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day on ovarian stimulation with gonadotropins in women undergoing intrauterine insemination
OVULATION INDUCTION Effect of letrozole at 2.5 mg or 5.0 mg/day on ovarian stimulation with gonadotropins in women undergoing intrauterine insemination Luis Noriega-Portella, M.D.,a Luis Noriega-Hoces, M.D.,a Andrea Delgado, M.D.,a Julio Rubio, Lic.,b Cynthia Gonzales-Casta~ neda, Lic.,b and Gustavo F. Gonzales, M.D., D.Sc.b a
PRANOR, group of Assisted Reproduction, and b Laboratory of Investigation and Development, Faculty of Sciences and Philosophy ‘‘Alberto Cazorla Talleri,’’ Universidad Peruana Cayetano Heredia, Lima, Peru
Objective: To determine the effect of combined therapy of letrozole (2.5 mg or 5.0 mg) with recombinant folliclestimulating hormone (FSH) in comparison with the administration of recombinant FSH alone in an intrauterine insemination (IUI) program. Design: Retrospective study. Setting: Assisted fertilization program in a specialized infertility center. Patient(s): 110 women undergoing IUI and gonadotropin therapy. Intervention(s): Recombinant FSH alone administered from day 3 or combined with letrozole, 2.5 or 5.0 mg/day, on days 3 to 7, and gonadotropins starting on day 7 of the menstrual cycle. Transvaginal ultrasound examinations were done until the dominant follicle reached 18 mm in diameter. Ovulation was triggered with 10,000 IU of human chorionic gonadotropin (hCG), and IUI performed 30 to 40 hours later. Main Outcome Measure(s): Recombinant FSH dose required, number of follicles greater than 14 mm and 18 mm, endometrial thickness, pregnancy rates, miscarriages, and characteristics of newborns. Result(s): Women treated with FSH and 5.0 mg/day of letrozole required a lower dose of FSH than the group cotreated with 2.5 mg/day of letrozole or with FSH alone. Throughout most of the follicular phase, the endometrial thickness was statistically significantly less in both letrozole cotreatment groups compared with the FSH control group. By the day of hCG administration, the endometrial thickness was comparable among all the groups. The pregnancy rates were the same with recombinant FSH alone or combined with letrozole. Conclusion(s): In terms of cost-effectiveness, 5.0 mg/day of letrozole is more effective than the 2.5 mg/day in cotreatment with no adverse effect on pregnancy rate or outcome. (Fertil Steril 2008;90:1818–25. 2008 by American Society for Reproductive Medicine.) Key Words: Letrozole, aromatase inhibitors, recombinant FSH, intrauterine insemination
Aromatase is the key enzyme that converts testosterone to estradiol (1). Aromatase inhibitors (AIs) are a series of compounds that were developed for the treatment of breast cancer, where AI was demonstrated to be more effective than the classic treatment with tamoxifen (1). More recently, AIs also have been used for applications outside cancer treatment (2–5), such as inducing ovulation in women who have failed with clomiphene citrate treatment (6). In this use, AIs have been demonstrated to be even superior to clomiphene citrate in effectiveness (3, 7); in fact, unlike clomiphene citrate, AIs have no antiestrogenic effect over the endometrium, which be an advantage for keeping the endometrium in optimal condition to maintain a pregnancy (8). Therefore, AIs constitute an excellent alternative to clomiphene citrate treatment as an Received May 20, 2007; revised and accepted August 28, 2007. Reprint requests: Gustavo F. Gonzales, MD, D.Sc., Av. Honorio Delgado 430, Lima, Peru (FAX: 00511-4821195; E-mail: [email protected]).
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ovulation inductor for the treatment of infertility, particularly in assisted reproduction programs. When combined with recombinant follicle-stimulating hormone (FSH), letrozole, a third-generation aromatase inhibitor, modifies the follicular, hormonal, and endometrial dynamic in comparison with treatment with recombinant FSH alone. Recombinant FSH plus letrozole could have beneficial effects on the ovulatory and endometrial cycle (9, 10). The combination of 2.5 mg/day of letrozole with recombinant FSH also reduces the dose of recombinant FSH required for superovulation, thus lowering the cost of assisted fertility programs (11), particularly in women who have had a poor response to hormonal stimuli (8, 12). A recent study demonstrated that the dose of 5.0 mg of letrozole resulted in a greater number of follicles and an improved rate of pregnancies per cycle than 2.5 mg/day (13). However, it is not known whether this pattern also would
Fertility and Sterility Vol. 90, No. 5, November 2008 Copyright ª2008 American Society for Reproductive Medicine, Published by Elsevier Inc.
0015-0282/08/$34.00 doi:10.1016/j.fertnstert.2007.08.060
be observed when a different dose of letrozole is co-administered with recombinant FSH. Our study was designed to determine the differences in the effect of combined therapy of letrozole (2.5 mg or 5.0 mg) with recombinant FSH when compared with the administration of recombinant FSH alone in an intrauterine insemination (IUI) program. In a developing country, the use of 5.0 mg/day of letrozole could be an optimal dose for reducing the cost to patients with no adverse impact on endometrial thickness or pregnancy rate. MATERIALS AND METHODS Patients This retrospective study is based on secondary data analysis obtained from the clinical records of women who attended a specialized infertility center (PRANOR, Assisted Reproduction Group) in Lima, Peru, from December 2004 to January 2007. The study was approved by the institutional review board of the Universidad Peruana Cayetano Heredia (code number 52252). Data were collected from records of patients who had had primary infertility of diverse etiology (anovulation, endometriosis, male factor infertility, and unexplained infertility) and who underwent controlled ovarian stimulation with recombinant FSH alone or combined with letrozole (2.5 or 5.0 mg/ day) for IUI. Criteria of Inclusion The criteria of inclusion were patients with infertility of at least 1 year duration, aged <40 years, with patent tubes on hysterosalpingogram or laparoscopic assessment, and a normal uterine cavity, plus the presence of at least 10 million of motile sperms/mL in the male partner’s assessment. Groups of Study This study comprised 110 women who underwent 124 IUI cycles. The patients were divided in three different treatment groups: group 1 received a combination of 2.5 mg/day of letrozole for 5 days plus recombinant FSH; group 2 received a combination of 5.0 mg/day of letrozole for 5 days plus recombinant FSH; and group 3 received recombinant FSH alone (control group). Group 1 was 26 patients who had 30 stimulation cycles. Group 2 was 48 patients who had 55 stimulation cycles. Group 3 was 36 patients who had 39 stimulation cycles. Endometriosis was diagnosed by laparoscopy, male factor infertility was diagnosed according to the World Health Organization (14) criteria for normal semen, and the diagnosis of unexplained infertility was based on exclusion of male and female factors. Ovulation was documented by follicular monitoring with transvaginal ultrasonography. All patients underwent a maximum of two cycles of insemination protocols. The dosage of medication was based on the clinical profile of the patient, including age, weight, and duration of infertility. The initial dose of recombinant FSH Fertility and Sterility
was of 50 IU in four cases, 100 IU in 34 cases, and 150 IU in one case, and in both letrozole treatment groups the dose was 50 IU in 18 cases, 100 IU in 64 cases, 150 IU in two cases, and 200 IU in one case. The dose was adjusted according to the follicular monitoring. None of the women had received letrozole before. Treatment In group 1, letrozole (Femara; Novartis Pharmaceuticals AG, Basel, Switzerland) was given at a dosage of 2.5 mg daily from day 3 to 7 of the menstrual cycle, followed by the recombinant follicle-stimulating hormone (FSH) injection starting on day 7 until the day of the human chorionic gonadotropin (hCG) administration. Group 2 received a combination of 5.0 mg of letrozole daily from day 3 to 7 of the cycle plus recombinant FSH starting on day 7 until the day of hCG administration. In group 3, recombinant FSH (Puregon; Organon Oss, The Netherlands) injections started on day 3 of the menstrual cycle until the day of hCG administration. Human chorionic gonadotropin (Pregnyl; Organon Oss, The Netherlands; 10,000 IU IM) was given to trigger ovulation when the mean diameter of the dominant follicle reached R18 mm. Examinations Transvaginal ultrasound examinations were done before the beginning of the treatment on day 3 of the cycle as well as on days 7, 9, 10, and 11 of the cycle and more often if needed. During the ultrasound examination, the number of follicles >8 mm, >14 mm, and >18 mm, the diameter of each visible follicle, and the endometrial thickness (maximum distance between the echogenic interfaces of the myometrium) were measured. The endometrial thickness was measured in the plane through the central longitudinal axis of the uterus. The ultrasound examination was done by the same ultrasonographer, using the same ultrasound equipment (General Electric; Voluson 730 Expert; General Electric Healthcare, Waukesha, WI). The values of serum FSH and estradiol were measured on day 3 of the cycle for all of the women. Information about sperm concentration, sperm motility, and sperm morphologic features of the partners also were collected. For this study, a concentration R20 106/mL, motility grade a þ b R50%, and morphology R30% normal forms were considered normal values (14). Intrauterine Insemination and Outcomes Intrauterine insemination was performed 30 to 40 hours after hCG administration. Clinical pregnancy was confirmed by fetal cardiac pulsation 5 weeks after a positive pregnancy test. The study concluded with the confirmation of the delivery of a newborn. The main outcome measures included the number of follicles >14 mm and >18 mm, the total dose of recombinant FSH used in all the groups, and the endometrial thickness 1819
