Progesterone signaling in uterine fibroids: Molecular mechanisms and therapeutic opportunities

Abstract

Progesterone (P4) is a vital female sex hormone involved in various physiological processes, including the maintenance of the endometrium, mammary gland development, and bone health. Beyond its reproductive roles, P4 is implicated in the pathogenesis of hormone-dependent conditions like uterine fibroids, the most common benign tumors in women, which can severely affect quality of life and fertility. Traditionally, estrogen was considered the primary driver of fibroid growth, but recent research highlights the significant role of P4 in fibroid growth. P4 interacts with progesterone receptors (PRs) and non-genomic membrane receptors (mPRs and PGRMCs) to activate signaling pathways that enhance tumor growth and survival. P4 promotes vascular changes that improve the blood supply to fibroids and modifies the extracellular matrix, a key component of fibroid structure. This understanding has led to the investigation of selective progesterone receptor modulators (SPRMs) as potential therapies for fibroids. Clinical trials have demonstrated the effectiveness of SPRMs like mifepristone, asoprisnil, and ulipristal acetate in reducing fibroid size and symptoms, though concerns about safety, particularly with long-term use, remain. Newer SPRMs, such as vilaprisan, show promise, but further research is necessary to assess the long-term safety and effectiveness. This review discusses the mechanisms by which progesterone contributes to fibroid growth and examines clinical effectiveness of SPRMs as potential treatments for uterine fibroids.

Graphical Abstract

INTRODUCTION

Progesterone (P4) is a female sex steroid hormone primarily produced by the corpus luteum during the luteal phase of the menstrual cycle, synthesized from cholesterol and metabolized mainly in the liver. Its primary role is to transition and maintain the endometrium in a secretory state for embryo implantation and pregnancy support. P4 also contributes to lobuloalveolar development in the mammary glands and promotes osteoblastic activity in bones, preventing bone loss [1]. Beyond reproductive functions, P4 signaling is implicated in hormone-dependent diseases like endometriosis, breast cancer, and uterine fibroids [2].

Uterine fibroids, the most common benign gynecological tumors, can significantly affect fertility and quality of life depending on their size, number, and location. While estrogen was traditionally seen as the main hormonal driver, recent research highlights the role of P4 in fibroid pathogenesis. Studies show P4 regulates angiogenic factors, extracellular matrix components, growth factors, and apoptotic agents in fibroid cells [3]. Given the role of P4 in promoting fibroid cell proliferation, selective progesterone receptor modulators (SPRMs) have been explored as potential therapies. This scoping review examines how P4 and its receptors contribute to fibroid pathogenesis and summarizes key findings on the efficacy and tolerability of SPRMs in treating uterine fibroids.

OVERVIEW OF PROGESTERONE RECEPTORS

The effects of progesterone in human cells are largely mediated by two main isoforms of the progesterone receptor (PR): PR-A (94 kDa) and PR-B (120 kDa). These receptors, products of the same gene (PGR on chromosome 11), act as ligand-dependent transcription factors. PR-A is a truncated version of PR-B, differing by 164 amino acids in the N-terminal region [4]. Beyond PR-A and PR-B, several other truncated PR isoforms (PR-C, PR-M, PR-S, PR-T) have been identified, though their roles remain unclear [5].

PRs contain four main domains: the N-terminal domain (NTD), DNA binding domain (DBD), hinge region, and ligand-binding domain (LBD), along with activation function (AF) regions critical for gene transcription [5]. The unique AF3 domain in the N-terminal domain (NTD) of PR-B enhances its transcriptional activity, leading to a stronger gene-activating function compared to PR-A, which can even act to repress PR-B activity [6].

Progesterone signaling also occurs outside the nucleus via membrane-bound receptors: membrane progesterone receptors (mPRs) and progesterone receptor membrane components (PGRMCs). These receptors facilitate rapid, non-genomic P4 signaling and belong to different receptor families, including PAQRs and MAPRs [7,8].

PROGESTERONE SIGNALING PATHWAYS

Classical progesterone signaling

Nuclear (genomic) PRs, PR-A and PR-B, mediate the classical or genomic P4 signaling pathway (Fig. 1). In the absence of P4, PRs are found in the cytoplasm in an inactive state, bound to chaperone proteins. Heat shock proteins (HSP90, HSP70 and HSP40), and cochaperone molecules, HSP90-binding protein p23 and HSP70/HSP90 organizing protein (Hop), prevent PRs from entering the nucleus [9]. Upon ligation with P4, PRs undergo a series of conformational changes, which leads to their release from the chaperone proteins and their translocation to the nucleus. Inside the nucleus, PRs dimerize and the dimers bind to specific promoter regions, called progesterone receptor elements (PREs) [10]. Along with other gene specific co-regulators and transcription factors, they form a complex which can induce or inhibit the expression of target genes. A large variety of physiological responses can be initiated, depending on whether PR dimers are in the form of homodimers (AA or BB) or heterodimers (AB). Activated PRs can also bind to DNA regions not identified as PREs by interacting with specific transcription factors, such as activating protein 1 (AP-1), specificity protein 1 (Sp1), signal transducer and activator of transcription5 (STAT5), and nuclear factor kappa-lightchain-enhancer of activated B cells (NF-κB) [5].

Figure 1.

Progesterone receptors and their signaling pathways. A. When progesterone is absent from the cytoplasm, genomic progesterone receptors (PR-A and PR-B) are attached to chaperone molecules. Genomic progesterone signaling is initiated when progesterone binds to PRs, which induces their detachment from the chaperone molecules and their translocation from the cytoplasm to the nucleus, where they form dimers and bind to specific DNA promoter regions called progesterone receptor elements (PREs). Then, the transcription of these target genes is either activated or suppressed, depending on the activity of other gene co-regulators that bind to the promoter region. B. Progesterone can also exert rapid non-genomic actions by binding to non-genomic receptors that are located on the cell membrane, called membrane progesterone receptors (mPRs) and progesterone receptor membrane components (PGRMCs). Activation of mPRs by progesterone ignites multiple intracellular signaling pathways, including the MAP kinase and PI3/AKT pathways. PGRMC1 is associated with SERBP1 forming a complex on the cell membrane. Binding of progesterone to this complex causes rise in cAMP levels and JAK/STAT activity.

