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Archive for the ‘Cardiomyopathy’ Category


Christopher J. Lynch, MD, PhD, the New Office of Nutrition Research, Director

Curator: Larry H. Bernstein, MD, FCAP

 

Christopher J. Lynch to direct Office of Nutrition Research

National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)

http://www.nih.gov/news-events/news-releases/christopher-j-lynch-direct-office-nutrition-research

 

Christopher J. Lynch, Ph.D., has been named the new director of the Office of Nutrition Research (ONR) and chief of the Nutrition Research Branch within the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK). Lynch officially assumed his new roles on Feb. 21, 2016. NIDDK is part of the National Institutes of Health.

Lynch will facilitate nutrition research within NIDDK and — through ONR — across NIH, in part by forming and leading a trans-NIH strategic working group. He will also continue and extend ongoing efforts at NIDDK to collaborate widely to advance nutrition research.

“Dr. Lynch is a leader in the nutrition community and his expertise will be vital to guiding the NIH strategic plan for nutrition research,” said NIH Director Francis S. Collins, M.D., Ph.D.  “As NIH works to expand nutrition knowledge, Dr. Lynch’s understanding of the field will help identify information gaps and create a framework to support future discoveries to ultimately improve human health.”

NIH supports a broad range of nutrition research, including studies on the effects of nutrient and dietary intake on human growth and disease, genetic influences on human nutrition and metabolism and other scientific areas. ONR was established in August 2015 to help NIH develop a strategic plan to expand mission-specific nutrition research.

NARRATIVE:
Our laboratory is dedicated to developing cures for metabolic diseases like Obesity, Diabetes and MSUD. We have several projects:
Project 1: How Antipsychotic Drugs Exert Obesity and Metabolic Disease Side effects
Project 2: Impact of Branched Chain Amino Acid (BCAA) signaling and metabolism in obesity and diabetes.
Project 3: Adipose tissue transplant as a treatment for Maple Syrup Urine Disease.
Project 4: How Gastric Bypass Surgery Provides A Rapid Cure For Diabetes And Other Obesity Co-Morbidities Like Hypertension
Project 5: Novel Mechanism Of Action Of Cannabinoid Receptor 1 Blockers For Improvement Of Diabetes

Timeline

  1. Klingerman CM, Stipanovic ME, Hajnal A, Lynch CJ. Acute Metabolic Effects of Olanzapine Depend on Dose and Injection Site. Dose Response. 2015 Oct-Dec; 13(4):1559325815618915.

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  1. Lynch CJ, Kimball SR, Xu Y, Salzberg AC, Kawasawa YI. Global deletion of BCATm increases expression of skeletal muscle genes associated with protein turnover. Physiol Genomics. 2015 Nov; 47(11):569-80.

View in: PubMed

  1. Lynch CJ, Xu Y, Hajnal A, Salzberg AC, Kawasawa YI. RNA sequencing reveals a slow to fast muscle fiber type transition after olanzapine infusion in rats. PLoS One. 2015; 10(4):e0123966.

View in: PubMed

  1. Shin AC, Fasshauer M, Filatova N, Grundell LA, Zielinski E, Zhou JY, Scherer T, Lindtner C, White PJ, Lapworth AL, Ilkayeva O, Knippschild U, Wolf AM, Scheja L, Grove KL, Smith RD, Qian WJ, Lynch CJ, Newgard CB, Buettner C. Brain Insulin Lowers Circulating BCAA Levels by Inducing Hepatic BCAA Catabolism. Cell Metab. 2014 Nov 4; 20(5):898-909.

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  1. Lynch CJ, Adams SH. Branched-chain amino acids in metabolic signalling and insulin resistance. Nat Rev Endocrinol. 2014 Dec; 10(12):723-36.

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  1. Olson KC, Chen G, Xu Y, Hajnal A, Lynch CJ. Alloisoleucine differentiates the branched-chain aminoacidemia of Zucker and dietary obese rats. Obesity (Silver Spring). 2014 May; 22(5):1212-5.

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  1. Zimmerman HA, Olson KC, Chen G, Lynch CJ. Adipose transplant for inborn errors of branched chain amino acid metabolism in mice. Mol Genet Metab. 2013 Aug; 109(4):345-53.

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  1. Olson KC, Chen G, Lynch CJ. Quantification of branched-chain keto acids in tissue by ultra fast liquid chromatography-mass spectrometry. Anal Biochem. 2013 Aug 15; 439(2):116-22.

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  1. She P, Olson KC, Kadota Y, Inukai A, Shimomura Y, Hoppel CL, Adams SH, Kawamata Y, Matsumoto H, Sakai R, Lang CH, Lynch CJ. Leucine and protein metabolism in obese Zucker rats. PLoS One. 2013; 8(3):e59443.

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  1. Lackey DE, Lynch CJ, Olson KC, Mostaedi R, Ali M, Smith WH, Karpe F, Humphreys S, Bedinger DH, Dunn TN, Thomas AP, Oort PJ, Kieffer DA, Amin R, Bettaieb A, Haj FG, Permana P, Anthony TG, Adams SH. Regulation of adipose branched-chain amino acid catabolism enzyme expression and cross-adipose amino acid flux in human obesity. Am J Physiol Endocrinol Metab. 2013 Jun 1; 304(11):E1175-87.

View in: PubMed

  1. Klingerman CM, Stipanovic ME, Bader M, Lynch CJ. Second-generation antipsychotics cause a rapid switch to fat oxidation that is required for survival in C57BL/6J mice. Schizophr Bull. 2014 Mar; 40(2):327-40.

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  1. Carr TD, DiGiovanni J, Lynch CJ, Shantz LM. Inhibition of mTOR suppresses UVB-induced keratinocyte proliferation and survival. Cancer Prev Res (Phila). 2012 Dec; 5(12):1394-404.

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  1. Lynch CJ, Zhou Q, Shyng SL, Heal DJ, Cheetham SC, Dickinson K, Gregory P, Firnges M, Nordheim U, Goshorn S, Reiche D, Turski L, Antel J. Some cannabinoid receptor ligands and their distomers are direct-acting openers of SUR1 K(ATP) channels. Am J Physiol Endocrinol Metab. 2012 Mar 1; 302(5):E540-51.

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  1. Albaugh VL, Singareddy R, Mauger D, Lynch CJ. A double blind, placebo-controlled, randomized crossover study of the acute metabolic effects of olanzapine in healthy volunteers. PLoS One. 2011; 6(8):e22662.

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  1. She P, Zhang Z, Marchionini D, Diaz WC, Jetton TJ, Kimball SR, Vary TC, Lang CH, Lynch CJ. Molecular characterization of skeletal muscle atrophy in the R6/2 mouse model of Huntington’s disease. Am J Physiol Endocrinol Metab. 2011 Jul; 301(1):E49-61.

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  1. Fogle RL, Hollenbeak CS, Stanley BA, Vary TC, Kimball SR, Lynch CJ. Functional proteomic analysis reveals sex-dependent differences in structural and energy-producing myocardial proteins in rat model of alcoholic cardiomyopathy. Physiol Genomics. 2011 Apr 12; 43(7):346-56.

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  1. Zhou Y, Jetton TL, Goshorn S, Lynch CJ, She P. Transamination is required for {alpha}-ketoisocaproate but not leucine to stimulate insulin secretion. J Biol Chem. 2010 Oct 29; 285(44):33718-26.

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  1. Agostino NM, Chinchilli VM, Lynch CJ, Koszyk-Szewczyk A, Gingrich R, Sivik J, Drabick JJ. Effect of the tyrosine kinase inhibitors (sunitinib, sorafenib, dasatinib, and imatinib) on blood glucose levels in diabetic and nondiabetic patients in general clinical practice. J Oncol Pharm Pract. 2011 Sep; 17(3):197-202.

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  1. Li J, Romestaing C, Han X, Li Y, Hao X, Wu Y, Sun C, Liu X, Jefferson LS, Xiong J, Lanoue KF, Chang Z, Lynch CJ, Wang H, Shi Y. Cardiolipin remodeling by ALCAT1 links oxidative stress and mitochondrial dysfunction to obesity. Cell Metab. 2010 Aug 4; 12(2):154-65.

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  1. Culnan DM, Albaugh V, Sun M, Lynch CJ, Lang CH, Cooney RN. Ileal interposition improves glucose tolerance and insulin sensitivity in the obese Zucker rat. Am J Physiol Gastrointest Liver Physiol. 2010 Sep; 299(3):G751-60.

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  1. Hajnal A, Kovacs P, Ahmed T, Meirelles K, Lynch CJ, Cooney RN. Gastric bypass surgery alters behavioral and neural taste functions for sweet taste in obese rats. Am J Physiol Gastrointest Liver Physiol. 2010 Oct; 299(4):G967-79.

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  1. Lang CH, Lynch CJ, Vary TC. BCATm deficiency ameliorates endotoxin-induced decrease in muscle protein synthesis and improves survival in septic mice. Am J Physiol Regul Integr Comp Physiol. 2010 Sep; 299(3):R935-44.

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  1. Albaugh VL, Vary TC, Ilkayeva O, Wenner BR, Maresca KP, Joyal JL, Breazeale S, Elich TD, Lang CH, Lynch CJ. Atypical antipsychotics rapidly and inappropriately switch peripheral fuel utilization to lipids, impairing metabolic flexibility in rodents. Schizophr Bull. 2012 Jan; 38(1):153-66.

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  1. Fogle RL, Lynch CJ, Palopoli M, Deiter G, Stanley BA, Vary TC. Impact of chronic alcohol ingestion on cardiac muscle protein expression. Alcohol Clin Exp Res. 2010 Jul; 34(7):1226-34.

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  1. Lang CH, Frost RA, Bronson SK, Lynch CJ, Vary TC. Skeletal muscle protein balance in mTOR heterozygous mice in response to inflammation and leucine. Am J Physiol Endocrinol Metab. 2010 Jun; 298(6):E1283-94.

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  1. Albaugh VL, Judson JG, She P, Lang CH, Maresca KP, Joyal JL, Lynch CJ. Olanzapine promotes fat accumulation in male rats by decreasing physical activity, repartitioning energy and increasing adipose tissue lipogenesis while impairing lipolysis. Mol Psychiatry. 2011 May; 16(5):569-81.

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  1. Lang CH, Lynch CJ, Vary TC. Alcohol-induced IGF-I resistance is ameliorated in mice deficient for mitochondrial branched-chain aminotransferase. J Nutr. 2010 May; 140(5):932-8.

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  1. She P, Zhou Y, Zhang Z, Griffin K, Gowda K, Lynch CJ. Disruption of BCAA metabolism in mice impairs exercise metabolism and endurance. J Appl Physiol (1985). 2010 Apr; 108(4):941-9.

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  1. Herman MA, She P, Peroni OD, Lynch CJ, Kahn BB. Adipose tissue branched chain amino acid (BCAA) metabolism modulates circulating BCAA levels. J Biol Chem. 2010 Apr 9; 285(15):11348-56.

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  1. Li P, Knabe DA, Kim SW, Lynch CJ, Hutson SM, Wu G. Lactating porcine mammary tissue catabolizes branched-chain amino acids for glutamine and aspartate synthesis. J Nutr. 2009 Aug; 139(8):1502-9.

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  1. Lu G, Sun H, She P, Youn JY, Warburton S, Ping P, Vondriska TM, Cai H, Lynch CJ, Wang Y. Protein phosphatase 2Cm is a critical regulator of branched-chain amino acid catabolism in mice and cultured cells. J Clin Invest. 2009 Jun; 119(6):1678-87.

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  1. Nairizi A, She P, Vary TC, Lynch CJ. Leucine supplementation of drinking water does not alter susceptibility to diet-induced obesity in mice. J Nutr. 2009 Apr; 139(4):715-9.

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  1. Meirelles K, Ahmed T, Culnan DM, Lynch CJ, Lang CH, Cooney RN. Mechanisms of glucose homeostasis after Roux-en-Y gastric bypass surgery in the obese, insulin-resistant Zucker rat. Ann Surg. 2009 Feb; 249(2):277-85.

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  1. Culnan DM, Cooney RN, Stanley B, Lynch CJ. Apolipoprotein A-IV, a putative satiety/antiatherogenic factor, rises after gastric bypass. Obesity (Silver Spring). 2009 Jan; 17(1):46-52.

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  1. She P, Van Horn C, Reid T, Hutson SM, Cooney RN, Lynch CJ. Obesity-related elevations in plasma leucine are associated with alterations in enzymes involved in branched-chain amino acid metabolism. Am J Physiol Endocrinol Metab. 2007 Dec; 293(6):E1552-63.

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  1. She P, Reid TM, Bronson SK, Vary TC, Hajnal A, Lynch CJ, Hutson SM. Disruption of BCATm in mice leads to increased energy expenditure associated with the activation of a futile protein turnover cycle. Cell Metab. 2007 Sep; 6(3):181-94.

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  1. Vary TC, Lynch CJ. Nutrient signaling components controlling protein synthesis in striated muscle. J Nutr. 2007 Aug; 137(8):1835-43.

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  1. Vary TC, Deiter G, Lynch CJ. Rapamycin limits formation of active eukaryotic initiation factor 4F complex following meal feeding in rat hearts. J Nutr. 2007 Aug; 137(8):1857-62.

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  1. Vary TC, Anthony JC, Jefferson LS, Kimball SR, Lynch CJ. Rapamycin blunts nutrient stimulation of eIF4G, but not PKCepsilon phosphorylation, in skeletal muscle. Am J Physiol Endocrinol Metab. 2007 Jul; 293(1):E188-96.

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  1. Vary TC, Lynch CJ. Meal feeding stimulates phosphorylation of multiple effector proteins regulating protein synthetic processes in rat hearts. J Nutr. 2006 Sep; 136(9):2284-90.

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  1. Lynch CJ, Gern B, Lloyd C, Hutson SM, Eicher R, Vary TC. Leucine in food mediates some of the postprandial rise in plasma leptin concentrations. Am J Physiol Endocrinol Metab. 2006 Sep; 291(3):E621-30.

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  1. Albaugh VL, Henry CR, Bello NT, Hajnal A, Lynch SL, Halle B, Lynch CJ. Hormonal and metabolic effects of olanzapine and clozapine related to body weight in rodents. Obesity (Silver Spring). 2006 Jan; 14(1):36-51.

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  1. Vary TC, Lynch CJ. Meal feeding enhances formation of eIF4F in skeletal muscle: role of increased eIF4E availability and eIF4G phosphorylation. Am J Physiol Endocrinol Metab. 2006 Apr; 290(4):E631-42.

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  1. Vary TC, Goodman S, Kilpatrick LE, Lynch CJ. Nutrient regulation of PKCepsilon is mediated by leucine, not insulin, in skeletal muscle. Am J Physiol Endocrinol Metab. 2005 Oct; 289(4):E684-94.

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  1. Vary TC, Lynch CJ. Biochemical approaches for nutritional support of skeletal muscle protein metabolism during sepsis. Nutr Res Rev. 2004 Jun; 17(1):77-88.

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  1. Lynch CJ, Halle B, Fujii H, Vary TC, Wallin R, Damuni Z, Hutson SM. Potential role of leucine metabolism in the leucine-signaling pathway involving mTOR. Am J Physiol Endocrinol Metab. 2003 Oct; 285(4):E854-63.

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  1. Lynch CJ, Hutson SM, Patson BJ, Vaval A, Vary TC. Tissue-specific effects of chronic dietary leucine and norleucine supplementation on protein synthesis in rats. Am J Physiol Endocrinol Metab. 2002 Oct; 283(4):E824-35.

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  1. Lynch CJ, Patson BJ, Anthony J, Vaval A, Jefferson LS, Vary TC. Leucine is a direct-acting nutrient signal that regulates protein synthesis in adipose tissue. Am J Physiol Endocrinol Metab. 2002 Sep; 283(3):E503-13.

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  1. Vary TC, Lynch CJ, Lang CH. Effects of chronic alcohol consumption on regulation of myocardial protein synthesis. Am J Physiol Heart Circ Physiol. 2001 Sep; 281(3):H1242-51.

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  1. Lynch CJ, Patson BJ, Goodman SA, Trapolsi D, Kimball SR. Zinc stimulates the activity of the insulin- and nutrient-regulated protein kinase mTOR. Am J Physiol Endocrinol Metab. 2001 Jul; 281(1):E25-34.

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Global deletion of BCATm increases expression of skeletal muscle genes associated with protein turnover.

