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The use of GnRH agonists and antagonists in infertility

Fidelis T. Akagbosu
In the early days of in vitro ferrilizarion (IVF), natural cydes were commonly employed1 and classic stimulation prorocols involved the use of clomiphene citrate and gonadotropins. The existence of elevated basal levels of luteinizing hormone (LH) were shown to have a poor correlation with the success of che IVF cycle2. Premacure LH surges, which occurred owing to the positive feedback ofrising estradiol levels induced by the gonadotro-pins, led to premature luteinizarion artd were considered reasons for cancellation of the rreatment cycle. The dis-covery that the continued administration ofgonadotro-pin releasing hormone (GnRH) analogs induced a state ofhypophyseal desensitization provided a welcome solu-tion ro the problems that had caused a large number of IVF cycle cancellations. The first clinical crials of the use of GnRH analogs showed thar they were effective in eliminacing the basal levels of LH3'4 and in reducing the incidence of low responders5'6. The incidence of cycle cancellations dim-inished significantly and the GnRH analogs were soon in routine use. The agonists led to desensitization of the pituitary gonadotropin cells and a reduction in the num-berofGnRH receptors, while the antagonists produced an immediate effect by competitive blockade of the GnRH receptors. Various protocols combining the use ofagonistic ana-logs and gonadotropins in IVF cycles have been des-cribed. Reports in the current literature have associared the use of GnRH agoniscs with high pregnancy rates after IVF and embryo rransfer (ET)7. Debatable facts include the suggesrion that agonist use has a harmful GnRH


effect on oocytes and embryo quality, and possible direct effects on the ovary8. The GnRH antagonists have recently been used in phase III clinical trials and should be available for routine use in IVF cycles in the near fut-ure. Ancagonist use has various advancages, such as short-term and low rnedication exposure during follicular development and immediate suppression ofLH and follicle stimulating hormone (FSH) levels. As there is no 'flare effect', the frequency ofcyst formation would be lower, pacient management would be easier and stress, cost and hyperstimulation may all be reduced. The pitukary response is preserved fol-lowing ovarian stimularion with human menopausal gonadotropin (hMG) and the GnRH ancago-nist cetrorelix9 and it may not be necessary to provide luteal supporc in IVF-ET cycles ifter antagonist use.

Gonadotropin releasing hormone is a decapeptide pro-duced in the mediobasal hypothalamus by neurons loc-ated in the arcuate nucleus. It was first postulated by Green and Harris in 194710 and first isolated and characterized by Schally and colleagues in 1971". In narive GnRH, the amino acid sequence in positions 2 and 3 facilitates the release of gonadotropins while the sequence in position 6 is involved in enzymatic cleavage; residues 1, 6 and 10 are important for the three-dimensional structure and thus for receptor binding. GnRH binds to specific transmernbrane receptors in the


On the basis of the rnechanism ofcor peritive binding. ten-fold rise in LH and four-fold elevation in estradiol15. This causes paradoxical suppression of the pituitary gonadotropin synthesis and liberation (desensinzation). recovery of gonadal function to Table 1 Structural formulae ofnative luteinizing hormone releasing hormone (GnRH) agonistic and antagonistic analogs •nces ofGnR}'.. normal menstrual cycle is re-established in approx mately 6 weeks. k is possible to modulate the degree hormone suppression by the dose of antagonist that administered. Within 12 h ofadministration. GnRH AGONISTS The agonists were developed soon after the idcntification ofthe structure ofnative GnRH. Knobil12 showed that GnRH givcn continuously causcd a decrease in LH and FSH levels. The agonists have a 100—200 times higher binding affinity for the GnRH receptors than does the native molecule. Reproduction gonadotropic cells.In Vitro Fertilization