Non-classical progesterone signaling

Progesterone (P4) can activate rapid, non-genomic effects through non-classical receptors, including membrane progesterone receptors (mPRs) and progesterone receptor membrane components (PGRMCs). This signaling pathway bypasses nuclear transcription, triggering intracellular responses within minutes [11]. mPR isoforms (mPRα, mPRβ, and mPRγ) are coupled with inhibitory G (Gi) proteins, leading to decreased cAMP and protein kinase A activity, while mPRδ and mPRε are coupled with stimulatory G (Gs) proteins, increasing cAMP and PKA activity [12,13]. mPRs also activate PI3K/AKT and MAP kinase pathways, impacting various reproductive functions [12,14].

PGRMC1, part of the PGRMC family, contains a cytochrome b5 domain that interacts with CYP enzymes involved in steroidogenesis, potentially influencing P4 synthesis [15]. PGRMC1 can associate with serpine 1 mRNA binding protein (SERBP1) on the cell membrane, forming a complex that, when activated by P4, reduces intracellular Ca²⁺ through increased cAMP and protein kinase G activation [16,17]. Additionally, PGRMC1 may enhance mPRα expression on the cell surface, indicating a cooperative role in non-genomic P4 signaling [18].

EXPRESSION OF PROGESTERONE RECEPTORS IN UTERINE FIBROIDS

P4 exerts its effect on fibroids mainly through interaction with classical PRs, since PR-A and PR-B expression has been shown to be higher in leiomyomas compared to normal myometrium [19,20]. The efficacy of selective progesterone receptor modulators (SPRMs) for the treatment of uterine fibroids is also strong indirect evidence that the interaction of P4 with classical PRs contributes to fibroid growth [21,22].

Research regarding the effect of mPRs on uterine leiomyomas remains limited, although a recent study examining the expression of PRs in uterine leiomyoma tissue samples suggested that both nuclear and membrane PRs contribute to leiomyoma pathobiology via distinct mechanisms [23]. It is evident that P4 and its receptors play a major role in uterine fibroid pathogenesis; however, further research is needed to understand the interactions between PRs and other regulators of tumor growth and to potentially discover new signaling pathways that can set new therapeutic targets. For instance, the crosstalk between P4 and E2 signaling has been shown, with studies indicating that E2 induced PR expression in leiomyoma cells [24]. Katzenellenbogen et al. demonstrated that E2 upregulated PR expression in breast cancer cells[25], suggesting a synergistic relationship between E2 and P4 signaling pathways. Moreover, mPRs regulate cAMP and PKA activity through Gi and Gs protein coupling, activating PI3K/AKT and MAPK pathways, while PGRMC1 may influence non-genomic P4 signaling via cAMP and protein kinase G [12–14,16,17]. These findings suggest that a complex regulatory network of P4 signaling may exist in fibroid pathogenesis and may influence the effectiveness of SPRMs.

FUNCTIONAL ROLE OF PROGESTERONE IN UTERINE FIBROIDS

Progesterone and extracellular matrix deposition in uterine fibroids

Although the exact underlying mechanisms that drive ECM deposition in uterine fibroids are still under investigation, many studies have shown that P4 may play a key role through its interaction with growth factors, ECM proteins, and MMPs [26]. Transforming growth factor-β (TGF-β) is one of the major growth factors that induce fibrosis in uterine fibroids. TGF-β3 expression was shown to be increased during the secretory phase (mediated mainly by P4), indicating the importance of P4 signaling on TGF-β function [27,28]. Recent studies proved that TGF-β1 and TGF-β3 transcripts were markedly decreased when uterine fibroid cells were treated with ulipristal acetate, a SPRM, indirectly suggesting that P4 mediates TGF-β signaling [23,29,30]. Moreover, P4 has been found to decrease the expression of decorin, a proteoglycan that inhibits TGF-β3, suggesting an alternative mechanism of P4 induced ECM deposition [31]. The expression of MMPs and TIMPs was also observed to be upregulated in leiomyoma cells when treated with the SPRM asoprisnil, proving that P4 can induce ECM accumulation by inhibiting its degradation by metalloproteinases [32]. Additionally, P4 was reported to enhance collagen synthesis, a major ECM component, through downregulation of mi-R29b in uterine leiomyoma xenografts [33].

The four mechanisms mentioned above (increased TGF-β expression, downregulation of decorin, upregulation of MMPs, enhanced collagen synthesis) provide strong evidence of P4’s influence on abnormal ECM synthesis and remodeling in uterine leiomyomas. More extended research on the molecular mechanisms mediating these interactions, and further exploring the effect of SPRMs on ECM deposition, will provide useful information and hopefully reveal new therapeutic targets for uterine fibroid shrinkage.

Progesterone and angiogenesis in uterine fibroids

The vascular pattern of uterine leiomyomas is characterized by a highly vascularized area in the periphery, also known as “perifibromal vascular capsule,” which surrounds a relatively hypovascular core [34]. Although uterine fibroids are mostly hypovascular, angiogenesis plays a key role in maintaining and promoting their growth, as implicated by the efficacy of uterine artery embolization in reducing fibroid size [35]. The difference in vascular architecture between uterine fibroid tissue and normal myometrium also indicates that dysregulated angiogenesis is part of fibroid pathophysiology and is potentially mediated by dysregulated expression of angiogenic and antiangiogenic factors [34,36].

Many studies have established that the expression of primary angiogenic growth factors, such as VEGF, EGF, FGF, PDGF, ADM and TGF-β, is higher in human leiomyomas compared to normal myometrial tissue [36]. In a recent study, aberrant expression of HMGA2, a known driver gene mutation of uterine fibroids, was linked to increased levels of angiogenic factor and their receptors compared to MED-12 mutant leiomyoma cells, suggesting that vascular alterations in uterine fibroids may be attributed to HMGA2 overexpression [37,38]. Another mechanism by which leiomyoma cells exhibit altered angiogenesis is their distinct reaction to hypoxia. In other tumors, hypoxia triggers the expression of hypoxia inducible factor-1 (HIF-1), which enhances angiogenesis and cell proliferation to promote tumor survival and growth [39]. However, HIF-1α and HIF-1β transcription has been repeatedly found to be low in leiomyoma cells despite being significantly hypoxic, suggesting a lack of response to hypoxia and potentially explaining why leiomyomas exhibit diminished vascularity and lack malignant features [36,40].

