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Zajičková T, Horváthová E, Kyzek S, Šályová E, Túryová E, Ševčovičová A, Gálová E. Comparison of Cytotoxic, Genotoxic, and DNA-Protective Effects of Skyrin on Cancerous vs. Non-Cancerous Human Cells. Int J Mol Sci 2022; 23:5339. [PMID: 35628149 PMCID: PMC9142076 DOI: 10.3390/ijms23105339] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2022] [Revised: 05/06/2022] [Accepted: 05/09/2022] [Indexed: 12/04/2022] Open
Abstract
Secondary metabolites as a potential source of anticancer therapeutics have been the subject of many studies. Since hypericin, a metabolite isolated from Hypericum perforatum L., shows several biomedical properties applicable in oncology, the aim of our study was to investigate its potential precursor skyrin in terms of genotoxic and DNA-protective effects. These skyrin effects were analyzed by cell-free methods, and cytotoxicity was estimated by an MTT assay and by a trypan blue exclusion test, while the genotoxic/antigenotoxic potential was examined by comet assay using non-cancerous human lymphocytes and the HepG2 cancer cell line. Skyrin did not show DNA-damaging effects but rather exhibited DNA-protectivity using a DNA-topology assay. However, we observed only weak antioxidant and chelating skyrin properties in other cell-free methods. Regarding the cytotoxic activity of skyrin, HepG2 cells were more prone to skyrin-induced death in comparison to human lymphocytes. Skyrin in non-cytotoxic concentrations did not exhibit elevated genotoxicity in both cell types. On the other hand, skyrin displayed moderate DNA-protective effects that were more noticeable in the case of non-cancerous human lymphocytes. The potential genotoxic effects of skyrin were not observed, and its DNA-protective capacity was more prominent in non-cancerous cells. Therefore, skyrin might be a promising agent used in anticancer therapy.
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Affiliation(s)
- Terézia Zajičková
- Department of Genetics, Faculty of Natural Sciences, Comenius University in Bratislava, Ilkovičova 6, 842 15 Bratislava, Slovakia; (T.Z.); (E.Š.); (E.T.); (A.Š.); (E.G.)
| | - Eva Horváthová
- Cancer Research Institute, Biomedical Research Centre of the Slovak Academy of Sciences, Dúbravská Cesta 9, 845 05 Bratislava, Slovakia;
| | - Stanislav Kyzek
- Department of Genetics, Faculty of Natural Sciences, Comenius University in Bratislava, Ilkovičova 6, 842 15 Bratislava, Slovakia; (T.Z.); (E.Š.); (E.T.); (A.Š.); (E.G.)
| | - Eva Šályová
- Department of Genetics, Faculty of Natural Sciences, Comenius University in Bratislava, Ilkovičova 6, 842 15 Bratislava, Slovakia; (T.Z.); (E.Š.); (E.T.); (A.Š.); (E.G.)
| | - Eva Túryová
- Department of Genetics, Faculty of Natural Sciences, Comenius University in Bratislava, Ilkovičova 6, 842 15 Bratislava, Slovakia; (T.Z.); (E.Š.); (E.T.); (A.Š.); (E.G.)
| | - Andrea Ševčovičová
- Department of Genetics, Faculty of Natural Sciences, Comenius University in Bratislava, Ilkovičova 6, 842 15 Bratislava, Slovakia; (T.Z.); (E.Š.); (E.T.); (A.Š.); (E.G.)
| | - Eliška Gálová
- Department of Genetics, Faculty of Natural Sciences, Comenius University in Bratislava, Ilkovičova 6, 842 15 Bratislava, Slovakia; (T.Z.); (E.Š.); (E.T.); (A.Š.); (E.G.)
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Sun EW, Martin AM, de Fontgalland D, Sposato L, Rabbitt P, Hollington P, Wattchow DA, Colella AD, Chataway T, Wewer Albrechtsen NJ, Spencer NJ, Young RL, Keating DJ. Evidence for Glucagon Secretion and Function Within the Human Gut. Endocrinology 2021; 162:6127286. [PMID: 33534908 DOI: 10.1210/endocr/bqab022] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/23/2020] [Indexed: 11/19/2022]
Abstract
Glucagon is secreted by pancreatic α cells in response to hypoglycemia and increases hepatic glucose output through hepatic glucagon receptors (GCGRs). There is evidence supporting the notion of extrapancreatic glucagon but its source and physiological functions remain elusive. Intestinal tissue samples were obtained from patients undergoing surgical resection of cancer. Mass spectrometry analysis was used to detect glucagon from mucosal lysate. Static incubations of mucosal tissue were performed to assess glucagon secretory response. Glucagon concentration was quantitated using a highly specific sandwich enzyme-linked immunosorbent assay. A cholesterol uptake assay and an isolated murine colonic motility assay were used to assess the physiological functions of intestinal GCGRs. Fully processed glucagon was detected by mass spectrometry in human intestinal mucosal lysate. High glucose evoked significant glucagon secretion from human ileal tissue independent of sodium glucose cotransporter and KATP channels, contrasting glucose-induced glucagon-like peptide 1 (GLP-1) secretion. The GLP-1 receptor agonist Exendin-4 attenuated glucose-induced glucagon secretion from the human ileum. GCGR blockade significantly increased cholesterol uptake in human ileal crypt culture and markedly slowed ex vivo colonic motility. Our findings describe the human gut as a potential source of extrapancreatic glucagon and demonstrate a novel enteric glucagon/GCGR circuit with important physiological functions beyond glycemic regulation.
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Affiliation(s)
- Emily W Sun
- Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Bedford Park, SA, Australia
| | - Alyce M Martin
- Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Bedford Park, SA, Australia
| | | | - Luigi Sposato
- Department of Surgery, Flinders Medical Centre, Bedford Park, SA, Australia
| | - Philippa Rabbitt
- Department of Surgery, Flinders Medical Centre, Bedford Park, SA, Australia
| | - Paul Hollington
- Department of Surgery, Flinders Medical Centre, Bedford Park, SA, Australia
| | - David A Wattchow
- Department of Surgery, Flinders Medical Centre, Bedford Park, SA, Australia
| | - Alexander D Colella
- Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Bedford Park, SA, Australia
| | - Tim Chataway
- Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Bedford Park, SA, Australia
| | | | - Nick J Spencer
- Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Bedford Park, SA, Australia
| | - Richard L Young
- Adelaide Medical School and NHMRC Centre of Research Excellence in Translating Nutritional Science to Good Health, University of Adelaide, Adelaide, SA, Australia
- Nutrition, Diabetes and Metabolism, Lifelong Health, South Australia Health and Medical Research Institute, Adelaide, SA, Australia
| | - Damien J Keating
- Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Bedford Park, SA, Australia
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Babinčák M, Jendželovský R, Košuth J, Majerník M, Vargová J, Mikulášek K, Zdráhal Z, Fedoročko P. Death Receptor 5 (TNFRSF10B) Is Upregulated and TRAIL Resistance Is Reversed in Hypoxia and Normoxia in Colorectal Cancer Cell Lines after Treatment with Skyrin, the Active Metabolite of Hypericum spp. Cancers (Basel) 2021; 13:1646. [PMID: 33916015 PMCID: PMC8036732 DOI: 10.3390/cancers13071646] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2020] [Revised: 03/26/2021] [Accepted: 03/29/2021] [Indexed: 02/08/2023] Open
Abstract
Skyrin (SKR) is a plant bisanthraquinone secondary metabolite from the Hypericum genus with potential use in anticancer therapy. However, its effect and mechanism of action are still unknown. The negative effect of SKR on HCT 116 and HT-29 cancer cell lines in hypoxic and normoxic conditions was observed. HCT 116 cells were more responsive to SKR treatment as demonstrated by decreased metabolic activity, cellularity and accumulation of cells in the G1 phase. Moreover, an increasing number of apoptotic cells was observed after treatment with SKR. Based on the LC-MS comparative proteomic data from hypoxia and normoxia (data are available via ProteomeXchange with the identifier PXD019995), SKR significantly upregulated Death receptor 5 (DR5), which was confirmed by real-time qualitative PCR (RT-qPCR). Furthermore, multiple changes in the Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-activated cascade were observed. Moreover, the reversion of TRAIL resistance was observed in HCT 116, HT-29 and SW620 cell lines, even in hypoxia, which was linked to the upregulation of DR5. In conclusion, our results propose the use of SKR as a prospective anticancer drug, particularly as an adjuvant to TRAIL-targeting treatment to reverse TRAIL resistance in hypoxia.
