Is home-based-monitoring of ovulation to time frozen embryo transfer a cost-effective alternative for hospital-based-monitoring of ovulation? Study protocol of the multicentre, non-inferiority Antarctica-2 randomised controlled trial

STUDY QUESTIONS The objective of this trial is to compare the effectiveness and costs of true natural cycle (true NC-) frozen embryo transfer (FET) using urinary LH tests to modified NC-FET using repeated ultrasound monitoring and ovulation trigger to time FET in the natural cycle. Secondary outcomes are the cancellation rates of FET (ovulation before hCG or no dominant follicle, no ovulation by LH urine test, poor embryo survival), pregnancy outcomes (miscarriage rate, clinical pregnancy rates, multiple ongoing pregnancy rates, live birth rates, costs) and neonatal outcomes (including gestational age, birthweight and sex, congenital abnormalities or diseases of babies born). WHAT IS KNOWN ALREADY FET is at the heart of modern IVF. To allow implantation of the thawed embryo, the endometrium must be prepared either by exogenous estrogen and progesterone supplementation (artificial cycle (AC)-FET) or by using the natural cycle to produce endogenous oestradiol before and progesterone after ovulation to time the transfer of the thawed embryo (NC-FET). During a NC-FET, women visit the hospital repeatedly and receive an ovulation trigger to time FET (i.e. modified (m)NC-FET or hospital-based monitoring). From the woman’s point of view, a more natural approach using home-based monitoring of the ovulation with LH urine tests to allow a natural ovulation to time FET may be desired (true NC-FET or home-based monitoring). STUDY DESIGN, SIZE, DURATION This is a multicentre, non-inferiority prospective randomised controlled trial design. Consenting women will undergo one FET cycle using either true NC-FET or mNC-FET based on randomisation. PARTICIPANTS/MATERIALS, SETTING, METHODS Based on our sample size calculation the study group will consist of 1464 women between 18 and 45 years old who are scheduled for FET. Women with anovulatory cycles, women who need ovulation induction and women with a contra indication for pregnancy will be excluded. The primary outcome is ongoing pregnancy. Secondary outcomes are cancellation rates of FET, pregnancy outcomes (including miscarriage rate, clinical pregnancy, multiple pregnancy rate and live birth rate). Costs will be estimated by counting resource use and calculating unit prices. STUDY FUNDING/COMPETING INTEREST(S) The study received a grant from The Dutch Organisation for Health Research and Development (ZonMw 843002807; www.zonmw.nl). ZonMw has no role in the design of the study, collection, analysis, and interpretation of data or writing of the manuscript. Dr. Broekmans reports personal fees from member of the external advisory board for Merck Serono, grants from Research support grant Merck Serono, outside the submitted work;. Dr. Cantineau reports and Unrestricted grant of Ferring B.V. to the Center for Reproductive medicine, no personal fee. Author up-to-date on Hyperthecosis. Congress meetings 2019 with Ferring B.V. and Theramex B.V. Dr. Goddijn reports Department research and educational grants from Guerbet, Merck and Ferring (location VUMC) outside the submitted work. Dr. Groenewoud reports personal fees from Titus Health Care, outside the submitted work; Dr. Lambalk reports grants from Ferring, grants from Merck, from Guerbet, outside the submitted work. The other authors have none to declare. TRIAL REGISTRATION NUMBER Dutch Trial Register (Trial NL6414 (NTR6590), https://www.trialregister.nl/). TRIAL REGISTRATION DATE 23 July 2017 DATE OF FIRST PATIENT’S ENROLMENT 10 April 2018

A 3D endometrial organotypic model simulating the acute inflammatory decidualisation initiation phase with epithelial induction of the key endometrial receptivity marker, integrin αVβ3

