Progesterone receptor modulators in contraception : clinical implications of ovarian and endometrial effects

Abstract: Background Emergency contraception (EC) offers a last chance to prevent an unwanted pregnancy after unprotected sexual intercourse (UPSI) but before pregnancy occurs, for example after contraceptive failure, non-use of contraception or after rape. Progesterone receptor modulators (PRMs) exert their effect through acting as partial antagonists and partial agonists of the progesterone receptor (PR) and have proven effective for EC in low doses by postponing or inhibiting ovulation if it was taken preovulatory in the menstrual cycle. They further affect the endometrium in a dose-dependent manner and high doses inhibit endometrial development and affect embryo implantation. There are concerns regarding thickening of the endometrium and the specific PRM associated endometrial changes (PAEC) that long-term PRM treatment can cause. However, there is no knowledge of the underlying molecular basis of this condition. It is further not known if the PRM ulipristal acetate (UPA) used as EC could interfere with the contraceptive effect of a regular combined oral contraceptive pill (COCP) initiated immediately after UPA intake or if UPA in the dose used for EC affects endometrial receptivity and embryo implantation. Low doses of mifepristone have not been explored regarding its effect on the embryo implantation process. Aim The overall aim of this thesis was to explore the effects of PRMs mifepristone and UPA when used for EC and further develop them for contraceptive purposes and to provide women with evidence-based information and advice on mechanisms of action of PRMs. The specific objectives were to investigate the clinical implications of their effects on ovaries and endometrium. This included short-term effects on endometrial receptivity and the embryo implantation process as well as long-term endometrial effects and further possible interaction with regular hormonal contraception. Materials, Methods and Results Study I was a multicenter, double-blind, randomized, placebo-controlled trial in which 76 healthy female volunteers between the ages of 18 and 35 with regular menstrual cycles were randomized to receive either 30 mg UPA or placebo at mid-cycle when the dominant ovarian follicle was ≥ 13 mm, followed by intake of a common COCP during 21 subsequent days. Hormonal measurements and transvaginal ultrasonography were performed regularly to assess ovarian activity. Ovarian quiescence was achieved after 7 days of COCP intake for most women, however in some women it took up to 14 days, irrespective of UPA administration. The proportion of women who ovulated in the study was approximately 30%, similar in both groups, which could be explained by follicle size at inclusion. Study II and III were exploratory studies on the human embryo implantation process and endometrial receptivity markers after treatment with UPA in an EC dose (Study II) and mifepristone in two different low concentrations (Study III) using a 3D human endometrial in vitro cell culture model. Endometrium was collected from proven fertile volunteers with regular menstrual cycles at cycle day LH+4 and endometrial stromal and epithelial cells were isolated and 3D constructs developed. Viable human blastocyst stage embryos, donated from couples that underwent IVF, were randomly allocated to the different treatment groups (UPA 200ng/ml ≈0.4μM, mifepristone 0.05μM, mifepristone 0.5μM and control). When the epithelial cells had grown into a confluent layer, the embryos were placed on top of the culture and further co-cultured for 5 days. The embryo attachment rates in different groups were: 5/10 in UPA, 4/10 in mifepristone 0.05μM, 0/8 in mifepristone 0.5μM and 7/10 in the control group. Some of the known endometrial receptivity markers in Study II were altered in the UPA group but most were unaffected. On the other hand, most of the receptivity markers examined in Study III were altered, with both the concentrations of mifepristone. However, they exerted functionally different effects on embryo implantation in a dose-dependent manner. Study IV was an exploratory study in which endometrial samples had been collected from fourteen healthy, pre-menopausal women, randomized to receive mifepristone during 3 months prior to surgical intervention of uterine leiomyoma in a previous placebo controlled trial. Endometrial biopsies obtained during surgery after the treatment period were assessed for the occurrence of PAEC and further explored regarding gene and protein expression in endometrium with PAEC compared to non- PAEC after mifepristone treatment. Methods used included histological evaluation, microarray analysis, Ingenuity Pathway Analysis, real-time PCR, protein extraction and mass spectrometry. Three genes relevant to functions and diseases of the uterus were upregulated in endometrium with PAEC; ADAM12, THY1 and TN-C and 25 different proteins were upregulated and five downregulated in endometrium with PAEC. Conclusion UPA-EC does not interfere with the ability of a COCP to induce ovarian quiescence when treatment is quickstarted immediately after EC intake and the treatment is safe and tolerable. Further, UPA in a concentration corresponding to EC dose does not inhibit embryo implantation, nor does a very low mifepristone concentration of 0.05μM, whereas mifepristone in a concentration of 0.5μM effectively inhibited attachment of the embryo to the endometrial construct in vitro. The specific morphological features of PAEC displayed after 3 months of mifepristone treatment may be explained by the altered expression of molecules affecting tissue architecture and extracellular matrix, however these molecules were not involved in endometrial cancer-signaling pathways based on IPA knowledge base.