TY - JOUR
T1 - Pure glucagon antagonists
T2 - Biological activities and camp accumulation using phosphodiesterase inhibitors
AU - Azizeh, Bassem Y.
AU - Van Tine, Brian A.
AU - Trivedi, Dev
AU - Hruby, Victor J.
N1 - Funding Information:
This research was supported by a grant from U.S. Public Health Service Grant DK 21085 and by HHMI Grant 71109-52130. Its contents are solely the responsibility of the authors and do not represent the official views of USPHS. We thank Susan Hitesman, Melissa Berren, Jason Phelps and Rael Caspari for their assistance with some of the bioassays.
PY - 1997
Y1 - 1997
N2 - Five new glucagon analogues have been designed, synthesized, characterized and their biological activities tested. The investigation was centered on modifications in the N-terminal region in particular, residues at Thr5, Phe6 and Tyr10 positions, with the goal of obtaining pure glucagon antagonists in our newly developed high sensitivity cAMP accumulation assay. The structures of the designed compounds are: [des-His1, des-Phe6. Glu9 ]glucagon-NH2 (1); [des-His1, des-Phe6, Glu9, Phe10] glucagon-NH2 (2): [des-His1, Tyr5, des-Phe6, Glu9] glucagon NH2 (3): [des-His1, Phe5, des-Phe6, Glu9]glucagon-NH2 (4) and [des-His1, des-Phe6, Glu9, D- Arg18]glucagon-NH2 (5). The binding potencies IC50 values in (nM) were 48.0, 27.4, 26.0, 20.0 and 416.0, respectively. All of these analogues when tested in the classical adenylate cyclase assay demonstrate antagonist properties, and in competition experiments, all caused a right-wardshift of the glucagon stimulated adenylate cyclase dose-response curve. The pA2 values for these analogues were 8.20 (1); 6.25 (2): 6.10 (3); 6.25 (4); and 6.08 (5), respectively. A newly revised assay has been developed to determine the intracellular cAMP accumulation levels in hepatocytes at the highest possible sensitivity. Four of the five glucagon analogues in this report (analogues 1, 2, 4 and 5), did not activate the adenylate cyclase in the presence of Rolipram up to a maximal physiological concentration of 1 μM, and thus are pure antagonists.
AB - Five new glucagon analogues have been designed, synthesized, characterized and their biological activities tested. The investigation was centered on modifications in the N-terminal region in particular, residues at Thr5, Phe6 and Tyr10 positions, with the goal of obtaining pure glucagon antagonists in our newly developed high sensitivity cAMP accumulation assay. The structures of the designed compounds are: [des-His1, des-Phe6. Glu9 ]glucagon-NH2 (1); [des-His1, des-Phe6, Glu9, Phe10] glucagon-NH2 (2): [des-His1, Tyr5, des-Phe6, Glu9] glucagon NH2 (3): [des-His1, Phe5, des-Phe6, Glu9]glucagon-NH2 (4) and [des-His1, des-Phe6, Glu9, D- Arg18]glucagon-NH2 (5). The binding potencies IC50 values in (nM) were 48.0, 27.4, 26.0, 20.0 and 416.0, respectively. All of these analogues when tested in the classical adenylate cyclase assay demonstrate antagonist properties, and in competition experiments, all caused a right-wardshift of the glucagon stimulated adenylate cyclase dose-response curve. The pA2 values for these analogues were 8.20 (1); 6.25 (2): 6.10 (3); 6.25 (4); and 6.08 (5), respectively. A newly revised assay has been developed to determine the intracellular cAMP accumulation levels in hepatocytes at the highest possible sensitivity. Four of the five glucagon analogues in this report (analogues 1, 2, 4 and 5), did not activate the adenylate cyclase in the presence of Rolipram up to a maximal physiological concentration of 1 μM, and thus are pure antagonists.
KW - Adenylate cyclase
KW - Amrinone
KW - Glucagon antagonists
KW - Hepatocytes
KW - IBMX
KW - Phosphodiesterase inhibitors
KW - Receptor binding
KW - Rolipram
KW - Structure-activity relationships
KW - cAMP accumulation
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U2 - 10.1016/S0196-9781(97)00131-9
DO - 10.1016/S0196-9781(97)00131-9
M3 - Article
C2 - 9213355
AN - SCOPUS:0030794042
SN - 0196-9781
VL - 18
SP - 633
EP - 641
JO - Peptides
JF - Peptides
IS - 5
ER -