at different stages of the follicular phase. Pregnancy rate, number of multiple gestations, and miscarriages were also investigated. The cost of the gonadotropins with or without letrozole also was calculated. In the recombinant FSH only group, three women had two insemination cycles; in the group who received recombinant FSH þ 2.5 mg of letrozole, four women had two insemination cycles; and in the group who received recombinant FSH þ 5.0 mg of letrozole, six women had two insemination cycles. The number of insemination cycles did not affect the results of the study. Newborn Analysis Data from newborns such as birthweight, Apgar score at the first and fifth minutes of life, and gestational age were collected. Statistical Analysis The results are presented as mean standard deviation. The Bartlett test was used to obtain homogeneity of variances. If the data were not normally distributed, they were transformed or a nonparametric analysis was performed. If variances were homogeneous, an analysis of variance (ANOVA) test was used; if differences were observed between groups, the Scheffe test was used to determine differences between a pair of means. The pregnancy rate was compared among the groups using the chi-square test (Fisher’s exact test). Data from the newborns also were compared among the treatment groups (birthweight, gestational age, and Apgar score). A logistic regression analysis was performed to assesses the probability of pregnancy in women undergoing IUI with recombinant FSH, recombinant FSH þ 2.5 mg of letrozole, or recombinant FSH þ 5.0 mg of letrozole. P<.05 was considered statistically significant. RESULTS Characteristics of the Study Participants A total of 110 patients underwent 124 IUI cycles. To determine the homogeneity of the treatment groups, we evaluated the baseline characteristics of the patients, which showed that there were no statistically significant differences in chrono-
logic age, infertility time, or body mass index. Similarly, the baseline serum hormone levels were not statistically significantly different among the study groups (Table 1). According to the type of infertility of the couple, we determined that there are no statistically significant differences for the infertility factors among the study groups. However, male factor infertility predominated over the other factors: 58.33% in the recombinant FSH only group, 57.69% in the recombinant FSH þ 2.5 of letrozole group, and 48.97% in the recombinant FSH þ 5.0 mg of letrozole group (P>.05). In the same groups, female factor infertility was found in 16.67%, 19.23%, and 14.28% (P>.05), respectively, and unexplained infertility in 25.0%, 23.07%, and 34.69% (P>.05), respectively. One couple in the recombinant FSH þ 5.0 mg letrozole group had both male and female factor infertility. The female factors included 10 cases of polycystic ovary syndrome; of these, one was in the group treated with recombinant FSH only, three were in the group treated with recombinant FSH þ 2.5 mg of letrozole, and six were in the group treated with recombinant FSH þ 5.0 mg of letrozole. Six cases of endometriosis were found in the recombinant FSH only group, four in the group receiving FSH þ 2.5 mg of letrozole, and four in the group receiving recombinant FSH þ 5.0 mg of letrozole. When evaluating the sperm parameters (sperm concentration, sperm motility grade 3, and sperm normal morphology) of the partners, differences were observed among the groups. Sperm concentration in the recombinant FSH þ 5.0 mg of letrozole group was statistically significantly (P<.05) lower compared with the recombinant FSH only group, but was similar to the recombinant FSH þ 2.5 mg letrozole group (P>.05). Sperm motility in the recombinant FSH þ 5.0 mg letrozole group was statistically significantly (P<.05) lower compared with the recombinant FSH only group. No differences were observed in sperm motility between the group receiving recombinant FSH only and the group receiving recombinant FSH þ 2.5 mg of letrozole (P>.05). Sperm morphology was similar among all of the groups (P>.05) (Table 2). Effect of Letrozole on Recombinant FSH Dose The total recombinant FSH dose needed during the treatment was statistically significantly (P<.05) lower in both of the
TABLE 1 Baseline characteristics of the patients according to treatment group. Treatment group (no. of cycles)
Age (years)
Infertility time (months)
Body mass index (kg/m2)
FSH levels (mIU/mL)
Estradiol levels (pg/mL)
rFSH (39) rFSH þ L 2.5 (30) rFSH þ L 5.0 (55)
34.05 3.62 33.13 4.22 33.76 3.02
45.49 26.29 37.20 21.91 47.02 34.62
24.52 2.56 24.51 2.81 24.86 3.04
6.58 1.74 7.55 1.90 7.86 2.82
46.49 20.61 38.99 10.75 42.05 27.96
Note: Data are mean standard deviation. P>.05 between groups. L 2.5 ¼ 2.5 mg/day of letrozole for 5 days; L 5.0 ¼ 5.0 mg/day of letrozole for 5 days; rFSH, recombinant follicle-stimulating hormone. Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
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TABLE 2 Sperm parameters according to the treatment groups. Treatment groups rFSH rFSH þ L 2.5 rFSH þ L 5.0
Sperm concentration (3 106/mL)
Sperm motility grade a D b (%)
Sperm normal morphology (%)
74.72 45.37 68.83 53.45 55.16 42.65a
54.89 12.70 50.30 12.24 48.94 11.47b
33.44 14.50 30.81 12.18 30.02 9.09
Note: Data are mean standard deviations. L 2.5 ¼ 2.5 mg/day of letrozole for 5 days; L 5.0 ¼ 5.0 mg/day of letrozole for 5 days; rFSH, recombinant follicle-stimulating hormone. Motility a: rapid and linear progressive sperm motility. Motility b: slow or sluggish linear or nonlinear sperm movement. rFSH, recombinant follicle-stimulating hormone. a P< .05 versus rFSH (Mann-Whitney test). b P< .05 with respect to rFSH (analysis of variance). Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
letrozole cotreatment groups. When comparing both letrozole dosage groups, we found that the group receiving recombinant FSH þ 5.0 mg of letrozole required a statistically significant (P<.05) lower dose of recombinant FSH during the overall treatment compared with the group receiving letrozole at a dosage of 2.5 mg/day (Table 3). The days of stimulation with recombinant FSH were also reduced in the groups cotreated with letrozole (2.5 mg or 5.0 mg). The lowest value was observed with letrozole at 5.0 mg/day.
mm were observed when comparing the recombinant FSH þ 2.5 mg letrozole group with the recombinant FSH þ 5.0 mg letrozole group (P>.05). The endometrial thickness measured on the day of the hCG administration showed no differences among the three treatment groups (P>.05). In average, the cost of treatment per cycle with recombinant FSH alone was US$ 619.68; with FSH þ 2.5 mg/day of letrozole was US$ 499.69; and with FSH þ 5.0 mg of letrozole was US$ 259.77 (Table 3).
The number of follicles with diameter >8 mm was similar in the groups of study (P>.05). The number of ovarian follicles with diameter >14 mm was statistically significantly (P<.05) higher in the recombinant FSH only group compared with the recombinant FSH þ 5.0 mg letrozole group. There was no statistically significant difference in the number of follicles with diameter >14 mm between the letrozole cotreatment groups. The number of ovarian follicles with diameter >18 mm was statistically significantly higher (P<.05) in the recombinant FSH only group than in the groups combined with letrozole. No differences in the number of follicles >18
Dynamic of the Endometrial Thickness The analysis of endometrial thickness before hCG administration showed that it was comparable among the three groups at the beginning of the treatment (recombinant FSH only: 3.22 0.75 cm; recombinant FSH þ 2.5 mg letrozole; 3.25 0.75 cm; recombinant FSH þ 5.0 mg letrozole: 3.18 0.67 cm; P>.05). However, throughout most of the follicular phase, the endometrial thickness was statistically significantly (P<.05) reduced in both letrozole cotreatment groups compared with the recombinant FSH control group.
