Evaluation of the parameters in predicting single‐dose methotrexate therapy success for ectopic pregnancy

Methotrexate has demonstrated efficacy in treating ectopic pregnancies. This study explores factors influencing treatment success, focusing on laboratory and ultrasonographic findings, particularly the day 4 to day 1 β‐hCG level ratio.


INTRODUCTION
Ectopic pregnancy (EP) manifests when conceptus material implants outside the endometrial cavity, predominantly observed in the fallopian tube, constituting 95% of cases.Its incidence, affecting about 1%-2% of pregnancies, underscores its clinical significance. 1,2Despite advancements in early diagnosis and treatment options, EP continues to impose substantial morbidity and mortality risks during the early stages of pregnancy.
Traditionally, surgical intervention has been the cornerstone of EP treatment.However, a pivotal paradigm shift occurred in the early 1980s with the emergence of methotrexate (MTX) as a successful medical alternative.Initially approved for gestational trophoblastic neoplasia, MTX's efficacy in treating EP has since been recognized, ushering in a new era in the management of this complex condition. 3TX operates by inhibiting dihydrofolic acid, disrupting DNA synthesis, particularly in rapidly proliferating tissues such as trophoblasts.The established regimen for MTX treatment entails a single intramuscular injection at a dose of 50 mg/m 2 .Despite its widespread adoption, the success of single-dose MTX treatment is not uniform.A meta-analysis has revealed varying success rates, spanning from 65% to 95%. 4 This observed variability underscores the importance of identifying predictive factors influencing treatment outcomes, prompting a closer examination of patient-specific parameters and their impact on the therapeutic efficacy of MTX.
The consistently high success rates associated with MTX have firmly established it as the primary treatment option for EP, with notable efficacy in cases not involving an acute abdomen or active intra-abdominal bleeding.This shift toward medical management has been particularly significant since the early 1980s, reflecting a departure from the traditional surgical approach. 5hile MTX has become a cornerstone in EP management, it is crucial to acknowledge relative contraindications that guide its judicious use.These include the presence of fetal cardiac activity, initial human chorionic gonadotropin (hCG) concentrations surpassing 5000 mIU/mL, and EP size exceeding 4 cm. 6Recognizing these contraindications is pivotal in refining patient selection and optimizing treatment outcomes, aligning with the evolving landscape of EP management.
MTX treatment is typically administered as a single dose, although variations such as a two-dose protocol or multiple-dose protocols have also been explored.The widespread adoption of a single-dose regimen reflects its simplicity and convenience, contributing to MTX's overall high success rate in treating ectopic pregnancies However, it is essential to acknowledge potential adverse effects associated with MTX administration.While generally well-tolerated, MTX can lead to adverse outcomes such as marrow suppression, kidney failure, or gastric perforation, each carrying the potential for life-threatening consequences.Recognizing these risks underscores the importance of careful patient selection and vigilant monitoring during and after MTX therapy. 7n addition to potential side effects, the post-treatment observation period often involves a hospital stay of 7-8 days.During this crucial timeframe, clinicians assess whether MTX has effectively reduced beta hCG levels.This monitoring period plays a pivotal role in gauging the response to treatment and informing subsequent decisions regarding the course of care.
The primary aim of this study is to evaluate factors influencing the success of MTX treatment for EP.Additionally, our secondary objective is to investigate conditions that contribute to the reduction of hospital stay and enhance MTX treatment success.Through a comprehensive analysis of patient characteristics, laboratory findings, and ultrasonographic observations, we seek to identify key determinants that can inform clinical decisions, ultimately optimizing outcomes in the medical management of ectopic pregnancies.

METHODS
A retrospective cohort study was conducted at Bursa Yuksek Ihtisas Training and Research Hospital, University of Health Sciences, following approval from the local ethics committee.The study focused on patients diagnosed with tubal EP within the timeframe of January 2018 to December 2021, identified through the hospital registry system.Comprehensive reviews of patients' records were performed, capturing essential characteristics such as age, gravidity, parity, and gestational date calculated from the first day of the menstrual period.Additionally, symptoms were meticulously documented.
Transvaginal ultrasonography findings, including measurements of endometrial thickness, adnexal mass size, and the maximum length of free fluid in the pouch of Douglas, were systematically recorded.Upon the diagnosis of EP, patients were admitted to the hospital.Surgical intervention was promptly conducted for those presenting with acute abdominal pain and/or active intraabdominal bleeding.Hemodynamically stable patients without acute abdomen and without contraindications for MTX treatment received a single intramuscular dose of MTX (50 mg/m 2 ).
The levels of β-hCG were systematically monitored on day 1, day 4, and day 7 following MTX administration. 8A decrease in β-hCG levels by more than 15% between day 4 and day 7 was deemed indicative of a successful response to the single MTX dose.Subsequently, patients entered a weekly monitoring phase until their β-hCG levels reached non-pregnant levels.
In instances where the decrease was <15%, a second MTX dose was administered.Notably, for some patients, a second MTX dose was administered on day 4. Throughout hospitalization, should patients develop acute abdominal pain or experience abdominal bleeding leading to a significant drop in hemoglobin levels, prompt surgical intervention was implemented.Receiver Operating Characteristic (ROC) curve analysis was employed to evaluate efficiency and determine cut-off values for β-hCG and the day 4 β-hCG level to day 1 β-hCG level ratio in predicting the success of single-dose MTX treatment and the timing of the second dose of MTX re-administration.Statistical significance was defined as a p-value <0.05.