aiid Assisted. the so-called 'flare effect' leads to a five-fold increase ofFSH. This is the bas for the clinical use ofthe agonists. The aim was . Aftcr cessarion of treacme with ancagonlsts. Rece tor blockade is immediate and prevents the acrion native GnRH. / GnRH Agonists Buserelin Goserelin Leuprorelin Triprorelin Nafarelin AntagomsR Cetrorelix Nal-Glu Antide Ganirelix Azaline B Antarelix D-Nal D-Nal D-Nal D-Nal D-Nal D-Nal D-Phe D-Phe D-Phe D-Phe D-Phe D^Phe D-Pal D-Pal D-Pal D-Pal D-Phe D-Pal 4 4 4 4 4 4 5 Arg NicLys 5 Phe Phe pGlu 2 His 3 Trp 4 Ser mino acid seqiii 5 Tyr GnRH ANTAGONISTS The GnRH ancagonists were developed in parallel wii the agonists. Their development hiscory has been pl gued mainly by the high incidence of histamine relea following use ofthe first-generation anragonists. Depending upon rhe indication for th rapy. Contin-uous (nor pulsatile) GnRH analog administration causes opposite effeccs: internalization of the agonist/receptor complex and a decrease in the number of receptors (down-regulatlon). Postmenopausal estradiol leve are commonly reached after 21 days16. rcsult in the arrest of follicular development and cause fall in sex steroid levels to castrate levels. which may have the advantage avoidlng hormone wlthdrawal effects such as vagir dryness or hot flushes17. this permits the physician for instance to mainta defined esrradiol levels. Table 1 shows the amino acid sequences ofGnRH and its analogs currently in use. Antag nists competitively bind to pitukary GnRH recepto and do not induce the release of gonadotropins.0 increase GnRH siability and binding affinity to its receptors by modifying the amino acid sequertce13'14. The modifi-cations were mainly at positions 6 and 10 and resuked in increased potency. In women. D-Ala D-Ala D-Ala . The decreased levels of FSH and LH A. Analog administrarion initially induces the llberation of high amounis of LH and FSH from the pitultary and an increase in the number ofGnRH receptors (up-regulation). 1 1 1 1 1 2 2 2 2 2 3 3 3 3 3 4 4 4 4 4 5 5 5 5 5 D-Ser D-Ser D-Leu D-Trp D-Nal D-Cit D-Glu D-NicLys D-hArg D-Phe D-Hcit 7 7 7 7 7 7 7 7 7 7 7 8 8 8 8 8 8 8 Lys(iPr) hArg Lys(iPr) Lys(iPr) 9 9 9 9 9 9 9 9 9^ 9r 9 ethylamii AzGly ethylamii Gly-NH: Gly-NH: D-Ala D-Ala D-Ala •'. 6 Gly ! analogs 7 8 Leu Arg 9 Pro 10 Gly-NH. The pituitary blockad persisrs during the agonist treatment but is completd reversible after therapy.

Klingmuller and colleagues20 scudied mean serum LH levels after subcutaneous injection of 3 mg cerrorelix in Agonist use in assisted reproduction techniques DifiFerenr treatment schedules using GnRH analogs s. generalized edema and anap.. including the so-csJled 'long' prococol..and fourth-generarion antagonisrs are the modern antagonists. female voluiitcers during three cdnsecutive cycles: a . 3. Com-pared to baseline values.6and 10 ofGnRH. a treatment cycl?and a posc-treatment co-itrol cycle. Th*e posp-treatment-cyclé was comparable to tKe ore-treatment cycle. It is not teraro-genic and is the most porenc GnRH ancagonist in monkeys19. and to allow reco^ ery. It has been well characterized in animal models and has reversible pharmacological action. GnRH analog' are used in IVF treacment cycles as well as in frozen embryo transfer cycles when hormone replacement is requ::ed. The invesrigational drug ancarelix has been used in pre-clinical trials while ganirelix and ancide have been used in phase 1 trials.5%. due to the existence ofGnRH rec-eptors on mast cells which degranulace after binding of basic antagonist18 and. Wirhin a period of 8 h.. The antagonist cetrorelix Cetrorelix is the most advanced of the GnRH antago-nists. gonadotropin actions such as 83% loss ofLH recepcors in rat granulosa^degrees is. the 'long' proiocol is generally ihe most effeccive and is most often used ac presenc.. depcnding on the duration of the preceding treatmenc. CLINICAL APPLICATIONS OF GnRH ANALOGS IN ASSISTED REPRODUCTION TECHNIQUES Premature LH surges have a negacive impact on the quality of oocytes and embryos and subsequentl'.af-ter* cetrorelix injection* (and somerimes also with a^onists!