TABLE 3 Total dose of recombinant follicle-stimulating hormone (rFSH), days of stimulation with rFSH, sonographic characteristics, and rFSH cost. Treatment groups Total rFSH dose (IU) Days of stimulation with rFSH Follicles >8 mm Follicles >14 mm Follicles >18 mm Endometrial thickness (mm) rFSH cost (US$)
rFSH (no. of cycles [ 39)
rFSH D L 2.5 (no. of cycles [ 30)
rFSH D L 5.0 (no. of cycles [ 55)a,b
723.08 196.30 7.85 1.76 4.26 1.97 3.39 1.66 2.08 0.98 8.96 0.87 619.68 168.22
558.33 294.51a 5.63 2.16a 4.77 2.10 2.77 1.63 1.37 0.72a 8.63 0.94 499.69 252.40a
253.64 145.57a,b 3.13 1.45a,b 4.22 1.86 2.56 1.44a 1.47 0.61b 8.51 1.42 259.77 124.75a,b
Note: Data on total rFSH dose are mean standard deviation. a P< .05 versus FSH. b P< .05 versus FSH þ 2.5 mg of letrozole. Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
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This difference began to decrease 3 days before hCG administration but was still statistically significant (P<.05). By the day of the hCG administration, endometrial thickness was comparable among all the groups (recombinant FSH only: 8.96 0.87 cm; recombinant FSH þ 2.5 mg letrozole; 8.63 0.94 cm; recombinant FSH þ 5.0 mg letrozole; 8.51 1.42 cm) (Fig. 1). Pregnancies, Multiple Gestations, Miscarriages, and Births Pregnancy rates were similar when comparing the group treated with recombinant FSH alone and both letrozole groups (P>.05). In the recombinant FSH only group, one multiple gestation (one triplet) and two miscarriages occurred, and one of the triplets was a stillbirth. No miscarriages occurred in either letrozole group (Table 4). No multiple gestations occurred in the 2.5 mg/day letrozole group. One multiple gestation (twin) and one ectopic pregnancy occurred in the 5.0 mg/day letrozole group. In the FSH-only group, one preterm delivery occurred; in the letrozole groups, one preterm delivery occurred in the 2.5 mg/day group and one in 5.0 mg/day group. The birthweight and Apgar score values, measured on the newborn’s first and fifth minutes, were similar (P>.05) among all treatment groups (Table 4). One preterm delivery occurred in the FSH-only group, the 2.5 mg/day letrozole group, and the 5.0 mg/day letrozole group; however, the gestational age was comparable (P>.05) among all groups. One woman from the FSH-only group had preeclampsia. In the 5.0 mg/day letrozole group, one case with gestational diabetes occurred as well as one case where the umbilical cord was encircled around portions of the fetus.
FIGURE 1
Endometrial Thickness (mm)
Endometrial thickness measured in the follicular phase of cycles stimulated with recombinant folliclestimulating hormone (FSH) alone, 2.5 mg/day of letrozole plus FSH, and 5.0 mg/day of letrozole plus FSH. Day 0 is the day of human chorionic gonadotropin administration. 8.5
6.5 FSH
DISCUSSION Follicle-stimulating hormone is widely used in ovarian stimulation during assisted reproduction treatment. Pregnancy rates for women who have received FSH are statistically significantly higher compared with women who did not undergo controlled ovarian stimulation before an intrauterine insemination (IUI) (15). However, the use of FSH has raised the cost of these therapies considerably, and it is not free of side effects such as ovarian hyperstimulation syndrome or multiple pregnancies (16). Over the past few years, several studies have proved the effectiveness of the aromatase inhibitors (AIs) in ovulation induction because they reduce the levels of estradiol. Compared with clomiphene citrate, AIs have a short half-life (45 hours); they can be eliminated rapidly from the body (17) without causing the side effects that have been observed in cycles with clomiphene citrate (9). Cotreatment of recombinant FSH plus 2.5 mg/day of the AI letrozole for 5 days significantly reduces the FSH dose required (11). In groups of women with comparable age, duration of infertility, body mass index, and basal FSH and estradiol levels, our study showed that both doses of letrozole (2.5 or 5.0 mg/day) reduced the dose of recombinant FSH required for a stimulation cycle in a program of IUI. Overall, the larger letrozole dose (5.0 mg/day) required less FSH and reduced the number of days of stimulation when compared with the 2.5 mg/day dose. A review of different studies showed that letrozole reduced the FSH dose required from 45% to 55% (18). In our study, 2.5 mg/day of letrozole reduced the FSH dose required 77.17% and 5.0 mg/day of letrozole reduced it 35.07%. This reduction in the required dose of FSH was also associated with a lower number of ovarian follicles >14 mm than is obtained with recombinant FSH alone.
FSH + L 2.5
4.5
FSH + L 5.0
2.5
0.5 -9
-8
-7
-6
-5
-4
-3
-2
-1
0
Days from hCG Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
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After controlling for age; time of infertility; presence of male factor infertility, endometriosis, polycystic ovarian syndrome, or unexplained infertility; body mass index; total dosage of recombinant FSH; and days of recombinant FSH stimulation, the logistic regression analysis showed that the probability of pregnancy was similar for treatments using recombinant FSH along, recombinant FSH þ 2.5 mg/day of letrozole, or recombinant FSH þ 5.0 mg/day of letrozole (P>.05) (Table 5). After controlling for sperm parameters in the multivariable analysis, the probability of pregnancy rate was similar between women treated with recombinant FSH only and those treated with recombinant FSH and letrozole (data not shown).
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This finding is very important because with the use of letrozole at a dose of 5.0 mg/day the cost of the treatment is significantly reduced. The cost of FSH plus 2.5 mg/day of letrozole has been suggested to be more than half the value of FSH alone (12). Our study demonstrated that the reduction in cost of using FSH with 2.5 mg/day of letrozole was 80.63% less than with using FSH alone; using 5.0 mg/day of letrozole reduces the cost 41.92%. In consequence, the cost of each cycle of IUI is lower. This is important for developing countries
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TABLE 4 Pregnancy, multiple gestation, and miscarriage rates and newborn characteristics according to treatment group. Treatment groups
rFSH (no. of cycles [ 39)
rFSH D L 2.5 (no. of cycles [ 30)
rFSH D L 5.0 (no. of cycles [ 55)
Pregnancies/cycle completeda,b,c Births/cycle completedb,d Miscarriages/cycle completedb Gestational age (weeks)e Birthweight (g)e Apgar score (min 1)e Apgar score (min 5)e
7/39 17.9% (6–30) 4/39 10.3% (1–19) 2/39 5.1% (–2–12) 37.20 2.77 (4) 2938.0 885.2 (4) 8.40 1.3 (4) 9.40 0.6 (4)
2/30 6.7% (–2–20) 2/30 6.7% (–2–20) 0/30 0 (0–0) 35.10 6.93 (2) 2150.0 1343.5 (2) 9.00 0.0 (2) 9.50 0.7 (2)
13/55 23.6% (13–35) 9/55 16.4% (6–26) 0/55 0 (0–0) 38.39 0.99 (9) 3094.5 542.5 (9) 9.00 0.0 (9) 9.67 0.5 (9)
Note: L 2.5 ¼ 2.5 mg/day of letrozole for 5 days; L 5.0 ¼ 5.0 mg/day of letrozole for 5 days; rFSH, recombinant folliclestimulating hormone. a Pregnancy rates are expressed as percentages. b Between parentheses is the 95% confidence interval. P>.05 for pregnancy, birth, and miscarriage rates. c One ectopic pregnancy occurred in the 5.0 mg/day letrozole group. d Three preterm deliveries occurred, one in each of the three groups. e Gestational age, birthweight and Apgar scores are expressed as mean standard deviation; P>.05 between groups. One pregnancy in the rFSH-only group and three from the rFSH þ L 5.0 group are still in progress. Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
because the cost of assisted reproduction programs precludes access for a large segment of the poor population. The use of FSH alone is clearly a risk for ovarian hyperstimulation syndrome (16); however, our FSH-only group had no cases of ovarian hyperstimulation syndrome because we used a lower dose of recombinant FSH than has been used in other studies (20). In fact, our average dose of 723 IU
of recombinant FSH has resulted in a pregnancy rate of 17.9% per completed cycle; to obtain a similar pregnancy rate, another recent study required 1352 IU of recombinant FSH (19). The low number of ovarian follicles recruited with letrozole and the low dose of FSH required, both did not affect negatively the endometrial thickness at the time of hCG
TABLE 5 Logistic regression analysis to assess the probability of pregnancy in women treated with recombinant follicle-stimulating hormone (rFSH), rFSH D 2.5 mg/day of letrozole, or rFSH D 5.0 mg/day of letrozole. Variables Treatment rFSH rFSH þ L 2.5 mg/day rFSH þ 5.0 mg/day Age (years) Time of infertility (months) Infertility Male factor Endometriosis Polycystic ovary Unexplained infertility Body mass index (kg/m2) Total rFSH dose (IU) Days of rFSH stimulation
Unadjusted ORa
Adjusted ORa
Confidence interval at 95%
1.00 0.326 1.416 0.938 1.009
1.00 0.130 0.948 0.919 1.005
0.012–1.382 0.137–6.837 0.788–1.072 0.989–1.021
0.779 0.496 2.456 1.00 1.115 0.998 0.926
0.821 0.271 4.171 1.00 1.154 0.999 1.062
