The rediscovery of estetrol and its implications for estrogen treatment

Abstract

Objectives:

To summarize the literature on the rediscovery and clinical exploration of the fourth natural estrogen estetrol (E4), more than 100 years after the discovery of estrone (E1), estradiol (E2), and estriol (E3).

Methods:

Literature review of E4 publications.

Results:

Preclinical and clinical research and development of E4 revealed that: (1) contrary to the other 3 natural estrogens, E4 has a high oral bioavailability, and (2) oral E4 has limited effect on hemostasis and other liver functions, and is, therefore, expected to be a safer estrogen. Since 2022, E4 combined with drospirenone has been registered worldwide as an oral contraceptive. E4 for menopausal hormone therapy is expected to become available in 2026. Clinical development of the use of E4 for the treatment of advanced breast cancer and advanced prostate cancer is ongoing, and E4 has been proposed as a component of an oral male contraceptive.

Conclusion:

The recognition of E4 as a useful and safe natural estrogen for human use is expected to change the scene of estrogen treatment in women and men significantly.

THE DISCOVERY OF NATURAL ESTROGENS

In 1923, Edgar Allen and Edward A. Doisy (Nobel Prize honoree 1943), demonstrated that ovarian follicles, which produce eggs in mammals, also produce a primary ovarian hormone, later named estrogen.^1,2 After hearing about the Ascheim-Zondek test for the diagnosis of pregnancy, Allen and Doisy realized there exists a rich and easily accessible source of hormones in urine from which they could develop a potent extract. Whereas the research of Allen and Doisy was sponsored by the Washington University School of Medicine (St Louis, MO), the work of their main rival, the German scientist Adolf Butenandt (Nobel Prize honoree 1939) of the University of Gottingen, was sponsored by a German pharmaceutical firm. In 1929, both teams announced the isolation of a pure crystal form of the female sex hormone, estrone (E1), but Allen and Doisy did so 2 months earlier than Butenandt. By 1931, E1 was commercially produced by Parke-Davis (Detroit, MI) and by Schering-Kahlbaum AG (Berlin, Germany). When Butenandt isolated E1 and analyzed its structure, he found that it was a steroid, the first hormone to have been identified in this molecular family. Estradiol (E2) and estriol (E3) were subsequently discovered in 1933. The most important male hormone, testosterone, was first isolated and synthesized in May of 1935 by Ernst Laqueur, a scientist working at Organon BV (Oss, The Netherlands), 3 months ahead of Butenandt, then working with Schering AG (Berlin, Germany).

THE IDENTIFICATION OF ESTETROL

Three decades after the discovery of E1, a fourth natural estrogen was identified by Egon Diczfalusy at the Karolinska Institute (Stockholm, Sweden) in 1965,³ and by Elio Gurpide at Rockefeller University (New York, NY) in 1966.⁴ Elio Gurpide named it estetrol (E4) because this estrogen has 4 hydroxyl (OH) groups, whereas E1 has 1 OH group, E2 has 2 OH groups, and E3 has 3 (Fig. 1). After its discovery, in the period from 1965-1985, biochemical and pharmacological studies were performed to elucidate the biosynthesis of E4 and characterize its properties.⁵ It was shown that E4 is synthesized by the fetoplacental unit in which 15α and 16α hydroxylation by the fetal liver play an exclusive role.^6,7 E4 concentrations were found to rise exponentially during the third trimester of pregnancy and reach the maternal bloodstream through the placenta. At term, the mean maternal E4 levels turned out to be about 1 ng/mL, which is high, but much lower than the levels of the other 3 natural estrogens. In fetal plasma, E4 levels were 10-20 times higher than in maternal plasma since E4 passes the placenta from the fetus to the mother.⁸ Further studies in human proliferative endometrium revealed that, compared with E2, E4 has a relative receptor binding affinity of about 5%.⁹ In immature rat uteri, E4 tends towards cell differentiation, in contrast to the typical mitotic responses observed with E2.¹⁰ In sheep, E4 is 15-30 times less potent than E2 and E3 for uterine vasodilation.¹¹ When tested in human MCF-7 breast cancer (BCa) cells, the E4 agonistic potency is only 2% of that of E2 in terms of in vitro proliferation.¹² An attempt to establish E4 as a marker of fetal well-being during pregnancy was made, but the effort was abandoned due to the large variations in blood and urine levels.⁵ In the 80s, it was concluded that E4 is a weak estrogen and no further research was performed. Estetrol was forgotten, as illustrated by a 1986 paper by Diczfalusy¹³ himself on human reproduction research, in which E4 is not mentioned at all.

FIG. 1.

The 4 estrogens highlighting structure and nomenclature.

THE REVIVAL OF ESTETROL

In 2001, the lead author of this paper, Herjan J.T. Coelingh Bennink at Pantarhei Bioscience (PRB; Zeist, The Netherlands), started a new research program on E4 with the aim to investigate whether E4 could be used as an estrogen for human therapeutic use. Historically crucial for the rediscovery of E4 was the first study on the bioavailability of E4 in rats, demonstrating a remarkable and much higher oral bioavailability than that of other natural estrogens. This finding was confirmed in humans, where E4 demonstrated an oral bioavailability of 70%-80%, a long elimination half-life of about 20-28 hours, and minimal binding to sex hormone-binding globulin. These characteristics made E4 notably more suitable for once-a-day oral dosing than other natural estrogens.¹⁴,¹⁵,¹⁶ These findings were confirmed in 2002 by Johnson & Johnson (J&J; New Brunswick, NJ). This company was planning to develop E4 for menopausal hormone therapy (MHT), but after the publication of the Woman’s Health Initiative study in 2002,¹⁷ J&J decided not to embark on MHT anymore. PRB then decided to discontinue efforts to develop E4 for MHT and shifted to the development of E4 for combined oral contraception (COC). The first presentation on “a new natural human estrogen” took place at the 10th World Congress on Menopause in Berlin (Germany) in 2002.