Lynch CJ1Kimball SR2Xu Y2Salzberg AC3Kawasawa YI4.   Author information
Physiol Genomics. 2015 Nov;47(11):569-80.  http://dx.doi.org:/10.1152/physiolgenomics.00055.2015

Consumption of a protein-containing meal by a fasted animal promotes protein accretion in skeletal muscle, in part through leucine stimulation of protein synthesis and indirectly through repression of protein degradation mediated by its metabolite, α-ketoisocaproate. Mice lacking the mitochondrial branched-chain aminotransferase (BCATm/Bcat2), which interconverts leucine and α-ketoisocaproate, exhibit elevated protein turnover. Here, the transcriptomes of gastrocnemius muscle from BCATm knockout (KO) and wild-type mice were compared by next-generation RNA sequencing (RNA-Seq) to identify potential adaptations associated with their persistently altered nutrient signaling. Statistically significant changes in the abundance of 1,486/∼39,010 genes were identified. Bioinformatics analysis of the RNA-Seq data indicated that pathways involved in protein synthesis [eukaryotic initiation factor (eIF)-2, mammalian target of rapamycin, eIF4, and p70S6K pathways including 40S and 60S ribosomal proteins], protein breakdown (e.g., ubiquitin mediated), and muscle degeneration (apoptosis, atrophy, myopathy, and cell death) were upregulated. Also in agreement with our previous observations, the abundance of mRNAs associated with reduced body size, glycemia, plasma insulin, and lipid signaling pathways was altered in BCATm KO mice. Consistently, genes encoding anaerobic and/or oxidative metabolism of carbohydrate, fatty acids, and branched chain amino acids were modestly but systematically reduced. Although there was no indication that muscle fiber type was different between KO and wild-type mice, a difference in the abundance of mRNAs associated with a muscular dystrophy phenotype was observed, consistent with the published exercise intolerance of these mice. The results suggest transcriptional adaptations occur in BCATm KO mice that along with altered nutrient signaling may contribute to their previously reported protein turnover, metabolic and exercise phenotypes.

 

RNA sequencing reveals a slow to fast muscle fiber type transition after olanzapine infusion in rats.

Lynch CJ1Xu Y1Hajnal A2Salzberg AC3Kawasawa YI4. Author information
PLoS One. 2015 Apr 20;10(4):e0123966. http://dx.doi.org:/10.1371/journal.pone.0123966. eCollection 2015.

Second generation antipsychotics (SGAs), like olanzapine, exhibit acute metabolic side effects leading to metabolic inflexibility, hyperglycemia, adiposity and diabetes. Understanding how SGAs affect the skeletal muscle transcriptome could elucidate approaches for mitigating these side effects. Male Sprague-Dawley rats were infused intravenously with vehicle or olanzapine for 24h using a dose leading to a mild hyperglycemia. RNA-Seq was performed on gastrocnemius muscle, followed by alignment of the data with the Rat Genome Assembly 5.0. Olanzapine altered expression of 1347 out of 26407 genes. Genes encoding skeletal muscle fiber-type specific sarcomeric, ion channel, glycolytic, O2- and Ca2+-handling, TCA cycle, vascularization and lipid oxidation proteins and pathways, along with NADH shuttles and LDH isoforms were affected. Bioinformatics analyses indicate that olanzapine decreased the expression of slower and more oxidative fiber type genes (e.g., type 1), while up regulating those for the most glycolytic and least metabolically flexible, fast twitch fiber type, IIb. Protein turnover genes, necessary to bring about transition, were also up regulated. Potential upstream regulators were also identified. Olanzapine appears to be rapidly affecting the muscle transcriptome to bring about a change to a fast-glycolytic fiber type. Such fiber types are more susceptible than slow muscle to atrophy, and such transitions are observed in chronic metabolic diseases. Thus these effects could contribute to the altered body composition and metabolic disease olanzapine causes. A potential interventional strategy is implicated because aerobic exercise, in contrast to resistance exercise, can oppose such slow to fast fiber transitions.

 

Brain insulin lowers circulating BCAA levels by inducing hepatic BCAA catabolism.

Shin AC1Fasshauer M1Filatova N1Grundell LA1Zielinski E1Zhou JY2Scherer T1Lindtner C1White PJ3Lapworth AL3,Ilkayeva O3Knippschild U4Wolf AM4Scheja L5Grove KL6Smith RD2Qian WJ2Lynch CJ7Newgard CB3Buettner C8. Author information
Cell Metab. 2014 Nov 4;20(5):898-909. http://dx.doi.org:/10.1016/j.cmet.2014.09.003   Epub 2014 Oct 9

Circulating branched-chain amino acid (BCAA) levels are elevated in obesity/diabetes and are a sensitive predictor for type 2 diabetes. Here we show in rats that insulin dose-dependently lowers plasma BCAA levels through induction of hepatic protein expression and activity of branched-chain α-keto acid dehydrogenase (BCKDH), the rate-limiting enzyme in the BCAA degradation pathway. Selective induction of hypothalamic insulin signaling in rats and genetic modulation of brain insulin receptors in mice demonstrate that brain insulin signaling is a major regulator of BCAA metabolism by inducing hepatic BCKDH. Short-term overfeeding impairs the ability of brain insulin to lower BCAAs in rats. High-fat feeding in nonhuman primates and obesity and/or diabetes in humans is associated with reduced BCKDH protein in liver. These findings support the concept that decreased hepatic BCKDH is a major cause of increased plasma BCAAs and that hypothalamic insulin resistance may account for impaired BCAA metabolism in obesity and diabetes.

 

Branched-chain amino acids in metabolic signalling and insulin resistance.

Lynch CJ1Adams SH2Author information
Nat Rev Endocrinol. 2014 Dec; 10(12):723-36. http://dx.doi.org:/10.1038/nrendo.2014.171

Branched-chain amino acids (BCAAs) are important nutrient signals that have direct and indirect effects. Frequently, BCAAs have been reported to mediate antiobesity effects, especially in rodent models. However, circulating levels of BCAAs tend to be increased in individuals with obesity and are associated with worse metabolic health and future insulin resistance or type 2 diabetes mellitus (T2DM). A hypothesized mechanism linking increased levels of BCAAs and T2DM involves leucine-mediated activation of the mammalian target of rapamycin complex 1 (mTORC1), which results in uncoupling of insulin signalling at an early stage. A BCAA dysmetabolism model proposes that the accumulation of mitotoxic metabolites (and not BCAAs per se) promotes β-cell mitochondrial dysfunction, stress signalling and apoptosis associated with T2DM. Alternatively, insulin resistance might promote aminoacidaemia by increasing the protein degradation that insulin normally suppresses, and/or by eliciting an impairment of efficient BCAA oxidative metabolism in some tissues. Whether and how impaired BCAA metabolism might occur in obesity is discussed in this Review. Research on the role of individual and model-dependent differences in BCAA metabolism is needed, as several genes (BCKDHA, PPM1K, IVD and KLF15) have been designated as candidate genes for obesity and/or T2DM in humans, and distinct phenotypes of tissue-specific branched chain ketoacid dehydrogenase complex activity have been detected in animal models of obesity and T2DM.

 

Leucine and protein metabolism in obese Zucker rats.

She P1Olson KCKadota YInukai AShimomura YHoppel CLAdams SHKawamata YMatsumoto HSakai RLang CHLynch CJAuthor information
PLoS One. 2013;8(3):e59443. http://dx.doi.org:/10.1371/journal.pone.0059443   Epub 2013 Mar 20.

Branched-chain amino acids (BCAAs) are circulating nutrient signals for protein accretion, however, they increase in obesity and elevations appear to be prognostic of diabetes. To understand the mechanisms whereby obesity affects BCAAs and protein metabolism, we employed metabolomics and measured rates of [1-(14)C]-leucine metabolism, tissue-specific protein synthesis and branched-chain keto-acid (BCKA) dehydrogenase complex (BCKDC) activities. Male obese Zucker rats (11-weeks old) had increased body weight (BW, 53%), liver (107%) and fat (∼300%), but lower plantaris and gastrocnemius masses (-21-24%). Plasma BCAAs and BCKAs were elevated 45-69% and ∼100%, respectively, in obese rats. Processes facilitating these rises appeared to include increased dietary intake (23%), leucine (Leu) turnover and proteolysis [35% per g fat free mass (FFM), urinary markers of proteolysis: 3-methylhistidine (183%) and 4-hydroxyproline (766%)] and decreased BCKDC per g kidney, heart, gastrocnemius and liver (-47-66%). A process disposing of circulating BCAAs, protein synthesis, was increased 23-29% by obesity in whole-body (FFM corrected), gastrocnemius and liver. Despite the observed decreases in BCKDC activities per gm tissue, rates of whole-body Leu oxidation in obese rats were 22% and 59% higher normalized to BW and FFM, respectively. Consistently, urinary concentrations of eight BCAA catabolism-derived acylcarnitines were also elevated. The unexpected increase in BCAA oxidation may be due to a substrate effect in liver. Supporting this idea, BCKAs were elevated more in liver (193-418%) than plasma or muscle, and per g losses of hepatic BCKDC activities were completely offset by increased liver mass, in contrast to other tissues. In summary, our results indicate that plasma BCKAs may represent a more sensitive metabolic signature for obesity than BCAAs. Processes supporting elevated BCAA]BCKAs in the obese Zucker rat include increased dietary intake, Leu and protein turnover along with impaired BCKDC activity. Elevated BCAAs/BCKAs may contribute to observed elevations in protein synthesis and BCAA oxidation.

 

Regulation of adipose branched-chain amino acid catabolism enzyme expression and cross-adipose amino acid flux in human obesity.

Lackey DE1Lynch CJOlson KCMostaedi RAli MSmith WHKarpe FHumphreys SBedinger DHDunn TNThomas APOort PJKieffer DAAmin RBettaieb AHaj FGPermana PAnthony TGAdams SH.
Am J Physiol Endocrinol Metab. 2013 Jun 1; 304(11):E1175-87. http://dx.doi.org:/10.1152/ajpendo.00630.2012

Elevated blood branched-chain amino acids (BCAA) are often associated with insulin resistance and type 2 diabetes, which might result from a reduced cellular utilization and/or incomplete BCAA oxidation. White adipose tissue (WAT) has become appreciated as a potential player in whole body BCAA metabolism. We tested if expression of the mitochondrial BCAA oxidation checkpoint, branched-chain α-ketoacid dehydrogenase (BCKD) complex, is reduced in obese WAT and regulated by metabolic signals. WAT BCKD protein (E1α subunit) was significantly reduced by 35-50% in various obesity models (fa/fa rats, db/db mice, diet-induced obese mice), and BCKD component transcripts significantly lower in subcutaneous (SC) adipocytes from obese vs. lean Pima Indians. Treatment of 3T3-L1 adipocytes or mice with peroxisome proliferator-activated receptor-γ agonists increased WAT BCAA catabolism enzyme mRNAs, whereas the nonmetabolizable glucose analog 2-deoxy-d-glucose had the opposite effect. The results support the hypothesis that suboptimal insulin action and/or perturbed metabolic signals in WAT, as would be seen with insulin resistance/type 2 diabetes, could impair WAT BCAA utilization. However, cross-tissue flux studies comparing lean vs. insulin-sensitive or insulin-resistant obese subjects revealed an unexpected negligible uptake of BCAA from human abdominal SC WAT. This suggests that SC WAT may not be an important contributor to blood BCAA phenotypes associated with insulin resistance in the overnight-fasted state. mRNA abundances for BCAA catabolic enzymes were markedly reduced in omental (but not SC) WAT of obese persons with metabolic syndrome compared with weight-matched healthy obese subjects, raising the possibility that visceral WAT contributes to the BCAA metabolic phenotype of metabolically compromised individuals.

 

Some cannabinoid receptor ligands and their distomers are direct-acting openers of SUR1 K(ATP) channels.

Lynch CJ1Zhou QShyng SLHeal DJCheetham SCDickinson KGregory PFirnges MNordheim UGoshorn SReiche D,Turski LAntel J.   Author information
Am J Physiol Endocrinol Metab. 2012 Mar 1;302(5):E540-51.
http://dx.doi.org:/10.1152/ajpendo.00258.2011

Here, we examined the chronic effects of two cannabinoid receptor-1 (CB1) inverse agonists, rimonabant and ibipinabant, in hyperinsulinemic Zucker rats to determine their chronic effects on insulinemia. Rimonabant and ibipinabant (10 mg·kg⁻¹·day⁻¹) elicited body weight-independent improvements in insulinemia and glycemia during 10 wk of chronic treatment. To elucidate the mechanism of insulin lowering, acute in vivo and in vitro studies were then performed. Surprisingly, chronic treatment was not required for insulin lowering. In acute in vivo and in vitro studies, the CB1 inverse agonists exhibited acute K channel opener (KCO; e.g., diazoxide and NN414)-like effects on glucose tolerance and glucose-stimulated insulin secretion (GSIS) with approximately fivefold better potency than diazoxide. Followup studies implied that these effects were inconsistent with a CB1-mediated mechanism. Thus effects of several CB1 agonists, inverse agonists, and distomers during GTTs or GSIS studies using perifused rat islets were unpredictable from their known CB1 activities. In vivo rimonabant and ibipinabant caused glucose intolerance in CB1 but not SUR1-KO mice. Electrophysiological studies indicated that, compared with diazoxide, 3 μM rimonabant and ibipinabant are partial agonists for K channel opening. Partial agonism was consistent with data from radioligand binding assays designed to detect SUR1 K(ATP) KCOs where rimonabant and ibipinabant allosterically regulated ³H-glibenclamide-specific binding in the presence of MgATP, as did diazoxide and NN414. Our findings indicate that some CB1 ligands may directly bind and allosterically regulate Kir6.2/SUR1 K(ATP) channels like other KCOs. This mechanism appears to be compatible with and may contribute to their acute and chronic effects on GSIS and insulinemia.

 

Transamination is required for {alpha}-ketoisocaproate but not leucine to stimulate insulin secretion.

Zhou Y1Jetton TLGoshorn SLynch CJShe PAuthor information
J Biol Chem. 2010 Oct 29;285(44):33718-26. http://dx.doi.org:/10.1074/jbc.M110.136846

It remains unclear how α-ketoisocaproate (KIC) and leucine are metabolized to stimulate insulin secretion. Mitochondrial BCATm (branched-chain aminotransferase) catalyzes reversible transamination of leucine and α-ketoglutarate to KIC and glutamate, the first step of leucine catabolism. We investigated the biochemical mechanisms of KIC and leucine-stimulated insulin secretion (KICSIS and LSIS, respectively) using BCATm(-/-) mice. In static incubation, BCATm disruption abolished insulin secretion by KIC, D,L-α-keto-β-methylvalerate, and α-ketocaproate without altering stimulation by glucose, leucine, or α-ketoglutarate. Similarly, during pancreas perfusions in BCATm(-/-) mice, glucose and arginine stimulated insulin release, whereas KICSIS was largely abolished. During islet perifusions, KIC and 2 mM glutamine caused robust dose-dependent insulin secretion in BCATm(+/+) not BCATm(-/-) islets, whereas LSIS was unaffected. Consistently, in contrast to BCATm(+/+) islets, the increases of the ATP concentration and NADPH/NADP(+) ratio in response to KIC were largely blunted in BCATm(-/-) islets. Compared with nontreated islets, the combination of KIC/glutamine (10/2 mM) did not influence α-ketoglutarate concentrations but caused 120 and 33% increases in malate in BCATm(+/+) and BCATm(-/-) islets, respectively. Although leucine oxidation and KIC transamination were blocked in BCATm(-/-) islets, KIC oxidation was unaltered. These data indicate that KICSIS requires transamination of KIC and glutamate to leucine and α-ketoglutarate, respectively. LSIS does not require leucine catabolism and may be through leucine activation of glutamate dehydrogenase. Thus, KICSIS and LSIS occur by enhancing the metabolism of glutamine/glutamate to α-ketoglutarate, which, in turn, is metabolized to produce the intracellular signals such as ATP and NADPH for insulin secretion.

 

Effect of the tyrosine kinase inhibitors (sunitinib, sorafenib, dasatinib, and imatinib) on blood glucose levels in diabetic and nondiabetic patients in general clinical practice.