STUDY QUESTION Is it possible to develop a simplified physiological in vitro system representing the key cell-types associated with a receptive endometrial phenotype? SUMMARY ANSWER We present a new concept to investigate endometrial receptivity, with a 3D organotypic co-culture model to simulate an early and transient acute autoinflammatory decidual status that resolves in the induction of a receptive endometrial phenotype. WHAT IS KNOWN ALREADY Embryo implantation is dependent on a receptive uterine environment. Ovarian steroids drive post-ovulation structural and functional changes in the endometrium, which becomes transiently receptive for an implanting conceptus, termed the ‘window of implantation’, and dysregulation of endometrial receptivity is implicated in a range of reproductive, obstetric, and gynaecological disorders and malignancies. The interactions that take place within the uterine microenvironment during this time are not fully understood, and human studies are constrained by a lack of access to uterine tissue from specific time-points during the menstrual cycle. Physiologically relevant in vitro model systems are therefore fundamental for conducting investigations to better understand the cellular and molecular mechanisms controlling endometrial receptivity. PARTICIPANTS/MATERIALS, SETTING, METHODS An endometrial stromal cell (ESC) line, and endometrial epithelial cells (EECs) isolated from uterine biopsy tissue and expanded in vitro by conditional reprogramming, were used throughout the study. Immunocytochemical and flow cytometric analyses were used to confirm epithelial phenotype following conditional reprogramming of EECs. To construct an endometrial organotypic co-culture model, ESCs were embedded within a 3D growth factor-reduced Matrigel structure, with a single layer of conditionally reprogrammed EECs seeded on top. Cells were stimulated with increasing doses of medroxyprogesterone acetate, cAMP and estradiol, in order to induce ESC decidual transformation and endometrial receptivity. Decidual response and the induction of a receptive epithelial phenotype were assessed by immunocytochemical detection and quantitative in-cell western analyses, respectively. MAIN RESULTS AND THE ROLE OF CHANCE A transient upregulation of the IL-33 receptor protein, ST2L, was observed in ESCs, indicating a transient autoinflammatory decidual response to the hormonal stimulation, known to induce receptivity gene expression in the overlying epithelium. Hormonal stimulation increased the EEC protein levels of the key marker of endometrial receptivity, integrin αVβ3 (n = 8; *P <0.05; ***P < 0.0001). To our knowledge, this is the first demonstration of a dedicated endometrial organotypic model, that has been developed to investigate endometrial receptivity, via the recapitulation of an early decidual transitory acute autoinflammatory phase and induction of an epithelial phenotypic change, to represent a receptive endometrial status. LIMITATIONS, REASONS FOR CAUTION This simplified in vitro ESC-EEC co-culture system may be only partly representative of more complex in vivo conditions. WIDER IMPLICATIONS OF THE FINDINGS The 3D endometrial organotypic model presented here may offer a valuable tool for investigating a range of reproductive, obstetric, and gynaecological disorders, to improve outcomes for assisted reproductive technologies, and for the development of advances in contraceptive methods. STUDY FUNDING/COMPETING INTEREST(S) This work was supported in part by an MRC Centre Grant (project reference MR/N022556/1). RF was the recipient of a Moray Endowment award and a Barbour Watson Trust award. C-JL is a Royal Society of Edinburgh Personal Research Fellow, funded by the Scottish Government. The authors have no conflicts of interest to declare.

A concise infertility work-up results in fewer pregnancies

STUDY QUESTION Is pregnancy success rate after a concise infertility work-up the same as pregnancy success rate after the traditional extensive infertility work-up? SUMMARY ANSWER The ongoing pregnancy rate within a follow-up of 1 year after a concise infertility work-up is significantly lower than the pregnancy success rate after the traditional and extensive infertility work-up. WHAT IS KNOWN ALREADY Based on cost-effectiveness studies, which have mainly focused on diagnosis, infertility work-up has become less comprehensive. Many centres have even adopted a one-stop approach to their infertility work-up. STUDY DESIGN, SIZE, DURATION We performed a historically controlled cohort study. In 2012 and 2013 all new infertile couples (n = 795) underwent an extensive infertility work-up (group A). In 2014 and 2015, all new infertile couples (n = 752) underwent a concise infertility work-up (group B). The follow-up period was 1 year for both groups. Complete follow-up was available for 99.0% of couples in group A and 97.5% in group B. PARTICIPANTS/MATERIALS, SETTING, METHODS The extensive infertility work-up consisted of history taking, a gynaecological ultrasound scan, semen analysis, ultrasonographic cycle monitoring, a timed postcoital test, a timed progesterone and chlamydia antibody titre. A hysterosalpingography (HSG) was advised routinely. The concise infertility work-up was mainly based on history taking, a gynaecological ultrasound scan and semen analysis. A HSG was only performed if tubal pathology was suspected or before the start of IUI. Laparoscopy and hormonal tests were only performed if indicated. Couples were treated according to the diagnosis with either expectant management (if the Hunault prognostic score was >30%), ovulation induction (in case of ovulation disorders), IUI in natural cycles (in case of cervical factor), IUI in stimulated cycles (if the Hunault prognostic score was <30%) or IVF/ICSI (in case of tubal factor, advanced female age, severe male factor and if other treatments remained unsuccessful). The primary outcomes were time to pregnancy and the ongoing pregnancy rates in both groups. The secondary outcomes were the number of investigations, the distribution of diagnoses made, the first treatment (started) after infertility work-up and the mode of conception. MAIN RESULTS AND THE ROLE OF CHANCE The descriptive data, such as age, duration of infertility, type of infertility and lifestyle habits, in both groups were comparable. In group A, more than twice the number of infertility investigations were performed, compared to group B. An HSG was made less frequently in group B (33% versus 42%) and at a later stage. A Kaplan–Meier curve shows a shorter time to pregnancy in group A. Also, a significantly higher overall ongoing pregnancy rate within a follow-up of 1 year was found in group A (58.7% versus 46.8%, respectively, P < 0.001). In group A, more couples conceived during the infertility work-up (14.7% versus 6.5%, respectively, P < 0.05). The diagnosis cervical infertility could only be made in group A (9.3%). The diagnosis unexplained infertility differed between groups, at 23.5% in group A and 32.2% in group B (P < 0.001). LIMITATIONS, REASONS FOR CAUTION This was a historically controlled cohort study; introduction of bias cannot be ruled out. The follow-up rate was similar in the two groups and therefore could not explain the differences in pregnancy rate. WIDER IMPLICATIONS OF THE FINDINGS Re-introduction of an extensive infertility work-up should be considered as it may lead to higher ongoing pregnancy rates within a year. The therapeutic effects of HSG and timing of intercourse may improve the fertility chance. This finding should be verified in a randomized controlled trial. STUDY FUNDING/COMPETING INTEREST(S) No funding was obtained for this study. No conflicts of interest were declared. TRIAL REGISTRATION NUMBER N/A.

