Superovulation strategies in assisted6 conception

Michael C. Macnamee and Peter R. bnnsden

I&'!K)BUGTQN.
The firsc- ;;.'- vitro fertilization ('B(SB)* pjegnancy-" was ecropic and the first child, Louise Brown, was conceived in a'natural' IVF cycle'. This birth in 1978 not only heralded an upsurge of interest in the treatment of human infertility, bur ushered in a period of particular interest in ovulation induction. The aim of'superovulationfbr assisted conception treatment techniques" is -ro induce. women to produce a larger number ofoocyces than the one or wo that are required for normal ovula-tlon-induction programs. Manv changes have occurred in the development of gonadotropins and in IVF superovulation protocols since 1978. These changes range from the use of clomiphene alone, through clomiphene and human menopausal gonadotropin (hMG, luteinizing hormone (LH), 75 IU and follicle stimulating hormone (FSH), 75 1U), ro the present use of highly purified urine-derived FSH (HPFSH). We now also have the latest and most sophisticated of the gonadotropins so far developed - recombinant human FSH (r-FSH), which is now the only preparation in use at Bourn Hall Clinic. The treatment options available to infertile couples have also expanded during this time to include simple ovulation induction, intrauterihe insemination (IUI) as well' as IVF, which remains the principal treatment for the most severe intenility problems. Many units are now also able to offer intracyroplasmic sperm injection (ICSI), ovum, sperm and embryo donation and IVF surrogacy. The most important factors governing the chances of successful IVF treatment are still: (1) Female age; (2) Response to superovulation; (3') t'tumber-andtquality-of-oocyres collected; (4) Quality of semen; (5) Number and quality of embryos transferred. Our aim with superovulation for IVF has always been ro produce a sufficient number of very high-quality embtyos so-as to enable two or. three t0"b"e transferred, and leave those remaining to be frozen for possible later use. In this way, the need for further stimulation cycles is reduced and the chances of success for each stimulation cycle is maximized. Clearly, if patients respond well to superovulation and produce high-quality oocytes, the chances of success are much greater than if they do not. In the late 1960s, oocyres maturing in the human ovary were identified by the presence of chromosomes in diakinesis of the first meioiic division, and the first demonstration that human eggs could be fertilized in vitro was made2. Subsequently it was shown that oocyte maturation and ovulation could be timed precisely after an injection of human chorionic gonadotropin (hCG) or by observing the onset of the endogenous LH surge in both gonadotropin-srimulated and 'natural' menstrual cycles3. Although far in advance of work in animal species, it was a full decade before this knowledge was successfully applied clinically, and the first live birth' following in. vitro fertilizat.ionnandembryo transfer (IVFET) was achieved. Louise Brown was the product of an embryo created following fertilization of a mature oocyte collected by timing the endogenous LH surge in a natural menstrual cycle. Her birth heralded the genesis of a new treatment,, for inferrility'and brought hope to countless thousands of couples. Following the founding of Bourn Hall Clinic

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In Vitro Fertilization and Assisted Reproduction in-1980, Stcptoe and Edwards continued to develop the techniques of IVF, which included using gonadotropins to stimulate follicular development4. This required the rigorous application of endocrine monitoring and the adoption of the then newly developed techniques of ultrasound ovarian imaging, which have led to major advances in the clinical management of supe avulation cycles. The early use of superc ilation cycles for IVF established that success was principally determined by the number of embryos replaced. This key finding, backed by mathematical models, demonstrated that the replacement of three or more healthy embryos optimized; the likelihood of achieving an ongoing pregnancy5"8. It was also found that replacement of more than three embryos increased the likelihood of higher-ore; ;r multiple pregnancies, while not increasing the pregnancy rate5'8. Given a reasonable sperm preparation, four or five good-quality oocyies must be recovered to be reasonably sure ofth? replacement of three morphologically normal embryos Thus, natural cycle IVF has largely been abandoned in favor of superovulation cycles for assisted conception, although there was a resurgence of interest in natural cycle IVF9 in the early 1990s, and it may still be of value in a limited number of patients. The purpose of this chapter is to review the history and theoretical back^ 'ind to the follicular stimulation protocols currently in use at Bourn Hall; we have not tried to give an exhaustive review of all the many different superovulation strategies employed in clinics throughout the world. Our review is supplemented by practical' notes in the form of information 'boxes' on each of the major regimes used. Information in this chapter may be enhanced by reading the chapters on LH releasing hormone (LHRH) agonists and antagonists and on the new gonadotropins elsewhere in this book.