P4 is a key mediator of uterine angiogenesis, mainly through its interaction with VEGF [41]. Although the specific mechanisms are not fully established, many studies have shown that P4 signaling also regulates angiogenesis in uterine fibroids [36,42,43]. P4 was observed to upregulate EGF expression in leiomyoma cells, while EGF transcripts significantly declined under the influence of asoprisnil, suggesting that P4 signaling regulates EGF expression and thus, angiogenesis [43,44]. Additionally, treatment of human leiomyoma cells with ulipristal acetate resulted in downregulation of ADM and VEGF as well as their receptors, indirectly indicating the role of P4 in promoting angiogenesis [42]. More thorough examination of the relationship between P4/PRs and angiogenic growth factors is needed to answer whether the dysregulated vascular pattern of uterine fibroids can be attributed to P4 stimulation.

Progesterone and apoptosis in uterine fibroids

Several studies have supported that Bcl-2, an antiapoptotic gene, is upregulated in uterine leiomyomas and significantly contributes to their pathogenesis [45–47]. Moreover, Bcl-2 protein was found to be abundant in leiomyoma cells but undetectable in myometrial tissue, indicating that Bcl-2 overexpression and subsequent escaping of the apoptosis is an essential feature of uterine fibroids [45].

Bcl-2 protein expression in leiomyomas was found to rise in accordance with P4 in the secretory phase, suggesting that P4 signaling can inhibit apoptosis through Bcl-2 protein activation [45]. Furthermore, in vivo leiomyoma tissue exhibited significantly greater levels of Bcl-2 mRNA when treated with P4, and liganded PR-A was found to strongly bind to the Bcl-2 promoter, suggesting that P4 can induce Bcl-2 gene expression through its interaction with PR-A [48]. The antiapoptotic effect of P4 was also validated in a more recent study, where P4 exhibited a dose-dependent inhibitory effect on apoptosis of uterine leiomyoma tissue cultures [49]. Additionally, telapristone acetate and asoprisnil, both potent SPRMs, were found to significantly decrease Bcl-2 protein levels while increasing apoptosis markers, indirectly proving the role of P4 on apoptosis evasion [50,51]. Another mechanism by which P4 may promote leiomyoma cell survival is by rapid activation of the AKT pathway, an essential signaling pathway that regulates cell proliferation, differentiation and survival and is commonly overactivated in many cancers [52,53]. In this study, phosphorylated AKT levels in leiomyoma cells significantly increased when treated with P4, an effect not observed in matched myometrial cells [53].

Escaping apoptosis is, without a doubt, an essential part of uterine fibroid survival. Exploring other possible antiapoptotic mechanisms of P4 signaling along with more extended research on the underlying molecular interactions between P4/PRs and proapoptotic/antiapoptotic factors will help deepen the understanding of the importance of P4 on uterine fibroid cell survival and potentially lead to new therapeutic targets.

TARGETING PROGESTERONE SIGNALING FOR FIBROID TREATMENT

Selective progesterone receptor modulators (SPRMs) are a new class of synthetic steroid drugs that act on PRs, inducing agonistic, antagonistic or combined effects on PR signaling, depending on the cell type [5] (Fig. 2). This divergent effect can be attributed to the difference in PR-A/PR-B ratio in each tissue and the affinity of each SPRM for every isoform, as well as the interaction of each SPRM with different gene co-activators or co-repressors [5,54]. Given the role of P4 in uterine fibroid pathogenesis, SPRMs have been suggested as a promising class of drugs for the treatment of this disease [54]. In this section, we discuss the mechanisms of action and the therapeutic effects of the main SPRMs, which include mifepristone, ulipristal acetate, asoprisnil and vilaprisan, on multiple P4-mediated effects that contribute to fibroid growth, such as cell proliferation, ECM synthesis, angiogenesis and apoptosis (Fig. 3 and Table 1).

Figure 2.

Mechanism of action of different SPRMs. SPRMs can exert agonistic, antagonistic or mixed effects on progesterone signaling by attaching to progesterone receptors with different affinities and recruiting specific coactivators or co-repressors leading to enhancement or silencing of target gene transcription, respectively.

Figure 3.

Molecular mechanisms and therapeutic potential of SPRMs in uterine fibroids. SPRMs* can reduce uterine fibroid size by over- or downregulating the expression of various molecules that are involved in different aspects of fibroid pathogenesis, including fibroid cell proliferation and apoptosis, angiogenesis and extracellular matrix (ECM) accumulation. All SPRMs exert potent antiproliferative, proapoptotic and antifibrotic effects. Ulipristal acetate exhibits more prominent antiangiogenic function compared to other SPRMs. *Vilaprisan is not mentioned since the molecular mechanisms which contribute to its clinical efficacy are still unclear.

Table 1.

Clinical efficacy of SPRMs in uterine fibroids.

