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Background Tubal ectopic pregnancies can cause significant morbidity or even death. Current treatment is with methotrexate or surgery. However, methotrexate treatment can fail in approximately 30% of women. Gefitinib, an epidermal growth factor receptor inhibitor, may improve the effects of methotrexate. We assessed the efficacy of administering oral gefitinib with methotrexate, versus methotrexate alone, to treat a tubal ectopic pregnancy. Objectives To test the hypothesis a combination of gefitinib with methotrexate can increase resolution of stable tubal ectopic pregnancy without the need for surgery, compared with methotrexate alone. Design A randomised, double-blind, placebo-controlled, multicentre, superiority trial. Setting Fifty UK hospitals. Participants A target of 328 women with a stable, tubal ectopic pregnancy. Intervention Women were randomised to combination of methotrexate and gefitinib or methotrexate and placebo. All participants received a single intramuscular dose of methotrexate 50 mg/m2 and were randomised in a 1:1 ratio of oral gefitinib (250 mg daily for 7 days) or placebo. Main outcome measures The primary outcome was surgical intervention for resolution of ectopic pregnancy. Secondary outcomes were the need for an additional dose of methotrexate, time to resolution of the ectopic pregnancy, number of treatment-associated hospital visits, safety and tolerability, acceptability of treatment and return to menses. Results Between 2 November 2016 and 6 October 2021, 328 women were randomly allocated to methotrexate and gefitinib (n = 165) or methotrexate and placebo (n = 163). Three women in the placebo group withdrew. Surgical intervention occurred in 30% (50/165) of the gefitinib group and in 29% (47/160) of the placebo group (adjusted risk ratio 1.15, 95% confidence interval 0.85 to 1.58; adjusted risk difference −0.01, 95% confidence interval −0.10 to 0.09; p = 0.37). Without surgical intervention, median time to resolution was 28.0 days in the gefitinib group and 28.0 days in the placebo group (subdistribution hazard ratio 1.03, 95% confidence interval 0.75 to 1.40). The need for additional methotrexate doses, number of additional hospital visits, participant acceptability, time to return of menses and serious adverse events were similar in both groups. Diarrhoea and rash were more common in the gefitinib group. Conclusions The addition of gefitinib to standard medical management with methotrexate to treat tubal ectopic pregnancy is not clinically effective as it does not reduce subsequent surgical intervention and is associated with higher rates of reported symptoms than placebo. Limitations We were unable to investigate how different gefitinib doses or modes of delivery would impact on the results. Future work Questions that remain unaddressed relate to the use of methotrexate and gefitinib combination treatment for other extrauterine and uterine ectopic pregnancy, such as caesarean scar pregnancies, or in the management of choriocarcinoma. Trial registration This trial is registered as ISRCTN 67795930 and EudraCT 2015-005013-76. Funding This project was funded by the National Institute for Health and Care Research (NIHR) Efficacy and Mechanism Evaluation (EME) programme and will be published in full in Efficacy and Mechanistic Evaluation; Vol. 10, No. 1. The gefitinib and placebo were supplied by Astra Zeneca. See the NIHR Journals Library website for further project information. Plain language summary What was the question? A tubal ectopic pregnancy is where a fertilised egg is not growing in the womb. The pregnancy cannot be saved and the woman is at risk of losing her fallopian tube and if this pregnancy is left to grow can even die. Current treatment is with methotrexate or surgery. An operation can happen because either the ectopic pregnancy has ruptured and caused internal bleeding, the medical treatment has not worked and the ectopic pregnancy needs to be removed or the patient can chose to have an operation. However, methotrexate treatment can fail in approximately 30% of women. We carried out research to see if the addition of a new drug (gefitinib, a drug used for lung cancer) to methotrexate could lower the number of women needing an operation to remove their ectopic pregnancy. What did we do? We involved 328 women with a stable tubal ectopic pregnancy, who were being treated medically with methotrexate, and randomly assigned them to have methotrexate alone or a combination of methotrexate and gefitinib. The gefitinib was taken in tablet form for 7 days, and the methotrexate was given as an injection. We followed the women up in line with their clinical care until their ectopic pregnancy resolved or they had surgery to remove the ectopic pregnancy. What did we find? The addition of gefitinib to methotrexate did not reduce the number of women who required surgery to remove their ectopic pregnancy. More women taking gefitinib experienced side effects, such as a facial rash or diarrhoea. What does this mean? Treatment with methotrexate remains the only drug treatment option for ectopic pregnancy. More research is needed. Scientific summary Background Tubal ectopic pregnancy (EP) can cause significant morbidity or even death. Current treatment is with methotrexate (MTX) or surgery. However, MTX treatment can fail in approximately 30% of women. Preclinical studies have shown that tubal implantation sites express high levels of epidermal growth factor receptor (EGFR) and that gefitinib (an EGFR antagonist) augments MTX-induced regression of pregnancy-like tissue. Clinical evidence from uncontrolled phase I and II trials has raised the possibility that a combination of MTX and gefitinib could be a more effective medical treatment than MTX alone to treat stable tubal EP. Objectives To test the hypothesis, a combination of gefitinib with MTX can increase resolution of stable tubal EP without the need for surgery, compared with MTX alone. Design A randomised, double-blind, placebo-controlled, multicentre, superiority trial. Setting This trial took place in 50 hospitals in the UK. Participants A target of 328 women with a stable, tubal EP. Intervention Participants were randomly assigned in a 1:1 ratio to receive either gefitinib and MTX or matched placebo and MTX with the use of minimisation to balance trial-group assignments according to baseline human chorionic gonadotropin levels (