0.2731–2.473 0.0227–3.253 0.6605–26.34 1.00 0.9495–1.403 0.9940–1.004 0.6198–1.82
Note: Data are presented as unadjusted and adjusted odds ratio (OR). a P>.05 for each variable in the unadjusted or in the adjusted model. Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
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administration. This is very important because a reduction of endometrial thickness is observed at the time of hCG administration after combined treatment with FSH and clomiphene citrate (9, 20). Similar (9, 21) or lower (20) pregnancy rates have been observed with clomiphene compared with AIs. Our study obtained similar pregnancy rates for the group receiving FSH alone compared with the groups receiving FSH plus 2.5 or 5.0 mg/day of letrozole. In fact, use of FSH with letrozole at 5.0 mg/day for 5 days resulted in a lower dose of FSH required, fewer ovarian follicles recruited, a similar endometrial thickness at the time of hCG administration, and similar pregnancy rate compared with FSH alone. Other investigators have obtained similar pregnancy rates when comparing groups treated with recombinant FSH alone and recombinant FSH þ 2.5 mg/day of letrozole in IUI programs (11, 19). Some investigators have found that the treatment with FSH plus clomiphene citrate results in similar pregnancy rates compared with FSH plus an AI such as letrozole (9, 21). However, it is also true that treatment with clomiphene citrate resulted in a higher number of miscarriages, ectopic gestations, and stillbirths (22) than treatment with letrozole. This is probably because clomiphene citrate has a long clearance half-life (of about 5 days to 3 weeks) and it may be stored in body fat (18), which means its antiestrogenic effect on the endometrium may be prolonged (23). In another study that used FSH with 5.0 mg/day of letrozole, FSH was administered from day 5 of the menstrual cycle, overlapping the letrozole treatment. Although the resulting pregnancy rates were not different among treatment groups, the addition of letrozole to gonadotropins was found to decrease the endometrial thickness (24), likely because the overlapping regimen advanced follicle growth and ovulation but did not allow time for clearance of the larger dose of letrozole by the time of hCG administration (18). In our protocol, in which FSH was administered from day 7 until the day of hCG administration, endometrial thickness was not affected by letrozole at the dose of 5.0 mg/day at the time of hCG administration. Endometrial thickness depends on estrogen, and AIs reduce the production of estradiol. Therefore, if letrozole inhibits the conversion of androstenedione to estradiol when administered and before clearance occurs, it may affect endometrial thickness. In fact, in our study, endometrial development was delayed in the first half of the follicular phase in both letrozole cotreated groups, as determined by the lower endometrial thickness found in these groups when compared with the results obtained in the group treated with FSH alone. When both letrozole groups are compared, the 5.0 mg/day dose delayed endometrial development even more than the 2.5 mg/day dose, and the endometrium required more days to recover from the estrogen reduction. However, the endometrium had completely recovered by the day of the hCG administration, indicating that the antiestrogen effect of letrozole had ended. This indicates that normal endometrial 1824
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development proceeds despite the lower estrogen levels, as confirmed by the comparable pregnancy rates observed among the study groups. Moreover, the fact that the pregnancies ended with live births suggests that the reduction in endometrial thickness in the first half of the cycle had no negative effect on pregnancy outcome. Although letrozole reduces estradiol levels during the follicular phase by inhibition of the aromatase enzyme, the pregnancy rate is not affected. Experimental studies have shown that adequate folliculogenesis is independent of estrogen but rather is conditioned on gonadotropin stimulation. Moreover, depletion of estradiol does not impair the developmental potential of the oocytes, including their fertilization and development into morula, blastocysts, and hatching blastocysts (25). Multiple gestations, a serious complication in assisted reproduction, derive from the use of gonadotropins. This has been confirmed in IUI programs in which FSH alone is used (19). Several studies have confirmed that the use of letrozole diminishes the incidence of multiple gestations (4). In our study, the use of FSH alone resulted in one multiple gestation of triplets, of which two were preterms and the other a stillbirth. One twin pregnancy occurred in the FSH þ letrozole groups, which completed at term without complications. Cotreatment of FSH with letrozole had no deleterious effect on the fetal development. The Apgar score is used to describe the condition of infants immediately after birth, and a 5-minute Apgar score of 7 to 10 is considered normal (26). In our study, all of the newborns had normal Apgar values, which were comparable among all the study groups. Our study was limited in that it was nonrandomized. However, all demographic variables were similar in our three groups (FSH alone, FSH þ 2.5 mg/day of letrozole, and FSH þ 5.0 mg/day of letrozole). Moreover, classification of the groups according to male factor, female factor, or unexplained infertility confirmed that groups were comparable and homogeneous. The only difference was that the sperm count and sperm motility of the male partners of the FSH þ letrozole 5.0 mg/day group were lower than in the other two groups; however, this was not a difficulty because the pregnancy rates did not differ. A number of adverse events are related to pregnancy, such as nutrition variations, preeclampsia, gestational diabetes, and labor complications. Our study had only one case of preeclampsia (in the FSH-only group) and one case of gestational diabetes (in the FSH þ 5.0 mg letrozole group). The 5.0 mg letrozole group also had a case of the umbilical cord encircling portions of the fetus. All three cases were treated by cesarean delivery. Irrespective of these factors, the rate of births per cycle completed was similar among the three groups. Women treated with recombinant FSH þ 5.0 mg/day of letrozole recruited fewer follicles and required a statistically significantly lower dosage of FSH, with no effect on the endometrial thickness at the time of hCG administration or the pregnancy rate. The treatment protocol of FSH þ letrozole
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at 5.0 mg/day should be considered a valid alternative for developing countries because the FSH required is reduced more than 50% over the 2.5 mg/day regimen, thus reducing the overall cost of the treatment with no impact on effectiveness. REFERENCES 1. Hong Y, Chen S. Aromatase inhibitors: structural features and biochemical characterization. Ann NY Acad Sci 2006;1089:237–51. 2. Dunkel L. Use of aromatase inhibitors to increase final height. Mol Cell Endocrinol 2006;254–5:207–16. 3. Holzer H, Casper R, Tulandi T. A new era in ovulation induction. Fertil Steril 2006;85:277–84. 4. Mitwally MF, Biljan MM, Casper RF. Pregnancy outcome after the use of an aromatase inhibitor for ovarian stimulation. Am J Obstet Gynecol 2005;192:381–6. 5. Bayar U, Basaran M, Kiran S, Coskun A, Gezer S. Use of an aromatase inhibitor in patients with polycystic ovary syndrome: a prospective randomized trial. Fertil Steril 2006;86:1447–51. 6. Begun MR, Quadir E, Begum A, Begum RA, Begum M. Role of aromatase inhibitor in ovulation induction in patients with poor response to clomiphene citrate. J Obstet Gynaecol Res 2006;32:502–6. 7. Jee BC, Ku SY, Suh CS, Kim KC, Lee WD, Kim KC, et al. Use of letrozole versus clomiphene citrate combined with gonadotropins in intrauterine insemination cycles: a pilot study. Fertil Steril 2006;85:1774–7. 8. Verpoest WM, Kolibianakis E, Papanikolaou E, Smitz J, Van Steirteghem A, Devroev P. Aromatase inhibitors in ovarian stimulation for IVF/ICSI: a pilot study. Reprod Biomed Online 2006;13:166–72. 9. Barroso G, Menocal G, Felix H, Rojas-Ruiz JC, Arsian M, Oehninger S. Comparison of the efficacy of the aromatase inhibitor letrozole and clomiphene citrate as adjuvants to recombinant follicle-stimulating hormone in controlled ovarian hyperstimulation: a prospective, randomized, blinded clinical trial. Fertil Steril 2006;86:1428–31. 10. Goswami SK, Das T, Chattopadhyay R, Sawhney V, Kumar J, Chaudhury K, et al. A randomized single-blind controlled trial of letrozole as a low-cost IVF protocol in women with poor ovarian response: a preliminary report. Hum Reprod 2004;19:2031–5. 11. Mitwally MF, Casper RF. Aromatase inhibition reduces gonadotrophin dose required for controlled ovarian stimulation in women with unexplained infertility. Hum Reprod 2003;18:1588–97. 12. Bedaiwy MA, Forman R, Mousa NA, Al Inany HG, Casper RF. Costeffectiveness of aromatase inhibitor cotreatment for controlled ovarian stimulation. Hum Reprod 2006;21:2838–44.