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Affiliation(s)
- Marián Babinčák
- Institute of Biology and Ecology, Faculty of Science, Pavol Jozef Šafárik University in Košice, Šrobárova 2, 041 54 Košice, Slovakia; (M.B.); (R.J.); (J.K.); (M.M.); (J.V.)
| | - Rastislav Jendželovský
- Institute of Biology and Ecology, Faculty of Science, Pavol Jozef Šafárik University in Košice, Šrobárova 2, 041 54 Košice, Slovakia; (M.B.); (R.J.); (J.K.); (M.M.); (J.V.)
| | - Ján Košuth
- Institute of Biology and Ecology, Faculty of Science, Pavol Jozef Šafárik University in Košice, Šrobárova 2, 041 54 Košice, Slovakia; (M.B.); (R.J.); (J.K.); (M.M.); (J.V.)
| | - Martin Majerník
- Institute of Biology and Ecology, Faculty of Science, Pavol Jozef Šafárik University in Košice, Šrobárova 2, 041 54 Košice, Slovakia; (M.B.); (R.J.); (J.K.); (M.M.); (J.V.)
| | - Jana Vargová
- Institute of Biology and Ecology, Faculty of Science, Pavol Jozef Šafárik University in Košice, Šrobárova 2, 041 54 Košice, Slovakia; (M.B.); (R.J.); (J.K.); (M.M.); (J.V.)
| | - Kamil Mikulášek
- Central European Institute of Technology, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic; (K.M.); (Z.Z.)
- National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic
| | - Zbyněk Zdráhal
- Central European Institute of Technology, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic; (K.M.); (Z.Z.)
- National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic
| | - Peter Fedoročko
- Institute of Biology and Ecology, Faculty of Science, Pavol Jozef Šafárik University in Košice, Šrobárova 2, 041 54 Košice, Slovakia; (M.B.); (R.J.); (J.K.); (M.M.); (J.V.)
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Bioactive Secondary Metabolites from Psychrophilic Fungi and Their Industrial Importance. Fungal Biol 2021. [DOI: 10.1007/978-3-030-85603-8_10] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/19/2022]
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Thappeta KRV, Zhao LN, Nge CE, Crasta S, Leong CY, Ng V, Kanagasundaram Y, Fan H, Ng SB. In-Silico Identified New Natural Sortase A Inhibitors Disrupt S. aureus Biofilm Formation. Int J Mol Sci 2020; 21:ijms21228601. [PMID: 33202690 PMCID: PMC7696255 DOI: 10.3390/ijms21228601] [Citation(s) in RCA: 27] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2020] [Revised: 11/09/2020] [Accepted: 11/12/2020] [Indexed: 12/12/2022] Open
Abstract
Sortase A (SrtA) is a membrane-associated enzyme that anchors surface-exposed proteins to the cell wall envelope of Gram-positive bacteria such as Staphylococcus aureus. As SrtA is essential for Gram-positive bacterial pathogenesis but dispensable for microbial growth or viability, SrtA is considered a favorable target for the enhancement of novel anti-infective drugs that aim to interfere with key bacterial virulence mechanisms, such as biofilm formation, without developing drug resistance. Here, we used virtual screening to search an in-house natural compound library and identified two natural compounds, N1287 (Skyrin) and N2576 ((4,5-dichloro-1H-pyrrol-2-yl)-[2,4-dihydroxy-3-(4-methyl-pentyl)-phenyl]-methanone) that inhibited the enzymatic activity of SrtA. These compounds also significantly reduced the growth of S. aureus but possessed moderate mammalian toxicity. Furthermore, S. aureus strains treated with these compounds exhibited reduction in adherence to host fibrinogen, as well as biofilm formation. Hence, these compounds may represent an anti-infective therapy without the side effects of antibiotics.
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Affiliation(s)
- Kishore Reddy Venkata Thappeta
- Singapore Institute of Food and Biotechnology Innovation (SIFBI), Agency for Science, Technology and Research (A*STAR), 31 Biopolis Way, #01-02 Nanos, Singapore 138669, Singapore; (K.R.V.T.); (C.E.N.); (S.C.); (C.Y.L.); (V.N.)
| | - Li Na Zhao
- Bioinformatics Institute (BII), Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, #07-01 Matrix, Singapore 138671, Singapore;
- Institute of Molecular and Cell Biology (IMCB), Agency for Science, Technology and Research (A*STAR), 61 Biopolis Drive, #3-09 Proteos, Singapore 138673, Singapore
| | - Choy Eng Nge
- Singapore Institute of Food and Biotechnology Innovation (SIFBI), Agency for Science, Technology and Research (A*STAR), 31 Biopolis Way, #01-02 Nanos, Singapore 138669, Singapore; (K.R.V.T.); (C.E.N.); (S.C.); (C.Y.L.); (V.N.)
| | - Sharon Crasta
- Singapore Institute of Food and Biotechnology Innovation (SIFBI), Agency for Science, Technology and Research (A*STAR), 31 Biopolis Way, #01-02 Nanos, Singapore 138669, Singapore; (K.R.V.T.); (C.E.N.); (S.C.); (C.Y.L.); (V.N.)
| | - Chung Yan Leong
- Singapore Institute of Food and Biotechnology Innovation (SIFBI), Agency for Science, Technology and Research (A*STAR), 31 Biopolis Way, #01-02 Nanos, Singapore 138669, Singapore; (K.R.V.T.); (C.E.N.); (S.C.); (C.Y.L.); (V.N.)
| | - Veronica Ng
- Singapore Institute of Food and Biotechnology Innovation (SIFBI), Agency for Science, Technology and Research (A*STAR), 31 Biopolis Way, #01-02 Nanos, Singapore 138669, Singapore; (K.R.V.T.); (C.E.N.); (S.C.); (C.Y.L.); (V.N.)
| | - Yoganathan Kanagasundaram
- Singapore Institute of Food and Biotechnology Innovation (SIFBI), Agency for Science, Technology and Research (A*STAR), 31 Biopolis Way, #01-02 Nanos, Singapore 138669, Singapore; (K.R.V.T.); (C.E.N.); (S.C.); (C.Y.L.); (V.N.)
- Bioinformatics Institute (BII), Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, #07-01 Matrix, Singapore 138671, Singapore;
- Correspondence: (Y.K.); (H.F.); (S.B.N.); Tel.: +65-6586-9508 (Y.K.); +65-6478-8500 (H.F.); +65-6478-8513 (S.B.N.)
| | - Hao Fan
- Bioinformatics Institute (BII), Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, #07-01 Matrix, Singapore 138671, Singapore;
- Correspondence: (Y.K.); (H.F.); (S.B.N.); Tel.: +65-6586-9508 (Y.K.); +65-6478-8500 (H.F.); +65-6478-8513 (S.B.N.)
| | - Siew Bee Ng
- Singapore Institute of Food and Biotechnology Innovation (SIFBI), Agency for Science, Technology and Research (A*STAR), 31 Biopolis Way, #01-02 Nanos, Singapore 138669, Singapore; (K.R.V.T.); (C.E.N.); (S.C.); (C.Y.L.); (V.N.)
- Bioinformatics Institute (BII), Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, #07-01 Matrix, Singapore 138671, Singapore;
- Correspondence: (Y.K.); (H.F.); (S.B.N.); Tel.: +65-6586-9508 (Y.K.); +65-6478-8500 (H.F.); +65-6478-8513 (S.B.N.)
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Obermaier S, Müller M. Biaryl-Forming Enzymes from Aspergilli Exhibit Substrate-Dependent Stereoselectivity. Biochemistry 2019; 58:2589-2593. [DOI: 10.1021/acs.biochem.9b00291] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Affiliation(s)
- Sebastian Obermaier
- Institute of Pharmaceutical Sciences, Albert-Ludwigs-Universität Freiburg, 79104 Freiburg, Germany
| | - Michael Müller
- Institute of Pharmaceutical Sciences, Albert-Ludwigs-Universität Freiburg, 79104 Freiburg, Germany
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Hou XM, Wang CY, Gerwick WH, Shao CL. Marine natural products as potential anti-tubercular agents. Eur J Med Chem 2019; 165:273-292. [PMID: 30685527 DOI: 10.1016/j.ejmech.2019.01.026] [Citation(s) in RCA: 47] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/16/2018] [Revised: 01/11/2019] [Accepted: 01/11/2019] [Indexed: 02/01/2023]
Abstract
Tuberculosis has been one of the greatest global health challenges of all time. Although the current first-line anti-tuberculosis (anti-TB) medicines used in the clinic have reduced mortality, multidrug-resistance and extensively drug-resistance forms of the disease have now spread worldwide and become a global problem. Even so, few new clinically approved drugs have emerged during the past 30 years. Highly biodiverse marine organisms have received considerable attention for drug discovery in the past couple of decades, and emerging TB drug resistance has motivated interest in assessing marine natural products (MNPs) in the treatment of this disease. So far, more than 170 compounds have been isolated from marine organisms with anti-TB properties, ten of which exhibit potent activity and have the potential for further development. This review systematically surveys MNPs with anti-TB activity and illustrates the impact of these compounds on drug discovery research against tuberculosis.
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Affiliation(s)
- Xue-Mei Hou
- Key Laboratory of Marine Drugs, The Ministry of Education of China, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, People's Republic of China; Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266200, People's Republic of China
| | - Chang-Yun Wang
- Key Laboratory of Marine Drugs, The Ministry of Education of China, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, People's Republic of China; Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266200, People's Republic of China
| | - William H Gerwick
- Center for Marine Biotechnology and Biomedicine, Scripps Institution of Oceanography and Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California San Diego, La Jolla, CA, 92093, United States.
| | - Chang-Lun Shao
- Key Laboratory of Marine Drugs, The Ministry of Education of China, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, People's Republic of China; Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266200, People's Republic of China.