Agostino NM1Chinchilli VMLynch CJKoszyk-Szewczyk AGingrich RSivik JDrabick JJ.
J Oncol Pharm Pract. 2011 Sep; 17(3):197-202. http://dx.doi.org:/10.1177/1078155210378913

Tyrosine kinase is a key enzyme activity utilized in many intracellular messaging pathways. Understanding the role of particular tyrosine kinases in malignancies has allowed for the design of tyrosine kinase inhibitors (TKIs), which can target these enzymes and interfere with downstream signaling. TKIs have proven to be successful in the treatment of chronic myeloid leukemia, renal cell carcinoma and gastrointestinal stromal tumor, and other malignancies. Scattered reports have suggested that these agents appear to affect blood glucose (BG). We retrospectively studied the BG concentrations in diabetic (17) and nondiabetic (61) patients treated with dasatinib (8), imatinib (39), sorafenib (23), and sunitinib (30) in our clinical practice. Mean declines of BG were dasatinib (53 mg/dL), imatinib (9 mg/dL), sorafenib (12 mg/dL), and sunitinib (14 mg/dL). All these declines in BG were statistically significant. Of note, 47% (8/17) of the patients with diabetes were able to discontinue their medications, including insulin in some patients. Only one diabetic patient developed symptomatic hypoglycemia while on sunitinib. The mechanism for the hypoglycemic effect of these drugs is unclear, but of the four agents tested, c-kit and PDGFRβ are the common target kinases. Clinicians should keep the potential hypoglycemic effects of these agents in mind; modification of hypoglycemic agents may be required in diabetic patients. These results also suggest that inhibition of a tyrosine kinase, be it c-kit, PDGFRβ or some other undefined target, may improve diabetes mellitus BG control and it deserves further study as a potential novel therapeutic option.

 

Cardiolipin remodeling by ALCAT1 links oxidative stress and mitochondrial dysfunction to obesity.

Li J1Romestaing CHan XLi YHao XWu YSun CLiu XJefferson LSXiong JLanoue KFChang ZLynch CJWang HShi Y.    Author information
Cell Metab. 2010 Aug 4;12(2):154-65. http://dx.doi.org:/10.1016/j.cmet.2010.07.003

Oxidative stress causes mitochondrial dysfunction and metabolic complications through unknown mechanisms. Cardiolipin (CL) is a key mitochondrial phospholipid required for oxidative phosphorylation. Oxidative damage to CL from pathological remodeling is implicated in the etiology of mitochondrial dysfunction commonly associated with diabetes, obesity, and other metabolic diseases. Here, we show that ALCAT1, a lyso-CL acyltransferase upregulated by oxidative stress and diet-induced obesity (DIO), catalyzes the synthesis of CL species that are highly sensitive to oxidative damage, leading to mitochondrial dysfunction, ROS production, and insulin resistance. These metabolic disorders were reminiscent of those observed in type 2 diabetes and were reversed by rosiglitazone treatment. Consequently, ALCAT1 deficiency prevented the onset of DIO and significantly improved mitochondrial complex I activity, lipid oxidation, and insulin signaling in ALCAT1(-/-) mice. Collectively, these findings identify a key role of ALCAT1 in regulating CL remodeling, mitochondrial dysfunction, and susceptibility to DIO.

 

BCATm deficiency ameliorates endotoxin-induced decrease in muscle protein synthesis and improves survival in septic mice.

Lang CH1Lynch CJVary TC.   Author information
Am J Physiol Regul Integr Comp Physiol. 2010 Sep; 299(3):R935-44.
http://dx.doi.org:/10.1152/ajpregu.00297.2010

Endotoxin (LPS) and sepsis decrease mammalian target of rapamycin (mTOR) activity in skeletal muscle, thereby reducing protein synthesis. Our study tests the hypothesis that inhibition of branched-chain amino acid (BCAA) catabolism, which elevates circulating BCAA and stimulates mTOR, will blunt the LPS-induced decrease in muscle protein synthesis. Wild-type (WT) and mitochondrial branched-chain aminotransferase (BCATm) knockout mice were studied 4 h after Escherichia coli LPS or saline. Basal skeletal muscle protein synthesis was increased in knockout mice compared with WT, and this change was associated with increased eukaryotic initiation factor (eIF)-4E binding protein-1 (4E-BP1) phosphorylation, eIF4E.eIF4G binding, 4E-BP1.raptor binding, and eIF3.raptor binding without a change in the mTOR.raptor complex in muscle. LPS decreased muscle protein synthesis in WT mice, a change associated with decreased 4E-BP1 phosphorylation as well as decreased formation of eIF4E.eIF4G, 4E-BP1.raptor, and eIF3.raptor complexes. In BCATm knockout mice given LPS, muscle protein synthesis only decreased to values found in vehicle-treated WT control mice, and this ameliorated LPS effect was associated with a coordinate increase in 4E-BP1.raptor, eIF3.raptor, and 4E-BP1 phosphorylation. Additionally, the LPS-induced increase in muscle cytokines was blunted in BCATm knockout mice, compared with WT animals. In a separate study, 7-day survival and muscle mass were increased in BCATm knockout vs. WT mice after polymicrobial peritonitis. These data suggest that elevating blood BCAA is sufficient to ameliorate the catabolic effect of LPS on skeletal muscle protein synthesis via alterations in protein-protein interactions within mTOR complex-1, and this may provide a survival advantage in response to bacterial infection.

 

Alcohol-induced IGF-I resistance is ameliorated in mice deficient for mitochondrial branched-chain aminotransferase.

Lang CH1Lynch CJVary TCAuthor information
J Nutr. 2010 May;140(5):932-8. http://dx.doi.org:/10.3945/jn.109.120501

Acute alcohol intoxication decreases skeletal muscle protein synthesis by impairing mammalian target of rapamycin (mTOR). In 2 studies, we determined whether inhibition of branched-chain amino acid (BCAA) catabolism ameliorates the inhibitory effect of alcohol on muscle protein synthesis by raising the plasma BCAA concentrations and/or by improving the anabolic response to insulin-like growth factor (IGF)-I. In the first study, 4 groups of mice were used: wild-type (WT) and mitochondrial branched-chain aminotransferase (BCATm) knockout (KO) mice orally administered saline or alcohol (5 g/kg, 1 h). Protein synthesis was greater in KO mice compared with WT controls and was associated with greater phosphorylation of eukaryotic initiation factor (eIF)-4E binding protein-1 (4EBP1), eIF4E-eIF4G binding, and 4EBP1-regulatory associated protein of mTOR (raptor) binding, but not mTOR-raptor binding. Alcohol decreased protein synthesis in WT mice, a change associated with less 4EBP1 phosphorylation, eIF4E-eIF4G binding, and raptor-4EBP1 binding, but greater mTOR-raptor complex formation. Comparable alcohol effects on protein synthesis and signal transduction were detected in BCATm KO mice. The second study used the same 4 groups, but all mice were injected with IGF-I (25 microg/mouse, 30 min). Alcohol impaired the ability of IGF-I to increase muscle protein synthesis, 4EBP1 and 70-kilodalton ribosomal protein S6 kinase-1 phosphorylation, eIF4E-eIF4G binding, and 4EBP1-raptor binding in WT mice. However, in alcohol-treated BCATm KO mice, this IGF-I resistance was not manifested. These data suggest that whereas the sustained elevation in plasma BCAA is not sufficient to ameliorate the catabolic effect of acute alcohol intoxication on muscle protein synthesis, it does improve the anabolic effect of IGF-I.

 

Impact of chronic alcohol ingestion on cardiac muscle protein expression.

Fogle RL1Lynch CJPalopoli MDeiter GStanley BAVary TCAuthor information
Alcohol Clin Exp Res. 2010 Jul;34(7):1226-34. http://dx.doi.org:/10.1111/j.1530-0277.2010.01200.x

BACKGROUND:

Chronic alcohol abuse contributes not only to an increased risk of health-related complications, but also to a premature mortality in adults. Myocardial dysfunction, including the development of a syndrome referred to as alcoholic cardiomyopathy, appears to be a major contributing factor. One mechanism to account for the pathogenesis of alcoholic cardiomyopathy involves alterations in protein expression secondary to an inhibition of protein synthesis. However, the full extent to which myocardial proteins are affected by chronic alcohol consumption remains unresolved.

METHODS:

The purpose of this study was to examine the effect of chronic alcohol consumption on the expression of cardiac proteins. Male rats were maintained for 16 weeks on a 40% ethanol-containing diet in which alcohol was provided both in drinking water and agar blocks. Control animals were pair-fed to consume the same caloric intake. Heart homogenates from control- and ethanol-fed rats were labeled with the cleavable isotope coded affinity tags (ICAT). Following the reaction with the ICAT reagent, we applied one-dimensional gel electrophoresis with in-gel trypsin digestion of proteins and subsequent MALDI-TOF-TOF mass spectrometric techniques for identification of peptides. Differences in the expression of cardiac proteins from control- and ethanol-fed rats were determined by mass spectrometry approaches.

RESULTS:

Initial proteomic analysis identified and quantified hundreds of cardiac proteins. Major decreases in the expression of specific myocardial proteins were observed. Proteins were grouped depending on their contribution to multiple activities of cardiac function and metabolism, including mitochondrial-, glycolytic-, myofibrillar-, membrane-associated, and plasma proteins. Another group contained identified proteins that could not be properly categorized under the aforementioned classification system.

CONCLUSIONS:

Based on the changes in proteins, we speculate modulation of cardiac muscle protein expression represents a fundamental alteration induced by chronic alcohol consumption, consistent with changes in myocardial wall thickness measured under the same conditions.

 

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Calcium Channel Blocker Potential for Angina

Larry H. Bernstein, MD, FCAP, Curator

LPBI

 

 

Pranidipine    

ANTHONY MELVIN CRASTO, PhD

str1

https://newdrugapprovals.files.wordpress.com/2015/12/str116.jpg

 

File:Pranidipine structure.svg

Pranidipine , OPC-13340, FRC 8411

Acalas®

NDA Filing in Japan

A calcium channel blocker potentially for the treatment of angina pectoris and hypertension.

 

CAS No. 99522-79-9

  • Molecular FormulaC25H24N2O6
  • Average mass 448.468

 

see dipine series………..http://organicsynthesisinternational.blogspot.in/p/dipine-series.html

manidipine

 

PAPER

Der Pharmacia Sinica, 2014, 5(1):11-17

https://newdrugapprovals.files.wordpress.com/2015/12/str113.jpg

pelagiaresearchlibrary.com/der-pharmacia-sinica/vol5-iss1/DPS-2014-5-1-11-17.pdf

 

Names
IUPAC name

methyl (2E)-phenylprop-2-en-1-yl 2,6-dimethyl-4-(3-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylate
Other names

2,6-dimethyl-4-(3-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylic acid O5-methyl O3-[(E)-3-phenylprop-2-enyl] ester
Identifiers
99522-79-9 Yes
ChEMBL ChEMBL1096842 
ChemSpider 4940726 
Jmol interactive 3D Image
MeSH C048161
PubChem 6436048
UNII 9DES9QVH58 Yes

 

 

 

PATENT SUBMITTED GRANTED
Process for the preparation of 1,4 – dihydropyridines and novel 1,4-dihydropyridines useful as therapeutic agents [US2003230478] 2003-12-18
Advanced Formulations and Therapies for Treating Hard-to-Heal Wounds [US2014357645] 2014-08-19 2014-12-04
METHODS OF TREATING CARDIOVASCULAR AND METABOLIC DISEASES [US2014322199] 2012-08-06 2014-10-30
Protein Carrier-Linked Prodrugs [US2014323402] 2012-08-10 2014-10-30
sGC STIMULATORS [US2014323448] 2014-04-29 2014-10-30
TREATMENT OF ARTERIAL WALL BY COMBINATION OF RAAS INHIBITOR AND HMG-CoA REDUCTASE INHIBITOR [US2014323536] 2012-12-07 2014-10-30
Agonists of Guanylate Cyclase Useful For the Treatment of Gastrointestinal Disorders, Inflammation, Cancer and Other Disorders [US2014329738] 2014-03-28 2014-11-06
METHODS, COMPOSITIONS, AND KITS FOR THE TREATMENT OF CANCER [US2014335050] 2012-05-25 2014-11-13
ROR GAMMA MODULATORS [US2014343023] 2012-09-18 2014-11-20
High-Loading Water-Soluable Carrier-Linked Prodrugs [US2014296257] 2012-08-10 2014-10-02 

 

 

Synthesis, isolation and use of a common key intermediate for calcium antagonist inhibitors

Neelakandan K.a,b, Manikandan H.b , B. Prabhakarana*, Santosha N.a , Ashok Chaudharia *, Mukund Kulkarnic , Gopalakrishnan Mannathusamyb and Shyam Titirmarea
a API Research Centre, Emcure Pharmaceutical Limited, Hinjawadi, Pune, India bDepartment of Chemistry, Annamalai University, Chidhambaram, India cDepartment of Chemistry, Pune University, Pune, India _________________________________________________________________________________

Pelagia Research Library     www.pelagiaresearchlibrary.com      Der Pharmacia Sinica, 2014, 5(1):11-17

 

The compound (3) synthesized from Nitrobenzaldehyde, tertiary butyl acetoacetate and piperidine can be used as a common intermediate for the production of calcium channel blockers like Nicardipine hydrochloride (1) and Pranidipine hydrochloride (2) with high purity.

 

The last twenty years have witnessed discoveries of calcium antagonists associated with multicoated pharmacodynamics potential which include not only antihypertensive and antiarrhythmic effects of the drugs but also action against excessive calcium entry in the cell of cardiovascular system and subsequent cell damage. Among many classes of calcium channel blockers, 1,4-dihydropyrimidine based drug molecules represented by Felodipine, Clevidipine, Benidipine, Nicardipine and Pranidipine are by far the best to reduce systemic vascular resistance and arterial pressure.

The reported synthetic approaches however proceed with complicated work ups, laborious purification procedures, highly expensive chemicals and low overall yields. (Scheme-I).

Synthetic scheme of Nicardipine and Pranidipine In view of the draw backs associated with previous synthetic approaches there is a strong need for environmentfriendly high yielding process applicable to the multi-kilogram production of calcium antagonist inhibitors. Herein, we report a scalable synthesis for Nicardipine hydrochloride (1) and Pranidipine hydrochloride (2) in fairly high overall yield using key intermediate 3-nitro benzylidene acid (3).Compound (3) was synthesized in two steps using 3-nitrobenzaldehyde, tertiary butyl acetoacetate and piperidine as a base to furnish tertiary butyl ester derivative (10). This was followed by hydrolysis of (10) in TFA and DCM to furnish compound (3) which would serve as a precursor for synthesis of versatile calcium antagonist inhibitors (Scheme-II).

Reported routes for synthesis of Benidipine,1,2 Lercanadipine,3-6 Nimodipine,7-11 Barnidipine12-14 and Manidipine15-16 were explored in our laboratory which involve reaction of nitro benzaldehyde with tertiary butyl acetoacetate using piperidine as a base to get tertiary butyl ester derivative (10). This is further treated with respective reagents to get various calcium channel blockers as shown in scheme 4. Since reported procedures involve in-situ generation of intermediate (3) and its reaction with corresponding fragments, it results in the formation of by-products which ultimately decrease the yield and increase the cost of API.

A novel process of manufacturing benzylidine acid derivative (3) was developed. Use of this intermediate was demonstrated by synthesis of Nicardipine and Pranidipine. This protocol may be employed for synthesis of other calcium channel blockers. In conclusion, a highly efficient, reproducible and scalable process for the synthesis of calcium channel blockers has been developed using (3) as the key intermediate.

 

[1] US 63 365 (Kyowa Hakko; appl.15.4.1982; J-prior.17.4.1981). [2] US 4 448 964 (Kyowa Hakko;15.5.1984; J-prior.17.4.1981). [3] Leonardi, A. et al.: Eur. J. Med.Chem. (EJMCA5) 33,399 (1988). [4] EP 153 016 (Recordati Chem. and Pharm.; appl. 21.1.1985; GB-prior. 14.2.1984). [5] US 4 705 797 (Recordati;10.11.1987; GB-prior. 14.2.1984). [6] WO 9 635 668 (Recordati Chem. and Pharm.; appl. 9.5.1996; I-prior. 12.5.1995). [7] DOS 2 117 571 (Bayer; appl. 10.4.1971). [8] DE 2 117 573 (Bayer; prior.10.4.1971) [9] US 3 799 934 (Bayer;26.3.1974;D-prior.10.4.1971). [10] US 3 932 645 (Bayer;13.1.1976;D-prior.10.4.1971). [11] Meyer, H. et al.: Arzneim.-Forsch. (ARZNAD) 31, 407 (1981); 33, 106 (1983). [12] DE 2 904 552 (Yamanouchi Pharm.; appl. 7.2.1979; J-prior.14.2.1978). [13] US 4 220 649 (Yamanouchi;2.9.1980; J-prior.14.2.1978). [14] CN 85 107 590( Faming Zhuanli Sheqing Gonhali S.; appl. 11.10.1985; J-prior.24.1.1985). [15] EP 94 159 (Takeda; appl. 15.4.1983; J-prior. 10.5.1982). [16] US 4 892 875 (Takeda;9.1.1990; J-prior. 10.5.1982, 11.1.1983).

