Dr.M.Aliyari. Sahand Medical Laboratory. IVF Stimulation protocols

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1 IVF Stimulation protocols IVF protocols Natural Cycle IVF In 1978 the first successful IVF procedure was performed without the use of drugs. This technique was abandoned due to low success rates in subsequent attempts. Since then, technology and laboratory techniques have improved dramatically, and the pregnancy rate after transfer of one embryo increased, enabling physicians to reintroduce the concept of "natural cycle IVF" with the retrieval of one oocyte. The advantages of natural cycle IVF are: Simplicity Can be performed in a clinic Less invasive Less stressful Can be repeated every month No use of stimulation drugs and, therefore, no risk of ovarian hyperstimulation syndrome (OHSS) The candidates for such treatment are usually: Patients <34 years old (pregnancy rate with older patients is very low) Low-responder patients in whom IVF failed and before offering them oocyte donation Day 2-3 follicle-stimulating hormone (FSH) <10, with regular menstrual cycles (range, days) More than 85% egg retrieval with >40% of embryo deaths before blastocyst stage Implantation rate per blastocyst is 40-50%. Major disadvantages are: Low pregnancy rates after transfer of individual embryos (around 7% per cycle) High cancellation rates (over 30%) due to numerous reasons, such as abnormal folliculogenesis, premature ovulation, unsuccessful oocyte retrieval, fertilization failure, poor embryo quality, or premature luteinizing hormone (LH) surge The need for frequent and repeated blood tests to monitor the increase in LH surge (if human chorionic gonadotropin [hcg] is not administered), with around-the-clock/24-hour availability of the IVF team for egg retrieval ١

2 Protocol for "natural cycle IVF" Drugs used in the various protocols Throughout the years diverse drugs have been developed to prepare the ovaries for egg collection. Gradual improvement in laboratory research significantly increased the number and quality of embryos developed, and this reflected on the stimulation protocols as the number of oocytes needed for successful treatment was significantly decreased. Several different drugs and protocols were developed over the years; however, the most important issue is still to individualize the drugs and protocol used. Clomiphene Citrate (CC) Alone, and Clomiphene Citrate with Gonadotropins for IVF Clomiphene citrate alone was used during the early days of IVF treatment, with doses of up to 150 mg per day for 5-8 days, starting between cycle days 2 and 5. Treatment was abandonded due to the high cancellation rates related to small numbers of developing follicles and premature LH surge. Combined treatment with gonadotropins has resulted in enhanced follicular development (compared to use of CC alone) but cancellation rates are still high compared to treatment with gonadotropins only. Protocols Using Gonadotropin-Releasing Hormone Agonist (GnRH-a) In 1984 the use of GnRH-a was introduced into infertility treatment. The logic behind administering the agonist is to temporarily suppress the female's natural hormones (down-regulation) and allow for greater control over the treatment cycle. Treatment with GnRH-a generates a short flare-up of gonadotropins followed by pituitary down-regulation. Clinical applications are derived from its capacity to cause rapid desensitization of the pituitary gland as a result of prolonged nonpulsatile administration, leading to a decrease in serum gonadotropin concentration which causes inhibition of ovarian steroidogenesis and follicular growth. Use of the drug induces a state of hypogonadotropic hypogonadism, which is also termed medical gonadectomy or medical hypophysectomy, a condition that is rapidly reversible. ٢

3 Schematic graph showing the first treatment protocol for IVF treatment. The analog used was HOE 766 which was subsequently termed Buserelin. This schematic graph demonstrates the flare-up phenomena, increased serum FSH concentration after administration of human menopausal gonadotropin (hmg), and the low LH serum concentration maintained throughout treatment until the administration of hcg. The major advantage offered by the administration of agonists is the efficient abolition of the spontaneous, or ill-timed LH surge. The incidence of premature LH surges and subsequent luteinization in cycles with exogenous gonadotropin stimulation, without the use of a GnRH-a, was observed by several investigators (range, approximately 20%), leading to an increased cancellation rate. A meta-analysis of randomized controlled trials has shown that the use of GnRH-a in the IVF treatment protocols reduced cancellation rates, increased the number of oocytes collected and ٣

