Vasily M Gelfanov

Publications (22) View all

• Source

  Available from: nfdx.org

  Article: A novel human-based receptor antagonist of sustained action reveals body weight control by endogenous GLP-1.

  James T Patterson, Nickki Ottaway, Vasily M Gelfanov, David L Smiley, Diego Perez-Tilve, Paul T Pfluger, Matthias H Tschöp, Richard D Dimarchi
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  ABSTRACT: Ex-4 (9-39)a is a well characterized GLP-1 receptor antagonist that suffers from two notable limitations, its nonhuman amino acid sequence and its relatively short in vivo duration of action. Comparable N-terminal shortening of human GLP-1 lessens agonism but does not provide a high potency antagonist. Through a series of GLP-1/Ex-4 hybrid peptides, the minimal structural changes required to generate a pure GLP-1-based antagonist were identified as Glu16, Val19, and Arg20, yielding an antagonist of approximately 3-fold greater in vitro potency compared with Ex-4 (9-39)a. The structural basis of antagonism appears to result from stabilization of the α helix combined with enhanced electrostatic and hydrophobic interactions with the extracellular domain of the receptor. Site-specific acylation of the human-based antagonist yielded a peptide of increased potency as a GLP-1 receptor antagonist and 10-fold greater selectivity relative to the GIP receptor. The acylated antagonist demonstrated sufficient duration of action to maintain inhibitory activity when administered as a daily subcutaneous injection. The sustained pharmacokinetics and enhanced human sequence combine to form an antagonist optimized for clinical study. Daily administration of this antagonist by subcutaneous injection to diet-induced obese mice for 1 week caused a significant increase in food intake, body weight, and glucose intolerance, demonstrating endogenous GLP-1 as a relevant hormone in mammalian energy balance in the obese state.
  ACS Chemical Biology 10/2010; 6(2):135-45. · 6.45 Impact Factor
• Article: Glucagon regulation of energy metabolism.

  Kristy M Heppner, Kirk M Habegger, Jonathan Day, Paul T Pfluger, Diego Perez-Tilve, Brian Ward, Vasily Gelfanov, Steve C Woods, Richard DiMarchi, Matthias Tschöp
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  ABSTRACT: Glucagon has long been known as a counter-regulatory hormone to insulin of fundamental importance to glucose homeostasis. Its prominent ability to stimulate glycogenolysis and gluconeogenesis, has historically cast this peptide as one hormone where the metabolic consequences of increasing blood glucose levels, especially in obesity, are viewed largely as being deleterious. This perspective may be changing in light of emerging data and reconsideration of historic studies, which suggest that glucagon has beneficial effects on body fat mass, food intake, and energy expenditure. In this review, we discuss the mechanisms of glucagon-mediated body weight regulation as well as possible novel therapeutic approaches in the treatment of obesity and glucose intolerance that may arise from these findings. The paper represents an invited review by a symposium, award winner or keynote speaker at the Society for the Study of Ingestive Behavior [SSIB] Annual Meeting in Portland, July 2009.
  Physiology & Behavior 04/2010; 100(5):545-8. · 2.87 Impact Factor
• Article: Functional association of the N-terminal residues with the central region in glucagon-related peptides.

  James T Patterson, Jonathan W Day, Vasily M Gelfanov, Richard D DiMarchi
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  ABSTRACT: GLP-1 is an incretin peptide involved in the regulation of glucose metabolism and the glucose-dependent stimulation of insulin secretion. Ex-4 is a paralog of GLP-1 that has comparable GLP-1R potency but extended physiological action. GLP-1 and Ex-4 are helical peptides that share ∼50% sequence homology but differ at several residues, notably the second amino acid which controls susceptibility to DPP-IV cleavage. This single amino acid difference yields divergent receptor potency when studied in the context of the two hormone sequences. Ex-4 uniquely tolerates Gly2 through select amino acid differences in the middle region of the peptide that are absent in GLP-1. We report that substitution of Ex-4 amino acids Glu16, Leu21, and Glu24 to the GLP-1 sequence enabled Gly2 tolerance. The coordination of the N-terminus with these central residues shows an interaction of substantial importance not only to DPP-IV stability but also to receptor activation. Extension of this observation to glucagon-based co-agonist peptides showed different structural requirements for effective communication between the N-terminus and the mid-section of these peptides in achieving high potency agonism at the GLP-1 and GCGRs.
  Journal of Peptide Science 06/2011; 17(10):659-66. · 1.80 Impact Factor
• Article: Charge inversion at position 68 of the glucagon and glucagon-like peptide-1 receptors supports selectivity in hormone action.

