FERTILITY AND STERILITY Copyright 1992 The American Fertility Society Printed on acid-free paper in U.S.A. Prospective study of short and ultrashort regimens of gonadotropinreleasing hormone agonist in an in vitro fertilization program Umesh Acharya, M.R.C.O.G.* Stewart Irvine, M.D. Mark Hamilton, M.D. Allan Templeton, M.D. Assisted Reproduction Unit, Department of Obstetrics and Gynaecology, University of Aberdeen, Aberdeen Maternity Hospital, Aberdeen, Scotland. Objective: To assess the usefulness of the ultrashort regimen of gonadotropin-releasing hormone agonist (GnRH-a) in ovulation induction in an in vitro fertilization (IVF) program. Design: A prospective randomized trial comparing short and ultrashort regimens of GnRH-a. Setting: Aberdeen Assisted Reproduction Unit. Patients: Forty-eight patients having IVF for the first time were randomized between the two protocols. Main Outcome Measures: Response to ovarian stimulation and occurrence of spontaneous luteinizing hormone (LH) surges. Results: In ovulation induction, fertilization, and pregnancy rates the ultrashort regimen produces results that were no different to the short regimen but it did not always prevent an LH surge. Conclusion: The ultrashort regimen can be a useful alternative for ovarian stimulation of patients undergoing IVF. Fertil Steril1992;58:1169-73 Key Words: In vitro fertilization, gonadotropin-releasing hormone agonist, ovarian stimulation, short and ultrashort regimen Induction of multiple ovulation followed by in vitro fertilization (IVF) and embryo transfer has become a widely accepted treatment of infertility. Initially' the anti-estrogen clomiphene citrate (CC) was used, followed by the combination of CC and human menopausal gonadotropin (hmg) for ovulation induction. Early attempts at stimulated IVF cycles were hampered by the occurrence of a premature luteinizing hormone (LH) surge and the inability to undertake oocyte recovery at convenient times. This problem has been partially overcome with the introduction of gonadotropin-releasing hormone agonists (GnRH-a). Initially GnRH-a was com- Received March 12, 1992; revised and accepted August 18, 1992. * Reprint requests: Umesh Acharya, M.R.C.O.G., Assisted Reproduction Unit, Department of Obstetrics and Gynaecology, University of Aberdeen, Aberdeen Maternity Hospital, Cornhill Road, Aberdeen AB9 2ZD, Scotland. menced in the luteal phase of the pretreatment cycle (1, 2) or during menstruation (3-5) and was continued until complete suppression of pituitary gonadotropin was achieved. This method results in down regulation in the space of 2 weeks and has been referred to as long course treatment. This was followed by the so-called short course treatment of GnRH -a whereby GnRH -a was given in the early follicular phase together with ovarian stimulation. The latter form of treatment achieves comparable results to the long course and is better tolerated by patients (2, 6, 7). More recently there have been reports of the ultrashort protocol in which the GnRH -a therapy is administered for 3 to 4 days only (8-10). The present study is the first prospective randomized study to compare the use of short and ultrashort protocols of GnRH -a therapy in achieving superovulation for purposes of IVF. This study Acharya et al. Short and ultrashort regimens in IVF 1169
aimed to examine whether the results obtained using the ultrashort protocol were similar to those obtained using the short protocol and to review any deleterious effects that the ultrashort protocol may have had, particularly the occurrence of a premature LH surge was monitored. Patients MATERIALS AND METHODS Patients having their first IVF treatment were recruited from the Aberdeen IVF program. Table 1 shows the characteristics of patients undergoing treatment. Forty-eight patients were recruited for the study and randomly allocated to one or other of the following protocols. Randomization was undertaken using sealed envelopes. There were 24 patients in each group. In group 1, (short protocol) norethisterone (Primolut N; Schering Pharmaceuticals, Burgess Hill, United Kingdom) 5 mg two times per day was commenced on day 21 of the cycle for 7 to 14 days. The use of norethisterone in the preceding cycle to delay menstruation has been previously described (11, 12). The use of norethisterone has been shown not to affect the outcome of IVF treatment (13). After the completion of norethisterone, a basal scan was performed and if the serum estradiol (E2) level was <70 pmoljl «19 pgjml), buserelin acetate (Suprefact; Hoechst UK Ltd., Middlesex, United Kingdom) 500 J.lgjSC was commenced the following day (day 1). This was continued until the day of human chorionic gonadotropin (hcg) administration. Superovulation was commenced initially at 300 IU of hmg (Pergonal; Serono Laboratories UK Ltd., Welwyn Garden City, United Kingdom) on day 2 for 3 days, followed by 225 IU on the fourth day and 150 IUjd for the rest of the cycle (till the day of hcg administration), the dose being adjusted according to ovarian response. Serial scans were performed using a Diasonics 250 ultrasound (US) machine (Diasonics Sonotron, Bedford, United Kingdom) equipped with a 7.5-MHz vaginal probe. Ten thousand international units of hcg was administered when three follicles were> 16 mm diameter and E2 > 2,000 pmoljl (545 pgjml). Vaginal oocyte recovery was carried out 34 hours later under US guidance. Embryo replacement was undertaken 48 hours after the oocyte recovery. Sperm were prepared by the swim-up technique and the motile sperm were used for insemination. In group 2 (ultrashort protocol) the procedure was similar except that buserelin acetate was adminis- Table 1 Patient Characteristics in Two Treatment Groups Ultrashort Short protocol protocol (n = 24) (n = 24) Female age (y) 34 (26 to 43)* 32 (26 to 43)* Duration of infertility (y) 5.0 (2-11)* 4.8 (1-20)' Etiology of infertility Tubal 10 (41. 