 

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Biochemistry and Dysmetabolism of Aging and Serious Illness

Curator: Larry H. Bernstein, MD, FCAP

 

White Matter Lipids as a Ketogenic Fuel Supply in Aging Female Brain: Implications for Alzheimer’s Disease

Lauren P. Klosinski, Jia Yao, Fei Yin, Alfred N. Fonteh, Michael G. Harrington, Trace A. Christensen, Eugenia Trushina, Roberta Diaz Brinton
http://www.ebiomedicine.com/article/S2352-3964(15)30192-4/abstract      DOI: http://dx.doi.org/10.1016/j.ebiom.2015.11.002
Highlights
  • Mitochondrial dysfunction activates mechanisms for catabolism of myelin lipids to generate ketone bodies for ATP production.
  • Mechanisms leading to ketone body driven energy production in brain coincide with stages of reproductive aging in females.
  • Sequential activation of myelin catabolism pathway during aging provides multiple therapeutic targets and windows of efficacy.

The mechanisms underlying white matter degeneration, a hallmark of multiple neurodegenerative diseases including Alzheimer’s, remain unclear. Herein we provide a mechanistic pathway, spanning multiple transitions of aging, that links mitochondrial dysfunction early in aging with later age white matter degeneration. Catabolism of myelin lipids to generate ketone bodies can be viewed as an adaptive survival response to address brain fuel and energy demand. Women are at greatest risk of late-onset-AD, thus, our analyses in female brain address mechanisms of AD pathology and therapeutic targets to prevent, delay and treat AD in the sex most affected with potential relevance to men.

 

White matter degeneration is a pathological hallmark of neurodegenerative diseases including Alzheimer’s. Age remains the greatest risk factor for Alzheimer’s and the prevalence of age-related late onset Alzheimer’s is greatest in females. We investigated mechanisms underlying white matter degeneration in an animal model consistent with the sex at greatest Alzheimer’s risk. Results of these analyses demonstrated decline in mitochondrial respiration, increased mitochondrial hydrogen peroxide production and cytosolic-phospholipase-A2 sphingomyelinase pathway activation during female brain aging. Electron microscopic and lipidomic analyses confirmed myelin degeneration. An increase in fatty acids and mitochondrial fatty acid metabolism machinery was coincident with a rise in brain ketone bodies and decline in plasma ketone bodies. This mechanistic pathway and its chronologically phased activation, links mitochondrial dysfunction early in aging with later age development of white matter degeneration. The catabolism of myelin lipids to generate ketone bodies can be viewed as a systems level adaptive response to address brain fuel and energy demand. Elucidation of the initiating factors and the mechanistic pathway leading to white matter catabolism in the aging female brain provides potential therapeutic targets to prevent and treat demyelinating diseases such as Alzheimer’s and multiple sclerosis. Targeting stages of disease and associated mechanisms will be critical.

3. Results

  1. 3.1. Pathway of Mitochondrial Deficits, H2O2 Production and cPLA2 Activation in the Aging Female Brain
  2. 3.2. cPLA2-sphingomyelinase Pathway Activation in White Matter Astrocytes During Reproductive Senescence
  3. 3.3. Investigation of White Matter Gene Expression Profile During Reproductive Senescence
  4. 3.4. Ultra Structural Analysis of Myelin Sheath During Reproductive Senescence
  5. 3.5. Analysis of the Lipid Profile of Brain During the Transition to Reproductive Senescence
  6. 3.6. Fatty Acid Metabolism and Ketone Generation Following the Transition to Reproductive Senescence

 

4. Discussion

Age remains the greatest risk factor for developing AD (Hansson et al., 2006, Alzheimer’s, 2015). Thus, investigation of transitions in the aging brain is a reasoned strategy for elucidating mechanisms and pathways of vulnerability for developing AD. Aging, while typically perceived as a linear process, is likely composed of dynamic transition states, which can protect against or exacerbate vulnerability to AD (Brinton et al., 2015). An aging transition unique to the female is the perimenopausal to menopausal conversion (Brinton et al., 2015). The bioenergetic similarities between the menopausal transition in women and the early appearance of hypometabolism in persons at risk for AD make the aging female a rational model to investigate mechanisms underlying risk of late onset AD.

Findings from this study replicate our earlier findings that age of reproductive senescence is associated with decline in mitochondrial respiration, increased H2O2 production and shift to ketogenic metabolism in brain (Yao et al., 2010, Ding et al., 2013, Yin et al., 2015). These well established early age-related changes in mitochondrial function and shift to ketone body utilization in brain, are now linked to a mechanistic pathway that connects early decline in mitochondrial respiration and H2O2 production to activation of the cPLA2-sphingomyelinase pathway to catabolize myelin lipids resulting in WM degeneration (Fig. 12). These lipids are sequestered in lipid droplets for subsequent use as a local source of ketone body generation via astrocyte mediated beta-oxidation of fatty acids. Astrocyte derived ketone bodies can then be transported to neurons where they undergo ketolysis to generate acetyl-CoA for TCA derived ATP generation required for synaptic and cell function (Fig. 12).

Thumbnail image of Fig. 12. Opens large image

http://www.ebiomedicine.com/cms/attachment/2040395791/2053874721/gr12.sml

Fig. 12

Schematic model of mitochondrial H2O2 activation of cPLA2-sphingomyelinase pathway as an adaptive response to provide myelin derived fatty acids as a substrate for ketone body generation: The cPLA2-sphingomyelinase pathway is proposed as a mechanistic pathway that links an early event, mitochondrial dysfunction and H2O2, in the prodromal/preclinical phase of Alzheimer’s with later stage development of pathology, white matter degeneration. Our findings demonstrate that an age dependent deficit in mitochondrial respiration and a concomitant rise in oxidative stress activate an adaptive cPLA2-sphingomyelinase pathway to provide myelin derived fatty acids as a substrate for ketone body generation to fuel an energetically compromised brain.

Biochemical evidence obtained from isolated whole brain mitochondria confirms that during reproductive senescence and in response to estrogen deprivation brain mitochondria decline in respiratory capacity (Yao et al., 2009, Yao et al., 2010, Brinton, 2008a, Brinton, 2008b, Swerdlow and Khan, 2009). A well-documented consequence of mitochondrial dysfunction is increased production of reactive oxygen species (ROS), specifically H2O2 (Boveris and Chance, 1973, Beal, 2005, Yin et al., 2014, Yap et al., 2009). While most research focuses on the damage generated by free radicals, in this case H2O2 functions as a signaling molecule to activate cPLA2, the initiating enzyme in the cPLA2-sphingomyelinase pathway (Farooqui and Horrocks, 2006, Han et al., 2003, Sun et al., 2004). In AD brain, increased cPLA2 immunoreactivity is detected almost exclusively in astrocytes suggesting that activation of the cPLA2-sphingomyelinase pathway is localized to astrocytes in AD, as opposed to the neuronal or oligodendroglial localization that is observed during apoptosis (Sun et al., 2004, Malaplate-Armand et al., 2006, Di Paolo and Kim, 2011, Stephenson et al., 1996,Stephenson et al., 1999). In our analysis, cPLA2 (Sanchez-Mejia and Mucke, 2010) activation followed the age-dependent rise in H2O2 production and was sustained at an elevated level.

Direct and robust activation of astrocytic cPLA2 by physiologically relevant concentrations of H2O2 was confirmed in vitro. Astrocytic involvement in the cPLA2-sphingomyelinase pathway was also indicated by an increase in cPLA2 positive astrocyte reactivity in WM tracts of reproductively incompetent mice. These data are consistent with findings from brains of persons with AD that demonstrate the same striking localization of cPLA2immunoreactivity within astrocytes, specifically in the hippocampal formation (Farooqui and Horrocks, 2004). While neurons and astrocytes contain endogenous levels of cPLA2, neuronal cPLA2 is activated by an influx of intracellular calcium, whereas astrocytic cPLA2 is directly activated by excessive generation of H2O2 (Sun et al., 2004, Xu et al., 2003, Tournier et al., 1997). Evidence of this cell type specific activation was confirmed by the activation of cPLA2 in astrocytes by H2O2 and the lack of activation in neurons. These data support that astrocytic, not neuronal, cPLA2 is the cellular mediator of the H2O2 dependent cPLA2-sphingomyelinase pathway activation and provide associative evidence supporting a role of astrocytic mitochondrial H2O2 in age-related WM catabolism.

The pattern of gene expression during the shift to reproductive senescence in the female mouse hippocampus recapitulates key observations in human AD brain tissue, specifically elevation in cPLA2, sphingomyelinase and ceramidase (Schaeffer et al., 2010, He et al., 2010, Li et al., 2014). Further, up-regulation of myelin synthesis, lipid metabolism and inflammatory genes in reproductively incompetent female mice is consistent with the gene expression pattern previously reported from aged male rodent hippocampus, aged female non-human primate hippocampus and human AD hippocampus (Blalock et al., 2003, Blalock et al., 2004, Blalock et al., 2010, Blalock et al., 2011, Kadish et al., 2009, Rowe et al., 2007). In these analyses of gene expression in aged male rodent hippocampus, aged female non-human primate hippocampus and human AD hippocampus down regulation of genes related to mitochondrial function, and up-regulation in multiple genes encoding for enzymes involved in ketone body metabolism occurred (Blalock et al., 2003, Blalock et al., 2004, Blalock et al., 2010, Blalock et al., 2011, Kadish et al., 2009, Rowe et al., 2007). The comparability across data derived from aging female mouse hippocampus reported herein and those derived from male rodent brain, female nonhuman brain and human AD brain strongly suggest that cPLA2-sphingomyelinase pathway activation, myelin sheath degeneration and fatty acid metabolism leading to ketone body generation is a metabolic adaptation that is generalizable across these naturally aging models and are evident in aged human AD brain. Collectively, these data support the translational relevance of findings reported herein.

Data obtained via immunohistochemistry, electron microscopy and MBP protein analyses demonstrated an age-related loss in myelin sheath integrity. Evidence for a loss of myelin structural integrity emerged in reproductively incompetent mice following activation of the cPLA2-sphingomyelinase pathway. The unraveling myelin phenotype observed following reproductive senescence and aging reported herein is consistent with the degenerative phenotype that emerges following exposure to the chemotherapy drug bortezomib which induces mitochondrial dysfunction and increased ROS generation (Carozzi et al., 2010, Cavaletti et al., 2007,Ling et al., 2003). In parallel to the decline in myelin integrity, lipid droplet density increased. In aged mice, accumulation of lipid droplets declined in parallel to the rise in ketone bodies consistent with the utilization of myelin-derived fatty acids to generate ketone bodies. Due to the sequential relationship between WM degeneration and lipid droplet formation, we posit that lipid droplets serve as a temporary storage site for myelin-derived fatty acids prior to undergoing β-oxidation in astrocytes to generate ketone bodies.

Microstructural alterations in myelin integrity were associated with alterations in the lipid profile of brain, indicative of WM degeneration resulting in release of myelin lipids. Sphingomyelin and galactocerebroside are two main lipids that compose the myelin sheath (Baumann and Pham-Dinh, 2001). Ceramide is common to both galactocerebroside and sphingomyelin and is composed of sphingosine coupled to a fatty acid. Ceramide levels increase in aging, in states of ketosis and in neurodegeneration (Filippov et al., 2012, Blazquez et al., 1999, Costantini et al., 2005). Specifically, ceramide levels are elevated at the earliest clinically recognizable stage of AD, indicating a degree of WM degeneration early in disease progression (Di Paolo and Kim, 2011,Han et al., 2002, Costantini et al., 2005). Sphingosine is statistically significantly elevated in the brains of AD patients compared to healthy controls; a rise that was significantly correlated with acid sphingomyelinase activity, Aβ levels and tau hyperphosphorylation (He et al., 2010). In our analyses, a rise in ceramides was first observed early in the aging process in reproductively incompetent mice. The rise in ceramides was coincident with the emergence of loss of myelin integrity consistent with the release of myelin ceramides from sphingomyelin via sphingomyelinase activation. Following the rise in ceramides, sphingosine and fatty acid levels increased. The temporal sequence of the lipid profile was consistent with gene expression indicating activation of ceramidase for catabolism of ceramide into sphingosine and fatty acid during reproductive senescence. Once released from ceramide, fatty acids can be transported into the mitochondrial matrix of astrocytes via CPT-1, where β-oxidation of fatty acids leads to the generation of acetyl-CoA (Glatz et al., 2010). It is well documented that acetyl-CoA cannot cross the inner mitochondrial membrane, thus posing a barrier to direct transport of acetyl-CoA generated by β-oxidation into neurons. In response, the newly generated acetyl-CoA undergoes ketogenesis to generate ketone bodies to fuel energy demands of neurons (Morris, 2005,Guzman and Blazquez, 2004, Stacpoole, 2012). Because astrocytes serve as the primary location of β-oxidation in brain they are critical to maintaining neuronal metabolic viability during periods of reduced glucose utilization (Panov et al., 2014, Ebert et al., 2003, Guzman and Blazquez, 2004).

Once fatty acids are released from myelin ceramides, they are transported into astrocytic mitochondria by CPT1 to undergo β-oxidation. The mitochondrial trifunctional protein HADHA catalyzes the last three steps of mitochondrial β-oxidation of long chain fatty acids, while mitochondrial ABAD (aka SCHAD—short chain fatty acid dehydrogenase) metabolizes short chain fatty acids. Concurrent with the release of myelin fatty acids in aged female mice, CPT1, HADHA and ABAD protein expression as well as ketone body generation increased significantly. These findings indicate that astrocytes play a pivotal role in the response to bioenergetic crisis in brain to activate an adaptive compensatory system that activates catabolism of myelin lipids and the metabolism of those lipids into fatty acids to generate ketone bodies necessary to fuel neuronal demand for acetyl-CoA and ATP.

Collectively, these findings provide a mechanistic pathway that links mitochondrial dysfunction and H2O2generation in brain early in the aging process to later stage white matter degeneration. Astrocytes play a pivotal role in providing a mechanistic strategy to address the bioenergetic demand of neurons in the aging female brain. While this pathway is coincident with reproductive aging in the female brain, it is likely to have mechanistic translatability to the aging male brain. Further, the mechanistic link between bioenergetic decline and WM degeneration has potential relevance to other neurological diseases involving white matter in which postmenopausal women are at greater risk, such as multiple sclerosis. The mechanistic pathway reported herein spans time and is characterized by a progression of early adaptive changes in the bioenergetic system of the brain leading to WM degeneration and ketone body production. Translationally, effective therapeutics to prevent, delay and treat WM degeneration during aging and Alzheimer’s disease will need to specifically target stages within the mechanistic pathway described herein. The fundamental initiating event is a bioenergetic switch from being a glucose dependent brain to a glucose and ketone body dependent brain. It remains to be determined whether it is possible to prevent conversion to or reversal of a ketone dependent brain. Effective therapeutic strategies to intervene in this process require biomarkers of bioenergetic phenotype of the brain and stage of mechanistic progression. The mechanistic pathway reported herein may have relevance to other age-related neurodegenerative diseases characterized by white matter degeneration such as multiple sclerosis.

