs, and lower triglyceride levels in hypertriglyceridemic patients. Statins are well tolerated and have an excellent safety record. Clinical trials in patients with and without coronary heart disease and with and without high cholesterol have demonstrated consistently that statins reduce the relative risk of major coronary events by approximately 30% and produce a greater absolute benefit in patients with higher baseline risk. Proposed mechanisms include favorable effects on plasma lipoproteins, endothelial function, plaque architecture and stability, thrombosis, and inflammation. Mechanisms independent of LDL lowering may play an important role in the clinical benefits conferred by these drugs and may ultimately broaden their indication from lipid-lowering to antiatherogenic agents.
The TallyHo (TH) mouse strain is a newly established model for non-insulin-dependent diabetes mellitus (NIDDM). TH mice show obesity, hyperinsulinemia, hyperlipidemia, and male-limited hyperglycemia. A genetic dissection of the diabetes syndrome has been carried out using male backcross 1 progeny obtained from crosses between (C57BL/6J x TH)F1 and TH mice or (CAST/Ei x TH)F1 and TH mice. A genome-wide scan reveals three quantitative trait loci (QTLs), Tanidd1-3 (TH-associated NIDDM) linked to hyperglycemia. The major QTL (common in both crosses), Tanidd1, maps to chromosome (Chr) 19. Additionally, gene-gene interactions contributing to hyperglycemia have been observed between Tanidd1 and a locus on Chr 18 as well as between Tanidd2 and a locus on Chr 16. The overt hyperglycemia in TH mice is, therefore, likely due to a mutation in a major diabetes susceptibility locus on Chr 19, which interacts with additional genes to lead to an observable phenotype.
Obesity is currently an exceptionally common problem in humans. The last several years have produced a significant number of breakthroughs in obesity related areas of investigation. Triglycerides are considered the main form of storage of excess calories in fat. A key enzyme in the synthesis of triglycerides is acylCoA: diacylglycerol acyltransferase (DGAT). Recent studies have shown that mice deficient in this enzyme are resistant to diet induced obesity and have increased insulin and leptin sensitivity. These effects suggest that inhibition of DGAT in vivo may be a novel therapeutic target not only for obesity but also for diabetes.
Most of the triacylglycerol (TAG) utilized for the assembly of very-low-density lipoprotein (VLDL) in the secretory apparatus of the hepatocyte is mobilized by lipolysis of the cytosolic TAG pool, followed by re-esterification. The lipases involved include arylacetamide deacetylase and/or triacylglycerol hydrolase. Some of the re-esterified products of lipolysis gain access to an apolipoprotein-B-rich VLDL precursor to form mature VLDL. Some, however, are returned to the cytosolic pool in a process that is stimulated by insulin and inhibited by microsomal triacylglycerol transfer protein (MTP). Phospholipids also contribute to VLDL TAG in a process which involves ADP-ribosylation factor-1 (ARF-1)-mediated activation of phospholipase D. The temporary storage of TAG in the liver, followed by its mobilization and secretion as VLDL, form part of a process by which the liver protects vulnerable body tissues from excess lipotoxic non-esterified ('free') fatty acids in the plasma.
The TallyHo (TH) mouse strain is a polygenic model for Type 2 diabetes with obesity. Genetic analysis in backcross progeny from a cross between F1 [C57BL/6J (B6) x TH] and TH mice mapped a quantitative trait locus (QTL) named TH-associated body weight 2 (tabw2) to chromosome 6. The TH-derived allele is associated with increased body weight. As a first step to identify the molecular basis of this obesity QTL, we constructed a congenic line of mice on the B6 genetic background that carries a genomic region from TH mice containing tabw2. Congenic mice homozygous for tabw2 (B6.TH-tabw2/tabw2) fed a chow diet exhibited slightly, but significantly, higher body weight and body fat and plasma leptin levels compared with controls (B6.TH-+/+). This difference was exacerbated when the animals were maintained on a high-fat and high-sucrose (HFS) diet. The diet-induced obesity in tabw2 congenic mice is accompanied by hyperleptinemia, mild hyperinsulinemia, impaired glucose tolerance, and reduced glucose uptake in adipose tissue in response to insulin administration. Using F2 progeny fed a HFS diet from an intercross of B6.TH-tabw2/+ mice, we were able to refine the map position of the tabw2 obesity susceptibility locus to a 15-cM region (95% confidence interval) extending distally from the marker D6Mit102. In summary, tabw2 congenic mice are a new animal model for diet-induced obesity that will be valuable for the study of gene-diet interactions.
