Ectopic pregnancies represent 1–2% of all pregnancies (100,000 per year in the United States1 and 12,000 per year in the United Kingdom2) and are a leading cause of maternal morbidity and mortality in the first trimester, responsible for 3–8% of all pregnancy-related deaths.2,3 Ectopic pregnancies can be treated surgically or medically by administering intramuscular methotrexate. A systematic meta-analysis concluded the cost-effectiveness of medical treatment with methotrexate drops significantly with higher pretreatment human chorionic gonadotrophin (hCG) levels (greater than 1,500 international units/L) and that laparoscopic excision remains the most effective treatment for ectopic pregnancy.3 Hence, many are treated surgically4 and there exists a need for more effective medical therapies to reduce operative intervention (and its inherent risks) in women diagnosed with ectopic pregnancy.
Human placenta expresses very high levels of the epidermal growth factor (EGF) receptor, a cellular signaling pathway that activates a potent cell survival response.5 The placenta has, by far, the highest expression of EGF receptor compared with all nonmalignant tissues in the body. A bioinformatics search on BioGPS suggests EGF receptor expression in human placenta is more than 30 times higher than average tissue expression,6,7 and there is good evidence that the placenta heavily relies on EGF receptor signaling for successful pregnancy outcomes.8–11 Therefore, EGF receptor inhibition may be a molecularly targeted approach to treat ectopic pregnancy.
Gefitinib is a small-molecule tyrosine kinase inhibitor that blocks EGF receptor signaling and is approved to treat non–small-cell lung cancer, in which 250 mg is taken daily on a continuing basis. It has good oral bioavailability and often causes transient diarrhea and skin rash.12,13 In preclinical studies,14 we have found gefitinib supra-additively augments methotrexate-induced regression of placental tissue. Given the encouraging results obtained from the preclinical studies,14 we translated our finding to humans, conducting a phase I trial to assess the safety, tolerability, and efficacy of combination gefitinib (once-daily 250 mg oral gefitinib taken up to 7 days) and intramuscular methotrexate to treat ectopic pregnancy.
MATERIALS AND METHODS
We performed a phase I single-arm (nonrandomized) open-label dose-escalation study, recruiting 12 women with an ultrasonographic diagnosis of a tubal ectopic pregnancy (trial registration: Australian New Zealand Clinical Trials Registry, ACTRN12610000684022). Women were recruited between October 2010 and October 2011 at two academic teaching hospitals: Monash Medical Centre (Melbourne, Australia) and The Royal Infirmary of Edinburgh (Edinburgh, United Kingdom). This was an investigator-initiated study with no funding from pharmaceutical companies. All drugs were purchased from the hospital pharmacy and none was provided from the manufacturer. Participants were not paid for their participation.
We recruited participants for the case group who were eligible for single-agent methotrexate treatment according to our local institutional protocols: hemodynamically stable, serum hCG less than 3,000 international units/L, minimal free fluid in the pelvis, gestational sac size of less than 4 cm and no fetal cardiac activity on ultrasonogram, and normal baseline (day 1) liver, renal, and hematologic indices. The upper cutoff of less than 3,000 international units/L was chosen because it is the current cutoff for eligibility for medical management at both enrolling institutions. We could not ethically propose a higher cutoff for this phase I study. We did not recruit women in whom there was a diagnosis or suspicion of a ruptured ectopic pregnancy.
The clinical definition of an ectopic pregnancy for this trial was the presence of a positive serum hCG and ultrasonographic findings of 1) a definite tubal ectopic pregnancy (extrauterine gestational sac with yolk sac, embryo, or both with or without cardiac activity and an empty uterus); or 2) a highly probable tubal ectopic pregnancy, defined as an inhomogeneous adnexal mass or extrauterine sac-like structure and an empty uterus.
Given the side effect profile of gefitinib,12 we excluded women with a history of significant respiratory, gastrointestinal, or dermatologic illnesses. We also excluded women of Japanese ethnicity because the incidence of interstitial lung disease has been reported to be higher among Japanese people. We initially excluded smokers when we started the study, but amended our protocol study to include them (amendment approved by our ethics committees). Written informed consent was obtained from all participants and human research ethics approvals were obtained at both sites before recruitment started (Monash Health Human Research Ethics Committee B, Monash Medical Centre, Melbourne, Australia, project number 10142B; and Scotland A Research Ethics Committee, Edinburgh, Scotland, project number 11/MRE00/2).
