Identification of inhibitors for MDM2 ubiquitin ligase activity from natural product extracts by a novel high-throughput electrochemiluminescent screen.
High-throughput screening technologies have revolutionized the manner in which potential therapeutics are identified. Although they are the source of lead compounds for ~65% of anticancer and antimicrobial drugs approved by the Food and Drug Administration between 1981 and 2002, natural products have largely been excluded from modern screening programs. This is due, at least in part, to the inherent difficulties in testing complex extract mixtures, which often contain nuisance compounds, in modern bioassay systems. In this article, the authors present a novel electrochemiluminescent assay system for inhibition of MDM2 activity that is suitable for testing natural product extracts in high-throughput screening systems. The assay was used to screen more than 144,000 natural product extracts. The authors identified 1 natural product, sempervirine, that inhibited MDM2 auto-ubiquitination, MDM2-mediated p53 degradation, and led to accumulation of p53 in cells. Sempervirine preferentially induced apoptosis in transformed cells expressing wild-type p53, suggesting that it could be a potential lead for anticancer therapeutics.
Sasiela
Christy A
CA
Molecular Targets Development Program, National Cancer Institute, Frederick, Maryland 21702, USA.
Stewart
David H
DH
Kitagaki
Jirouta
J
Safiran
Yassamin J
YJ
Yang
Yili
Y
Weissman
Allan M
AM
Oberoi
Pankaj
P
Davydov
Ilia V
IV
Goncharova
Ekaterina
E
Beutler
John A
JA
McMahon
James B
JB
O'Keefe
Barry R
BR
D000818:Animals; D001681:Biological Assay; D001688:Biological Products / Q000494:pharmacology*; D053148:Caspase 3 / Q000378:metabolism; D016923:Cell Death; D002461:Cell Line, Transformed; D045424:Complex Mixtures / Q000494:pharmacology*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D008163:Luminescent Measurements / Q000379:methods*; D051379:Mice; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D051736:Proto-Oncogene Proteins c-mdm2 / Q000037:antagonists & inhibitors*; D046948:Secologanin Tryptamine Alkaloids / Q000737:chemistry; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D054875:Ubiquitination / Q000187:drug effects
D001688:Biological Products; D045424:Complex Mixtures; D046948:Secologanin Tryptamine Alkaloids; D016159:Tumor Suppressor Protein p53; D051736:Proto-Oncogene Proteins c-mdm2; D011065:Poly(ADP-ribose) Polymerases; D053148:Caspase 3; D046988:Proteasome Endopeptidase Complex; C009234:sempervirine