				Clinical efficacy of SPRMs in uterine fibroids
SPRMs	Clinical Trial	Treatment regimen	Amenorrhea rates	Fibroid size	Uterine size	Adverse Events (AEs)
Mifepristone	Phase II [66]	5 mg or 10 mg daily for 12 months	40–70% in both groups	-	↓52–53% in both groups	-Simple endometrial hyperplasia in 13.9% (all in the 10 mg group) at 6 months
	[67]	5 mg or 10 mg daily for 3 months	89.8% in the 10 mg and 90% in the 5mg group	↓ 45% in the 10 mg and ↓57% in the 5 mg group	↓40% in the 10-mg and ↓36% 5-mg group	-Simple endometrial hyperplasia in 1 of 50 patients in the 10 mg group
	[113]	10 mg or placebo daily for 3 months	84.2% in the 10 mg group	↓26–32% in the 10 mg group versus none in the placebo	↓26–32% in the 10 mg group versus none in the placebo	-Simple endometrial hyperplasia in 63.1% in the 10 mg group
	Phase II [70] (NCT00881140)	10 mg vaginally daily for 3 months	44.8%	↓26.4%	-	-Hot flushes (10.3%), abdominal pain (24.1%), vaginal discharge (20.7%)
	[71]	25 mg daily (group 1) or 50 mg biweekly (group 2) for 3 months	97.8% in group 1 and 97.9% in group 2	↓21.7% in the 25 mg and ↓19.7% in the 50 mg group	↓18.5% in the 25 mg and ↓15.7% in the 50 mg group	-Nausea 6.7% and 4.2%, vomiting in 2.2% and 2.1%, hot flushes in 4.4% and 6.3%, fatigue in 4.4% and none, and diarrhea in 2.2% and none of the patients in Group 1 and 2, respectively
	Phase III [114] NCT00133705	5 mg or placebo daily for 26 weeks	41% in the 5 mg group	-	↓47% in the 5 mg group	-
	Phase III [115] (NCT01786226)	2.5 mg or 5 mg daily for 3 months	-	↓27.9% in the 2.5 mg and ↓45.5% and 5 mg group	↓18.2% in the 2.5 mg and ↓22.1% and 5 mg group	-No significant differences between the treatment groups
Ulipristal acetate	Phase II [86] (NCT00290251)	10 mg or 20 mg or placebo daily for 3 or 6 months	8/13 and 12/13 women in the 10 mg and 20 mg groups, respectively, versus none in the placebo group -	↓17% in the 10 mg and ↓24% in the 20 mg group vs ↓7% in the placebo group	-	-Vaginal spotting/discharge, fatigue, breast pain, hot flushes more common in the 10 mg group -PRL elevation in 17 women -ALT or AST abnormal levels in 9 women in the UPA groups
	Phase III [87] PEARL I (NCT00755755)	5 mg or 10 mg or placebo daily for 13 weeks	73% and 82% in the 5 mg and 10 mg groups, respectively, versus 6% in the placebo group	↓21% in the 5 mg and ↓12% in the 10 mg group vs ↑6% in the placebo group	> 25% reduction in more patients in the UPA group compared to placebo	-PAECs in 62% and 57% of women in the 5 mg and 10 mg groups, respectively, versus 6% in the placebo group
	Phase III [88] PEARL II (NCT00740831)	5 mg or 10 mg oral UPA daily or once-monthly IM leuprolide acetate (LA) injection for 3 months	-	36% in the 5 mg and 42% and 10 mg group vs 47% in the LA group (Fibroid volume decrease was maintained longer post treatment with UPA compared to LA)	-	-Hot flushes in 11% and 10% in the 5 mg and 10 mg groups, respectively, versus 40% in the LA group -PAECs in 58% and 59% in the 5 mg and 10 mg groups, respectively, versus 12% in the LA group
	Phase III [21] PEARL III and PEARL III extension (NCT01156857, NCT01252069)	10 mg daily for 12 months (four 3-month courses)	-Amenorrhea rates were 79%, 89%, 88% and 90% after treatment courses 1, 2, 3 and 4, respectively	45%, 63%, 67% and 72% for treatment courses 1, 2, 3 and 4, respectively	-	-Headache (16.3%), nasopharyngitis (6.7%), abdominal pain (5.3%), hot flushes (4.8%), fatigue (4.8%) -PAECs in 26% and 25% after treatment 1 and 4, respectively
	Phase III [89] PEARL IV (NCT01629563)	5 mg or 10 mg daily for two repeated 12-week treatment courses	62% and 73% in the 5 mg and 10 mg groups, respectively, in both treatment courses	54% in the 5 mg and 58% in the 10 mg group	-	-Headache and hot flushes in >3% of patients in both treatment groups -PAECs in 8% and 19% after the second treatment course with 5 mg and 10 mg UPA, respectively -3 cases of endometrial hyperplasia
	Phase III [90] PREMYA (NCT01635452)	5 mg daily for 3 months (plus 12-month follow-up)	-60% of patients reported symptomatic improvement at 3 months -Reduced pain and improved QoL were sustained during follow-up			N/A
	Phase III [22] VENUS I (NCT02147197)	5 mg or 10 mg or placebo daily for 12 weeks (plus 12-week follow-up)	47.2% and 58.3% in the 5 mg and 10 mg groups, respectively, versus 1.8% in the placebo group	-	-	-TEAEs in 48.5% of patients in the UPA groups versus 28.6% in the placebo group -Hot flushes, blood CPK elevation and hypertension -PAECs in 26.2% and 29.7% in the 5 mg and 10 mg groups, respectively, versus 13.6% in the placebo group -1 case of simple endometrial hyperplasia in the 10 mg group
	Phase III [91] VENUS II (NCT02147158)	5 mg or 10 mg or placebo daily for two 12-week courses, separated by a drug-free interval of two menses	42% and 54.8% in the 5 mg and 10 mg groups, respectively, versus 0% in the placebo group in the first course	-	-	-Hot flushes in 7.5% and 11.6% in the 5 mg and 10 mg groups, respectively, versus 1.7% in the placebo group in the first course
	Phase III [92] MYOMEX	Group 1: 5 mg oral UpA daily + single placebo injection Group 2: single leuprolide acetate injection (11.25 mg in 1mL) + daily placebo for 12 weeks prior to laparoscopic myomectomy	-	↓7.2% in group 1 versus ↓38.4% in group 2 preoperatively	↓6.4% in group 1 versus ↓26.2% in group 2	-Hot flushes (Group 1: 7% versus 43%, Group 2: 12% versus 65%) -4 postoperative complications in group 1 and 2 in group 2
	Phase III [96] UCON	Group 1:5 mg UPA daily for three 12-week treatment courses (separated by 4-week drug-free intervals) Group 2: levonorgestrel-releasing intrauterine system	64% in group 1 versus 25% in group 2	-	-	-PAECs in 8% of patients in group 1 --Bladder symptoms, abdominal pain, musculoskeletal pain, nausea and upper respiratory tract infection by ≥ 3% of patients in group 1
	Phase IV[116]	5 mg daily for 3 months	-	↓50%	-	-No major complications
Asoprisnil	Phase II [105] NCT00160459	5 mg or 10 mg or 25 mg or placebo daily for 12 weeks	16%, 36%, and 70% in the 5, 10, and 25 mg groups, respectively, versus 0% in the placebo group	-	↓14%, ↓9%, and ↓17% in the 5, 10, and 25 mg groups, respectively, versus ↑1% in the placebo group	-PAECs in 43%, 58%, and 58% in the 5, 10, and 25 mg groups, respectively -Bloating, breast pain, and vasomotor symptoms (including hot flashes or night sweats) more common in the asoprisnil groups
	Phase II [117]	10 mg or 25 mg or placebo for 12 weeks prior to surgery	-	↓25.8% in the 25 mg group versus ↑4.9% in the placebo group		-Headache, nasopharyngitis, nausea, back pain, perioperative complications, and abdominal pain
	Phase III [100] (NCT00152269, NCT00160381)	10 mg or 26 mg or placebo daily for 12 months	66–78% and 83–93% in the 10 mg and 25 mg groups, respectively, versus 3–12% in the placebo grou	↓48% in the 10 mg and ↓63% in the 25 mg groups versus ↑16 % in the placebo group	↓ 28% in the 10 mg and ↓ 39% in the 25 mg group versus ↑ 13 % in the placebo group	-Hot flushes (14% in the 10 mg group versus 7% in the placebo) -Increase in mean endometrial thickness by ~2 mm with asoprisnil -PAECs in 8–19% of patients in the 10 mg group versus 1–4% in the placebo)
Vilaprisan	Phase I [108] (NCT01816815)	0.1 mg, 0.5 mg, 1 mg, 2 mg, 5 mg or placebo daily for 12 weeks	-Non-bleeding rates increased with increasing VPR dose and were 0%, 27.3%, 80%, 100% and 90.9% for 0.1 mg, 0.5 mg, 1 mg, 2 mg, 5 mg groups, respectively, versus 0% in the placebo group -Mean time to first menstrual bleeding after VPR discontinuation was 10.3–25.8 days for doses 1–5 mg and 12.2–14.8 days for doses ≤ 0.5 mg -Follicular growth was not suppressed and minimum average E2 levels remained above 40 pg/ml			-PAECs increased with increasing VPR doses (9%, 60%, 83.3%, 66.7% and 100% for 0.1 mg, 0.5 mg, 1 mg, 2 mg, 5 mg groups at day 84, respectively, versus 10% in the placebo group) -Headache, ovarian cyst, fatigue and abdominal pain were reported by ≥ 5% of all participants
	Phase I [109] (NCT02262663)	0.1 mg, 1 mg, 2 mg or 4 mg daily for 12 weeks	75% in all groups			-Maximum endometrial thickness increased slightly in all treatment groups -PAECs in 70–95% of patients treated with doses ≥ 1mg -Headache, endometrial disorder, ovarian cyst, cervical cyst, pelvic pain, acne, hemorrhagic, nausea, proteinuria present, hot flush, and dizziness in >10% of subjects
	Phase II [110] ASTEROID 1 (NCT02131662)	0.5 mg, 1 mg, 2 mg or 4 mg or placebo daily for 12 weeks (plus 24-week followup)	65%, 85%, 89% and 83% in the 0.5, 1.0, 2.0, and 4.0 mg VPR groups, respectively, versus 9% in the placebo group	↓14.9%–41.4% with VPR versus ↑4.9% with placebo	-	-Ovarian cyst (11.0%), headache (9.7%), and host flush (9.3%) were the most common TEAEs -PAECs in 38.5%, 58.3%, 45.5%, and 33.3% of women in the 0.5, 1.0, 2.0, and 4.0 mg VPR groups, respectively, versus 20% in the placebo group
	Phase II [111] ASTEROID 2 (NCT02465814)	2 mg or placebo or UPA daily for two 12-week treatment courses	-	↓29.9% with VPR and ↓23.8% with UPA treatment, versus ↑6.3% with placebo	-	No serious AEs were reported
	Phase III [112] ASTEROID 3 (NCT03400943)	2 mg or placebo daily for two 12-week treatment courses	83.3% in the VPR group versus 0% in the placebo group			-Serious AEs in 27.8% of the total study population

