Optimization of Tetrahydroindazoles as Inhibitors of Human Dihydroorotate Dehydrogenase and Evaluation of Their Activity and In Vitro Metabolic Stability.

Human dihydroorotate dehydrogenase (DHODH), an enzyme in the de novo pyrimidine synthesis pathway, is a target for the treatment of rheumatoid arthritis and multiple sclerosis and is re-emerging as an attractive target for cancer therapy. Here we describe the optimization of recently identified tetrahydroindazoles (HZ) as DHODH inhibitors. Several of the HZ analogues synthesized in this study are highly potent inhibitors of DHODH in an enzymatic assay, while also inhibiting cancer cell growth and viability and activating p53-dependent transcription factor activity in a reporter cell assay. Furthermore, we demonstrate the specificity of the compounds toward the de novo pyrimidine synthesis pathway through supplementation with an excess of uridine. We also show that induction of the DNA damage marker γ-H2AX after DHODH inhibition is preventable by cotreatment with the pan-caspase inhibitor Z-VAD-FMK. Additional solubility and in vitro metabolic stability profiling revealed compound 51 as a favorable candidate for preclinical efficacy studies.

Overeating Saturated Fat Promotes Fatty Liver and Ceramides Compared With Polyunsaturated Fat: A Randomized Trial.

CONTEXT: Saturated fatty acid (SFA) vs polyunsaturated fatty acid (PUFA) may promote nonalcoholic fatty liver disease by yet unclear mechanisms. OBJECTIVE: To investigate if overeating SFA- and PUFA-enriched diets lead to differential liver fat accumulation in overweight and obese humans. DESIGN: Double-blind randomized trial (LIPOGAIN-2). Overfeeding SFA vs PUFA for 8 weeks, followed by 4 weeks of caloric restriction. SETTING: General community. PARTICIPANTS: Men and women who are overweight or have obesity (n = 61). INTERVENTION: Muffins, high in either palm (SFA) or sunflower oil (PUFA), were added to the habitual diet. MAIN OUTCOME MEASURES: Lean tissue mass (not reported here). Secondary and exploratory outcomes included liver and ectopic fat depots. RESULTS: By design, body weight gain was similar in SFA (2.31 ± 1.38 kg) and PUFA (2.01 ± 1.90 kg) groups, P = 0.50. SFA markedly induced liver fat content (50% relative increase) along with liver enzymes and atherogenic serum lipids. In contrast, despite similar weight gain, PUFA did not increase liver fat or liver enzymes or cause any adverse effects on blood lipids. SFA had no differential effect on the accumulation of visceral fat, pancreas fat, or total body fat compared with PUFA. SFA consistently increased, whereas PUFA reduced circulating ceramides, changes that were moderately associated with liver fat changes and proposed markers of hepatic lipogenesis. The adverse metabolic effects of SFA were reversed by calorie restriction. CONCLUSIONS: SFA markedly induces liver fat and serum ceramides, whereas dietary PUFA prevents liver fat accumulation and reduces ceramides and hyperlipidemia during excess energy intake and weight gain in overweight individuals.

Effects of free omega-3 carboxylic acids and fenofibrate on liver fat content in patients with hypertriglyceridemia and non-alcoholic fatty liver disease: A double-blind, randomized, placebo-controlled study.

BACKGROUND: Treatment with omega-3 fatty acids and fenofibrates reduces serum triglyceride levels, but few studies have compared the effect of these agents on liver fat. OBJECTIVE: The aim of the EFFECT I trial (NCT02354976) was to determine the effects of free omega-3 carboxylic acids (OM-3CA) and fenofibrate on liver fat in overweight or obese individuals with non-alcoholic fatty liver disease and hypertriglyceridemia. METHODS: Seventy-eight patients were randomized to receive oral doses of 4 g OM-3CA (n = 25), 200 mg fenofibrate (n = 27), or placebo (n = 26) for 12 weeks in a double-blind, parallel-group study. Liver proton density fat fraction (PDFF) and volume, pancreas volume, and adipose tissue volumes were assessed by magnetic resonance imaging. RESULTS: Changes in liver PDFF at 12 weeks were not significantly different across treatment groups (relative changes from baseline: placebo, +4%; OM-3CA, -2%; and fenofibrate, +17%). The common PNPLA3 genetic polymorphism (I148M) did not significantly influence the effects of OM-3CA or fenofibrate on liver PDFF. Fenofibrate treatment significantly increased liver and pancreas volumes vs placebo treatment, and the changes in liver and pancreas volumes were positively correlated (rho 0.45, P = .02). Total liver fat volume increased significantly in patients using fenofibrate vs OM-3CA (+23% vs -3%, P = .04). Compared with OM-3CA, fenofibrate increased total liver fat and liver volume. Serum triglycerides decreased with OM-3CA (-26%, P = .02) and fenofibrate (-38%, P < .001) vs placebo. In contrast to OM-3CA, fenofibrate reduced plasma docosahexaenoic acid levels and increased plasma acetylcarnitine and butyrylcarnitine levels, estimated delta-9 desaturase activity and the concentration of urine F2-isoprostanes. CONCLUSIONS: OM-3CA and fenofibrate reduced serum triglycerides but did not reduce liver fat. Fenofibrate increased total liver volume and total liver fat volume vs OM-3CA, indicating a complex effect of fenofibrate on human hepatic lipid metabolism.

