Biological Activities of Glucagon-Like Peptide-1 Analogues in Vitro and in Vivo

Studies support a role for glucagon-like peptide 1 (GLP-1) as a potential treatment for diabetes. However, since GLP-1 is rapidly degraded in the circulation by cleavage at Ala2, its clinical application is limited. Hence, understanding the structure−activity of GLP-1 may lead to the development of more stable and potent analogues. In this study, we investigated GLP-1 analogues including those with N-, C-, and midchain modifications and a series of secretin-class chimeric peptides. Peptides were analyzed in CHO cells expressing the hGLP-1 receptor (R7 cells), and in vivo oral glucose tolerance tests (OGTTs) were performed after injection of the peptides in normal and diabetic (db/db) mice. [d-Ala2]GLP-1 and [Gly2]GLP-1 showed normal or relatively lower receptor binding and cAMP activation but exerted markedly enhanced abilities to reduce the glycemic response to an OGTT in vivo. Improved biological effectiveness of [d-Ala2]GLP-1 was also observed in diabetic db/db mice. Similarly, improved biological activity of acetyl- and hexenoic-His1-GLP-1, glucagon( 1 - 5 )-, glucagon( 1 - 10 )-, PACAP( 1 - 5 )-, VIP( 1 - 5 )-, and secretin( 1 - 10 )-GLP-1 was observed, despite normal or lower receptor binding and activation in vitro. [Ala8/11/12/16] substitutions also increased biological activity in vivo over wtGLP-1, while C-terminal truncation of 4−12 amino acids abolished receptor binding and biological activity. All other modified peptides examined showed normal or decreased activity in vitro and in vivo. These results indicate that specific N- and midchain modifications to GLP-1 can increase its potency in vivo. Specifically, linkage of acyl-chains to the α-amino group of His1 and replacement of Ala2 result in significantly increased biological effects of GLP-1 in vivo, likely due to decreased degradation rather than enhanced receptor interactions. Replacement of certain residues in the midchain of GLP-1 also augment biological activity.