Mifepristone

Mifepristone (RU-486) is a SPRM initially developed for its anti-glucocorticoid properties. Later, it was found to bind strongly to PRs, with an affinity more than twice that of progesterone itself in human endometrial and myometrial tissues [5,55]. Due to its anti-glucocorticoid and anti-progesterone actions, mifepristone is FDA-approved for managing Cushing’s syndrome and, in combination with misoprostol, for early pregnancy termination [56]. However, its potential therapeutic uses have expanded to include several gynecological conditions, such as endometriosis, adenomyosis, labor induction, and uterine fibroids [57].

Mechanisms of action in uterine fibroids:

The mechanism of action of mifepristone in treating uterine fibroids involves multiple pathways, which collectively inhibit fibroid growth and alleviate symptoms. Mifepristone targets the insulin-like growth factor-1 (IGF-1) signaling pathway by suppressing the ERK1/2 pathway, contributing to fibroid shrinkage [58]. The drug also downregulates TGF-β and the AKT pathway, which are known to support fibroid cell growth and ECM accumulation [53,59]. Mifepristone promotes fibroid cell apoptosis by reducing Bcl-2 levels, thus presenting another therapeutic mechanism [60]. Furthermore, mifepristone reduces ECM synthesis by downregulating ECM-related genes, including COL1A1, fibronectin, and syndecan-1 [61,62].

In fibroid cells, mifepristone has been shown to block the progesterone-induced upregulation of LAT-2, an amino acid transporter with oncogenic properties [63]. The drug also increases the expression of KLF11, a tumor suppressor gene typically downregulated in fibroids, leading to further fibroid regression [64]. Additionally, recent studies have associated mifepristone treatment with upregulated expression of GSTM1, a gene linked to tumor restriction, adding another mechanism to its tumor-suppressive actions [65]. Together, these findings indicate that mifepristone efficiently promotes fibroid regression by regulating genes involved in cell proliferation, ECM deposition, and apoptosis (Fig. 3).

