Estrogen Receptor Structures & Functions

Steroid or nuclear hormone receptors (NRs) constitute an important superfamily of transcription regulators that are involved in widely diverse physiological functions, including control of embryonic development, cell differentiation and homeostasis. Members of the superfamily include the steroid hormone receptors and receptors for thyroid hormone, retinoids, 1,25-dihydroxy-vitamin D3 and a variety of other ligands. The proteins function as dimeric molecules in nuclei to regulate the transcription of target genes in a ligand-responsive manner. In addition to C-terminal ligand-binding domains, these nuclear receptors contain a highly-conserved, N-terminal zinc-finger that mediates specific binding to target DNA sequences, termed ligand-responsive elements. In the absence of ligand, steroid hormone receptors are thought to be weakly associated with nuclear components; hormone binding greatly increases receptor affinity.

NRs are extremely important in medical research, a large number of them being implicated in diseases such as cancer, diabetes, hormone resistance syndromes, etc. While several NRs act as ligand-inducible transcription factors, many do not yet have a defined ligand and are accordingly termed "orphan" receptors. During the last decade, more than 300 NRs have been described, many of which are orphans.

The oestrogen receptor consists of 3 functional and structural domains: an N-terminal (modulatory) domain; a DNA binding domain (DBD) that mediates specific binding to target DNA sequences, termed ligand-responsive elements; and a ligand binding domain (LBD). The N-terminal domain is unique to the oestrogen receptors and spans approximately the first 180 residues; the highly-conserved DNA-binding domain is smaller (around 65 residues) and occupies the central portion of the protein; and the ligand binding domain lies at the receptor C-terminus.

The oestrogen receptor stimulates transcription via 2 distinct transcriptional activation domains: TAF-1 in the N-terminal domain, and TAF-2 in the hormone-binding domain. TAF-2 activity requires a region in the C-terminus of the hormone-binding domain (between residues 538-552 in the mouse oestrogen receptor) that is conserved in many nuclear hormone receptors. It is therefore suggested that this region may be essential for ligand-dependent transcriptional activation by other members of the nuclear receptor family. The hormone-receptor complex appears to recognize discrete DNA sequences upstream of transcriptional start sites.

Figure 2. Structure of estrogen receptor (ER)
A to F represent different domains of the ER. Numbers
represent amino acids from amino to carboxy termini.

Comparison of the amino acid sequences of the ERa and ERb- receptors:
GustafssonF2adapted
The separate domains are identified in the ERa diagram; the numbers in the ERb diagram show the sequence identity as %.A subtle difference between the two in their ligand-binding pockets is the substituiton of Leu 338 in ERa with Met 384 in ERb

Coactivators and Corepressors

Transactivation requires the recruitment of coactivators, such as SRC-1, that posess histone acetyltransferase activity or can recruit a histone acetyl transferase. This complex can decompact the chromatin, enabling a transcription initiation complex to form. Silencing involves the recruitment of corepressors, such as SMRT, and histone deacetyltransferases. The transconformation of Helix12, along with other associated structural changes, creates a surface on the receptor that can bind coactivators such as SRC-1. These coactivators contain one or more "LXXLL boxes" that are responsible for nuclear receptor binding, where L is leucine and X is any amino acid in the sequence motif.

Expression

Expression of the estrogen receptor (ER) has a key role in breast cancer; the ER gene is located at chromosome 6q24-q27 (6q25.1) and is made up of 8 exons with a total of 140 kb. The polymorphism in codon 325 of exon 4 (ER325) is a transition CCC-->CCG. There is a relationship between the ER325 polymorphism and susceptibility to breast cancer.

The estrogen-related receptors (ERR alpha, -beta and -gamma), a subfamily of orphan nuclear receptors closely related to the ERs, have shown that the ERRs share target genes, coregulatory proteins, ligands and sites of action with the ERs.

Pharmocology

Selective Estrogen Receptor Modulators (SERMs) are a new class of drugsthat bind to estrogen receptor (ER) and elicit agonistic or antagonistic responses, depending on the target tissue.

SERMs' potential as the next generation in menopausal treatment is that it exhibits different effects depending on the tissue; this is likely due to the way the SERM interacts with estrogen receptor(ER)a and ERb subtypes and the ER subtype distribution. Studies have found, for example, that the SERM Tamoxifene exhibits antiestrogenic effects in the breast thus inhibiting growth of breast tumor cells that have estrogen-receptors. In the uterus Tamoxifene is estrogenic and stimulatory, thereby increasing the risk of endometrial cancer. Raloxifene, another SERM, has been shown to increase bone mineral density in postmenopausal women but has antiestrogenic effects on both breast tissue and endometrial tissue. Both SERMs have estrogenic effects on serum lipoproteins and coagulation. Raloxifene has been recently shown to reduce cardiovascular events but ,as do Tamoxifene and estrogen, increase the incidence of thromboembolic events, such as strokes. SERMs exacerbate vasomotor instability such as hot flashes but have unknown effects on the central nervous system.

Ideal SERM:

Commercialy available SERMs:

SERM

Compagny

Tamoxifene

Raloxifene

Zeneca

Eli Lilly

Idoxifene

SmithKline Beecham

Droloxifene

Levormeloxifene

Pfizer

Novo Nordisk

RALOXIFENE

Raloxifene is being described as the first multifunctional drug. The molecule, raloxifene hydrochloride was developed in an effort to find a treatment for breast cancer and osteoporosis. Tissue selectivity of raloxifene may be achieved through several mechanisms: the ligand structure, interaction of the ligand with different estrogen receptor subtypes in various tissues, and intracellular events after ligand binding. Raloxifene has estrogen-agonistic effects on bone, lipids and clotting factors, and estrogen-antagonistic effects on the breast and uterus. It appears to prevent bone fractures and to reduce the risk of breast cancer.

Structure

Raloxifene is a nonsteroidal benzothiophene compound. Several modifications to the 2-arylbenzothiophene core have shown that the 6-hydroxy and the 4'-hydroxy substituents of raloxifene are important for estrogen receptor binding and for mimicking the corresponding 3- and 17beta-hydroxy groups of 17beta-estradiol . An estrogen-antagonistic region of raloxifene is characterized by a piperidine side chain. The orthogonal orientation of this basic side chain may contribute to raloxifene's lack of uterotrophic effects. Replacement of the 6-hydroxy substituent with various functional groups has resulted in a reduction in cholesterol-lowering effect and antiproliferative action, whereas substitution of the 4`-hydroxy group with small electronegative substituents did not change the in vivo profile . Substitution of a larger group at the 4` position, however, resulted in increased uterine stimulation. Several raloxifene analogues are under investigation for their potential applications in a variety of diseases in which estrogen is thought to play a pathogenic role, such as uterine leiomyoma and endometriosis .

MECHANISM OF ACTION

The SERMs compete with endogenous estrogens for binding to the receptor and may either activate or block estrogen action. The detailed molecular mechanism by which estrogens and SERMs exert their biological effects on different tissues has not been fully elucidated and remains an area of intensive research. Selective responses are thought to be mediated at three different sites: ligand (estrogen compared with SERM), receptor (estrogen receptor subtypes), and effector site (specific intracellular events in different tissues) .

Raloxifene may interact with unique subsets of coactivators and co-repressors; interfere with the formation of estrogen receptor-associated proteins with the estrogen receptor; or display estrogen receptor-independent nongenomic effects, as tamoxifen does.