Activities of a non-classical estrogen, Z-bis-dehydrodoisynolic acid, with ERalpha and ERbeta

Area of Science:

• Endocrinology
• Molecular Pharmacology
• Estrogen Receptor Signaling

Background:

• (+/-)-Z-bis-Dehydrodoisynolic acid [(+/-)-Z-BDDA] exhibits high in vivo estrogenicity despite poor estrogen receptor (ER) binding.
• This discrepancy suggests potential involvement of alternative ERs or non-classical molecular mechanisms.
• Previous studies highlighted this paradox with ERalpha, necessitating investigation into ERbeta's role.

Purpose of the Study:

• To investigate the activity of individual Z-BDDA enantiomers, (+)-Z-BDDA and (-)-Z-BDDA, on ERalpha and ERbeta.
• To compare the in vitro and cell-based activities of these enantiomers with their known in vivo effects.
• To determine if ERbeta can resolve the activity-binding paradox observed with Z-BDDA.

Main Methods:

• Transfectional analysis in Hela cells to assess gene activation and repression mediated by ERalpha and ERbeta.
• In vitro binding assays (competition and direct binding) to quantify the relative affinities of Z-BDDA enantiomers for ERalpha and ERbeta.
• Comparison of enantiomer activities and affinities against 17beta-estradiol (E2) as a reference.

Main Results:

• (-)-Z-BDDA demonstrated significant agonist activity for gene activation and repression with both ERalpha (EC50 ≈ 0.3nM and 0.2nM, respectively) and ERbeta (EC50 ≈ 5nM and 0.2nM, respectively).
• (+)-Z-BDDA exhibited minimal to no activity in all cell-based assays.
• In vitro binding assays revealed that (-)-Z-BDDA had approximately 1.7-7% relative affinity for ERalpha and ERbeta compared to E2, while (+)-Z-BDDA had significantly lower affinities (0.08-0.3%).

Conclusions:

• The potent in vivo activity of Z-BDDA is primarily attributed to the (-)-enantiomer, which acts as an agonist for both ERalpha and ERbeta.
• The observed activity-binding paradox, characterized by high in vivo potency and low in vitro receptor affinity, is now associated with both ERalpha and ERbeta.
• ERbeta does not resolve the activity-binding paradox, indicating that additional in vivo metabolic, pharmacokinetic studies, or alternative mechanisms are required to fully explain Z-BDDA's potent estrogenic effects.