Cloning, stable expression of human phosphodiesterase 7A and development of an assay for screening of PDE7 selective inhibitors.
Phosphodiesterases (PDEs) constitute a superfamily of enzymes that plays an important role in signal transduction by catalysing the hydrolysis of cAMP and cGMP. cDNA encoding PDE7A1 subtype was cloned and a stable recombinant HEK 293 cell line expressing high levels of PDE7A1 was generated. Transient transfection of pCRE-Luc plasmid, harboring luciferase reporter gene into the stable recombinant cell line and subsequent treatment with PDE7 inhibitor, resulted in a dose-dependent increase in luciferase activity. This method provides a simple and sensitive cell-based assay for screening of PDE7 selective inhibitors for the treatment of T cell mediated diseases.
Malik
Renu
R
Department of Biotechnology, Ranbaxy Research Laboratories, Plot No. 20, Sector 18, Udyog Vihar Industrial Area, Gurgaon, 122001, Haryana, India.
Bora
Roop Singh
RS
Gupta
Dikshi
D
Sharma
Pratibha
P
Arya
Ranjana
R
Chaudhary
Shiwani
S
Saini
Kulvinder Singh
KS
D002460:Cell Line; D003001:Cloning, Molecular; D054708:Cyclic Nucleotide Phosphodiesterases, Type 7 / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D004353:Drug Evaluation, Preclinical / Q000379:methods; D015870:Gene Expression; D017930:Genes, Reporter; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology*
D010726:Phosphodiesterase Inhibitors; D008156:Luciferases; D054708:Cyclic Nucleotide Phosphodiesterases, Type 7; C517458:PDE7A protein, human