Blood. 2015 Oct 15;126(16):1925-9.    http://dx.doi.org:/10.1182/blood-2014-12-617498. Epub 2015 Aug 14.
Targeting the leukemia cell metabolism by the CPT1a inhibition: functional preclinical effects in leukemias.
Cancer cells are characterized by perturbations of their metabolic processes. Recent observations demonstrated that the fatty acid oxidation (FAO) pathway may represent an alternative carbon source for anabolic processes in different tumors, therefore appearing particularly promising for therapeutic purposes. Because the carnitine palmitoyl transferase 1a (CPT1a) is a protein that catalyzes the rate-limiting step of FAO, here we investigated the in vitro antileukemic activity of the novel CPT1a inhibitor ST1326 on leukemia cell lines and primary cells obtained from patients with hematologic malignancies. By real-time metabolic analysis, we documented that ST1326 inhibited FAO in leukemia cell lines associated with a dose- and time-dependent cell growth arrest, mitochondrial damage, and apoptosis induction. Data obtained on primary hematopoietic malignant cells confirmed the FAO inhibition and cytotoxic activity of ST1326, particularly on acute myeloid leukemia cells. These data suggest that leukemia treatment may be carried out by targeting metabolic processes.
Oncogene. 2015 Oct 12.   http://dx.doi.org:/10.1038/onc.2015.394. [Epub ahead of print]
Tumour-suppression function of KLF12 through regulation of anoikis.
Suppression of detachment-induced cell death, known as anoikis, is an essential step for cancer metastasis to occur. We report here that expression of KLF12, a member of the Kruppel-like family of transcription factors, is downregulated in lung cancer cell lines that have been selected to grow in the absence of cell adhesion. Knockdown of KLF12 in parental cells results in decreased apoptosis following cell detachment from matrix. KLF12 regulates anoikis by promoting the cell cycle transition through S phase and therefore cell proliferation. Reduced expression levels of KLF12 results in increased ability of lung cancer cells to form tumours in vivo and is associated with poorer survival in lung cancer patients. We therefore identify KLF12 as a novel metastasis-suppressor gene whose loss of function is associated with anoikis resistance through control of the cell cycle.
Mol Cell. 2015 Oct 14. pii: S1097-2765(15)00764-9. doi: 10.1016/j.molcel.2015.09.025. [Epub ahead of print]
PEPCK Coordinates the Regulation of Central Carbon Metabolism to Promote Cancer Cell Growth.
Phosphoenolpyruvate carboxykinase (PEPCK) is well known for its role in gluconeogenesis. However, PEPCK is also a key regulator of TCA cycle flux. The TCA cycle integrates glucose, amino acid, and lipid metabolism depending on cellular needs. In addition, biosynthetic pathways crucial to tumor growth require the TCA cycle for the processing of glucose and glutamine derived carbons. We show here an unexpected role for PEPCK in promoting cancer cell proliferation in vitro and in vivo by increasing glucose and glutamine utilization toward anabolic metabolism. Unexpectedly, PEPCK also increased the synthesis of ribose from non-carbohydrate sources, such as glutamine, a phenomenon not previously described. Finally, we show that the effects of PEPCK on glucose metabolism and cell proliferation are in part mediated via activation of mTORC1. Taken together, these data demonstrate a role for PEPCK that links metabolic flux and anabolic pathways to cancer cell proliferation.
Mol Cancer Res. 2015 Oct;13(10):1408-20.   http://dx.doi.org:/10.1158/1541-7786.MCR-15-0048. Epub 2015 Jun 16.
Disruption of Proline Synthesis in Melanoma Inhibits Protein Production Mediated by the GCN2 Pathway.
Many processes are deregulated in melanoma cells and one of those is protein production. Although much is known about protein synthesis in cancer cells, effective ways of therapeutically targeting this process remain an understudied area of research. A process that is upregulated in melanoma compared with normal melanocytes is proline biosynthesis, which has been linked to both oncogene and tumor suppressor pathways, suggesting an important convergent point for therapeutic intervention. Therefore, an RNAi screen of a kinase library was undertaken, identifying aldehyde dehydrogenase 18 family, member A1 (ALDH18A1) as a critically important gene in regulating melanoma cell growth through proline biosynthesis. Inhibition of ALDH18A1, the gene encoding pyrroline-5-carboxylate synthase (P5CS), significantly decreased cultured melanoma cell viability and tumor growth. Knockdown of P5CS using siRNA had no effect on apoptosis, autophagy, or the cell cycle but cell-doubling time increased dramatically suggesting that there was a general slowdown in cellular metabolism. Mechanistically, targeting ALDH18A1 activated the serine/threonine protein kinase GCN2 (general control nonderepressible 2) to inhibit protein synthesis, which could be reversed with proline supplementation. Thus, targeting ALDH18A1 in melanoma can be used to disrupt proline biosynthesis to limit cell metabolism thereby increasing the cellular doubling time mediated through the GCN2 pathway.  This study demonstrates that melanoma cells are sensitive to disruption of proline synthesis and provides a proof-of-concept that the proline synthesis pathway can be therapeutically targeted in melanoma tumors for tumor inhibitory efficacy. Mol Cancer Res; 13(10); 1408-20. ©2015 AACR.
SDHB-Deficient Cancers: The Role of Mutations That Impair Iron Sulfur Cluster Delivery.
BACKGROUND:  Mutations in the Fe-S cluster-containing SDHB subunit of succinate dehydrogenase cause familial cancer syndromes. Recently the tripeptide motif L(I)YR was identified in the Fe-S recipient protein SDHB, to which the cochaperone HSC20 binds.
METHODS:   In order to characterize the metabolic basis of SDH-deficient cancers we performed stable isotope-resolved metabolomics in a novel SDHB-deficient renal cell carcinoma cell line and conducted bioinformatics and biochemical screening to analyze Fe-S cluster acquisition and assembly of SDH in the presence of other cancer-causing SDHB mutations.

RESULTS:

We found that the SDHB(R46Q) mutation in UOK269 cells disrupted binding of HSC20, causing rapid degradation of SDHB. In the absence of SDHB, respiration was undetectable in UOK269 cells, succinate was elevated to 351.4±63.2 nmol/mg cellular protein, and glutamine became the main source of TCA cycle metabolites through reductive carboxylation. Furthermore, HIF1α, but not HIF2α, increased markedly and the cells showed a strong DNA CpG island methylator phenotype (CIMP). Biochemical and bioinformatic screening revealed that 37% of disease-causing missense mutations in SDHB were located in either the L(I)YR Fe-S transfer motifs or in the 11 Fe-S cluster-ligating cysteines.

CONCLUSIONS:

These findings provide a conceptual framework for understanding how particular mutations disproportionately cause the loss of SDH activity, resulting in accumulation of succinate and metabolic remodeling in SDHB cancer syndromes.

 

SR4 Uncouples Mitochondrial Oxidative Phosphorylation, Modulates AMPK-mTOR Signaling, and Inhibits Proliferation of HepG2 Hepatocarcinoma Cells

  1. L. Figarola, J. Singhal, J. D. Tompkins, G. W. Rogers, C. Warden, D. Horne, A. D. Riggs, S. Awasthi and S. S. Singhal.

J Biol Chem. 2015 Nov 3, [epub ahead of print]

 

CD47 Receptor Globally Regulates Metabolic Pathways That Control Resistance to Ionizing Radiation

  1. W. Miller, D. R. Soto-Pantoja, A. L. Schwartz, J. M. Sipes, W. G. DeGraff, L. A. Ridnour, D. A. Wink and D. D. Roberts.

J Biol Chem. 2015 Oct 9, 290 (41): 24858-74.

 

Knockdown of PKM2 Suppresses Tumor Growth and Invasion in Lung Adenocarcinoma

  1. Sun, A. Zhu, L. Zhang, J. Zhang, Z. Zhong and F. Wang.

Int J Mol Sci. 2015 Oct 15, 16 (10): 24574-87.

 

EglN2 associates with the NRF1-PGC1alpha complex and controls mitochondrial function in breast cancer

  1. Zhang, C. Wang, X. Chen, M. Takada, C. Fan, X. Zheng, H. Wen, Y. Liu, C. Wang, R. G. Pestell, K. M. Aird, W. G. Kaelin, Jr., X. S. Liu and Q. Zhang.

EMBO J. 2015 Oct 22, [epub ahead of print]

 

Mitochondrial Genetics Regulate Breast Cancer Tumorigenicity and Metastatic Potential.

Current paradigms of carcinogenic risk suggest that genetic, hormonal, and environmental factors influence an individual’s predilection for developing metastatic breast cancer. Investigations of tumor latency and metastasis in mice have illustrated differences between inbred strains, but the possibility that mitochondrial genetic inheritance may contribute to such differences in vivo has not been directly tested. In this study, we tested this hypothesis in mitochondrial-nuclear exchange mice we generated, where cohorts shared identical nuclear backgrounds but different mtDNA genomes on the background of the PyMT transgenic mouse model of spontaneous mammary carcinoma. In this setting, we found that primary tumor latency and metastasis segregated with mtDNA, suggesting that mtDNA influences disease progression to a far greater extent than previously appreciated. Our findings prompt further investigation into metabolic differences controlled by mitochondrial process as a basis for understanding tumor development and metastasis in individual subjects. Importantly, differences in mitochondrial DNA are sufficient to fundamentally alter disease course in the PyMT mouse mammary tumor model, suggesting that functional metabolic differences direct early tumor growth and metastatic efficiency. Cancer Res; 75(20); 4429-36. ©2015 AACR.

 

Cancer Lett. 2015 Oct 29. pii: S0304-3835(15)00656-4.    http://dx.doi.org:/10.1016/j.canlet.2015.10.025. [Epub ahead of print]
Carboxyamidotriazole inhibits oxidative phosphorylation in cancer cells and exerts synergistic anti-cancer effect with glycolysis inhibition.

Targeting cancer cell metabolism is a promising strategy against cancer. Here, we confirmed that the anti-cancer drug carboxyamidotriazole (CAI) inhibited mitochondrial respiration in cancer cells for the first time and found a way to enhance its anti-cancer activity by further disturbing the energy metabolism. CAI promoted glucose uptake and lactate production when incubated with cancer cells. The oxidative phosphorylation (OXPHOS) in cancer cells was inhibited by CAI, and the decrease in the activity of the respiratory chain complex I could be one explanation. The anti-cancer effect of CAI was greatly potentiated when being combined with 2-deoxyglucose (2-DG). The cancer cells treated with the combination of CAI and 2-DG were arrested in G2/M phase. The apoptosis and necrosis rates were also increased. In a mouse xenograft model, this combination was well tolerated and retarded the tumor growth. The impairment of cancer cell survival was associated with significant cellular ATP decrease, suggesting that the combination of CAI and 2-DG could be one of the strategies to cause dual inhibition of energy pathways, which might be an effective therapeutic approach for a broad spectrum of tumors.

 

Cancer Immunol Res. 2015 Nov;3(11):1236-47.    http://dx.doi.org:/10.1158/2326-6066.CIR-15-0036. Epub 2015 May 29.
Inhibition of Fatty Acid Oxidation Modulates Immunosuppressive Functions of Myeloid-Derived Suppressor Cells and Enhances Cancer Therapies.

Myeloid-derived suppressor cells (MDSC) promote tumor growth by inhibiting T-cell immunity and promoting malignant cell proliferation and migration. The therapeutic potential of blocking MDSC in tumors has been limited by their heterogeneity, plasticity, and resistance to various chemotherapy agents. Recent studies have highlighted the role of energy metabolic pathways in the differentiation and function of immune cells; however, the metabolic characteristics regulating MDSC remain unclear. We aimed to determine the energy metabolic pathway(s) used by MDSC, establish its impact on their immunosuppressive function, and test whether its inhibition blocks MDSC and enhances antitumor therapies. Using several murine tumor models, we found that tumor-infiltrating MDSC (T-MDSC) increased fatty acid uptake and activated fatty acid oxidation (FAO). This was accompanied by an increased mitochondrial mass, upregulation of key FAO enzymes, and increased oxygen consumption rate. Pharmacologic inhibition of FAO blocked immune inhibitory pathways and functions in T-MDSC and decreased their production of inhibitory cytokines. FAO inhibition alone significantly delayed tumor growth in a T-cell-dependent manner and enhanced the antitumor effect of adoptive T-cell therapy. Furthermore, FAO inhibition combined with low-dose chemotherapy completely inhibited T-MDSC immunosuppressive effects and induced a significant antitumor effect. Interestingly, a similar increase in fatty acid uptake and expression of FAO-related enzymes was found in human MDSC in peripheral blood and tumors. These results support the possibility of testing FAO inhibition as a novel approach to block MDSC and enhance various cancer therapies. Cancer Immunol Res; 3(11); 1236-47. ©2015 AACR.

 

Ionizing radiation induces myofibroblast differentiation via lactate dehydrogenase

  1. L. Judge, K. M. Owens, S. J. Pollock, C. F. Woeller, T. H. Thatcher, J. P. Williams, R. P. Phipps, P. J. Sime and R. M. Kottmann.

Am J Physiol Lung Cell Mol Physiol. 2015 Oct 15, 309 (8): L879-87.

 

Vitamin C selectively kills KRAS and BRAF mutant colorectal cancer cells by targeting GAPDH

  1. Yun, E. Mullarky, C. Lu, K. N. Bosch, A. Kavalier, K. Rivera, J. Roper, Chio, II, E. G. Giannopoulou, C. Rago, A. Muley, J. M. Asara, J. Paik, O. Elemento, Z. Chen, D. J. Pappin, L. E. Dow, N. Papadopoulos, S. S. Gross and L. C. Cantley.

Science. 2015 Nov 5, [epub ahead of print]

 

Down-regulation of FBP1 by ZEB1-mediated repression confers to growth and invasion in lung cancer cells

  1. Zhang, J. Wang, H. Xing, Q. Li, Q. Zhao and J. Li.

Mol Cell Biochem. 2015 Nov 6, [epub ahead of print]

 

J Mol Cell Cardiol. 2015 Oct 23. pii: S0022-2828(15)30073-0.     http://dx.doi.org:/10.1016/j.yjmcc.2015.10.002. [Epub ahead of print]
GRK2 compromises cardiomyocyte mitochondrial function by diminishing fatty acid-mediated oxygen consumption and increasing superoxide levels.

The G protein-coupled receptor kinase-2 (GRK2) is upregulated in the injured heart and contributes to heart failure pathogenesis. GRK2 was recently shown to associate with mitochondria but its functional impact in myocytes due to this localization is unclear. This study was undertaken to determine the effect of elevated GRK2 on mitochondrial respiration in cardiomyocytes. Sub-fractionation of purified cardiac mitochondria revealed that basally GRK2 is found in multiple compartments. Overexpression of GRK2 in mouse cardiomyocytes resulted in an increased amount of mitochondrial-based superoxide. Inhibition of GRK2 increased oxygen consumption rates and ATP production. Moreover, fatty acid oxidation was found to be significantly impaired when GRK2 was elevated and was dependent on the catalytic activity and mitochondrial localization of this kinase. Our study shows that independent of cardiac injury, GRK2 is localized in the mitochondria and its kinase activity negatively impacts the function of this organelle by increasing superoxide levels and altering substrate utilization for energy production.

 

Br J Pharmacol. 2015 Oct 27. doi: 10.1111/bph.13377. [Epub ahead of print]
All-trans retinoic acid protects against doxorubicin-induced cardiotoxicity by activating the Erk2 signalling pathway.
BACKGROUND AND PURPOSE:

Doxorubicin (Dox) is a powerful antineoplastic agent for treating a wide range of cancers. However, doxorubicin cardiotoxicity of the heart has largely limited its clinical use. It is known that all-trans retinoic acid (ATRA) plays important roles in many cardiac biological processes, however, the protective effects of ATRA on doxorubicin cardiotoxicity remain unknown. Here, we studied the effect of ATRA on doxorubicin cardiotoxicity and underlying mechanisms.

EXPERIMENTAL APPROACHES:

Cellular viability assays, western blotting and mitochondrial respiration analyses were employed to evaluate the cellular response to ATRA in H9c2 cells and primary cardiomyocytes. Quantitative PCR (Polymerase Chain Reaction) and gene knockdown were performed to investigate the underlying molecular mechanisms of ATRA’s effects on doxorubicin cardiotoxicity.

KEY RESULTS:

ATRA significantly inhibited doxorubicin-induced apoptosis in H9c2 cells and primary cardiomyocytes. ATRA was more effective against doxorubicin cardiotoxicity than resveratrol and dexrazoxane. ATRA also suppressed reactive oxygen species (ROS) generation, and restored the expression level of mRNA and proteins in phase II detoxifying enzyme system: Nrf2 (nuclear factor-E2-related factor 2), MnSOD (manganese superoxide dismutase), HO-1 (heme oxygenase1) as well as mitochondrial function (mitochondrial membrane integrity, mitochondrial DNA copy numbers, mitochondrial respiration capacity, biogenesis and dynamics). Both Erk1/2 (extracellular signal-regulated kinase1/2) inhibitor (U0126) and Erk2 siRNA, but not Erk1 siRNA, abolished the protective effect of ATRA against doxorubicin-induced toxicity in H9c2 cells. Remarkably, ATRA did not compromise the anticancer efficacy of doxorubicin in gastric carcinoma cells.

CONCLUSION AND IMPLICATION:

ATRA protected cardiomyocytes against doxorubicin-induced toxicity by activating the Erk2 pathway without compromising the anticancer efficacy of doxorubicin. Therefore, ATRA may be a promising candidate as a cardioprotective agent against doxorubicin cardiotoxicity.

 

Proteomic and Biochemical Studies of Lysine Malonylation Suggest Its Malonic Aciduria-associated Regulatory Role in Mitochondrial Function and Fatty Acid Oxidation

  1. Colak, O. Pougovkina, L. Dai, M. Tan, H. Te Brinke, H. Huang, Z. Cheng, J. Park, X. Wan, X. Liu, W. W. Yue, R. J. Wanders, J. W. Locasale, D. B. Lombard, V. C. de Boer and Y. Zhao.

Mol Cell Proteomics. 2015 Nov 1, 14 (11): 3056-71.

