- Email: [email protected]
Effect of 17α-hydroxyprogesterone caproate before embryo transfer on the outcome of in vitro fertilization and embryo transfer: a randomized trial
IN VITRO FERTILIZATION Effect of 17a-hydroxyprogesterone caproate before embryo transfer on the outcome of in vitro fertilization and embryo transfer: a randomized trial Antoine Abu-Musa, M.D., Ph.D., Ihab Usta, M.D., Anwar Nassar, M.D., Fatiha Hajami, M.D., and Antoine Hannoun, M.D. Department of Obstetrics and Gynecology, American University of Beirut Medical Center, Beirut, Lebanon
Objective: To assess the effect of 17a-hydroxyprogesterone caproate (17-HPC) given before embryo transfer on the pregnancy outcome of IVF–embryo transfer (ET) cycles. Design: Randomized controlled study. Setting: A university-based hospital IVF unit. Patient(s): One hundred twenty-five consecutive patients undergoing IVF-ET were randomly assigned into treatment and control groups. Intervention(s): In the treatment group, 63 patients received 17-HPC (250 mg, IM), 1 day before ET. The control group consisted of 62 patients who did not receive any injections. Main Outcome Measure(s): Pregnancy and multiple-pregnancy rates. Result(s): The two groups were similar with respect to the age of patients, total dose of FSH, number of oocytes and embryos obtained, and number and quality of embryos transferred. There was no significant difference in the pregnancy rate (34.9% vs. 38.7%) or in the rate of multiple gestation (15.9% vs. 9.7%) between cases and controls, respectively. Conclusion(s): The use of 17-HPC before ET does not appear to affect the outcome of IVF-ET. (Fertil Steril 2008;89:1098–102. 2008 by American Society for Reproductive Medicine.) Key Words: 17a-hydroxyprogesterone caproate, IVF-ET, pregnancy outcome
The attempt to improve the implantation process in IVF– embryo transfer (ET) programs has been the subject of numerous studies. Most of these studies have focused on the induction and selection of the best quality embryos and on the improvement in uterine receptivity. Several developments in controlled ovarian hyperstimulation (COH), fertilization, and embryo culture techniques have contributed to the improvement of the quality of embryos that are available for ET. In contrast, little has been achieved in improving uterine receptivity. The evaluation and improvement of uterine receptivity are limited by the fact that the process of implantation is regulated by multiple and complex morphological and biochemical factors (1). Thus, all of these factors should be investigated simultaneously to clearly assess the receptivity status of the endometrium. In addition, tissue sampling for the direct assessment of markers of uterine receptivity is inherently impossible in the actual ET cycles. Recently, the direct visualization of the uterine contractile activity by using high-resolution ultrasound has been evaluReceived April 25, 2007; revised and accepted May 29, 2007. Reprint requests: Antoine Abu-Musa, M.D., Ph.D., P.O. Box 113-6044-6A, Department of Obstetrics and Gynecology, American University of Beirut Medical Center, Beirut, Lebanon (E-mail: [email protected]).
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ated as a noninvasive prognostic factor of uterine receptivity (2, 3). The uterus has typically three patterns of contractility throughout the menstrual cycle that influence embryo implantation (4–7). Increased uterine contractility at the time of ET has been shown to adversely affect embryo implantation and pregnancy rates in IVF, probably because of the mechanical expulsion of embryos from the uterine cavity (8). This suggests that uterine contractility can be an important factor in determining endometrial receptivity. Embryo transfer itself can provoke a uterine response as a result of stimulation of the cervix and intracavitary canalization. In this respect, it has been suggested that the use of adjuvants, such as uterine relaxants, to decrease uterine contractility during ET should be considered to prime a uterus and make it suitable for embryo implantation (9). Progesterone acts to relax smooth muscles in many organs, including the uterus. 17a-Hydroxyprogesterone caproate is a P derivative. Esterification of 17a- hydroxyprogesterone with caproic acid provides much greater and more prolonged progestational activity after IM injection. The aim of this study was to examine the effect of 17-HPC treatment for priming of the uterus on the pregnancy outcome in IVF-ET cycles. The hypothesis was that 17-HPC would improve the pregnancy rate in IVF-ET cycles.
Fertility and Sterility Vol. 89, No. 5, May 2008 Copyright ª2008 American Society for Reproductive Medicine, Published by Elsevier Inc.
0015-0282/08/$34.00 doi:10.1016/j.fertnstert.2007.05.043
MATERIALS AND METHODS This study was performed on 125 consecutive patients undergoing IVF-ET at the IVF Unit of the American University of Beirut Medical Center. All patients who presented for their first IVF cycle at our center were eligible to participate in the study. The study included women who underwent IVF between June and December 2006 because of tubal, malefactor infertility, unexplained, polycystic ovary syndrome, or endometriosis factors. Tubal-factor infertility was based on laparoscopic diagnosis of severe pelvic adhesions or blocked tubes. Male-factor infertility was considered when at least two semen analyses were abnormal according to the World Health Organization laboratory manual (10). Patients were considered to have unexplained infertility if their workup was negative, including normal semen analysis, patent tubes confirmed by either a hysterosalpingogram or laparoscopy, and ovulatory cycles. The diagnosis of polycystic ovary syndrome was made according to the Rotterdam consensus (11). None of the patients who were eligible for the study declined to participate. After the two IVF specialists (A.A.-M. and A.H.) checked them for eligibility, the 125 patients were randomly divided into treatment and control groups by using computer-generated random tables (generated by A.N., a non-IVF specialist). Randomization was performed after oocyte pickup by using sealed envelopes. The infertility specialist attending to the patients was not blinded to the arm to which patients were assigned; however, the research assistant who entered the data and performed the statistical analysis was. The primary outcome of the study was to assess the difference in pregnancy rate between patients and controls. The study was institutional review board approved, and patients were not entitled to any financial reimbursement.
