The long-acting amylin/calcitonin receptor agonist ZP5461 suppresses food intake and body weight in male rats.

The peptide hormone amylin reduces food intake and body weight and is an attractive candidate target for novel pharmacotherapies to treat obesity. However, the short half-life of native amylin and amylin analogs like pramlintide limits these compounds' potential utility in promoting sustained negative energy balance. Here, we evaluate the ability of the novel long-acting amylin/calcitonin receptor agonist ZP5461 to reduce feeding and body weight in rats, and also test the role of calcitonin receptors (CTRs) in the dorsal vagal complex (DVC) of the hindbrain in the energy balance effects of chronic ZP5461 administration. Acute dose-response studies indicate that systemic ZP5461 (0.5-3 nmol/kg) robustly suppresses energy intake and body weight gain in chow- and high-fat diet (HFD)-fed rats. When HFD-fed rats received chronic systemic administration of ZP5461 (1-2 nmol/kg), the compound initially produced reductions in energy intake and weight gain but failed to produce sustained suppression of intake and body weight. Using virally mediated knockdown of DVC CTRs, the ability of chronic systemic ZP5461 to promote early reductions in intake and body weight gain was determined to be mediated in part by activation of DVC CTRs, implicating the DVC as a central site of action for ZP5461. Future studies should address other dosing regimens of ZP5461 to determine whether an alternative dose/frequency of administration would produce more sustained body weight suppression.

Characterization of signalling and regulation of common calcitonin receptor splice variants and polymorphisms.

The calcitonin receptor (CTR) is a class B G protein-coupled receptor that is a therapeutic target for the treatment of hypercalcaemia of malignancy, Paget's disease and osteoporosis. In primates, the CTR is subject to alternative splicing, with a unique, primate-specific splice variant being preferentially expressed in reproductive organs, lung and kidney. In addition, humans possess a common non-synonymous single-nucleotide polymorphism (SNP) encoding a proline/leucine substitution in the C-terminal tail. In low power studies, the leucine polymorphism has been associated with increased risk of osteoporosis in East Asian populations and, independently, with increased risk of kidney stone disease in a central Asian population. The CTR is pleiotropically coupled, though the relative physiological importance of these pathways is poorly understood. Using both COS-7 and HEK293 cells recombinantly expressing human CTR, we have characterized both splice variant and polymorphism dependent response to CTs from several species in key signalling pathways and competition binding assays. These data indicate that the naturally occurring changes to the intracellular face of CTR alter ligand affinity and signalling, in a pathway and agonist dependent manner. These results further support the potential for these primate-specific CTR variants to engender different physiological responses. In addition, we report that the CTR exhibits constitutive internalization, independent of splice variant and polymorphism and this profile is unaltered by peptide binding.

Effects of Roux-en-Y gastric bypass on fasting and postprandial inflammation-related parameters in obese subjects with normal glucose tolerance and in obese subjects with type 2 diabetes.

BACKGROUND: Obesity is characterized by low grade inflammation and an altered secretion of inflammatory cytokines from the adipose tissue. Weight loss has shown to reduce inflammation; however, changes in cytokine profiles during massive weight loss are not well described. The present study explored the hypothesis that Roux-en-Y gastric bypass (RYGB) reduces circulating levels of pro-inflammatory cytokines, while increasing anti-inflammatory cytokines in obese subjects with type 2 diabetes (T2D) and in obese normal glucose tolerant (NGT) subjects. METHODS: Thirteen obese subjects with T2D [weight; 129 ± 14 kg, glycated hemoglobin (HbA1c); 7.0 ± 0.9%, body mass index (BMI); 43.2 ± 5.3 kg/m(2), mean ± SD] and twelve matched obese NGT subjects [weight; 127 ± 15 kg, HbA1c; 5.5 ± 0.4%, BMI; 41.5 ± 4.8 kg/m(2), mean ± SD] were examined before, one week, three months, and one year after surgery. Interleukin (IL)-6, leptin, adiponectin, IL-8, transforming growth factor beta (TGF-ß), and the incretin hormone glucagon-like peptide-1 (GLP-1) were measured in the fasting state and during a liquid meal. Insulin resistance was evaluated by HOMA-IR. RESULTS: Weight loss did not differ between the two groups. Before surgery, HbA1c was higher and HOMA-IR lower in T2D patients, however, converged to the values of NGT subjects one year after surgery. Circulating cytokine concentrations did not differ between the two groups at any time point. One week after surgery, circulating IL-6 and IL-8 were increased, while adiponectin and leptin were reduced compared with pre-surgical concentrations. Three months after surgery, IL-8 was increased, leptin was reduced, and no change was observed for IL-6, TGF-ß, and adiponectin. One year after surgery, concentrations of IL-6, TGF-ß, and leptin were significantly reduced compared to before surgery, while adiponectin was significantly increased. CONCLUSIONS: One year after RYGB, fasting concentrations of IL-6 and leptin were reduced, while no changes were observed in IL-8. TGF-ß was decreased and adiponectin increased in both T2D and NGT obese subjects. This study is the first to examine IL-8 and TGF-ß in obese subject after RYGB. Resolution of inflammation could offer a potential explanation for the health improvement associated with major weight loss after bariatric surgery. TRIAL REGISTRATION: http://www.clinicaltrials.gov (NCT01579981).

Amylin(1-8) is devoid of anabolic activity in bone.

Amylin(1-8), a cyclic peptide consisting of the eight N-terminal amino acids of the 37-amino acid peptide amylin, has been shown to induce proliferation of primary osteoblasts and to induce bone formation in healthy male mice, whereas no data on efficacy in bone disease-related models have been reported. Therefore, we evaluated any effects of amylin(1-8) in ovariectomized rats with established osteopenia, a model for postmenopausal osteoporosis. At doses up to 100 nmol/kg/day, a dose highly effective in healthy mice, amylin(1-8) was unable to increase bone mineral density in ovariectomized rats during an 8-week treatment period. Histomorphometric analysis of the tibia indicated that amylin(1-8) did not change bone histomorphometric parameters. In an attempt to verify any potential biological effects of amylin(1-8), we investigated the efficacy of this peptide in various in vitro assays. Experiments designed to confirm previously published results on the proliferative effects of amylin(1-8) on primary osteoblasts failed to show any response. Amylin(1-8) was able to partially displace (125)I-rat amylin(1-37) from amylin receptors composed of the calcitonin receptor and RAMP1, indicating specific interaction of the peptide with the amylin binding site. However, in vitro efficacy assays with amylin(1-8) in calcitonin receptor-RAMP-positive HEK293T and MCF7 cells failed to reveal any agonist activity of amylin(1-8), whereas amylin(1-37) showed the expected agonist activity. In conclusion, our results indicate that amylin(1-8) does not show agonist activity on amylin receptors, does not affect osteoblast proliferation, and is devoid of anabolic activity in bone.