Prospective study of a modified gonadotropin-releasing hormone agonist long protocol in an in vitro fertilization program

FERTILITY AND STERILITY Copyright

©

Vol. 61, No.4, April 1994

1994 The American Fertility Society

Printed on acid-free paper in U.

s.

A.

Prospective study of a modified gonadotropin-releasing hormone agonist long protocol in an in vitro fertilization program

Kostas Pantos, M.D.* Toula Meimeth-Damianaki, M.S. Terpsi Vaxevanoglou, M.S. Emmanuel Kapetanakis, M.D. Infertility Centre of Athens, Athens, Greece

Objective: To determine if pituitary suppression is still maintained if GnRH agonist (GnRH -a) is discontinued as gonadotropin stimulation is begun in a long protocol. Design: Prospective, randomized study. Setting: An outpatient IVF-GIFT program. Patients: One hundred seventy-three patients entering an IVF-GIFT program. Interventions: Gonadotropin-releasing hormone agonist in long protocol was either discontinued or continued as gonadotropin stimulation was begun. Main Outcome Measures: Luteinizing hormone (LH), E 2 , and P levels, egg numbers, fertilization rate, number of embryos transferred, day of gonadotropin stimulation, and pregnancy rates (PRs). Results: Pituitary suppression was maintained although GnRH -a was discontinued as gonadotropin stimulation was begun. No spontaneous LH surge was seen, and PRs were increased in the IVF patients. Conclusion: This study indicates that the advantages gained from use of GnRH-a in the long protocol are not compromised by its early discontinuation. Fertil Steril 1994;61:709-13 Key Words: Gonadotropin-releasing hormone agonist, in vitro fertilization, gamete intrafallopian transfer

An increase in pregnancy rates (PRs) in IVF programs by the combined use of GnRH agonists (GnRH-a) and hMG has been reported (1, 2). In addition, the use of GnRH-a has been associated with a reduction in cancellation rates, prevention of premature endogenous LH surges, avoidance of premature luteinization, and an increase in multiple follicular growth (3). Various types of protocols for GnRH -a use have been developed. In the "short protocol" (4), the advantage of the initial stimulation phase of the go-

Received April 5, 1993; revised and accepted November 19, 1993. * Reprint requests: Kostas Pantos, M.D., Infertility Centre of Athens, 282 Kifissias Avenue, Halandri, Athens, Greece GR15232.

nadotropins is used to recruit follicles. In the "long protocol" (3), the ovarian stimulation is initiated after pituitary desensitization with GnRH-a. The GnRH-a is then generally continued up to heG administration to ensure constant pituitary suppression. Both of the above protocols have been widely used in IVF programs. Further, an "ultrashort" protocol has been described (5) in which the agonist is administered for only 3 days during the menstrual phase and is immediately followed by hMG. This protocol is characterized by its simplicity and cost reduction. Most recently, a new protocol (6) has been described in which the GnRH-a in a long protocol was discontinued as hMG was started, and a favorable description of a cost analysis was presented. Macnamee et al. (7), in his description of an

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Modified GnRH-a long protocol in IVF

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ultrashort protocol, reported no evidence of an endogenous LH surge, although they stopped GnRHa administration early in their cycle. This provided the incentive for us to devise a prospective trial comparing the cessation of GnRH -a in the long protocol at the time of hMG stimulation with the continuation of GnRH-a during hMG stimulation. This was done to examine if any of the advantages gained from use of GnRH -a in the long protocol would be compromised by halting its administration at the time of hMG stimulation and what the clinical significance of such a measure would be. MATERIALS AND METHODS

In a prospective study during the 6-month period between January 1992 and June 1992, 173 women aged 21 to 42 years were scheduled for IVF-ET or G 1FT using the long buserelin acetate protocol at the Infertility Centre of Athens, Athens, Greece. All patients had prior complete infertility investigation and treatment. Indications for IVF-ET were those of tubal infertility, male factor infertility, and antibody-related infertility. Indications for GIFT were unexplained and endometriosis-related infertility. The women in this study were randomly assigned into two groups based on alternate IVF -program numbers on commencement of treatment, regardless of cause of infertility or method of assisted conception. Group 1 consisted of 82 patients who received buserelin acetate (Suprefact; Hoechst, AG, Frankfurt am Main, Germany) 0.5 mgjd SC injection for 10 days commencing on day 20 or 21 of an idealized 28-day cycle (Fig. 1). A satisfactory suppression was checked by assaying E2 serum levels. If E2 was >50pgjmL (0.18 nmoljL), then the busere-

E2

!