Controlled Ovarian Stimulation and Intracytoplasmic Sperm Injection Procedures Controlled ovarian hyperstimulation was performed by a long standard protocol with GnRH agonist (GnRH-a; Decapeptyl, Beaufour Ipsen Pharma, France) and recombinant FSH (Puregon, Organon, the Netherlands). In brief, GnRHa was started in the mid-luteal phase at a daily dose of 0.05 mg until the day of hCG injection. Recombinant FSH (200 U/d) was started on the 3rd day of the cycle. Follicular development was assessed by transvaginal ultrasonography. When at least two dominant follicles were R18 mm, hCG (10,000 IU; Pregnyl, Organon) was administered. Oocyte retrieval by transvaginal ultrasonographic guidance was performed approximately 36 hours after the hCG administration. Intracytoplasmic sperm injection was used almost routinely in all our patients regardless of the cause of infertility; however, regular IVF was used in a few patients according to the preference of the attending physician. The ET was performed with a Wallace transfer catheter (Smiths Medical, Hythe, Kent, UK) 2 days after oocyte recovery under ultrasound guidance. The embryos were scored according to a grading Fertility and Sterility
system published elsewhere (12). All patients received vaginal micronized P (200 mg) every 8 hours, starting the afternoon of the oocyte pickup and continuing up to the day of b-hCG measurement; this was maintained throughout the first trimester if the b-hCG was positive. There were two outcome variables in this study. The clinical pregnancy rate was defined as the presence of a positive fetal heart rate, detected by vaginal ultrasound that was performed 20 days after a positive pregnancy test. Multiple pregnancy was defined as the presence of two or more gestational sacs with positive fetal heart rates. Treatment With 17-HPC Patients in the treatment group received a dose of 17-HPC (Proluton depot, 250 mg, IM; Schering, Germany) 24 hours before ET, whereas patients in the control group did not receive any injections. Statistical Analysis Data entry was performed by using SPSS software (SPSS for Windows XP, version 12; SPSS, Chicago, IL), and statistical analysis was performed by using the Mann-Whitney test for nonparametric variables. For discrete variables, the c2-test was used, and the Fisher’s exact test was used if the expected number in a cell was less than five or when >20% of the expected counts were <5 or if any were <1. Multiple logistic regression was used to identify significant factors affecting clinical pregnancy rates. A P value of < .05 was considered statistically significant. Odds ratios (ORs) for outcome variables, with 95% confidence intervals (CIs), were cited where appropriate. RESULTS Cases and controls were similar with respect to the age of patients, total dose of FSH, number of oocytes and embryos obtained, and number and quality of embryos transferred (Table 1). However, the percentage of patients undergoing intracytoplasmic sperm injection was significantly lower in controls, and the percentage of patients R40 years of age tended to be higher in controls. This is believed to be entirely due to chance, because the study was a randomized trial. The two groups had similar pregnancy rates (OR, 0.85; 95% CI, 0.41–1.75; Table 2). Although there was a tendency for an increase in the rate of multiple gestation in the 17-HPC group (45.5% vs. 25.0%), this difference did not reach statistical significance (OR, 1.76; 95% CI, 0.62–5.00). When the pregnancy rates were compared according to the causes of infertility, there was no significant difference between the treatment and control groups. There was also no significant difference in total pregnancy rates between the two groups when they were subgrouped according to age (Table 3). Multiple logistic regression including factors of age of R40 years, Proluton, and intracytoplasmic sperm injection showed no significant association between Proluton 1099
TABLE 1 Patient characteristics. Characteristic
Treatment group (n [ 63)
Control group (n [ 62)
33 (19–46) 2,400 (1,200–5,850) 10 (2–36) 6 (1–24) 3 (1–4) 3 (0–12)
33.5 (19–47) 2,400 (1,100–5,850) 9 (1–33) 6 (1–18) 3 (1–4) 3 (0–9)
Age (y) Dose of FSH (IU) No. of oocytes No. of embryos No. of embryos transferred No. of grade A embryos
Note: Data are presented as median (range). Medians were compared by using the Mann-Whitney test, and percentages were compared by using the c2 test or Fisher’s exact tests. ICSI ¼ intracytoplasmic sperm injection. Abu-Musa. 17a-hydroxyprogesterone caproate in IVF. Fertil Steril 2008.
injection (OR, 0.72; 95% CI, 0.33–1.59; P¼.417) or intracytoplasmic sperm injection (OR, 1.07; 95% CI, 0.36–3.20; P¼.91) and clinical pregnancy rate. Woman’s age of R40 years was the only significant factor that was associated with decreased clinical pregnancy (OR, 0.19; 95% CI, 0.04–0.90; P¼.036). Twelve of the 63 patients (19.0%) who received 17-HPC had minimal local pain and irritation at the site of the injection. This did not require medical attention or intervention. DISCUSSION Implantation failure remains the main limiting factor for success in IVF cycles. This failure is due, in part, to uterus receptivity at the time of ET. Uterine contractility has been shown to be an important factor in determining uterine receptivity. The effect of uterine contractility on human fecundity and, notably, on embryo implantation in IVF-ET cycles, has been evaluated in recent studies. Uterine contractions (UCs) follow three characteristic patterns during the men-
TABLE 2 Clinical pregnancies.
