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Published on: February 21, 2014
Evidence for estrogenic contamination of the MAPK inhibitor PD98059
1Department of Obstetrics and Gynecology, Indiana University School of Medicine, Indianapolis, IN 46202, USA.
Abstract:
PD98059 blocks phosphorylation and activation of MAPK proteins, ERK1 and ERK2. In the course of examining the effect of PD98059 on estrogen-induced transcription of reporter genes in a human breast cancer cell line and in yeast, we found that two of four different batches of PD98059 produced estrogenic effects in a dose-dependent manner. In a competitive binding assay, these preparations of PD98059 displaced radiolabeled estradiol from ER alpha. Furthermore, in the yeast assay, addition of a coactivator protein, AIB1, enhanced the transcriptional effect of PD98059, indicating that it induces receptor-coactivator interactions. Although concentrations of PD98059 required to activate ER alpha in these experimental systems are 10(4)- to 10(5) higher than the concentration of estradiol required to do the same, the concentrations required to block MAPK activation are well above those which would produce maximal estrogenic effects. Thus, when PD98059 is used in estrogen-responsive cells, contaminating estrogenic activity may confound interpretation of experimental results.
Insights
PD98059, a MAPK inhibitor, exhibits unexpected estrogenic activity. This contamination can interfere with experiments involving estrogen-responsive cells and estrogen receptors.
Area of Science:
- Molecular Biology
- Endocrinology
- Cell Biology
Background:
- PD98059 is a known inhibitor of mitogen-activated protein kinase (MAPK) pathway proteins, ERK1 and ERK2.
- Estrogen signaling is crucial in various biological processes, including breast cancer development.
Purpose of the Study:
- To investigate the potential estrogenic effects of PD98059.
- To determine if PD98059 interacts with estrogen receptors (ERs) and affects estrogen-induced transcription.
Main Methods:
- Reporter gene assays in a human breast cancer cell line and yeast.
- Competitive binding assays using radiolabeled estradiol and ER alpha.
- Assessment of coactivator protein (AIB1) effects on PD98059-mediated transcription.
Main Results:
- Two of four PD98059 batches displayed dose-dependent estrogenic effects.
- PD98059 preparations displaced estradiol from ER alpha in binding assays.
- AIB1 enhanced the estrogenic transcriptional activity of PD98059 in yeast.
Conclusions:
- Contaminating estrogenic activity in PD98059 can confound experimental results in estrogen-responsive systems.
- The effective concentrations for estrogenic effects are lower than those required for MAPK inhibition.
- Researchers should be cautious when using PD98059 in studies involving estrogen signaling.
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