3-PEP promotes bone regeneration by up regulating BCL-2 expression via ERK phosphorylation

Bone healing and regeneration is a complex process that recapitulates embryonic skeletal development and is delayed in diseases like osteoporosis. Bone healing therapies like recombinant bone morphogenetic-2 protein (rhBMP-2) and parathyroid hormone (PTH), an approved bone anabolic therapy reduces fracture risks but are fraught with high cost and several side effects. Thus, there is an unmet need for cost-effective bone healing agents. In this study, we have synthesized 3-piperidinylethoxypterocarpan (3-PEP) which is a hybrid of bone supplement ipriflavone and anti-resorptive drug raloxifene and evaluated its bone regeneration and healing potential. Prior to studies in animal models, the potency of 3-PEP was confirmed in calvarial osteoblast cells. Bromodeoxy uridine cell proliferation and cell viability assay revealed that 3-PEP at 100 pM concentration increased the proliferation and survival of osteoblasts simultaneously inhibiting the apoptosis by involving activation of BCL-2 by phosphorylation at Ser70 site through MEK-ERK pathway. In vivo studies were conducted in estrogen-deficient ovariectomized Balb/c mice and drill hole injury was generated in the mid diaphysis of the femur in all the animals. Treatment with 3-PEP commenced the next day onward and terminated at 7 and 15 days. Micro-CT analysis and calcein labeling of newly generated bone at the drill hole injury site showed that 3-PEP promotes bone healing and new bone formation at a dose of 5 mg/kg at the injury site. These data were also corroborated in non-ovariectomized Balb/c mice cortical defect model. Owing to the side effects associated with rhBMP-2 and PTH, along with the expenses involved, our study proposes an alternative therapeutic option for bone healing.