this regimen is that the escrogen-driven negative feed back on endogenous gonadotropin secretion is defeated Acting at the hypothalamic and pituitary levels, CC pro vokes a mild hypersecretion of pituitary gonadotropins If administered shortly after menstruation begins, CC will stimulate the growth of a number of small follicles The administration of exogenous gonadotropins wil then sustain the growth of this cohort of recruited folli cles. Thus, synchronous multiple follicle developmen can be induced in an otherwise monoovular species. The best success rates using this combination wer achieved when+extensive monitoring of endogenous Lr-secretion in, the late follicular phase was employed. I undetected, the endogenous LH surge, although severer attenuated10, can lead to ovularion prior to surgica oocyte recovery, or to the collection of postmatur oocytes with a reduced capacity to fertilize. The onset o the LH surge is not predictable by observation of follici size alone or the absolute level of estrogen or the rate c estrogen rise10. However, if detected and supported b the administration ofhCG, and if the onset of the surg is used to time oocyte recovery, the patient's clinics performance through an assisted conception cycle i not compromised. An increasing proportion of patient will surge if ovulation is induced in the later stages c follicular growth10. The early results with CC/hMG stimulation demon strated its efficacy, achieving a clinical pregnancy rate c 35% following the transfer of three embryos. Irrespectiv of the number of embryos transferred, overall clinic; pregnancy rates per transfer of 25% have been reporte from several centers6"8. These figures equate with 'take-home baby rate' of around 15%. Up to the time c publication of the first edition of this book in 1992, th majority of the estimated 10 000 babies born world-wid following IVF were the result of CC/hMG treacmen more than 1200 of these were from Bourn Hall alone. The extensive endocrine and ultrasound monitorin of IVF patients stimulated with CC/hMG has allowe detailed analysis of possible endocrine indicators < reproductive success that occur in the late follicul; phase10. No difference was found between those patien who became pregnant after embryo transfer and tho" whose embryos failed to implant in terms of the fin: estrogen level, the diameter of the four largest follicles; the time ofhCG administration, the number ofoocyti

CLOMIPHENE CITRATE AND GONADOTROPINS IN SUPEROVULATION

Until the late 1980s, the most common ovarian stimulation regimen used in assisted conception was the combination of clomiphene citrate (CC), a partial agonist of estrogen and exogenous gonadotropin preparations in the form ofhMG (see Box 1 for details). The rationale of

92

0.30

*/><0.01

Superovulation strategies in assisted conception Non-pregnant 0.40-r (rt=100) Discharged prior to oocyte retrieval due to poor response (n = 50) Pregnant (n=100) Failed fertilization due to oocyte problem (n = 25)

oS

51
b^: 33 0.-16 0.20

Days-prior to oi'ul>ftion"in<i<fe(ion5; Figure 1 lyofilcs- ofc urinary, LH;. outgut* in. [regnant- and-nonpregnant Wf patients10* collected, their fertilization and cleavage rates, or the average number of embryos transferred per patient. However, patients who established, a clinical pregnancy had a significantly lower urinary LH output in thc*2 days prior to the induction ofovulation (Figure 1). A similar pattern was demonstrated for plasma levels of LH in the late follicular phase. More detailed analysis has shown [hat a link exists between oocyte quality and the mean tonic urinary LH output in the late follicular phase. Failure of fertilization solely attributable to poor oocyte quality (assessed by light microscopy) correlates with higher late follicular phase LH levels than those solely attributable to sperm factors (Table 1). Further evidence of a detrimental effect of high LH on ovarian function comes from the finding that patients discharged prior to oocyte recovery because of inadequate follicular development had high tonic levels of LH production. Principal among these findings is the fact that high follicular phase levels ofLH are incompatible with implantation and are predictive of early pregnancy loss following ovularion Tible 1 Mean ( SEM) late follicular phase luteinizing hormone (LH) levels (1U/1 per hour) and ascribed reason for failure of fertilization10 Ascribtd cause Poor oocyte quality Poor sperm quality Other non-oocyte problems Unknown /< 0,01; ^< 0.01 n 25 25 10 25 Urinary LH 0.3110.04^ 0.20 0.03' 0.22 0.03 0.2\0.0^

0.