STUDY QUESTION What is the capacity of the change between Day 1 and Day 4 post-treatment serum human chorionic gonadotropin (hCG) levels for predicting single-dose methotrexate treatment success in tubal ectopic pregnancy? SUMMARY ANSWER Any fall in Days 1–4 serum hCG signified an 85% (95% CI 76.8–90.6) likelihood of treatment success for women with tubal ectopic pregnancy (initial hCG of ≥1000 and ≤5000 IU/l) managed with single-dose methotrexate. WHAT IS KNOWN ALREADY For those with tubal ectopic pregnancy managed by single-dose methotrexate, current guidelines advocate intervention if Days 4–7 hCG fails to fall by >15%. The trajectory of hCG over Days 1–4 has been proposed as an early indicator that predicts treatment success, allowing early reassurance for women. However, almost all prior studies of Days 1–4 hCG changes have been retrospective. STUDY DESIGN, SIZE, DURATION This was a prospective cohort study of women with tubal ectopic pregnancy (pre-treatment hCG of ≥1000 and ≤5000 IU/l) managed with single-dose methotrexate. The data were derived from a UK multicentre randomized controlled trial of methotrexate and gefitinib versus methotrexate and placebo for treatment of tubal ectopic pregnancy (GEM3). For this analysis, we include data from both treatment arms. PARTICIPANTS/MATERIALS, SETTING, METHODS Participants were categorized according to single-dose methotrexate treatment success or failure. Treatment success for this analysis was defined as complete and uneventful resolution of tubal ectopic pregnancy to serum hCG <30 IU/l following single-dose methotrexate treatment without additional treatment. Patient characteristics of the treatment success and failure groups were compared. Changes in Days 1–4, 1–7, and 4–7 serum hCG were evaluated as predictors of treatment success through receiver operating characteristic curve analysis. Test performance characteristics were calculated for percentage change ranges and thresholds including optimal classification thresholds. MAIN RESULTS AND THE ROLE OF CHANCE A total of 322 women with tubal ectopic pregnancy were treated with single-dose methotrexate. The overall single-dose methotrexate treatment success rate was 59% (n = 189/322). For any fall in serum hCG on Days 1–4, likelihood ratios were >3, while for any fall of serum hCG >20% on Days 1–7, likelihood ratios reached 5. Any rise of serum hCG on Days 1–7 and 4–7 strongly reduced the chance of success. Any fall in Days 1–4 hCG predicted single-dose methotrexate treatment success with a sensitivity of 58% and specificity 84%, resulting in positive and negative predictive values of 85% and 57%, respectively. Any rise in Days 1–4 serum hCG <18% was identified as an optimal test threshold that predicted treatment success with 79% sensitivity and 74% specificity, resulting in 82% positive predictive value and 69% negative predictive value. LIMITATIONS, REASONS FOR CAUTION Our findings may be limited by intervention bias resulting from existing guidelines which influences evaluation of hCG changes reliant on Day 7 serum hCG levels. WIDER IMPLICATIONS OF THE FINDINGS Examining a large prospective cohort, we show the value of Days 1–4 serum hCG changes in predicting single-dose methotrexate treatment success in tubal ectopic pregnancy. We recommend that clinicians provide early reassurance to women who have a fall or only a modest (<18%) rise in Days 1–4 serum hCG levels, that their treatment will likely be effective. STUDY FUNDING/COMPETING INTEREST(S) This project was supported by funding from the Efficacy and Mechanism Evaluation programme, a Medical Research Council and National Institute for Health Research partnership (grant reference number 14/150/03). A.W.H. has received honoraria for consultancy for Ferring, Roche, Nordic Pharma and AbbVie. W.C.D. has received honoraria from Merck and Guerbet and research funding from Galvani Biosciences. L.H.R.W. has received research funding from Roche Diagnostics. B.W.M. is supported by a NHMRC Investigator grant (GNT1176437). B.W.M. also reports consultancy for ObsEva and Merck and travel support from Merck. The other authors declare no competing interests. TRIAL REGISTRATION NUMBER This study is a secondary analysis of the GEM3 trial (ISRCTN Registry ISRCTN67795930). [ABSTRACT FROM AUTHOR]

Tubal ectopic pregnancies can cause substantial morbidity or even death. Current treatment is with methotrexate or surgery. Methotrexate treatment fails in approximately 30% of women who subsequently require rescue surgery. Gefitinib, an epidermal growth factor receptor inhibitor, might improve the effects of methotrexate. We assessed the efficacy of oral gefitinib with methotrexate, versus methotrexate alone, to treat tubal ectopic pregnancy. We performed a multicentre, randomised, double-blind, placebo-controlled trial across 50 UK hospitals. Participants diagnosed with tubal ectopic pregnancy were administered a single dose of intramuscular methotrexate (50 mg/m2) and randomised (1:1 ratio) to 7 days of additional oral gefitinib (250 mg daily) or placebo. The primary outcome, analysed by intention to treat, was surgical intervention to resolve the ectopic pregnancy. Secondary outcomes included time to resolution of ectopic pregnancy and serious adverse events. This trial is registered at the ISRCTN registry, ISCRTN 67795930. Between Nov 2, 2016, and Oct 6, 2021, 328 participants were allocated to methotrexate and gefitinib (n=165) or methotrexate and placebo (n=163). Three participants in the placebo group withdrew. Surgical intervention occurred in 50 (30%) of 165 participants in the gefitinib group and in 47 (29%) of 160 participants in the placebo group (adjusted risk ratio 1·15, 95% CI 0·85 to 1·58; adjusted risk difference –0·01, 95% CI –0·10 to 0·09; p=0·37). Without surgical intervention, median time to resolution was 28·0 days in the gefitinib group and 28·0 days in the placebo group (subdistribution hazard ratio 1·03, 95% CI 0·75 to 1·40). Serious adverse events occurred in five (3%) of 165 participants in the gefitinib group and in six (4%) of 162 participants in the placebo group. Diarrhoea and rash were more common in the gefitinib group. In women with a tubal ectopic pregnancy, adding oral gefitinib to parenteral methotrexate does not offer clinical benefit over methotrexate and increases minor adverse reactions. National Institute of Health Research. [ABSTRACT FROM AUTHOR]