Parameter
Treatment group (n [ 63)
Control group (n [ 62)
Clinical 22 (34.9) 24 (38.7) pregnancy (%) No. of sacs (%) Singleton 12 (19.0) 18 (29.0) Twins 9 (14.3) 4 (6.5) Triplets — — Quadruplets 1 (1.6) 2 (3.2) Multiple 10/22 (45.5) 6/24 (25.0) gestation (%)
P value .80
.19 .24 .62 .25
strual cycle. During the early follicular phase (menses), UCs are vigorous, with predominant antegrade (fundusto-cervix) displacement. This pattern is instrumental in the forward emptying of uterine contents after sloughing of the endometrium upon P withdrawal (13). During the late follicular phase, uterine contractility increases under the influence of rising E2 levels, with progressive predominance of retrograde (cervix-to-fundus) displacement. This pattern of uterine contractility has been implicated in the rapid transport of sperm toward the distal end of the fallopian tubes that occurs after intercourse (14). During the luteal phase, uterine contractility characteristically decreases, which is a phenomenon that is crucial for maximizing uterine receptivity to embryo implantation (8). IJland et al. (5) found in spontaneous cycles that conceptional cycles had less endometrial wavelike activity compared with nonconception cycles. Increased uterine contractility at the time of ET is associated with poorer implantation (7) and poorer IVF-ET outcome (15), probably because of the mechanical expulsion of embryos from the uterine cavity. These results suggest that uterine contractility influences embryo implantation. Therefore, it is logical to seek a rapid relaxation of the myometrium, which may prevent the premature expulsion of embryos from the uterine cavity at the time of ET and potentially improve pregnancy rates. A recent
TABLE 3 Comparison of pregnancy rates with age of patients. Age (y)
Treatment Control group (%) group (%)
%30 13/25 (52.0) 9/19 (47.4) 31–35 5/15 (33.3) 8/14 (57.1) 35–40 4/19 (21.1) 5/17 (29.4) R40 0/4 (0) 2/12 (16.7)
OR (95% CI) 1.20 (0.37–3.90) 0.38 (0.09–1.64) 0.64 (0.15–2.75) 0.00 (0.00–6.41)
Note: c2 or Fisher’s exact tests were used.
Note: c2 or Fisher’s exact tests were used.
Abu-Musa. 17a-hydroxyprogesterone caproate in IVF. Fertil Steril 2008.
Abu-Musa. 17a-hydroxyprogesterone caproate in IVF. Fertil Steril 2008.
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study showed that the administration of piroxicam, a nonsteroidal anti-inflammatory drug, just before ET increased the implantation and pregnancy rates after IVF-ET (9). Prostaglandin, which is synthesized from arachidonic acid by cyclooxygenase, stimulates UCs. Nonsteroidal anti-inflammatory drugs block the action of cyclooxygenase and inhibit the production of prostaglandin (16), which may cause a reduction in uterine contractility. Few studies have evaluated the role of vaginal P on UC and IVF outcome (17, 18). These researchers administered vaginal P starting on the day of oocyte retrieval to relax uterine contractility at the time of ET. As a result, they found a decrease in UC frequency on the day of ET, suggesting that a uterine-relaxing effect by P administration before ET is likely to improve IVF-ET outcome. When vaginal P was given on the evening of ET as opposed to on the day of oocyte pickup, uterine contractility remained exacerbated despite high mean serum P levels observed at the time of ET. This indicates that during the early luteal phase of COH, the uterinerelaxation process that is mediated by P may take longer than during the corresponding phase of the natural menstrual cycle (19–21). Hence, in COH, the achievement of rapid myometrial relaxation appears to require a more intense uterine exposure to P than is the case in the regular menstrual cycle. This suggests a degree of resistance to the uterine-relaxing action of endogenous P that may be attributable to the persisting uterine stimulation by the supraphysiological E2 levels of COH (17). In our study, the aim of giving a 17-HPC, which has a potent and prolonged progestational activity, was to achieve this intense uterine exposure to P and thus a rapid relaxation of the myometrium, which may prevent the premature expulsion of embryos from the uterine cavity at the time of ET and improve pregnancy rates. The role of 17HPC for luteal support in patients undergoing IVF-ET cycles has been evaluated in very few studies (22–25). Some of these studies have shown no beneficial effect of 17-HPC on the outcome of IVF-ET (22, 23). However, other investigators have reported a higher pregnancy rate when 17-HPC was used, compared with placebo or intravaginal P (24, 25). In our study, although there was no difference in