-or native GnRH).Allêrgic side-cff(fei:s»wiilT*"the"'' early antagonists included local erythema and in-duption. luteal phase supporr with progesterone or human chori-onic gonadotropin (hCG) is required after analog us. but very high doses were requircd.on-trol cycle. In the IVF cycles. As a result of the pituirary desensirizatibn.-in positio. a long duration of action and no clinically relevant side-effects.nd gonadotropins are now in use.ic has-the-disadvantages-of^a-loi'ig^treatment pe:i6d uiipil"dësensitization is achieved and relatively high i:osts due to an increased requiremenr for hMG24. Maximum plasma concencration of cecrorelix is reached in 1-2 h after the injecrion with all doses tesred.hylactoid reacrionï The observed side-effeccs were caused by the inducrion ofhistamine release. esrradiol reachcd postmenopausal values. Sincc the antagonists do noc cause pituitary exhaustion. These were developed in search for the ideal antagonist with high potency. Local erythema of vary. Specific effects of GnRH agonist use in IVF have been documented.seen. No sysremic side-effeccs have been observed in trials. also of naciveGnRH19. About 85% protein binding occurs in human albumin and human plasma and no differences in binding have been documented berween males and females16. 2. in which the 'flare-up' effect is urilized. approximacely 20% ofsrimulated IVF c. Antide srudies in ovariectomized monkeys showed prolonged suppression ofgonadotropins. to a much lesser excent. an LH surge followed by a rost-ovulatory rise in progesterone was found in all women.' on pregnancy rate21'22. The firsc-generarion antagoniscs were charaaerized by modifications in positions 2 and 3. the iriyoduction of-D-arginiiic. Pre-treatment with agonists signifi-cantly reduces the occurrence of LH surges. similar effeccs have been demonsrraced in humans after subcucaneous injecrion. Hsueh and co-workers25 noced snri.The nse ofGnRH agonists and antagonists in infc'^'ility normal state occurs. these cells can respond ro an ade-quate stimulus almost immediately. The rhird. in a pa-aJlel manner. How-e'<'eE. and the 'short' and 'ultra-short' protocol.1°c and FSH by 63. They usually have modifications in amino acid positions 1. The second-generarion antagonists had modifications in positions 1 and* 6 and. which aims at complete pitukary sup-pression. LH was reduced by l6.'cles were cancelled. this percentage has decreased to 2% anc rer-tilization and implantarion rates have increased23. decreased aromacase activity and 85 . The in vitro assay systems were effective.»6-:..'ith agonist use. At different time intervals after the ter-mination oftreatment. owing to premature LH surges. Before rhe agonist era. later. LH levels decreased and subsequencly were deeply suppressed. V'.

Albano and collcagues27 compared differeni doses of thc GnRH antagonist cetrorelix during comrolled ovarinn hyperstimulation (COH). Furcher clinical scudk will show whether luteal phase support is necessar in IVF-ET cycles where antagonists had been used. but fell again thereafter. No pre-maturc endogenous LH surge occurred in parients treatcd wkh 0. As a result ofthe antagonist treatmenc. This observation might be explainec by the rapid increase in estradiol levels in the combinec treatment schedule. which up to now has been the least costly rreatmcnt26. this stimulates metabolism of progesterone co 20a-OH progescerone with little progestational activity. starring on day 2 of the menstrual cycle.25 mg ofcecrorelix. Pltuitary responsiveness following cetrorelix use Felberbaum and associates9 demonscrated pituirary responsiveness following cetrorelix use in IVF by applying a GnRH test. whereas 40-50 ampules werc required in the 'long' protocol of GnRH agonists in thc center. Cetrorelix was administered daily subcucaneously starting on cycle day 7. This complication is frequen in PCOD patiencs and necesskates careful monitorin. The mean number of oocyres retrieved was.5 and 0. Three hours before ovularion induction with hCG. To date.8. of 3 mg and to 25 mlU/ml in those given 1 mg < cecrorelix.25 rng administered daily. 