Fibroblast growth factor 21 (FGF21), a hormone with short half-life, has consistently shown strong pharmacological efficacy. We first assessed the efficacy of murine recombinant FGF21 in C57BL6 lean mice for 5 wk. We then generated a long-acting FGF21 molecule by fusing a Fc to a variant of human recombinant FGF21 (hrFGF21) that contained two engineered mutations [L98R, P171G; Fc-FGF21(RG)] and tested it in C57BL6 diet-induced obese mice and obese rhesus monkeys. We compared its metabolic properties with those of the hrFGF21. Groups of diet-induced obese mice were treated for 36 d with different doses of hrFGF21 (01, 0.3, and 1 mg/kg twice daily) and with Fc-FGF21(RG) (2.3 mg/kg, every 5 d). Body weight, glucose, insulin, cholesterol, and triglyceride levels were decreased after treatment with either compound. A glucose tolerance test (GTT) was also improved. Obese rhesus monkeys were treated with hrFGF21 (once a day) and Fc-FGF21(RG) (once a week) in a dose-escalation fashion. Doses started at 0.1 and 0.3 mg/kg and ended at 3 and 5 mg/kg for hrFGF21 and Fc-FGF21(RG), respectively. Doses were escalated every 2 wk, and animals were followed up for a washout period of 3 wk. Body weight, glucose, insulin, cholesterol, and triglyceride levels and the GTT profile were decreased to a greater extent with Fc-FGF21(RG) than with hrFGF21. The PK-PD relationship of Fc-FGF21(RG) exposure and triglyceride reduction was also conducted with a maximum response model. In conclusion, in more than one species, Fc-FGF21(RG) chronically administered once a week showed similar or greater efficacy than hrFGF21 administered daily.

Antagonizing the glucagon signaling pathway represents an attractive therapeutic approach for reducing excess hepatic glucose production in patients with type 2 diabetes. Despite extensive efforts, there is currently no human therapeutic that directly inhibits the glucagon/glucagon receptor pathway. We undertook a novel approach by generating high-affinity human monoclonal antibodies (mAbs) to the human glucagon receptor (GCGR) that display potent antagonistic activity in vitro and in vivo. A single injection of a lead antibody, mAb B, at 3 mg/kg, normalized blood glucose levels in ob/ob mice for 8 days. In addition, a single injection of mAb B dose-dependently lowered fasting blood glucose levels without inducing hypoglycemia and improved glucose tolerance in normal C57BL/6 mice. In normal cynomolgus monkeys, a single injection improved glucose tolerance while increasing glucagon and active glucagonlike peptide-1 levels. Thus, the anti-GCGR mAb could represent an effective new therapeutic for the treatment of type 2 diabetes.

Uncontrolled hepatic glucose output (HGO) contributes significantly to the pathological hyperglycemic state of patients with type 2 diabetes. Glucagon, through action on its receptor, stimulates HGO, thereby leading to increased glycemia. Antagonizing the glucagon signaling pathway represents an attractive therapeutic approach for the treatment of type 2 diabetes. We previously reported the generation and characterization of several high-affinity monoclonal antibodies (mAbs) targeting the glucagon receptor (GCGR). In the present study, we demonstrate that a 5-week treatment of diet-induced obese mice with mAb effectively normalized nonfasting blood glucose. Similar treatment also reduced fasting blood glucose without inducing hypoglycemia or other undesirable metabolic perturbations. In addition, no hypoglycemia was found in db/db mice that were treated with a combination of insulin and mAb. Long-term treatment with the mAb caused dose-dependent hyperglucagonemia and minimal to mild ␣-cell hyperplasia in lean mice. There was no evidence of pancreatic ␣-cell neoplastic transformation in mice treated with mAb for as long as 18 weeks. Treatment-induced hyperglucagonemia and ␣-cell hyperplasia were reversible after treatment withdrawal for periods of 4 and 10 weeks, respectively. It is noteworthy that pancreatic ␤-cell function was preserved, as demonstrated by improved glucose tolerance throughout the 18-week treatment period. Our studies further support the concept that long-term inhibition of GCGR signaling by a mAb could be an effective approach for controlling diabetic hyperglycemia.

A series of conformationally constrained derivatives of glucagon-like peptide-1 (GLP-1) were designed and evaluated. By use of [Gly (8)]GLP-1(7-37)-NH2 (2) peptide as a starting point, 17 cyclic derivatives possessing i to i + 4, i to i + 5, or i to i + 7 side chain to side chain lactam bridges from positions 18 to 30 were prepared. The effect of a helix-promoting alpha-amino-isobutyric acid (Aib) substitution at position 22 was also evaluated. The introduction of i to i + 4 glutamic acid-lysine lactam constraints in c[Glu (18)-Lys (22)][Gly (8)]GLP-1(7-37)-NH2 (6), c[Glu (22)-Lys (26)][Gly (8)]GLP-1(7-37)-NH2 (10), and c[Glu (23)-Lys (27)][Gly (8)]GLP-1(7-37)-NH2 (11) resulted in potent functional activity and receptor affinities comparable to native GLP-1. Selected GLP-1 peptides were chemoselectively PEGylated in order to prolong their in vivo activity. PEGylated peptides [Gly (8),Aib (22)]GLP-1(7-37)-Cys ((PEG))-Ala-NH2 (23) and c[Glu (22)-Lys (26)][Gly (8)]GLP-1(7-37)-Cys ((PEG))-Ser-Gly-NH2 (24) retained picomolar functional potency and avid receptor binding properties. Importantly, PEGylated GLP-1 peptide 23 exhibited sustained in vivo efficacy with respect to blood glucose reduction and decreased body weight for several days in nonhuman primates.

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