pregnancy rate between cases and controls, there was a tendency for increase in the rate of twin gestation in patients who received 17-HPC; however, this did not reach statistical significance. The main limitation of this study is the small sample size. When pregnancy rate is taken as the primary outcome and assuming a pregnancy rate of 35% with IVF treatment, a 10% increase in pregnancy rate with 17-HPC treatment would require a sample size of 376 subjects in each arm, with an a of 0.05 and b of 80%. To detect a significant effect of 17-HPC on the implantation rate, assuming an implantation rate of 19% in untreated women and an effect associated with 17HPC of 10%, 286 women in each arm would be required to achieve a power of 80% and a level of significance of 95%. Another limitation of this study is that uterine contractility and 17-HPC levels (using high-pressure liquid chromatography or RIA) were not assessed on the day of ET. In one study, the mean serum level of 17-HPC after a single IM injection of Fertility and Sterility
1,000 mg reached its maximum (44–81 nmol/L) after 2–7 days (26). Costabile et al. (23) have demonstrated a slow increase in plasma level until the 5th day that was caused by a retarded absorption from the injection site. The timing of administration may need to be modified to R48 hours before ET, allowing for 17-HPC to reach significant blood levels. Alternatively, the dose of 17-HPC may need to be increased to achieve the required uterine relaxation. We did not use the same doses of 17-HPC that were used by other investigators (23–25), because in these studies, 17-HPC was used alone for the luteal support. In our study, 17-HPC was used as a bolus dose of P in addition to the standard vaginal P dose that we use for luteal support. It may be speculated that the benefit of 17-HPC may be reduced or masked by concurrent administration of vaginal P, which is given routinely to IVF patients in our center. In conclusion, preliminary evidence from this study shows that the addition of an IM progestational agent that has potent smooth-muscular relaxant properties to vaginal micronized P before ET does not appear to improve the pregnancy rate in IVF-ET cycles. REFERENCES 1. Tabibzadeh S, Babaknia A. The signals and molecular pathways involved in implantation, a symbiotic interaction between blastocyst and endometrium involving adhesion and tissue invasion. Hum Reprod 1995;10:1579–602. 2. Oike K, Obata S, Tagaki K, Matsuo K, Ishihara K, Kikuchi S. Observation of endometrial movement with transvaginal sonography. J Ultrasound Med 1988;7:S99. 3. Zaidi J, Pittrof R, Shaker A, Kyei-Mensah A, Campbell S, Tan SL. Assessment of uterine artery blood flow on the day of human chorionic gonadotropin administration by transvaginal color Doppler ultrasound in an in vitro fertilization program. Fertil Steril 1996;65:377–81. 4. IJland MM, Evers JLH, Dunselman GAJ, Van Katwijk C, Lo CR, Hoogland HJ. Endometrial wavelike movements during menstrual cycle. Fertil Steril 1996;65:746–9. 5. IJland MM, Evers JLH, Dunselman GAJ, Volovics L, Lo CR, Hoogland HJ. Relation between endometrial wavelike activity and fecundability in spontaneous cycles. Fertil Steril 1997;67:492–6. 6. IJland MM, Evers JLH, Dunselman GAJ, Lo CR, Hoogland HJ. Endometrial wavelike activity, endometrial thickness, and ultrasound texture in controlled ovarian hyperstimulation cycles. Fertil Steril 1998;70:279–83. 7. Bulletti C, Prefetto RA, Bazzocchi G, Romero R, Mimmi P, Polli V, et al. Electromechanical activities of human uteri during extra-corporeal perfusion with ovarian steroids. Hum Reprod 1993;8:1558–63. 8. Fanchin R, Righini C, Olivennes F, Taylor S, Ziegler D, Frydman R. Uterine contractions at the time of embryo transfer alter pregnancy rates after in-vitro fertilization. Hum Reprod 1998;13:1968–74. 9. Moon HS, Park SH, Lee JO, Kim KS, Joo BS. Treatment with piroxicam before embryo transfer increases the pregnancy rate after in vitro fertilization and embryo transfer. Fertil Steril 2004;82:816–20. 10. World Health Organization. WHO laboratory manual for the examination of human semen and sperm-cervical mucus interaction. 4th ed. New York: Cambridge University Press, 1999. 11. The Rotterdam ESHRE/ASRM-sponsored PCOS consensus workshop group. Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome (PCOS). Hum Reprod 2004;19:41–7. 12. Steer CV, Mills CL, Tan SL, Campbell S, Edwards RG. The cumulative embryo score: a predictive embryo scoring technique to select the optimal number of embryos to transfer in an in-vitro fertilization and embryo transfer programme. Hum Reprod 1992;7:117–9.