We administered 50 mg/m2 intramuscular methotrexate on day 1 and daily oral 250 mg gefitinib using a dose-escalation protocol. The first three participants received 250 mg gefitinib on day 1, the next three received 250 mg per day of gefitinib for 3 days, and the last six participants received 250 mg per day of gefitinib for 7 days.
Our primary outcome was safety and tolerability. Women were assessed clinically (history and examination) and biochemically (full blood count [hemoglobin, white cell count, and platelet count], renal function test [serum electrolytes, urea, and creatinine], and liver enzymes) on days 4, 7, and 11, then weekly until the ectopic pregnancy resolved (hCG<5 international units/L). All adverse events were documented and classified according to the Common Terminology Criteria for Adverse Events version 4.03 (National Cancer Institute, National Institutes of Health, U.S. Department of Health and Human Services, June 14, 2010). Participants were contacted at 3 months and 6 months posttreatment to document the return of menstrual cycles and any subsequent pregnancies. We offered screening for chlamydia and gonorrhea by polymerase chain reaction of the urine sample at the time of recruitment.
To monitor treatment response, we followed protocols commonly used for single-dose methotrexate. Serum hCG levels were measured on days 4, 7, and 11 then weekly until serum hCG levels resolved (<5 international units/L). No further treatment was administered if serum hCG fell 15% or more between days 4 and 7 of treatment and continued to decline thereafter. If the serum hCG did not fall 15% or more between days 4 and 7, we administered a second dose of methotrexate. If ectopic pregnancy ruptured or uncontrolled internal bleeding was suspected, we offered prompt surgical management.
We performed a planned subgroup analysis in which pretreatment hCG levels were between 1,000 and 3,000 international units/L, comparing participants successfully treated and women treated at our institutions with single-agent methotrexate (historic controls). These historic controls were women presenting to our institutions over the past 5 years and treated with methotrexate in which there was strict adherence to the treatment protocol, including a record of all serum hCGs until resolution of the ectopic pregnancy (including a record of serum hCG levels at days 1, 4, and 7). We compared pretreatment serum hCG levels and at days 4 and 7 and time to resolution (in days) between trial participants and historic controls.
The treatment protocol used for the historic controls (including inclusion criteria, dose of methotrexate, and hCG monitoring) was the same as that used for trial participants, except no gefitinib was administered and biochemical tests of organ function (full blood count, renal function tests, and liver enzymes) were performed only on days 1 and 7.
We used descriptive statistics to describe our data. The Mann-Whitney U test was used to compare serum hCG levels between participants and historic controls at days 1, 4, and 7 and time taken for the ectopic pregnancies to resolve. We used parametric tests or Fisher’s exact test to compare baseline clinical characteristics between participants and those in the control group. Statistical analysis was performed using GraphPad Prism Version 5 (La Jolla, CA).
One hundred eighty-four women were diagnosed with ectopic pregnancies at the two recruiting institutions during the period of this study. Fifty-seven of these were referred for medical treatment and potentially suitable for recruitment (ie, fulfilled the protocol criteria for medical management with methotrexate at our institutions, the same criteria we used for possible recruitment to our study).
Participation depended on whether any of the three investigators involved in recruiting were available for both recruitment and the anticipated length of follow-up. Of the 57 patients referred for medical management, an investigator was available to approach 25 women (32 were not approached). Of these 25, six declined, seven were excluded, and 12 were recruited. We therefore recruited 12 out of 44 possible participants (27%). Figure 1 provides further details regarding the reasons for exclusions and nonrecruitment.
The mean (±standard deviation) age of participants was 31.5 (5.9) years, and the mean (±standard deviation) body mass index (calculated as weight (kg)/[height (m)]2) was 26.7 (6.3). Nine (75%) had previously been pregnant, two (17%) had previously been treated for an ectopic pregnancy, and two (17%) were smokers. All conceptions were spontaneous and no participants reported a history of endometriosis or sexually transmitted infections. Chlamydia and gonorrhea screening performed during the study was negative for all participants.