7)t 13 (54.2) Unexplained 8 (33.3) 4 (16.7) Male 3 (12.5) 5 (20.8) Endometriosis 3 (12.5) 2 (8.3) Outcome Menstruation 17 (70.8)t 13 (54.2) Pregnancies 2 (8.3) 5 (20.8) Failed fertilization 3 (12.5) 1 (4.2) Poor stimulation 1 (4.2) 1 (4.2) Overstimulated 1 (4.2) 3 (12.5) Other 0 1 (4.2), Values are medians with ranges in parentheses. t Values in parentheses are percents. tered for only 3 days (days 1 to 3). In both groups blood was taken on the 5th, 10th, and 15th day after hcg administration (hcg +5, hcg + 10, and hcg + 15) for progesterone (P) and E2 levels to monitor the luteal phase. Luteal support was given by administering 2,500 IU of hcg on the day of embryo replacement. Assays Serum E2 and P levels were measured by radioimmunoassay (RIA, Estradiol Coat-A-Count; Diagnostic Products Corporation Ltd., Oxon, United Kingdom and P using Amerlex-M Kit; Amerlite Diagnostics Ltd., Amersham, United Kingdom), the details of which have been previously reported (14). Serum LH was also measured by an RIA (LH MAlA clone; Serono Diagnostics, Fleet, Hampshire, United Kingdom). The LH intra-assay coefficient of variation (CV) was 4.4% and the interassay CV was 6.3%. Statistics No assumptions were made regarding the distribution of the data and consequently a nonparametric method (Mann-Whitmey U-test) was used for comparison between groups. Analysis of variance was used to compare outcome measures between the two groups. RESULTS Both groups were similar with respect to age, duration of infertility, and indication for IVF as can 1170 Acharya et al. Short and ultrashort regimens in IVF Fertility and Sterility
be seen from Table 1. Analysis of variance showed that the distribution of etiology of infertility in these two groups was not significantly different and the percentage of patients with primary infertility (83.3%, short; 75%, ultrashort) and secondary infertility (16.7%, short; 25%, ultrashort) was similar in both groups. No difference was found in the stimulation requirements (Table 2) or in the response to stimulation as measured by E2 values (Fig. 1) in the two groups. There was no difference in the number of follicles aspirated or number of oocytes obtained per patient; however, a higher number of oocytes cleaved per patient in the ultrashort group but this did not reach significance. This did not appear to be related to the semen quality as there was no difference in the sperm concentration or grade III and grade IV motility of sperm used for insemination. The median sperm concentrations were 9 X 10 6 /ml and 5 X 10 6 1 ml in the first and second groups respectively, the median grade III motilities were 77.5% and 76.5%, respectively, and grade IV motilities were 10% and 8.5%, respectively. However, when the embryos did cleave, the grading distribution was the same in both groups, as was the number of patients having three embryos transferred (we follow the Human Fertilization and Embryology Authority's guidelines in the "Code of Practice" of transferring no more than three embryos in each cycle). The embryos were graded on a scale of 0 to 10 according to recommended embryological criteria (15, 16). Table 2 Stimulation, Ovarian, and Embryologic Details in Two Groups Duration of hmg administration (d) Amount of hmg (ampules 75 IU FSH; 75 IV LH) No. of days from basal scan to oocyte recovery No. of follicles aspirated per patient No. of oocytes obtained per patient No. of oocytes cleaved per patient Grade of embryo transferred 4 6 8 10 No. of patients having three embryos transferred Short protocol 11 (9-16)" 28 (17-72) 14 (12-18) 9 (1-26) 7 (1-17) 3.5 (0-13) 1 (2.2)t 6 (13.0) 22 (47.8) 17 (37.0) 13 (54.2) * Values are medians with ranges in parentheses. t Values in parentheses are percents. Ultrashort protocol 11 (7-18) 28 (14-80) 15 (12-18) 12 (1-24) 8 (1-20) 6 (0-15) 1 (2.0) 5 (9.8) 21 (41.2) 24 (47.1) 15 (62.5) 1300 1100 900 S 0 700 E Q. N W 500 300 100 100-18 -15-12 -9-6 -3 a Days from oocyte recovery Figure 1 Graph of E2 values with days from oocyte recovery in ultrashort (- 0 - median and 95% confidence intervals; CI) and short regimens (-+- median and 95% Cn. There were five pregnancies in the ultrashort group and two in the short group. Clearly this result did not reach statistical significance and could not be expected to do so in a study of this size. A much larger study would be required to achieve this. The pregnancy rate per embryo transfer was 