Towards an alternative Treatment of Diabetes

While insulin remains the dominant treatment for diabetes, in recent years have so-called incretin-based therapies become available for the treatment of type 2 diabetes (T2D). The “incretin effect” refers to the observation that insulin secretion increases substantially in response to orally ingested glucose compared to intravenous glucose administration. Especially as T2D continues to affect ever larger parts of the world’s populations, it is highly desirable to have different regimes for treatment available. Also, to different patients, different treatments could be most suitable. Two main incretin hormones have been identified; one of these being Glucagon-Like Peptide-1 (GLP-1). The receptor for GLP-1 (GLP-1R) is the focus of this thesis.

GLP-1R is a G-protein coupled receptor (GPCR). Also known as seven-transmembrane (7TM) receptors, GPCRs constitute one of the largest families of proteins in the human genome. They have become important drug targets, which make elucidation of their molecular structure and functional domains increasingly important for designing new and better therapeutic agents.

The thesis investigates different aspects of GLP-1R interactions with GLP-1 and other receptor agonists.

Firstly, the crystal structure of GLP-1 in complex with the extracellular domain (ECD) of GLP-1R was established. The crystal structure solved in the project showed that GLP-1 is a continuous α-helix from Thr13* to Val33* when bound to the ECD, but the helix has a distortion of the backbone around Gly22*.

Secondly, a combination of crystallography and site-directed mutagenesis supported the existence of different binding modes of GLP-1 and exendin-4. The work demonstrated a ligand-supported effect of a Leu32-Ala mutation in the ECD of the full-length GLP-1R. Whether the ligand-specific effect is affected by a kink in the α-helix of GLP-1 remains to be investigated.

Thirdly, a cysteine-deprived and C-terminally truncated GLP-1R established that seven cysteine residues and more than half of the C-terminal tail are not required for GLP-1 binding or function.

Finally, real-time cAMP-measurements indicated that exendin-4 has a prolonged effect on GLP-1R compared to other peptide agonists; and receptor domains and specific residues involved in small molecule-mediated activation of GLP-1R were identified.

It is probable that these findings could help in improving the design and development of small molecule GLP-1R agonists that are suitable for oral administration. It is obvious that rational drug design, regardless of whether it is applied to peptide- or small molecule ligands, would benefit immensely from a crystal structure of the full-length GLP-1R. The results obtained with the cysteine-deprived and C-terminally truncated GLP-1R may guide the design of stable receptor constructs that can ultimately lead to structural characterization of the full-length GLP-1R.