It has been hypothesized that activation of peroxisome-proliferator-activated receptor-gamma (PPAR gamma) by thiazolidinedione drugs can increase adipogenesis at the expense of osteogenesis, leading to bone loss. However, the reported skeletal effects of these compounds are varied and their effects on cortical bone are unknown. In this study, we examined the changes in both cancellous and cortical bone of 6-month-old male mice treated with darglitazone, a potent and selective PPAR gamma agonist, at 10 mg/kg/day by dosing the compound in a food mixture for 2 or 8 weeks. At 2 weeks, we observed significantly increased marrow adipose tissue area, decreased trabecular bone density of distal femur, and decreased surface referent bone formation rate of lumbar vertebrae in the mice treated with darglitazone compared with controls. At 8 weeks, lower cancellous bone mass was seen at both distal femurs and lumbar vertebrae of the mice treated with darglitazone. In addition, mineralizing surface was significantly lower, whereas osteoclast surface and number were significantly higher in the lumbar vertebrae of darglitazone-treated mice. At the femoral diaphysis, darglitazone treatment caused bone loss on the endocortical surface. Interestingly, periosteal mineral apposition rate and surface referent bone formation rate were significantly increased in darglitazone-treated mice. In bone marrow cell cultures, darglitazone suppressed alkaline phosphatase activity, ostcoblastic gene expression, and mineralized nodule formation while increasing adipogenic gene expression and lipid accumulation. In summary, darglitazone enhanced adipogenesis and caused cancellous bone loss by increasing bone resorption and decreasing bone formation in mice. In addition, darglitazone induced cortical bone loss on the endocortical surface but increased bone formation on the periosteal surface. These data suggest that PPAR gamma plays a role in regulating bone resorption and formation and reveal surface-specific effects of a PPAR gamma agonist on bone. (c) 2006 Elsevier Inc. All rights reserved.

CP432 is a newly discovered, nonprostanoid EP4 receptor selective prostaglandin E-2 agonist. CP432 stimulates trabecular and cortical bone formation and restores bone mass and bone strength in aged ovariectomized rats with established osteopenia.Introduction: The purpose of this Study was to determine whether a newly discovered, nonprostanoid EP4 receptor selective prostaglandin E-2 (PGE(2)) agonist, CP432, could produce bone anabolic effects in aged, ovariectomized (OVX) rats with established osteopenia.Materials and Methods: CP432 at 0.3, 1, or 3 mg/kg/day was given for 6 weeks by subcutaneous injection to 12-month-old rats that had been OVX for 8.5 months. The effects on bone mass, bone formation, bone resorption, and bone strength were determined.Results: Total femoral BMD increased significantly in OVX rats treated with CP432 at all doses. CP432 completely restored trabecular bone volume of the third lumbar vertebral body accompanied with a dose-dependent decrease in osteoclast number and osteoclast surface and a dose-dependent increase in mineralizing surface, mineral apposition rate, and bone formation rate-tissue reference in OVX rats. CP432 at 1 and 3 mg/kg/day significantly increased total tissue area, cortical bone area, and periosteal and endocortical bone formation in the tibial shafts compared with both sham and OVX controls. CP432 at all doses significantly and dose-dependently increased ultimate strength in the fifth lumber vertebral body compared with both sham and OVX controls. At 1 and 3 mg/kg/day, CP432 significantly increased maximal load in a three-point bending test of femoral shaft compared with both sham and OVX controls.Conclusions: CP432 completely restored trabecular and cortical bone mass and strength in established osteopenic, aged OVX rats by stimulating bone formation and inhibiting bone resorption on trabecular and cortical surfaces.