27.8% (51 18) in the ultrashort group and 10.5% (2/19) in the short group. With regard to the pregnancy outcomes, of the 5 in the ultrashort group, 4 resulted in live births (2 singletons and 2 sets of twins) and 1 resulted in a missed abortion. In the short group both pregnancies resulted in live singleton births. A premature LH surge, resulting in cancellation of the cycle, occurred once in the ultrashort group but did not occur in the short group. The LH values on the day of hcg administration ranged from 0.5 to 1.6 lull. In the patient with the LH surge, the surge occurred on day 15. The LH values before this day did not exceed 3.0 lull. The LH values on day 14, 15, and 16 were 6.3, 21.2, and 8.9 lull, respectively. The corresponding P values were 1.3, 2.2, and 5.5 nmol/l (0.41, 0.69, and 1.73 ng/ml), and the E2 values were 2,970, 4,790, and 2,750 pmoljl (809.0, 1,304.8, and 749.1 pg/ml), respectively, demonstrating that this was a true LH surge. This patient had 10 days of pretreatment with norethisterone. This cycle was cancelled because at the onset of the LH surge the three largest follicles had a mean diameter of 15.3 mm (16.3, 14.7, and 14.5 mm) only. It can be seen that although this patient's response was slower than the median response, it would be within the range for that particular group of patients. There were no biochemical or US evidence of pre- Acharya et al. Short and ultrashort regimens in IVF 1171
mature luteinization in any of the other patients in the study. The number of patients with overstimulation was the same in both groups. There was no difference in the luteal phase E2 levels. On hcg +5, E2 was 2,300 pmol/l (626.5 pg/ml) in the short group and 1,350 pmol/l (367.8 pg/ml) in the ultrashort group. Similarly on hcg +10, E2 values were 4,500 pmol/l (1,225.8 pg/ml) and 4,200 pmol/l (1,144.1 pg/ml), and on hcg + 15, E2 values were 160 pmol/l (43.6 pg/ml) and 350 pmol/l (95.4 pg/ml). Progesterone values 5 and 15 days after hcg administration were not significantly different, i.e., 127 nmol/l (39.9 ng/ml) and 117.6 nmol/l (37.0 ng/ml) on hcg +5, in the ultrashort and short protocols, respectively, and 6.5 nmol/l (2.0 ng/ml) and 5.5 nmol/l (1.7 ng/ml), respectively, on hcg + 15. In the midluteal phase (hcg + 10) the P value was significantly raised (P < 0.05) in the ultrashort group, 274.3 nmol/l (86.2 ng/ml), compared with the short group, 127.0 nmol/l (39.9 ng/ml). DISCUSSION This study compares the use of the short and ultrashort protocols of GnRH -a administration in the context of IVF ovulation induction regimens. Although the long and short protocols have been shown to be useful (2,4,6,7,17), this study confirms that treatment using the ultrashort protocol is a satisfactory alternative. The follicular response and the number of oocytes obtained were very similar for both regimens. The cleavage and the pregnancy rate were slightly higher in the ultrashort group, but this did not reach statistical significance (a much larger study would be required to address this question). Similarly, there was no difference in the incidence of ovarian overstimulation or the number of patients reaching embryo replacement. It would appear that the ultrashort regimen is more convenient for the patient, GnRH-a being injected for only 3 days compared with a median of 12 days for the short protocol. This shorter duration of agonist administration would also have implications for the cost of and compliance with treatment. There was also a higher pregnancy rate with the ultrashort protocol but this did not reach significance in this small study. However the ultrashort regimen did not totally prevent the occurrence of a spontaneous LH surge. Smitz et al. (10) using the agonist (500 Ilg/d SC, day 3 to 5) in 10 endocrino- 1172 Acharya et al. Short and ultrashort regimens in IVF logically normal patients also found that it did not always prevent an LH surge. Howles et al. (8) studied seven patients who had responded poorly to CC and hmg previously and had high tonic LH levels. In their small study they found that administering GnRH-a (500 Ilg/d SC) on days 1 to 3 of menstrual cycle significantly reduced the urinary LH output in the latter stages of follicular development. Martikainen et al. (9) administered GnRH-a (200 Ilg IN three times per day) and found that it prevented the LH surge although in their study it was administered for 4 days (days 1 to 4). In our study, we observed that the midluteal P was significantly raised in the ultrashort group, possibly because of the fact that more follicles were aspirated in this group (12 versus 9). This study is the first prospective randomized controlled trial of short versus ultrashort GnRH-a regimens. The efficacy of the ultrashort regimen is confirmed. It is simpler to use and is more convenient for the patient, requiring fewer subcutaneous injections. The shorter duration of GnRH-a administration makes it more cost effective and no significant compromise is apparent in terms of IVF, outcome, embryo quality, and pregnancy rates although premature luteinization is not always prevented by this regimen. 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