The effect of a progesterone receptor modulator on the endometrium and breast in premenopausal women

Abstract

Background: The levonorgestrel intrauterine system, LNG-IUS 52 mg, is a highly effective and cost-effective contraceptive, entailing minimal patient effort. Irregular bleeding patterns are common during the first months of use and constitutes one of the main reasons for discontinuation. Up to date, no standard treatment approach has been proven to resolve this problem. Mifepristone is a synthetic steroid hormone, acting mainly as an inhibitor of the progesterone receptor (PR) thereby preventing the effect of progesterone. Administration of mifepristone has been proven beneficial in numerous fields in reproductive medicine and exerts various effects depending on the dosage and stage of the menstrual cycle at treatment. Administration of low daily doses of mifepristone results in anovulation and endometrial suppression with subsequent amenorrhea. Continuous treatment with PRMs cause endometrial alterations previously thought to be similar to unopposed estrogen exposure, given the PR antagonistic effects of PRMs. These alterations are today recognized as progesterone receptor modulator associated changes (PAEC). They are considered to be benign and dissolves with the cessation of treatment. The molecular alterations resulting in their development are still unknown. The mechanism of action of steroidal hormones on breast tissue remains largely unidentified. Epidemiological studies show a positive correlation between number of menstrual cycle exposure and hormone therapy to the risk of breast cancer. While this increased risk has been believed to be mainly caused by estrogen, a growing body of literature suggest progesterone and progestins to play a central role. PRMs can be used as a tool to study the effects of progesterone and holds potential to prevent breast epithelial cell proliferation.

Aim: The overall aim of this thesis is to explore the effects of the PRM mifepristone on the endometrium and on human breast tissue in premenopausal women. The specific objectives were to assess whether inducing amenorrhea with mifepristone, prior to placement of the LNG-IUS, could reduce the bleeding irregularities during the first months of use. Another objective was to evaluate the endometrial morphology after continuous treatment with mifepristone following insertion of the LNG-IUS, without prior endometrial shedding. Furthermore, we sought to explore how mifepristone alters the transcriptomic landscape in human beast in vivo and the epigenetic alterations observed in the breast tissue following PRM treatment.

Materials, methods and results: Study I was a prospective, randomized, placebo controlled, double-blind trial including healthy women with regular menstrual cycles opting for the LNG-IUS 52 mg for contraceptive purposes. Fifty-eight women were randomized whereof 29 to the mifepristone and 29 to the comparator group. Study participants received mifepristone, 50 mg every other day or a comparator. The pretreatment period with mifepristone was 2 months, followed by the LNG-IUS insertion. Women kept bleeding diaries as per instruction for the pretreatment period and until 6 months after placement of the device. After removing drop outs and exclusions, 19 women in the mifepristone and 19 in the comparator arm contributed to the final analysis. Bleeding diary data were analyzed as rates of bleeding and spotting days (B/S%) per treatment cycle. The results showed a significant reduction of B/S% during the pretreatment period in the mifepristone arm compared to placebo. Following insertion of the device, no statistical difference could be seen between the two groups. Women in Study II originated from Study I. Endometrial biopsies were retrieved at baseline, prior to the pretreatment period with mifepristone or the comparator. A second biopsy was retrieved at 3 months following LNG-IUS placement, with the IUS in situ. Nine paired biopsies from the mifepristone and 8 from the comparator group, contributed to the final analysis. The specimens were analyzed by an expert pathologist who was blinded to the treatment. All baseline biopsies where benign. The second biopsies were all benign and showed, as expected, changes due to progestin effect on the endometrium. There was no presence of PAEC. Participants in Study III originated from Study I. Core needle breast biopsies were collected at baseline and after 2 months treatment with mifepristone or the comparator. Paired biopsies from 16 women in the mifepristone group contributed to the analysis. The changes on mRNA expression level at baseline compared to after mifepristone treatment were screened using RNA sequencing. Functional annotation and pathway enrichment analysis of the differentially expressed genes (DEGs) revealed genes mainly involved in extracellular matrix (ECM) remodeling. In Study IV, patient cohorts and databases were used to generate and validate a breast tissue specific epigenetic index. That index was subsequently used to assess breast tissue samples from three clinical trials, including Study I. Based on the results from this specific epigenetic index, PRM treatment could exhibit favorable results in the mammary gland from healthy women as well as women with increased risk for developing breast cancer.

Conclusion: The applied mifepristone treatment regimen could not demonstrate any significant improvement in bleeding disturbances following placement of the LNG-IUS compared to placebo. Continuous treatment with mifepristone and subsequent LNG-IUS insertion without prior endometrial shedding, could represent a safe alternative regarding PAEC endometrial safety. Transcriptomic alterations in the breast after treatment with mifepristone revealed pathways mainly involved in ECM remodeling. Furthermore, epigenetic and genetic alterations in the breast following PRM treatment seem promising and suggestive of further investigations regarding the potential beneficial effects of these compounds in the prevention of breast cancer.