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13. Salamanca A, Beltran E. Subendometrial contractility in menstrual phase visualized by transvaginal sonography in patients with endometriosis. Fertil Steril 1995;64:193–5. 14. Leyendecker G, Kunz G, Wildt L, Beil D, Deininger H. Uterine hyperperistalsis and dysperistalsis as dysfunctions of the mechanism of rapid sperm transport in patients with endometriosis and infertility. Hum Reprod 1996;11:1542–51. 15. Fanchin R, Righini C, Ayoubi JM, Olivennes F, Ziegler D, Frydman R. New look at endometrial echogenicity: objective computer-assisted measurements predict endometrial receptivity in in vitro fertilization embryo transfer. Fertil Steril 2000;74:274–81. 16. Dawood MY. Nonsteroidal antiinflammatory drugs and reproduction. Am J Obstet Gynecol 1993;169:1255–65. 17. Fanchin R, Righini C, Ziegler D, Olivennes F, Ledee N, Frydman R. Effects of vaginal progesterone administration on uterine contractility at the time of embryo transfer. Fertil Steril 2001;75:1136–40. 18. Ayoubi JM, Fanchin R, Kaddouz D, Frydman R, Ziegler D. Uterorelaxing effects of vaginal progesterone: comparison of two methodologies for assessing uterine contraction frequency on ultrasound scans. Fertil Steril 2001;76:736–40. 19. Abramowicz JS, Archer DF. Uterine endometrial peristalsis-a transvaginal ultrasound study. Fertil Steril 1990;54:451–4. 20. Lyons EA, Taylor PJ, Zheng XH, Ballard G, Levi CS, Kredentser JV. Characterization of subendometrial myometrial contractions throughout
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the menstrual cycle in normal fertile women. Fertil Steril 1991;55: 771–4. Fanchin R, Ayoubi JM, Olivennes F, Righini C, de Ziegler D, Frydman R. Hormonal influence on the uterine contractility during controlled ovarian hyperstimulation. Hum Reprod 2000;15(Suppl 1):90–100. Leeton J, Trounson A, Jessup D. Support of the luteal phase in in vitro fertilization programs: results of a controlled trial with intramuscular Proluton. J In Vitro Fert Embryo Transf 1985;2:166–9. Costabile L, Gerli S, Manna C, Rossetti D, Di Renzo GC, Unfer V. A prospective randomized study comparing intramuscular progesterone 17 a-hydroxyprogesterone caproate in patients undergoing in vitro fertilization-embryo transfer cycles. Fertil Steril 2001;76:394–6. Abate A, Brigandi A, Abate FG, Manti F, Unfer V, Perino M. Luteal phase support with 17alpha-hydroxyprogesterone versus unsupported cycles in in vitro fertilization: a comparative randomized study. Gynecol Obstet Invest 1999;48:78–80. Unfer V, Casini ML, Costabile L, Gerli S, Baldini D, Di Renzo GC. 17 alpha-hydroxyprogesterone versus intravaginal progesterone in IVF-embryo transfer cycles: a prospective randomized study. Reprod Biomed Online 2004;9:17–21. Onsrud M, Paus E, Haug E, Kjorstad K. Intramuscular administration of hydroxyprogesterone caproate in patients with endometrial carcinoma. Pharmacokinetics and effects on adrenal function. Acta Obstet Gynecol Scand 1985;64:519–23.
17a-hydroxyprogesterone caproate in IVF
Vol. 89, No. 5, May 2008
Objective: To assess the effect of 17a-hydroxyprogesterone caproate (17-HPC) given before embryo transfer on the pregnancy outcome of IVF–embryo transfer (ET) cycles. Design: Randomized controlled study. Setting: A university-based hospital IVF unit. Patient(s): One hundred twenty-five consecutive patients undergoing IVF-ET were randomly assigned into treatment and control groups. Intervention(s): In the treatment group, 63 patients received 17-HPC (250 mg, IM), 1 day before ET. The control group consisted of 62 patients who did not receive any injections. Main Outcome Measure(s): Pregnancy and multiple-pregnancy rates. Result(s): The two groups were similar with respect to the age of patients, total dose of FSH, number of oocytes and embryos obtained, and number and quality of embryos transferred. There was no significant difference in the pregnancy rate (34.9% vs. 38.7%) or in the rate of multiple gestation (15.9% vs. 9.7%) between cases and controls, respectively. Conclusion(s): The use of 17-HPC before ET does not appear to affect the outcome of IVF-ET. (Fertil Steril 2008;89:1098–102. 2008 by American Society for Reproductive Medicine.) Key Words: 17a-hydroxyprogesterone caproate, IVF-ET, pregnancy outcome
The attempt to improve the implantation process in IVF– embryo transfer (ET) programs has been the subject of numerous studies. Most of these studies have focused on the induction and selection of the best quality embryos and on the improvement in uterine receptivity. Several developments in controlled ovarian hyperstimulation (COH), fertilization, and embryo culture techniques have contributed to the improvement of the quality of embryos that are available for ET. In contrast, little has been achieved in improving uterine receptivity. The evaluation and improvement of uterine receptivity are limited by the fact that the process of implantation is regulated by multiple and complex morphological and biochemical factors (1). Thus, all of these factors should be investigated simultaneously to clearly assess the receptivity status of the endometrium. In addition, tissue sampling for the direct assessment of markers of uterine receptivity is inherently impossible in the actual ET cycles. Recently, the direct visualization of the uterine contractile activity by using high-resolution ultrasound has been evaluReceived April 25, 2007; revised and accepted May 29, 2007. Reprint requests: Antoine Abu-Musa, M.D., Ph.D., P.O. Box 113-6044-6A, Department of Obstetrics and Gynecology, American University of Beirut Medical Center, Beirut, Lebanon (E-mail: [email protected]).
1098
ated as a noninvasive prognostic factor of uterine receptivity (2, 3). The uterus has typically three patterns of contractility throughout the menstrual cycle that influence embryo implantation (4–7). Increased uterine contractility at the time of ET has been shown to adversely affect embryo implantation and pregnancy rates in IVF, probably because of the mechanical expulsion of embryos from the uterine cavity (8). This suggests that uterine contractility can be an important factor in determining endometrial receptivity. Embryo transfer itself can provoke a uterine response as a result of stimulation of the cervix and intracavitary canalization. In this respect, it has been suggested that the use of adjuvants, such as uterine relaxants, to decrease uterine contractility during ET should be considered to prime a uterus and make it suitable for embryo implantation (9). Progesterone acts to relax smooth muscles in many organs, including the uterus. 17a-Hydroxyprogesterone caproate is a P derivative. Esterification of 17a- hydroxyprogesterone with caproic acid provides much greater and more prolonged progestational activity after IM injection. The aim of this study was to examine the effect of 17-HPC treatment for priming of the uterus on the pregnancy outcome in IVF-ET cycles. The hypothesis was that 17-HPC would improve the pregnancy rate in IVF-ET cycles.