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13. Al-Fadhli R, Silvestre C, Buckett W, Tan SL, Tulandi T. A randomized trial of superovulation with two different doses of letrozole. Fertil Steril 2006;85:161–4. 14. World Health Organization. Laboratory manual for the examination of human semen and sperm-cervical mucus interaction. 4th ed. New York: Cambridge University Press, 1998. 15. Hughes EG. The effectiveness of ovulation induction and intrauterine insemination in the treatment of persistent infertility: a meta-analysis. Hum Reprod 1997;12:1865–72. 16. Vegetti W, Alagna F. FSH and folliculogenesis: from physiology to ovarian stimulation. Reprod Biomed Online 2006;12:684–94. 17. Sioufi A, Sandrenan N, Godbillon J, Trunet P, Czendlik C, Howald H, et al. Comparative bioavailability of letrozole under fed and fasting conditions in 12 healthy subjects after a 2.5 mg single oral administration. Biopharm Drug Dispos 1997;18:489–97. 18. Casper RF, Mitwally MF. Review: aromatase inhibitors for ovulation induction. J Clin Endocrinol Metab 2006;91:760–71. 19. Bedaiwy MA, Mousa NA, Esfandiari N, Forman R, Casper RF. Follicular phase dynamics with combined aromatase inhibitor and follicle stimulating hormone treatment. J Clin Endocrinol Metab 2007;92:825–33. 20. Wu HH, Wang NM, Cheng ML, Hsieh JN. Randomized comparison of ovulation induction and hormone profile between the aromatase inhibitor anastrozole and clomiphene citrate in women with infertility. Gynecol Endocrinol 2007;23:76–81. 21. Sipe CS, Davis WA, Maifeld M, Van Voorhis BJ. A prospective randomized trial comparing anastrozole and clomiphene citrate in an ovulation induction protocol using gonadotropins. Fertil Steril 2006;86: 1676–81. 22. Jain JK, Kuo J. Pregnancy outcomes with increased clomiphene citrate dose. Gynecol Endocrinol 2004;19:141–5. 23. Unfer V, Casini ML, Costabile L, Mignosa M, Gerli S, Di Renzo GC. High dose of phytoestrogens can reverse the antiestrogenic effects of clomiphene citrate on the endometrium in patients undergoing intrauterine insemination: a randomized trial. J Soc Gynecol Investig 2004;11: 323–8. 24. Healey S, Tan SL, Tulandi T, Biljan MM. Effects of letrozole on superovulation with gonadotropins in women undergoing intrauterine insemination. Fertil Steril 2003;80:1325–9. 25. Fatum M, Gyo Y, Diana P, Laufer N, Simon A. Is estradiol mandatory for an adequate follicular and embryo development? A mouse model using aromatase inhibitor (anastrozole). J Assist Reprod Genet 2006;23: 407–12. 26. Papile LA. The Apgar score in the 21st century. N Engl J Med 2001;344: 519–20.
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PRANOR, group of Assisted Reproduction, and b Laboratory of Investigation and Development, Faculty of Sciences and Philosophy ‘‘Alberto Cazorla Talleri,’’ Universidad Peruana Cayetano Heredia, Lima, Peru
Objective: To determine the effect of combined therapy of letrozole (2.5 mg or 5.0 mg) with recombinant folliclestimulating hormone (FSH) in comparison with the administration of recombinant FSH alone in an intrauterine insemination (IUI) program. Design: Retrospective study. Setting: Assisted fertilization program in a specialized infertility center. Patient(s): 110 women undergoing IUI and gonadotropin therapy. Intervention(s): Recombinant FSH alone administered from day 3 or combined with letrozole, 2.5 or 5.0 mg/day, on days 3 to 7, and gonadotropins starting on day 7 of the menstrual cycle. Transvaginal ultrasound examinations were done until the dominant follicle reached 18 mm in diameter. Ovulation was triggered with 10,000 IU of human chorionic gonadotropin (hCG), and IUI performed 30 to 40 hours later. Main Outcome Measure(s): Recombinant FSH dose required, number of follicles greater than 14 mm and 18 mm, endometrial thickness, pregnancy rates, miscarriages, and characteristics of newborns. Result(s): Women treated with FSH and 5.0 mg/day of letrozole required a lower dose of FSH than the group cotreated with 2.5 mg/day of letrozole or with FSH alone. Throughout most of the follicular phase, the endometrial thickness was statistically significantly less in both letrozole cotreatment groups compared with the FSH control group. By the day of hCG administration, the endometrial thickness was comparable among all the groups. The pregnancy rates were the same with recombinant FSH alone or combined with letrozole. Conclusion(s): In terms of cost-effectiveness, 5.0 mg/day of letrozole is more effective than the 2.5 mg/day in cotreatment with no adverse effect on pregnancy rate or outcome. (Fertil Steril 2008;90:1818–25. 2008 by American Society for Reproductive Medicine.) Key Words: Letrozole, aromatase inhibitors, recombinant FSH, intrauterine insemination
Aromatase is the key enzyme that converts testosterone to estradiol (1). Aromatase inhibitors (AIs) are a series of compounds that were developed for the treatment of breast cancer, where AI was demonstrated to be more effective than the classic treatment with tamoxifen (1). More recently, AIs also have been used for applications outside cancer treatment (2–5), such as inducing ovulation in women who have failed with clomiphene citrate treatment (6). In this use, AIs have been demonstrated to be even superior to clomiphene citrate in effectiveness (3, 7); in fact, unlike clomiphene citrate, AIs have no antiestrogenic effect over the endometrium, which be an advantage for keeping the endometrium in optimal condition to maintain a pregnancy (8). Therefore, AIs constitute an excellent alternative to clomiphene citrate treatment as an Received May 20, 2007; revised and accepted August 28, 2007. Reprint requests: Gustavo F. Gonzales, MD, D.Sc., Av. Honorio Delgado 430, Lima, Peru (FAX: 00511-4821195; E-mail: [email protected]).
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ovulation inductor for the treatment of infertility, particularly in assisted reproduction programs. When combined with recombinant follicle-stimulating hormone (FSH), letrozole, a third-generation aromatase inhibitor, modifies the follicular, hormonal, and endometrial dynamic in comparison with treatment with recombinant FSH alone. Recombinant FSH plus letrozole could have beneficial effects on the ovulatory and endometrial cycle (9, 10). The combination of 2.5 mg/day of letrozole with recombinant FSH also reduces the dose of recombinant FSH required for superovulation, thus lowering the cost of assisted fertility programs (11), particularly in women who have had a poor response to hormonal stimuli (8, 12). A recent study demonstrated that the dose of 5.0 mg of letrozole resulted in a greater number of follicles and an improved rate of pregnancies per cycle than 2.5 mg/day (13). However, it is not known whether this pattern also would
Fertility and Sterility Vol. 90, No. 5, November 2008 Copyright ª2008 American Society for Reproductive Medicine, Published by Elsevier Inc.
0015-0282/08/$34.00 doi:10.1016/j.fertnstert.2007.08.060
be observed when a different dose of letrozole is co-administered with recombinant FSH. Our study was designed to determine the differences in the effect of combined therapy of letrozole (2.5 mg or 5.0 mg) with recombinant FSH when compared with the administration of recombinant FSH alone in an intrauterine insemination (IUI) program. In a developing country, the use of 5.0 mg/day of letrozole could be an optimal dose for reducing the cost to patients with no adverse impact on endometrial thickness or pregnancy rate. MATERIALS AND METHODS Patients This retrospective study is based on secondary data analysis obtained from the clinical records of women who attended a specialized infertility center (PRANOR, Assisted Reproduction Group) in Lima, Peru, from December 2004 to January 2007. The study was approved by the institutional review board of the Universidad Peruana Cayetano Heredia (code number 52252). Data were collected from records of patients who had had primary infertility of diverse etiology (anovulation, endometriosis, male factor infertility, and unexplained infertility) and who underwent controlled ovarian stimulation with recombinant FSH alone or combined with letrozole (2.5 or 5.0 mg/ day) for IUI. Criteria of Inclusion The criteria of inclusion were patients with infertility of at least 1 year duration, aged <40 years, with patent tubes on hysterosalpingogram or laparoscopic assessment, and a normal uterine cavity, plus the presence of at least 10 million of motile sperms/mL in the male partner’s assessment. Groups of Study This study comprised 110 women who underwent 124 IUI cycles. The patients were divided in three different treatment groups: group 1 received a combination of 2.5 mg/day of letrozole for 5 days plus recombinant FSH; group 2 received a combination of 5.0 mg/day of letrozole for 5 days plus recombinant FSH; and group 3 received recombinant FSH alone (control group). Group 1 was 26 patients who had 30 stimulation cycles. Group 2 was 48 patients who had 55 stimulation cycles. Group 3 was 36 patients who had 39 stimulation cycles. Endometriosis was diagnosed by laparoscopy, male factor infertility was diagnosed according to the World Health Organization (14) criteria for normal semen, and the diagnosis of unexplained infertility was based on exclusion of male and female factors. Ovulation was documented by follicular monitoring with transvaginal ultrasonography. All patients underwent a maximum of two cycles of insemination protocols. The dosage of medication was based on the clinical profile of the patient, including age, weight, and duration of infertility. The initial dose of recombinant FSH Fertility and Sterility
