• Piskopou, Anastasia
• Fili, Stavroula
• Margiolaki, Irene
• Reinle-Schmitt, Mathilde
• Christopoulou, Magda
• Beckers, Detlef
• Morin, Mickael
• Valmas, Alexandros
• Spiliopoulou, Maria
• Degen, Thomas
• Papadea, Polyxeni
• Rosmaraki, Eleftheria
• Chasapis, Christos T.
• Triandafillidis, Dimitris P.
• Karavassili, Fotini
• Filopoulou, Aikaterini
• Gozzo, Fabia

Abstract

<jats:p>Regular injections of insulin provide life-saving benefits to millions of diabetics. Apart from native insulin and insulin analogue formulations, microcrystalline insulin suspensions are also commercially available. The onset of action of the currently available basal insulins relies on the slow dissociation of insulin hexamers in the subcutaneous space due to the strong binding of small organic ligands. With the aim of identifying insulin–ligand complexes with enhanced pharmacokinetic and pharmacodynamic profiles, the binding affinity of two resorcinol-based molecules (4-chlororesorcinol and 4-bromoresorcinol) and the structural characteristics of insulin upon co-crystallization with them were investigated in the present study. `In solution' measurements were performed <jats:italic>via</jats:italic> saturation transfer difference (STD) NMR. Co-crystallization upon pH variation resulted in the production of polycrystalline precipitates, whose structural characteristics (<jats:italic>i.e.</jats:italic> unit-cell symmetry and dimension) were assessed. In both cases, different polymorphs (four and three, respectively) of monoclinic symmetry (<jats:italic>P</jats:italic>2<jats:sub>1</jats:sub> and <jats:italic>C</jats:italic>2 space groups) were identified via X-ray powder diffraction. The results demonstrate the efficiency of a new approach that combines spectroscopy and diffraction techniques and provides an innovative alternative for high-throughput examination of insulin and other therapeutic proteins.</jats:p>

Topics

• precipitate
• Nuclear Magnetic Resonance spectroscopy
• crystallization
• space group