Clinical trials and efficacy in uterine fibroids:

Clinically, mifepristone has been extensively studied for fibroid treatment, particularly at low doses. A 2005 study showed that daily doses of 5 mg and 10 mg mifepristone for 12 months led to significant fibroid shrinkage (48%) and symptom relief (52–53%) without excessive endometrial hyperplasia, although some regrowth occurred after treatment cessation [66]. A similar study in 2008 confirmed that a 5 mg daily dose effectively reduced fibroid size and symptoms, while additional evidence suggested 5 mg as the optimal dose when compared to lower doses [67,68].

A large randomized, double-blind trial conducted over 3 months with 124 patients compared daily 5 mg mifepristone to placebo, showing a 28.5% reduction in fibroid size in the mifepristone group versus a slight increase in the placebo group. Amenorrhea was significantly more common in the mifepristone group (93.1% vs. 4.3%), along with marked improvements in pelvic pain, bleeding, and quality of life [69].

Alternative regimens have also been explored. For example, vaginal mifepristone administration led to fibroid volume reduction and symptom improvement without causing significant endometrial changes [70]. Another study compared daily versus biweekly dosing and found that both regimens resulted in similar efficacy, suggesting a cost-effective alternative [71].

Overall, these clinical findings suggest mifepristone as a promising treatment option for uterine fibroids, particularly for symptom management and preoperative fibroid reduction. With its mechanisms targeting proliferation, apoptosis, and ECM synthesis, mifepristone provides an effective, non-surgical treatment alternative for symptomatic fibroids.

Ulipristal acetate

Ulipristal acetate (UPA) (CDB-2914) is a SPRM approved by the European Medicines Agency (EMA) in 2009 and the Food and Drug Administration (FDA) in 2010 for emergency postcoital contraception as a single dose [5]. UPA functions by binding to PRs, thereby blocking the activation of PR-mediated gene transcription by endogenous P4 [5]. Unlike mifepristone, UPA has minimal effects on glucocorticoid receptors and no known activity on estrogen or mineralocorticoid receptors [72]. Due to its potent anti-progesterone effects, UPA has shown benefits in treating several gynecological conditions, especially uterine fibroids. In 2012, the EMA approved UPA for fibroid management at a daily dose of 5 mg, following positive outcomes from phase III/IV trials [73]. However, reports of severe liver complications led to its withdrawal from the market in 2020, pending further safety assessments [74].

Mechanisms of action in uterine fibroids:

Preclinical studies have demonstrated multiple actions of UPA, including antifibrotic, antiangiogenic, antiproliferative, and proapoptotic effects (Fig. 3). UPA promotes apoptosis in fibroid cells by downregulating the anti-apoptotic protein Bcl-2 while upregulating cleaved caspase-3 and PARP expression. This effect is complemented by reduced proliferation through decreased expression of proliferating cell nuclear antigen (PCNA) [75]. More recent research has shown that UPA induces autophagy in cultured leiomyoma cells, indicated by increased levels of autophagy markers such as LC3-II, p62/SQSTM1, Atg7, and Atg4D [76]. UPA also modulates the PR isoform ratio by upregulating PR-A and downregulating PR-B expression, a mechanism that restricts fibroid growth [42]. Additionally, UPA has antiangiogenic effects, as evidenced by the downregulation of VEGF-A, VEGF-B and adrenomedullin (ADM) in leiomyoma cells, along with their receptors [42,77]. UPA further limits ECM synthesis through the increased activity of matrix metalloproteinases (MMP-1, MMP-2, MMP-3, and MMP-9) and decreased production of tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) in leiomyoma cells, but not in normal myometrium [78–80]. Tinelli et al. identified specific molecular mechanisms of UPA in uterine fibroids, showing that UPA treatment reduced the expression of cofilin, Erk and Src phosphorylation, p27, and ezrin, while not affecting Akt phosphorylation, cyclin D1, or β-catenin levels [81]. These changes may play a role in modulating cytoskeleton remodeling and cell cycle regulation in uterine fibroids.

Additional antifibrotic mechanisms include the inhibition of TGF-β signaling, evidenced by reduced levels of TGF-β3, p-TGFR2, p-Smad2, and p-Smad3 in UPA-treated fibroid tissue [29,30]. UPA also downregulates various profibrotic factors, including activin A, fibronectin, versican, collagen type I and II, procollagen type I, and fibrillin [23,29,30,77,82]. UPA affects osmoregulation in fibroid cells by inhibiting the transcription factor NFAT5 and the osmolyte transporters AKR1B1 and SLC5A3, resulting in decreased production of proteoglycans such as versican, aggrecan, and brevican [83]. Additionally, UPA downregulates ECM structural components, such as integrin subunit beta 4 and tenascin-C, as well as the A-kinase anchoring protein 13 (AKAP13), which influences mechanical signaling in fibroids [84,85].

Clinical trials and efficacy in uterine fibroids:

UPA has been the focus of numerous clinical trials for fibroid treatment. A phase II study compared daily doses of 10 mg and 20 mg UPA to placebo, demonstrating fibroid volume reduction of 17% and 24% in the UPA groups, while the placebo group showed a 7% increase. Amenorrhea was achieved in 20 out of 26 women in the UPA groups, while none of the placebo-treated participants experienced amenorrhea. Quality of life (QoL) scores and hemoglobin levels also significantly improved in the UPA groups. Notably, 9 participants exhibited abnormal liver function tests, and some showed cystic glandular changes in endometrial biopsies [86].

The PEARL trials (I-IV) contributed substantial evidence on efficacy and safety of UPA in treating symptomatic uterine fibroids. In PEARL I, patients were assigned to either 5 mg or 10 mg UPA or placebo daily for 13 weeks preoperatively [87]. UPA at 5 mg and 10 mg decreased fibroid size by 21% and 12%, respectively, while placebo led to a 3% increase. Bleeding control was achieved in 91% of women on 5 mg UPA and 92% on 10 mg, compared to 19% with placebo. Amenorrhea rates were significantly higher in the UPA groups (73% and 82%) compared to placebo (6%). Although some endometrial changes were observed, they were reversible post-treatment [87].