Abstract Background The current measure of treatment efficacy of single-dose methotrexate for ectopic pregnancy, is a fall in serum hCG of ≥15% between days 4–7 of treatment, which has a positive predictive value of 93% for treatment success. Two small studies have proposed a fall in serum hCG between days 0–4 after treatment confers similar, earlier prognostic information, with positive predictive values of 100% and 88% for treatment success. We sought to validate this in a large, independent cohort because of the potentially significant clinical implications. Methods We conducted a retrospective study of women (n=206) treated with single-dose methotrexate for ectopic pregnancy (pre-treatment serum hCG levels ≤3000 IU/L) at Scottish hospitals between 2006–2011. Women were divided into two cohorts based on whether their serum hCG levels rose or fell between days 0–4 after methotrexate. Treatment outcomes of women in each cohort were compared, and the test performance characteristics calculated. This methodology was repeated for the current measure (≥15% fall in serum hCG between days 4–7 of treatment) and an alternate early measure ( Results In our cohort, the positive predictive value of the current clinical measure was 89% (95% CI 84-94%) (121/136). A falling serum hCG between days 0–4 predicted treatment success in 85% (95% CI 79-92%) of cases (94/110) and a Conclusions We have verified that a decline in serum hCG between days 0–4 after methotrexate treatment for ectopic pregnancies, with pre-treatment serum hCG levels ≤3000 IU/L, provides an early indication of likelihood of treatment success, and performs just as well as the existing measure, which only provides prognostic information on day 7.

Abstract Background There is a debate about the cost-efficiency of methotrexate for the management of ectopic pregnancy (EP), especially for patients presenting with serum human chorionic gonadotrophin levels of >1500 IU/L. We hypothesised that further experience with methotrexate, and increased use of guideline-based protocols, has reduced the direct costs of management with methotrexate. Methods We conducted a retrospective cost analysis on women treated for EP in a large UK teaching hospital to (1) investigate whether the cost of medical management is less expensive than surgical management for those patients eligible for both treatments and (2) to compare the cost of medical management for women with hCG concentrations 1500–3000 IU/L against those with similar hCG concentrations that elected for surgery. Three distinct treatment groups were identified: (1) those who had initial medical management with methotrexate, (2) those who were eligible for initial medical management but chose surgery (‘elected’ surgery) and (3) those who initially ‘required’ surgery and did not meet the eligibility criteria for methotrexate. We calculated the costs from the point of view of the National Health Service (NHS) in the UK. We summarised the cost per study group using the mean, standard deviation, median and range and, to account for the skewed nature of the data, we calculated 95% confidence intervals for differential costs using the nonparametric bootstrap method. Results Methotrexate was £1179 (CI 819–1550) per patient cheaper than surgery but there were no significant savings with methotrexate in women with hCG >1500 IU/L due to treatment failures. Conclusions Our data support an ongoing unmet economic need for better medical treatments for EP with hCG >1500 IU/L.

Chlamydia trachomatis (CT), the most common bacterial sexually transmitted infection worldwide, has been widely researched for its involvement in many disease pathologies in the reproductive tract, including pelvic inflammatory disease, ectopic pregnancy, and tubal factor infertility. Recent findings, through the efforts to understand the pathogenesis of CT, suggest that CT can induce the process of epithelial-to-mesenchymal transition (EMT) through epigenetic changes in the epithelium of the female reproductive tract. This literature review aims to analyze the evidence for CT's ability to promote EMT and to pinpoint the areas that merit further investigation. [ABSTRACT FROM AUTHOR]

Abstract: Aim: We have assessed the potential predictive ability of the biomarkers activin B and fibronectin (FN1) alone and when added to established markers for triaging patients as being at low or high risk of ectopic pregnancy (EP). We also assessed their use as predictors of viability at 12 weeks gestation. Methods: Exploratory secondary analysis of a prospective study including all women classified as a pregnancy of known location (PUL) based on transvaginal ultrasonography between January and December 2007 at the early pregnancy unit of St Georges’ Hospital (London). We used multinomial logistic regression to assess the diagnostic potential of the biomarkers to triage PUL at high risk of complications (EP or persistent PUL), and standard binary logistic regression to predict first trimester viability at 12 weeks. Results: For discriminating high‐risk (n = 16) from low‐risk PUL (n = 93), the area under the receiver operating characteristic curve (AUC) was 0.75 (95% confidence interval 0.60–0.85) for activin B and 0.55 (0.41–0.68) for FN1. Adding activin B to a multinomial logistic regression model incorporating β‐hCG ratio and initial progesterone yielded odds ratios of 0.16 (0.05–0.55) for failing vs high‐risk PUL and 0.29 (0.07–1.19) for intrauterine vs high‐risk PUL and increased the model's AUC from 0.84 to 0.89. At a risk threshold of 5% for high‐risk PUL, sensitivity increased from 84% to 87% and specificity from 48% to 64%. For discriminating viable (n = 28) from non‐viable (n = 81) pregnancies at 12 weeks, both markers had an AUC of 0.54. Conclusions: Our results suggested that activin B may be a promising marker to improve PUL triage in addition to established markers. [ABSTRACT FROM AUTHOR]