Fertility and Sterility Vol. 89, No. 5, May 2008 Copyright ª2008 American Society for Reproductive Medicine, Published by Elsevier Inc.
0015-0282/08/$34.00 doi:10.1016/j.fertnstert.2007.05.043
MATERIALS AND METHODS This study was performed on 125 consecutive patients undergoing IVF-ET at the IVF Unit of the American University of Beirut Medical Center. All patients who presented for their first IVF cycle at our center were eligible to participate in the study. The study included women who underwent IVF between June and December 2006 because of tubal, malefactor infertility, unexplained, polycystic ovary syndrome, or endometriosis factors. Tubal-factor infertility was based on laparoscopic diagnosis of severe pelvic adhesions or blocked tubes. Male-factor infertility was considered when at least two semen analyses were abnormal according to the World Health Organization laboratory manual (10). Patients were considered to have unexplained infertility if their workup was negative, including normal semen analysis, patent tubes confirmed by either a hysterosalpingogram or laparoscopy, and ovulatory cycles. The diagnosis of polycystic ovary syndrome was made according to the Rotterdam consensus (11). None of the patients who were eligible for the study declined to participate. After the two IVF specialists (A.A.-M. and A.H.) checked them for eligibility, the 125 patients were randomly divided into treatment and control groups by using computer-generated random tables (generated by A.N., a non-IVF specialist). Randomization was performed after oocyte pickup by using sealed envelopes. The infertility specialist attending to the patients was not blinded to the arm to which patients were assigned; however, the research assistant who entered the data and performed the statistical analysis was. The primary outcome of the study was to assess the difference in pregnancy rate between patients and controls. The study was institutional review board approved, and patients were not entitled to any financial reimbursement.
Controlled Ovarian Stimulation and Intracytoplasmic Sperm Injection Procedures Controlled ovarian hyperstimulation was performed by a long standard protocol with GnRH agonist (GnRH-a; Decapeptyl, Beaufour Ipsen Pharma, France) and recombinant FSH (Puregon, Organon, the Netherlands). In brief, GnRHa was started in the mid-luteal phase at a daily dose of 0.05 mg until the day of hCG injection. Recombinant FSH (200 U/d) was started on the 3rd day of the cycle. Follicular development was assessed by transvaginal ultrasonography. When at least two dominant follicles were R18 mm, hCG (10,000 IU; Pregnyl, Organon) was administered. Oocyte retrieval by transvaginal ultrasonographic guidance was performed approximately 36 hours after the hCG administration. Intracytoplasmic sperm injection was used almost routinely in all our patients regardless of the cause of infertility; however, regular IVF was used in a few patients according to the preference of the attending physician. The ET was performed with a Wallace transfer catheter (Smiths Medical, Hythe, Kent, UK) 2 days after oocyte recovery under ultrasound guidance. The embryos were scored according to a grading Fertility and Sterility
system published elsewhere (12). All patients received vaginal micronized P (200 mg) every 8 hours, starting the afternoon of the oocyte pickup and continuing up to the day of b-hCG measurement; this was maintained throughout the first trimester if the b-hCG was positive. There were two outcome variables in this study. The clinical pregnancy rate was defined as the presence of a positive fetal heart rate, detected by vaginal ultrasound that was performed 20 days after a positive pregnancy test. Multiple pregnancy was defined as the presence of two or more gestational sacs with positive fetal heart rates. Treatment With 17-HPC Patients in the treatment group received a dose of 17-HPC (Proluton depot, 250 mg, IM; Schering, Germany) 24 hours before ET, whereas patients in the control group did not receive any injections. Statistical Analysis Data entry was performed by using SPSS software (SPSS for Windows XP, version 12; SPSS, Chicago, IL), and statistical analysis was performed by using the Mann-Whitney test for nonparametric variables. For discrete variables, the c2-test was used, and the Fisher’s exact test was used if the expected number in a cell was less than five or when >20% of the expected counts were <5 or if any were <1. Multiple logistic regression was used to identify significant factors affecting clinical pregnancy rates. A P value of < .05 was considered statistically significant. Odds ratios (ORs) for outcome variables, with 95% confidence intervals (CIs), were cited where appropriate. RESULTS Cases and controls were similar with respect to the age of patients, total dose of FSH, number of oocytes and embryos obtained, and number and quality of embryos transferred (Table 1). However, the percentage of patients undergoing intracytoplasmic sperm injection was significantly lower in controls, and the percentage of patients R40 years of age tended to be higher in controls. This is believed to be entirely due to chance, because the study was a randomized trial. The two groups had similar pregnancy rates (OR, 0.85; 95% CI, 0.41–1.75; Table 2). Although there was a tendency for an increase in the rate of multiple gestation in the 17-HPC group (45.5% vs. 25.0%), this difference did not reach statistical significance (OR, 1.76; 95% CI, 0.62–5.00). When the pregnancy rates were compared according to the causes of infertility, there was no significant difference between the treatment and control groups. There was also no significant difference in total pregnancy rates between the two groups when they were subgrouped according to age (Table 3). Multiple logistic regression including factors of age of R40 years, Proluton, and intracytoplasmic sperm injection showed no significant association between Proluton 1099
TABLE 1 Patient characteristics. Characteristic
Treatment group (n [ 63)
Control group (n [ 62)
33 (19–46) 2,400 (1,200–5,850) 10 (2–36) 6 (1–24) 3 (1–4) 3 (0–12)
33.5 (19–47) 2,400 (1,100–5,850) 9 (1–33) 6 (1–18) 3 (1–4) 3 (0–9)
Age (y) Dose of FSH (IU) No. of oocytes No. of embryos No. of embryos transferred No. of grade A embryos
Note: Data are presented as median (range). Medians were compared by using the Mann-Whitney test, and percentages were compared by using the c2 test or Fisher’s exact tests. ICSI ¼ intracytoplasmic sperm injection. Abu-Musa. 17a-hydroxyprogesterone caproate in IVF. Fertil Steril 2008.
