title: Effects of non-covalent self-association on the subcutaneous absorption of a therapeutic peptide.
abstract: OBJECTIVE
  To utilize an acylated peptide as a model system to investigate the relationships among solution peptide conformation, non-covalent self-association, subcutaneous absorption and bioavailability under pharmaceutically relevant solution formulation conditions.
  
  
  METHODS
  CD spectroscopy, FTIR spectroscopy, equilibrium sedimentation, dynamic light scattering, and size exclusion chromatography were employed to characterize the effects of octanoylation on conformation and self-association of the 31 amino acid peptide derivative des-amino-histidine(7) arginine(26) human glucagon-like peptide (7-37)-OH (IP(7)R(26)GLP-1). Hyperglycemic clamp studies were performed to compare the bioavailability, pharmacokinetics, and pharmacodynamics of solution formulations of oct-IP(7)R(26)GLP-1 administered subcutaneously to normal dogs.
  
  
  RESULTS
  Octanoylation of IP(7)R(26)GLP-1 was shown to confer the propensity for a major solvent-induced conformational transition with an accompanying solvent- and temperature-dependent self-association behavior. Formulations were characterized that give rise to remarkably different pharmacodynamics and pharmacokinetics that correlate with distinct peptide conformational and self-association states. These states correspond to: (i) a minimally associated alpha-helical form (apparent molecular weight = 14 kDa), (ii) a highly associated, predominantly beta-sheet form (effective molecular diameter 20 nm), and (iii) an unusually large, micelle-like soluble beta-sheet aggregate (effective molecular diameter 50 nm).
  
  
  CONCLUSIONS
  Bioavailability and pharmacokinetics of a self-associating peptide can be influenced by aggregate size and the ease of disruption of the non-covalent intermolecular interactions at the subcutaneous site. Hydrophobic aggregation mediated by seemingly innocuous solution formulation conditions can have a dramatic effect on the subcutaneous bioavailability and pharmacokinetics of a therapeutic peptide and in the extreme, can totally preclude its absorption. A size exclusion chromatographic method is identified that distinguishes subcutaneously bioavailable aggregated oct-IP(7)R(26)GLP-1 from non-bioavailable aggregated oct-IP(7)R(26)GLP-1.
authors.affiliation: Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285, USA.
authors.forename: D K
authors.initials: DK
authors.lastname: Clodfelter
authors.forename: A H
authors.initials: AH
authors.lastname: Pekar
authors.forename: D M
authors.initials: DM
authors.lastname: Rebhun
authors.forename: K A
authors.initials: KA
authors.lastname: Destrampe
authors.forename: H A
authors.initials: HA
authors.lastname: Havel
authors.forename: S R
authors.initials: SR
authors.lastname: Myers
authors.forename: M L
authors.initials: ML
authors.lastname: Brader
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002850:Chromatography, Gel; D002942:Circular Dichroism; D004285:Dogs; D005260:Female; D005934:Glucagon / Q000008:administration & dosage* / Q000737:chemistry / Q000493:pharmacokinetics*; D052216:Glucagon-Like Peptide 1; D007279:Injections, Subcutaneous; D007328:Insulin / Q000097:blood; D008027:Light; D008297:Male; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000008:administration & dosage* / Q000737:chemistry / Q000493:pharmacokinetics*; D011498:Protein Precursors / Q000008:administration & dosage* / Q000737:chemistry; D012542:Scattering, Radiation; D013055:Spectrophotometry, Infrared
chemical_list: D007328:Insulin; D010446:Peptide Fragments; D011498:Protein Precursors; C111566:imidazopropionyl(7)-arginyl(26)-N(epsilon)-octanoyl-lysyl(34)-glucaon-like peptide-1(7-37)-OH; D052216:Glucagon-Like Peptide 1; D005934:Glucagon