 

Foxg1 localizes to mitochondria and coordinates cell differentiation and bioenergetics

  1. Pancrazi, G. Di Benedetto, L. Colombaioni, G. Della Sala, G. Testa, F. Olimpico, A. Reyes, M. Zeviani, T. Pozzan and M. Costa.

Proc Natl Acad Sci U S A. 2015 Oct 27, 112(45): 13910-5.

 

Evidence of Mitochondrial Dysfunction within the Complex Genetic Etiology of Schizophrenia

  1. E. Hjelm, B. Rollins, F. Mamdani, J. C. Lauterborn, G. Kirov, G. Lynch, C. M. Gall, A. Sequeira and M. P. Vawter.

Mol Neuropsychiatry. 2015 Nov 1, 1 (4): 201-219.

 

Metabolic Reprogramming Is Required for Myofibroblast Contractility and Differentiation

  1. Bernard, N. J. Logsdon, S. Ravi, N. Xie, B. P. Persons, S. Rangarajan, J. W. Zmijewski, K. Mitra, G. Liu, V. M. Darley-Usmar and V. J. Thannickal.

J Biol Chem. 2015 Oct 16, 290 (42): 25427-38.

 

J Biol Chem. 2015 Oct 23;290(43):25834-46.    http://dx.doi.org:/10.1074/jbc.M115.658815. Epub 2015 Sep 4.
Kinome Screen Identifies PFKFB3 and Glucose Metabolism as Important Regulators of the Insulin/Insulin-like Growth Factor (IGF)-1 Signaling Pathway.

The insulin/insulin-like growth factor (IGF)-1 signaling pathway (ISP) plays a fundamental role in long term health in a range of organisms. Protein kinases including Akt and ERK are intimately involved in the ISP. To identify other kinases that may participate in this pathway or intersect with it in a regulatory manner, we performed a whole kinome (779 kinases) siRNA screen for positive or negative regulators of the ISP, using GLUT4 translocation to the cell surface as an output for pathway activity. We identified PFKFB3, a positive regulator of glycolysis that is highly expressed in cancer cells and adipocytes, as a positive ISP regulator. Pharmacological inhibition of PFKFB3 suppressed insulin-stimulated glucose uptake, GLUT4 translocation, and Akt signaling in 3T3-L1 adipocytes. In contrast, overexpression of PFKFB3 in HEK293 cells potentiated insulin-dependent phosphorylation of Akt and Akt substrates. Furthermore, pharmacological modulation of glycolysis in 3T3-L1 adipocytes affected Akt phosphorylation. These data add to an emerging body of evidence that metabolism plays a central role in regulating numerous biological processes including the ISP. Our findings have important implications for diseases such as type 2 diabetes and cancer that are characterized by marked disruption of both metabolism and growth factor signaling.

 

FASEB J. 2015 Oct 19.    http://dx.doi.org:/pii: fj.15-276360. [Epub ahead of print]
Perm1 enhances mitochondrial biogenesis, oxidative capacity, and fatigue resistance in adult skeletal muscle.

Skeletal muscle mitochondrial content and oxidative capacity are important determinants of muscle function and whole-body health. Mitochondrial content and function are enhanced by endurance exercise and impaired in states or diseases where muscle function is compromised, such as myopathies, muscular dystrophies, neuromuscular diseases, and age-related muscle atrophy. Hence, elucidating the mechanisms that control muscle mitochondrial content and oxidative function can provide new insights into states and diseases that affect muscle health. In past studies, we identified Perm1 (PPARGC1- and ESRR-induced regulator, muscle 1) as a gene induced by endurance exercise in skeletal muscle, and regulating mitochondrial oxidative function in cultured myotubes. The capacity of Perm1 to regulate muscle mitochondrial content and function in vivo is not yet known. In this study, we use adeno-associated viral (AAV) vectors to increase Perm1 expression in skeletal muscles of 4-wk-old mice. Compared to control vector, AAV1-Perm1 leads to significant increases in mitochondrial content and oxidative capacity (by 40-80%). Moreover, AAV1-Perm1-transduced muscles show increased capillary density and resistance to fatigue (by 33 and 31%, respectively), without prominent changes in fiber-type composition. These findings suggest that Perm1 selectively regulates mitochondrial biogenesis and oxidative function, and implicate Perm1 in muscle adaptations that also occur in response to endurance exercise.-Cho, Y., Hazen, B. C., Gandra, P. G., Ward, S. R., Schenk, S., Russell, A. P., Kralli, A. Perm1 enhances mitochondrial biogenesis, oxidative capacity, and fatigue resistance in adult skeletal muscle.

 

A conserved MADS-box phosphorylation motif regulates differentiation and mitochondrial function in skeletal, cardiac, and smooth muscle cells.
Exposure to metabolic disease during fetal development alters cellular differentiation and perturbs metabolic homeostasis, but the underlying molecular regulators of this phenomenon in muscle cells are not completely understood. To address this, we undertook a computational approach to identify cooperating partners of the myocyte enhancer factor-2 (MEF2) family of transcription factors, known regulators of muscle differentiation and metabolic function. We demonstrate that MEF2 and the serum response factor (SRF) collaboratively regulate the expression of numerous muscle-specific genes, including microRNA-133a (miR-133a). Using tandem mass spectrometry techniques, we identify a conserved phosphorylation motif within the MEF2 and SRF Mcm1 Agamous Deficiens SRF (MADS)-box that regulates miR-133a expression and mitochondrial function in response to a lipotoxic signal. Furthermore, reconstitution of MEF2 function by expression of a neutralizing mutation in this identified phosphorylation motif restores miR-133a expression and mitochondrial membrane potential during lipotoxicity. Mechanistically, we demonstrate that miR-133a regulates mitochondrial function through translational inhibition of a mitophagy and cell death modulating protein, called Nix. Finally, we show that rodents exposed to gestational diabetes during fetal development display muscle diacylglycerol accumulation, concurrent with insulin resistance, reduced miR-133a, and elevated Nix expression, as young adult rats. Given the diverse roles of miR-133a and Nix in regulating mitochondrial function, and proliferation in certain cancers, dysregulation of this genetic pathway may have broad implications involving insulin resistance, cardiovascular disease, and cancer biology.

 

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Empagliflozin

Larry H. Bernstein, MD, FCAP, Curator

LPBI

 

Empagliflozin Benefits in EMPA-REG Explored in Diabetics Initially With or Without Heart Failure

Marlene Busko

http://www.medscape.com/viewarticle/854542

 

ORLANDO, FL — Patients with type 2 diabetes and established CVD who received the antidiabetic sodium-glucose cotransporter 2 (SGLT2) inhibitor empagliflozin (Jardiance, Lilly/Boehringer Ingelheim), as opposed to placebo, had a reduced risk of being hospitalized for heart failure or dying from CVD during a median follow-up of 3.1 years. The finding was strongest in patients without heart failure at baseline[1]. The finding is noteworthy in part because associated heart failure has been a concern, justified or not, with some other diabetes medications.

In these high-risk patients, empagliflozin resulted in a “consistent benefit” in these outcomes, Dr Silvio E Inzucchi (Yale University School of Medicine, New Haven, CT) said, presenting these findings from a prespecified secondary analysis of the EMPA-REG OUTCOME trial at theAmerican Heart Association (AHA) 2015 Scientific Sessions.

Unlike the gasps and applause that greeted him when he presented the trial’s primary outcome results at the European Association for the Study of Diabetes (EASD) 2015 Meeting in Stockholm in mid-September, the audience reaction this time was more measured. The trial had also been published at about the time of its EASD presentation [2].

The principal findings showed that compared with patients who took placebo, those who were randomized to empagliflozin had a 38% (P<0.001) reduced risk of CV death and a 35% P=0.002) reduced risk of hospitalization for HF, at a median follow-up of 3.1 years.

In the current secondary analysis, the 90% of patients who were free of heart failure at study entry showed a steep and significant drop in HF hospitalizations during the trial. There was also a drop in HF hospitalizations with active therapy in the minority who had HF at baseline, but it failed to reach significance.

“I think metformin is likely to remain our first-line oral therapy for patients with type 2 diabetes,” Dr Donald M Lloyd-Jones (Northwestern University Feinberg School of Medicine, Chicago, IL), cochair at an AHA press briefing, told heartwire from Medscape. “There is an alphabet soup of diabetes medications,” with multiple agents that effectively lower blood glucose and reduce patients’ risk of retinopathy, nephropathy, and neuropathy.

However, “it was . . . unexpected that [empagliflozin], as reported recently [at the EASD meeting and] in the New England Journal of Medicine [has an] effect on CV death and other CV events.” This is still an early stage of research, he cautioned, and it is not known how the drug exerts its CV effects and whether there is a class effect. “But [this] could be a game changer, because we would love to have [antidiabetic] medications that not only control blood sugar but also reduce death and [other] hard events,” he said.

 

First CV Outcomes Trial in this Drug Class

Until now, none of the antiglycemic medications has also been shown to improve HF outcomes, Inzucchi explained. “We’ve actually been searching decades for a diabetes medicine that will not only lower blood glucose but also reduce cardiovascular complications,” he said in a press briefing. “And I would remind you that based on the 2008 FDA guidance to industry, all new diabetes medications need to be tested for cardiovascular safety before being allowed on the market,” he added.

EMPA-REG OUTCOME is the first published, large CV-outcome trial of an SGLT-2 inhibitor.

As previously described, the trial randomized 7028 adult patients who had type 2 diabetes and established CVD to receive 10 mg/day or 25 mg/day empagliflozin or placebo. The CVD included prior MI (46.6%), CABG (24.8%), stroke (23.3%), and peripheral artery disease (PAD) (20.8%).

The patients were also required to have an HbA1c level between 7% and 10%, body-mass index (BMI) <45, and, because the drug exerts its effects via the kidney, estimated glomerular filtration rate (eGFR) >30 mL/min/1.73 m2.

“Importantly, study medication was given upon a backdrop of standard care—antihyperglycemia therapy, as well as other evidence-based cardiovascular therapies such as statins, ACE inhibitors, and aspirin,” Inzucchi stressed.

 

Spotlight on HF Outcomes

The current analysis dove deeper into the heart-failure outcomes in the trial.

The risk of hospitalization for HF or CV death was consistently significantly lower in patients who received empagliflozin vs placebo, in subgroup analyses related to age, kidney function, and medication use (ACE inhibitors/angiotensin receptor blockers [ARBs], diuretics, beta-blockers, or mineralocorticoid-receptor antagonists).

Overall, the patients who received empagliflozin had a 34% reduced risk of being hospitalized for HF or dying from CV causes and a 39% reduced risk of being hospitalized for or dying from HF.

Risk of Hospitalization or Death, Empagliflozin vs Placebo

Outcome HR (95% CI) P
Hospitalization for HF or CV death 0.66 (0.55–0.79) <0.00001
Hospitalization for or death from HF 0.61 (0.47–0.79) <0.00001

Most patients (90%) did not have HF at baseline.

In the patients without HF at baseline, “as you might expect, [HF] hospitalizations were relatively small in number” (1.8% of patients on the study drug and 3.1% of patients on placebo), said Inzucchi. There was a statistically significant 41% reduced risk of HF hospitalization in patients without HF at baseline on the study drug vs placebo (HR 0.59, 95% CI 0.43–0.82).

In the smaller number of patients who did have HF at baseline, the rate of hospitalizations for HF was much higher (10.4% of patients on the study drug and 12.3% of patients on placebo). But in this case, the difference between patients on the study drug vs placebo was not statistically significant (HR 0.75, 95% CI 0.48–1.19).

The results were similar when the analysis was repeated for the combined outcome of hospitalization for HF or CV death.

“Not surprisingly,” adverse events were more common in sicker patients with baseline HF; genital infections, a well-known adverse event in drugs that increase glucose in the urine, were three times more common in those patients, said Inzucchi.

“I think these are very compelling data, but early days,” said Lloyd-Jones.

Inzucchi receives research grants from Genzyme and honoraria from Boehringer Ingelheim, Merck Sharp & Dome, Sanofi, Amgen, and Genzyme, and he is a consultant on advisory boards for Boehringer Ingelheim, Sanofi, and Amgen. Disclosures for the coauthors are listed in the abstract. Lloyd-Jones has no relevant financial relationships.

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MYBPC3 gene and the heart

Larry H. Bernstein, MD, FCAP, Curator

LPBI

 

MYBPC3 myosin binding protein C, cardiac [ Homo sapiens (human) ]

http://www.ncbi.nlm.nih.gov/gene/4607

MYBPC3 provided by HGNC
Official Full Name – myosin binding protein C, cardiac provided by HGNC
Primary source – HGNC:HGNC:7551 ;
See related Ensembl:ENSG00000134571; HPRD:02980; MIM:600958; Vega:OTTHUMG00000166986
Gene type protein coding RefSeq status
REVIEWED Organism Homo sapiens
LineageEukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Primates; Haplorrhini; Catarrhini; Hominidae; Homo
Also known asFHC; CMH4; CMD1MM; LVNC10; MYBP-C

SummaryMYBPC3 encodes the cardiac isoform of myosin-binding protein C. Myosin-binding protein C is a myosin-associated protein found in the cross-bridge-bearing zone (C region) of A bands in striated muscle. MYBPC3, the cardiac isoform, is expressed exclussively in heart muscle. Regulatory phosphorylation of the cardiac isoform in vivo by cAMP-dependent protein kinase (PKA) upon adrenergic stimulation may be linked to modulation of cardiac contraction. Mutations in MYBPC3 are one cause of familial hypertrophic cardiomyopathy. [provided by RefSeq, Jul 2008]

 

 

 

What is the official name of the MYBPC3 gene?

The official name of this gene is “myosin binding protein C, cardiac.”

MYBPC3 is the gene’s official symbol. The MYBPC3 gene is also known by other names, listed below.

Read more about gene names and symbols on the About page.

 

What is the normal function of the MYBPC3 gene?

The MYBPC3 gene provides instructions for making the cardiac myosin binding protein C (cardiac MyBP-C), which is found in heart (cardiac) muscle cells. In these cells, cardiac MyBP-C is associated with a structure called the sarcomere, which is the basic unit of muscle contraction. Sarcomeres are made up of thick and thin filaments. The overlapping thick and thin filaments attach to each other and release, which allows the filaments to move relative to one another so that muscles can contract. Regular contractions of cardiac muscle pump blood to the rest of the body.

In cardiac muscle sarcomeres, cardiac MyBP-C attaches to thick filaments and keeps them from being broken down. Cardiac MyBP-C has chemical groups called phosphate groups attached to it; when the phosphate groups are removed, cardiac MyBP-C is broken down, followed by the breakdown of the proteins of the thick filament. Cardiac MyBP-C also regulates the rate of muscle contraction, although the mechanism is not fully understood.

 

Does the MYBPC3 gene share characteristics with other genes?

The MYBPC3 gene belongs to a family of genes called fibronectin type III domain containing(fibronectin type III domain containing). It also belongs to a family of genes called immunoglobulin superfamily, I-set domain containing (immunoglobulin superfamily, I-set domain containing). It also belongs to a family of genes called MYBP (myosin binding proteins).

A gene family is a group of genes that share important characteristics. Classifying individual genes into families helps researchers describe how genes are related to each other. For more information, see What are gene families? in the Handbook.

http://ghr.nlm.nih.gov/gene/MYBPC3

 

Aliases for MYBPC3 Gene

http://www.genecards.org/cgi-bin/carddisp.pl

  • Myosin Binding Protein C, Cardiac 2 3
  • C-Protein, Cardiac Muscle Isoform 3 4
  • CMD1MM 3 6
  • LVNC10 3 6
  • CMH4 3 6
  • Myosin-Binding Protein C, Cardiac-Type 3
  • Myosin-Binding Protein C, Cardiac 2
  • Cardiac MyBP-C 4
  • MYBP-C 3
  • FHC 3

 

GO – Molecular functioni

http://www.uniprot.org/uniprot/Q14896

GO – Biological processi

Keywords – Molecular functioni

Muscle protein

Keywords – Biological processi

Cell adhesion

Keywords – Ligandi

Actin-binding, Metal-binding, Zinc

Enzyme and pathway databases

 

Organization and Sequence of Human Cardiac Myosin Binding Protein C Gene (MYBPC3) and Identification of Mutations Predicted to Produce Truncated Proteins in Familial Hypertrophic Cardiomyopathy

Lucie CarrierGisele BonneEllen BahrendBing YuPascale RichardFlorence NielBernard Hainque, et al.

