AOA Versus Non-AOA in Low Prognosis Patients by the POSEIDON Criteria

Study Overview
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Brief Summary
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Poor ovarian response (POR) remains one of the significant challenges of Assisted Reproductive Technology (ART). Facing difficulties related to clinical practice, optimizing the embryo culture process is necessary to improve the embryo number and quality in this group of patients. Potential techniques mentioned in the current literature include follicular size at trigger, dual trigger, artificial oocyte activation (AOA), blastocyst transfer, and the role of preimplantation genetic testing for aneuploidy (PGT-A). AOA is currently expected to improve treatment outcomes in poor ovarian responders with the potential for clinical efficacy. However, this issue has not been evaluated before.

Detailed Description
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Poor ovarian response (POR) remains one of the significant challenges of Assisted Reproductive Technology (ART). Patients with POR yield a low number of oocytes, leading to a low number of useable embryos and a decline in the live birth rate. According to the consensus of the European Society of Human Reproduction and Embryology (ESHRE) in 2011, POR was diagnosed using Bologna criteria. However, some recent studies show the classification by Bologna is not efficient, because the oocyte number should be combined with female age since the likelihood of achieving a live birth among patients with similar oocyte yield ultimately depends on the age of the patient. In 2016, POSEIDON (Patient-Oriented Strategies Encompassing IndividualizeD Oocyte Number) Group was established and released the new criteria. The POSEIDON criteria proposed a shift from the terminology of POR to the concept of low prognosis. According to POSEIDON criteria, low prognosis account for 30-40% of all stimulated in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) cycles. The low prognosis patient is classified into four groups according to the results of ovarian reserve markers (AMH, AFC, or both), female age, and the number of oocytes retrieved in previous cycles, such as: maternal age < 35, AMH ≥ 1,2 ng/ml and AFC ≥ 5 (subgroup 1a: < 4 oocytes; subgroup 1b: 4-9 oocytes); maternal age ≥ 35, AMH ≥ 1,2 ng/ml and AFC ≥ 5 (subgroup 2a: < 4 oocytes; subgroup 2b: 4-9 oocytes); maternal age < 35, AMH < 1,2 ng/ml and AFC < 5; maternal age ≥ 35, AMH < 1,2 ng/mL and AFC < 5. Although many efforts have been made to improve treatment outcomes in this group of patients, such as researching, understanding, and modifying clinical ovarian stimulation regimens, the results are still not feasible. Especially, group 4, which have advanced maternal age (≥ 35) and seized for 14.4% of low prognosis, has a low cumulative live birth rate (11% in group 4). Female age is a critical element in the POSEIDON classification because age is crucially related to embryo ploidy and more importantly live birth outcome. The probability of having embryo ploidy sharply declined after the age of 34 and was lower than 50% in women aged 35 years and over. Therefore, patients in group 4 will have an increased risk of aneuploidy embryos, decreasing the live birth rate in these groups of patients. A recent study evaluated cumulative live birth rates per cycle, there was a remarkable difference between POSEIDON patients (21, 43, 10, 25, 29, and 17% in groups 1a, 1b, 2a, 2b, 3, and 4, respectively) and non-POSEIDON counterparts (52%). Facing difficulties related to clinical practice, optimizing the embryo culture process is necessary to improve the embryo number and quality in the POR group. Potential techniques include follicular size at the trigger, dual trigger, artificial oocyte activation (AOA), blastocyst transfer, and the role of preimplantation genetic testing for aneuploidy (PGT-A). In Vietnam, AOA was first reported in 2011, performing on 1588 oocytes, and said the fertilization rate was higher in the ICSI - AOA than in the ICSI group (80.8% vs 74.3%, respectively; p<0.002). AOA is expected to improve treatment outcomes for low prognosis patients, especially in group 4 by the POSEIDON criteria with the potential for clinical efficacy and safety. Therefore, this study aims to evaluate the effectiveness and safety of AOA on treatment outcomes in low prognosis patients defined by the POSEIDON criteria (2016).