  Jonathan W Day, Pengyun Li, James T Patterson, Joe Chabenne, Maria Dimarchi Chabenne, Vasily M Gelfanov, Richard D Dimarchi
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  ABSTRACT: Glucagon and glucagon-like peptide-1 (GLP-1)are two structurally related hormones that acutely regulate glucose control in opposite directions through homologous receptors. The molecular basis for selectivity between these two hormones and their receptors is of physiological and medicinal importance. The application of co-agonists to enhance body weight reduction and correct multiple abnormalities associated with the metabolic syndrome has recently been reported. Substitution of amino acids 16, 18, and 20 in glucagon with those found in GLP-1 and exendin-4 were identified as partial contributors to balanced, high potency receptor action. The amidation of the C-terminus was an additional glucagon-based structural change observed to be of seminal importance to discriminate recognition by both receptors. In this work, the molecular basis for receptor selectivity associated with differences in C-terminal peptide sequence has been determined. A single charge inversion in glucagon and GLP-1 receptor sequence at position 68* was determined to significantly alter hormone action. Changing E68* in GLP-1R to the corresponding Lys of GCGR reduced receptor activity for natural GLP-1 hormones by eightfold. The enhanced C-terminal positive charges in GLP-1 peptides favor the native receptor's negative charge at position 68*, while the unfavorable interaction with the C-terminal acid of native glucagon is minimized by amidation. The extension of these observations to other glucagon-related hormones such as oxyntomodulin and exendin, as well as other related receptors such as GIPR, should assist in the assembly of additional hormones with broadened pharmacology.
  Journal of Peptide Science 03/2011; 17(3):218-25. · 1.80 Impact Factor
• Source

  Available from: journalofdst.org

  Article: Optimization of the native glucagon sequence for medicinal purposes.

  Joseph R Chabenne, Maria A DiMarchi, Vasily M Gelfanov, Richard D DiMarchi
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  ABSTRACT: Glucagon is a life-saving medication used in the treatment of hypoglycemia. It possesses poor solubility in aqueous buffers at or near physiological pH values. At low and high pH, at which the peptide can be formulated to concentrations of a milligram or more per milliliter, the chemical integrity of the hormone is limited, as evidenced by the formation of multiple degradation-related peptides. Consequently, the commercial preparation is provided as a lyophilized solid with an acidic diluent and directions for rendering it soluble at the time of use. Any unused material is recommended for disposal immediately after initial use. A set of glucagon analogs was prepared by solid-phase peptide synthesis to explore the identification of a glucagon analog with enhanced solubility and chemical stability at physiological pH. The physical properties of the peptide analogs were studied by solubility determination, high-performance chromatography, and mass spectral analysis. The biochemical properties were determined in engineered human embryonic kidney cell line 293 (HEK293) cells that overexpressed either the human glucagon or glucagon-like peptide-1 (GLP-1) receptors linked to a luciferase reporter gene. We observed the previously characterized formation of glucagon degradation products upon incubation of the peptide in dilute acid for extended periods or elevated temperature. Lowering the isoelectric point of the hormone through the substitution of asparagine-28 with aspartic acid significantly increased the solubility at physiological pH. Similarly, the C-terminal extension (Cex) of the hormone with an exendin-based, 10-residue, C-terminal sequence yielded a peptide of dramatically enhanced solubility. These two glucagon analogs, D28 and Cex, maintained high potency and selectivity for the glucagon receptor relative to GLP-1 receptor. Glucagon presents unique structural challenges to the identification of an analog of high biological activity and selectivity that also possesses sufficient aqueous solubility and stability such that it might be developed as a ready-to-use medicine. The glucagon analogs D28 and Cex demonstrated all of the chemical, physical, and biochemical properties supportive of further study as potential clinical candidates for treatment of hypoglycemia.
  Journal of diabetes science and technology 01/2010; 4(6):1322-31.