OnRH·a

E2. LH. P

!

g~up1 ~r----------~h:M:O;~;<;
(821

I

lin acetate injections were continued until E2 was <50 pgjmL (0.18 nmoljL). Once suppression had been achieved, 300 IUjd 1M of hMG (Humegon; N.V. Organon, Oss, Holland) was commenced and continued until at least three follicles> 17 mm diameter were seen by vaginal ultrasound (US). At this point, both hM G and GnRH -a were halted, and ovulation was induced with 10,000 IU 1M of hCG (Pregnyl; N.V. Organon). Oocyte retrieval was performed 36 hours after the hCG injection with the aid of US and light IV sedation. Group 2 consisted of 91 patients who were treated similarly except that the GnRH -a suppression was halted on the day of hMG stimulation and the hMG was continued alone (Fig. 1). In both groups, the treatment was monitored by vaginal US scans only. On the morning of hCG administration, a blood sample was taken and E 2 , P, and LH were measured. All measurements were performed by a local clinical laboratory using standard immunoassay techniques. The resulting values were kept aside for the purposes of this study and were not used to determine the day of hCG administration. If IVF was performed, a maximum of four embryos were transferred into the uterus 48 hours after oocyte retrieval. The GIFT procedures were performed in the operating theater by laparoscopy under general anesthesia, and four to five oocytes (depending on oocyte quality) were replaced in the tubes. Supernumerary eggs were then exposed to fertilization and subsequently were cryopreserved for future use. All patients received luteal support in the form of hCG 1,500 IU 1M on days 3, 6, and 9 after oocyte retrieval. Pregnancy was detected by rising {J-hCG blood levels 14 days after IVF-ET or 16 days after GIFT was performed. All pregnancies were confirmed by the demonstration of gestational sac by US scan 4 weeks after oocyte retrieval. Statistical methods included Mann -Whitney nonparametric analysis, and paired comparisons were made with Student's t-test, x2 , and Fisher's exact tests.

E2. LH. P

!

OnRH·a g~up2

(911

~

hMO~hCO

I

~--------. x 10 days

I

--------i

Figure 1 Treatment protocols for group 1 (continuing GnRH-a) and group 2 (discontinuing GnRH-a).

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Modified GnRH-a long protocol in IVF

RESULTS

One hundred seventy-three women took part in this study. Eighty-two were included in group 1 and 91 in group 2. These women were between 21 and 42 years of age with no difference in mean age by group (Table 1). There was no significant difference in the distribution of causes of infertility between the two groups. Fertility and Sterility

Table 1

Comparison of Group 1 and Group 2 Parameters

Age (y) E2 baseline (pg/mL) E2 day ofhCG (pg/mL) LH day ofhCG (mIU/mL) P day ofhCG (ng/mL) Eggs retrieved Embryos replaced (IVF) HMG days

Group 1

Group 2

Statistical significance*

34.4 ± 4.28t

35.0 ± 4.09

NC:j:

33.8 ± 9.84

30.9 ± 11.29

NC

1,727 ± 1,288

1,549 ± 1,070

NC

6.1 ± 5.84

5.7 ± 4.79

NC

0.96 ± 0.62 13.2 ± 9.6

0.90 ± 0.50 12.5 ± 8.2

NC NC

3.3 ± 1.2 9.8 ± 1.7

3.3 ± 1.3 9.6 ± 1.9

NC NC

• Comparison between groups via the Mann-Whitney nonparametric test and independent samples t-test. t Values are means ± SD. NC, no change.

+

NUL*

Although initial baseline E z levels after suppressjon were higher in group 1, there was no statistical difference between the two groups (Table 1). Of course, the hMG was commenced only if E z was <50 pg/mL «0.18 nmol/L). Furthermore, there was no difference between the two groups of E z , LH, and P levels on the day of hCG administration (Table 1). It should be noted that there was no spontaneous LH surge in any of the patients in group 2 in which the GnRH-a was discontinued when hMG was commenced. No woman in either group experienced hyperstimulation syndrome. Furthermore, the number of eggs retrieved, the number of eggs or embryos replaced, and the number ofhMG stimulation days were not significantly different in the two groups studied. There was no difference by group in the method of treatment (Fig. 2). The number of patients in group 1 versus group 2 having GIFT performed (11 % versus 15.3%), the number of patients having ET after IVF (64% versus 62.6%), and the number of patients not having ET after IVF because of zero fertilization (24.2% versus 22%) are similar. Pregnancy rates in the GIFT procedure (Table 2) are similar between the two groups, although the numbers are too small for conclusions. Pregnancy rates per transfer in the IVF-ET procedure are higher in group 2 (35.2%) than in group 1 (19.4%) but only with borderline significance (P < 0.10; P = 0.07). Of these pregnancies, 16.7% in group 1 and 16% in group 2 aborted. The ongoing rate was similar in both groups, and there were two twins in group 1 and six twins and one triplet (with subsequent selective reduction to twin) in group 2. All other pregnancies were singletons. Vol. 61, No.4, April 1994

Figure 2 Method of treatment by group. Group 1 (Illll); group 2 (1!!lI). *P > 0.10 (not significant) between groups. NUL, No ET because of zero fertilization.