RESULTS
Over a 4-year period, a total of 465 patients were diagnosed with tubal EP.Following the exclusion of women with inadequate follow-up examinations and those with inappropriate timing of β-hCG assessments, 439 patients met the inclusion criteria for the study.
Among these patients, 259 required immediate surgery due to acute abdominal symptoms and active abdominal bleeding.The mean gestational age at diagnosis was 43 days, with an average largest adnexal mass size of 30 mm.Approximately 34% of women presented with β-hCG levels exceeding 5000 mIU/mL, while the mean β-hCG level at admission was 3223 mIU/mL.Detailed data are presented in Table 1.
A total of 143 patients achieved successful treatment with MTX, distributed as follows: single-dose MTX (n = 102), day 1 and 7 administered MTX (n = 22), and day 1 and day 4 administered MTX (n = 19).Additionally, 37 women required surgery after MTX administration, with 31 undergoing surgery within 7 days of MTX treatment and 6 patients requiring surgery after 7 days and onwards.
Comparisons between women successfully treated with MTX and those undergoing surgery after MTX utilization revealed no significant differences in age, gravidity, parity, vaginal bleeding, endometrial thickness, longest adnexal mass dimension, and the width of free fluid in the pouch of Douglas.Approximately 14% of women had adnexal masses larger than 4 cm.However, significant differences were observed in specific parameters, including day 1 β-hCG levels, gestational date, and the proportion of women with β-hCG levels exceeding 5000 and 10 000 mIU/mL.Notably, the mean day 4 to day 1 β-hCG level ratio was higher in the group of women who underwent surgery after MTX administration (0.91 to 1.25), and this difference was statistically significant.Detailed data and parameter comparisons are presented in Table 2.
The overall success rate of MTX treatment was 79%.Notably, when patients presented with β-hCG levels <1000 mIU/mL, the success rate significantly increased to 93%, and for those with β-hCG levels below 2000 mIU/mL, the success rate remained relatively high at 89%.However, the success rate gradually decreased with higher β-hCG levels.Specifically, when β-hCG levels exceeded 5000 mIU/mL, the success rate dropped to 50%.A detailed breakdown of success rates corresponding to β-hCG levels is provided in Table 3.
Considering the lower occurrence of adverse side effects and the potential for cost-effectiveness, we conducted an evaluation of the factors influencing the success of single-dose MTX treatment.For comparison, we separately analyzed the single-dose MTX successful group with the two-dose MTX groups (day 1/4 and day 1/7), and specific parameters are presented in Table 4. Throughout the study, patients were diligently monitored for potential side effects of MTX.Importantly, no serious side effects, such as liver toxicity, bone marrow suppression, or stomach perforation, were observed in any patient.
Notably, there were no statistically significant differences in terms of gestational date, symptoms, or ultrasonographic evaluations between these groups.Although the day 1 β-hCG level was higher in the two-dose group, it is worth highlighting that the day 4 to day 1 β-hCG level ratio was also higher in both two-dose regimens.
The patients successfully treated with a single dose of MTX were compared with those who underwent surgery within 7 days after MTX treatment group, consisting of 31 patients.The comparison between these groups revealed a significant difference in the day 4 to day 1 β-hCG level ratio, which was higher in that group.However, no significant difference was detected in the number of women with adnexal masses larger than 4 cm.Furthermore, we assessed the same parameters to determine differences when comparing the two-dose MTX groups.However, no statistical differences were observed between the day 1/4 MTX treatment group and the day 1/7 MTX treatment group, including day 1 β-hCG levels and the day 4 to day 1 β-hCG level ratio.
Women who received MTX and subsequently underwent surgery within 7 days constituted one of the pivotal groups in our study.It is crucial to minimize the occurrence of such cases, despite the fact that all patients in this group underwent surgery while hospitalized.This subgroup comprised 31 patients.When comparing these patients with women treated with a single dose of MTX, we found that they had similar mean gestational age, gravidity, and parity.Although these women tended to have larger adnexal masses and showed the presence of free fluid and increased endometrial thickness, these differences were not statistically significant.
As previously mentioned, women who underwent surgery within the first week of MTX treatment not only had higher day 1 β-hCG levels but also exhibited a significantly elevated day 4 to day 1 β-hCG level ratio.A detailed evaluation of these parameters is provided in Table 5.
To optimize the success of MTX treatment and determine the ideal timing for the second dose of MTX, a ROC analysis was conducted for β-hCG levels at day 1 and the day 4 to day 1 β-hCG level ratio.Figure 1 illustrates the cut-off values for β-hCG levels indicative of MTX treatment success.A specific cut-off value of 2255 mIU/mL was identified, with a specificity of 68.5% and a sensitivity of 70.3%.
To optimize the success of MTX treatment and determine the ideal timing for the second dose of MTX, a ROC analysis was conducted for β-hCG levels at day 1 and the day 4 to day 1 β-hCG level ratio.Figure 1 illustrates the cut-off values for β-hCG levels indicative of MTX treatment success.A specific cut-off value of 2255 mIU/mL was identified, with a specificity of 68.5% and a sensitivity of 70.3%.
The day 4 to day 1 β-hCG level ratio exhibited significant differences when comparing the single-dose MTX group with all other groups.Figure 2 illustrates the ROC analysis for determining the appropriate threshold indicating the failure of single-dose MTX treatment.A cutoff value of 95.5% was identified, signifying that the decrease in the day 1 β-hCG level was only 4.5% by  facilitated early detection.Timely diagnosis is pivotal, as it opens the door to medical treatment options.
MTX, a medication with a successful history in treating hydatidiform moles and choriocarcinomas for over four decades, has revolutionized the management of ectopic pregnancies. 9While certain contraindications for MTX are universally accepted, 10 the concept of relative contraindications was introduced in a 2013 committee opinion by The Practice Committee of the American Society for Reproductive Medicine. 6Notably, high β-hCG levels (>5000 mIU/mL) and the presence of larger ectopic pregnancies (as determined by transvaginal ultrasonography) were among the factors considered as relative contraindications.Consequently, physicians have increasingly favored MTX as a treatment option for these patients.
The primary objective of this study is to elucidate the success of MTX treatment and identify the factors influencing this success.Age has been a subject of investigation regarding its potential association with the success of MTX treatment.However, existing studies have not reached a consensus on whether age plays a significant role in MTX treatment success. 11,12In our present study, after comparing age across various groups, no significant differences were observed.
Vaginal bleeding is a common symptom of EP.A recent study suggested that vaginal bleeding at admission could potentially signal a higher risk of medical treatment failure.The authors of this study proposed that inadequate invasion of the tubal serosa, which is thinner than the endometrial cavity, might lead to reduced progesterone levels, resulting in vaginal bleeding prior to tubal rupture. 13However, in our study with a larger participant pool, we did not find a significant association between vaginal bleeding and the success of MTX treatment.
Active intra-abdominal bleeding and acute abdomen are well-established indications for surgical intervention in EP.Some patients may experience abdominal bleeding, detectable via transvaginal or transabdominal ultrasonography, and some gynecologists might consider the presence of abdominal bleeding as an indication for surgery.The key question we aimed to answer was whether the presence of intraabdominal free fluid was indicative of MTX treatment failure.Our analyses revealed that neither the presence of free fluid nor the length of free fluid in the pouch of Douglas were associated with MTX treatment failure.
Therefore, women detected to have free fluid during transvaginal ultrasonographic examinations may still be suitable candidates for MTX treatment unless they present with acute abdomen.However, it is essential for these patients to undergo careful monitoring with serial ultrasonographic examinations and complete blood count analyses.Based on our study, the decision for surgical intervention should be considered in the presence of acute abdomen or a dramatic decrease in complete blood count.
The size of EP, particularly when exceeding 4 cm, has been considered a relative contraindication for MTX The ROC curve analysis of the β-hCG level for the MTX treatment success.For a cut-off value of 2255 mIU/mL, specificity was 68,5% and sensitivity was 70.3%.Area under the curve 0.764; p < 0,001; standard error 0.043.ROC, receiver operating characteristic; MTX, methotrexate.

