A reporter gene assay for screening of PDE4 subtype selective inhibitors
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- 作者：Roop Singh Bora ; Renu Malik ; Ranjana Arya ; Dikshi Gupta ; Vibhuti Singh ; Neeraj Aggarwal ; Sun ; a Dastidar ; Abhijit Ray ; Kulvinder Singh Saini
- 关键词：Phosphodiesterase ; Stable cell lines ; Cellular localization ; Reporter gene assay
- 刊名：Biochemical and Biophysical Research Communications
- 出版年：2007
- 出版时间：27 April 2007
- 年：2007
- 卷：356
- 期：1
- 页码：153-158
- 全文大小：986 K
文摘
Phosphodiesterase (PDE) constitutes a superfamily of enzymes that catalyze the hydrolysis of cAMP and cGMP into their corresponding monophosphates and play an important role in diverse physiological functions. The present study provides a process for identifying PDE4 subtypes selective inhibitors using a reporter gene assay. Stable recombinant HEK-293 cell lines expressing high levels of PDE4A4B, PDE4B2A, and PDE4D3 subtypes individually were generated. Transient transfection of pCRE-Luc plasmid, harboring luciferase reporter gene under the control of cAMP response element (CRE)-binding sequence, into these stable recombinant cell lines followed by treatment with PDE4 inhibitor, resulted in a dose dependent increase in luciferase activity. This methods provide a novel, simple and sensitive assay for high throughput screening of PDE4 subtype selective inhibitors for treatment of asthma and COPD.