injection (OR, 0.72; 95% CI, 0.33–1.59; P¼.417) or intracytoplasmic sperm injection (OR, 1.07; 95% CI, 0.36–3.20; P¼.91) and clinical pregnancy rate. Woman’s age of R40 years was the only significant factor that was associated with decreased clinical pregnancy (OR, 0.19; 95% CI, 0.04–0.90; P¼.036). Twelve of the 63 patients (19.0%) who received 17-HPC had minimal local pain and irritation at the site of the injection. This did not require medical attention or intervention. DISCUSSION Implantation failure remains the main limiting factor for success in IVF cycles. This failure is due, in part, to uterus receptivity at the time of ET. Uterine contractility has been shown to be an important factor in determining uterine receptivity. The effect of uterine contractility on human fecundity and, notably, on embryo implantation in IVF-ET cycles, has been evaluated in recent studies. Uterine contractions (UCs) follow three characteristic patterns during the men-
TABLE 2 Clinical pregnancies.
Parameter
Treatment group (n [ 63)
Control group (n [ 62)
Clinical 22 (34.9) 24 (38.7) pregnancy (%) No. of sacs (%) Singleton 12 (19.0) 18 (29.0) Twins 9 (14.3) 4 (6.5) Triplets — — Quadruplets 1 (1.6) 2 (3.2) Multiple 10/22 (45.5) 6/24 (25.0) gestation (%)
P value .80
.19 .24 .62 .25
strual cycle. During the early follicular phase (menses), UCs are vigorous, with predominant antegrade (fundusto-cervix) displacement. This pattern is instrumental in the forward emptying of uterine contents after sloughing of the endometrium upon P withdrawal (13). During the late follicular phase, uterine contractility increases under the influence of rising E2 levels, with progressive predominance of retrograde (cervix-to-fundus) displacement. This pattern of uterine contractility has been implicated in the rapid transport of sperm toward the distal end of the fallopian tubes that occurs after intercourse (14). During the luteal phase, uterine contractility characteristically decreases, which is a phenomenon that is crucial for maximizing uterine receptivity to embryo implantation (8). IJland et al. (5) found in spontaneous cycles that conceptional cycles had less endometrial wavelike activity compared with nonconception cycles. Increased uterine contractility at the time of ET is associated with poorer implantation (7) and poorer IVF-ET outcome (15), probably because of the mechanical expulsion of embryos from the uterine cavity. These results suggest that uterine contractility influences embryo implantation. Therefore, it is logical to seek a rapid relaxation of the myometrium, which may prevent the premature expulsion of embryos from the uterine cavity at the time of ET and potentially improve pregnancy rates. A recent
TABLE 3 Comparison of pregnancy rates with age of patients. Age (y)
Treatment Control group (%) group (%)
%30 13/25 (52.0) 9/19 (47.4) 31–35 5/15 (33.3) 8/14 (57.1) 35–40 4/19 (21.1) 5/17 (29.4) R40 0/4 (0) 2/12 (16.7)
OR (95% CI) 1.20 (0.37–3.90) 0.38 (0.09–1.64) 0.64 (0.15–2.75) 0.00 (0.00–6.41)
Note: c2 or Fisher’s exact tests were used.
Note: c2 or Fisher’s exact tests were used.
Abu-Musa. 17a-hydroxyprogesterone caproate in IVF. Fertil Steril 2008.
