EFFECTS OF NONCOVALENT SELF-ASSOCIATION ON THE SUBCUTANEOUS ABSORPTION OF A THERAPEUTIC PEPTIDE

Purpose. To utilize an acylated peptide as a model system to investiga te the relationships among solution peptide conformation, noncovalent self-association, subcutaneous absorption and bioavailability under ph armaceutically relevant solution formulation conditions. Methods, CD s pectroscopy, FTIR spectroscopy, equilibrium sedimentation, dynamic lig ht scattering, and size exclusion chromatography were employed to char acterize the effects of octanoylation on conformation and self-associa tion of the 31 amino acid peptide derivative des-amino-histidine(7) ar ginine(26) human glucagon-like peptide (7-37)-OH (IP(7)R(26)GLP-1). Hy perglycemic clamp studies were performed to compare the bioavailabilit y, pharmacokinetics, and pharmacodynamics of solution formulations of oct-IP(7)R(26)GLP-1 administered subcutaneously to normal dogs. Result s. Octanoylation of IP(7)R(26)GLP-1 was shown to confer the propensity for a major solvent-induced conformational transition with an accompa nying solvent-and temperature-dependent self-association behavior. For mulations were characterized that give rise to remarkably different ph armacodynamics and pharmacokinetics that correlate with distinct pepti de conformational and self-association states. These states correspond to: (i) a minimally associated alpha-helical form (apparent molecular weight = IJ kDa), (ii) a highly associated, predominantly beta-sheet form (effective molecular diameter 20 nm), and (iii) an unusually larg e, micelle-like soluble beta-sheet aggregate (effective molecular diam eter 50 nm). Conclusions, Bioavailability and pharmacokinetics of a se lf-associating peptide can be influenced by aggregate size and the eas e of disruption of the non-covalent intermolecular interactions at the subcutaneous site. Hydrophobic aggregation mediated by seemingly inno cuous solution formulation conditions can have a dramatic effect on th e subcutaneous bioavailability and pharmacokinetics of a therapeutic p eptide and in the extreme, can totally preclude its absorption. A size exclusion chromato graphic method is identified that distinguishes su bcutaneously bioavailable aggregated oct-IP(7)R(26)GLP-1 from non-bioa vailable aggregated oct-IP(7)R(26)GLP-1.