25 p. The amount cff hMG needed after cerrorclix treatment is comparable with the 'ukra-short' prorocol. initial LH leve were very low. which results from the fast anc synchronized development ofmulriple follicles16. in the treacment cycle. Luteinizing hormone me surcments were made 30 and 80 min after the injectio. no evidence for disturbance of the lure phase has beerr detected witli the use of? antagonists2 Lin and coworkers29 reported that the granulo' cells from women regulated by the GnRH antagoni. It appears thac the incideno of severe OHSS is higher in hMG-scimulated cycle of 'down-regulared' patients than in those treacec wirh hMG alone. cetrorelix showed a higher degree of steroidogenes in all cultures (tescosterone.In Vitro Fertilization anii Assisted Reproduction decrcascd progescerone accumulation. The minimal effective dose of cetrorelix able to prevent an LH surge in COH patiencs was 0. Diedrich and coll-eagues23 were able ro stimulare parients successfully after pre-trearment with cerrorelix by daily administration" scarting on cycle day 7 until induction of ovulacion. LH levels increasc to about 10 mlU/ml in parients who received a do. In 42% of the embryos. In 69 parients COH v. ovulation may be induced with GnRH (nath or agonist) insread ofhCG. hCG and cAMP) cha the granulosa cells obtained from the women regulace by the GnRH agonisc buserelin. This could be beneficial'i patients with polycystic ovarian disease (PCOD) or . The beneficial effec results from interference with the pathological LH/FSF rario commonly found in these women. owing ti the inhibition of gonadotropins. Analog use also leads to the activation ot 20a-OH progesterone dehydro-genase. Six pregnancies were achieved in the first 42 parients trearcd by this regimen and three healthy babies were delivered. risk ofovarian hyperstimularionsyndrome (OUSS).g ofGnRH was injected. and cetrorelix was adminiscered from day 6 of the hMG creatment every day up to and includlng the last day ofthe hMG injection.'as carried out with hMG. After 30 min. Long-term treatmen with agonists has been shown to result in the suppressioi of escradiol and ovarian androgen levels. Afrer fzill sup pression of the endogenous gonadotropins is reached normal follicular developrnent and synchronizatioi can be established by administration of exogenous LP and FSH or even FSH alone. The mean amount of hMG required per parient was 27 ampules.1 per patient and the fertillzation rate was 61. 86 . Since the pituitai response is preserved following the administration i cetrorelix. Antagonist use in assisted reproduction techniques The simpliciry offered by ancagonists in regulating the menstrual cycle has made them porentially useful in assisted reproducrion cycles. Polycystic ovarian disease The rreatment of PCOD parients with GnRH agonisi has been well established30'31. Pretreatment with an agonist cannot preven the occurrence ofOHSS.5%. Patients in this scudy were srimulated with hMG starring on day 2 ofthe cycle. embryo quality was judged as being 'excellent' by morphological criteria.

s. Nakamura and' colleagucs36 reported rwo protocols combining GnRH analogs and gonadotro-pins for IVF-ET in patients with various scages ofendo-metriosis who were resiscant to conventional therapies. the-Gn'R'H an'alog was scarted'at the mid-luteal phase and exogenous gonado-tropin was commenced betweerfth'e' 3rd"and the 7th"day of the mensrrual cvcle after pituirary. suffering from PCOD with high estradiol values (> 5000 pg/ml). 27%). GnRH analog. The-clinical pregRan6y-»rai.compadson* to the agonists. ovulation Is induced by the administration of hCG. Endometribsis and'uterihe myorna . attleast. for. Ih general during hMG-stimulated cycles.suppre. OHSS may be avoided.orper'transtér \va. Successful attempts have been made to in-duce ovulation by a single administration of an agonist. At Bourn Hall Clinic. In both endometriosis and urerine fibroids the thera-peutlc use ofGnRH agonists is well established but again the initial 'flare effects' are undesirable.