Ectopic Pregnancy (EP) occurs in around 1–2% of all pregnancies, and is associated with significant morbidity and mortality. Over 98% implant in the Fallopian tube. Women with risk factors, or early pregnancy symptoms of abdominal pain or vaginal bleeding are assessed by Early Pregnancy Assessment Services. The mainstay of diagnosis is by transvaginal ultrasound supported by serial serum human chorionic gonadotrophin (hCG) measurements. Management of tubal EP has moved away from surgery with growing experience with medical (methotrexate) and expectant management for selected women. Surgery will always have a role in the management of women with EP who are acutely unwell, where medical management is not likely to work, and for failed medical management. Ultrasound diagnostic criteria for non-tubal EP have been established and these cases are best managed on an individual basis. Future areas for research are the need to shorten the time to diagnosis an EP and the use of novel combination medical treatments. Future areas of medical education are the critical need to teach healthcare professionals to consider pregnancy related causes of collapse in all women of reproductive age. [ABSTRACT FROM AUTHOR]

Galectin-1 and galectin-3 are abundantly expressed at implantation sites in the uterus, suggesting their involvement in the establishment of pregnancy. In this study, we examined the expression and localization of galectin-1 and galectin-3 in fallopian tubes from nonpregnant women, and in those presenting with tubal ectopic pregnancy. There was no significant difference in the expression of either galectin-1 (LGALS1) or galectin-3 (LGALS3) transcripts in the fallopian tube across the menstrual cycle. Their expressions in the fallopian tube were inversely correlated to each other (r = −0.5134, p < 0.0001) and differentially localized. Galectin-1 protein was abundant in the stroma of nonpregnant fallopian tubes, whereas galectin-3 was mainly localized to the epithelium, notably to the cilia of ciliated cells and the apical cytoplasm of secretory cells. In ectopic pregnancies, LGALS3 expression was significantly reduced (p < 0.0001), but LGALS1 expression did not change when compared to nonpregnant fallopian tubes collected during the mid-secretory phase. The percentage of fallopian tube epithelial cells expressing galectin-3 in cilia tended to be reduced (p = 0.0685), with an accompanying loss of a normal ciliary structure, while nuclear galectin-3 increased (p < 0.05) in ectopic pregnancies. Epithelial immunostaining for galectin-1 tended to be elevated in fallopian tubes from women with ectopic pregnancy. Coculture of human trophoblast origin SW71 cells significantly increased LGALS1 expression in human fallopian tube epithelial OE-E6/E7 cells, suggesting that trophoblast-derived products regulate LGALS1 expression in the oviductal epithelium. These findings imply a differential contribution of galectin-1 and galectin-3 in the homeostasis of human fallopian tubes and in the pathophysiology of ectopic pregnancy. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Abstract: Ectopic pregnancy (EP) occurs in 1–2% of pregnancies, and is associated with significant morbidity and mortality. Women with abdominal pain or vaginal bleeding in early pregnancy, or risk factors for EP, are generally assessed by Early Pregnancy Assessment Units. Diagnosis is predominantly by trans-vaginal ultrasound supported by quantified serum human chorionic gonadotrophin (hCG). The resolution limit of trans-vaginal ultrasound means not all EPs can be identified, leaving women with a ‘Pregnancy of Unknown Location’. Management for EP has moved away from surgery with growing experience in medical management, and evidence-based recognition of expectant management for selected women. Surgery will always have a role in the management of women with EP who are acutely unwell, when medical management is not likely to work, or has failed. On-going areas of research include improvements in women's risk stratification at their first attendance with symptoms, shortening time until diagnosis of EP, and combination medical treatments. [Copyright &y& Elsevier]

Epidemiological studies have shown that cigarette smoking is a major risk factor for tubal ectopic pregnancy but the reason for this remains unclear. Here, we set out to determine the effect of smoking on Fallopian tube gene expression. An oviductal epithelial cell line (OE-E6/E7) and explants of human Fallopian tubes from non-pregnant women (n = 6) were exposed to physiologically relevant concentrations of cotinine, the principle metabolite of nicotine, and changes in gene expression analyzed using the Illumina Human HT-12 array. Cotinine sensitive genes identified through this process were then localized and quantified in Fallopian tube biopsies from non-pregnant smokers (n = 10) and non-smokers (n = 11) using immunohistochemistry and TaqMan RT-PCR. The principle cotinine induced change in gene expression detected by the array analysis in both explants and the cell line was significant down regulation (P<0.05) of the pro-apoptotic gene BAD. We therefore assessed the effect of smoking on cell turnover in retrospectively collected human samples. Consistent with the array data, smoking was associated with decreased levels of BAD transcript (P<0.01) and increased levels of BCL2 transcript (P<0.05) in Fallopian tube biopsies. BAD and BCL2 specific immunolabelling was localized to Fallopian tube epithelium. Although no other significant differences in levels of apoptosis or cell cycle associated proteins were observed, smoking was associated with significant changes in the morphology of the Fallopian tube epithelium (P<0.05). These results suggest that smoking may alter tubal epithelial cell turnover and is associated with structural, as well as functional, changes that may contribute to the development of ectopic pregnancy. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To determine the safety, tolerability, and efficacy of combination gefitinib and methotrexate to treat ectopic pregnancy. METHODS: We performed a phase I, single-arm (nonrandomized), open-label study. Twelve women with ectopic pregnancies were administered methotrexate (50 mg/m², intramuscular) and 250 mg oral gefitinib in a dose-escalation protocol: one dose (day 1) n=3; three doses (days 1-3) n=3; seven doses (days 1-7) n=6. Efficacy was examined by comparing human chorionic gonadotrophin (hCG) decline and time to resolution with historic controls administered methotrexate only. RESULTS: Common side effects were transient acneiform rash in 67% (8/12) and diarrhea in 42% (5/12) of participants. There was no clinical or biochemical evidence of serious pulmonary, renal, hepatic, or hematologic toxicity. Of six participants with a pretreatment serum hCG level between 1,000 and 3,000 international units/L, hCG levels declined significantly faster than in the control group. Median serum hCG levels by day 7 after treatment were less than one fifth of levels observed among 71 historic controls treated with methotrexate alone (median [interquartile range] hCG in participants 261 [55-1,445] international units/L compared with controls 1,426 [940-2,573]; P=.008). Median time for the ectopic pregnancies to resolve with combination therapy was 34% shorter compared with methotrexate alone (21 days compared with 32 days; P=.018). CONCLUSION: Combination gefitinib and methotrexate has potential as a treatment for ectopic pregnancy but is commonly associated with minor side effects such as transient rash and diarrhea. The treatment requires validation of safety and efficacy in a larger trial. [ABSTRACT FROM AUTHOR]