The EP4 receptor, one of the subtypes of the prostaglandin E2 (PGE2) receptor, plays a critical role in the anabolic effects of PGE2 on bone. However, its role in the maintenance of bone mass in aged animals and its role in fracture healing is not well known. Our studies addressed these issues by characterizing the skeletal phenotype of aged, EP4 receptor knockout (KO) mice, and by comparing fracture healing in aged KO mice versus wild type (WT) mice. There was no significant difference in body weight and femoral length between KO and WT mice at 15 to 16 months of age. Lower bone mass was seen radiographically in both axial and long bones of KO mice relative to WT mice. Micro-CT images of the distal femurs showed thinner cortices, fewer trabeculae, and a deteriorated trabecular network in KO mice. Total bone content, trabecular content, and cortical content, as assessed by pQCT in the distal femur, were lower in KO mice than WT controls. Histomorphometric measurements showed that trabecular bone volume and bone formation rate were significantly decreased whereas osteoclast number on trabecular surface and eroded surface on endocortical surface were significantly increased in KO mice. These data indicated that deleting the EP4 receptor resulted in an imbalance in bone resorption over formation, leading to a negative bone balance. The lower bone formation rate in EP4 KO mice was primarily due to decreased mineralizing surface, suggesting that the defect in overall bone formation was mainly due to the defect in osteoblastogenesis. Fracture healing was examined in KO and WT mice subjected to a transverse femoral fracture. Callus formation was significantly delayed as evidenced both radiographically and histologically in the fractured femurs of KO mice compared with those of WT mice. KO mice had significant decreases in total callus area, cartilaginous callus area, and bony callus area 2 weeks after fracture. By 4 weeks, complete bony bridging was seen in WT mice but not in KO mice. These data demonstrate that the absence of the EP4 receptor decreases bone mass and impairs fracture healing in aged male mice. Our findings indicate that the EP4 receptor is a positive regulator in the maintenance of bone mass and fracture healing.

Novel pyrazolinone-piperidine dipeptide derivatives were synthesized and evaluated as growth hormone secretagogues (GHSs). Two analogues, capromorelin (5, CP-424391-18, hGHS-R1a K(i)=7 nM, rat pituicyte EC(50)=3 nM) and the des-methyl analogue 5c (hGHS-R1a K(i)=17 nM, rat pituicyte EC(50)=3 nM), increased plasma GH levels in an anesthesized rat model, with ED(50) values less than 0.05 mg/kg iv. Capromorelin showed enhanced intestinal absorption in rodent models and exhibited superior pharmacokinetic properties, including high bioavailabilities in two animal species [F(rat)=65%, F(dog)=44%]. This short-duration GHS was orally active in canine models and was selected as a development candidate for the treatment of musculoskeletal frailty in elderly adults.

New tert-butyl, picolyl and fluorinated analogues of capromorelin (3), a short-acting growth hormone secretagogue (GHS), were prepared as part of a program to identify long-acting GHSs that increase 24-h plasma IGF-1 levels. Compounds 4c and 4d (ACD LogD values ≥2.9) displayed extended plasma elimination half-lives in dogs, primarily due to high volumes of distribution, but showed weak GH secretagogue activities in rats (ED50s>10 mg/kg). A less lipophilic derivative 4 (ACD LogD=1.6) exhibited a shorter canine half-life, but stimulated GH secretion in two animal species. Repeat oral dosing of 4 in dogs for 29 days (6 mg/kg) resulted in a significant down-regulation of the post dose GH response and a 60 and 40% increase in IGF-1 levels relative to pre-dose levels at the 8- and 24-h post dose time points. Compound 4 (CP-464709-18) has been selected as a development candidate for the treatment of frailty.

{abs}Background —Exogenous administration of growth hormone (GH) and subsequently increased production of insulin-like growth factor-1 can influence left ventricular (LV) myocardial growth and geometry in the setting of congestive heart failure (CHF). This study determined the effects of an orally active GH secretagogue (GHS) treatment that causes a release of endogenous GH on LV function and myocyte contractility in a model of developing CHF. Methods and Results —Pigs were randomly assigned to the following treatment groups: (1) chronic rapid pacing at 240 bpm for 3 weeks (n=11); (2) chronic rapid pacing and GHS (CP-424,391 at 10 mg·kg −1 ·d −1 , n=9); and (3) sham controls (n=8). In the untreated pacing CHF group, LV fractional shortening was reduced (21±2% versus 47±2%) and peak wall stress increased (364±21 versus 141±5 g/cm 2 ) from normal control values ( P <0.05). In the GHS group, LV fractional shortening was higher (29±2%) and LV peak wall stress lower (187±126 g/cm 2 ) than untreated CHF values ( P <0.05). With GHS treatment, the ratio of LV mass to body weight increased by 44% from untreated values. Steady-state myocyte velocity of shortening was reduced with pacing CHF compared with controls (38±1 versus 78±1 μm/s, P <0.05) and was increased from pacing CHF values with GHS treatment (55±7 μm/s, P <0.05). Conclusions —The improved LV pump function that occurred with GHS treatment in this model of CHF was most likely a result of favorable effects on LV myocardial remodeling and contractile processes. On the basis of these results, further studies are warranted to determine the potential role of GH secretagogues in the treatment of CHF.