Ovarian response in three consecutive in vitro fertilization cycles

FERTILITY AND STERILITY VOL. 77, NO. 4, APRIL 2002 Copyright 2002 American Society for Reproductive Medicine Published by Elsevier Science Inc. Printed on acid-free paper in U.S.A. Ovarian response in three consecutive in vitro fertilization cycles Fatemeh Hoveyda, M.B.B.S., M.R.C.O.G., a Lawrence Engmann, M.Bch.B., M.R.C.O.G., b Jo Steele, B.Sc., c Andres Lopez Bernal, M.D., c and David H. Barlow, F.R.C.O.G. c The Oxford Fertility Unit, John Radcliffe Hospital, University of Oxford, Oxford, United Kingdom Received May 30, 2001; revised and accepted September 27, 2001. Presented as a poster at the Annual Congress of British Fertility Society, Belfast, Northern Ireland, United Kingdom, April 24 27, 2001. Reprint requests: Fatemeh Hoveyda, M.R.C.O.G., Department of Obstetrics and Gynecology, Wycombe General Hospital, High Wycombe, Buckinghamshire, HB112TT, United Kingdom (FAX: 018 6576 9141, Professor Barlow, John Radcliffe Hospital, Level 3; E-mail: fhoveyda@doctors. org.uk). a Department of Obstetrics and Gynecology, Wycombe General Hospital, High Wycombe, Buckinghamshire, United Kingdom. b Department of Obstetrics and Gynecology, University of Connecticut Health Center School of Medicine, Farmington, Connecticut. c The Oxford Fertility Unit, John Radcliffe Hospital. 0015-0282/02/$22.00 PII S0015-0282(01)03237-X Objective: This study was designed to assess the ovarian response in the same patient in consecutive IVF cycles. Design: Retrospective study. Setting: Assisted reproductive unit at a university hospital. Patient(s): One hundred ninety women who underwent three consecutive cycles of IVF. Intervention(s): All women used a combination of pituitary desensitization and gonadotropin stimulation protocol and underwent oocyte retrieval. Main Outcome Measure(s): Number of follicles produced and number of oocytes retrieved. Result(s): There were no significant differences in the number of follicles produced, number of oocytes retrieved, and number of embryos created by the same woman among the three cycles of treatment. Conclusion(s): Consistent ovarian response can be achieved during the first three consecutive IVF cycles. (Fertil Steril 2002;77:706 10. 2002 by American Society for Reproductive Medicine.) Key Words: Ovarian response, predictive value, and consecutive cycles, in vitro fertilization In vitro fertilization (IVF) treatment is now widely used for the treatment of infertility, but there is constant debate regarding its effectiveness, availability, consequences, and the cost of treatment. The total number of IVF cycles undertaken in the world in 1995 was reported as 250,000 (1). The live birth rate after a single cycle of IVF is low and varies between 6% and 25% per cycle (1). Previous studies have confirmed that multiple cycles of treatment improve the overall chances of having a live birth after assisted conception treatment, and the cumulative probability of success is higher after several cycles of IVF treatment (2, 3). Therefore, most couples enter an IVF cycle with the intention of having more than one cycle. Unfortunately, IVF treatment may be associated with short-term complications, emotional stress (4), and potential long-term health consequences. Ovarian response to stimulation is an important determinant of success in IVF treatment. Hence, when a couple is facing the prospect of several IVF cycles of treatment it would be valuable in advising the couple if the response in the first treatment cycle was predictive of the response in subsequent cycles. It is unclear in the literature whether ovarian response deteriorates with subsequent cycles of ovarian stimulation. Although previous studies have assessed ovarian response after several cycles of ovarian stimulation (5, 6), most of these were performed during ovulation induction without the pituitary desensitization (6) or were not performed in consecutive cycles (5). The aim of our study was to compare the ovarian performance among three consecutive IVF cycles in the same woman to determine the consistency of the ovarian response in women who show responsiveness. MATERIALS AND METHODS Study Population Patients data were obtained retrospectively from the Oxford Fertility Unit databases. The local ethics committee accepts that the analysis of clinical data by clinicians for the purpose of 706