The objective of this study was to examine the effect of a high fat meal and hyperlipidemia on the pharmacokinetic behavior of amiodarone. To evaluate these effects, single doses of amiodarone were administered to rats i.v. (25 mg/kg) or orally (50 mg/kg). Some rats were rendered hyperlipidemic by intraperitoneal doses of poloxamer 407 followed by amiodarone i.v. In other normolipidemic rats, amiodarone was administered i.v. in a fasted state or after the administration of 1% cholesterol in peanut oil. Amiodarone plasma concentrations were considerably (>11-fold) increased in hyperlipidemia. Substantial decreases were noted in the clearance, volume of distribution and unbound fraction (11.6, 23 and 24.7-fold, respectively) in plasma of hyperlipidemic rats. Oral lipid caused a significant increase in plasma AUC(0-infinity) (1.38-fold) and a significant decrease in clearance (1.5-fold) of amiodarone after intravenous doses. Oral consumption of 1% cholesterol in peanut oil significantly increased the plasma AUC (1.83-fold) and bioavailability of amiodarone (1.31-fold) after oral doses. In determining oral bioavailability of lipophilic drugs such as amiodarone in food effect studies, in addition to the increase in absorption of drugs, other factors such as a decrease in clearance due to increases in lipoprotein levels should be taken into account.
Statins are the most commonly prescribed agents for the treatment of hypercholesterolaemia. This is due to their efficacy in reducing low-density lipoprotein cholesterol (LDL) level which is the primary goal of the treatment especially for patients with multiple risk factors or with established coronary heart diseases. The purpose of this study was to develop a pharmacokinetic/pharmacodynamic (PK/PD) model that describes the LDL-lowering process in patients with hypercholesterolaemia treated with atorvastatin, fluvastatin or simvastatin. A total of 100 patients were studied retrospectively. They received atorvastatin (n = 57), fluvastatin (n = 26) or simvastatin (n = 17). As no pharmacokinetic data were available, the absorption rate was fixed to 1/h and atorvastatin, simvastatin and fluvastatin elimination half-lives were fixed to 14, 2 and 2.5 h respectively. A total of 309 LDL levels were measured and the data were analysed by nonmem v. The time course of the LDL-lowering effect of statins was described by an indirect-response model with precursor (LDL synthesis, input rate K(in)) and response (circulating LDL, input and output rates K) compartments. The following parameters were estimated: LDL input rate (K(in)) 0.14 +/- 0.015 g/L/day (mean +/- SD); inhibition fraction of K(in) (INH) 0.21 +/- 0.017; and dose producing 50% increase of LDL removal (D50), 26 +/- 7.8, 1.3 +/- 0.48 and 15 +/- 5.25 mg for atorvastatin, simvastatin and fluvastatin, respectively. Gender, bodyweight, age, calories/day, sugar/day, lipids/day, hyperlipidaemia types and waist/hip circumference, renal and hepatic functions had no effect on the pharmacodynamic parameters. The pharmacodynamic parameters for the three statins were accurately estimated. The PK/PD model developed successfully predicted the time course of the LDL-lowering effect of statins.
A highly potent and selective DGAT-1 inhibitor was identified and used in rodent models of obesity and postprandial chylomicron excursion to validate DGAT-1 inhibition as a novel approach for the treatment of metabolic diseases. Specifically, compound 4a conferred weight loss and a reduction in liver triglycerides when dosed chronically in DIO mice and depleted serum triglycerides following a lipid challenge in a dose-dependent manner, thus, reproducing major phenotypical characteristics of DGAT-1(-/-) mice.
Niacin therapy in atherosclerosis
- C D Meyers
- V S Kamanna
- M L Kashyap
Characteristics of the newly established diabetic model mice
- S D Rhee
- W H Jeong
- Y Y Sung