Anti-adhesion Gel versus No gel following Operative Hysteroscopy prior to Subsequent fertility Treatment or timed InterCourse (AGNOHSTIC), a randomised controlled trial: protocol

STUDY QUESTIONS Does the application of anti-adhesion gel, compared to no gel, following operative hysteroscopy to treat intrauterine pathology in women wishing to conceive increase the chance of conception leading to live birth? WHAT IS KNOWN ALREADY Intrauterine adhesions (IUAs) following operative hysteroscopy may impair reproductive success in women of reproductive age. Anti-adhesion barrier gels may decrease the occurrence of IUAs, but the evidence on their effectiveness to improve reproductive outcomes is sparse and of low quality. STUDY DESIGN, SIZE, DURATION This multicentre, parallel group, superiority, blinded and pragmatic randomised controlled trial is being carried out in seven participating centres in Belgium. Recruitment started in April 2019. Women will be randomly allocated to treatment with anti-adhesion gel (intervention group) or no gel (control group). Sterile ultrasound gel will be applied into the vagina as a mock-procedure in both treatment arms. The patient, fertility physician and gynaecologist performing the second-look hysteroscopy are unaware of the allocated treatment. Power analysis, based on a target improvement of 15% in conception leading to live birth using anti-adhesion gel, a power of 85%, a significance level of 5%, and a drop-out rate of 10%, yielded a number of 444 patients to be randomised. The baseline rate of conception leading to live birth in the control group is expected to be 45%. PARTICIPANTS/MATERIALS, SETTING, METHODS Women of reproductive age (18–47 years), wishing to conceive (spontaneously or by fertility treatment) and scheduled for operative hysteroscopy to treat intrauterine pathology (endometrial polyps, myomas with uterine cavity deformation, uterine septa, IUAs or retained products of conception) are eligible for recruitment. Women may try to conceive from 3 to 6 weeks after receiving allocated treatment with follow-up ending at 30 weeks after treatment. If the woman fails to conceive within this timeframe, a second-look hysteroscopy will be scheduled within 2–6 weeks to check for IUAs. The primary endpoint is conception leading to live birth, measured at 30 weeks after randomisation. The secondary endpoints are time to conception, clinical pregnancy, miscarriage and ectopic pregnancy rates, measured at 30 weeks after receiving allocated treatment. The long-term follow-up starts when the patient is pregnant and she will be contacted every trimester. STUDY FUNDING/COMPETING INTEREST(S) This work is funded by the Belgian Healthcare Knowledge Centre (KCE). The anti-adhesion gel is supplied at no cost by Nordic Pharma and without conditions. Dr. Tomassetti reports grants and non-financial support from Merck SA, non-financial support from Ferring SA, personal fees and non-financial support from Gedeon-Richter, outside the submitted work. None of the other authors have a conflict of interest.