—iound*ro be superiorwitH the ultra-long prococol (67 vs. In theultra-longprotocolgrftup (21^patients).ucerine fibroid volume and reversing the related symp-tomarology. Women of less than 35 years bf age. The fibroids tend to return to their pre-treatment size about 6 months after creatment is discon-rinued. Beneficial efTects of such use include some reduction in fibroid volume and thus easier access ro che ovaries at crans-vaginal oocyte recrieval. Romer 37 reported the safety of hysteroscopic myoma resection of submucous myomas with largely intramural components following pre-treatment with rwo or three rnonthly injections of GnRH analogs. the combinarion of both treacment modalities (antagonist and agon-ist) might result in an additional benefit. Since the pituitary responsivcness is preserved during the co-administration of an antagonist.»LH/F-SH-ratio found in PCOD patients16.i. using the flare-up effect. It is imporcant to increase the dose of gonadotropins in the stimulatory phase of the IVF cycle to achieve adequate ovarian fbllicular develop-menr. ®i»aaian.Thc use ofGnRH agonists and antagonists in infertility Phc cype oft&tcal supporc recomiTicndéd'''is important in the presencc ofOHSS.h^perstHTHilaiaoi»syB'Efe'onie Ovarian hvperstimulation is a well-known and poten-tially lifethreatening iatrogenic-complication'ofovula-tion induction32. These features may be responsible for the occurrence of the complication of OHSS after ovarian stimulation and ovulation induction with hCG. injectiansQi» hCG—In-. GnRH ancagonist use would have the advantages of immediate suppression of gonadotro-pins and a greater suppression of LH comparcd to FSH secrerion. which has a high dcgree of homology and synergisric effecrs with LH33 and thus can mimic the normally occurring mid-cycle LH peak.60..sion. Under these conditions. in spire of irs high similarity to LH. it has a prolbnged half-life and does not induce the typical mid-cycle increase in FSH 34.'YiilïQLigftiAe overalAc^lrsiwictfrIV>p(*iEC-*good-*in end&^ metriosis. The immediate suppression produced by the administration ofa GnRH-ancagonist would offer advantages in reducing the dura-tion of treatment and leading to a faster improvement of subjective symptoms. seem ro be at particularly HigK risk fdï th'e development of' OHSS. With thc use ofprogesterone. che frequency of complications seems to bc much lower than aftcr mulriple. In the long protocol group" (l'l' parierits). was administered. luteal phase support may not be necessary. If hCG is not used for ovulation induction. Future clinical trials would f'" '. However. GiïR^ft analogs» aTe efficient' a» reducing. the optimal protocol of treacment has not yet been détermined.days prior'co ovarian stimularion along with menotropin.s. This difference is clear during-«a short-term treatment and might be advantageous with regard to the hig^(. Pro-longed GnRH analog suppression of ovarian function before superovularion may thus overcome some. In none of the patients treated in this way was severe OHSS observed35. 87 . hCG does not cause idenrical physio-logical reaccions. in cases in which an antagonist had been used in the preceding stimulatory phase.causes of mferCTltÉ^iw-pauentSAvithendometriosT?'*. The beneficial effeccs of pre-surgical GnRH analog use are well established. GnRH agonists have been used for 2—3 months before IVF treacment in women with moderate-sized uterine fibroids.and multiple early-scage follicles.

Smith \.fWomno/1947.2: 309 7. In HollanderJR. Isolation and properties ol the FSH and LH-releasing hormone. Gynakol Geburtsh Rundsch 1992. CaratyA. Hum Reprod 1994. Baba Y. Diedrich K.the development ofGnRH antagoniscs witl-sufficient oral bioavailability represent present and futur< challenges16. Bast RC. Agonists are still preferred in dinicaL situations where the use of a single monthly injection is required. Nieschlag E. Hum Reprod 1995. Bauer 0. Gonadotrophin-releasing-Hormon und Analoga — Physiologic and Pharmakologie. "raft IL. Caraty A. Schatz B. GnRH-Analogs. Maheux R. Annual Progress in Reproductive Medicine.9(Suppl4):13 10. Reversible hypogonadism