K. L. Iresha Sampathi Perera’s research while affiliated with Marquette University and other places

Activation of estrogen receptor-beta (ERβ) has potential benefits for vasomotor symptoms as well as memory consolidation in menopausal women. Ten 4-(4-substituted cyclohexyl)phenols were synthesized and evaluated as ERβ agonists in a TR-FRET ligand displacement assay. Four of the more potent compounds, all of which contained fluoroalkyl groups as the 4-substituent, were further assessed for agonist and antagonist activity in cell-based functional assays for ERβ and ERα. The 4-(4-fluoroalkyl-cyclohexyl)phenols were all >50-fold selective for ERβ over ERα, and none were found to be antagonists for either ERβ or ERα at 7,250 nM. Detection of 200 μM of trans-4-(4-(fluoromethyl)cyclohexyl)phenol in the presence of mouse brain homogenate was possible by 19F NMR spectroscopy.

Estrogens in the brain are critical in the protection of neural pathways, and in the enhancement of hippocampal memory consolidation. The estrogen receptor exists in two forms: alpha (ERα) and beta (ERβ), the latter of which is predominant in the hippocampus and a drug target for several cognitive disorders. Thus, we have developed a selective ERβ agonist to combat memory decline in post‐menopausal women affected by significantly lowered estrogen levels. The molecule reported is an A‐C estrogen that shows high potency and selectivity for ERβ. ERα agonist activity has been linked to increased risk of diseases such as breast cancer and stroke, among others. Selectivity for ERβ over ERα is therefore critical in the success of a drug molecule that acts like estrogen. TR‐FRET displacement assays were utilized to determine the most promising candidates from a library of lead molecules by detecting binding of the molecules to the ERβ ligand binding domain. ISP358‐2 was one of few potent molecules that displayed an EC 50 < 30 nM, and also showed a 12‐fold selectivity of ERβ (EC 50 = 24 ± 5 nM) over ERα (EC 50 = 289 ± 92 nM). ERβ cell‐based assays further confirmed the molecule's potency, and ERα cell‐based potency showed ERβ selectivity. To verify our cell‐based assay results, ISP358‐2 was tested for selectivity in seven other nuclear hormone receptors in cell‐based assays. We saw no significant activity in any nuclear hormone receptor other than the estrogen receptor. We hypothesized that the selectivity observed was due to selective coactivator recruitment present only in the native full length ERβ. This hypothesis was supported by data obtained from the cell‐based transcriptional activation assay, which employs a full length and native estrogen receptor and most closely mimic the in vivo situation. The results obtained in this assay were profound in that ISP358‐2 showed an ERβ agonist potency of 31 ± 7 nm, and an ERα agonist potency of 20,400 ± 859 nM. This equates to a selectivity of ERβ of ~750 fold over ERα. These results suggest that the coactivator plays a role in the selectivity of ISP358‐2. In a TR‐FRET coactivator binding assay the results showed a 15 fold selectivity for ERβ (EC 50 =191 ± 15 nM) over ERα (EC 50 =2,940 ± 390 nM). However, the coactivator assay utilizes a PPARɣ derived peptide. Using docking studies, we showed that the brain‐relevant SRC‐1 peptide docks more favorably. Modeling studies also demonstrate that the conformational change is favorable for coactivator recruitment for ERβ when agonist ISP358‐2 is bound, but not for the agonist‐bound ERα. Support or Funding Information This work was supported by NIH grants R15GM118304 and R01DA038042. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .

A short and efficient route to 4-(4-hydroxyphenyl)cycloheptanemethanol was developed, which resulted in the preparation of a mixture of 4 stereoisomers. The stereoisomers were separated by preparative HPLC, and two of the stereoisomers identified by X-ray crystallography. The stereoisomers, as well as a small family of 4-cycloheptylphenol derivatives, were evaluated as estrogen receptor-beta agonists. The lead compound, 4-(4-hydroxyphenyl)cycloheptanemethanol was selective for activating ER relative to seven other nuclear hormone receptors, with 300-fold selectivity for the β over α isoform and with EC50 of 30-50 nM in cell-based and direct binding assays.

Estrogen receptor-beta (ERβ) is a drug target for memory consolidation in post-menopausal women. Herein is reported a series of potent and selective ERβ agonists (SERBAs) with in vivo efficacy that are A-C estrogens, lacking the B and D estrogen rings. The most potent and selective A-C estrogen is selective for activating ER relative to seven other nuclear hormone receptors, with a surprising 750-fold selectivity for the beta over alpha isoform, and with EC50’s of 20-30 nM in cell-based and direct binding assays. Comparison of potency in different assays suggests that the ER isoform selectivity is related to the compound’s ability to drive the productive conformational change needed to activate transcription. The compound also shows in vivo efficacy after microinfusion into the dorsal hippocampus, and after intraperitoneal injection (0.5 mg/kg) or oral gavage delivery (5 mg/kg). This simple yet novel A-C estrogen is selective, brain penetrant, and facilitates memory consolidation.