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Demant M, Bagger JI, Suppli MP, Lund A, Gyldenløve M, Hansen KB, Hare KJ, Christensen M, Sonne DP, Holst JJ, Vilsbøll T, Knop FK. Determinants of Fasting Hyperglucagonemia in Patients with Type 2 Diabetes and Nondiabetic Control Subjects. Metab Syndr Relat Disord 2018; 16:530-536. [PMID: 30325692 DOI: 10.1089/met.2018.0066] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
BACKGROUND Fasting hyperglucagonemia can be detrimental to glucose metabolism in patients with type 2 diabetes (T2D) and may contribute to metabolic disturbances in obese and/or prediabetic subjects. However, the mechanisms underlying fasting hyperglucagonemia remain elusive. METHODS We evaluated the interrelationship between fasting hyperglucagonemia and demographic and biochemical parameters in 106 patients with T2D (31% female, age: 57 ± 9 years [mean ± standard deviation; body mass index (BMI): 30.1 ± 4.4 kg/m2; fasting plasma glucose (FPG): 9.61 ± 2.39 mM; hemoglobin A1c (HbA1c): 57.1 ± 13.1 mmol/mol] and 163 nondiabetic control subjects (29% female; age: 45 ± 17 years; BMI: 25.8 ± 4.1 kg/m2; FPG: 5.2 ± 0.4 mM; and HbA1c: 35.4 ± 3.8 mmol/mol). Multiple linear regression analysis was applied using a stepwise approach with fasting plasma glucagon as dependent parameter and BMI, waist-to-hip ratio (WHR), blood pressure, hemoglobin A1c, FPG, and insulin concentrations as independent parameters. RESULTS Fasting plasma glucagon concentrations were significantly higher among patients with T2D (13.5 ± 6.3 vs. 8.5 ± 3.8 mM, P < 0.001) together with HbA1c (P < 0.001), FPG (P < 0.001), and insulin (84.9 ± 56.4 vs. 57.7 ± 35.3 mM, P < 0.001). When adjusted for T2D, HbA1c and insulin were significantly positive determinants for fasting plasma glucagon concentrations. Furthermore, WHR comprised a significant positive determinant. CONCLUSIONS We confirm that fasting plasma glucagon concentrations are abnormally high in patients with T2D, and show that fasting plasma glucagon concentrations are influenced by WHR (in addition to glycemic control and fasting plasma insulin concentrations), which may point to visceral fat deposition as an important determinant of increased fasting plasma glucagon concentrations.
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Affiliation(s)
- Mia Demant
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - Jonatan I Bagger
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - Malte P Suppli
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - Asger Lund
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - Mette Gyldenløve
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - Katrine B Hansen
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - Kristine J Hare
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - Mikkel Christensen
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - David P Sonne
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark
| | - Jens J Holst
- 2 Department of Biomedical Sciences and Faculty of Health and Medical Sciences, University of Copenhagen , Copenhagen, Denmark
| | - Tina Vilsbøll
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark .,3 Department of Clinical Medicine, Faculty of Health and Medical Sciences, University of Copenhagen , Copenhagen, Denmark
| | - Filip K Knop
- 1 Clinical Metabolic Physiology, Steno Diabetes Center Copenhagen, Gentofte Hospital, Hellerup, Denmark .,3 Department of Clinical Medicine, Faculty of Health and Medical Sciences, University of Copenhagen , Copenhagen, Denmark .,4 Novo Nordisk Foundation Center for Basic Metabolic Research, Faculty of Health and Medical Sciences, University of Copenhagen , Copenhagen, Denmark
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Xie XS, Fang XW, Huang R, Zhang SP, Wei HX, Wu SH. A new dimeric anthraquinone from endophytic Talaromyces sp. YE3016. Nat Prod Res 2016; 30:1706-11. [PMID: 26815015 DOI: 10.1080/14786419.2015.1136888] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/22/2022]
Abstract
A new unsymmetrical dimeric anthraquinone, 3-demethyl-3-(2-hydroxypropyl)-skyrin (1) was isolated from the solid-state fermentation extract of an endophytic fungal strain Talaromyces sp. YE 3016, together with five known compounds, skyrin (2), oxyskyrin (3), emodin (4), 1,3,6-trihydroxy-8-methyl-anthraquinone (5) and ergosterol (6). The structure of the new compound was elucidated on the basis of spectroscopic analysis. Compounds 1-3 exhibited moderate cytotoxic activities against MCF-7 cell line.
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Affiliation(s)
- Xiao-Song Xie
- a Key Laboratory for Microbial Resources of the Ministry of Education, Yunnan Institute of Microbiology , Yunnan University , Kunming , P.R. China
| | - Xiao-Wei Fang
- a Key Laboratory for Microbial Resources of the Ministry of Education, Yunnan Institute of Microbiology , Yunnan University , Kunming , P.R. China
| | - Rong Huang
- b School of Chemical Science and Technology , Yunnan University , Kunming , P.R. China
| | - Shou-Peng Zhang
- a Key Laboratory for Microbial Resources of the Ministry of Education, Yunnan Institute of Microbiology , Yunnan University , Kunming , P.R. China
| | - Hong-Xia Wei
- a Key Laboratory for Microbial Resources of the Ministry of Education, Yunnan Institute of Microbiology , Yunnan University , Kunming , P.R. China
| | - Shao-Hua Wu
- a Key Laboratory for Microbial Resources of the Ministry of Education, Yunnan Institute of Microbiology , Yunnan University , Kunming , P.R. China
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Han J, Zhang M, Froese S, Dai FF, Robitaille M, Bhattacharjee A, Huang X, Jia W, Angers S, Wheeler MB, Wei L. The Identification of Novel Protein-Protein Interactions in Liver that Affect Glucagon Receptor Activity. PLoS One 2015; 10:e0129226. [PMID: 26075596 PMCID: PMC4468146 DOI: 10.1371/journal.pone.0129226] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2015] [Accepted: 05/06/2015] [Indexed: 11/18/2022] Open
Abstract
Glucagon regulates glucose homeostasis by controlling glycogenolysis and gluconeogenesis in the liver. Exaggerated and dysregulated glucagon secretion can exacerbate hyperglycemia contributing to type 2 diabetes (T2D). Thus, it is important to understand how glucagon receptor (GCGR) activity and signaling is controlled in hepatocytes. To better understand this, we sought to identify proteins that interact with the GCGR to affect ligand-dependent receptor activation. A Flag-tagged human GCGR was recombinantly expressed in Chinese hamster ovary (CHO) cells, and GCGR complexes were isolated by affinity purification (AP). Complexes were then analyzed by mass spectrometry (MS), and protein-GCGR interactions were validated by co-immunoprecipitation (Co-IP) and Western blot. This was followed by studies in primary hepatocytes to assess the effects of each interactor on glucagon-dependent glucose production and intracellular cAMP accumulation, and then in immortalized CHO and liver cell lines to further examine cell signaling. Thirty-three unique interactors were identified from the AP-MS screening of GCGR expressing CHO cells in both glucagon liganded and unliganded states. These studies revealed a particularly robust interaction between GCGR and 5 proteins, further validated by Co-IP, Western blot and qPCR. Overexpression of selected interactors in mouse hepatocytes indicated that two interactors, LDLR and TMED2, significantly enhanced glucagon-stimulated glucose production, while YWHAB inhibited glucose production. This was mirrored with glucagon-stimulated cAMP production, with LDLR and TMED2 enhancing and YWHAB inhibiting cAMP accumulation. To further link these interactors to glucose production, key gluconeogenic genes were assessed. Both LDLR and TMED2 stimulated while YWHAB inhibited PEPCK and G6Pase gene expression. In the present study, we have probed the GCGR interactome and found three novel GCGR interactors that control glucagon-stimulated glucose production by modulating cAMP accumulation and genes that control gluconeogenesis. These interactors may be useful targets to control glucose homeostasis in T2D.