F I G U R E 2
The ROC curve analysis of the Day 4 to day 1 β-hCG level ratio for the prediction of single-dose MTX treatment.For a cutoff value of 95,5%, specificity was 72,5% and sensitivity was 84,7% with a positive predictive value of 75,4%.Area under the curve 0.815; p < 0,001; standard error 0.033.ROC, receiver operating characteristic; MTX, methotrexate.
treatment. 6In our present study, we found no statistically significant differences, not only in terms of the longest adnexal mass length but also in the number of women with adnexal masses larger than 4 cm.This suggests that adnexal mass length may not be directly associated with MTX treatment failure unless the patient presents with acute abdominal symptoms.
It is worth noting that in some cases, the entire fallopian tube may become filled with a blood clot, which can appear as a mass during diagnostic imaging.In such instances, careful monitoring and assessment of the abdominal examination, as well as tracking free fluid levels in the abdomen and changes in hemogram levels, can help guide clinical management, as previously mentioned.
MTX has consistently demonstrated its effectiveness in the medical treatment of EP when administered at a dose of 50 mg/m 2 .5][16] One of the most critical predictors of success has consistently been the β-hCG level.
In the present study, we observed an overall MTX treatment success rate of 79%, consistent with findings in the existing literature. 17As expected, the success rate gradually decreased as β-hCG levels increased.Notably, nearly 93% of patients with β-hCG levels below 1000 mIU/mL were successfully treated without encountering adverse effects.In contrast, approximately half of the women with β-hCG levels exceeding 5000 mIU/mL either experienced MTX treatment failure or required surgery.Consequently, one of the primary objectives of our study was to identify a cut-off value for MTX treatment.
The ROC curve analysis identified a crucial cut-off value of 2255 mIU/mL, which holds significant clinical implications for our clinic.Traditionally, nearly all women diagnosed with EP were hospitalized for 7 days to monitor day 4 and day 7 β-hCG levels.However, this newly established cut-off value may allow us to consider outpatient management after explaining potential scenarios and obtaining written consent from these patients.
We evaluated the day 4 to day 1 β-hCG level ratio as a determinant of the success of single-dose MTX treatment with the aim of reducing the MTX failure rate and potentially sparing patients from a second dose.It is worth noting that a contemporary study mentioned the possibility of β-hCG levels increasing on day 4 due to the lysis of trophoblastic tissue. 180][21] In our study, we introduced a novel approach by using the ratio of day 4 to day 1 β-hCG levels (calculated by dividing the day 4 β-hCG level by the day 1 β-hCG level) to determine the likelihood of single-dose MTX treatment success.
According to our calculations, if the result of the day 4 β-hCG level to day 1 β-hCG level ratio was <95.5%, there was a 75% predictive value for the success of single-dose MTX treatment.
It is important to highlight that this calculation method, involving the day 4 to day 1 β-hCG level ratio, has not been widely employed in existing literature.This evidence is of paramount significance, as a ratio exceeding 95.5% could potentially prompt the administration of a second dose of MTX to enhance the success rate and mitigate the risk of surgery.
To sum up, MTX stands as a suitable treatment option for EP.The establishment of an optimal cutoff value for MTX treatment provides valuable guidance for patient follow-up and identifying those at greater risk of treatment failure.The day 4 to day 1 β-hCG level ratio emerges as a valuable marker for predicting the success of single-dose MTX treatment.This marker may enable the earlier administration of a second dose of MTX, potentially leading to higher treatment success rates.

