The use of biopharmaceuticals in the treatment of diseases such as diabetes, cancer, and hemophilia has increased dramatically over the past decades. Despite their many advantages such as high potency, specificity, and low toxicity, many biopharmaceuticals suffer from inherent chemical and physical instabilities and short plasma half-lives, which make their formulation development and delivery challenging.
Lipidation is a successful strategy for extending the half-lives of peptide drugs through lipidation-induced self-association and association to albumin. Though albumin-association is exploited by several approved lipidated peptide drugs, structural knowledge about the albumin-peptide complexes and their interactions on the atomic level is limited. This thesis treat selfassociation and albumin-association of two lipidated insulin analogues, insulin detemir and insulin degludec, through an interdisciplinary approach using small angle X-ray scattering (SAXS) and molecular dynamics (MD) simulations.
The first solution structures of a detemir trihexamer, and albumin–insulin analogue complexes in 1:6, 1:12, and 2:12 stoichiometries based on SAXS data were modeled, and equilibria for albumin-detemir and albumin-degludec mixtures were proposed. The albumin-detemir hexamer solution structure shows four possible detemir binding sites. These binding sites were investigated by MD simulations and molecular mechanics Poisson-Boltzmann surface area free energy calculations. The overlapping FA3-FA4 binding site on albumin was found to be the most favorable detemir binding site.
The presence of albumin was found to enhance detemir’s stability against freeze-thaw and agitation stresses almost independently on complex formation, suggesting that albumin-detemir complex formation does not lead to further stabilization.
Schematic representations of the sequences of native insulin, deglutec and detemir.