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Affiliation(s)
- Junfeng Han
- Department of Physiology and Medicine, University of Toronto, Toronto, Ontario, Canada
- Department of Endocrinology and Metabolism, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai, China
| | - Ming Zhang
- Department of Physiology and Medicine, University of Toronto, Toronto, Ontario, Canada
| | - Sean Froese
- Department of Physiology and Medicine, University of Toronto, Toronto, Ontario, Canada
| | - Feihan F. Dai
- Department of Physiology and Medicine, University of Toronto, Toronto, Ontario, Canada
| | - Mélanie Robitaille
- Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, M5S1A8, Canada
| | - Alpana Bhattacharjee
- Department of Physiology and Medicine, University of Toronto, Toronto, Ontario, Canada
| | - Xinyi Huang
- Department of Physiology and Medicine, University of Toronto, Toronto, Ontario, Canada
| | - Weiping Jia
- Department of Endocrinology and Metabolism, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai, China
| | - Stéphane Angers
- Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, M5S1A8, Canada
| | - Michael B. Wheeler
- Department of Physiology and Medicine, University of Toronto, Toronto, Ontario, Canada
- * E-mail: (MW); (LW)
| | - Li Wei
- Department of Endocrinology and Metabolism, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai, China
- * E-mail: (MW); (LW)
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11
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Lotfy M, Kalasz H, Szalai G, Singh J, Adeghate E. Recent Progress in the Use of Glucagon and Glucagon Receptor Antago-nists in the Treatment of Diabetes Mellitus. THE OPEN MEDICINAL CHEMISTRY JOURNAL 2014; 8:28-35. [PMID: 25674162 PMCID: PMC4321206 DOI: 10.2174/1874104501408010028] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/06/2014] [Revised: 10/08/2014] [Accepted: 10/12/2014] [Indexed: 12/25/2022]
Abstract
Glucagon is an important pancreatic hormone, released into blood circulation by alpha cells of the islet of
Langerhans. Glucagon induces gluconeogenesis and glycogenolysis in hepatocytes, leading to an increase in hepatic glucose
production and subsequently hyperglycemia in susceptible individuals. Hyperglucagonemia is a constant feature in
patients with T2DM. A number of bioactive agents that can block glucagon receptor have been identified. These glucagon
receptor antagonists can reduce the hyperglycemia associated with exogenous glucagon administration in normal as well
as diabetic subjects. Glucagon receptor antagonists include isoserine and beta-alanine derivatives, bicyclic 19-residue peptide
BI-32169, Des-His1-[Glu9] glucagon amide and related compounds, 5-hydroxyalkyl-4-phenylpyridines, N-[3-cano-6-
(1,1 dimethylpropyl)-4,5,6,7-tetrahydro-1-benzothien-2-yl]-2-ethylbutamide, Skyrin and NNC 250926. The absorption,
dosage, catabolism, excretion and medicinal chemistry of these agents are the subject of this review. It emphasizes the
role of glucagon in glucose homeostasis and how it could be applied as a novel tool for the management of diabetes mellitus
by blocking its receptors with either monoclonal antibodies, peptide and non-peptide antagonists or gene knockout
techniques.
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Affiliation(s)
- Mohamed Lotfy
- Department of Biology, College of Science, United Arab Emirates University; School of Forensic and Investigative Sciences, University of Central Lancashire, Preston PR1 2HE, England, UK; National Research Centre, Hormones Department, Cairo, Egypt
| | - Huba Kalasz
- Department of Pharmacology and Pharmacotherapy, Semmelweis University, Budapest, Hungary
| | - Gyorgy Szalai
- ENT Department, St. Janos Hospital, Budapest, Hungary
| | - Jaipaul Singh
- School of Forensic and Investigative Sciences and School of Pharmacy and Biomedical Science, University of Central Lancashire, Preston PR1 2HE, England, UK
| | - Ernest Adeghate
- Department of Anatomy, College of Medicine & Health Sciences, United Arab Emirates University, Al Ain, United Ar-ab Emirates
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12
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Abstract
Glucose homeostasis is precisely regulated by glucagon and insulin, which are released by pancreatic α- and β-cells, respectively. While β-cells have been the focus of intense research, less is known about α-cell function and the actions of glucagon. In recent years, the study of this endocrine cell type has experienced a renewed drive. The present review contains a summary of established concepts as well as new information about the regulation of α-cells by glucose, amino acids, fatty acids and other nutrients, focusing especially on glucagon release, glucagon synthesis and α-cell survival. We have also discussed the role of glucagon in glucose homeostasis and in energy and lipid metabolism as well as its potential as a modulator of food intake and body weight. In addition to the well-established action on the liver, we discuss the effects of glucagon in other organs, where the glucagon receptor is expressed. These tissues include the heart, kidneys, adipose tissue, brain, small intestine and the gustatory epithelium. Alterations in α-cell function and abnormal glucagon concentrations are present in diabetes and are thought to aggravate the hyperglycaemic state of diabetic patients. In this respect, several experimental approaches in diabetic models have shown important beneficial results in improving hyperglycaemia after the modulation of glucagon secretion or action. Moreover, glucagon receptor agonism has also been used as a therapeutic strategy to treat obesity.
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13
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Lu RL, Bao GH, Hu FL, Huang B, Li CR, Li ZZ. Comparison of cytotoxic extracts from fruiting bodies, infected insects and cultured mycelia of Cordyceps formosana. Food Chem 2014; 145:1066-71. [DOI: 10.1016/j.foodchem.2013.09.001] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/10/2013] [Revised: 05/23/2013] [Accepted: 09/02/2013] [Indexed: 10/26/2022]
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14
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Podlesny EE, Kozlowski MC. Divergent approach to the bisanthraquinone natural products: total synthesis of (S)-bisoranjidiol and derivatives from binaphtho-para-quinones. J Org Chem 2013; 78:466-76. [PMID: 23249414 PMCID: PMC3560291 DOI: 10.1021/jo302364h] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023]
Abstract
The development of the first asymmetric synthesis of a chiral anthraquinone dimer is outlined, resulting in the first total synthesis of (S)-bisoranjidiol. Rather than a biomimetic dimerization retrosynthetic disconnection, the anthracenyl ring systems are generated after formation of the axially chiral binaphthalene framework. This synthetic strategy has enabled the synthesis of several analogues. Key features of the synthesis include the enantioselective coupling of a hindered 2-naphthol containing substitution peri to the site of C-C bond formation, the regioselective oxidation of 8,8'-hydroxylated binaphthols to binaphtho-para-quinones, and a tandem regioselective Diels-Alder/aromatization reaction.
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Affiliation(s)
- Erin E. Podlesny
- Roy and Diana Vagelos Laboratories, Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6323
| | - Marisa C. Kozlowski
- Roy and Diana Vagelos Laboratories, Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6323
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15
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Jackson MA, Caputo N, Castle JR, David LL, Roberts CT, Ward WK. Stable liquid glucagon formulations for rescue treatment and bi-hormonal closed-loop pancreas. Curr Diab Rep 2012; 12:705-10. [PMID: 22972416 PMCID: PMC3970213 DOI: 10.1007/s11892-012-0320-5] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 10/27/2022]
Abstract
Small doses of glucagon given subcutaneously in the research setting by an automated system prevent most cases of hypoglycemia in persons with diabetes. However, glucagon is very unstable and cannot be kept in a portable pump. Glucagon rapidly forms amyloid fibrils, even within the first day after reconstitution. Aggregation eventually leads to insoluble gels, which occlude pump catheters. Fibrillation occurs rapidly at acid pH, but is absent or minimal at alkaline pH values of ~10. Glucagon also degrades over time; this problem is greater at alkaline pH. Several studies suggest that its primary degradative pathway is deamidation, which results in a conversion of asparagine to aspartic acid. A cell-based assay for glucagon bioactivity that assesses glucagon receptor (GluR) activation can screen promising glucagon formulations. However, mammalian hepatocytes are usually problematic as they can lose GluR expression during culture. Assays for cyclic AMP (cAMP) or its downstream effector, protein kinase A (PKA), in engineered cell systems, are more reliable and suitable for inexpensive, high-throughput assessment of bioactivity.
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Affiliation(s)
- Melanie A Jackson
- Oregon Health and Science University (OHSU), 3181 SW Sam Jackson Park Road, OP05DC, Portland, OR 97239, USA.
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16
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Cho YM, Merchant CE, Kieffer TJ. Targeting the glucagon receptor family for diabetes and obesity therapy. Pharmacol Ther 2012; 135:247-78. [DOI: 10.1016/j.pharmthera.2012.05.009] [Citation(s) in RCA: 93] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2012] [Accepted: 05/15/2012] [Indexed: 12/11/2022]
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17
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Unger RH, Cherrington AD. Glucagonocentric restructuring of diabetes: a pathophysiologic and therapeutic makeover. J Clin Invest 2012; 122:4-12. [PMID: 22214853 DOI: 10.1172/jci60016] [Citation(s) in RCA: 532] [Impact Index Per Article: 40.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022] Open
Abstract
The hormone glucagon has long been dismissed as a minor contributor to metabolic disease. Here we propose that glucagon excess, rather than insulin deficiency, is the sine qua non of diabetes. We base this on the following evidence: (a) glucagon increases hepatic glucose and ketone production, catabolic features present in insulin deficiency; (b) hyperglucagonemia is present in every form of poorly controlled diabetes; (c) the glucagon suppressors leptin and somatostatin suppress all catabolic manifestations of diabetes during total insulin deficiency; (d) total β cell destruction in glucagon receptor-null mice does not cause diabetes; and (e) perfusion of normal pancreas with anti-insulin serum causes marked hyperglucagonemia. From this and other evidence, we conclude that glucose-responsive β cells normally regulate juxtaposed α cells and that without intraislet insulin, unregulated α cells hypersecrete glucagon, which directly causes the symptoms of diabetes. This indicates that glucagon suppression or inactivation may provide therapeutic advantages over insulin monotherapy.
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Affiliation(s)
- Roger H Unger
- Touchstone Center for Diabetes Research, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, Texas 75390-8854, USA.