^. 0.30 &-. SS

At'orted%l'Z:weeks'gestation,(n=30) Failed implantation .following-embryo replacement (n = 95)

ST

0.20 CHI 0.16-L Blocks represent mean SEM Figure 2 Outcome of IVF treatment in relation to late follicular phaseoutput of luteinizing hormone. The blocks represent mean SEM induction, IVF11'12 and natural conception13. Our data on urinary LH output and success following IVF-ET are summarized in Figure 2. Only in cases where mean late follicular phase urinary LH levels were above 0.3 IU/1 per hour was a decrease in oocyte quality obvious by routine light microscopy. The less discernible consequences of high late follicular phase LH levels may be explained by consideration of the control mechanism of oocyte maturation and ovarian paracrinology. Broadly, there are two schools of thought explaining the effect of high LH levels. One maintains that the natural mid-cycle LH surge promotes the degradation of the gap-junctional communication system in the granulosa cell layers, which maintains meiotic arrest by allowing the passage of oocyte maturation inhibitor(s) from the follicle wall to the oocyte14. The other view is that the LH surge stimulates the expression of maturation-inducing factors by the granulosa cells 15. It is possible that both these systems may work together 16. Recent evidence also suggests a close relationship between ovarian growth factors, their binding proteins and circulating levels of the gonadotropins17. The role of these growth factors and their control mechanism(s) within the ovary are yet to be fully understood. The pathophysiological basis of elevated late follicular phase LH levels may rest partly with the individual but Pregnant (n= 80)

93

In Vitro Fertilization and Assisted Reproduction most likely with the individual's response to CC administration. The degree of gonadotropin hypersecretion induced with clomiphene varies between cycles within the same individual and between individuals. The hyper-secrction of endogenous gonadotropin may be sustained into the late follicular phase in some individuals, to the detriment of their success. Pathological ek ations in circulating LH, such as are found in son.e patients with polycystic ovarian syr drome, may also contribute to their infertility, and to du ir likelihood of early spontaneous abortion13.

retrospective analyses and concern specific groups of. patients and are therefore temporally disconnected and non-randomized. However, two large, well-controlled studies show a doubling of the 'take-home baby rate' on both the long and the ultra-short-^protocols when compared with CC/hMG stimulation18'19. This increase in success is due to improved oocyte quality, achieved by the reduction of LH levels and greater consistency of"* oocyte quality. In conclusion, the use of LHRH analogs, in combination with exogenous gonadotropins, significantly increases the chance of pregnancy after embryo replacement when compared to treatment with CC/hMG. The physiological basis for this incremental success is attributable to the suppression of endogenous LH in the late follicular phase. The result is improved oocyte quality and the generation of 'fitter' embryos. However, endo-metrial factors are as important in implantation as embryo quality. To this end, a case can be made that the estrogensparing effect of LHRH analog treatment may also be important.

USE OF THE LHRH ANALOGS IN SUPEROVULATION FOR IVF

In common with others18, we have shown inprovements in follicular recruitment and growth, fertilization rates and pregnancy rates when endogenous LH secretion is reduced in stimulation cycles using LHRH analogs19"21. Initially, these reductions were achieved by the use of the LHRH agonist buserelin and more recently we have also been using nafarelin by nasal spray. A derailed review of the use of the LHRH agonists and antagonists is given elsewhere in this book (see Box 2). Superovularion strategies combining the use of LHRH agonists and hMG originally, and now recombi-nant FSH (r-FSH), '11 into three categories: the so-called 'ultra-short', the 'short' (see Box 3) and the 'long' (see Box 4) protocols. The rationale of the long protocol is to induce a temporary state of hypogonadotropic hypogonadism. Follicular recruitment and growth are then stimulated by the administration of exogenous FSH. Both the short and the ultra-short protocols make use of the initial stimulatory phase of LHRH analogs to encourage the recruitment of follicles and maintain a degree ofhyposecretion of endogenous gonadotropins in the later part of the follicular phase. Exogenous FSH is concurrently administered to stimulate follicular growth. Loumaye22 has summarized the data concerning the advantages of LHRH analog therapy when compared with CC/hMG or gonadotropins alone. The major studies which he cites all claim a higher clinical pregnancy rate with the LHRH analog/gonadotropin treatments, but the magnitude of the difference is questioned when overall statistics are considered23'24. The problem lies with the quality of the data. Many of the studies are