Ectopic pregnancy (EP) is difficult to diagnose early and accurately. Women often present at emergency departments in early pregnancy with a ‘pregnancy of unknown location’ (PUL), and diagnosis and exclusion of EP is challenging due to a lack of reliable biomarkers. The objective of this study was to identify novel diagnostic biomarkers for EP. Shotgun proteomics, incorporating combinatorial-ligand library pre-fractionation, was used to interrogate pooled sera (n = 40) from women undergoing surgery for EP, termination of viable intrauterine pregnancy and management of non-viable intrauterine pregnancy. Western blot was used to validate results in individual sera. ELISAs were developed to interrogate sera from women with PUL (n = 120). Sera were collected at time of first symptomatic presentation and categorized according to pregnancy outcome. The main outcome measures were differences between groups and area under the receiver operating curve (ROC). Proteomics identified six biomarker candidates. Western blot detected significant differences in levels of two of these candidates. ELISA of sera from second cohort revealed that these differences were only significant for one of these candidates, fibronectin. ROC analysis of ability of fibronectin to discriminate EP from other pregnancy outcomes suggested that fibronectin has diagnostic potential (ROC 0.6439; 95% CI 0.5090 to 0.7788; P>0.05), becoming significant when ‘ambiguous’ medically managed PUL excluded from analysis (ROC 0.6538; 95% CI 0.5158 to 0.7918; P<0.05). Fibronectin may make a useful adjunct to future multiplex EP diagnostic tests. [ABSTRACT FROM AUTHOR]

Background: Ectopic pregnancy (EP) occurs in 1–2% of pregnancies, but is over-represented as a leading cause of maternal death in early pregnancy. It remains a challenge to diagnose early and accurately. Women often present in early pregnancy with a ‘pregnancy of unknown location’ (PUL) and the diagnosis and exclusion of EP is difficult due to a lack of reliable biomarkers. A serum biomarker able to clearly distinguish between EP and other pregnancy outcomes would greatly assist clinicians in diagnosing and safely managing PULs. This study evaluates the ability of maternal serum macrophage inhibitory cytokine-1 (MIC-1) levels to differentiate between EP and other pregnancy outcomes in women with a PUL. Methods: Sera were collected from 120 women with a PUL at first clinical presentation and assayed for MIC-1 by ELISA. Results were classified according to ultimate pregnancy outcome and the discriminatory ability of MIC-1 to diagnose EP was assessed. Results: Serum MIC-1 levels were lower in women with histologically confirmed (definite) EP (dEP) (median 552 ng/mL; interquartile range (IQR) 414–693 ng/mL) compared to women with definite viable intra-uterine pregnancies (dVIUPs) (722 ng/mL; IQR 412–1122 ng/mL), and higher when compared to women with definite non-viable intra-uterine pregnancies (dNVIUPs) (465 ng/mL; IQR 341–675 ng/mL). MIC-1 levels were significantly higher in women with dEP compared to women whose PULs resolved without medical intervention (srPUL) (401 ng/mL; IQR 315–475 ng/mL) (p<0.003). There were no women with an ectopic pregnancy where serum MIC-1>1000 ng/mL. Conclusion: Serum MIC-1 levels in PUL were not able to categorically diagnose EP, however, MIC-1 could distinguish women with an EP that required medical intervention and those women whose PULs spontaneously resolved. A single serum MIC-1 measurement also excluded EP at levels above 1000 ng/mL. MIC-1 may play a role in the development of a combined assay of biomarkers for the diagnosis of EP. [ABSTRACT FROM AUTHOR]

