N‐terminal alterations turn the gut hormone GLP‐2 into an antagonist with gradual loss of GLP‐2 receptor selectivity towards more GLP‐1 receptor interaction

Abstract

Background and PurposeTo fully elucidate the regulatory role of the GLP‐2 system in the gut and the bones, potent and selective GLP‐2 receptor (GLP‐2R) antagonists are needed. Searching for antagonist activity, we performed systematic N‐terminal truncations of human GLP‐2(1‐33).Experimental ApproachCOS‐7 cells were transfected with the human GLP‐2R and assessed for cAMP accumulation or competition binding using 125I‐GLP‐2(1‐33)[M10Y]. To examine selectivity, COS‐7 cells expressing human GLP‐1 or GIP receptors were assessed for cAMP accumulation.Key ResultsAffinity of the N‐terminally truncated GLP‐2 peptides for the GLP‐2 receptor decreased with reduced N‐terminal peptide length (Ki 6.5–871 nM), while increasing antagonism appeared with inhibitory potencies (IC50) values from 79 to 204 nM for truncation up to GLP‐2(4‐33) and then declined. In contrast, truncation‐dependent increases in intrinsic activity were observed from an Emax of only 20% for GLP‐(2‐33) up to 46% for GLP‐2(6‐33) at 1 μM, followed by a decline. GLP‐2(9‐33) had the highest intrinsic efficacy (Emax 65%) and no antagonistic properties. Moreover, with truncations up to GLP‐2(8‐33), a gradual loss in selectivity for the GLP‐2 receptor appeared with increasing GLP‐1 receptor (GLP‐1R) inhibition (up to 73% at 1 μM). Lipidation of the peptides improved antagonism (IC50 down to 7.9 nM) for both the GLP‐2 and the GLP‐1R.Conclusion and ImplicationsThe N‐terminus of GLP‐2 is crucial for GLP‐2R activity and selectivity. Our observations form the basis for the development of tool compounds for further characterization of the GLP‐2 system.