Combination treatment did not cause significant toxicities (Table 1). The most common side effects were transient acneiform rash in 67% (8/12) and diarrhea in 42% (5/12), known side effects of gefitinib. There was no clinical or biochemical evidence of serious pulmonary, renal, hepatic, or hematologic toxicity. All participants promptly resumed their menstrual cycles.
Combination gefitinib and methotrexate resolved the ectopic pregnancy in 10 of 12 (83%) participants (Table 2). Of the two participants with abdominal pain and offered laparoscopic salpingectomy for possible tubal rupture, one was confirmed as ruptured at operation and on histopathology. Only one of the 10 women successfully treated with the combination treatment required a second dose of methotrexate.
In a planned subgroup analysis, we compared treatment efficacy between trial participants whose pretreatment serum hCG levels were between 1,000 and 3,000 international units/L (and successfully treated without requiring surgery, n=6) and 71 women presenting to our institutions with similar pretreatment serum hCGs levels (1,000–3,000 international units/L) and successfully managed with single-agent methotrexate without requiring surgery (historic controls).
There were no significant differences in baseline characteristics between the subgroup of six participants and the 71 historic controls. The groups were similar in maternal age (mean±standard error of the mean 32.8±3.0 years in participants compared with 30.7±0.7 years in participants in the control group; P=.5), parity (33% primigravid women among participants compared with 56% among participants in the control group; P=.4), previous ectopic pregnancy (33% among participants compared with 16% participants in the control group; P=.3), history of pelvic inflammatory disease (0% among participants compared with 18% participants in the control group; P=.6), and smoking status (17% among participants compared with 28% participants in the control group; P=1.0).
The serum hCG levels over time are shown in Figure 2. There were no differences in pretreatment or day 1 serum hCG levels (median [interquartile range] hCG in participants was 2,190 [1,811–2,507] international units/L compared with 1,646 [1,357–2,212] international units/L for historic controls; P=.17). Median hCG levels among participants dropped by almost one third between days 1 and 4. In contrast, hCG levels among historic controls rose between days 1 and 4. At day 4, median hCG levels among participants were significantly lower compared with participants in the control group (median [interquartile range] hCG in participants 785 [204–2,047] international units/L compared with participants in the control group 1,838 [1,500–2,649] international units/L; P=.017). By day 7, median serum hCGs were less than one fifth of levels observed among participants in the control group (median [interquartile range] hCG in participants 261 [55–1,445] international units/L compared with participants in the control group 1,426 [940–2,573]; P=.008).
The median time for the ectopic pregnancies to resolve (serum hCG<5 international units/L) was 11 days shorter (34% shorter) among trial participants compared with participants in the control group (participants 21 days (interquartile range 17–27 days) compared with participants in the control group 32 days (interquartile range 25–49 days, P=.018). One participant successfully treated for an ectopic pregnancy in her only remaining fallopian tube (previous unilateral salpingectomy for ruptured ectopic pregnancy) later conceived spontaneously and delivered a healthy neonate at term.
Here we report a phase I study, administering the combination of gefitinib and methotrexate to 12 women with ectopic pregnancies. The combination commonly caused transient diarrhea and rash but did not cause significant biochemical abnormalities to the liver, hematologic, and renal systems. Notably, one participant with an ectopic pregnancy in her only remaining fallopian tube was successfully treated and subsequently conceived a spontaneous intrauterine pregnancy. This provides encouraging evidence that fallopian tubes affected by ectopic pregnancy and exposed to combination gefitinib and methotrexate treatment remain functional. If so, it compares favorably with surgical excision where often, the entire tube is removed (salpingectomy).
Furthermore, we have obtained encouraging efficacy data suggesting the addition of oral gefitinib to methotrexate may considerably enhance its efficacy. In a planned subgroup analysis, trial participants with pretreatment serum hCG levels between 1,000 and 3,000 international units/L had a significantly more rapid decline in hCG levels and were cured significantly faster (34% less time to cure) than a historic cohort receiving methotrexate alone.
