Engineering a novel protease-based Exendin-4 derivative for type 2 antidiabetic therapeutics.

To develop an effective long-acting antidiabetic agent, we designed a novel Exendin-4 derivative (termed LEx4) containing an albumin-binding domain (ABD), a protease-cleavable linker and a native Exendin-4. Here, we present the LEx4 with balanced glucoregulatory activity and prolonged in vivo activity. As a first step, the LEx4 with purity more than 99% was prepared. Microscale thermophoresis (MST) results demonstrated that LEx4 associates with rat and monkey serum albumin with high-affinity (Ka = 1.26 × 106 M−1 and 1.52 × 106 M−1, respectively). Then the stability test in vitro showed the enhanced antiproteolytic ability of LEx4 in rat and human plasma compared to native Exendin-4. Oral glucose tolerance test (OGTT) in type 2 diabetic mice showed the glucose-lowering efficacy of LEx4 was clearly dosage-dependent within 25–250 nmol/kg. In addition, the protracted antidiabetic effects of LEx4 were further confirmed by both multiple OGTTs and hypoglycemic efficacies test in type 2 diabetic mice. In Sprague Dawley (SD) rats, LEx4 also showed 3.3-fold longer elimination half-life (t1/2) than native Exendin-4. Furthermore, once daily injection of LEx4 to db/db mice achieved long-term beneficial effects on body weight, blood biochemical values, glucose tolerance and pancreatic tissue. We believe LEx4 has superior pharmaceutical potential as a therapeutic drug to against type-2 diabetes mellitus (T2DM) based on these results. This strategy of albumin binding is also applicable to other bioactive peptides for development of long-acting therapeutic drugs.