Background: Ectopic pregnancy (EP) is an important cause of morbidity and mortality amongst women of reproductive age. Tubal EP is well described in industrialised countries, but less is known about its impact in low-resource countries, in particular in the South Pacific Region. Methods: We undertook a retrospective review of women with tubal EP treated at a provincial referral hospital in coastal Papua New Guinea over a period of 56 months. Demographic and clinical variables were obtained from patients' medical records and analysed. The institutional rate of tubal EP was calculated, and diagnosis and management reviewed. Potential risk factors for tubal EP were identified, and delays contributing to increased morbidity described. Results: A total of 73 women had tubal EP. The institutional rate of tubal EP over the study period was 6.3 per 1,000 deliveries. There were no maternal deaths due to EP. The mean age of women was 31.5+/-5.7 years, 85% were parous, 67% were rural dwellers and 62% had a history of sub-fertility. The most commonly used diagnostic aid was culdocentesis. One third of women had clinical evidence of shock on arrival. All women with tubal EP were managed by open salpingectomy. Tubal rupture was confirmed for 48% of patients and was more common amongst rural dwellers. Forty-three percent of women had macroscopic evidence of pelvic infection. Two-thirds of patients received blood transfusions, and post-operative recovery lasted six days on average. Late presentation, lack of clinical suspicion, and delays with receiving appropriate treatments were observed. Conclusions: Tubal EP is a common gynaecological emergency in a referral hospital in coastal PNG, and causes significant morbidity, in particular amongst women residing in rural areas. Sexually transmitted infections are likely to represent the most important risk factor for tubal EP in PNG. Interventions to reduce the morbidity due to tubal EP include the prevention, detection and treatment of sexually transmitted infections, identification and reduction of barriers to prompt presentation, increasing health workers' awareness of ectopic pregnancy, providing pregnancy test kits to rural health centres, and strengthening hospital blood transfusion services, including facilities for autotransfusion. [ABSTRACT FROM AUTHOR]

Background: Ectopic pregnancy (EP) remains the most life-threatening acute condition in modern gynaecology. It remains difficult to diagnose early and accurately. Women often present at emergency departments in early pregnancy with a 'pregnancy of unknown location' (PUL) and diagnosis/exclusion of EP is challenging due to a lack of reliable biomarkers. Recent studies suggest that serum levels of a disintegrin and metalloprotease protein-12 (ADAM-12) can be used differentiate EP from viable intrauterine pregnancy (VIUP). Here we describe a prospective study evaluating the performance of ADAM-12 in differentiating EP from the full spectrum of alternative PUL outcomes in an independent patient cohort. Methodology/Principal Findings: Sera were collected from 120 patients at their first clinical presentation with a PUL and assayed for ADAM-12 by ELISA. Patients were categorized according to final pregnancy outcomes. Serum ADAM-12 concentrations were increased in women with histologically-confirmed EP (median 442 pg/mL; 25%-75% percentile 232-783 pg/mL) compared to women with VIUP (256 pg/mL; 168-442 pg/mL) or miscarriage (192 pg/mL; 133-476 pg/mL). Serum ADAM-12 did not differentiate histologically- confirmed EP from spontaneously resolving PUL (srPUL) (416 pg/mL; 154-608 pg/mL). The diagnostic potential of ADAM-12 was only significant when 'ambiguous' PUL outcomes were excluded from the analysis (AROC = 0.6633; P = 0.03901). Conclusions/Significance: When measured in isolation, ADAM-12 levels had limited value as a diagnostic biomarker for EP in our patient cohort. The development of a reliable serum biomarker-based test for EP remains an ongoing challenge. [ABSTRACT FROM AUTHOR]

Abstract: As part of successful human reproduction, the Fallopian tube must provide a suitable environment for pre-implantation development of the embryo and for efficient transport of the embryo to the uterus for implantation. These functions are coordinated by paracrine interactions between tubal epithelial, smooth muscle and immune cells and the cells of the developing embryo. Alterations in these signals can lead to a tubal microenvironment encouraging of embryo implantation and to dysregulated tubal motility, ultimately resulting in inappropriate and early implantation of the embryo in the Fallopian tube. Here, we highlight novel and emerging concepts in tubal physiology and pathobiology, such as the induction of a receptive phenotype within the Fallopian tube, leading to ectopic implantation. Chlamydia trachomatis infection is a risk factor for tubal ectopic pregnancy. Activation of toll-like receptor 2 (TLR-2) in the Fallopian tube epithelium, by C. trachomatis has recently been demonstrated, leading to the dysregulation of factors involved in implantation and smooth muscle contractility, such as prokineticins (PROK), activin A and interleukin 1 (IL-1). The Fallopian tube has also recently been shown to harbour a unique population of immune cells, compared to the endometrium. In addition, the complement of immune cells in the Fallopian tube has been reported to be altered in Fallopian tube from women with ectopic pregnancy. There are increasing data suggesting that vascularisation of the Fallopian tube, by the embryo during ectopic pregnancy, differs from that initiated in the uterus during normal pregnancy. This too, is likely the result of paracrine signals between the embryo and the tubal microenvironment. [Copyright &y& Elsevier]

The endometrium in early pregnancy undergoes decidualization and functional changes induced by local trophoblast, which are not fully understood. We hypothesized that endometrium from tubal ectopic pregnancy (EP) could be interrogated to identify novel genes and pathways involved in these processes. Gestation-matched endometrium was collected from women with EP (n = 11) and intrauterine pregnancies (IUP) (n = 13). RNA was extracted from the tissue. In addition, tissues were prepared for histological analysis for degree of decidualization. We compared a) the samples from EP that were decidualized (n = 6) with non-decidualized samples (n = 5), and b) the decidualized EP (n = 6) with decidualizationmatched IUP (n = 6) samples using an Affymetrix gene array platform, with Ingenuity Pathway Analysis, combined with quantitative RT-PCR. Expression of PRL and IGFBP1 was used to confirm the degree of decidualization in each group. There were no differences in PRL or IGFBP1 expression in the decidualization-matched samples but a marked reduction (P<0.001) in the non-decidualized samples. Decidualization was associated with increased expression of 428 genes including SCARA5 (181-fold), DKK1 (71-fold) and PROK1 (32-fold), and decreased expression of 230 genes including MMP-7 (35-fold) and SFRP4 (21-fold). The top canonical pathways associated with these differentially expressed genes were Natural Killer Cell and Wnt/ b-Catenin signaling. Local trophoblast was associated with much less alteration of endometrial gene expression with an increase in 56 genes, including CSH1 (8-fold), and a reduction in 29 genes including CRISP3 (8-fold). The top associated canonical pathway was Antigen Presentation. The study of endometrium from tubal EP may promote novel insights into genes involved in decidualization and those influenced by factors from neighboring trophoblast. This has afforded unique information not highlighted by previous studies and adds to our understanding of the endometrium in early pregnancy. [ABSTRACT FROM AUTHOR]