THE NEW (30NADOTROPINS

The most recently developed r-FSH (Gonal-F; Serono Laboratories UK Ltd., Welwyn Garden City, UK) is now used in all our stimulation cycles at Bourn Hall Clinic, combined with an LHRH analog administered by nasal spray (Synarel; Searle, High Wycombe, UK) or self-administered subcutaneous injection (Suprecur; Shire Pharmaceuricals Ltd., Andover, UK). We usually start r-FSH at 150 IU subcutaneously daily after a baseline ultrasound scan and after baseline serum estimations of LH, estradiol and progesterone have shown that down-regulation has been achieved. The initial dose of r-FSH may be varied between 150 and 450 IU subcutaneously daily, depending upon the age of the patient, the presence of polycystic ovaries or a'hiscory of poor ovarian response. When a reasonable number of follicles have appeared and the leading one or two follicles are of at least 18 mm in diameter, hCG is self-administered subcutaneously. There is no doubt that the future of gonadotropins for assisted conception treatments belongs to the recently developed recombinant products, including FSH, LH and hCG. Bourn Hall Clinic has been using r-FSH sinct

94

1994, initially in clinical trials 25, and now as the preparation of choice. We have also participated in trials with r-LH. We believe that r-LH may in the future be useful if titrated in small amounts with rFSH in women who need small amounts of added LH when undergoing stimulation26, e.g. in hypogonadotropic hypogonadal anovulation27. Our own experience and that of others 28'29 is that recombinant FSH preparations, which are totally devoid of LH, are able to stimulate follicular development in normal women in a more consistent manner than the older preparations. One of our aims is to simplify the treatment as much as possible and thereby make it easier, cheaper and more convenient for couples. The development of the most highly purified subcutaneous preparation possible, in the form of r-FSH, is a very important advance in achieving this aim. It enables patients to self-administer their drugs and ensures that they do not receive the extraneous proteins contained in the older hMG preparations. Another simplification of treatment cycles would be to discontinue all endocrine monitoring. At Bourn Hall, however, we continue to monitor serum estradiol and LH levels, as We believe the information is still useful in assisting the decision-making process. Also, when we are looking back on any failed treatment cycles, it is of help in deciding how to modify future treatment cycles. It also provides useful research data.

Superovulation strategies in assisted conception the induction ofovulation in polycyscic ovary syndrome following LHRH analog administration have come together to produce the most successful Superovulation regimes so far devised. The use of LHRH analogs in combination with exogenous gonadotropins has led to a more consistent range ofoocyte quality and the generation of more, and fitter, embryos. Our ability to reduce the detrimental influence of late follicular phase LH has been significantly enhanced by the availability of recombinant FSH preparations. Although we are now replacing consistently better-quality embryos and overall success rates have improved, the failure rate ofIVF is still unacceptably high at between 75 and 90% of women starting treatment. The problem is no longer to generate goodquality embryos, but to create the optimum conditions under which the embryos can implant. Drug-induced changes in the physiology of follicular recruitment and development will have direct and indirect influences upon the ability of the endometrium to sustain implantation. The supraphysiological levels of estrogen and progesterone from multiple follicles and corpora lutea must influence endometrial development. The integrity of the corpus luceum from Superovulation cvcles, particularly those using the LHRH agonises, must also be questioned. So far, luteal phase support, either with progesterone supplementation or hCG to stimulate endogenous steroid production, has been characterized bv a singular lack of success. A much more fundamental understanding of the processes regulating endometrial function and the language and content of the communication between embryo and endomerrium must be achieved before a rational approach

CONCLUSIONS
One of the most significant advances in assisted conception techniques has been the adoption of Superovulation strategies, which control the endogenous secretion of LH. The description of the inverse relationship between basal LH secretion in the follicular phase and reproductive success in endocrinologically 'normal* women and

to luteal phase management can be adopted. However, we are at a more fundamental stage in this research and clinical applications of the most recent findings regarding Implantation are still some distance in the future.