TABLE 1 Baseline characteristics of women undergoing three consecutive IVF cycles. Characteristic Cycle 1 Cycle 2 Cycle 3 P value Age (years) 33.9 (22.5 40.9) 34.6 (22.9 41.7) 35.5 (23.9 42.3).05 Baseline FSH level (miu/ml) 6.0 (1.3 14.0) 6.1 (1.3 14.5) 6.3 (1.0 13.5).05 Note: Values are medians with ranges in parentheses. improving the quality of the service does not require approval. Using this database, 265 women were identified who started three consecutive IVF cycles during the 4-year period between January 1995 and December 1998. These were couples who had never undergone IVF treatment previously. We excluded 75 women from the analysis because they did not undergo oocyte retrieval in all three consecutive IVF cycles in that time period. This study focuses primarily on the 190 women who underwent a total of 570 cycles. The indications for treatment were unexplained infertility (67, 35.2%), tubal disease (54, 28.4%), male factor (49, 25.7%), endometriosis (5, 2.6%), polycystic ovarian syndrome (2, 1.1%), multiple factors (11, 5.8%), and other (2, 1.1%). Treatment Protocol A long protocol of pituitary desensitization was used, starting with a GnRH agonist (nafarelin; Synarel, Searle, High Wycombe, UK) in the luteal phase at a dose of 400 g intranasal, twice daily (7). Controlled ovarian stimulation with urinary follicle-stimulating hormone (FSH) or urinary human menopausal gonadotropin (hmg) was commenced once pituitary desensitization was confirmed. The starting dose of FSH was one to eight ampules (75 600 IU of FSH activity), depending on the patient s age, previous response to ovarian stimulation, early follicular phase serum FSH level, history of endometriosis, and the presence or absence of polycystic ovaries on ultrasound assessment. Follicular growth was monitored with serial ultrasound scans and serum estradiol (E 2 ) levels, and the dose of gonadotropin was adjusted according to the follicular response. When the diameters of two or three leading follicles were at least 16 mm by ultrasound scan, hcg (Profasi; Serono, Welwyn Garden City, UK) in a dose of 5,000 IU or 10,000 IU was administered as a single injection. Transvaginal ultrasound guided oocyte retrieval was performed approximately 36 hours after hcg administration, and embryo transfer was performed 48 hours after oocyte retrieval. Progestogen pessaries (Cyclogest, Hoechst, Hounslow, UK) were given for luteal support starting on the day of embryo transfer, and were continued until a positive pregnancy was obtained. A clinical pregnancy was defined as a positive pregnancy test with fetal heart motion demonstrated on ultrasound scan at 6 weeks gestation. Outcome Measures The main outcome variables were the number of follicles produced and the number of oocytes retrieved. The other variables assessed were the total number of embryos, number of grade A embryos, estradiol level on the day of hcg administration, and the number of ampules of gonadotropin used and number of ampules of gonadotropin used per follicle aspirated. Embryos were graded as A, B, C, D, and E according to criteria used in the Oxford Fertility Unit. Grade A and B embryos were considered as good quality embryos. Grade A embryos had the appearance of regular blastomeres, with or without minor fragments. Grade B had irregular blastomeres, with or without minor fragments. Statistical Analysis The data were tabulated and analyzed using the Statistical Package for the Social Sciences (Release 6.0, SPSS Inc., Chicago, IL). Repeated measurements of each outcome variable were analyzed using the Friedman s test for nonparametric data. The results are presented as median and range. Values of P.05 were considered to be statistically significant. RESULTS A total of 190 women completed three consecutive IVF cycles, undergoing a total of 570 cycles. The baseline characteristics of these patients in the study are presented in Table 1. There was a significant increase in early follicular phase serum FSH level over the three cycles (P.05). The women were on average a year older by the time they commenced their third cycle (P.05). The body mass index (kg/m 2 ) was measured prior to the first cycle, with the median value being 22.5 with a range of 17.3 to 33.6. Characteristics of ovarian stimulation and response are presented in Tables 2 and 3, respectively. The starting dose of gonadotropin and total number of ampules used increased significantly over the three consecutive cycles (P.05), with eight more ampules of gonadotropin required in the third cycle compared to the first cycle. An increased number of ampules of gonadotropins were required per follicle retrieved in each successive cycle (P.05). There were no significant differences in the number of FERTILITY & STERILITY 707