ESTETROL FOR COMBINED ORAL CONTRACEPTION AND MENOPAUSAL HORMONE THERAPY

During the new E4 research and development by PRB from 2002 to 2015, 2 essential discoveries were made in human studies: First, the favorable pharmacokinetic properties of E4 after oral administration mentioned previously;^14,15 Second, the low interference of oral E4 with liver function and hemostasis demonstrated in phase 1 and phase 2 clinical studies for women’s health applications such as COC’s and MHT.^16,18,19 The combination of these 2 properties of E4 means that E4 can be given safely and effectively at high oral doses. Dose-finding studies with E4 have shown that 15-20 mg per day is suitable for COC and MHT (ie, 10 times the dose used for E2), and up to 60 mg/d seems safe and effective for oncological use in advanced BCa and advanced prostate cancer (PCa).^20,21 Fetal exposure to E4 at term pregnancy is comparable to an oral dose of 50-60 mg E4 as demonstrated by kinetic simulation.¹⁶ Instead of speaking about an estrogenic drug as weak or strong, we prefer to talk about its potency. Potency is the result of the pharmacokinetic properties of the drug and its interaction with the receptor, in this case, the estrogen receptor. In the case of comparing the potency of E4 with E2, both absorption and metabolism are important. Whereas E4 has an oral bioavailability of 70%-80% and has no metabolites, E2 is heavily metabolized by the liver after oral administration, resulting in only 5% of the oral dose becoming bioavailable as E2, whereas the remaining 95% enters the body as E1 and E1 sulfates.^16,22 This means that oral treatment with E2 actually is oral treatment with E1. Transdermal and vaginal use of E2 does not have metabolic liver complications and provides primarily E2 to the body.

Mode-of-action studies showed that, while E2 activates both the nuclear and membrane estrogen receptor alpha (ERα), E4 binds to nuclear ERα, but does not activate membrane ERα in specific tissues (resulting in, eg, improving endothelial healing, and a minimal impact on liver function and on normal and malignant breast tissue).²³ In addition, when coincubated with E2 in vitro, E4 was able to partially antagonize the E2-induced proliferation of human breast epithelium.²⁴

In 2015, E4 was out-licensed to the Belgian company Mithra Pharmaceuticals SA (Liege, Belgium) for further development in nononcological women’s health applications. Phase 3 thirteen-cycle studies in 3,027 participants revealed that 15 mg E4 combined with 3 mg drospirenone is a safe and effective COC with a predictable bleeding pattern and Pearl Indices of 0.44 (European/Russian study) and 2.42 (USA/Canadian study).²⁵ Moreover, it has limited effects on hemostasis, and beneficial effects on lipids, carbohydrate metabolism and bone turnover.²⁵,²⁶,²⁷ Its adverse event profile aligns with other COCs.²⁵ Compared with ethinylestradiol COCs, E2-COC and E4-COC appear to have a safer thrombotic profile, based on pharmacovigilance data on reported venous thromboembolism.²⁸ Since 2022, the E4/drospirenone COC is available worldwide. In a phase 2 trial, 15 mg E4 was effective for the relief of vasomotor symptoms, as shown by a significantly decreased frequency and severity of hot flushes, and decreased symptoms related to signs of the genitourinary syndrome of menopause.^29,30 This has been confirmed in phase 3 studies investigating 15 mg and 20 mg daily doses.³¹ It is expected that E4 for MHT will become available in 2026.

ESTETROL FOR REPRODUCTIVE ENDOCRINE ONCOLOGY

Since 2015, clinical studies in BCa and PCa have been performed by Pantarhei Oncology BV (an affiliate of PRB). A dose of 20 mg E4 induces apoptosis in BCa tumor tissue in women with advanced BCa,³² and higher doses (40 mg and 60 mg) have antitumor effects in patients with end-stage BCa.³³ In a phase 2 study in patients with advanced PCa, E4 was used at a dose of 40 mg per day as a cotreatment of androgen deprivation therapy.³⁴ No treatment-related cardiovascular events are observed, and hot flushes and bone turnover are substantially reduced. Enhanced disease control seems likely by E4 androgen deprivation therapy cotreatment by further suppressing free testosterone, prostate-specific antigen, follicle-stimulating hormone, and insulin-like growth factor-1.

ESTETROL FOR OTHER CLINICAL APPLICATIONS

The rediscovery of E4 and the new research and development data stimulated many other researchers to evaluate the clinical potential of E4 for a range of new applications. Estetrol research is presently ongoing in dysmenorrhea,³⁵ endometriosis,³⁶ female sexual function,³⁷ wound healing,³⁸ neonatal hypoxic-ischemic encephalopathy,³⁹ and hair loss.⁴⁰ In addition, E4 has been proposed as a component of an oral male contraceptive.⁴¹

CONCLUSIONS

A century after the discovery of the natural estrogens estrone, estradiol, and estriol, the human fetal estrogen estetrol has been rediscovered and investigated extensively. The journey of estetrol from its identification in 1965 and its rediscovery and successful clinical development since 2001 confirms its potential as a unique and most likely safer estrogen for human use.