Effects of dapagliflozin and n-3 carboxylic acids on non-alcoholic fatty liver disease in people with type 2 diabetes: a double-blind randomised placebo-controlled study.

AIMS/HYPOTHESIS: The EFFECT-II study aimed to investigate the effects of dapagliflozin and omega-3 (n-3) carboxylic acids (OM-3CA), individually or combined, on liver fat content in individuals with type 2 diabetes and non-alcoholic fatty liver disease (NAFLD). METHODS: This randomised placebo-controlled double-blind parallel-group study was performed at five clinical research centres at university hospitals in Sweden. 84 participants with type 2 diabetes and NAFLD were randomly assigned 1:1:1:1 to four treatments by a centralised randomisation system, and all participants as well as investigators and staff involved in the study conduct and analyses were blinded to treatments. Each group received oral doses of one of the following: 10 mg dapagliflozin (n = 21), 4 g OM-3CA (n = 20), a combination of both (n = 22) or placebo (n = 21). The primary endpoint was liver fat content assessed by MRI (proton density fat fraction [PDFF]) and, in addition, total liver volume and markers of glucose and lipid metabolism as well as of hepatocyte injury and oxidative stress were assessed at baseline and after 12 weeks of treatment (completion of the trial). RESULTS: Participants had a mean age of 65.5 years (SD 5.9), BMI 31.2 kg/m2 (3.5) and liver PDFF 18% (9.3). All active treatments significantly reduced liver PDFF from baseline, relative changes: OM-3CA, -15%; dapagliflozin, -13%; OM-3CA + dapagliflozin, -21%. Only the combination treatment reduced liver PDFF (p = 0.046) and total liver fat volume (relative change, -24%, p = 0.037) in comparison with placebo. There was an interaction between the PNPLA3 I148M polymorphism and change in liver PDFF in the active treatment groups (p = 0.03). Dapagliflozin monotherapy, but not the combination with OM-3CA, reduced the levels of hepatocyte injury biomarkers, including alanine aminotransferase, aspartate aminotransferase, γ-glutamyl transferase (γ-GT), cytokeratin (CK) 18-M30 and CK 18-M65 and plasma fibroblast growth factor 21 (FGF21). Changes in γ-GT correlated with changes in liver PDFF (ρ = 0.53, p = 0.02). Dapagliflozin alone and in combination with OM-3CA improved glucose control and reduced body weight and abdominal fat volumes. Fatty acid oxidative stress biomarkers were not affected by treatments. There were no new or unexpected adverse events compared with previous studies with these treatments. CONCLUSIONS/INTERPRETATION: Combined treatment with dapagliflozin and OM-3CA significantly reduced liver fat content. Dapagliflozin monotherapy reduced all measured hepatocyte injury biomarkers and FGF21, suggesting a disease-modifying effect in NAFLD. TRIAL REGISTRATION: ClinicalTrials.gov NCT02279407 FUNDING: The study was funded by AstraZeneca.

A concept for holistic whole body MRI data analysis, Imiomics.

PURPOSE: To present and evaluate a whole-body image analysis concept, Imiomics (imaging-omics) and an image registration method that enables Imiomics analyses by deforming all image data to a common coordinate system, so that the information in each voxel can be compared between persons or within a person over time and integrated with non-imaging data. METHODS: The presented image registration method utilizes relative elasticity constraints of different tissue obtained from whole-body water-fat MRI. The registration method is evaluated by inverse consistency and Dice coefficients and the Imiomics concept is evaluated by example analyses of importance for metabolic research using non-imaging parameters where we know what to expect. The example analyses include whole body imaging atlas creation, anomaly detection, and cross-sectional and longitudinal analysis. RESULTS: The image registration method evaluation on 128 subjects shows low inverse consistency errors and high Dice coefficients. Also, the statistical atlas with fat content intensity values shows low standard deviation values, indicating successful deformations to the common coordinate system. The example analyses show expected associations and correlations which agree with explicit measurements, and thereby illustrate the usefulness of the proposed Imiomics concept. CONCLUSIONS: The registration method is well-suited for Imiomics analyses, which enable analyses of relationships to non-imaging data, e.g. clinical data, in new types of holistic targeted and untargeted big-data analysis.

Dietary intake of eicosapentaenoic and docosahexaenoic acids is linked to gray matter volume and cognitive function in elderly.

