Development of a High-Throughput Screening Strategy for Upregulators of the OPG/RANKL Ratio with the Potential for Antiosteoporosis Effects.
The ratio between osteoprotegerin (OPG) and the receptor activator of NF-κB ligand (RANKL) in the bone microenvironment indicates the level of osteoclastogenesis, and upregulation of this ratio would improve osteoporosis. In this study, we established a novel high-throughput screening (HTS) system using two stably transfected monoclonal cell lines that either express firefly luciferase under the OPG promoter control or concurrently express firefly and renilla luciferases under control of the OPG and RANKL promoters, respectively. With this system, we can conveniently and rapidly detect the effects of compounds on the expression of OPG and RANKL through changes in firefly and renilla luciferase activities. A total of 8160 compounds were screened using this system, yielding five compounds without previously reported activity. The compound with greatest potential is E05657 with high activity and low effective concentration in the HTS system. It increases the OPG/RANKL ratio and OPG secretion, decreases the NFATc1 expression, and reduces osteoclastogenesis in vitro. These results indicate that this novel HTS system can be used to identify small molecules with potential antiosteoporosis effects, and E05657 is a promising lead compound as a novel antiosteoporosis drug.
Institute of Medicinal Biotechnology, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing, China.
Shiqiang
S
Gong
Institute of Medicinal Biotechnology, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing, China.
Xiaowan
X
Han
Institute of Medicinal Biotechnology, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing, China.
Xuehong
X
Li
Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and School of Basic Medicine, Peking Union Medical College, Beijing, China.
Jun
J
Yang
Institute of Medicinal Biotechnology, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing, China.
Xiaobo
X
He
Institute of Medicinal Biotechnology, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing, China sisyimb@hotmail.com.
Shuyi
S
Si
D002460:Cell Line; D060833:Cellular Microenvironment / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D049409:Luciferases, Firefly / Q000235:genetics; D050778:NFATC Transcription Factors / Q000235:genetics; D010024:Osteoporosis / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D053244:Osteoprotegerin / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D053245:RANK Ligand / Q000235:genetics* / Q000378:metabolism; D054852:Small Molecule Libraries / Q000737:chemistry / Q000494:pharmacology
D050778:NFATC Transcription Factors; C494003:NFATC1 protein, human; D053244:Osteoprotegerin; D053245:RANK Ligand; D054852:Small Molecule Libraries; C505688:TNFSF11 protein, human; D049409:Luciferases, Firefly
high-throughput screening (HTS); osteoporosis; osteoprotegerin (OPG); receptor activator of NF-κB ligand (RANKL)