Pharmacological Inhibition of Monoacylglycerol O-Acyltransferase 2 Improves Hyperlipidemia, Obesity, and Diabetes by Change in Intestinal Fat Utilization

RESEARCH ARTICLE Pharmacological Inhibition of Monoacylglycerol O-Acyltransferase 2 Improves Hyperlipidemia, Obesity, and Diabetes by Change in Intestinal Fat Utilization a11111 Kazumi Take1*, Taisuke Mochida1, Toshiyuki Maki1, Yoshinori Satomi2, Megumi Hirayama2, Masanori Nakakariya3, Nobuyuki Amano3, Ryutaro Adachi4, Kenjiro Sato1, Tomoyuki Kitazaki1, Shiro Takekawa1 1 Cardiovascular and Metabolic Drug Discovery Unit, Takeda Pharmaceutical Company Limited, Fujisawa, Japan, 2 Integrated Technology Research Laboratories, Takeda Pharmaceutical Company Limited, Fujisawa, Japan, 3 DMPK Research Laboratories, Takeda Pharmaceutical Company Limited, Fujisawa, Japan, 4 Biomolecular Research Laboratories, Takeda Pharmaceutical Company Limited, Fujisawa, Japan OPEN ACCESS Citation: Take K, Mochida T, Maki T, Satomi Y, Hirayama M, Nakakariya M, et al. (2016) Pharmacological Inhibition of Monoacylglycerol OAcyltransferase 2 Improves Hyperlipidemia, Obesity, and Diabetes by Change in Intestinal Fat Utilization. PLoS ONE 11(3): e0150976. doi:10.1371/journal. pone.0150976 Editor: Hiroyuki Itabe, Showa University School of Pharmacy, JAPAN Received: December 13, 2015 Accepted: February 22, 2016 Published: March 3, 2016 Copyright: © 2016 Take et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability Statement: All relevant data are within the paper and its Supporting Information files. Funding: This study was financially supported by Takeda Pharmaceutical Company Limited. Employees of Takeda played roles in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: All authors are employees of Takeda Pharmaceutical Company Limited. CompA used in the authors' study is a product in drug * Abstract Monoacylglycerol O-acyltransferase 2 (MGAT2) catalyzes the synthesis of diacylglycerol (DG), a triacylglycerol precursor and potential peripheral target for novel anti-obesity therapeutics. High-throughput screening identified lead compounds with MGAT2 inhibitory activity. Through structural modification, a potent, selective, and orally bioavailable MGAT2 inhibitor, compound A (compA), was discovered. CompA dose-dependently inhibited postprandial increases in plasma triglyceride (TG) levels. Metabolic flux analysis revealed that compA inhibited triglyceride/diacylglycerol resynthesis in the small intestine and increased free fatty acid and acyl-carnitine with shorter acyl chains than originally labelled fatty acid. CompA decreased high-fat diet (HFD) intake in C57BL/6J mice. MGAT2-null mice showed a similar phenotype as compA-treated mice and compA did not suppress a food intake in MGAT2 KO mice, indicating that the anorectic effects were dependent on MGAT2 inhibition. Chronic administration of compA significantly prevented body weight gain and fat accumulation in mice fed HFD. MGAT2 inhibition by CompA under severe diabetes ameliorated hyperglycemia and fatty liver in HFD-streptozotocin (STZ)-treated mice. Homeostatic model assessments (HOMA-IR) revealed that compA treatment significantly improved insulin sensitivity. The proximal half of the small intestine displayed weight gain following compA treatment. A similar phenomenon has been observed in Roux-en-Y gastric bypass-treated animals and some studies have reported that this intestinal remodeling is essential to the anti-diabetic effects of bariatric surgery. These results clearly demonstrated that MGAT2 inhibition improved dyslipidemia, obesity, and diabetes, suggesting that compA is an effective therapeutic for obesity-related metabolic disorders. PLOS ONE | DOI:10.1371/journal.pone.0150976 March 3, 2016 1 / 18 MGAT2 Inhibitor Improves Obesity and Related Comorbidities by Change in Intestinal Fat Utilization development, and the authors will be filing a patent. There are no further patents, products in development or marketed products to declare. This does not alter the authors’ adherence to all the PLOS ONE policies on sharing data and materials. Introduction Obesity is a major risk factor for type 2 diabetes and cardiovascular disease and is associated with an increase in energy intake relative to energy expenditure [1, 2]. Postprandial dyslipidemia in response to overfeeding with a high-fat diet (HFD) increases fat accumulation, primarily in adipose tissue, and results in obesity [3]. Excess blood lipid levels cause triglyceride (TG) deposition in the skeletal muscle, liver, and pancreas [4]. Ectopic fat storage is closely linked to systemic lipotoxicity, a critical mediator in decreased energy expenditure, insulin resistance, and impaired insulin secretion [5, 6]. TG biosynthesis occurs via 2 major pathways: the monoacylglycerol (MG) pathway and glycerol 3-phosphate pathway [7–9]. In the small intestinal mucosa, the MG pathway accounts for 70%–80% of postprandial TG synthesis [10], with subsequent incorporation of resynthesized TGs into chylomicrons for secretion into the blood and transport to peripheral tissues. Monoacylglycerol O-acyltransferase (MGAT) catalyzes the formation of diacylglycerol (DG), a TG and phospholipid precursor, from 2-monoacylglycerol (2-MG) and fatty acyl-CoA [11]. There are three reported MGAT isoforms in human and rodent genomes [12–15]: MGAT1, mainly expressed in the stomach and kidney but not the small intestine [12], and MGAT2 (rodents and humans) and MGAT3 (humans only), both highly expressed in the small intestine [13–16]. MGAT2 mediates the rate-limiting step in intestinal TG absorption, and MGAT2-null mice have demonstrated that MGAT2 plays an important role in systemic lipid and glucose metabolism [11, 17, 18]. Mice lacking MGAT2 are protected from obesity and insulin resistance induced by HFD [18, 19]. These mice exhibit increased energy expenditure, suggesting that MGAT2 also influences systemic fat utilization [18, 19]. Therefore, MGAT2 is considered a promising pharmacological target for treating obesity and its associated diseases. Here we present the first pharmacological profiling of compound A (compA), a novel and orally active inhibitor of MGAT2 enzymatic activity (IC50 = 7.8 and 2.4 nmol/L for human and mouse MGAT2, respectively) with a good pharmacokinetic profile. Our results support MGAT2 activity inhibition as a potential therapeutic strategy to counteract human obesity, diabetes, and comorbidities related to abnormal lipid metabolism. Methods Materials Structure of compA was shown in Fig 1A. Synthesis of compA was reported previously [20]. This compound exhibited selectivity (greater than 30,000-fold) against related acyltransferases (MGAT3, DGAT1, DGAT2, and ACAT1) [20]. Glycerol-labeled MG (2-oleyl-[1, 1, 2, 3, 3 d5]glycerol) and fatty acid-labeled MG (2-[17, 17, 18, 18, 18 d5]-oleoylglycerol) were purchased fr (...truncated)