TABLE 2 Characteristics of ovarian stimulation in three consecutive IVF cycles. Characteristics Cycle 1 Cycle 2 Cycle 3 Starting no. of ampules of hmg 3.0 (1 8) 3.0 (1.5 8.0) 3.5 (1.5 8.0) Total no. of ampules of hmg a 26.0 (12 108) 30.0 (12 96) 34.0 (12 120) Duration of ovarian stimulation (days) 9.0 (6 21) 9.0 (6 16) 10.0 (6 19) No. of ampules of hmg per follicle a 2.2 (6 23) 2.4 (5 14) 2.8 (4 50) Note: Values are medians with ranges in parentheses. P.05. follicles produced or the number of oocytes retrieved over the three cycles, with an average of 12 follicles and 8 oocytes produced per cycle. Further, the number of embryos produced did not change significantly over the three cycles. The proportion of embryos classified as grade A did not change significantly in successive cycles. There was an increase in the median number of embryos transferred in the third cycle. The clinical pregnancy rate was 3.1%, 6.8%, and 24.2% in the first, second, and third cycles, respectively. The live birth rate was 1.0%, 3.7%, and 19.5% in the first, second, and third cycles, respectively. There were no multiple births in the first two cycles but there were eight sets of twins in third cycle. To establish whether there was any influence of age on the ovarian response in three consecutive cycles, we stratified and analyzed the data according to two age groups, 35 years and 35 years at the start of the first cycle (Table 4). Thus, 112 women were aged 35 and 78 women were 35 years of age. The number of follicles and oocytes did not change significantly in the three consecutive cycles for women 35. However, there was an increase in the proportion of embryos that were grade A. The number of ampules of gonadotropin required per follicle in this age group also increased significantly over three consecutive cycles. For those women 35 years of age, the number of follicles produced, total number of embryos created, and proportion of embryos which were grade A over consecutive cycles did not change significantly. However, the number of ampules of gonadotropins required per follicle increased significantly over three consecutive cycles. We noted that to achieve consistent ovarian response in women 35 years of age, there was a progressive increase from a higher baseline in the total number of gonadotropins used as well as the number of ampoules of gonadotropins required per follicle. Thus, the total number of ampules of gonadotropin increased from 24 to 32 in the first and third cycles for women 35 years as compared to an increase from 27 to 44 in the first and third cycles for women 35 years. DISCUSSION It is a conundrum that a study aiming at examining IVF response and embryo yield in consecutive IVF treatments must restrict its observation to couples who actually progress to those end points. Defining that couples would have three cycles of IVF within 4 years will exclude couples achieving success to treatment, because these couples will be unlikely to have three cycles within this time period. Conversely, women who show such poor ovarian response that oocyte TABLE 3 Characteristics of ovarian response in three consecutive IVF cycles. Characteristics Cycle 1 Cycle 2 Cycle 3 No. of follicles 12 (3 34) 12 (3 33) 12 (1 33) No. of oocytes 8 (1 25) 8 (0 23) 8 (1 26) No. of embryos produced 4 (0 16) 5 (0 15) 5 (0 15) No. of embryos transferred a 2(0 3) 2 (0 3) 3 (0 3) No. of grade A embryos produced 1 (0 12) 2 (0 9) 2 (0 10) Proportion of embryos of grade A quality (%) 38 (0 100) 40 (0 100) 42 (0 100) Serum E 2 on day of hcg (pg/ml) a 1050 (205 3176) 1103 (177 4890) 1236 (219 3901) Note: Values are medians with ranges in parentheses, unless otherwise stated. a P.05. 708 Hoveyda et al. Ovarian response in three consecutive IVF cycles Vol. 77, No. 4, April 2002