PEARL II compared 5 mg and 10 mg daily UPA to monthly leuprolide acetate injections over 3 months [88]. Uterine bleeding control was achieved in 90% and 98% of women receiving UPA, and 89% with leuprolide acetate. While leuprolide caused greater fibroid shrinkage (47% reduction) compared to UPA (20–22%), it was associated with more hot flashes (40% vs. 10–11% with UPA). UPA, however, had a higher rate of benign endometrial changes, which were reversible post-treatment [88].

PEARL III and its extension examined the long-term efficacy of UPA [21]. Patients received up to four 3-month cycles of 10 mg UPA with intermittent norethisterone acetate (NETA) or placebo for 10 days. Fibroid volume decreased by 45% after the first cycle, and up to 72% after the fourth. Amenorrhea rates reached 79% initially and rose to 90% after multiple cycles. PAECs (benign endometrial changes) were observed but decreased after treatment, indicating UPA’s safety and efficacy over repeated cycles [21].

In PEARL IV, 451 women received two 12-week courses of either 5 mg or 10 mg UPA. By the end of treatment, amenorrhea was achieved in 62% (5 mg) and 73% (10 mg) groups, with fibroid volume reduction of 54% and 58%, respectively [89]. PAECs were more common in the 10 mg group but did not indicate malignancy. The study demonstrated that both doses were effective for symptom relief and fibroid reduction, with no fibroid regrowth observed during follow-up [89].

The PREMYA trial, a large multicenter study, involved 1473 women receiving 5 mg of UPA for 3 months [90]. At the end of the treatment, 60% of participants reported significant symptom improvement, and 38.8% required less invasive surgical interventions. QoL scores remained improved over a 12-month follow-up, suggesting long-lasting effects of UPA treatment [90].

The VENUS I and II trials, conducted primarily among African American women with higher BMI, also confirmed the efficacy of UPA [22,91]. In VENUS I, fibroid volume was reduced by 9.6% and 16.3% in the 5 mg and 10 mg groups, respectively, compared to a 7.2% increase in placebo. Amenorrhea rates were significantly higher in the UPA groups (47.2% and 58.3%) compared to placebo (1.8%). VENUS II demonstrated further fibroid volume reduction during a second treatment cycle, with minor adverse events, such as hot flashes and fatigue, and no endometrial malignancy [22,91].

Other trials and studies on UPA in fibroid treatment:

The MYOMEX trial compared UPA to leuprolide acetate as preoperative treatments before myomectomy. Although leuprolide resulted in greater fibroid reduction and shorter surgery times, UPA was associated with less intense intraoperative blood loss, although it made myomectomies somewhat more challenging. Both pre-treatments led to similar improvements in QoL and bleeding patterns six months post-surgery [92,93]. The MYOMEX-2 trial aims to assess the long-term cost-effectiveness of UPA versus surgery, with results expected soon [94].

The effect of UPA on in vitro fertilization (IVF) outcomes was studied retrospectively, showing a 49% decrease in mean fibroid volume, with improvements in uterine cavity distortion and symptomatic relief. IVF success rates were similar in the UPA-pretreated and control groups, indicating that UPA might optimize uterine conditions for IVF candidates with fibroids [95].

The UCON study compared UPA to the levonorgestrel-releasing intrauterine system in women with fibroids and heavy bleeding. UPA achieved higher amenorrhea rates (64% vs. 25%) and significant QoL improvements without hepatotoxicity or endometrial malignancy, suggesting that it may be more effective in achieving amenorrhea than the levonorgestrel device [96]. Overall, UPA shows efficiency as a SPRM for managing symptomatic uterine fibroids.

Asoprisnil

Asoprisnil (J-867) is a unique SPRM with high specificity for PRs, particularly in the endometrium [97,98]. Unlike other SPRMs, it belongs to the 11β-benzaldoxime substituted steroidal family and demonstrates both agonist and antagonist effects on P4 signaling by recruiting gene coactivators and corepressors when bound to PRs [5,99]. In rabbit uterine cells, PR binding affinity of asoprisnil is approximately three times higher than that of P4, while it exhibits medium affinity for glucocorticoid receptors, low affinity for androgen receptors, and no affinity for estrogen or mineralocorticoid receptors [98]. Initial promising preclinical data led to its development for treating uterine fibroids and endometriosis [98], but concerns over endometrial changes limited its clinical use [100,101].

Mechanisms of action in uterine fibroids:

Preclinical studies have shown that asoprisnil can selectively inhibit fibroid cell growth by downregulating growth factors like TGF-β3, EGF, and IGF-I and their receptors, while sparing normal myometrial cells [44,102]. It also induces apoptosis in fibroid cells, marked by reduced Bcl-2 levels, increased cleaved caspase-3 and PARP expression, and elevated TUNEL-positive cell rates [51,102]. Interestingly, PR-B expression is higher in asoprisnil-treated leiomyoma cells, suggesting that PR-B may contribute to selective actions of asoprisnil on fibroid tissue [51]. Additionally, asoprisnil increases ER-stress proteins and activates apoptosis-related genes such as GADD153, leading to the upregulation of pro-apoptotic proteins Bax and Bak and further reducing Bcl-2 levels [103]. Other studies have shown that asoprisnil activates the TRAIL-mediated apoptotic pathway and suppresses X-linked inhibitor of apoptosis in fibroid cells [104]. Furthermore, asoprisnil reduces ECM production in fibroids by decreasing collagen types I and III, TIMP-1, and TIMP-2, while enhancing the synthesis of ECM-degrading proteins like MMP-1 and MT1-MMP [32].

Clinical trials and efficacy in uterine fibroids:

In a Phase II clinical trial with 129 women, asoprisnil at doses of 5, 10, or 25 mg daily significantly reduced fibroid-related bleeding and induced amenorrhea in a dose-dependent manner, with the highest efficacy observed in the 25 mg group, which also achieved a 36% reduction in fibroid volume [105]. However, endometrial biopsies revealed non-physiologic changes in over 50% of women on asoprisnil, and some patients developed asymptomatic ovarian cysts. Another study noted structural changes in the endometrium and decreased mitotic activity in the fibroid tissue of asoprisnil-treated women [106].