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18
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Christensen M, Bagger JI, Vilsbøll T, Knop FK. The alpha-cell as target for type 2 diabetes therapy. Rev Diabet Stud 2011; 8:369-81. [PMID: 22262074 DOI: 10.1900/rds.2011.8.369] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Glucagon is the main secretory product of the pancreatic alpha-cells. The main function of this peptide hormone is to provide sustained glucose supply to the brain and other vital organs during fasting conditions. This is exerted by stimulation of hepatic glucose production via specific G protein-coupled receptors in the hepatocytes. Type 2 diabetic patients are characterized by elevated glucagon levels contributing decisively to hyperglycemia in these patients. Accumulating evidence demonstrates that targeting the pancreatic alpha-cell and its main secretory product glucagon is a possible treatment for type 2 diabetes. Several lines of preclinical evidence have paved the way for the development of drugs, which suppress glucagon secretion or antagonize the glucagon receptor. In this review, the physiological actions of glucagon and the role of glucagon in type 2 diabetic pathophysiology are outlined. Furthermore, potential advantages and limitations of antagonizing the glucagon receptor or suppressing glucagon secretion in the treatment of type 2 diabetes are discussed with a focus on already marketed drugs and drugs in clinical development. It is concluded that the development of novel glucagon receptor antagonists are confronted with several safety issues. At present, available pharmacological agents based on the glucose-dependent glucagonostatic effects of GLP-1 represent the most favorable way to apply constraints to the alpha-cell in type 2 diabetes.
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Affiliation(s)
- Mikkel Christensen
- Diabetes Research Division, Department of Internal Medicine F, Gentofte Hospital, University of Copenhagen, Denmark
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19
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Bagger JI, Knop FK, Holst JJ, Vilsbøll T. Glucagon antagonism as a potential therapeutic target in type 2 diabetes. Diabetes Obes Metab 2011; 13:965-71. [PMID: 21615669 DOI: 10.1111/j.1463-1326.2011.01427.x] [Citation(s) in RCA: 95] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
Glucagon is a hormone secreted from the alpha cells of the pancreatic islets. Through its effect on hepatic glucose production (HGP), glucagon plays a central role in the regulation of glucose homeostasis. In patients with type 2 diabetes mellitus (T2DM), abnormal regulation of glucagon secretion has been implicated in the development of fasting and postprandial hyperglycaemia. Therefore, new therapeutic agents based on antagonizing glucagon action, and hence blockade of glucagon-induced HGP, could be effective in lowering both fasting and postprandial hyperglycaemia in patients with T2DM. This review focuses on the mechanism of action, safety and efficacy of glucagon antagonists in the treatment of T2DM and discusses the challenges associated with this new potential antidiabetic treatment modality.
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Affiliation(s)
- J I Bagger
- Diabetes Research Division, Department of Internal Medicine F, Gentofte Hospital, University of Copenhagen, Hellerup, Denmark
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20
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Abstract
Excessive production of glucose by the liver contributes to fasting and postprandial hyperglycaemia, hallmarks of type 2 diabetes. A central feature of this pathologic response is insufficient hepatic insulin action, due to a combination of insulin resistance and impaired β-cell function. However, a case can be made that glucagon also plays a role in dysregulated hepatic glucose production and abnormal glucose homeostasis. Plasma glucagon concentrations are inappropriately elevated in diabetic individuals, and α-cell suppression by hyperglycaemia is blunted. Experimental evidence suggests that this contributes to greater rates of hepatic glucose production in the fasting state and attenuated reduction after meals. Recent studies in animal models indicate that reduction of glucagon action can have profound effects to mitigate hyperglycaemia even in the face of severe hypoinsulinaemia. While there are no specific treatments for diabetic patients yet available that act specifically on the glucagon signalling pathway, newer agents including glucagon-like peptide-1 (GLP-1) receptor agonists and dipeptidyl peptidase-4 (DPP-4) inhibitors reduce plasma glucagon and this is thought to contribute to their action to lower blood glucose. The α-cell and glucagon receptor remain tempting targets for novel diabetes treatments, but it is important to understand the magnitude of benefit new strategies would provide as preclinical models suggest that chronic interference with glucagon action could entail adverse effects as well.
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Affiliation(s)
- D D'Alessio
- Department of Medicine, Division of Endocrinology, Metabolism and Diabetes, University of Cincinnati and Cincinnati VA Medical Center, Cincinnati, OH 45267-0547, USA.
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21
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Abstract
OBJECTIVE To determine the role of glucagon action in the metabolic phenotype of untreated insulin deficiency. RESEARCH DESIGN AND METHODS We compared pertinent clinical and metabolic parameters in glucagon receptor-null (Gcgr(-/-)) mice and wild-type (Gcgr(+/+)) controls after equivalent destruction of β-cells. We used a double dose of streptozotocin to maximize β-cell destruction. RESULTS Gcgr(+/+) mice became hyperglycemic (>500 mg/dL), hyperketonemic, polyuric, and cachectic and had to be killed after 6 weeks. Despite comparable β-cell destruction in Gcgr(-/-) mice, none of the foregoing clinical or laboratory manifestations of diabetes appeared. There was marked α-cell hyperplasia and hyperglucagonemia (~1,200 pg/mL), but hepatic phosphorylated cAMP response element binding protein and phosphoenolpyruvate carboxykinase mRNA were profoundly reduced compared with Gcgr(+/+) mice with diabetes--evidence that glucagon action had been effectively blocked. Fasting glucose levels and oral and intraperitoneal glucose tolerance tests were normal. Both fasting and nonfasting free fatty acid levels and nonfasting β-hydroxy butyrate levels were lower. CONCLUSIONS We conclude that blocking glucagon action prevents the deadly metabolic and clinical derangements of type 1 diabetic mice.
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Affiliation(s)
- Young Lee
- Touchstone Center for Diabetes Research, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, Texas
| | - May-Yun Wang
- Touchstone Center for Diabetes Research, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, Texas
| | - Xiu Quan Du
- Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York
| | - Maureen J. Charron
- Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York
| | - Roger H. Unger
- Touchstone Center for Diabetes Research, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, Texas
- VA North Texas Health Care System, Dallas, Texas
- Corresponding author: Roger H. Unger,
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22
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Nyenwe EA, Jerkins TW, Umpierrez GE, Kitabchi AE. Management of type 2 diabetes: evolving strategies for the treatment of patients with type 2 diabetes. Metabolism 2011; 60:1-23. [PMID: 21134520 PMCID: PMC3746516 DOI: 10.1016/j.metabol.2010.09.010] [Citation(s) in RCA: 216] [Impact Index Per Article: 15.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/10/2010] [Accepted: 09/18/2010] [Indexed: 01/08/2023]
Abstract
The prevalence of type 2 diabetes continues to increase at an alarming rate around the world, with even more people being affected by prediabetes. Although the pathogenesis and long-term complications of type 2 diabetes are fairly well known, its treatment has remained challenging, with only half of the patients achieving the recommended hemoglobin A(1c) target. This narrative review explores the pathogenetic rationale for the treatment of type 2 diabetes, with the view of fostering better understanding of the evolving treatment modalities. The diagnostic criteria including the role of hemoglobin A(1c) in the diagnosis of diabetes are discussed. Due attention is given to the different therapeutic maneuvers and their utility in the management of the diabetic patient. The evidence supporting the role of exercise, medical nutrition therapy, glucose monitoring, and antiobesity measures including pharmacotherapy and bariatric surgery is discussed. The controversial subject of optimum glycemic control in hospitalized and ambulatory patients is discussed in detail. An update of the available pharmacologic options for the management of type 2 diabetes is provided with particular emphasis on newer and emerging modalities. Special attention has been given to the initiation of insulin therapy in patients with type 2 diabetes, with explanation of the pathophysiologic basis for insulin therapy in the ambulatory diabetic patient. A review of the evidence supporting the efficacy of the different preventive measures is also provided.
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Affiliation(s)
- Ebenezer A. Nyenwe
- Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, The University of Tennessee Health Science Center, Memphis, TN 38163, USA
| | | | | | - Abbas E. Kitabchi
- Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, The University of Tennessee Health Science Center, Memphis, TN 38163, USA
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23
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Ichiba H, Ogawa T, Yajima T, Fukushima T. Analysis of hydroxyl radical-induced oxidation process of glucagon by reversed-phase HPLC and ESI-MS/MS. Biomed Chromatogr 2009; 23:1051-8. [PMID: 19402179 DOI: 10.1002/bmc.1222] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Structural modification of a polypeptide hormone, glucagon, by a hydroxyl radical in vitro was investigated by reversed-phase high-performance liquid chromatography (RP-HPLC), and the oxidized site of glucagon was detected by electrospray ionization tandem mass spectrometry (ESI-MS/MS). It was shown that (27)methionine (Met) was oxidized to (27)Met sulfoxide by hydroxyl radical, and the production rate of (27)Met sulfoxide was faster than that by hydrogen peroxide. In addition, production of (27)Met sulfoxide enantiomer was confirmed by RP-HPLC analysis. cAMP production in a HepG2 cell induced by (27)Met sulfoxide glucagon was reduced to approximately 75% as compared with that induced by the native form of glucagon.