was of 50 IU in four cases, 100 IU in 34 cases, and 150 IU in one case, and in both letrozole treatment groups the dose was 50 IU in 18 cases, 100 IU in 64 cases, 150 IU in two cases, and 200 IU in one case. The dose was adjusted according to the follicular monitoring. None of the women had received letrozole before. Treatment In group 1, letrozole (Femara; Novartis Pharmaceuticals AG, Basel, Switzerland) was given at a dosage of 2.5 mg daily from day 3 to 7 of the menstrual cycle, followed by the recombinant follicle-stimulating hormone (FSH) injection starting on day 7 until the day of the human chorionic gonadotropin (hCG) administration. Group 2 received a combination of 5.0 mg of letrozole daily from day 3 to 7 of the cycle plus recombinant FSH starting on day 7 until the day of hCG administration. In group 3, recombinant FSH (Puregon; Organon Oss, The Netherlands) injections started on day 3 of the menstrual cycle until the day of hCG administration. Human chorionic gonadotropin (Pregnyl; Organon Oss, The Netherlands; 10,000 IU IM) was given to trigger ovulation when the mean diameter of the dominant follicle reached R18 mm. Examinations Transvaginal ultrasound examinations were done before the beginning of the treatment on day 3 of the cycle as well as on days 7, 9, 10, and 11 of the cycle and more often if needed. During the ultrasound examination, the number of follicles >8 mm, >14 mm, and >18 mm, the diameter of each visible follicle, and the endometrial thickness (maximum distance between the echogenic interfaces of the myometrium) were measured. The endometrial thickness was measured in the plane through the central longitudinal axis of the uterus. The ultrasound examination was done by the same ultrasonographer, using the same ultrasound equipment (General Electric; Voluson 730 Expert; General Electric Healthcare, Waukesha, WI). The values of serum FSH and estradiol were measured on day 3 of the cycle for all of the women. Information about sperm concentration, sperm motility, and sperm morphologic features of the partners also were collected. For this study, a concentration R20 106/mL, motility grade a þ b R50%, and morphology R30% normal forms were considered normal values (14). Intrauterine Insemination and Outcomes Intrauterine insemination was performed 30 to 40 hours after hCG administration. Clinical pregnancy was confirmed by fetal cardiac pulsation 5 weeks after a positive pregnancy test. The study concluded with the confirmation of the delivery of a newborn. The main outcome measures included the number of follicles >14 mm and >18 mm, the total dose of recombinant FSH used in all the groups, and the endometrial thickness 1819
at different stages of the follicular phase. Pregnancy rate, number of multiple gestations, and miscarriages were also investigated. The cost of the gonadotropins with or without letrozole also was calculated. In the recombinant FSH only group, three women had two insemination cycles; in the group who received recombinant FSH þ 2.5 mg of letrozole, four women had two insemination cycles; and in the group who received recombinant FSH þ 5.0 mg of letrozole, six women had two insemination cycles. The number of insemination cycles did not affect the results of the study. Newborn Analysis Data from newborns such as birthweight, Apgar score at the first and fifth minutes of life, and gestational age were collected. Statistical Analysis The results are presented as mean standard deviation. The Bartlett test was used to obtain homogeneity of variances. If the data were not normally distributed, they were transformed or a nonparametric analysis was performed. If variances were homogeneous, an analysis of variance (ANOVA) test was used; if differences were observed between groups, the Scheffe test was used to determine differences between a pair of means. The pregnancy rate was compared among the groups using the chi-square test (Fisher’s exact test). Data from the newborns also were compared among the treatment groups (birthweight, gestational age, and Apgar score). A logistic regression analysis was performed to assesses the probability of pregnancy in women undergoing IUI with recombinant FSH, recombinant FSH þ 2.5 mg of letrozole, or recombinant FSH þ 5.0 mg of letrozole. P<.05 was considered statistically significant. RESULTS Characteristics of the Study Participants A total of 110 patients underwent 124 IUI cycles. To determine the homogeneity of the treatment groups, we evaluated the baseline characteristics of the patients, which showed that there were no statistically significant differences in chrono-
logic age, infertility time, or body mass index. Similarly, the baseline serum hormone levels were not statistically significantly different among the study groups (Table 1). According to the type of infertility of the couple, we determined that there are no statistically significant differences for the infertility factors among the study groups. However, male factor infertility predominated over the other factors: 58.33% in the recombinant FSH only group, 57.69% in the recombinant FSH þ 2.5 of letrozole group, and 48.97% in the recombinant FSH þ 5.0 mg of letrozole group (P>.05). In the same groups, female factor infertility was found in 16.67%, 19.23%, and 14.28% (P>.05), respectively, and unexplained infertility in 25.0%, 23.07%, and 34.69% (P>.05), respectively. One couple in the recombinant FSH þ 5.0 mg letrozole group had both male and female factor infertility. The female factors included 10 cases of polycystic ovary syndrome; of these, one was in the group treated with recombinant FSH only, three were in the group treated with recombinant FSH þ 2.5 mg of letrozole, and six were in the group treated with recombinant FSH þ 5.0 mg of letrozole. Six cases of endometriosis were found in the recombinant FSH only group, four in the group receiving FSH þ 2.5 mg of letrozole, and four in the group receiving recombinant FSH þ 5.0 mg of letrozole. When evaluating the sperm parameters (sperm concentration, sperm motility grade 3, and sperm normal morphology) of the partners, differences were observed among the groups. Sperm concentration in the recombinant FSH þ 5.0 mg of letrozole group was statistically significantly (P<.05) lower compared with the recombinant FSH only group, but was similar to the recombinant FSH þ 2.5 mg letrozole group (P>.05). Sperm motility in the recombinant FSH þ 5.0 mg letrozole group was statistically significantly (P<.05) lower compared with the recombinant FSH only group. No differences were observed in sperm motility between the group receiving recombinant FSH only and the group receiving recombinant FSH þ 2.5 mg of letrozole (P>.05). Sperm morphology was similar among all of the groups (P>.05) (Table 2). Effect of Letrozole on Recombinant FSH Dose The total recombinant FSH dose needed during the treatment was statistically significantly (P<.05) lower in both of the
TABLE 1 Baseline characteristics of the patients according to treatment group. Treatment group (no. of cycles)
Age (years)
Infertility time (months)
Body mass index (kg/m2)
FSH levels (mIU/mL)
Estradiol levels (pg/mL)
rFSH (39) rFSH þ L 2.5 (30) rFSH þ L 5.0 (55)
34.05 3.62 33.13 4.22 33.76 3.02
45.49 26.29 37.20 21.91 47.02 34.62
24.52 2.56 24.51 2.81 24.86 3.04
6.58 1.74 7.55 1.90 7.86 2.82
46.49 20.61 38.99 10.75 42.05 27.96
Note: Data are mean standard deviation. P>.05 between groups. L 2.5 ¼ 2.5 mg/day of letrozole for 5 days; L 5.0 ¼ 5.0 mg/day of letrozole for 5 days; rFSH, recombinant follicle-stimulating hormone. Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
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TABLE 2 Sperm parameters according to the treatment groups. Treatment groups rFSH rFSH þ L 2.5 rFSH þ L 5.0
Sperm concentration (3 106/mL)
Sperm motility grade a D b (%)
Sperm normal morphology (%)
74.72 45.37 68.83 53.45 55.16 42.65a
54.89 12.70 50.30 12.24 48.94 11.47b
33.44 14.50 30.81 12.18 30.02 9.09
Note: Data are mean standard deviations. L 2.5 ¼ 2.5 mg/day of letrozole for 5 days; L 5.0 ¼ 5.0 mg/day of letrozole for 5 days; rFSH, recombinant follicle-stimulating hormone. Motility a: rapid and linear progressive sperm motility. Motility b: slow or sluggish linear or nonlinear sperm movement. rFSH, recombinant follicle-stimulating hormone. a P< .05 versus rFSH (Mann-Whitney test). b P< .05 with respect to rFSH (analysis of variance). Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
letrozole cotreatment groups. When comparing both letrozole dosage groups, we found that the group receiving recombinant FSH þ 5.0 mg of letrozole required a statistically significant (P<.05) lower dose of recombinant FSH during the overall treatment compared with the group receiving letrozole at a dosage of 2.5 mg/day (Table 3). The days of stimulation with recombinant FSH were also reduced in the groups cotreated with letrozole (2.5 mg or 5.0 mg). The lowest value was observed with letrozole at 5.0 mg/day.
mm were observed when comparing the recombinant FSH þ 2.5 mg letrozole group with the recombinant FSH þ 5.0 mg letrozole group (P>.05). The endometrial thickness measured on the day of the hCG administration showed no differences among the three treatment groups (P>.05). In average, the cost of treatment per cycle with recombinant FSH alone was US$ 619.68; with FSH þ 2.5 mg/day of letrozole was US$ 499.69; and with FSH þ 5.0 mg of letrozole was US$ 259.77 (Table 3).
The number of follicles with diameter >8 mm was similar in the groups of study (P>.05). The number of ovarian follicles with diameter >14 mm was statistically significantly (P<.05) higher in the recombinant FSH only group compared with the recombinant FSH þ 5.0 mg letrozole group. There was no statistically significant difference in the number of follicles with diameter >14 mm between the letrozole cotreatment groups. The number of ovarian follicles with diameter >18 mm was statistically significantly higher (P<.05) in the recombinant FSH only group than in the groups combined with letrozole. No differences in the number of follicles >18
Dynamic of the Endometrial Thickness The analysis of endometrial thickness before hCG administration showed that it was comparable among the three groups at the beginning of the treatment (recombinant FSH only: 3.22 0.75 cm; recombinant FSH þ 2.5 mg letrozole; 3.25 0.75 cm; recombinant FSH þ 5.0 mg letrozole: 3.18 0.67 cm; P>.05). However, throughout most of the follicular phase, the endometrial thickness was statistically significantly (P<.05) reduced in both letrozole cotreatment groups compared with the recombinant FSH control group.