A pooled analysis of two Phase III trials in 907 women confirmed the efficacy of asoprisnil in reducing uterine bleeding, fibroid volume, and improving quality of life, with effects maintained over a 6-month follow-up [100]. However, non-physiologic endometrial changes occurred frequently in asoprisnil-treated women, and several cases of breast and endometrial cancer were observed during an extension study, raising concerns about the long-term safety of the drug [101]. These findings suggest that while asoprisnil is effective in reducing fibroid symptoms, its endometrial safety profile warrants caution, limiting its clinical utility.

Vilaprisan

Vilaprisan (BAY 1002670) is a novel SPRM with potent antagonistic activity on PRs, estimated to be stronger than mifepristone or UPA [107]. Although highly selective for PRs, preclinical studies suggest it has moderate binding affinity to GR and low affinity to AR [107]. Given its strong anti-progesterone effect, vilaprisan entered clinical trials in 2017 to assess its efficacy and safety for treating uterine fibroids, demonstrating promising early-phase results.

Clinical trials and efficacy in uterine fibroids:

In initial Phase I trials, the effects of vilaprisan on menstrual cycle regulation were evaluated [108]. A randomized, placebo-controlled study with 73 healthy women administered doses of 0.1–5 mg daily for 12 weeks. The 2 mg dose showed significantly increased non-bleeding rates, with menstrual bleeding resuming in all women within 52 days of stopping treatment. Follicular growth was unaffected, while estrogen levels (E2) stayed above 40 pg/mL. Some participants on higher doses, particularly 5 mg, developed PAECs and reported adverse events (AEs) like headache, ovarian cyst formation, fatigue, and abdominal pain [108].

Another Phase I study evaluated the impact of vilaprisan on ovarian function [109]. Seventy women received doses between 0.5–4 mg daily for 12 weeks. Ovulation was inhibited in over 80% of women taking 1 mg or more, and E2, P4, FSH, and LH levels were markedly decreased, although follicular growth continued. Amenorrhea rates peaked at 2 mg, and endometrial thickness increased but normalized after treatment ended. As in the first study, PAECs occurred in a dose-dependent manner, affecting up to 95% of women on doses of 1 mg or more [109].

With ovarian function effects established, Phase II and III trials assessed the efficacy and safety of vilaprisan in women with fibroids and heavy menstrual bleeding. The ASTEROID 1 trial, a Phase II study of 300 women, tested daily doses of 0.5–4 mg over 12 weeks, followed by a 24-week follow-up [110]. Complete cessation of bleeding occurred in up to 60% of patients in the vilaprisan groups, while only 1.7% in the placebo group achieved this. Amenorrhea was achieved in over 83% of patients taking at least 1 mg, and median menstrual blood loss dropped to 0 mL in all vilaprisan groups. Fibroid volume decreased significantly (14.9%–41.4%) in a dose-dependent manner, with most women reporting improved symptoms. Common AEs included ovarian cysts, headache, and hot flashes, with no significant difference in AE frequency across groups. These findings suggested vilaprisan is effective in controlling fibroid-related bleeding, with 2 mg identified as the optimal dose [110].

The ASTEROID 2 trial compared vilaprisan to UPA and placebo in 100 women with fibroids and heavy menstrual bleeding [111]. At week 12, 62.9% of vilaprisan-treated women achieved complete bleeding cessation compared to none in the placebo group. Fibroid volume decreased by 29.9% with vilaprisan and 23.8% with UPA, while placebo saw a slight increase. The study indicated the comparable efficacy of vilaprisan to UPA in reducing bleeding and fibroid size, with no serious safety concerns [111].

Finally, the ASTEROID 3 trial, a Phase III study, assigned 75 women to 2 mg vilaprisan or placebo over two 12-week courses [112]. Amenorrhea was achieved in 83.3% of vilaprisan-treated patients compared to none in the placebo group. Vilaprisan effectively reduced menstrual blood loss, and although serious AEs were reported in 27.8% of participants, they did not differ significantly between groups. These results highlight the potential of vilaprisan for rapid bleeding reduction, but further trials are needed to assess long-term safety and efficacy [112].

CONCLUSIONS AND FUTURE PERSPECTIVES

Uterine fibroids are a complex, multifactorial disease involving various pathophysiological mechanisms that contribute to fibroid cell growth and survival. P4 plays a central role in fibroid pathogenesis, activating pathways that promote leiomyoma cell proliferation, upregulating antiapoptotic factors, and disrupting vascular patterns. P4 also drives excessive ECM synthesis while inhibiting ECM resorption by suppressing matrix metalloproteinases. The recognition of P4 as a key mediator in fibroid growth has led to interest in SPRMs for treatment. Mifepristone has shown efficacy in reducing fibroid size and symptoms, but its long-term use is limited by endometrial proliferation concerns. Asoprisnil effectively controls bleeding but raises safety concerns with prolonged use. Ulipristal acetate, approved by the EMA for fibroid management, was effective in reducing fibroid size and bleeding but was suspended by the European Medicines Agency (EMA) in February 2018 due to reports of liver toxicity. In May of 2018 the EMA issued restrictive measures, and a Pharmacovigilance Risk Assessment Committee made recommendations to reduce the risks in May 2018 to resume treatment of premenopausal women with ulipristal acetate for whom surgical procedures are inappropriate or unsuccessful suspended. Ulipristal acetate was not approved for use in the U.S. Vilaprisan, a newer SPRM, has shown promise in recent trials for controlling bleeding and reducing fibroid volume. However, Bayer Healthcare Pharmaceuticals issued a pause in December 2018 based on preliminary findings from long-term rodent carcinogenicity studies. Despite the absence of findings indicating safety concerns for humans, further development was suspended due to a change in development plans. Though no SPRMs are approved for treatment of fibroids in the U.S., the class of compounds demonstrated rapid reduction in bleeding and a remarkably durable reduction in fibroid size. It is possible that new compounds could be developed, or intermittent treatment regimens could be devised with existing SPRMs to serve as a nonsurgical option for this complex disease.