To Bmp2 As A Target For New Anti-osteoporosis Drug Screening Model Construction And Applied Research

Bone Morphogenetic Protein II (BMP2), a member of the TGF-βsuper-family, is highly expressed in osteoblasts and is a crucial regulator of osteogenic differentiation which has been demonstrated both in vitro and in vivo. Many observations clearly indicate the high potency of BMP2 as an inducer of osteogenesis and it may be a novel therapeutic target for diseases associated with bone loss, especially in menopausal and postmenopausal women.In order to discover new agents that enhance the expression of the mouse BMP2, a high-throughput assay was developed to screen a synthetic and natural compound library. The cell-based high-throughput screen was conducted in 96 well plates by using the clonal murine calvarial MC3T3-E1 cells. These cells were stably transfected with mouse bmp2 promoter-luciferase reporter vector and calibrated with the known anti-osteoporosis compound Genistein. Among 3192 compounds screened, 7 positive agents were identified as up-regulators of BMP2 expression, and Z' factor of 40 assays is in accordance with the standard of HTS and ensuring the credibility of the model.Activity of 3 positive compounds, Daidzein, Formononetin and 2-Acetyldibenzothiophene (2-ABT), on BMP2 expression were confirmed by real-time RT-PCR and flow cytometry in MG63 cells. MTT assay of 2-ABT showed its hypotoxicity in vitro and AKP assay showed its positive effect on bone formation. 76 compounds were designed and synthesized on the base of 2-ABT, their BMP2 up-regulatory activity were detected in our model.Thus, it is demonstrated that this screening model is useful for discovering lead compounds to treat osteoporosis and maintain the body bone metabolism balance.