Assisted Conception Unit, St James's University Hospital, Leeds LS9 7TF, UK
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Abstract |
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Key words: GnRHa/hormonal suppression/IVF/pregnancy
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Introduction |
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The aim of this study was to compare three GnRH analogues: buserelin as a s.c. injection, nafarelin as an intranasal spray, both used routinely in this unit, and leuprorelin, as a long-acting depot preparation, all commenced in the early follicular phase in a long protocol regime. We particularly wanted to compare any evidence of escape from suppression between the long-acting depot with the shorter-acting analogues and also to investigate whether leuprorelin would have any detrimental effects in the luteal phase, because of its longer duration of action.
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Materials and methods |
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GnRHa regime
Group A patients (n = 20), received s.c. buserelin acetate (Hoechst, Hounslow, Middlesex, UK), at a dose of 0.5 mg daily. Group B patients (n = 20), received nafarelin intranasal spray (Searle, High Wycombe, UK), 100 µg 8-hourly. Both drugs were started on the first day of the menstrual period (day 1), and continued throughout the cycle until the day human chorionic gonadotrophin (HCG) was administered. Patients in group C (n = 20) received a single s.c. depot injection of leuprorelin acetate (Prostap SR; Searle, Lederle Laboratories, Gosport, Hants, UK) 3.75 mg on day 1. Ovarian stimulation in all groups commenced after 21 days of pituitary desensitization. Patients were considered desensitized when an ultrasound scan confirmed an endometrial thickness <3 mm.
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Clinical protocol |
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A venous blood sample was taken from each patient on the first day of GnRHa administration and every third day of the down-regulation and stimulation phases of the cycle, up until the day of HCG administration. These samples were analysed for serum concentrations of LH, follicle stimulating hormone (FSH), oestradiol and progesterone.
Hormonal assays
Serum FSH and LH were measured in all samples using a Technicon Immuno 1 automated clinical analyser employing a sandwich immunoassay format, with a detection limit of 0.1 mIU/l. Progesterone and oestradiol assays used a similar kit employing a competitive immunoassay format with a detection limit of 0.4 nmol/l and 37 pmol/l respectively. Serum HCG concentrations were determined with a commercial kit (AXSYM; Abbot Laboratories, Abbot Park, IL, USA) based on microparticle enzyme immunoassay (MEIA) technology. Sensitivity of the assay was 2.0 mIU/ml.
Statistical analysis
All data were expressed as mean ± SEM, and analysed using SPSS version 7.0. One-way analysis of variance (ANOVA), or multiple ANOVA (MANOVA) were used to assess results. Statistical significance was defined as P < 0.05 for all tests.
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Results |
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Figure 1ad, show serum hormone concentrations of oestradiol, FSH, LH and progesterone, expressed as mean ± SEM against time (days), in each group, in the down-regulation phase of the cycle. The curve for each group was analysed independently and then compared at each time-point with the curves for the other two groups. Cut-off limits for satisfactory pituitary/ovarian suppression were set at 2.5 U/l, 0.5 U/l, 130 pmol/l and 2 nmol/l for FSH, LH, oestradiol and progesterone respectively. There was no significant difference between the curves for FSH, LH or progesterone. However, for oestradiol (Figure 1a
), the suppression curve of oestradiol with buserelin was significantly different (P < 0.05) when compared with the curves for nafarelin or leuprorelin. Patients on buserelin took longer for their oestradiol to be suppressed to the cut-off limit of 130 pmol/l, though by day 21 of the down-regulation phase, all three analogues had achieved comparable suppression.