TABLE 3 Total dose of recombinant follicle-stimulating hormone (rFSH), days of stimulation with rFSH, sonographic characteristics, and rFSH cost. Treatment groups Total rFSH dose (IU) Days of stimulation with rFSH Follicles >8 mm Follicles >14 mm Follicles >18 mm Endometrial thickness (mm) rFSH cost (US$)
rFSH (no. of cycles [ 39)
rFSH D L 2.5 (no. of cycles [ 30)
rFSH D L 5.0 (no. of cycles [ 55)a,b
723.08 196.30 7.85 1.76 4.26 1.97 3.39 1.66 2.08 0.98 8.96 0.87 619.68 168.22
558.33 294.51a 5.63 2.16a 4.77 2.10 2.77 1.63 1.37 0.72a 8.63 0.94 499.69 252.40a
253.64 145.57a,b 3.13 1.45a,b 4.22 1.86 2.56 1.44a 1.47 0.61b 8.51 1.42 259.77 124.75a,b
Note: Data on total rFSH dose are mean standard deviation. a P< .05 versus FSH. b P< .05 versus FSH þ 2.5 mg of letrozole. Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
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This difference began to decrease 3 days before hCG administration but was still statistically significant (P<.05). By the day of the hCG administration, endometrial thickness was comparable among all the groups (recombinant FSH only: 8.96 0.87 cm; recombinant FSH þ 2.5 mg letrozole; 8.63 0.94 cm; recombinant FSH þ 5.0 mg letrozole; 8.51 1.42 cm) (Fig. 1). Pregnancies, Multiple Gestations, Miscarriages, and Births Pregnancy rates were similar when comparing the group treated with recombinant FSH alone and both letrozole groups (P>.05). In the recombinant FSH only group, one multiple gestation (one triplet) and two miscarriages occurred, and one of the triplets was a stillbirth. No miscarriages occurred in either letrozole group (Table 4). No multiple gestations occurred in the 2.5 mg/day letrozole group. One multiple gestation (twin) and one ectopic pregnancy occurred in the 5.0 mg/day letrozole group. In the FSH-only group, one preterm delivery occurred; in the letrozole groups, one preterm delivery occurred in the 2.5 mg/day group and one in 5.0 mg/day group. The birthweight and Apgar score values, measured on the newborn’s first and fifth minutes, were similar (P>.05) among all treatment groups (Table 4). One preterm delivery occurred in the FSH-only group, the 2.5 mg/day letrozole group, and the 5.0 mg/day letrozole group; however, the gestational age was comparable (P>.05) among all groups. One woman from the FSH-only group had preeclampsia. In the 5.0 mg/day letrozole group, one case with gestational diabetes occurred as well as one case where the umbilical cord was encircled around portions of the fetus.
FIGURE 1
Endometrial Thickness (mm)
Endometrial thickness measured in the follicular phase of cycles stimulated with recombinant folliclestimulating hormone (FSH) alone, 2.5 mg/day of letrozole plus FSH, and 5.0 mg/day of letrozole plus FSH. Day 0 is the day of human chorionic gonadotropin administration. 8.5
6.5 FSH
DISCUSSION Follicle-stimulating hormone is widely used in ovarian stimulation during assisted reproduction treatment. Pregnancy rates for women who have received FSH are statistically significantly higher compared with women who did not undergo controlled ovarian stimulation before an intrauterine insemination (IUI) (15). However, the use of FSH has raised the cost of these therapies considerably, and it is not free of side effects such as ovarian hyperstimulation syndrome or multiple pregnancies (16). Over the past few years, several studies have proved the effectiveness of the aromatase inhibitors (AIs) in ovulation induction because they reduce the levels of estradiol. Compared with clomiphene citrate, AIs have a short half-life (45 hours); they can be eliminated rapidly from the body (17) without causing the side effects that have been observed in cycles with clomiphene citrate (9). Cotreatment of recombinant FSH plus 2.5 mg/day of the AI letrozole for 5 days significantly reduces the FSH dose required (11). In groups of women with comparable age, duration of infertility, body mass index, and basal FSH and estradiol levels, our study showed that both doses of letrozole (2.5 or 5.0 mg/day) reduced the dose of recombinant FSH required for a stimulation cycle in a program of IUI. Overall, the larger letrozole dose (5.0 mg/day) required less FSH and reduced the number of days of stimulation when compared with the 2.5 mg/day dose. A review of different studies showed that letrozole reduced the FSH dose required from 45% to 55% (18). In our study, 2.5 mg/day of letrozole reduced the FSH dose required 77.17% and 5.0 mg/day of letrozole reduced it 35.07%. This reduction in the required dose of FSH was also associated with a lower number of ovarian follicles >14 mm than is obtained with recombinant FSH alone.
FSH + L 2.5
4.5
FSH + L 5.0
2.5
0.5 -9
-8
-7
-6
-5
-4
-3
-2
-1
0
Days from hCG Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
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After controlling for age; time of infertility; presence of male factor infertility, endometriosis, polycystic ovarian syndrome, or unexplained infertility; body mass index; total dosage of recombinant FSH; and days of recombinant FSH stimulation, the logistic regression analysis showed that the probability of pregnancy was similar for treatments using recombinant FSH along, recombinant FSH þ 2.5 mg/day of letrozole, or recombinant FSH þ 5.0 mg/day of letrozole (P>.05) (Table 5). After controlling for sperm parameters in the multivariable analysis, the probability of pregnancy rate was similar between women treated with recombinant FSH only and those treated with recombinant FSH and letrozole (data not shown).
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This finding is very important because with the use of letrozole at a dose of 5.0 mg/day the cost of the treatment is significantly reduced. The cost of FSH plus 2.5 mg/day of letrozole has been suggested to be more than half the value of FSH alone (12). Our study demonstrated that the reduction in cost of using FSH with 2.5 mg/day of letrozole was 80.63% less than with using FSH alone; using 5.0 mg/day of letrozole reduces the cost 41.92%. In consequence, the cost of each cycle of IUI is lower. This is important for developing countries
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TABLE 4 Pregnancy, multiple gestation, and miscarriage rates and newborn characteristics according to treatment group. Treatment groups
rFSH (no. of cycles [ 39)
rFSH D L 2.5 (no. of cycles [ 30)
rFSH D L 5.0 (no. of cycles [ 55)
Pregnancies/cycle completeda,b,c Births/cycle completedb,d Miscarriages/cycle completedb Gestational age (weeks)e Birthweight (g)e Apgar score (min 1)e Apgar score (min 5)e
7/39 17.9% (6–30) 4/39 10.3% (1–19) 2/39 5.1% (–2–12) 37.20 2.77 (4) 2938.0 885.2 (4) 8.40 1.3 (4) 9.40 0.6 (4)
2/30 6.7% (–2–20) 2/30 6.7% (–2–20) 0/30 0 (0–0) 35.10 6.93 (2) 2150.0 1343.5 (2) 9.00 0.0 (2) 9.50 0.7 (2)
13/55 23.6% (13–35) 9/55 16.4% (6–26) 0/55 0 (0–0) 38.39 0.99 (9) 3094.5 542.5 (9) 9.00 0.0 (9) 9.67 0.5 (9)
Note: L 2.5 ¼ 2.5 mg/day of letrozole for 5 days; L 5.0 ¼ 5.0 mg/day of letrozole for 5 days; rFSH, recombinant folliclestimulating hormone. a Pregnancy rates are expressed as percentages. b Between parentheses is the 95% confidence interval. P>.05 for pregnancy, birth, and miscarriage rates. c One ectopic pregnancy occurred in the 5.0 mg/day letrozole group. d Three preterm deliveries occurred, one in each of the three groups. e Gestational age, birthweight and Apgar scores are expressed as mean standard deviation; P>.05 between groups. One pregnancy in the rFSH-only group and three from the rFSH þ L 5.0 group are still in progress. Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
because the cost of assisted reproduction programs precludes access for a large segment of the poor population. The use of FSH alone is clearly a risk for ovarian hyperstimulation syndrome (16); however, our FSH-only group had no cases of ovarian hyperstimulation syndrome because we used a lower dose of recombinant FSH than has been used in other studies (20). In fact, our average dose of 723 IU
of recombinant FSH has resulted in a pregnancy rate of 17.9% per completed cycle; to obtain a similar pregnancy rate, another recent study required 1352 IU of recombinant FSH (19). The low number of ovarian follicles recruited with letrozole and the low dose of FSH required, both did not affect negatively the endometrial thickness at the time of hCG
TABLE 5 Logistic regression analysis to assess the probability of pregnancy in women treated with recombinant follicle-stimulating hormone (rFSH), rFSH D 2.5 mg/day of letrozole, or rFSH D 5.0 mg/day of letrozole. Variables Treatment rFSH rFSH þ L 2.5 mg/day rFSH þ 5.0 mg/day Age (years) Time of infertility (months) Infertility Male factor Endometriosis Polycystic ovary Unexplained infertility Body mass index (kg/m2) Total rFSH dose (IU) Days of rFSH stimulation
Unadjusted ORa
Adjusted ORa
Confidence interval at 95%
1.00 0.326 1.416 0.938 1.009
1.00 0.130 0.948 0.919 1.005
0.012–1.382 0.137–6.837 0.788–1.072 0.989–1.021
0.779 0.496 2.456 1.00 1.115 0.998 0.926
0.821 0.271 4.171 1.00 1.154 0.999 1.062
0.2731–2.473 0.0227–3.253 0.6605–26.34 1.00 0.9495–1.403 0.9940–1.004 0.6198–1.82
Note: Data are presented as unadjusted and adjusted odds ratio (OR). a P>.05 for each variable in the unadjusted or in the adjusted model. Noriega-Portella. Letrozole and recombinant FSH in IUI program. Fertil Steril 2008.