Background: Embryo retention in the Fallopian tube (FT) is thought to lead to ectopic pregnancy (EP), a considerable cause of morbidity. In mice, genetic/pharmacological silencing of cannabinoid receptor Cnr1, encoding CB1, causes retention of embryos in the oviduct. The role of the endocannabinoids in tubal implantation in humans is not known. Methods and Findings: Timed FT biopsies (n = 18) were collected from women undergoing gynecological procedures for benign conditions. Endometrial biopsies and whole blood were collected from women undergoing surgery for EP (n = 11); management of miscarriage (n = 6), and termination of pregnancy (n = 8). Using RT-PCR and immunohistochemistry, CB1 mRNA and protein expression levels/patterns were examined in FT and endometrial biopsies. The distribution of two polymorphisms of CNR1 was examined by TaqMan analysis of genomic DNA from the whole blood samples. In normal FT, CB1 mRNA was higher in luteal compared to follicular-phase (p<0.05). CB1 protein was located in smooth muscle of the wall and of endothelial vessels, and luminal epithelium of FT. In FT from women with EP, CB1 mRNA expression was low. CB1 mRNA expression was also significantly lower (p<0.05) in endometrium of women with EP compared to intrauterine pregnancies (IUP). Although of 1359G/A (rs1049353) polymorphisms of CNR1 gene suggests differential distribution of genotypes between the small, available cohorts of women with EP and those with IUP, results were not statistically significant. Conclusions: CB1 mRNA shows temporal variation in expression in human FT, likely regulated by progesterone. CB1 mRNA is expressed in low levels in both the FT and endometrium of women with EP. We propose that aberrant endocannabinoidsignaling in human FT leads to EP. Furthermore, our finding of reduced mRNA expression along with a possible association between polymorphism genotypes of the CNR1 gene and EP, suggests a possible genetic predisposition to EP that warrants replication in a larger sample pool. [ABSTRACT FROM AUTHOR]

1–2% of pregnancies are ectopic, the majority implanting in the Fallopian tube. A single, systemic dose of methotrexate, a DNA-synthesis (S phase) inhibitor, has been used since 1991 for outpatient treatment of women with stable EP. However, methotrexate has limited clinical and cost effectiveness, restricting its use to 25–30% of these women. There is an unmet need for better medical treatment for EP. Colony stimulating factor-1 (CSF-1) promotes placentation and creates a pro-inflammatory environment that is fundamental for the maintenance of a normal pregnancy. We hypothesised that CSF-1 is also involved in the placentation and maintenance of an EP. Herein, we demonstrate the immunolocalisation of the CSF-1 receptor (CSF-1R) as well as its ligand (CSF-1) in immortalised first trimester trophoblast cells. We show that a specific CSF-1R kinase inhibitor, GW2580, abolishes CSF-1 induced trophoblast cell proliferation and migration and can be cytotoxic. We then demonstrate the expression of CSF-1R and CSF-1 in the cytotrophoblast and syncytiotrophoblast within ectopic implantation sites from women with EP. Our data suggests that CSF-1 is involved in the survival and proliferation of trophoblast cells in EP. This suggests that pharmacological disruption of CSF-1/CSF-1R signaling axis could be the basis of a new therapeutic for EP. [ABSTRACT FROM AUTHOR]

Ectopic pregnancy remains a considerable cause of maternal morbidity and mortality worldwide. Currently, it is diagnosed using a combination of transvaginal ultrasound and serial serum β-human chorionic gonadotrophin levels. Diagnosis is often delayed and these tests are time-consuming and costly, both psychologically to the patient and financially to health services. The development of a biomarker that can differentiate a tubal ectopic from an intrauterine implantation is therefore important. In the pre-genomic era, a one-by-one scientific approach has revealed over 20 candidate biomarkers that could be used as a test to diagnose ectopic pregnancy although at present their clinical utility is very limited. These biomarkers cluster into themes: markers of abnormal embryo/trophoblast growth, markers of abnormal corpus luteum function, markers of a growing pregnancy in the Fallopian tube, markers of inflammation and peritoneal irritation, and uterine markers of normal implantation. It is likely that this thematic approach will facilitate the identification of newer biomarkers using microarray technology and inform the development of investigative paradigms using multiple markers at the time of presentation. [ABSTRACT FROM AUTHOR]