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hi Vitro Fertilization and Assisted Reproduction

BOX 1 Clomiphene citrate and gonadotropins

(a) Schematic representation of CC/hMG regimen

Day of stimi. 'ation 56 hCG

2
CC hMG FSH

CC = clomiphene citrate 50 mg b.d. hMG = 2 ampoules/day FSH = 2 ampoules/day

(b) The conversion of urinary LH levels (uLH) into excretion rate

uLH (IU/1 per hour) = X (IU/1) x volume of sample (I) time since last passed urine (h) where X= LH value of sample

(c) Programming the onset of menstruation

Day relative to day of oocyte recovery -16 -2 0 -12 -8

NET

CC hMG
FSH

t start hMG FSH hCG withdraw start CC NET FSH NET = norethisterone 10 mg t.d.s. CC = clomiphene citrate 50 mg b.d. hMG/FSH = doses at 150-300 lU/day hCG = 5000 IU human chorionic gonadotrophin oocyte recovery

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Superovulation strategies in assisted conception

BOX 1 Continued The basic regimen

Clomiphene citrate (CC) is administered at a dose of 100 mg/day from day 2 to day 6 inclusive of the cycle. Human menopausal gonadotropins (hMG) or follicle Stimulating hormone (FSH) are administered at 150 IU (two ampules)/day from day 5 of the cycle until the day of human chorionic gonadotropin (hCG) administration.

Modifications

The following modifications may be considered in the light of the response to previous stimulation regimes: (1) The dose ofCC may be reduced to 50 mg/day; (2) Tamoxifen (30 mg/day) for 5 days may be substituted for CC; (3) High-purity FSH (HP-FSH) or recombinant FSH (rFSH) may be added from day 2 of the cycle; (4) The daily doses of gonadotropins may be titrated to the response, i.e. increased if the response is poor or decreased if the response is exaggerated; (5) The administration of gonadotropins may be omitted on the day ofhCG administration.

daily ultrasound and endocrine monitoring from day 9 of the cycle). Daily estimations of serum LH, estradiol and progesterone (combined with 4-hourly urinary LH estimations in the earlier days), carried out once the leading follicle is greater than 14 mm, guarantee the timely detection of the onset of the LH surge. Once detected, the weakened surge should be supported with hCG (5000 or 10 000 IU) and oocyte collection timed for 24-27 h after the LH secretion first doubles10.

Ovulation induction

The criteria for the administration ofhCG for ovula-tion induction in CC/hMG cycles are the occurrence of the spontaneous LH surge or one or more follicles reaching 18-20 mm in mean diameter. The larger the chosen follicle size, the more likely it is that the endogenous LH surge will be triggered. A single dose of 5000-10 000 IU of hCG is administered to induce ovularion and oocyte recovery is timed for 34.5 hours later.

Programming in CC/hMG cycles

Monitoring

-Because, of" the strong* likelihood (?f*an endogenous luieinizing hormone (LH) surge in patients treated ^vith 6Q4TMG, -close monitoring of the-endocrine profile is required during the late follicular phase, as well as regular measurement of follicle size by ultra-sonography. (The safest monitoring strategy involves

The spontaneous LH surge bedevils the prescheduling of oocyte retrieval in patients on this type of super- ovularion strategy. However, a degree of prescheduling can be achieved in most cases. The best way is to work back from the anticipated day of oocyte retrieval and program the onset of menstruation to occur 12 days before by extending the preceding luteal phase by administration of norcthisterone (30 mg/day in divided doses, see (c)).