TABLE 4 Characteristics of ovarian stimulation in three consecutive IVF cycles in women 35 and 35 years of age. Characteristics Cycle 1 Cycle 2 Cycle 3 Cycle 1 Cycle 2 Cycle 3 Total no. of ampules of hmg a 24 (12 108) 27 (12 84) 32 (14 120) 27 (12 96) 34 (12 96) 44 (12 114) No. of ampules/follicles a 1.9 (0.6 21.0) 2.1 (0.5 13.2) 2.5 (0.6 20.0) 2.7 (0.6 23.2) 3.3 (90.6 13.7) 4.0 (0.4 50.0) No. of follicles 13 (3 34) 12 (3 33) 13 (5 29) 11 (4 31) 12 (3 21) 12 (1 33) No. of oocytes 8 (1 25) 9 (2 23) 9 (2 24) 8 (1 20) 7 (0 19) 8 (1 26) No. of embryos produced 4.0 a (0 16) 5.0 a (0 15) 5.5 a (0 15) 4.0 (0 14) 5.0 (0 11) 4.0 (0 13) No. of grade A embryos produced 1 (0 8) 2 (0 8) 2 (0 10) 1 (0 12) 1 (0 9) 1 (0 8) Proportion of embryos of grade A quality (%) 40.0 (0 100) 40.0 (0 100) 45.0 (0 100) 37.5 (0 100) 33.3 (0 100) 33.3 (0 100) Note: Values are medians with ranges in parentheses, unless otherwise stated. a P.05. retrieval does not consistently occur will not contribute to the defined end points. We are concerned with the couples who are showing sufficient response as to achieve oocyte retrieval, and wish to address what advice they should be given regarding how they might expect to perform in repeated cycles. By taking all 265 couples who commenced three consecutive IVF cycles over a 4-year period we found that 81% (215 out of 265) did achieve oocyte retrieval in their first cycle and that 88% of these (190 women) maintained this performance over the three cycles. What was the consistency of response achieved for this group? The results of our study have confirmed that follicular recruitment and oocyte and embryo production during the first three consecutive cycles can be consistent. The data indicate that it was possible to achieve a consistent ovarian response in these women over a 4-year study period during which stimulation protocols, methods of oocyte retrieval, and laboratory procedures remained unchanged. This observation is based on a relatively large group of women, all of whom underwent their first three consecutive IVF cycles and reached oocyte retrieval on each of these cycles. Clinicians should feel confident in recommending at least three cycles of IVF treatment, as the ovarian response is unlikely to deteriorate during subsequent cycles of treatment although the dosage of gonadotropin may need to be adjusted. Although previous studies have been performed to assess ovarian response in subsequent cycles of ovarian stimulation, none have followed the same women through three consecutive cycles of IVF. The findings of Ebbiary et al (6) and Diamond et al (8) are consistent with those of our study, but these studies were performed with women undergoing ovulation induction or intrauterine insemination and not with women undergoing controlled ovarian stimulation with the use of GnRH agonists in IVF cycles. Yovel et al. (5) assessed women who had failed to conceive after three consecutive embryo transfer cycles and who underwent from four to eight IVF cycles. They showed that there was no effect of cycle number on the number of oocytes retrieved and total number of embryos produced. However, their study was not performed on consecutive cycles and the same women were not followed through all the four to eight cycles. Their treatment protocol and that of Ron-El et al. (9) also consisted of a heterogeneous protocol with or without the use of GnRH agonist treatment. Further, the use of independent analysis rather than repeated measures in the statistical analysis makes no allowance for any missing observations, so the data from different cycles may not relate to exactly the same group of women. We believe that the findings of our study are reliable in view of the more robust design of the study. We assessed women who had three consecutive cycles and used similar protocols in subsequent cycles. In view of the nature of the repeated measures we used in the statistical analysis, the women also served as their own controls in subsequent cycles. The context of the consistency of ovarian response observed was one of significant increase in the total number of ampules used