4 embryos developed, facilitating better selection of embryos for replacement, and thereby increasing pregnancy rates. Route of Administration Administration routes are intramuscular, subcutaneous or intranasal. Treatment protocols with GnRH-a Several protocols were designed with the combination of GnRH-a and gonadotropins. The most common are the "Long Protocol" the "Short Protocol" and the "Ultra-short Protocol". Long GnRH-a Protocol The "Long Protocol" which became the most popular procedure is based on a prolonged duration of GnRH -a treatment. There is still much controversy regarding the optimal GnRH-a protocol. In this protocol agonist treatment is either initiated in the early follicular (day 2 of the cycle), or the mid-luteal phase (day 21) of the preceding cycle. The use of GnRH-a causes an initial rise in pituitary secretion of FSH and LH. This increase in gonadotropins may cause/create a cyst formation that interferes with the treatment. The elevated concentrations of serum E2 (estradiol) and the high concentration of E2 (estradiol) in the fluids aspirated from these cysts indicate that the cysts originate from the ovarian follicles and have been termed "functional follicular cysts". The incidence of such cyst formations is reported to be 2 40%. In order to decrease the frequency of this "functional cyst" formation, it is recommended that GnRH-a be administered when the pituitary is less sensitive to gonadotropins. This can occur in the presence of progesterone in the serum of patients, and therefore it is suggested that GnRH-a be given during the mid-luteal phase, or when the patient is receiving other progesterone treatment, such as oral contraception (OC). The above protocol is/this figure demonstrates the "Long Protocol" using GnRH- a starting on day 2 of the cycle (follicular phase protocol) ۴

5 GnRH-a can be administered with a "depo-preparation" or via daily injections. Although there is an advantage for the patient when using a depo preparation, which needs only one injection, the effect of the drug can last up to 108 days (3 months) after the injection. Therefore, if the patient did not conceive in this particular cycle a "hypogonadotropic" state may ensue due to the continuous "down-regulation" of the pituitary, and menopausal symptoms may occur, such as hot flushes, headaches, mood changes, depression and night sweats. Some of these patients may need estrogen supplement to overcome the symptoms. The response of patients to gonadotropins after treatment with GnRH-a varies and depends on the agonist used. Different agonists vary in their potency. It was shown that the ovary possesses receptors to GnRH, therefore, there is a direct influence of the GnRH-a on the ovary as well. In some protocols, instead of using recombinant-fsh alone, hmg (which contains some LH activities) is administered, or additional recombinant-lh is given. Use of a depo-preparation is associated with increased gonadotropin requirements and longer stimulation periods and should, therefore, not be advocated in terms of cost effectiveness. Absorption of GnRH-a fluctuates inter- and intra-individually when administered via the intranasal route, producing an unpredictable desensitization level, but generally this is sufficient to prevent premature LH surges. Since the treatment is effective, the clinician should decide which protocol is suitable for each patient. Stimulation process Stimulation with gonadotropins usually starts when down-regulation is achieved. In most cases this occurs approximately 10 days from the first injection of GnRH-a. In patients with polycystic ovarian disease (PCOD) it may take longer (approximately 14 days). Pituitary down-regulation can be verified by measuring the endometrial lining which should be 5 mm or lower. Serum E2 (estradiol), if measured, is usually <200 pmol/l. Vaginal ultrasound examination is also needed to confirm the absence of an ovarian cyst. Should functional ovarian cysts, that secrete estrogen, be detected, three options are available: To continue GnRH-a and wait for down-regulation (this may take another 1-2 weeks) To aspirate the cysts through the vagina using an ultrasound vaginal probe (which does not require any sedation); down-regulation is usually achieved after 2-3 days Cancel the cycle and start again the following month. Dose of gonadotropin varies among centers, age of patients, body weight, and previous treatment experience if such exists. Human Chorionic Gonadotropins (hcg) Human chorionic gonadotropins (hcg) are usually administered when at least three follicles have reached 18 mm in diameter. In some instances when the patient s response is inadequate, the treating physicians should decide on the method of treatment continuation. Egg collection is performed between 34 and 40 hours after hcg injection, before ovulation occurs. Long GnRH protocol starting at the mid-luteal phase ۵