<)7

In Vitro Fertilization and Assisted Reproduction

BOX 2 The LHRH analogs and gonadotropins

The LHRH agonists and endogenous gonadotropin levels

ment

Phase 1

Phase 2

Phase 3 -\\-

.---- Gonadotropin receptors Pituitary -- LH stores

Plasma LH Plasma FSH

LHRH analog administration

Principles of action

On first exposure to the LHRH agonist, a massive (release of pituitary-igonadotropins is. provoked (phase 1). As a result, there is an inhibition ofLH synthesis, which results in a depletion of the readily releasable, lightly glycosylated forms of gonadotropin, as well as *the more stable'highly glycosylated. reserve pools. A decrease in pituitary gonadotropin LHRH receptor numbers and a decrease in gonadotropin sensitivity to endogenous LHRH is a further consequence of the dynamics of the membrane-bound LHRH receptor, the necessity for their clustering, prior to response Initiation andotheir internalization.and degradation

once activated. The continued occupation of newly synthesized receptors maintains lowered sensitivity,' although pituitary stores of gonadotropin are re-established. The consequence is lowered circulating levels ofLH and a less strong reduction in circulating FSH (phase 2). After stopping the administration of the LHRH analogs, normal pituitary function is recovered only as de novo receptor synthesis occurs. As the gonadotrophs become sensitive to endogenous LHRH, then circulating levels of LH and FSH are restored (phase 3).

r"f

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SuperouuLition strategies in assisted conception

BOX 3 Schematic representation of the 'short' and 'ultra-short' LHRH analog (LHRH-a) regimens

Menstruation 234

hCG

t ttt
LHRH-a (SOOug/day)

J
r-FSH

The 'short' protocol

The 'ultra-short' protocol

LHRH-a
r-FSH
Both the short and the ultra-short strategies make use of the initial release phase of LHRH agonisr action (see Box 2) to assist follicular recruitment and early development. Continued follicular growth is then achieved by the administration of high doses of r-FSH (225 ILJ/day) as the pituitary gonadotrophs become desensitized. To take full advantage of the release phase of LHRH agonist action, both of these regimens must be started early in the follicular phase, i.e. by day 2 of menstruation or exceptionally day 3. However, as the onset of menstruation is not necessarily correlated with the full demise of the corpus luteum, it is essential that endocrine and ultrasound checks are made to ensure that the corpus luceum is not rescued by the endogenous gonadotropin release following agonist administration. Recombinant FSH administration should be started 1 or 2 days following agonist administration, when circulating levels of gonadotropin begin to decline. Ovulation induction

Aberrations of endogenous LH activity and LH surges do not occur often with these regimens (< 1% of cases) but are usually associated with high circulating levels of estradiol from multicystic or polycystic ovaries. The comparatively less aggressive nature of the ultra-short regimen means that normal pituitary sensitivity is regained approximately 9-14 days following the last dose of LHRHa. In normally ovu-lating women, the criteria for hCG administration are usually met before this, but, with slow responders, the LH surge may be reinstated. It is recommended that LH monitoring in blood and urine be instituted if treatment exceeds 12 days.

Ovulation induction should be considered when the leading follicle reaches 17 mm in diameter. Again 5000-10 000 IU hCG should be administered and oocyte retrieval scheduled for 34-39 h thereafter.

Monitoring As with the long LHRH-a protocols, once ovarian stimulation has begun, follicular growth can be adequately monitored by regular ultrasonography and estradiol determinations in the late follicular phase.

Programming These regimes can be programmed successfully by scheduling the onset of menstruation by extending the preceding luteal phase (see Box 1).

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In Vitro Fertilization and Assisted Reproduction