Circulation Research.1997; 80: 427-434   http://dx.doi.org:/10.1161/01.res.0000435859.24609.b3

Cardiac myosin binding protein C (MyBP-C) is a sarcomeric protein belonging to the intracellular immunoglobulin superfamily. Its function is uncertain, but for a decade evidence has existed for both structural and regulatory roles. The gene encoding cardiac MyBP-C (MYBPC3) in humans is located on chromosome 11p11.2, and mutations have been identified in this gene in unrelated families with familial hypertrophic cardiomyopathy (FHC). Detailed characterization of the MYBPC3 gene is essential for studies on gene regulation, analysis of the role of MyBP-C in cardiac contraction through the use of recombinant DNA technology, and mutational analyses of FHC. The organization of human MYBPC3 and screening for mutations in a panel of French families with FHC were established using polymerase chain reaction, single-strand conformation polymorphism, and sequencing. The MYBPC3 gene comprises >21 000 base pairs and contains 35 exons. Two exons are unusually small in size, 3 bp each. We found six new mutations associated with FHC in seven unrelated French families. Four of these mutations are predicted to produce truncated cardiac MyBP-C polypeptides. The two others should each produce two aberrant proteins, one truncated and one mutated. The present study provides the first organization and sequence for an MyBP-C gene. The mutations reported here and previously in MYBPC3 result in aberrant transcripts that are predicted to encode significantly truncated cardiac MyBP-C polypeptides. This spectrum of mutations differs from the ones previously observed in other disease genes causing FHC. Our data strengthen the functional importance of MyBP-C in the regulation of cardiac work and provide the basis for further studies.

Cardiac MyBP-C is a member of a family comprising isoforms specific for slow-skeletal, fast-skeletal, and cardiac muscles. The skeletal isoforms were initially described in 1971 [1] and later came to be recognized as proteins with specific myosin- and titin-binding properties located in the A bands of the thick filaments of all vertebrate cross-striated muscle and forming a series of seven to nine transverse stripes, 43 nm apart, in the crossbridgebearing region. [2-5] Subsequent cloning of the three isoforms showed them to belong to the intracellular immunoglobulin superfamily and to share a conserved domain pattern consisting of IgI set domains and fn-3 domains. [6-11]

Comparison of the cardiac and the skeletal MyBP-C isoform sequences reveals three distinct regions that are specific to the cardiac isoform: the N-terminal domain C0 IgI containing 101 residues, the MyBP-C motif (a 105-residue stretch linking the C1 and C2 IgI domains), and a 28-residue loop inserted in the C5 IgI domain. [7,12,13] The MyBP-C motif is not specific to the cardiac isoform, but the alignment of skeletal and cardiac sequences revealed the addition of a nine-residue loop in the cardiac variant, which is the key substrate site for phosphorylation by both protein kinase A and a calmodulin-dependent protein kinase associated with the native protein. [7] As for the 28-residue loop, it is strictly cardiac specific. [14,15] The major myosin-binding site of MyBP-C resides in the C-terminal C10 IgI domain and is mainly restricted to the last 102 amino acids. [16-18] The titin-binding site is also located in the C-terminal region, spanning the C8 to C10 IgI domains of the molecule. [6,13]

The function of MyBP-C is uncertain, but for a decade evidence has existed to indicate both structural and regulatory roles. It should be stressed, however, that most studies were performed on skeletal muscles and that very little functional data exist for cardiac muscle. Several investigators have shown that MyBP-C modulates in vitro the shape and the length of sarcomeric thick filaments [19-21] and that depending on ionic strength and the molar ratio of actin and myosin in solution, the addition of MyBP-C can modulate the actin-activated ATPase activity of skeletal and cardiac myosins. [3,22,23] Partial extraction of the MyBP-C from rat skinned cardiac myocytes and rabbit skeletal muscle fibers alters Ca2+-sensitive tension, supporting the view that contractile function is affected by MyBP-C. [24] This view was very recently strengthened by the elegant studies of Weisberg and Winegrad. [25] These authors showed that phosphorylation of cardiac MyBP-C alters myosin crossbridges in native thick filaments isolated from rat ventricles and suggested that MyBP-C can modify force production in activated cardiac muscles.

The gene encoding the cardiac isoform in humans (MYBPC3) was assigned to the chromosomal location 11p11.2 [7] in a region where we had identified the CMH4 disease locus in FHC. [26] Recently, three mutations in MYBPC3 have been identified in unrelated families with FHC by our group [27] and others. [28] FHC is a genetically and phenotypically heterogeneous disease, transmitted as an autosomal-dominant trait. None of the previous hypotheses of the pathophysiological mechanisms would have predicted that defects in sarcomeric protein genes could be a possible molecular basis for the disease. The results of molecular genetic studies have nevertheless shown that many forms of the disease involve mutations in genes encoding sarcomeric proteins (for reviews, see [29-31]), and the findings that MYBPC3 is one of these disease genes are consistent with the view that cardiac MyBP-C may play a more important role in the regulation of cardiac contraction than was previously thought.

Detailed characterization of the MYBPC3 gene is essential for studies of gene regulation, analysis of the role of cardiac MyBP-C in the sarcomere structure and function through the use of recombinant DNA technology, and, finally, mutational analyses and further studies in FHC. In the present work, we have determined the organization and sequence of the human MYBPC3 gene and shown it to exceed 21 000 bp in size and to contain 35 exons, out of which 34 are coding. We also report that six new mutations in the MYBPC3 gene are associated with FHC in seven unrelated French families. Four of these mutations are predicted to produce truncated cardiac MyBP-C polypeptides in these families. The two others should each produce two aberrant proteins, one truncated and the other mutated or deleted.

 

Screening the Human MYBPC3 Gene for Mutations

The primers were constructed on the basis of flanking intron sequences and were used to amplify each exon (see Table 1). The touchdown PCR was performed (as described above according to the conditions reported in Table 1) on genomic DNA from unrelated FHC patients. For SSCP, PCR products were denatured for 5 minutes at 96 degrees C in a standard denaturing buffer, kept on ice for 5 minutes, loaded onto 6% to 10% polyacrylamide gels, and then run at 6 mA and at 7 degrees C or 20 degrees C in a Hoeffer apparatus. The bands were visualized after silver staining of the gels (Bio-Rad). Sequencing was performed as described above.

Table 1.

Oligonucleotide Primers and PCR Conditions for Detection of Mutations in Human MYBPC3 Gene

RNA Isolation, cDNA Synthesis, and MYBPC3 cDNA Amplifications

Total cellular RNA was isolated from human lymphoblastoid cell lines using RNA Plus (Bioprobe Systems), and the cDNA synthesis was performed as previously described. [27] The cDNA products were amplified in a 50-micro L PCR reaction using two outer primers (see Table 2). A second round of PCR was performed with a final dilution of 1:100 of the first round products, using nested primers (see Table 2). The primers were determined according to the cDNA sequence (EMBL accession number X84075), and cDNA fragments were amplified using a touchdown PCR protocol between 70 degrees C and 60 degrees C. Sizes of normal and mutated cDNA-PCR fragments were assessed, followed by size-fractionation on agarose gels. After extraction and purification of the normal and the putative mutated cDNAs, they were cloned using pGEM-T System II (Promega) and then sequenced as described above.

Table 2.

Oligonucleotide Primers for MYBPC3 cDNA Amplifications

Genomic Organization and Sequence of Human MYBPC3

The size of introns was first estimated by PCR amplification of DNA segments between exons from control genomic DNA, followed by size-fractionation of the PCR products on agarose gels. The exon/intron boundaries and the entire intronic sequences were then determined by sequencing. The sequences have been deposited with EMBL (accession number Y10129). The schematic organization of the human MYBPC3 gene and the alignment of exons with structural domains in the protein are shown in Figure 1. The gene comprises >21 000 bp and contains 35 exons, out of which 34 are coding. A (GT) repeat was found in intron 20 (data not shown). The 101-residue N-terminal extra IgI domain is encoded by exons 1 to 3; the proline-rich domain (51 residues), by exons 3 and 4; the C1 IgI domain (104 residues), by exons 4 to 6; the MyBP-C motif (105 residues), by exons 6 to 12; the C2 IgI domain (91 residues), by exons 12 to 16; the C3 IgI domain (91 residues), by exons 16 to 18; the C4 IgI domain (90 residues), by exons 18 to 20; the linker (11 residues), by exons 20 and 21; the C5 IgI domain (127 residues), by exons 21 to 24; the C6 fn-3 domain (98 residues), by exons 24 to 26; the C7 fn-3 domain (101 residues), by exons 26 to 28; the C8 IgI domain (95 residues), by exons 28 to 30; the C9 fn-3 domain (115 residues), by exons 30 to 32; and the C-terminal C10 IgI domain (94 residues), by exons 32 to 34.

Figure 1.

Schematic organization of the human MYBPC3 gene and alignment of exons with structural domains of the protein. Top, The structural domains of cardiac MyBP-C. The high-affinity myosin heavy chain domain (confined to the C10 IgI repeat), the titin binding site (C8 to C10), and the phosphorylation sites are indicated. Middle, The mRNA with the limits of exons. Bottom, the schematic organization of the gene with locations of exons shown by boxes and introns shown by horizontal lines. The exons are numbered from the 5 prime end of the gene, with exon 1 containing the first codon ATG. The exons coding for structural domains are indicated by interrupted lines.

The sizes of exons and introns are summarized in Table 3. The exon sizes, excluding the 5 prime and 3 prime untranslated regions, vary between 3 and 267 bp. Two of the exons, ie, exons 10 and 14, are unusually small and contain three nucleotides each. The remaining 32 exons vary in size between 18 and 267 bp. Twenty-seven exons finish with a split codon (see Table 3). The intron sizes vary between 85 and [nearly =]2000 bp. The major consensus donor splice site is GTGAG in 53% of the cases, and the major consensus acceptor splice site is CAG in 91% of the cases. Twenty-seven of the 34 introns contain putative branch point sequences located -14 to -51 upstream from each splice acceptor site. Introns 1, 4, 11, 14, 16, 24, and 31 do not contain any known consensus branch point sequence.

Table 3.

Exon-Intron Boundaries in the Human MYBPC3 Gene Identification of Mutations in MYBPC3 Gene Associated With FHC

Because the families were not large enough to assess linkage on the basis of a statistically significant Lod score, we used haplotype analysis to define the disease locus responsible for FHC in each family. Linkage was established on the basis of the transmission of a common haplotype in affected individuals and exclusion on the basis of affected recombinant individuals. Families 717 and 740 presented linkage only to CMH4, and the other five families (families 702, 716, 731, 750, and 754) were less informative but at least potentially linked to CMH4 (data not shown).

All the exon-intron boundaries were analyzed by PCRSSCP according to the conditions described in Table 2. A total of six new mutations were identified in MYBPC3 associated with FHC in seven unrelated French families (Figure 2 andFigure 3, Table 4).

Table 4.

Consequences at mRNA Level of MYBPC3 Mutations

Figure 2.

Pedigrees of families with MYBPC3 gene mutations. Clinical affection status is indicated: darkened, affected; clear, unaffected; and clear with a cross, indeterminate. Genetically affected status is indicated by an asterisk. The mutations (M) are as follows: M1, GTGAG[arrow right]GTGAA splice donor site mutation in intron 7; M2, GAA[arrow right]CAA mutation in exon 17; M3, GT[arrow right]AT splice donor site mutation in intron 23; M4, TGAT[arrow right]TGGT transversion in the branch point consensus sequence of intron 23; M5, [-GCGTC] deletion in exon 25; and M6, duplication [+TTCAAGAATGGC]/deletion [-ACCT] in exon 33.

Figure 3.

Normal and mutated cardiac MyBP-C polypeptides. N indicates the normal structure of human cardiac MyBP-C; M1 to M6 correspond to the predicted products of the aberrant MyBP-C cDNAs resulting from the different mutations.

M1 is a GTGAG[arrow right]GTGAA transition in the 3 prime splice donor site of intron 7 in family 717. The G residue at position +5 in the intron is a highly conserved nucleotide in the splice donor consensus sequence. [33] The G[arrow right]A mutation inactivates this donor site. Amplification of MYBPC3 cDNA from patients’ lymphocytes identified the skipping of the 49-bp exon 7 that produces a frameshift. No alternative splice donor site was found in intron 7. The aberrant cDNA encodes 258 normal cardiac MyBP-C residues, followed by 25 new amino acids, and a premature termination of translation. This should produce a large truncated protein (-80%) lacking the MyBP-C motif containing the phosphorylation sites and the titin and myosin binding sites.

M2 is a G[arrow right]C transversion at position 1656 in exon 17 in families 702 and 750 that produces a mutated polypeptide in the C3 domain at the position 542 (Glu[arrow right]Gln). Otherwise, this mutation affects the last nucleotide of the exon, which is part of the consensus splicing site. [34] A common feature in human exon-intron boundaries is that 80% of exons finish with a guanine (85% in MYBPC3). This mutation also results in an aberrant transcript in lymphocytes (with the skipping of exon 17) that directly introduces a stop codon. The aberrant cDNA encodes 486 normal cardiac MyBP-C residues, leading to a truncated protein (-62%) that lacks the titin and myosin binding sites.

M3 is a GT[arrow right]AT transition in the 3 prime splice donor site of intron 23 in family 716 that inactivates this splicing site. This mutation produces the skipping of the 160-bp exon 23. No alternative splice donor site was found in lymphocytes. The mutated cDNA identified in lymphocytes encodes 717 normal residues and then 51 novel amino acids, followed by premature termination of the translation in the C5 domain, leading to a potential truncated protein (-44%) that loses the titin and myosin binding domains.

M4 is a TGAT[arrow right]TGGT transition in intron 23 in family 740. This A[arrow right]G mutation inactivates a potential branch point consensus sequence (URAY). Although three potential branch points exist upstream from the mutation, they do not seem to be used, since analysis of the transcripts in lymphocytes indicates the existence of two aberrant cDNAs. One corresponds to the skipping of the 105-bp exon 24 without frameshift and encodes a polypeptide depleted of 35 amino acids in the C6 domain (-50% of C6). The other still contains the 724-bp intron 23. This mutant cDNA is associated with a frameshift: it encodes 770 normal cardiac MyBP-C residues and then 100 novel amino acids, followed by a stop codon, and the corresponding truncated protein (-40%) should not interact with either titin or myosin.

M5 is a 5-bp deletion (-GCGTC) in exon 25 in family 731. This deletion also produces a frameshift: the aberrant cDNA identified in the lymphocytes encodes 845 normal MyBP-C residues and then 35 novel amino acids, followed by a premature stop codon in the C6 domain that should produce a truncated protein (-34%), losing the C-terminal region containing both the titin- and myosin-binding sites.

M6 is a 12-bp duplication (+TTCAAGAATGGC)/4-bp deletion (-ACCT) in exon 33 in family 754. This modification introduces a frameshift at position 3691 that leads to 1220 normal MyBP-C residues and then 19 novel amino acids, followed by a premature stop codon in the last third part of the C10 domain. The predicted truncated protein (-4%) should also lose part of its myosin binding site.

All these six mutations were absent in 200 samples from control unrelated subjects without FHC and also in 42 unrelated probands with FHC (out of which 8 have mutations in MYBPC3, 8 have mutations in the beta-myosin heavy chain gene [MYH7], 1 has a mutation in the cardiac troponin T gene, and 25 have presently undefined mutations).

Discussion

The present work describes the first genomic organization for an MyBP-C. The gene is over 21 000 bp and contains 35 exons. An interesting feature of the organization of this gene is that there is a striking correspondence between the limits of the exons and those of structural domains (Figure 1). The IgI and fn-3 domains are encoded by two or three exons. The linker region between the IgI C4 and IgI C5 domains corresponds to exon 20. Twenty-six of the 28 cardiac-specific amino acids of the IgI C5 domain correspond to exon 22. Finally, the MyBP-C motif is encoded by the most complex exon structure: the nine cardiac-specific amino acids correspond to exon 8, and the four phosphorylation sites described by Gautel et al [7] are encoded by six exons and are located at the end or at the junction of two exons (phosphorylation sites: A, junction of exons 7 and 8; B, end of exon 8; C, end of exon 9, exon 10, and beginning of exon 11; and D, end of exon 12). The correlation between exonic organization and protein structure has also recently been described concerning the titin, [35] suggesting a common feature for the intracellular immunoglobulin superfamily.

