This article is part of the series Aromatase inhibitors in reproductive biology and endocrinology.

Open Access Research

Hormonal, follicular and endometrial dynamics in letrozole-treated versus natural cycles in patients undergoing controlled ovarian stimulation

Mohamed A Bedaiwy12, Mahmoud A Abdelaleem2, Mostafa Hussein2, Noha Mousa3, Lisa N Brunengraber1 and Robert F Casper3*

Author Affiliations

1 Department of Obstetrics and Gynecology, University Hospitals Case Medical Center, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA

2 Assiut University, Assiut, Egypt

3 Reproductive Sciences Division, Department of Obstetrics and Gynecology, University of Toronto, Toronto, Canada

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Reproductive Biology and Endocrinology 2011, 9:83  doi:10.1186/1477-7827-9-83


The electronic version of this article is the complete one and can be found online at: http://www.rbej.com/content/9/1/83


Received:19 August 2010
Accepted:21 June 2011
Published:21 June 2011

© 2011 Bedaiwy et al; licensee BioMed Central Ltd.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

The objective of this study was to compare letrozole-stimulated cycles to natural cycles in 208 patients undergoing intrauterine insemination (IUI) between July of 2004 and January of 2007. Group I (n = 47) received cycle monitoring only (natural group), Group II (n = 125) received letrozole 2.5 mg/day on cycle days three to seven, and Group III (n = 36) received letrozole 5 mg/day on cycle days three to seven. There were no differences between the groups in endometrial thickness or P4 on the day of hCG. Estradiol levels had higher variation in the second half of the follicular phase in both letrozole-treated groups compared to the control group. Estradiol per preovulatory follicle was similar in both letrozole cycles to that observed in the natural cycles. LH was lower on the day of hCG administration in the letrozole 2.5 mg/day group vs. the natural group. In summary, letrozole results in some minor changes in follicular, hormonal and endometrial dynamics compared to natural cycles. Increased folliculogenesis and pregnancy rates were observed in the letrozole-treated groups compared to the natural group. These findings need to be confirmed in larger, prospective studies.

Background

For over 40 years, the first-line therapy for ovulation induction (OI) has been clomiphene citrate (CC) [1]. Its inherent properties such as low price, oral route of administration and high ovulation success rate (60-90%) make it an attractive therapy. However, the pregnancy rate is [2] disappointing. Sub-optimal pregnancy rates with CC have been attributed to peripheral anti-estrogenic effects, mainly on the endometrium and the cervical mucus[3] Gonadotropins are more effective in ovulation induction and are associated with higher pregnancy rates than CC, but are expensive and carry higher risk for ovarian hyperstimulation syndrome and multiple gestations [4].

Newer options for ovulation induction are the third-generation aromatase inhibitors (AIs), the most commonly used being letrozole. Initially introduced to treat postmenopausal breast cancer, AIs are now also being used for ovulation induction or enhancement. A recent meta-analysis addressing the use of letrozole in assisted conception concluded that letrozole is as effective as other methods of ovulation induction [5]. When letrozole is used in combination with gonadotropins, it leads to lower gonadotropin requirements and pregnancy rates similar to gonadotropin treatment alone [6]. In a study comparing combined therapy of letrozole (2.5 mg/day or 5.0 mg/day) and recombinant FSH with recombinant FSH alone in an intrauterine insemination (IUI) program, 5 mg/day of letrozole was more cost-effective than the 2.5 mg/day in co-treatment with no adverse effect on pregnancy rate or outcome [7].

Aromatase inhibitors for ovulation induction are orally administered, and are relatively inexpensive with minor side effects such as very infrequent headaches and leg cramps[8]. Aromatase inhibitors increase endogenous FSH production in response to decreased estrogen biosynthesis in the ovary and extraovarian tissues, including the brain [9]. Because they do not deplete estrogen receptors like CC, normal central feedback mechanisms remain intact[10]. As the dominant follicle grows and estrogen levels rise, normal negative feedback occurs centrally resulting in suppression of FSH and atresia of the smaller growing follicles. Therefore, a single dominant follicle, and mono-ovulation, is the rule in most cases[11] with the clear advantage of reducing multiple-gestation pregnancies.

Compared to CC, letrozole has been associated with lower preovulatory estradiol (E2) levels [12], as well as thicker endometrium and a trend towards higher pregnancy rates[13]. Standard ovarian stimulation protocols often produce high preovulatory E2 levels that could adversely affect the development of the endometrium, the follicles, and the embryo. Therefore, the lower E2 when using AIs may lead to an improvement of implantation [14,15].