In the present study, we tested whether elderly with a high dietary intake of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) would have higher cognitive test scores and greater brain volume than those with low dietary intake of these fatty acids. Data were obtained from the Prospective Investigation of the Vasculature in Uppsala Seniors (PIVUS) cohort. The dietary intake of EPA and DHA was determined by a 7-day food protocol in 252 cognitively healthy elderly (122 females) at the age of 70 years. At age 75, participants' global cognitive function was examined, and their brain volumes were measured by magnetic resonance imaging (MRI). Three different multivariate linear regression models were applied to test our hypothesis: model A (adjusted for gender and age), model B (additionally controlled for lifestyle factors, e.g., education), and model C (further controlled for cardiometabolic factors, e.g., systolic blood pressure). We found that the self-reported 7-day dietary intake of EPA and DHA at the age of 70 years was positively associated with global gray matter volume (P < 0.05, except for model C) and increased global cognitive performance score (P < 0.05). However, no significant associations were observed between the dietary intake of EPA and DHA and global white matter, total brain volume, and regional gray matter, respectively. Further, no effects were observed when examining cognitively impaired (n = 27) elderly as separate analyses. These cross-sectional findings suggest that dietary intake of EPA and DHA may be linked to improved cognitive health in late life but must be confirmed in patient studies.

Whole-body screening of atherosclerosis with magnetic resonance angiography.

With whole-body magnetic resonance angiography (WBMRA), it is possible to examine the whole arterial tree except intracranial and coronary vessels in a single examination without the risks involved in ionizing radiation or arterial cannulation. Whole-body magnetic resonance angiography is well suited for repeated clinical examinations in patients with systemic diseases such as vasculitis or atherosclerosis and can also be used for scientific purposes. On the basis of the WBMRA overview, a possible further development of the WBMRA concept can be to perform further acquisitions at sites with atherosclerotic plaques with higher-resolution scans to determine the degree of stenosis more accurately or to achieve plaque characterization. A total validation of WBMRA compared with digital subtraction angiography (DSA) is not possible owing to the hazards of ionizing radiation. Studies have shown a high sensitivity and specificity for the pelvic and lower limb arteries in comparison with DSA. No systematic validation against DSA has been performed for the renal, aortic, and carotid arteries. Various methods have been used, however, for confirmation of vascular abnormalities found on WBMRA such as ultrasonography, dedicated MRA, or DSA, with reasonably high agreement. The WBMRA method has not been studied with regard to prediction of future cardiovascular (CV) events, as have intima media thickness, coronary artery calcium scoring, and the ankle-brachial index. The full usefulness of WBMRA in an epidemiological setting and as a complementary screening tool for assessing CV risk still needs to be validated against future CV events.

Oxazolones as potent inhibitors of 11beta-hydroxysteroid dehydrogenase type 1.

2,5,5-Trisubstituted oxazolones were identified as potent inhibitors of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1). The synthesis, structure-activity relationship and metabolic stability of these compounds are presented.

Clearance of iron oxide particles in rat liver: effect of hydrated particle size and coating material on liver metabolism.

OBJECTIVES: We sought to evaluate the effect of the particle size and coating material of various iron oxide preparations on the rate of rat liver clearance. MATERIALS AND METHODS: The following iron oxide formulations were used in this study: dextran-coated ferumoxide (size = 97 nm) and ferumoxtran-10 (size = 21 nm), carboxydextran-coated SHU555A (size = 69 nm) and fractionated SHU555A (size = 12 nm), and oxidized-starch coated materials either unformulated NC100150 (size = 15 nm) or formulated NC100150 injection (size = 12 nm). All formulations were administered to 165 rats at 2 dose levels. Quantitative liver R2* values were obtained during a 63-day time period. The concentration of iron oxide particles in the liver was determined by relaxometry, and these values were used to calculate the particle half-lives in the liver. RESULTS: After the administration of a high dose of iron oxide, the half-life of iron oxide particles in rat liver was 8 days for dextran-coated materials, 10 days for carboxydextran materials, 14 days for unformulated oxidized-starch, and 29 days for formulated oxidized-starch. CONCLUSIONS: The results of the study indicate that materials with similar coating but different sizes exhibited similar rates of liver clearance. It was, therefore, concluded that the coating material significantly influences the rate of iron oxide clearance in rat liver.

High in-plane resolution T2-weighted magnetic resonance imaging of acute myocardial ischemia in pigs using the intravascular contrast agent NC100150 injection.

RATIONALE AND OBJECTIVES: The intravascular contrast agent NC100150 injection was tested for its ability to demarcate nonperfused myocardium in a porcine model of coronary occlusion. MATERIALS AND METHODS: A T2-weighted fast spin echo sequence was acquired ex vivo and in vivo during first pass and steady-state circulation of the contrast agent in 2 dosages (2 and 5 mg Fe/kg bw) or saline. RESULTS: Ex vivo, in the high-dose group, the volume of nonperfused myocardium determined from T2-weighted images was 99% of that determined from photographs where perfused myocardium stained with fluorescein. A significantly higher contrast to noise ratio between perfused and nonperfused myocardium was found (both ex and in vivo in steady state) compared with the control group. During first pass, a significant reduction in signal intensity (74 +/- 18%) was found in perfused myocardium after contrast injection. CONCLUSION: NC100150 injection, combined with T2-weighted turbo spin echo imaging, allowed detailed visualization of non-perfused myocardium in the steady state, which corresponded to the area at risk as determined by fluorescein.