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Affiliation(s)
- Hideaki Ichiba
- Department of Analytical Chemistry, Faculty of Pharmaceutical Sciences, Toho University, 2-2-1 Miyama, Funabashi-shi, Chiba 274-8510, Japan
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24
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Brunati M, Rojas JL, Sponga F, Ciciliato I, Losi D, Göttlich E, de Hoog S, Genilloud O, Marinelli F. Diversity and pharmaceutical screening of fungi from benthic mats of Antarctic lakes. Mar Genomics 2009; 2:43-50. [PMID: 21798171 DOI: 10.1016/j.margen.2009.04.002] [Citation(s) in RCA: 62] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2008] [Revised: 04/01/2009] [Accepted: 04/08/2009] [Indexed: 10/20/2022]
Abstract
During the MICROMAT project, the fungal diversity of microbial mats growing in the benthic environment of Antarctic lakes was accessed for the discovery of novel antibiotics and anticancers. In all, 160 filamentous fungi belonging to fifteen different genera and 171 yeasts were isolated from 11 lakes, classified and cultivated in different media and at different temperatures. Filamentous fungi were then screened to discover novel antimicrobial and cytotoxic compounds. A total of 1422 extracts were prepared by solid phase extraction of the culture broths or by biomass solvent extraction. 47 (29%) filamentous fungi showed antimicrobial activity; most of them inhibited the growth of gram-positive Staphyloccus aureus (14%), gram-negative E. coli (10%), and of yeasts Candida albicans (11%) and Cryptococcus neoformans (8%). Less activity was detected against representatives of enterobacteria and filamentous fungi. The most productive in terms of bioactivities were cold-tolerant cosmopolitan hyphomycetes such as Penicillium, Aspergillus, Beauveria and Cladosporium. Two bioactive bis-anthraquinones (rugulosin and skyrin) were identified by LC-MS as the main products in a strain of Penicillium chrysogenum isolated from a saline lake in the Vestfold Hills. LC-MS fractionation of extracts from two diverse species of Aspergillus, that exhibited relatively potent antimicrobial activities, evidenced a chemical novelty that was further investigated. To our knowledge, this is the first report of new antibiotics produced by fungi from benthic microbial mats from Antarctic lakes. It can be concluded that these microbial assemblages represent an extremely rich source for the isolation of new strains producing novel bioactive metabolites with the potential to be developed as drugs.
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Affiliation(s)
- Mara Brunati
- Vicuron Pharmaceuticals (formerly Biosearch Italia S.p.A), Via R. Lepetit 34, 21040 Gerenzano Varese, Italy
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25
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Madsen P, Kodra JT, Behrens C, Nishimura E, Jeppesen CB, Pridal L, Andersen B, Knudsen LB, Valcarce-Aspegren C, Guldbrandt M, Christensen IT, Jørgensen AS, Ynddal L, Brand CL, Bagger MA, Lau J. Human Glucagon Receptor Antagonists with Thiazole Cores. A Novel Series with Superior Pharmacokinetic Properties. J Med Chem 2009; 52:2989-3000. [DOI: 10.1021/jm8016249] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
Affiliation(s)
- Peter Madsen
- Novo Nordisk A/S, Novo Nordisk Park, DK-2760 Maaloev, Denmark
| | - János T. Kodra
- Novo Nordisk A/S, Novo Nordisk Park, DK-2760 Maaloev, Denmark
| | - Carsten Behrens
- Novo Nordisk A/S, Novo Nordisk Park, DK-2760 Maaloev, Denmark
| | - Erica Nishimura
- Novo Nordisk A/S, Novo Nordisk Park, DK-2760 Maaloev, Denmark
| | | | - Lone Pridal
- Novo Nordisk A/S, Novo Nordisk Park, DK-2760 Maaloev, Denmark
| | | | | | | | | | | | | | - Lars Ynddal
- Novo Nordisk A/S, Novo Nordisk Park, DK-2760 Maaloev, Denmark
| | | | | | - Jesper Lau
- Novo Nordisk A/S, Novo Nordisk Park, DK-2760 Maaloev, Denmark
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26
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Burcelin R, Knauf C, Cani P. Pancreatic α-cell dysfunction in diabetes. DIABETES & METABOLISM 2008; 34 Suppl 2:S49-55. [DOI: 10.1016/s1262-3636(08)73395-0] [Citation(s) in RCA: 50] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
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27
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Zhang Y, Ling S, Fang Y, Zhu T, Gu Q, Zhu WM. Isolation, Structure Elucidation, and Antimycobacterial Properties of Dimeric Naphtho-γ-pyrones from the Marine-Derived FungusAspergillus carbonarius. Chem Biodivers 2008; 5:93-100. [DOI: 10.1002/cbdv.200890017] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
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28
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Abstract
Type-2 diabetes is associated with impaired glucose clearance by the liver in the postprandial state, and with elevated glucose production in the post-absorptive state. New targets within the liver are currently being investigated for development of antihyperglycaemic drugs for type-2 diabetes. They include glucokinase, which catalyses the first step in glucose metabolism, the glucagon receptor, and enzymes of gluconeogenesis and/or glycogenolysis such as glucose 6-phosphatase, fructose 1,6-bisphosphatase and glycogen phosphorylase. Preclinical studies with candidate drugs on animal models or cell-based assays suggest that these targets have the potential for pharmacological glycaemic control. Data from clinical studies is awaited. Further work is required for better understanding of the implications of targeting these sites in terms of possible side-effects or tachyphylaxis. The advantage of combined targeting of two or more sites within the liver for minimizing side-effects and tachyphylaxis caused by single-site targeting is discussed.
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Affiliation(s)
- Loranne Agius
- Institute of Cellular Medicine, School of Clinical Medical Sciences, Newcastle University, Newcastle upon Tyne NE2 4HH, UK.
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29
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Rivera N, Everett-Grueter CA, Edgerton DS, Rodewald T, Neal DW, Nishimura E, Larsen MO, Jacobsen LO, Kristensen K, Brand CL, Cherrington AD. A novel glucagon receptor antagonist, NNC 25-0926, blunts hepatic glucose production in the conscious dog. J Pharmacol Exp Ther 2007; 321:743-52. [PMID: 17308040 DOI: 10.1124/jpet.106.115717] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023] Open
Abstract
Elevated glucagon is associated with fasting hyperglycemia in type 2 diabetes. We assessed the effects of the glucagon receptor antagonist (2R)-N-[4-({4-(1-cyclohexen-1-yl)[(3,5-dichloroanilino)carbonyl]anilino}methyl)benzoyl]-2-hydroxy-b-alanine (NNC 25-0926) on hepatic glucose production (HPG) in vivo, using arteriovenous difference and tracer techniques in conscious dogs. The experiments consisted of equilibration (-140 to -40 min), control (40-0 min), and experimental [0-180 min, divided into P1 (0-60 min) and P2 (60-180 min)] periods. In P1, NNC 25-0926 was given intragastrically at 0 (veh), 10, 20, 40, or 100 mg/kg, and euglycemia was maintained. In P2, somatostatin, basal intraportal insulin, and 5-fold basal intraportal glucagon (2.5 ng/kg/min) were infused. Arterial plasma insulin levels remained basal throughout the study in all groups. Arterial plasma glucagon levels remained basal during the control period and P1 and then increased to approximately 70 pg/ml in P2 in all groups. Arterial plasma glucose levels were basal in the control period and P1 in all groups. In P2, the arterial glucose level increased to 245+/-22 and 172+/-15 mg/dl in the veh and 10 mg/kg groups, respectively, whereas in the 20, 40, and 100 mg/kg groups, there was no rise in glucose. Net hepatic glucose output was approximately 2 mg/kg/min in all groups during the control period. In P2, it increased by 9.4+/-2 mg/kg/min in the veh group. In the 10, 20, 40, and 100 mg/kg groups, the rise was only 4.1+/-0.9, 1.6+/-0.6, 2.4+/-0.7, and 1.5+/-0.3 mg/kg/min, respectively, due to inhibition of glycogenolysis. In conclusion, NNC 25-0926 effectively blocked the ability of glucagon to increase HGP in the dog.
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Affiliation(s)
- Noelia Rivera
- Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, 704 Robinson Research Bldg., Nashville, TN 37232-0615, USA
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Gromada J, Franklin I, Wollheim CB. Alpha-cells of the endocrine pancreas: 35 years of research but the enigma remains. Endocr Rev 2007; 28:84-116. [PMID: 17261637 DOI: 10.1210/er.2006-0007] [Citation(s) in RCA: 433] [Impact Index Per Article: 24.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
Glucagon, a hormone secreted from the alpha-cells of the endocrine pancreas, is critical for blood glucose homeostasis. It is the major counterpart to insulin and is released during hypoglycemia to induce hepatic glucose output. The control of glucagon secretion is multifactorial and involves direct effects of nutrients on alpha-cell stimulus-secretion coupling as well as paracrine regulation by insulin and zinc and other factors secreted from neighboring beta- and delta-cells within the islet of Langerhans. Glucagon secretion is also regulated by circulating hormones and the autonomic nervous system. In this review, we describe the components of the alpha-cell stimulus secretion coupling and how nutrient metabolism in the alpha-cell leads to changes in glucagon secretion. The islet cell composition and organization are described in different species and serve as a basis for understanding how the numerous paracrine, hormonal, and nervous signals fine-tune glucagon secretion under different physiological conditions. We also highlight the pathophysiology of the alpha-cell and how hyperglucagonemia represents an important component of the metabolic abnormalities associated with diabetes mellitus. Therapeutic inhibition of glucagon action in patients with type 2 diabetes remains an exciting prospect.