There have been several studies comparing letrozole to CC. However, there is a paucity of research comparing letrozole with natural cycles. Larger studies comparing CC, letrozole, and natural cycles in a single study are necessary to further characterize the effect of letrozole on hormonal dynamics and pregnancy rates. Our hypothesis is that letrozole-treated cycles mimic natural cycles in hormonal and endometrial parameters. The aim of this study was to compare cycle dynamics in letrozole-treated versus natural cycles in infertile patients undergoing intrauterine insemination (IUI).

Methods

Patient recruitment and counseling

We conducted a retrospective cohort study of 208 consecutive infertile patients who were recruited to participate. Briefly, patients underwent IUI between July of 2004 and January of 2007 at the Toronto Center For Assisted Reproductive Technology, Toronto, Canada. Patients were divided into three groups. The first group (n = 47) received cycle monitoring only (natural cycle; group I). The 2 remaining groups received letrozole on cycle days three to seven at either 2.5 mg/day (n = 125) (group II) or 5 mg/day (n = 36) (group III). An informed consent was obtained from all participants clearly denoting the off-label use of the medication prior to treatment. Subsequently IRB approval was obtained to use their data for the purposes of this study.

Causative factors of infertility were investigated and defined as follows. Tubal patency was confirmed by sonohysterography with contrast, hysterosalpingography, and or pelvic laparoscopy. Mild male factor infertility was diagnosed according to the World Health Organization (WHO) criteria for normal semen[16]. Endometriosis was diagnosed by pelvic laparoscopy. Unexplained infertility was based on the exclusion of known factors of infertility.

Cycle monitoring and insemination

All patients were followed with serial measurements of serum estradiol (E2), progesterone (P4), and luteinizing hormone (LH) using a radioimmunoassay (RIA) kit. Transvaginal ultrasonography (TVS) was performed for follicular diameter tracking and measurement of endometrial thickness. We measured follicular diameter in 2 perpendicular planes and calculated the mean, while endometrial thickness was measured in the sagittal plane at the widest part of endometrial cavity. Serum samples were obtained and TVS performed on cycle days three, seven, once between days 9 and 11, the day of human chorionic gonadotropin (hCG) administration, and when a follicle achieved a diameter of ≥16 mm. Serum follicle stimulating hormone (FSH) was measured on day 3 only. The results for E2 and P4 levels were reported as picomoles per liter and nanomoles per liter, respectively, and LH and FSH levels were reported in international units per liter.

An LH surge was defined as an increase in LH level greater than 100% over the mean of the preceding 2 measures. IUI was performed 36-40 hours after hCG administration if no endogenous LH surge occurred. If an endogenous LH surge was detected on the day of hCG administration, two IUIs were performed at 24 and 48 h. Pregnancy was diagnosed by quantitative β-hCG assay two weeks after the insemination. Clinical pregnancy was confirmed by observing fetal cardiac pulsation four weeks after positive pregnancy test by TVS.

Outcome parameters

Hormonal outcome measures were E2, P4, LH and FSH. Non-hormonal outcomes were number and size of growing follicles, endometrial thickness, and pregnancy. All three treatment groups were compared to each other for hormonal, endometrial and follicular dynamics, while the two groups receiving letrozole were combined for comparison against the natural cycle group for pregnancy rates.

Statistical analysis

Data management was done using a preprepared Excel data spreadsheet. Outcome measures are expressed as mean ± standard deviation from the mean (SD). Statistical significance for continuous variables was calculated using ANOVA test. Categorical variables were compared using the χ2 and Fisher's Exact Test. P < 0.05 was considered statistically significant. Statistical analysis was performed with SPSS (Release 14.01, SPSS Inc., Chicago, IL).

Results

Patients

The study included 208 patients who underwent a total of 300 consecutive IUI cycles. There were 71 cycles in the natural cycle group, 179 cycles in the letrozole 2.5 mg/day group, and 50 cycles in the letrozole 5 mg/day group (table 1). Male factor and unexplained infertility were the most common indications for IUI. Other identified causes of infertility included endometriosis, and polycystic ovarian syndrome. The most frequent cause of cycle cancellation differed among the patient groups. The most common cause of cancellation in the natural cycle group was anovulation, while in the letrozole 2.5 mg/day group, the main cause was presence of ovarian cysts on cycle day 3 before the start of stimulation. The most common cause of cancellation in the letrozole 5 mg/day group was an elevated serum FSH on cycle day 3 before the start of stimulation.

