Characterisation and biological activity of Glu3 amino acid substituted GIP receptor antagonists

Glucose-dependent insulinotropic polypeptide (GIP) is an important gastrointestinal hormone, which regulates insulin release and glucose homeostasis, but is rapidly inactivated by enzymatic N-terminal truncation. Here we report the enzyme resistance and biological activity of several Glu3-substituted analogues of GIP namely; (Ala3)GIP, (Lys3)GIP, (Phe3)GIP, (Trp3)GIP and (Tyr3)GIP. Only (Lys3)GIP demonstrated moderately enhanced resistance to DPP-IV (p < 0.05 to p < 0.01) compared to native GIP. All analogues demonstrated a decreased potency in cAMP production (EC50 1.47 to 11.02 nM; p < 0.01 to p < 0.001) with (Lys3)GIP and (Phe3)GIP significantly inhibiting GIP-stimulated cAMP production (p < 0.05). In BRIN-BD11 cells, (Lys3)GIP, (Phe3)GIP, (Trp3)GIP and (Tyr3)GIP did not stimulate insulin secretion with both (Lys3)GIP and (Phe3)GIP significantly inhibiting GIP-stimulated insulin secretion (p < 0.05). Injection of each GIP analogue together with glucose in ob/ob mice significantly increased the glycaemic excursion compared to control (p < 0.05 to p < 0.001). This was associated with lack of significant insulin responses. (Ala3)GIP, (Phe3)GIP and (Tyr3)GIP, when administered together with GIP, significantly reduced plasma insulin (p < 0.05 to p < 0.01) and impaired the glucose-lowering ability (p < 0.05 to p < 0.01) of the native peptide. The DPP-IV resistance and GIP antagonism observed were similar but less pronounced than (Pro3)GIP. These data demonstrate that position 3 amino acid substitution of GIP with (Ala3), (Phe3), (Tyr3) or (Pro3) provides a new class of functional GIP receptor antagonists.