We suggest that the new mutations described here cause FHC because they segregate with the disease, are not present in controls, and result in aberrant transcripts that are predicted to encode significantly altered cardiac MyBP-C polypeptide structure and/or function. They are all transcribed into mRNAs in lymphocytes. However, because most, if not all, genes in humans are thought to be transcribed at very low levels in lymphocytes (“illegitimate transcription”), [36] these results do not address the hypothesis that these mutations are expressed in the diseased myocardium. Since cardiac MyBP-C is specifically expressed in heart, ventricular tissue is needed to address this issue, and we had no access to any myocardial specimens. One study documented the expression of a missense mutation in the mRNA for the beta-myosin heavy chain in myocardial tissue from an affected patient with FHC. [37] Because the beta-myosin heavy chain is normally expressed in slow-twitch skeletal fibers, skeletal muscle biopsies can also be used to show that the mutated myosin is produced in the muscle and that the mutation alters the function of the beta-myosin and the contractile properties of the muscle fibers. [38,39] One might thus reasonably assume that the MYBPC3 gene mutations are expressed in the myocardium and that they exert their effect by altering the multimeric complex assembly of the cardiac sarcomere via at least one of these mechanisms: (1) They can act as “poison polypeptides” through a dominant-negative effect. The altered proteins would be incorporated in the sarcomere and would alter the assembly of the sarcomeric filaments, since most truncated MyBP-Cs are unable to cross-link the titin and/or myosin molecules. (2) They can act as “null alleles,” potentially leading to haplo insufficiency; the production of insufficient quantities of normal cardiac MyBP-C would produce an imbalance in stoichiometry of the thick-filament components that would be sufficient to alter the sarcomeric structure and function. (3) Since myosin, titin, and MyBP-C might be translated and assembled cotranslationally, one can also assume that the misfolded, mutated MYBPC3 mRNAs may disturb the translation of the other sarcomeric components that would interfere with the proper assembly of sarcomeric structures.

The full spectrum of mutations of the FHC disease genes is far from known, but it is intriguing to note that most mutations found so far in MYH7 are missense ones, whereas most of those in MYBPC3 disrupt the reading frame and produce premature stop codons. Both genes are large ones, composed of [nearly =]40 exons, and there are no reasons for different types of mutations in the two genes. Thus, one might hypothesize that mutations leading to truncated proteins exist also for MYH7 in humans but have no deleterious effect. In support of this are the reports of two deletions in the C-terminal part of the beta-myosin heavy chain molecule with almost no phenotype. One is a 2.4-kbp deletion including part of intron 39 and exon 40 containing the 3 prime untranslated region and the polyadenylation signal, which was reported in a small pedigree. [40] Only the proband had developed clinically diagnosed hypertrophic cardiomyopathy at a very late onset (age, 59 years), and the other genotypically affected family members had not developed the disease at 10, 32, and 33 years. The other one is a large deletion leaving only a short variant of the beta-myosin heavy chain constituting only the first 53 residues of the molecule (out of 1935). This deletion was found by chance in an unaffected individual. [41] For MYBPC3, in contrast, the majority of the mutations described so far produce the C-terminal truncation of the cardiac MyBP-C polypeptides and are associated with an FHC phenotype. However, no definitive conclusion can be drawn at this stage concerning the pathogenic mechanisms of mutations in these two genes. The present work provides the molecular basis for the production of transgenic animals for cardiac MyBP-C that will help to resolve some of these issues.

Footnotes
  • Received December 2, 1996; accepted January 10, 1997.

  • This manuscript was sent to Laurence Kedes, Consulting Editor, for review by expert referees, editorial decision, and final disposition.

  • Selected Abbreviations and Acronyms
    EMBL
    European Molecular Biology Laboratory
    FHC
    familial hypertrophic cardiomyopathy
    fn-3
    fibronectin III
    MyBP-C
    myosin binding protein C
    PCR
    polymerase chain reaction
    SSCP
    single-strand conformation polymorphism analysis
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MYBPC3 – Hypertrophic Cardiomyopathy Testing

http://www.cincinnatichildrens.org/workarea/downloadasset.aspx?id=90111
Hypertrophic Cardiomyopathy (HCM) is relatively common, with a prevalence of 1 in 500 adults (1). HCM is a primary disorder of heart muscle characterized by left ventricular hypertrophy. The most classic finding in HCM is asymmetric septal hypertrophy, with or without left ventricular outflow tract obstruction. The disease demonstrates extensive clinical variability with regard to age of onset, severity and progression of disease. HCM can affect infants and children although it is more typically identified in adolescence or adulthood (2,3).

The MYBPC3 gene codes for cardiac myosin binding protein C. Phosphorylation of this protein modulates contraction and is an important component of the sarcomere (4). The MYBPC3 gene contains 35 exons and is located at chromosome 11p11.2. Up to 40% of individuals with a clinical diagnosis of HCM have MYBPC3 mutations (2). MYBPC3 mutations are inherited in an autosomal dominant manner. The majority of individuals inherit the MYBPC3 from a parent, although de novo mutations do occur. Mutations in MYBPC3 and MYH7 genes are the most common causes of HCM. However, the disease is genetically heterogeneous and sequencing additional genes should be considered if familial HCM is suspected or the underlying etiology remains unknown. Approximately 50-65% of individuals with a known or suspected diagnosis of familial HCM have a mutation in one of a number of genes encoding components of the sarcomere and cytoskeleton (3). Compound heterozygous mutations have been reported in MYBPC3 and other genes associated with HCM (5). Mutations in the MYBPC3 gene have been primarily associated with HCM, but can also be associated with other types of heart muscle disease including dilated cardiomyopathy, restrictive cardiomyopathy and left-ventricular non-compaction (6).
Indication MYBPC3 testing is utilized to confirm a diagnosis of HCM in patients with clinically evident disease. Genetic testing also allows for early identification and diagnosis of individuals at greatest risk prior to the expression of typical clinical manifestations. If a mutation is identified in an asymptomatic individual, regular and routine outpatient follow up is indicated. If clinically unaffected members of a family with an identified mutation for HCM are found not to carry that mutation, they can be definitely diagnosed as unaffected and reassured that neither they nor their children will be at higher risk compared to the general population to develop symptoms related to HCM. A negative test result in an individual with a known familial mutation also eliminates the need for routine follow up.
Methodology:
All 35 exons of the MYBPC3 gene, as well as the exon/intron boundaries and a portion of untranslated regions of the gene are amplified by PCR. Genomic DNA sequences from both forward and reverse directions are obtained by automatic fluorescent detection using an ABI PRISM® 3730 DNA Analyzer. Sequence variants different from National Center for Biotechnology Information GenBank references are further evaluated for genetic significance. If a mutation is identified, a known familial mutation analysis will be available for additional family members.
Sensitivity & Accuracy:
Greater than 98.5% of the mutations in exon 1-35 of MYBPC3 are detectable by sequence based methods. Sequencing does not detect deletions or duplications. Mutations in MYBPC3 account for up to 40% of cases of idiopathic hypertrophic cardiomyopathy.
References:
1. Maron BJ, Gardin JM, Flack JM, Gidding SS, Kurosaki TT, Bild DE. Prevalence of hypertrophic cardiomyopathy in a general population of young adults. Echocardiographic analysis of 4111 subjects in the cardia study. Coronary artery risk development in (young) adults. Circulation. 1995;92:785-789.
2. Kaski JP, Syrris P, Esteban MT, Jenkins S, Pantazis A, Deanfield JE, McKenna WJ, Elliott PM. Prevalence of sarcomere protein gene mutations in preadolescent children with hypertrophic cardiomyopathy. Circulation Cardiovascular Genetics. 2009;2:436441.
3. Morita H, Rehm HL, Menesses A, McDonough B, Roberts AE, Kucherlapati R, Towbin JA, Seidman JG, Seidman CE. Shared genetic causes of cardiac hypertrophy in children and adults. The New England Journal of Medicine. 2008;358:1899-1908.
4. van Dijk SJ, Dooijes D, dos Remedios C, Michels M, Lamers JM, Winegrad S, Schlossarek S, Carrier L, ten Cate FJ, Stienen GJ, van der Velden J. Cardiac myosin-binding protein c mutations and hypertrophic cardiomyopathy: Haploinsufficiency, deranged phosphorylation, and cardiomyocyte dysfunction. Circulation. 2009;119:1473-1483.
5. Van Driest SL, Vasile VC, Ommen SR, Will ML, Tajik AJ, Gersh BJ, Ackerman MJ. Myosin binding protein c mutations and compound heterozygosity in hypertrophic cardiomyopathy. Journal of the American College of Cardiology. 2004;44:1903-1910.
6. Hershberger RE, Norton N, Morales A, Li DX, Siegfried JD, Gonzalez-Quintana J. Coding sequence rare variants identified in MYBPC3, MYH6, TPM1, TNNC1, and TNNI3 from 312 patients with familial or idiopathic dilated cardiomyopathy. CirculationCardiovascular Genetics. 2010;3:155-161.

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Cardiac Resynchronization Therapy (CRT) Improves Symptoms and Reduces Mortality and Readmission among Selected Patients with Heart Failure and Left Ventricular Systolic Dysfunction

Reporter: Aviva Lev-Ari, PhD, RN

QRS Duration, Bundle-Branch Block Morphology, and Outcomes Among Older Patients With Heart Failure Receiving Cardiac Resynchronization Therapy

Pamela N. Peterson, MD, MSPH1,2,3; Melissa A. Greiner, MS4; Laura G. Qualls, MS4; Sana M. Al-Khatib, MD, MHS4,5; Jeptha P. Curtis, MD6; Gregg C. Fonarow, MD7; Stephen C. Hammill, MD8; Paul A. Heidenreich, MD9; Bradley G. Hammill, MS4; Jonathan P. Piccini, MD, MHS4,5; Adrian F. Hernandez, MD, MHS4,5; Lesley H. Curtis, PhD4,5; Frederick A. Masoudi, MD, MSPH2,3

Importance  The benefits of cardiac resynchronization therapy (CRT) in clinical trials were greater among patients with left bundle-branch block (LBBB) or longer QRS duration.

Objective  To measure associations between QRS duration and morphology and outcomes among patients receiving a CRT defibrillator (CRT-D) in clinical practice.

Design, Setting, and Participants  Retrospective cohort study of Medicare beneficiaries in the National Cardiovascular Data Registry’s ICD Registry between 2006 and 2009 who underwent CRT-D implantation. Patients were stratified according to whether they were admitted for CRT-D implantation or for another reason, then categorized as having either LBBB or no LBBB and QRS duration of either 150 ms or greater or 120 to 149 ms.

Main Outcomes and Measures  All-cause mortality; all-cause, cardiovascular, and heart failure readmission; and complications. Patients underwent follow-up for up to 3 years, with follow-up through December 2011.

Results  Among 24 169 patients admitted for CRT-D implantation, 1-year and 3-year mortality rates were 9.2% and 25.9%, respectively. All-cause readmission rates were 10.2% at 30 days and 43.3% at 1 year. Both the unadjusted rate and adjusted risk of 3-year mortality were lowest among patients with LBBB and QRS duration of 150 ms or greater (20.9%), compared with LBBB and QRS duration of 120 to 149 ms (26.5%; adjusted hazard ratio [HR], 1.30 [99% CI, 1.18-1.42]), no LBBB and QRS duration of 150 ms or greater (30.7%; HR, 1.34 [99% CI, 1.20-1.49]), and no LBBB and QRS duration of 120 to 149 ms (32.3%; HR, 1.52 [99% CI, 1.38-1.67]). The unadjusted rate and adjusted risk of 1-year all-cause readmission were also lowest among patients with LBBB and QRS duration of 150 ms or greater (38.6%), compared with LBBB and QRS duration of 120 to 149 ms (44.8%; adjusted HR, 1.18 [99% CI, 1.10-1.26]), no LBBB and QRS duration of 150 ms or greater (45.7%; HR, 1.16 [99% CI, 1.08-1.26]), and no LBBB and QRS duration of 120 to 149 ms (49.6%; HR, 1.31 [99% CI, 1.23-1.40]). There were no observed associations with complications.

Conclusions and Relevance  Among fee-for-service Medicare beneficiaries undergoing CRT-D implantation in clinical practice, LBBB and QRS duration of 150 ms or greater, compared with LBBB and QRS duration less than 150 ms or no LBBB regardless of QRS duration, was associated with lower risk of all-cause mortality and of all-cause, cardiovascular, and heart failure readmissions.

Clinical trials have shown that cardiac resynchronization therapy (CRT) improves symptoms and reduces mortality and readmission among selected patients with heart failure and left ventricular systolic dysfunction. Following broad implementation of CRT, it was recognized that one-third to one-half of patients receiving the therapy for heart failure do not improve.1 Identification of patients likely to benefit from CRT is particularly important, because CRT defibrillator (CRT-D) implantation is expensive, invasive, and associated with important procedural risks.

A primary question regarding optimal patient selection for CRT is whether patients with longer QRS duration or left bundle-branch block (LBBB) morphology derive greater benefit than others. Current guidelines recommend selection of patients primarily on the basis of QRS duration and morphology based predominantly on meta-analyses and subgroup analyses of clinical trials evaluating either QRS duration or morphology. Only 1 study specifically evaluated the combination of QRS duration and morphology but did not assess meaningful patient outcomes.2 Thus, the role of QRS duration and morphology in the selection of patients for CRT in contemporary clinical practice remains unclear.

The objectives of this study were to determine the long-term outcomes of unselected patients undergoing CRT-D implantation in real-world settings and associations between combinations of QRS duration and presence of LBBB and longitudinal outcomes, including mortality, readmission, and complications following CRT-D implantation in a large population of Medicare beneficiaries who received CRT-Ds.

SOURCE

http://jama.jamanetwork.com/article.aspx?articleid=1728715&utm_content=sidebar-related&utm_term=alsolike&utm_source=Silverchair%20Information%20Systems&utm_medium=email&utm_campaign=JAMA:OnlineFirst08/30/2015

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Metabolic Genomics and Pharmaceutics, Vol. 1 of BioMed Series D available on Amazon Kindle


Metabolic Genomics and Pharmaceutics, Vol. 1 of BioMed Series D available on Amazon Kindle

Reporter: Stephen S Williams, PhD

 

Leaders in Pharmaceutical Business Intelligence would like to announce the First volume of their BioMedical E-Book Series D:

Metabolic Genomics & Pharmaceutics, Vol. I

SACHS FLYER 2014 Metabolomics SeriesDindividualred-page2

which is now available on Amazon Kindle at

http://www.amazon.com/dp/B012BB0ZF0.

This e-Book is a comprehensive review of recent Original Research on  METABOLOMICS and related opportunities for Targeted Therapy written by Experts, Authors, Writers. This is the first volume of the Series D: e-Books on BioMedicine – Metabolomics, Immunology, Infectious Diseases.  It is written for comprehension at the third year medical student level, or as a reference for licensing board exams, but it is also written for the education of a first time baccalaureate degree reader in the biological sciences.  Hopefully, it can be read with great interest by the undergraduate student who is undecided in the choice of a career. The results of Original Research are gaining value added for the e-Reader by the Methodology of Curation. The e-Book’s articles have been published on the Open Access Online Scientific Journal, since April 2012.  All new articles on this subject, will continue to be incorporated, as published with periodical updates.

We invite e-Readers to write an Article Reviews on Amazon for this e-Book on Amazon.

All forthcoming BioMed e-Book Titles can be viewed at:

https://pharmaceuticalintelligence.com/biomed-e-books/

Leaders in Pharmaceutical Business Intelligence, launched in April 2012 an Open Access Online Scientific Journal is a scientific, medical and business multi expert authoring environment in several domains of  life sciences, pharmaceutical, healthcare & medicine industries. The venture operates as an online scientific intellectual exchange at their website http://pharmaceuticalintelligence.com and for curation and reporting on frontiers in biomedical, biological sciences, healthcare economics, pharmacology, pharmaceuticals & medicine. In addition the venture publishes a Medical E-book Series available on Amazon’s Kindle platform.

Analyzing and sharing the vast and rapidly expanding volume of scientific knowledge has never been so crucial to innovation in the medical field. WE are addressing need of overcoming this scientific information overload by:

  • delivering curation and summary interpretations of latest findings and innovations on an open-access, Web 2.0 platform with future goals of providing primarily concept-driven search in the near future
  • providing a social platform for scientists and clinicians to enter into discussion using social media
  • compiling recent discoveries and issues in yearly-updated Medical E-book Series on Amazon’s mobile Kindle platform

This curation offers better organization and visibility to the critical information useful for the next innovations in academic, clinical, and industrial research by providing these hybrid networks.

Table of Contents for Metabolic Genomics & Pharmaceutics, Vol. I

Chapter 1: Metabolic Pathways

Chapter 2: Lipid Metabolism

Chapter 3: Cell Signaling

Chapter 4: Protein Synthesis and Degradation

Chapter 5: Sub-cellular Structure

Chapter 6: Proteomics

Chapter 7: Metabolomics

Chapter 8:  Impairments in Pathological States: Endocrine Disorders; Stress

                   Hypermetabolism and Cancer

Chapter 9: Genomic Expression in Health and Disease 

 

Summary 

Epilogue

 

 

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