Our analysis of efficacy in this trial is a preliminary finding based on small numbers and was compared with a historic control group rather than a randomized comparison trial arm. As such, it requires validation. However, we believe our preliminary observations on potential efficacy are considerably strengthened when considered together with our preclinical data.14 We have generated laboratory data, both in vitro and in animal models, suggesting adding gefitinib to methotrexate enhances its ability to induce placental cell death. Therefore, the possibility that gefitinib may enhance the ability of methotrexate to resolve ectopic pregnancy is biologically plausible. We have started a phase II trial to further investigate efficacy where we are including ectopic pregnancies of larger size that are currently managed surgically (Australian Trials Registration No. 12611001056987).
If verified in future trials, a significantly faster time to cure with gefitinib and methotrexate (compared with methotrexate alone) may not only be clinically beneficial, but could make medical treatment of ectopic pregnancies the preferred option economically. In a systematic meta-analysis, Mol et al3 found when compared with laparoscopic surgery, the single-dose methotrexate protocol resulted in significant cost savings (of approximately $518 per patient) if only one dose was required. These savings resulted from both reduced operating room use and hospital stay. However, savings for the medical option were lost if the pretreatment serum hCG levels were higher than 1,500 international units/L as a result of the need for prolonged follow-up and a higher rate of surgical reinterventions.
It is well known that transient diarrhea and skin rash are common side effects of gefitinib.15,16 These side effects were commonly observed among our cohort and probably occur more often than treatment with single-agent methotrexate. These side effects were transient and were responsive to treatments for symptomatic relief such as topical skin emollients or antidiarrheal therapies. It is possible the inconvenience of enduring these side effects may be acceptable to patients if it were confirmed in future trials that combination therapy provides a faster time to cure and is more efficacious in resolving larger ectopic pregnancies compared with single-agent methotrexate. Because these minor side effects occurred commonly, future randomized studies should include evaluations of participant satisfaction.
Gefitinib is associated with a rare but serious side effect of interstitial lung disease, a thickening of the lung parenchyma. Postmarketing surveillance data of 31,045 patients in the United States reported to the Food and Drug Administration suggests the incidence of interstitial lung disease among those taking gefitinib indefinitely is 0.3%.12 Interestingly, there appears to be a 1% incidence of interstitial lung disease among Japanese people, and we therefore excluded women of this ethnic background.12 Of note, the median time to develop interstitial lung disease while taking gefitinib is 42 days, and risk factors for developing interstitial lung disease include male sex and age older than 55 years.12 Importantly, it is possible coexistent lung pathology needs to be present for gefitinib to induce interstitial lung disease. This premise is supported by the fact trials examining cetuximab treatment (monoclonal antibody that blocks the EGF receptor) for colon cancer did not report the occurrence of interstitial lung disease.17 For these reasons, we believe the administration of a short course of gefitinib to women (non-Japanese descent) of reproductive age without coexisting lung pathology avoids these five risk factors and may be safe. Although we did not observe adverse effects in major organs (lungs, kidney, liver, and hematologic systems), we caution that we have not conclusively shown this regimen is safe in this small trial. In a phase II trial (in progress), we are continuing to collect data on safety and tolerability.
Combination gefitinib and methotrexate may have other pregnancy-related indications such as improving the treatment of persistent gestational trophoblastic disease,18 aiding in the conservative management of placenta accreta (placenta pathologically adherent to the myometrium), and in the treatment of ectopic pregnancies occurring at sites other than the fallopian tube (eg, the cesarean delivery scar in the uterus, the cervix, or the uterine cornua). These are particularly dangerous situations and notoriously difficult to manage even surgically, often requiring hysterectomy.19
In conclusion, we have undertaken a phase I trial suggesting the combination of gefitinib and methotrexate is potentially more effective in resolving ectopic pregnancies than methotrexate alone. The combination could prove to be a potent therapy that not only improves the medical treatment of ectopic pregnancy, but also other disorders of trophoblastic growth. However, this treatment should not be used until its efficacy and safety is confirmed in larger trials.