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Affiliation(s)
- Jesper Gromada
- Novartis Institutes for BioMedical Research, 100 Technology Square, Cambridge, Massachusetts 02139, USA.
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31
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Sloop KW, Michael MD, Moyers JS. Glucagon as a target for the treatment of Type 2 diabetes. Expert Opin Ther Targets 2007; 9:593-600. [PMID: 15948676 DOI: 10.1517/14728222.9.3.593] [Citation(s) in RCA: 56] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
Glucagon is the key counter-regulatory hormone that opposes the action of insulin. In states of relative hypoglycaemia, glucagon acts to increase blood glucose by stimulating hepatic glycogen breakdown and gluconeogenesis to achieve euglycaemia. Type 2 diabetes is characterised by inappropriate regulation of hepatic glucose production, which is due, at least in part, to an imbalance in the bihormonal relationship between plasma levels of glucagon and insulin. The glucose-lowering effects of glucagon peptide antagonists and antiglucagon neutralising antibodies first demonstrated the potential of glucagon receptor (GCGR) antagonism as a treatment for hyperglycaemia. In recent years, the development of GCGR antisense oligonucleotides and small molecular weight GCGR antagonists have been pursued as possible therapeutic agents to target glucagon action as a treatment for Type 2 diabetes.
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Affiliation(s)
- Kyle W Sloop
- Endocrine Discovery, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285, USA.
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Yang X, Yates ML, Candelore MR, Feeney W, Hora D, Kim RM, Parmee ER, Berger JP, Zhang BB, Qureshi SA. Cloning and expression of canine glucagon receptor and its use to evaluate glucagon receptor antagonists in vitro and in vivo. Eur J Pharmacol 2007; 555:8-16. [PMID: 17140563 DOI: 10.1016/j.ejphar.2006.10.033] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2006] [Revised: 10/09/2006] [Accepted: 10/11/2006] [Indexed: 10/24/2022]
Abstract
Glucose homeostasis is maintained by the combined actions of insulin and glucagon. Hyperglucagonemia and/or elevation of glucagon/insulin ratio have been reported in diabetic patients and in animal models of diabetes. Therefore, antagonizing glucagon receptor function has long been considered a useful approach to lower hyperglycemia. Dogs serve as an excellent model for studying glycemic control and various aspects of glucagon biology in vivo; however, the amino acid sequence of the dog glucagon receptor has not been reported. To better understand the pharmacology of the dog glucagon receptor and to characterize glucagon receptor antagonists, we cloned a cDNA corresponding to the glucagon receptor from dog liver RNA. The dog glucagon receptor shares a significant (>75%) homology at both nucleotide and amino acid levels with the glucagon receptor from human, monkey, mouse, and rat. The protein is highly conserved among all species in areas corresponding to the 7 trans-membrane domains. However, it shows significant divergence at the carboxy terminus such that the receptor from dog has the longest cytoplasmic tail among all species examined. When expressed in chinese hamster ovary cells, the dog glucagon receptor bound [125I]Glucagon with a K(d) of 477+/-106 pM. Glucagon stimulated the rise of intracellular cAMP levels in these cells with an EC(50) of 9.6+/-1.7 nM and such effects could be blocked by known peptidyl and non-peptidyl small molecule antagonists. In addition we show that a small molecule glucagon receptor antagonist with significant activity in cell based assays also blocked the ability of glucagon to induce elevation in blood glucose in beagle dogs. These data demonstrate that the cloned cDNA encodes a functional dog glucagon receptor. The availability of the dog cDNA will facilitate the understanding of glucagon pharmacology and aid in the characterization of novel glucagon antagonists that may serve as anti-hyperglycemic treatment for type 2 diabetes mellitus.
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Affiliation(s)
- Xiaodong Yang
- Department of Metabolic Disorder-Molecular Endocrinology, Merck Research Laboratories, Rahway, NJ 07065, USA
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Raju B, Cryer PE. Maintenance of the postabsorptive plasma glucose concentration: insulin or insulin plus glucagon? Am J Physiol Endocrinol Metab 2005; 289:E181-6. [PMID: 16014355 DOI: 10.1152/ajpendo.00460.2004] [Citation(s) in RCA: 31] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
The prevalent view is that the postabsorptive plasma glucose concentration is maintained within the physiological range by the interplay of the glucose-lowering action of insulin and the glucose-raising action of glucagon. It is supported by a body of evidence derived from studies of suppression of glucagon (and insulin, among other effects) with somatostatin in animals and humans, immunoneutralization of glucagon, defective glucagon synthesis, diverse mutations, and absent or reduced glucagon receptors in animals and glucagon antagonists in cells, animals, and humans. Many of these studies are open to alternative interpretations, and some lead to seemingly contradictory conclusions. For example, immunoneutralization of glucagon lowered plasma glucose concentrations in rabbits, but administration of a glucagon antagonist did not lower plasma glucose concentrations in healthy humans. Evidence that the glycemic threshold for glucagon secretion, unlike that for insulin secretion, lies below the physiological range, and the finding that selective suppression of insulin secretion without stimulation of glucagon secretion raises fasting plasma glucose concentrations in humans underscore the primacy of insulin in the regulation of the postabsorptive plasma glucose concentration and challenge the prevalent view. The alternative view is that the postabsorptive plasma glucose concentration is maintained within the physiological range by insulin alone, specifically regulated increments and decrements in insulin, and the resulting decrements and increments in endogenous glucose production, respectively, and glucagon becomes relevant only when glucose levels drift below the physiological range. Although the balance of evidence suggests that glucagon is involved in the maintenance of euglycemia, more definitive evidence is needed, particularly in humans.
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Affiliation(s)
- Bharathi Raju
- Division of Endocrinology, Metabolism and Lipid Research, Washington Univ. School of Medicine, 660 South Euclid Ave., St. Louis, MO 63110, USA
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34
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Dallas-Yang Q, Shen X, Strowski M, Brady E, Saperstein R, Gibson RE, Szalkowski D, Qureshi SA, Candelore MR, Fenyk-Melody JE, Parmee ER, Zhang BB, Jiang G. Hepatic glucagon receptor binding and glucose-lowering in vivo by peptidyl and non-peptidyl glucagon receptor antagonists. Eur J Pharmacol 2005; 501:225-34. [PMID: 15464082 DOI: 10.1016/j.ejphar.2004.08.023] [Citation(s) in RCA: 34] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2004] [Revised: 08/05/2004] [Accepted: 08/10/2004] [Indexed: 11/22/2022]
Abstract
Glucagon receptor antagonists have been actively pursued as potential therapeutics for the treatment of type 2 diabetes. Peptidyl and non-peptidyl glucagon receptor antagonists have been shown to block glucagon-induced blood glucose elevation in both animals and humans. How the antagonists and the glucagon receptor interact in vivo has not been reported and is the subject of the current study. Using (125)I-labeled glucagon as a radiotracer, we developed an in vivo glucagon receptor occupancy assay in mice expressing a human glucagon receptor in place of the endogenous mouse glucagon receptor (hGCGR mice). Using this assay, we first showed that the glucagon receptor is expressed predominantly in liver, to a much lesser extent in kidney, and is below detection in several other tissues/organs in the mice. We subsequently showed that, at 2 mg/kg body weight (mg/pk) dosed intraperitoneally (i.p.), peptidyl glucagon receptor antagonist des-His-glucagon binds to approximately 78% of the hepatic glucagon receptor and blocks an exogenous glucagon-induced blood glucose elevation in the mice. Finally, we also showed that, at 10 and 30 mg/kg dosed orally (p.o.), compound A, a non-peptidyl small molecule glucagon receptor antagonist, occupied 65-70% of the hepatic glucagon receptor, and significantly diminished exogenous glucagon-induced blood glucose elevation in the mice. At 3 mg/kg, however, compound A occupied only approximately 39% of the hepatic glucagon receptor and did not affect exogenous glucagon-induced blood glucose elevation in the mice. Taken together, the results confirmed previous reports that glucagon receptors are present predominantly in the liver, and provide the first direct evidence that peptidyl and non-peptidyl glucagon receptor antagonists bind to the hepatic glucagon receptor in vivo, and that at least 60% receptor occupancy correlates with the glucose lowering efficacy by the antagonists in vivo.