BOX 4 Pituitary suppression followed by gonadotropins

(a) Schematic representation of the regimen Ovulation Menstruation hCG

t
- 500 yg -

1
LHRH-a
-| h ?M M9 -

Jt

Luteal phase start

r-FSH

f
Menstruation
Down-regulation period Stimulation period

~\ Follicular phase start hCG

(b) Endocrine correlates of pituitary gonadotroph suppression LH < 5 IU/1 regulation has been achieved before beginning gonadotropins. Adequate superovulation can be achieved by the daily administration Estradiol < 50 pg/ml Progesterone < 1 ng/ml
The 'long' regimen Both the strategies depicted in this figure entail a period of LHRH analog (LHRH-a) administration, resulting in desensitization of the pituitary gonado-rrophs to endogenous LHRH released from the hypo-thalamus. This results in ovarian quiescence and suppression of circulating gonadotropins. The time taken to achieve this temporary state ofhypogonado-rropic hypogonadism is variable, ranging from 8 to 21 days. Once desensitization has been achieved, the daily dose of LHRH-a can be reduced to a maintenance level, preventing an LH surge and reducing but not eliminating basal secretion of LH. In normally ovulating women, the downregulation is usually complete after 7 days of LHRH-a, while in oligo-menorrheic women, particularly those with polycysric ovary syndrome, it may take considerably longer. It is always advisable to confirm by both endocrine and ultrasound testing that a suitable degree of downContinued of 225 IU of r-FSH once down-regulation has been achieved. In our experience, beginning LHRH agonist treatment In the early to midluteal phase of the preceding cycle (i.e. 710 days prior to the anticipated onset of menstruation) is the most efficacious. This method of treatment takes advantage of the progesterone-dependent lowering of LH tone seen in the luteal-phase. It also avoids the rare but not unknown consequences of starting agonist administration in the early follicular phase; namely, resurrection of the waning corpus luteum, the disruption of normal folliculo-genesis, the expansion of cystic structures within the ovary and the stimulation of recruitment and growth of follicles by the initial release phase of agonist action. While the former can be avoided by ^

100

Superovulation strategies in assisted cone'-: don

BOX 4 Continued assessment of the circulating progesterone levels and delaying agonist administration until they fall below 1 ng/ml, the latter cannot be avoided, but can be remedied by the administration of hCG and delaying r-FSH administration until the following menstrual period.

monitored for up to 3 or 4 days. When the estradiol level has dropped to 3000-3500 pg/ml, then hCG may be given. This will reduce the chance of seve.-i OHSS developing30 and is known as 'coasting'.

OvuldtiVffinduction^ Monitoring The most critical aspect of monitoring for these regimes is the assessment of pituitary desensitizarion. Although ultrasound imaging can assess the degree of ovarian quiescence, only the additional assessmentof; circulating levels of LH, estradiol and progesterone can confirm adequate down-regulation prior to the start of r-FSH. As with all other superovulation regimes, the daily dose of r-FSH should be 'titrated' to the response. Once gonadotropin administration has begun, fol-' licle growth can be monitored solely with ultrasound imaging, although the reassurance provided by estrad-iol determinations in the late follicular phase is desirable. As endogenous LH secretion is suppressed and the LH surge is abolished, lureinization rarely occurs until the ovularion-inducing dose ofhCG is given. In the presence of high levels of estradiol and the evidence of pending ovarian hyperstimulation syndrome (OHSS), the daily dose of r-FSH may be stopped, the analog continued and the patient Programming Once down-regulation has been achieved, then the scheduling of oocyte retrieval can be achieved b'.' delaying the administration of gonadotropins. In normo-ovularory women, 10 days of gonadotropin stimulation is usually sufficient to achieve the criteri for ovulation induction and ensures sufficient estrc-gen priming for the endometrium. Therefore, stimulation should be commenced 12 days prior to the scheduled day ofoocyce retrieval. The principal criterion for the administration ofhCG is the leading two follicles having reached 18 mm or more in diameter. There is a considerable amount or flexibility of timing of the administration ofhCG; v.i often do not give it until the leading follicle(s) s.:s 10 mm or 22 mm in diameter, depending on estradiol levels and the size of the other follicles. hCG is given in a dose of 5000-10 000 IU and ooc\-i recovery timed for 3439 h later.

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In Vitro Fertilization and Assisted Reproduction

References
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14. Dekcl N. Spatial relationship of follicular cells in the control ofmeiosis. In Haseltine FB, First NL, eds. Progress in Clinical and Biological Research. New York: Alan R Liss, 1988:267: 87-151 15. EppigJJ, Downs SM. The effect ofhypoxanthine on mouse oocyte growth and development in vitro: maintenance of meiotic arrest and gonadotropin-induced oocyte maturation. Dev 5/0/1987; 119:313-21 16. EppigJJ. Oocyte maturation in mammals. In Edwards RG, ed. Establishing a Successful Human Pregnancy, Serono Symposia. New York: Raven Press, 1990:66:77-86 17. Hartshorne CM, Bell SC, Waites GT. Binding proteins for insulinlike growth factors in the human ovary: identification,

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