Table 1. Study demographics

Hormonal dynamics

Values for E2, P4 and LH are shown in table 2. The letrozole 5 mg/day group had significantly lower E2 on day seven, but significantly higher E2 on the day of hCG administration when compared to the natural group (P = 0.025 and 0.041, respectively). However, the E2 per preovulatory follicle was similar in all three groups on the day of hCG. On days 9 to 11 and the day of hCG administration, E2 had a larger variability, as evidenced by larger standard deviations, in the letrozole groups compared to the natural group (Table 2). LH was significantly lower on the day of hCG administration in the letrozole 2.5 mg/day group compared to the natural group (p = 0.000). P4 showed no differences between any treatment groups.

Table 2. Hormonal dynamics: follicular phase levels of serum estradiol, progesterone, and LH

Follicular dynamics

There were no statistically significant differences in follicular diameter between the three groups until cycle day seven (table 3).Compared to the natural group, both the letrozole 2.5 mg/day and 5 mg/day groups had significantly more follicles ≥ 10 mm on day 7 (p = 0.0001 and 0.0001, respectively), more follicles ≥ 12 mm on days 9-11 (p = 0.0001 and 0.005), and more follicles ≥ 15 mm on the day of hCG administration (p = 0.0001 and 0.0001).

Table 3. Follicular dynamics in study groups

Endometrial dynamics

When comparing endometrial thickness, neither letrozole group differed significantly from the natural group on any day (table 4).

Table 4. Endometrial thickness (cm) in study groups

Pregnancy rate

Three out of 63 natural cycles (4.8%) and 22 out of 185 letrozole cycles (12%) resulted in pregnancy (table 5). This difference was statistically significant (p = = 0.02). Although only a small number of cycles were repeated as part of this study, the cumulative pregnancy rate was 13.7% in the letrozole groups, significantly higher than the natural group rate of 6.4% (p = 0.01). There were 3 twin pregnancies in the 2 letrozole groups and none in the natural cycle group.

Table 5. Pregnancy rate in study groups

Discussion

Endometrial sparing, increased folliculogenesis, and an increase in pregnancy rate were observed in our letrozole-treated patients as compared to naturally-cycling patients. The endometrial-sparing effect of letrozole is well-demonstrated in this study by the lack of significant difference in endometrial thickness compared to the natural cycle group, and also has been confirmed by previous studies [17,18]. In addition, our ovulation monitoring findings agree with a previous study showing multifollicular development and better pregnancy outcomes with the use of letrozole 5 mg/day [14,17]. Our finding that serial serum progesterone levels were comparable between the three groups agrees with a previous study [18], and suggests that letrozole does not have a premature luteinizing effect on the developing follicle. Table 6 summarizes the results of some trials using letrozole as an ovulation inducing agent in different clinical scenaria with different results and conclusions.

Table 6. Summary of randomized trials assessing the efficacy of letrozole

Although basal LH levels were comparable among the 3 groups, there was a trend towards higher LH (although non-significant) on day 7 in the letrozole groups. This rise might be due to the release of the anterior pituitary from the negative feedback of E2. Three important observations deserve mentioning. First, the increase in LH is still well-below the levels for definition of premature LH surge. Second, after discontinuation of letrozole (days 9-11), serum LH returned to levels similar to the non-stimulated group reflecting the short half life of the letrozole. Thirdly, a natural LH surge was observed more frequently in non-stimulated cycles than in the letrozole stimulated cycles. This finding is likely artifactual since we suggest hCG administration to all women undergoing IUI so that timing of insemination can be optimized. The women undergoing natural cycle monitoring usually requested everything to be natural including no hCG trigger so the difference between the groups is unlikely to be related to the letrozole. Spontaneous LH surges do occur with letrozole for ovulation induction.

This study has some limitations, the most important of which include that it is retrospective, non-powered, non-randomized and not blinded. These issues are inherent in retrospective studies. However, we believe the results are of interest since there are few studies comparing natural cycles to letrozole-stimulated cycles. The main distinguishing features of letrozole as an ovulation inducing agent is its endometrial sparing effect and the early cycle multi-follicular development that may have translated into a better pregnancy rate in the present study. In addition, a recent study demonstrated that letrozole improves blood flow compared to CC and this observation may also be associated with improved pregnancy rates [19]. A well-designed and powered randomized clinical trial will be needed to confirm this result.