1. Centers for Disease Control and Prevention (CDC). Ectopic pregnancy: United States, 1990–1992. MMWR Morb Mortal Wkly Rep 1995;44:46–8.
2. The Confidential Enquiry into Maternal and Child Health (CEMACH). Saving mother's lives: reviewing maternal deaths to make motherhood safer 2003–2005. The Seventh Report on Confidential Enquiries into Maternal Deaths in the United Kingdom. London (United Kingdom): 2007:8.
3. Mol F, Mol BW, Ankum WM, van der Veen F, Hajenius PJ. Current evidence on surgery, systemic methotrexate and expectant management in the treatment of tubal ectopic pregnancy: a systematic review and meta-analysis. Hum Reprod Update 2008;14:309–19.
4. Jurkovic D, Wilkinson H. Diagnosis and management of ectopic pregnancy. BMJ 2011;342:d3397.
5. Herbst RS, Fukuoka M, Baselga J. Gefitinib—a novel targeted approach to treating cancer. Nat Rev Cancer 2004;4:956–65.
6. Su AI, Cooke MP, Ching KA, Hakak Y, Walker JR, Wiltshire T, et al.. Large-scale analysis of the human and mouse transcriptomes. Proc Natl Acad Sci U S A 2002;99:4465–70.
7. Wu C, Orozco C, Boyer J, Leglise M, Goodale J, Batalov S, et al.. BioGPS: an extensible and customizable portal for querying and organizing gene annotation resources. Genome Biol 2009;10:R130.
8. LaMarca HL, Dash PR, Vishnuthevan K, Harvey E, Sullivan DE, Morris CA, et al.. Epidermal growth factor-stimulated extravillous cytotrophoblast motility is mediated by the activation of PI3-K, Akt and both p38 and p42/44 mitogen-activated protein kinases. Hum Reprod 2008;23:1733–41.
9. Johnstone ED, Mackova M, Das S, Payne SG, Lowen B, Sibley CP, et al.. Multiple anti-apoptotic pathways stimulated by EGF in cytotrophoblasts. Placenta 2005;26:548–55.
10. Wolff GS, Chiang PJ, Smith SM, Romero R, Armant DR. Epidermal growth factor-like growth factors prevent apoptosis of alcohol-exposed human placental cytotrophoblast cells. Biol Reprod 2007;77:53–60.
11. Sibilia M, Wagner EF. Strain-dependent epithelial defects in mice lacking the EGF receptor. Science 1995;269:234–8.
12. Cohen MH, Williams GA, Sridhara R, Chen G, McGuinn WD Jr, Morse D, et al.. United States Food and Drug Administration Drug Approval summary: gefitinib (ZD1839; Iressa) tablets. Clin Cancer Res 2004;10:1212–8.
13. Ciardiello F, Tortora G. EGFR antagonists in cancer treatment. N Engl J Med 2008;358:1160–74.
14. Nilsson UW, Johns TG, Wilmann T, Kaitu’u-Lino T, Whitehead C, Dimitriadis E, et al.. Effects of gefitinib, an epidermal growth factor receptor inhibitor, on human placental cell growth. Obstet Gynecol 2013;122:737–44.
15. van Zandwijk N. Tolerability of gefitinib in patients receiving treatment in everyday clinical practice. Br J Cancer 2003;89(suppl 2):S9–14.
16. Cataldo VD, Gibbons DL, Perez-Soler R, Quintas-Cardama A. Treatment of non-small-cell lung cancer with erlotinib or gefitinib. N Engl J Med 2011;364:947–55.
17. Hoag JB, Azizi A, Doherty TJ, Lu J, Willis RE, Lund ME. Association of cetuximab with adverse pulmonary events in cancer patients: a comprehensive review. J Exp Clin Cancer Res 2009;28:113.
18. Seckl MJ, Sebire NJ, Berkowitz RS. Gestational trophoblastic disease. Lancet 2010;376:717–29.
19. Chetty M, Elson J. Treating non-tubal ectopic pregnancy. Best Pract Res Clin Obstet Gynaecol 2009;23:529–38.