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Affiliation(s)
- Qing Dallas-Yang
- Metabolic Disorders-Diabetes, Merck Research Laboratories, Rahway, NJ 07065, USA
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35
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36
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Qureshi SA, Rios Candelore M, Xie D, Yang X, Tota LM, Ding VDH, Li Z, Bansal A, Miller C, Cohen SM, Jiang G, Brady E, Saperstein R, Duffy JL, Tata JR, Chapman KT, Moller DE, Zhang BB. A novel glucagon receptor antagonist inhibits glucagon-mediated biological effects. Diabetes 2004; 53:3267-73. [PMID: 15561959 DOI: 10.2337/diabetes.53.12.3267] [Citation(s) in RCA: 84] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/13/2022]
Abstract
Glucagon maintains glucose homeostasis during the fasting state by promoting hepatic gluconeogenesis and glycogenolysis. Hyperglucagonemia and/or an elevated glucagon-to-insulin ratio have been reported in diabetic patients and animals. Antagonizing the glucagon receptor is expected to result in reduced hepatic glucose overproduction, leading to overall glycemic control. Here we report the discovery and characterization of compound 1 (Cpd 1), a compound that inhibits binding of 125I-labeled glucagon to the human glucagon receptor with a half-maximal inhibitory concentration value of 181 +/- 10 nmol/l. In CHO cells overexpressing the human glucagon receptor, Cpd 1 increased the half-maximal effect for glucagon stimulation of adenylyl cyclase with a KDB of 81 +/- 11 nmol/l. In addition, Cpd 1 blocked glucagon-mediated glycogenolysis in primary human hepatocytes. In contrast, a structurally related analog (Cpd 2) was not effective in blocking glucagon-mediated biological effects. Real-time measurement of glycogen synthesis and breakdown in perfused mouse liver showed that Cpd 1 is capable of blocking glucagon-induced glycogenolysis in a dosage-dependent manner. Finally, when dosed in humanized mice, Cpd 1 blocked the rise of glucose levels observed after intraperitoneal administration of exogenous glucagon. Taken together, these data suggest that Cpd 1 is a potent glucagon receptor antagonist that has the capability to block the effects of glucagon in vivo.
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Affiliation(s)
- Sajjad A Qureshi
- Department of Metabolic Disorder and Molecular Endocrinology, Merck Research Laboratories, Rahway, New Jersey, USA.
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37
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Potterat O, Wagner K, Gemmecker G, Mack J, Puder C, Vettermann R, Streicher R. BI-32169, a bicyclic 19-peptide with strong glucagon receptor antagonist activity from Streptomyces sp. JOURNAL OF NATURAL PRODUCTS 2004; 67:1528-1531. [PMID: 15387654 DOI: 10.1021/np040093o] [Citation(s) in RCA: 59] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/24/2023]
Abstract
A new bicyclic 19-peptide, BI-32169, has been isolated from the culture broth of Streptomyces sp. (DSM 14996). Its structure has been established by amino acid analysis, mass spectrometry, and 2D NMR analysis. BI-32169 consists exclusively of protein amino acids and is cyclized from the side chain of Asp(9) to the N-terminus of Gly(1). One disulfide bond between Cys(6) and Cys(19) forms a bicyclic structure. BI-32169 and its methyl ester derivative showed potent inhibitory activity against the human glucagon receptor (IC(50) 440 and 320 nM, respectively) in a functional cell-based assay.
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Affiliation(s)
- Olivier Potterat
- Boehringer Ingelheim Pharma GmbH and Co. KG, Birkendorfer Strasse 65, D-88397 Biberach an der Riss, Germany.
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38
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Sapse AM, Lawton S, Rothchild R, Unson CG. An ab initio study of non-peptide glucagon receptor antagonists. ACTA ACUST UNITED AC 2003. [DOI: 10.1016/j.theochem.2003.07.001] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/27/2022]
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Abstract
As a counterregulatory hormone for insulin, glucagon plays a critical role in maintaining glucose homeostasis in vivo in both animals and humans. To increase blood glucose, glucagon promotes hepatic glucose output by increasing glycogenolysis and gluconeogenesis and by decreasing glycogenesis and glycolysis in a concerted fashion via multiple mechanisms. Compared with healthy subjects, diabetic patients and animals have abnormal secretion of not only insulin but also glucagon. Hyperglucagonemia and altered insulin-to-glucagon ratios play important roles in initiating and maintaining pathological hyperglycemic states. Not surprisingly, glucagon and glucagon receptor have been pursued extensively in recent years as potential targets for the therapeutic treatment of diabetes.
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Affiliation(s)
- Guoqiang Jiang
- Department of Metabolic Disorders and Molecular Endocrinology, Merck Research Laboratory, Rahway, New Jersey 07065, USA
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40
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Yao YQ, Zhang DF, Huang AL, Luo Y, Zhang DZ, Wang B, Zhou WP, Ren H, Guo SH. Effects of electroporation on primary rat hepatocytes in vitro. World J Gastroenterol 2002; 8:893-6. [PMID: 12378637 PMCID: PMC4656582 DOI: 10.3748/wjg.v8.i5.893] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/09/2001] [Revised: 05/10/2002] [Accepted: 05/15/2002] [Indexed: 02/06/2023] Open
Abstract
AIM To investigate the effects of electroporation on primary rat hepatocyte and to optimize the electroporation conditions introducing foreign genes into primary hepatocytes. METHODS A single-pulse procedure was performed at low voltage (220-400 V) but with high capacitance (500-950 microF). Hepatocytes were divided into 4 groups according to the electroporation conditions: group I, 220 V and 500 microF; group II, 220 V and 950 microF; group III, 400 V and 950 microF,and group IV. The control group was freshly isolated hepatocytes and directly cultured under the same conditions as those of electroporation groups. The effects of electroporation on primary rat hepatocytes were detected by trypan blue exclusion (TBE) and MTT analysis. Besides, albumin (Alb), alanine transaminase (ALT) and lactate dehydrogenase (LDH) in the supernatants of cultured hepatocytes were measured by biochemical assay. RESULTS Between day 1 and day 15 after incubation, primary rat hepatocytes of each electroporation group appeared normal, being the same with those of control group. TBE staining showed that slight hepatocyte damage and high survival rate were found in the electroporation groups and the control group. Cultured for 3, 7, 11 and 15 days, hepatocyte viability was approximately 92.6+/-2.5 %, 89.5+/-3.3 %, 82.0+/-3.5 % and 74.3+/-1.2 %, respectively. MTT analysis indicated that the viabilities of hepatocytes had no significant difference between each electroporation group, and those were similar to that of control group. At the 36th hour after electroporation, Alb, ALT and LDH in the supernatants of control group were 5.3+/-0.1 g x L(-1), 183.7+/-8.4 nkat x L(-1) and 896.8+/-58.5 nkat x L(-1); those of group II were 5.7+/-0.1 g x L(-1), 215.4+/-16.7 nkat x L(-1) and 1063.8+/-51.8 nkat x L(-1); and those of group III were 5.8+/-0.2 g x L(-1), 217.1+/-8.4 nkat x L(-1) and 1063.8+/-10.0 nkat x L(-1). Statistically, the proteins of group II and group III were significantly higher than those of control group (P<0.05), whereas the protein production of group I, Alb, ALT and LDH were 5.3+/-0.2 g x L(-1), 205.4+/-3.3 nkat x L(-1) and 1035.4+/-116.9 nkat x L(-1), were similar to those of control group. At the same time, TBE and MTT analysis indicated that there was no significant cell viability difference between electroporation groups and control group. CONCLUSION This single-pulse electroporation procedure performed at low voltage (220-400 V) but with high capacitance (950 microF) is one of the optimal choices to introduce foreign genes into primary rat hepatocyte.
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Affiliation(s)
- Yun-Qing Yao
- Department of Infectious Diseases of the First Affiliated Hospital, Chongqing University of Medical Sciences, Chongqing 400016, China.
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Dallas-Yang Q, Qureshi SA, Xie D, Zhang BB, Jiang G. Detection of glucagon-dependent GTPgammaS binding in high-throughput format. Anal Biochem 2002; 301:156-9. [PMID: 11811981 DOI: 10.1006/abio.2001.5483] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Affiliation(s)
- Qing Dallas-Yang
- Department of Molecular Endocrinology, Merck Research Laboratories, RY80N-C31, Rahway, New Jersey 07065, USA
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Parker JC, Andrews KM, Allen MR, Stock JL, McNeish JD. Glycemic control in mice with targeted disruption of the glucagon receptor gene. Biochem Biophys Res Commun 2002; 290:839-43. [PMID: 11785978 DOI: 10.1006/bbrc.2001.6265] [Citation(s) in RCA: 121] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
The action of glucagon in the liver is mediated by G-coupled receptors. To examine the role of glucagon in glucose homeostasis, we have generated mice in which the glucagon receptor was inactivated (GR(-/-) mice). Blood glucose levels were somewhat reduced in GR(-/-) mice relative to wild type, in both the fed and fasted state. Plasma insulin levels were not significantly affected. There was no significant effect on fasting plasma cholesterol or triglyceride levels associated with deletion of the glucagon receptor. Glucose tolerance, as assessed by an oral glucose tolerance test, improved. Plasma glucagon levels were strikingly elevated in both fed and fasted animals. Despite a total absence of glucagon receptors, these animals maintained near-normal glycemia and normal lipidemia, in the presence of circulating glucagon concentrations that were elevated by two orders of magnitude.
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Affiliation(s)
- Janice C Parker
- Pfizer Global Research & Development, Groton Laboratories, Groton, Connecticut 06340, USA.
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