DISCUSSION

Currently, the GnRH-a are widely used in IVF programs for various reasons. Their use has been associated with prevention of premature endogenous LH surge, avoidance of premature luteinization and resulting follicular atresia, reduction of cancellation rates, and increase in multiple follicular growth with improved follicular maturation and, finally, increased PRs (1-3). Another reason for GnRH -a administration is to allow flexibility in hCG administration. In our study, we did not allow for flexibility of hCG administration to suit unit function so that we could keep all parameters constant for both groups without introducing more variables. It has been assumed that for continued pituitary

Table 2

Pregnancies per Method of Treatment per Group

GIFT Pregnancies Pregnancies/GIFT (%) IVF-ET Pregnancies Pregnancies/ET (%) Abortions

Group 1

Group 2

9 6 66 62 12 19.4 2 (16.7)*

14 8 57 71 25 35.2 4 (16)

Statistical significance*

NCt

P < 0.10 P

=

0.07

* Comparisons between groups via x2 test. t NC, no change. Values in parentheses are percents.

*

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suppression the GnRH -a has to be administered until the day of hCG injection. It is known, however, that in GnRH-a treated cycles there is a luteal phase defect (LPD) (8) with unacceptably low luteal phase P levels (10). Although GnRH-a has a short half-life, after its discontinuation the return of gonadotropin secretion to normal levels may take a few weeks (9, 10). Plasma levels of FSH return to levels that are found before the administration of GnRH-a earlier than LH levels do (9). The luteolytic effect of GnRH-a may be reversed after hCG administration (11). This indicates that the GnRH -a luteolytic effect is via a reduction of pituitary secretion of gonadotropins, namely LH. It is accepted that LH is luteotrophic (12), and its absence is a cause of corpus luteum disfunction. Because the suppression of endogenous gonadotropin release seems to continue for at least 12 days after the arrest of buserelin acetate (10), one would expect that halting GnRH -a administration at the time of hMG stimulation would have no adverse outcome on the cycle. The results of our prospective study do actually show no LH surge in the group of patients in whom GnRH -a was halted at the time of hMG administration. In fact, there seems no difference in mean LH, P, and E2 levels at the time of hCG administration in the two groups studied. The combination of events that in no patient was there an increased LH result on the day of hCG plus the fact that no luteinization occurred (as would have been expected if LH had risen much earlier) as seen by low P levels on the day of hCG plus the fact that no ovulation had occurred at oocyte retrieval 36 hours after hCG were enough for us to deduce that no LH surge had occurred. It seems that GnRH-a does not need to be continued until the day of hCG injection to maintain the pituitary suppression during the stimulation phase. Furthermore, the total number of oocytes retrieved, the fertilization rate, and the mean number of embryos (IVF -ET) or . eggs (GIFT) transferred seems to be the same for each group studied. The PR, however, is higher in group 2 patients than in group 1 patients, although with minor statistical significance. This could be related to two factors. The first concerns the lessened impact of GnRH -a on the luteal phase of group 2 patients as its administration has been halted at least 12 days before oocyte retrieval. The second concerns the local impact of GnRH-a on the human follicle. It has been shown that significant concentrations of intact bioactive buserelin acetate is present in follicular fluid at least 35 hours after the last nasal admin712

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istration (13). This could mean that GnRH-a might by itself have a direct intrafollicular effect. One effect of GnRH -a on the rat ovary is the inhibition of FSH action on granulosa cells (14). Of course, there is always the theoretical concern of teratogenetic risk when maturing oocytes are directly exposed to the GnRH -a analogue. In gonads of rats specific high-affinity receptors for GnRH have been found (15). The rat ovarian GnRH receptor has been demonstrated to exhibit similar characteristics to the pituitary GnRH receptor in terms of affinity and specificity (16). Furthermore, in humans the presence of low affinity receptors in the ovary has been reported (17). Considering the absence of adverse effects by the premature halting of GnRH -a administration but also the possible intrafollicular effects plus the LPD, one would expect that discontinuing GnRH-a early in the cycle can only have a beneficial effect on IVF-ET cycle outcome. Of course, the early cessation of GnRH -a administration also means a lessened cost for the patient. Several investigators (18) have reported a substantial increase in gonadotropin requirements in women suppressed with GnRH-a before the initiation of ovarian stimulation. One could expect that halting early GnRH-a could mean a reduction in the amount of hMG required for ovarian stimulation. This was not investigated in this report because we wanted to keep ovarian stimulation constant for both groups. However, we noted that the length of stimulation (as seen by the number of hMG days) required for ovarian stimulation was identical in both groups studied. Further studies are required to determine if the amount of hMG can be reduced in combination with early cessation of GnRH-a. This could further benefit the patients by reducing also the risk of hyperstimulation syndrome. It is still controversial whether GnRH-a is the most effective protocol for routine ovarian stimulation in IVF programs. Our study does show that there is still room for simplification in a cost-effective manner of existing GnRH-a protocols. Further studies are needed to confirm the absence of LH surge, with this modified GnRH-a long protocol and to determine if further simplifications can be made and for what group of patients their use would be most appropriate. REFERENCES 1. Abdalla HI, Ahuja KK, Leonard T, Morris NN, Honour JW, Jacobs HS. Comparative trial of luteinizing hormone-

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