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Discussion |
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Our results agree with those of Penzias et al. (1992), Balasch et al. (1992), Tarlatzis et al. (1994a) and Porcu et al. (1995), i.e. long-acting depot preparations are as effective in pituitary suppression as the short-acting analogues. We found no significant difference in suppression of FSH, LH, oestradiol or progesterone with any of the three analogues after 21 days of administration. We also found that the number of patients suppressed as well as the percentage suppression of serum oestradiol to defined cut-off values, by day 15, was superior with nafarelin or leuprorelin when compared to patients on buserelin. The incidence of cyst formation in all three groups was similar (nafarelin = 10%; buserelin = 10%; leuprorelin = 15%). Tarlatzis et al. (1994b) have shown that cyst formation with GnRHa use is associated with elevated serum oestradiol concentrations. The delayed suppression with persistence of cysts observed with the patients on buserelin could indicate failure of buserelin to deal effectively with this adjunctive oestrogen production. We found no evidence of escape from suppression with any of the three analogues, comparing the number of viable oocytes retrieved or fertilized. In addition, leuprorelin did not cause profound suppression by comparable gonadotrophin dose and duration of stimulation.
One of the recognized complications of analogue use in a long protocol is the risk of luteal phase dysfunction from premature luteolysis. Inhibition of luteal steroidogenesis by GnRHa has been described by Smitz et al. (1987). Short-acting analogues such as buserelin or nafarelin have been traditionally favoured because of their brevity of action. At cessation of administration they rapidly disappear from the circulation and a normal pituitary response can be shown within a few days (Porcu et al., 1995). Any effect on the luteal phase is thus minimal. Depot preparations have a longer duration of action, ~7 weeks (Broekmans et al., 1992
; Porcu et al., 1995
), with potentially adverse consequences on corpus luteal function. A report by Bourgain et al. (1994) investigated the influence of GnRHa on the endometrium and endometrial development. Endometrial histological maturation, ultrastructure, oestrogen and progesterone receptor status were analysed in the mid-luteal phase in GnRHa/HMG cycles with and without luteal supplementation. They found that supplementation with HCG or progesterone produced fewer signs of luteal phase deficiency compared to non-supplemented cycles. All the patients in this study received luteal phase support with either HCG or progesterone, and we found no difference in the miscarriage rates of patients on leuprorelin as compared with either nafarelin or buserelin. Filicori et al. (1996) have noted that depot GnRHa administration that provides measurable drug concentrations in early pregnancy is not associated with an increased abortion rate. Moreover we did not find any difference in implantation rates or pregnancy rates between the three agonists. Although the presence of GnRH and its receptor has been established in endometrium, until recently it has been unknown whether these receptors are present on preimplantation embryos. Inadvertent exposure of concepti to GnRHa does not appear to have a deleterious effect (Tolis et al., 1981
; Skarin et al., 1982
; Testart et al., 1993
). Gartner et al. (1997) have even suggested that analogues may be beneficial to implantation. A recent study by Raga et al. (1998) has demonstrated the presence of GnRHa receptors on preimplantation embryos at the mRNA and protein levels. Their randomized study between administration or omission of GnRHa until 6 weeks of gestation, with luteal support, showed that administration of GnRHa throughout the luteal phase and early pregnancy had a positive influence on pregnancy and implantation outcome. Direct effects of long-acting depot preparations on the embryo can now be viewed in a fuller context and seem unlikely to be detrimental. Furthermore we noted no difference in the mean numbers of embryos formed or cleaved in those patients on leuprorelin compared to the shorter acting analogues.
Our study shows that leuprorelin, nafarelin and buserelin are equally effective when pituitary suppression, ovarian stimulation and IVF outcome are compared. Although satisfactory suppression with nafarelin and leuprorelin occurred after 2 weeks, buserelin required an additional week to achieve equivalent results. However, after 3 weeks there was no significant difference between the three analogues.
GnRH antagonists, particularly the cyclic compounds which have a potency comparable with the linear agonists, may become more widespread in general clinical use. Their advantages include no initial flare response due to an immediate competitive pituitary receptor occupancy (Rivier et al., 1996). Further work in rat models may ultimately lead to the availability of GnRH antagonists with oral activity and minimal histamine-releasing side-effects (Rivier et al., 1996
), making them eminently more attractive.
At present, however, the decision on which GnRH agonist to use will ultimately depend on patient and clinician preference, side-effects, availability and cost.
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Notes |
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References |
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Submitted on March 27, 1998; accepted on October 28, 1998.