T A B L E 1
Abbreviation: β-HCG, beta human chorionic gonadotropin.

T A B L E 2 Age
Demonstration and the comparison of the parameters between the MTX successful group and women who were operated after MTX administration.MTX successful (n = 143)Surgery after MTX (n = 37) p The comparison of the parameters between the single-dose group and two-dose groups.
Note: Bold indicates statistically significant data.Abbreviations: β-HCG, beta human chorionic gonadotropin; MTX, methotrexate.T A B L E 3The success rate of MTX treatment in accordance with day 1 β-HCG level.Abbreviations: β-HCG, beta human chorionic gonadotropin; MTX, methotrexate.theday4β-hCGlevel.This threshold demonstrated a specificity of 72.5% and sensitivity of 84.7%.These results indicate that if the decrease between day 1 and day 4 is less than 4.5%, administering the second dose of MTX may be warranted without waiting for the day 7 β-hCG level, as depicted in Figure2.DISCUSSIONEP remains a life-threatening condition, especially when diagnosis and treatment are delayed.Fortunately, advances in laboratory testing and screening methods, such as transvaginal ultrasonography, have T A B L E 4 Note: Bold indicates statistically significant data.Abbreviations: β-HCG, beta human chorionic gonadotropin; Day 1/7, day 1 and day 7; Day 1/4, day 1 and day 4; MTX, methotrexate.T A B L E 5 Demonstration of the parameters in women who had been treated with single-dose MTX and who had undergone operation within the first week after MTX treatment.