6 In this protocol, GnRH-a is initiated on day 21 of the cycle. Usually, patients will commence menses within 7 days, and the gonadotropin treatment will start on day 2 of the menstrual cycle if pituitary down-regulation is achieved. Both follicular and luteal phase initiation of GnRH-a (long-protocol down-regulation) are equally efficacious. However, when GnRH-a is administered during the mid-luteal phase, down-regulation seems to be achieved more rapidly, is more profound and, therefore, additional gonadotropin is needed for further ovarian stimulation. There is also the possibility that GnRH-ag may be administered when the patient is pregnant; however, several studies have shown no deleterious effect in such an event. Use of oral contraceptives (OCs) prior to GnRH-a administration The main advantage of a long agonist protocol with OCs is the fact that the treatment does not need synchronization with the patient's cycle. This permits time for oocyte recovery to be planned several weeks in advance. In general, a long GnRH-a protocol in combination with OCs seems to be advantageous in: 1. Prevention of functional cysts and spontaneous pregnancy 2. Prevention of LH surges 3. Improved response to gonadotropins 4. Facilitates programming of the IVF cycle and the prevention of unwanted administration of agonists during a spontaneous pregnancy. The short-term GnRH-a protocol or the flare-up protocol ۶

7 This type of simulation exploits the initial rise (flare-up) of serum gonadotropins on follicular recruitment. Therefore, GnRH-a is administered on cycle day 2 followed by gonadotropin treatment started on day 3. Pituitary desensitization will occur several days later while the patients are still on gondatropin treatment. A shorter period of GnRH-a administration for 3 days (ultra-short protocol), or for 7 days, was also suggested and was termed Ultra-Short GnRH-a Protocol. This protocol is based on the assumption that suppression of the endogenous LH surge may be obtained through a very short course of GnRH-a administration. ٧

8 Various modifications to the aforementioned protocols were suggested, such as: A micro-dose GnRH-a flare protocol This protocol was developed (with several modifications) for poor ovarian responders who received OC s during the month prior to stimulation, micro-doses of GnRH-a during the first days of the follicular phase, and gonadotropins that were introduced several days thereafter. Other strategies used with poor-responders include: Increasing the dose of gonadotropins administered Reducing the amount of GnRH-a given Cotreatment with growth hormone The low- or no-dose GnRH-a appears to be most effective for older women and those with an unexplained poor response to gonadotropins. Ovarian Stimulation Using GnRH-antagonists GnRH antagonists, as well as GnRH-a prevent the woman from experiencing an LH surge. The GnRH antagonist acts by competing with native GnRH molecules at their binding sites in the pituitary, whereas the action of GnRH-a is to "down-regulate" the pituitary's ability to produce the LH surge. The GnRH antagonist is usually initiated on (approximately) the 6th day of FSH administration that stimulates the development of follicles in the ovary. The GnRH antagonist is usually given subcutaneously once daily and continued until hcg is given. The long-acting GnRH antagonist "Cetrotide" can also be given as a single injection (in a higher 3 mg dose), rather than as daily injections of the lower 0.25 mg dose. When the single 3 mg dose is administered, Cetrotide should provide 4 days of suppression (no LH surge for 96 hours). If the patient needs more days of stimulation beyond the 96 hours, the daily 0.25mg Cetrotide injections are given until stimulation is completed and hcg is introduced. Compared to agonists, the GnRH antagonist decreases the FSH dosages required for ovarian stimulation. However, published literature has provided evidence that slightly fewer eggs are retrieved (on average), slightly less embryos are available on the day of transfer (on average), and slightly lower pregnancy rates were achieved with antagonists than with the use of agonists. Some clinics have reported that women who are low-responders to ovarian stimulation protocols that involve use of GnRH-a may receive better stimulation if an antagonist is used instead. ٨

9 Nevertheless, several advantages have been clearly identified with these protocols. The compliance of patients with the GnRH antagonist protocols was excellent due to the shortened exposure to GnRH analog administration, and to the good clinical tolerance of this third generation of antagonists. Furthermore, the amount of exogenous gonadotropins needed for ovarian stimulation was reduced, as well as the occurrence of OHSS (ovarian hyperstimulation syndrome). Finally, the overall cost of this regimen was significantly lower than that of the GnRH-a protocol. The new GnRH antagonists also permit the design of milder stimulation schemes, with the return to the use of clomiphene citrate, minimal stimulation, or even natural cycles. ٩

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