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study showed that the administration of piroxicam, a nonsteroidal anti-inflammatory drug, just before ET increased the implantation and pregnancy rates after IVF-ET (9). Prostaglandin, which is synthesized from arachidonic acid by cyclooxygenase, stimulates UCs. Nonsteroidal anti-inflammatory drugs block the action of cyclooxygenase and inhibit the production of prostaglandin (16), which may cause a reduction in uterine contractility. Few studies have evaluated the role of vaginal P on UC and IVF outcome (17, 18). These researchers administered vaginal P starting on the day of oocyte retrieval to relax uterine contractility at the time of ET. As a result, they found a decrease in UC frequency on the day of ET, suggesting that a uterine-relaxing effect by P administration before ET is likely to improve IVF-ET outcome. When vaginal P was given on the evening of ET as opposed to on the day of oocyte pickup, uterine contractility remained exacerbated despite high mean serum P levels observed at the time of ET. This indicates that during the early luteal phase of COH, the uterinerelaxation process that is mediated by P may take longer than during the corresponding phase of the natural menstrual cycle (19–21). Hence, in COH, the achievement of rapid myometrial relaxation appears to require a more intense uterine exposure to P than is the case in the regular menstrual cycle. This suggests a degree of resistance to the uterine-relaxing action of endogenous P that may be attributable to the persisting uterine stimulation by the supraphysiological E2 levels of COH (17). In our study, the aim of giving a 17-HPC, which has a potent and prolonged progestational activity, was to achieve this intense uterine exposure to P and thus a rapid relaxation of the myometrium, which may prevent the premature expulsion of embryos from the uterine cavity at the time of ET and improve pregnancy rates. The role of 17HPC for luteal support in patients undergoing IVF-ET cycles has been evaluated in very few studies (22–25). Some of these studies have shown no beneficial effect of 17-HPC on the outcome of IVF-ET (22, 23). However, other investigators have reported a higher pregnancy rate when 17-HPC was used, compared with placebo or intravaginal P (24, 25). In our study, although there was no difference in pregnancy rate between cases and controls, there was a tendency for increase in the rate of twin gestation in patients who received 17-HPC; however, this did not reach statistical significance. The main limitation of this study is the small sample size. When pregnancy rate is taken as the primary outcome and assuming a pregnancy rate of 35% with IVF treatment, a 10% increase in pregnancy rate with 17-HPC treatment would require a sample size of 376 subjects in each arm, with an a of 0.05 and b of 80%. To detect a significant effect of 17-HPC on the implantation rate, assuming an implantation rate of 19% in untreated women and an effect associated with 17HPC of 10%, 286 women in each arm would be required to achieve a power of 80% and a level of significance of 95%. Another limitation of this study is that uterine contractility and 17-HPC levels (using high-pressure liquid chromatography or RIA) were not assessed on the day of ET. In one study, the mean serum level of 17-HPC after a single IM injection of Fertility and Sterility
1,000 mg reached its maximum (44–81 nmol/L) after 2–7 days (26). Costabile et al. (23) have demonstrated a slow increase in plasma level until the 5th day that was caused by a retarded absorption from the injection site. The timing of administration may need to be modified to R48 hours before ET, allowing for 17-HPC to reach significant blood levels. Alternatively, the dose of 17-HPC may need to be increased to achieve the required uterine relaxation. We did not use the same doses of 17-HPC that were used by other investigators (23–25), because in these studies, 17-HPC was used alone for the luteal support. In our study, 17-HPC was used as a bolus dose of P in addition to the standard vaginal P dose that we use for luteal support. It may be speculated that the benefit of 17-HPC may be reduced or masked by concurrent administration of vaginal P, which is given routinely to IVF patients in our center. In conclusion, preliminary evidence from this study shows that the addition of an IM progestational agent that has potent smooth-muscular relaxant properties to vaginal micronized P before ET does not appear to improve the pregnancy rate in IVF-ET cycles. REFERENCES 1. Tabibzadeh S, Babaknia A. The signals and molecular pathways involved in implantation, a symbiotic interaction between blastocyst and endometrium involving adhesion and tissue invasion. Hum Reprod 1995;10:1579–602. 2. Oike K, Obata S, Tagaki K, Matsuo K, Ishihara K, Kikuchi S. Observation of endometrial movement with transvaginal sonography. J Ultrasound Med 1988;7:S99. 3. Zaidi J, Pittrof R, Shaker A, Kyei-Mensah A, Campbell S, Tan SL. Assessment of uterine artery blood flow on the day of human chorionic gonadotropin administration by transvaginal color Doppler ultrasound in an in vitro fertilization program. Fertil Steril 1996;65:377–81. 4. IJland MM, Evers JLH, Dunselman GAJ, Van Katwijk C, Lo CR, Hoogland HJ. Endometrial wavelike movements during menstrual cycle. Fertil Steril 1996;65:746–9. 5. IJland MM, Evers JLH, Dunselman GAJ, Volovics L, Lo CR, Hoogland HJ. Relation between endometrial wavelike activity and fecundability in spontaneous cycles. Fertil Steril 1997;67:492–6. 6. IJland MM, Evers JLH, Dunselman GAJ, Lo CR, Hoogland HJ. Endometrial wavelike activity, endometrial thickness, and ultrasound texture in controlled ovarian hyperstimulation cycles. Fertil Steril 1998;70:279–83. 7. Bulletti C, Prefetto RA, Bazzocchi G, Romero R, Mimmi P, Polli V, et al. Electromechanical activities of human uteri during extra-corporeal perfusion with ovarian steroids. Hum Reprod 1993;8:1558–63. 8. Fanchin R, Righini C, Olivennes F, Taylor S, Ziegler D, Frydman R. Uterine contractions at the time of embryo transfer alter pregnancy rates after in-vitro fertilization. Hum Reprod 1998;13:1968–74. 9. Moon HS, Park SH, Lee JO, Kim KS, Joo BS. Treatment with piroxicam before embryo transfer increases the pregnancy rate after in vitro fertilization and embryo transfer. Fertil Steril 2004;82:816–20. 10. World Health Organization. WHO laboratory manual for the examination of human semen and sperm-cervical mucus interaction. 4th ed. New York: Cambridge University Press, 1999. 11. The Rotterdam ESHRE/ASRM-sponsored PCOS consensus workshop group. Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome (PCOS). Hum Reprod 2004;19:41–7. 12. Steer CV, Mills CL, Tan SL, Campbell S, Edwards RG. The cumulative embryo score: a predictive embryo scoring technique to select the optimal number of embryos to transfer in an in-vitro fertilization and embryo transfer programme. Hum Reprod 1992;7:117–9.
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