for ovarian stimulation in subsequent cycles. Other studies have shown that increasing the dose of gonadotropin in subsequent cycles does not improve the outcome of the cycle (10, 11). Again, these studies did not assess the same group of women over several consecutive IVF cycles. A future prospective study would need to be done to establish whether the ovarian response is maintained in the face of a constant dose of gonadotropin in subsequent cycles. The median number of embryos transferred remained at two for the first two cycles, but rose to three in the third cycle. The couples initially complied with our voluntary two-embryo transfer policy, but following two failed attempts to become pregnant, many couples opted for a threeembryo transfer in the subsequent cycle. However, this practice is questionable and should be discouraged, as previous studies have shown that the live birth rate during first three IVF attempts is constant (12, 13). FERTILITY & STERILITY 709

The live birth rate per cycle in this study does not reflect the actual live birth rate per cycle for all women in our unit. This is because few women who achieved a pregnancy in their first or second cycle would have returned to complete three cycles of stimulation within the 4 years studied. The data from our study may be useful in counseling patients and planning their future treatment regarding the likelihood of successful ovarian stimulation in future cycles if adequate stimulation was achieved during the initial cycle. This also would enable us to modulate the number of follicles to a certain extent, knowing the individual ovarian response from the previous treatment cycle. The results of this study may also be useful for counseling women who undergo multiple cycle packages in certain units (3). References 1. De Mouzon J, Lancaster P. World collaborative report on in vitro fertilization. Preliminary data from 1995. J Assist Reprod Genet 1997; 5(Suppl):251S 65S. 2. Hull MGR, Eddowes H, Fahy U. Expectations of assisted conception for infertility. BMJ 1992;304:1465 9. 3. Engmann L, Maconochie N, Bekir J, Jacobs H, Tan S. Cumulative probability of clinical pregnancy and live birth after a multiple cycle IVF package: a more realistic assessment of overall and age specific success rates? Br J Obstet Gynaecol 1999;106:165 70. 4. Seibel M. Infertility: the impact of stress, the benefit of counseling. J Assist Reprod Genet 1997;14:181 3. 5. Yovel I, Geva E, Lessing J, Yaron Y, Botchan A, Amit A. Analysis of the fourth to eight in-vitro fertilization treatments after three previously failed attempts. Hum Reprod 1994;9:738 41. 6. Ebbiary N, Morgan C, Martin K, Afnan M, Newton JR. Ovarian response in consecutive cycles of ovarian stimulation in normally ovulating women. Hum Reprod 1995;10:536 43. 7. Lockwood G, Pinkerton S, Barlow D. A prospective randomized single blind comparative trial of nafarelin acetate with buserelin in longprotocol gonadotrophin-releasing hormone analogue controlled in-vitro fertilization cycles. Hum Reprod 1995;10:293 8. 8. Diamond M, De Cherney A, Baretto P, Lunenfeld B. Multiple consecutive cycles of ovulation induction with human menopausal gonadotropins. Gynecol Endocrinol 1989;3:237 40. 9. Ron-El R, Herman A, Golan A, Nahum H, Soffer Y, Caspi E. Ovarian response in repetitive cycles induced by menotrophin alone or combined with gonadotrophin releasing hormone analogue. Hum Reprod 1990;5:427 30. 10. Stadmauer L, Diktoff EC, Session D, Kelly A. High dosages of gonadotropins are associated with poor pregnancy outcomes after in vitro fertilization-embryo transfer. Fertil Steril 1994;61:1058 64. 11. Land JA, Yarmolinskaya MI, Dumoulin JCM, Evers JL. High-dose human menopausal gonadotropin stimulation in poor responders does not improve in vitro fertilization outcome. Fertil Steril 1996;65:961 5. 12. Croucher C, Lass A, Margara R, Winston R. Predictive value of the results of a first in-vitro fertilization cycle on the outcome of subsequent cycles. Hum Reprod 1999;14:2417 8. 13. Meldrum D, Silverberg K, Bustillo M, Stokes L. Success rate with repeated cycles of in vitro fertilization embryo transfer. Fertil Steril 1998;69:1005 9. 710 Hoveyda et al. Ovarian response in three consecutive IVF cycles Vol. 77, No. 4, April 2002