To address methodological deficiencies in published randomized controlled trials and systematic reviews, this study has developed a core outcome set to guide future research in ectopic pregnancy (EP). To identify potential outcomes, we performed a comprehensive literature review and interviews with individuals with lived experience in EP. Potential core outcomes were then entered into a 3-round Delphi survey. A total of 154 participants from 6 continents, comprising health care professionals, researchers, and individuals with lived experience in EP, completed all 3 rounds of the Delphi survey. Outcomes were prioritized at 3 consensus development meetings, and recommendations were developed on how to report these outcomes where possible. Not applicable. Health care professionals, researchers, and individuals with lived experience in EP. Not applicable. Consensus for inclusion in core outcome set. Six outcomes reached full consensus, including treatment success, resolution time, the number of additional interventions, adverse events, mortality and severe morbidity, and treatment satisfaction. The core outcome set with 6 outcomes for EP will help standardize reporting of clinical trials, facilitate implementation of findings into clinical practice, and enhance patient-centered care. [ABSTRACT FROM AUTHOR]

Placenta has the highest expression of epidermal growth factor (EGF) receptor of all tissues, a cell signaling pathway promoting survival and growth. Therefore, EGF receptor inhibition could potentially treat ectopic pregnancy. We undertook preclinical studies to examine whether gefitinib (orally available EGF receptor inhibitor) with or without methotrexate inhibits placental cell growth. Gefitinib and methotrexate were added to placental cells and their ability inhibit cell growth, block EGF receptor signaling, and induce apoptosis (programmed cell death) was examined. They were also administered to two animal mouse models to examine their effects on placental tissue in vivo. Epidermal growth factor receptor was highly expressed in placental tissue from ectopic pregnancies. Combining gefitinib with methotrexate potently inhibited growth of placental cells, including placental cell lines (JEG3, BeWo cells) and cells isolated from first-trimester placenta. These drugs were additive in blocking EGF receptor signaling and inducing apoptosis. Gefitinib and methotrexate administered together were more potent in decreasing the volume of human placental cells xenografted subcutaneously onto mice compared with either alone. By day 19 after xenografting, mean (±standard error of the mean), xenograft volumes were: 821 (±68) mm after gefitinib treatment, 901 (±204) mm after methotrexate treatment, and 345 (±137) mm after both drugs were given (P<.01 for both comparisons of single therapy compared with combination therapy). Combining these agents doubled rates of fetal resorption in pregnant mice compared with each drug alone. Combining gefitinib with methotrexate potently inhibits placental cell growth in vitro and in mouse models. The combination may have potential in treating ectopic pregnancies. [ABSTRACT FROM AUTHOR]

Objective: To investigate whether gonadotropin-releasing hormone (GnRH) and GnRH receptor (GnRHR) are expressed at tubal ectopic pregnancy sites, and to study the potential role of GnRH signaling in regulating immortalized human trophoblast cell viability.Design: Immunohistochemical and experimental studies.Setting: Academic research laboratory.Patient(s): Fallopian tube implantation sites (n = 25) were collected from women with ectopic pregnancy. First-trimester human placenta biopsies (n = 5) were obtained from elective terminations of pregnancy.Intervention(s): None.Main Outcome Measure(s): GnRH and GnRHR expression was examined by means of immunohistochemistry and histoscoring. Trophoblastic BeWo choriocarcinoma and immortalized extravillous trophoblast (HTR-8/SVneo) cell viability was examined by means of cell counting after incubation with GnRH and/or GnRH antagonist (Antide).Result(s): GnRH and GnRHR immunoreactivity was detected in cytotrophoblast, syncytiotrophoblast, and extravillous trophoblast in all women with tubal pregnancy. GnRH immunoreactivity was higher and GnRHR immunoreactivity lower in syncytiotrophoblast compared with cytotrophoblast. GnRH and GnRHR immunoreactivity was detected in adjacent fallopian tube epithelium. Whereas neither GnRH nor Antide altered HTR-8/SVneo cell viability, treatment with GnRH significantly increased the overall cell viability of BeWo cells at 48 and 72 hours, and these effects were abolished by pretreatment with Antide.Conclusion(s): GnRH and GnRHR are expressed in trophoblast cell populations and fallopian tube epithelium at tubal ectopic pregnancy sites. GnRH increases BeWo cell viability, an effect mediated by the GnRHR. Further work is required to investigate the potential role of GnRH signaling in ectopic pregnancy. [ABSTRACT FROM AUTHOR]

Objective: To demonstrate expression and regulation of prokineticins (PROKs) and their receptors (PROKRs) in fallopian tube (FT) from women who are not pregnant and women with ectopic pregnancy (EP). Design: Tissue analysis. Setting: Large United Kingdom teaching hospital. Patient(s): Women undergoing hysterectomy for benign gynecological conditions (n = 15) and surgery for EP (n = 16). Intervention(s): Quantitative reverse transcriptase–polymerase chain reaction (RT-PCR) and immunohistochemistry were used to determine FT PROK/PROKR messenger RNA (mRNA) expression and protein localization, respectively. The PROK/PROKR levels were measured in tubal explant cultures stimulated with estrogen (E) and progestogen. Main Outcome Measure(s): Differential expression of PROK and PROKR. Result(s): The FT PROK2 and PROKR1 mRNA levels were up-regulated during the P-dominant midluteal phase of the menstrual cycle. Increased PROKR1 expression was observed in tubal explant cultures treated with medroxy-progesterone acetate (MPA). The PROK and PROKR proteins were localized to the epithelium and smooth muscle layers of the FT. The PROKR1 and PROKR2 mRNA levels were lower in FT from women with EP compared with nonpregnant FT from the midluteal phase. Conclusion(s): These data suggest a potential role for PROKs in FT function. The PROKs are known to affect smooth muscle contraction in the gut. Dysregulated PROK expression in FT could affect FT smooth muscle contractility and embryo–tubal transport, providing a potential cause for EP. [ABSTRACT FROM AUTHOR]