The authors declare that they have no competing interests.

Competing interests

The authors declare that they have no competing interests.

Authors' contributions

MAB collected and analyzed the data and drafted the manuscript. MAA Collected the data and drafting the manuscript. MH participated in the data collection. NM participated in the design of the study and performed the statistical analysis. LNB: participated in the data collection and helped to draft the manuscript. RC Conceived the idea of the study, and participated in its design and coordination and helped to draft the manuscript. All authors read and approved the final manuscript.

References

  1. Casper RF: Aromatase inhibitors in ovarian stimulation.

    J Steroid Biochem Mol Biol 2007, 106:71-75. PubMed Abstract | Publisher Full Text OpenURL

  2. Neveu NGL, St.-Michel P, Lavoie HB: Comparison of clomiphene citrate, metformin, or the combination of both for first-line ovulation induction and achievement of pregnancy in 154 women with polycystic ovary syndrome.

    Fertil Steril 2007, 87:113-120. PubMed Abstract | Publisher Full Text OpenURL

  3. Gorlitsky GAKN, Speroff L: Ovulation and pregnancy rates with clomiphene citrate.

    Obstet Gynecol 1978, 51:265-269. PubMed Abstract | Publisher Full Text OpenURL

  4. Guzick DSCS, Coutifaris C, Overstreet JW, Factor-Litvak P, Steinkampf MP, Hill JA, Mastroianni L, Buster JE, Nakajima ST, Vogel DL, Canfield RE: Efficacy of superovulation and intrauterine insemination in the treatment of infertility. National Cooperative Reproductive Medicine Network.

    N Engl J Med 1999, 340:177-183. PubMed Abstract | Publisher Full Text OpenURL

  5. Requena A, Herrero J, Landeras J, Navarro E, Neyro JL, Salvador C, Tur R, Callejo J, Checa MA, Farre M, Espinós JJ, Fábregues F, Graña-Barcia M: Use of letrozole in assisted reproduction: a systematic review and meta-analysis.

    Hum Reprod Update 2008, 14:571-582. PubMed Abstract | Publisher Full Text | PubMed Central Full Text OpenURL

  6. Holzer H, Casper R, Tulandi T: A new era in ovulation induction.

    Fertil Steril 2006, 85:277-284. PubMed Abstract | Publisher Full Text OpenURL

  7. Noriega-Portella L, Noriega-Hoces L, Delgado A, Rubio J, Gonzales-Castaneda C, Gonzales GF: Effect of letrozole at 2.5 mg or 5.0 mg/day on ovarian stimulation with gonadotropins in women undergoing intrauterine insemination.

    Fertil Steril 2008, 90:1818-1825. PubMed Abstract | Publisher Full Text OpenURL

  8. Hamilton A, Piccart M: The third-generation non-steroidal aromatase inhibitors: a review of their clinical benefits in the second-line hormonal treatment of advanced breast cancer.

    Ann Oncol 1999, 10:377-384. PubMed Abstract | Publisher Full Text OpenURL

  9. Naftolin : Brain aromatization of androgens.

    J Reprod Med 1994, 39:257-261. PubMed Abstract OpenURL

  10. Robert F, Casper MFMM: Aromatase Inhibitors for Ovulation Induction.

    J Clin Endocrinol Metab 2006, 91:760-771. PubMed Abstract | Publisher Full Text OpenURL

  11. Casper RF: Letrozole versus clomiphene citrate: which is better for ovulation induction?

    Fertil Steril 2009, 92:858-9. PubMed Abstract | Publisher Full Text OpenURL

  12. 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, 86:1447-1451. PubMed Abstract | Publisher Full Text OpenURL

  13. Serdar B: Ovulation induction in women with infertility: a new indication for aromatase inhibitors.

    Fertil Steril 2003, 80(6):1338. PubMed Abstract | Publisher Full Text OpenURL

  14. Simón CCF, Valbuena D, Remohí J, Pellicer A: Clinical evidence for a detrimental effect on uterine receptivity of high serum oestradiol concentrations in high and normal responder patients.

    Hum Reprod 1995, 10:2432-2437. PubMed Abstract | Publisher Full Text OpenURL

  15. Simon C, Cano F, Valbuena D, Remohi J, Pellicer A: Clinical evidence for a detrimental effect on uterine receptivity of high serum oestradiol concentrations in high and normal responder patients.