indaced by a luteinizing hormone-releasing hormone (LH-RH) agonist (buserelin) as a new therapeutic approach for endometriosis./. Howles CM. Biochem Biophys Res Commun 1997. would exert side-efFects during long-term treacment based on the pharma-cological action ofthese compounds. Arimiira A. Porter RN. Edwards RG. Hollander V. Hum Reprod 1987. The ancagonist seem to offer many advantages compared to the agonists but their efficacy and safety have to be proved in long term studies. Hypotha-lamic and other peptide hormones. In Asch RH. Coelingh-Bennink HJT. Felberbaum R. reduced require-rnents for gonadotropins and reduced'cost-. Use of GnRH analogs in IVF.36:53-88 13.. At presenc. Kupker W. The neurovascular link between the neurohypophysis and adenohypophysis.5: 11. 3rd edn.2:521 3. Remohi J. et al.47: 639^3 6. Runnebaum B. Lancet 1984. Purdy JM. Hum Refirod 1987. such as decrease ir libido or loss ofbone density. As with GnRH agonists. Biochem Biophys Re: Commun 1971. Comaru-Schally AM. In Bouchard P. et al. Bouchard P. Carnforth. Green JD.4l:863-71 16. Hedon B. Development and applications of luteinising hormone-releasing hormone anragonists in the trcatment of infertility: an overview. Howlcs CM. Knobil E. ii has to be expected that GnRH anragonisrs. Reissrnann T. GnRH agonists are well-tested and easily available pharmaceutical agents in sex steroid-dependent diseases and in the management of IVF cycles. Palou SN. CONCLUSIONS It is scill coo early co speculate about. Carnforth. etal.In Vitro Fertilization and Assisted Refiroduction hopefully address thc place ofantagonisrs in suppressing endoiTietriosis or moderate-sized uterine fibroids before Wf treatment. 10:1974-81 17. Steptoe PC. have not yet been established. Fertil Steril 1987. but the exacc clinica regimens for GnRH antagonists in IVF trcatment cycle. Contraception. LemayA. eds. etal.24:694-9 8. There has been much progress. Suitable susrained deliver) systems and. Mechanism ofaction and clinical uses ofGnRH-antagonists in women. Edwards RG. Felberbaum R. eds. Haour F. et al. when appliec for complete hormone suppression. Parls: Elsevier. Philadelphia: Lea & Febiger.tWo fertilisarion. Behre HM. Cancer Medicine.the possible end of the 'agonisr era'16. Effects of high ronic levelsofluteinisirighormoneonoutcorneofw.74:488-92 14. eds. Hormone profiles and pitukary response under ovarian stimulation with HMG and GnRHantagonist (Cetrorelix). expeccations would include a shorter per-iod of stimulation.2:569-71 5. Macnamec M. Kiescl L. Br JObstet Gynaecol 1980. eds. Ovarian stimulation by a combination ofgonadotropin releasing hormonc agonist and gonadotropins for in vitro fertilisation. BringerJ. et al. Macnamee MC. Harris GW. Smitz J.43:393-9 12. Baram T. Fertil Steril 1984. Devroey P. Resisrance ro enzymatic degradation ofLH-RH analogs possessing increased biologi-cal actlviry. Braeckmans P. Gonadotrophins and Gonadal Peptides. clinics or in the context ofclinical trials. Pellicer A.32:22-30 19. UK: Parthenon Publishing. In Bouchard P.2:1284 4. Schally AV. Pregnancy after IVF when high ronic LH is reduced by long-term treatment with GnRH agoniscs. Neveu S. Lancet '986. Koch Y. Recent Prog Horm Res 1980. Frei E. fewer side-effects. etal. Studd JWW. Edwards RG. Fertilite. Faure N. 1993:827 15. Recent Progress on GnRHand Gonadal Peptides. Managemenr of failed cycles in an IVF/GIFT programme with the combina-tion ofa GnRH analogue and HMG. There are ongoing clinical scudies involving cetrorelix and antide in this field. Gonadotrophin re-leasing hormone analogue-induced regulation of cesticular function in monkeys and men. 1993:211 136-46 88 . Inducrion ofovulation for m vitro fertilisation using buserclin and gonadotrophins. Medalie D. GnRH. 1990:209 18. the use of aniagonists is restricted to a few References 1. Hazum E. Establishing full term human pregnancies using cleaving embryos grown in vitro. French IVF Registry FIVNAT 1995 Report. Schally AV. UK: Parthenon Publishing. 1993: 107-25 9. Franchimont P. Sexualïte 1996. The neuroendocrinecontrolofthe menstrual cycle.87:737-56 2. Weinbauer GF.