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administration. This is very important because a reduction of endometrial thickness is observed at the time of hCG administration after combined treatment with FSH and clomiphene citrate (9, 20). Similar (9, 21) or lower (20) pregnancy rates have been observed with clomiphene compared with AIs. Our study obtained similar pregnancy rates for the group receiving FSH alone compared with the groups receiving FSH plus 2.5 or 5.0 mg/day of letrozole. In fact, use of FSH with letrozole at 5.0 mg/day for 5 days resulted in a lower dose of FSH required, fewer ovarian follicles recruited, a similar endometrial thickness at the time of hCG administration, and similar pregnancy rate compared with FSH alone. Other investigators have obtained similar pregnancy rates when comparing groups treated with recombinant FSH alone and recombinant FSH þ 2.5 mg/day of letrozole in IUI programs (11, 19). Some investigators have found that the treatment with FSH plus clomiphene citrate results in similar pregnancy rates compared with FSH plus an AI such as letrozole (9, 21). However, it is also true that treatment with clomiphene citrate resulted in a higher number of miscarriages, ectopic gestations, and stillbirths (22) than treatment with letrozole. This is probably because clomiphene citrate has a long clearance half-life (of about 5 days to 3 weeks) and it may be stored in body fat (18), which means its antiestrogenic effect on the endometrium may be prolonged (23). In another study that used FSH with 5.0 mg/day of letrozole, FSH was administered from day 5 of the menstrual cycle, overlapping the letrozole treatment. Although the resulting pregnancy rates were not different among treatment groups, the addition of letrozole to gonadotropins was found to decrease the endometrial thickness (24), likely because the overlapping regimen advanced follicle growth and ovulation but did not allow time for clearance of the larger dose of letrozole by the time of hCG administration (18). In our protocol, in which FSH was administered from day 7 until the day of hCG administration, endometrial thickness was not affected by letrozole at the dose of 5.0 mg/day at the time of hCG administration. Endometrial thickness depends on estrogen, and AIs reduce the production of estradiol. Therefore, if letrozole inhibits the conversion of androstenedione to estradiol when administered and before clearance occurs, it may affect endometrial thickness. In fact, in our study, endometrial development was delayed in the first half of the follicular phase in both letrozole cotreated groups, as determined by the lower endometrial thickness found in these groups when compared with the results obtained in the group treated with FSH alone. When both letrozole groups are compared, the 5.0 mg/day dose delayed endometrial development even more than the 2.5 mg/day dose, and the endometrium required more days to recover from the estrogen reduction. However, the endometrium had completely recovered by the day of the hCG administration, indicating that the antiestrogen effect of letrozole had ended. This indicates that normal endometrial 1824
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development proceeds despite the lower estrogen levels, as confirmed by the comparable pregnancy rates observed among the study groups. Moreover, the fact that the pregnancies ended with live births suggests that the reduction in endometrial thickness in the first half of the cycle had no negative effect on pregnancy outcome. Although letrozole reduces estradiol levels during the follicular phase by inhibition of the aromatase enzyme, the pregnancy rate is not affected. Experimental studies have shown that adequate folliculogenesis is independent of estrogen but rather is conditioned on gonadotropin stimulation. Moreover, depletion of estradiol does not impair the developmental potential of the oocytes, including their fertilization and development into morula, blastocysts, and hatching blastocysts (25). Multiple gestations, a serious complication in assisted reproduction, derive from the use of gonadotropins. This has been confirmed in IUI programs in which FSH alone is used (19). Several studies have confirmed that the use of letrozole diminishes the incidence of multiple gestations (4). In our study, the use of FSH alone resulted in one multiple gestation of triplets, of which two were preterms and the other a stillbirth. One twin pregnancy occurred in the FSH þ letrozole groups, which completed at term without complications. Cotreatment of FSH with letrozole had no deleterious effect on the fetal development. The Apgar score is used to describe the condition of infants immediately after birth, and a 5-minute Apgar score of 7 to 10 is considered normal (26). In our study, all of the newborns had normal Apgar values, which were comparable among all the study groups. Our study was limited in that it was nonrandomized. However, all demographic variables were similar in our three groups (FSH alone, FSH þ 2.5 mg/day of letrozole, and FSH þ 5.0 mg/day of letrozole). Moreover, classification of the groups according to male factor, female factor, or unexplained infertility confirmed that groups were comparable and homogeneous. The only difference was that the sperm count and sperm motility of the male partners of the FSH þ letrozole 5.0 mg/day group were lower than in the other two groups; however, this was not a difficulty because the pregnancy rates did not differ. A number of adverse events are related to pregnancy, such as nutrition variations, preeclampsia, gestational diabetes, and labor complications. Our study had only one case of preeclampsia (in the FSH-only group) and one case of gestational diabetes (in the FSH þ 5.0 mg letrozole group). The 5.0 mg letrozole group also had a case of the umbilical cord encircling portions of the fetus. All three cases were treated by cesarean delivery. Irrespective of these factors, the rate of births per cycle completed was similar among the three groups. Women treated with recombinant FSH þ 5.0 mg/day of letrozole recruited fewer follicles and required a statistically significantly lower dosage of FSH, with no effect on the endometrial thickness at the time of hCG administration or the pregnancy rate. The treatment protocol of FSH þ letrozole
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at 5.0 mg/day should be considered a valid alternative for developing countries because the FSH required is reduced more than 50% over the 2.5 mg/day regimen, thus reducing the overall cost of the treatment with no impact on effectiveness. REFERENCES 1. Hong Y, Chen S. Aromatase inhibitors: structural features and biochemical characterization. Ann NY Acad Sci 2006;1089:237–51. 2. Dunkel L. Use of aromatase inhibitors to increase final height. Mol Cell Endocrinol 2006;254–5:207–16. 3. Holzer H, Casper R, Tulandi T. A new era in ovulation induction. Fertil Steril 2006;85:277–84. 4. Mitwally MF, Biljan MM, Casper RF. Pregnancy outcome after the use of an aromatase inhibitor for ovarian stimulation. Am J Obstet Gynecol 2005;192:381–6. 5. Bayar U, Basaran M, Kiran S, Coskun A, Gezer S. Use of an aromatase inhibitor in patients with polycystic ovary syndrome: a prospective randomized trial. Fertil Steril 2006;86:1447–51. 6. Begun MR, Quadir E, Begum A, Begum RA, Begum M. Role of aromatase inhibitor in ovulation induction in patients with poor response to clomiphene citrate. J Obstet Gynaecol Res 2006;32:502–6. 7. Jee BC, Ku SY, Suh CS, Kim KC, Lee WD, Kim KC, et al. Use of letrozole versus clomiphene citrate combined with gonadotropins in intrauterine insemination cycles: a pilot study. Fertil Steril 2006;85:1774–7. 8. Verpoest WM, Kolibianakis E, Papanikolaou E, Smitz J, Van Steirteghem A, Devroev P. Aromatase inhibitors in ovarian stimulation for IVF/ICSI: a pilot study. Reprod Biomed Online 2006;13:166–72. 9. Barroso G, Menocal G, Felix H, Rojas-Ruiz JC, Arsian M, Oehninger S. Comparison of the efficacy of the aromatase inhibitor letrozole and clomiphene citrate as adjuvants to recombinant follicle-stimulating hormone in controlled ovarian hyperstimulation: a prospective, randomized, blinded clinical trial. Fertil Steril 2006;86:1428–31. 10. Goswami SK, Das T, Chattopadhyay R, Sawhney V, Kumar J, Chaudhury K, et al. A randomized single-blind controlled trial of letrozole as a low-cost IVF protocol in women with poor ovarian response: a preliminary report. Hum Reprod 2004;19:2031–5. 11. Mitwally MF, Casper RF. Aromatase inhibition reduces gonadotrophin dose required for controlled ovarian stimulation in women with unexplained infertility. Hum Reprod 2003;18:1588–97. 12. Bedaiwy MA, Forman R, Mousa NA, Al Inany HG, Casper RF. Costeffectiveness of aromatase inhibitor cotreatment for controlled ovarian stimulation. Hum Reprod 2006;21:2838–44.
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