    Hum Reprod 1995, 10:2432-2437. PubMed Abstract | Publisher Full Text OpenURL

  16. Organization WH: WHO laboratory manual for the examination of human semen and sperm-cervical mucus interaction. 4th edition. Cambridge: Cambridge University Press; 1999. OpenURL

  17. Bedaiwy MA, Mousa NA, Casper RF: Aromatase inhibitors prevent the estrogen rise associated with the flare effect of gonadotropins in patients treated with GnRH agonists.

    Fertil Steril 2009, 91:1574-1577. PubMed Abstract | Publisher Full Text OpenURL

  18. Bedaiwy MA, Shokry M, Mousa N, Claessens A, Esfandiari N, Gotleib L, Casper R: Letrozole co-treatment in infertile women 40 years old and older receiving controlled ovarian stimulation and intrauterine insemination.

    Fertil Steril 2009, 91:2501-2507. PubMed Abstract | Publisher Full Text OpenURL

  19. Baruah J, Roy KK, Rahman SM, Kumar S, Sharma JB, Karmakar D: Endometrial effects of letrozole and clomiphene citrate in women with polycystic ovary syndrome using spiral artery Doppler.

    Arch Gynecol Obstet 2009, 279:311-314. PubMed Abstract | Publisher Full Text OpenURL

  20. Al-Fozan H, Al-Khadouri M, Tan SL, Tulandi T: A randomized trial of letrozole versus clomiphene citrate in women undergoing superovulation.

    Fertil Steril 2004, 82:1561-1563. PubMed Abstract | Publisher Full Text OpenURL

  21. Barroso G, Menocal G, Felix H, Rojas-Ruiz JC, Arslan 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, 86:1428-1431. PubMed Abstract | Publisher Full Text OpenURL

  22. Bayar U, Tanriverdi HA, Barut A, Ayoglu F, Ozcan O, Kaya E: Letrozole vs. clomiphene citrate in patients with ovulatory infertility.

    Fertil Steril 2006, 85:1045-1048. PubMed Abstract | Publisher Full Text OpenURL

  23. Atay V, Cam C, Muhcu M, Cam M, Karateke A: Comparison of letrozole and clomiphene citrate in women with polycystic ovaries undergoing ovarian stimulation.

    J Int Med Res 2006, 34:73-76. PubMed Abstract | Publisher Full Text OpenURL

  24. Badawy A, Abdel Aal I, Abulatta M: Clomiphene citrate or letrozole for ovulation induction in women with polycystic ovarian syndrome: a prospective randomized trial.

    Fertil Steril 2009, 92:849-52. PubMed Abstract | Publisher Full Text OpenURL

  25. Badawy A, Shokeir T, Allam AF, Abdelhady H: Pregnancy outcome after ovulation induction with aromatase inhibitors or clomiphene citrate in unexplained infertility.

    Acta Obstet Gynecol Scand 2009, 88:187-191. PubMed Abstract | Publisher Full Text OpenURL

  26. Badawy A, Elnashar A, Totongy M: Clomiphene citrate or aromatase inhibitors for superovulation in women with unexplained infertility undergoing intrauterine insemination: a prospective randomized trial.

    Fertil Steril 2009, 92:1355-1359. PubMed Abstract | Publisher Full Text OpenURL

  27. Sh Tehrani Nejad E, Abediasl Z, Rashidi BH, Azimi Nekoo E, Shariat M, Amirchaghmaghi E: Comparison of the efficacy of the aromatase inhibitor letrozole and clomiphen citrate gonadotropins in controlled ovarian hyperstimulation: a prospective, simply randomized, clinical trial.

    J Assist Reprod Genet 2008, 25:187-190. PubMed Abstract | Publisher Full Text | PubMed Central Full Text OpenURL

  28. Topipat C, Choktanasiri W, Jultanmas R, Weerakiet S, Wongkularb A, Rojanasakul A: A comparison of the effects of clomiphene citrate and the aromatase inhibitor letrozole on superovulation in Asian women with normal ovulatory cycles.

    Gynecol Endocrinol 2008, 24:145-150. PubMed Abstract | Publisher Full Text OpenURL

  29. Begum MR, Ferdous J, Begum A, Quadir E: Comparison of efficacy of aromatase inhibitor and clomiphene citrate in induction of ovulation in polycystic ovarian syndrome.

    Fertil Steril 2009, 92:853-7. PubMed Abstract | Publisher Full Text OpenURL