Official Title
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The Effectiveness and Safety of Artificial Oocyte Activation With Calcium Ionophore in Low Prognosis Women Classified as POSEIDON Group 4: a Randomized Clinical Trial

Conditions
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in Vitro Fertilization

Intervention / Treatment
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* Procedure: ICSI with AOA
* Procedure: ICSI without AOA


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Was diagnosed as low prognosis patients by the POSEIDON criteria in group 4: maternal age ≥ 35 years old, AMH < 1,2 ng/ml and AFC < 5 Cycles ≤ 3 Oocytes could be collected with OPU procedure Ovarian stimulation with GnRH antagonist protocol Agree to participate in the randomization Exclusion Criteria: Uterine abnormalities such as unicornuate, bicornuate uterus, didelphys and adenomyosis Recent history of any current untreated endocrine abnormality Gonadotropin resistance syndrome Contraindications of gonadotropins Absolute asthernozoospermia Cryptozoospermia Surgical sperm retrieval Previous low fertilization (< 30%) Globozoospermia Cycles using donor oocytes Preimplantation Genetic Testing (PGT) cycles

Ages Eligible for Study
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Minimum Age: 35 Years
Maximum Age: 45 Years

Sexes Eligible for Study
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Female

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Treatment
Allocation: Randomized
Intervention Model: Parallel Assignment
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Active Comparator: ICSI with AOA<br>The oocytes were transferred into the calcium ionophore activation solution for two times of post-ICSI AOA (Ionomycin concentration of 10 µM). Then the oocytes will be washed several times using the medium drops in dish AOA and dish ICSI, then divided into drops with maximum 3 oocytes per drop for culturing. After that, the culture dish will be put in the K-system G185 incubator at 37oC, 6% CO2, and 5% O2. | Procedure: ICSI with AOA<br>* Step 1: All post-ICSI oocytes will be added in the medium drop in AOA dish, these oocytes will be placed in the incubator at 37oC, 6% CO2, and 5% O2 for 20 minutes. Step 2: After 20 minutes, calcium ionophore stock (Sigma-Aldrich - USA) will be diluted into 10µM drops in AOA dish. Step 3: After 10 minutes of incubation, all post-ICSI oocytes will be moved to the AOA drop, then this dish will be put back in the incubator for 10 minutes. Step 4: After 10 minutes, all the oocytes in drop AOA will be moved to another drop for rinsing and then put back in the incubator for 20 minutes. Step 5: After 20 minutes, the oocytes will be moved into drop AOA and then put back in the incubator for 10 minutes. Step 6: After 10 minutes, all post-ICSI oocytes will be added in the medium drop, then divided into drops with maximum 3 oocytes per drop for culturing. After that, the culture dish will be put in the K-system G185 incubator at 37oC, 6% CO2, and 5% O2<br>|
| Active Comparator: ICSI without AOA<br>Post-ICSI oocytes will be cultured in drops containing the Sage - 1 - StepSM medium (maximum 3 oocytes per drop) at 37oC, 6% CO2 and 5% O2 in K-system G185 incubator. | Procedure: ICSI without AOA<br>* Conventional ICSI procedure will be performed without the application of AOA.<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Live birth rate | The complete expulsion or extraction from a woman of a product of fertilisation, after 24 completed weeks of gestational age; which, after such separation, breathes or shows any other evidence of life, such as heart beat, umbilical cord pulsation or definite movement of voluntary muscles, irrespective of whether the umbilical cord has been cut or the placenta is attached. A birth weight of 350 grams or more can be used if gestational age is unknown (twins are a single count). | At 24 weeks of gestation | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Fertilization rate | The percentage of transformation of micro injected oocytes into two pronuclei | One day after oocyte retrieval | 
| Fertilization failure rate | The percentage transformation of micro injected oocytes into no pronuclei | One day after oocyte retrieval | 
| Abnormal fertilization rate | The percentage transformation of micro injected oocytes into more than two pronuclei | One day after oocyte retrieval | 
| Total embryos on day 3 | Number of embryos on day 3 | Three days after oocyte retrieval | 
| Good quality day 3 embryo rate | Number of good quality embryos on day 3 | Three days after oocyte retrieval | 
| Total embryos blastocyst | Number of embryos on day 5 | Five days after oocyte retrieval | 
| Good quality blastocyst rate | Number of good quality embryos on day 5 | Five days after oocyte retrieval | 
| Positive pregnancy test | Serum ß-hCG ≥25mIU/mL | At 2 weeks after embryo placement | 
| Implantation rate | Implantation rate is explained as as the number of gestational sacs per number of embryos transferred. | At 3 weeks after embryo placement | 
| Cumulative implantation rate at 12 months | After 12 months, most patients doing IVF have finished all their frozen embryos; therefore, we consider this time point for analyzing the cumulative implantation rate. Cumulative implantation is total case of gestational sacs per total patient to be enrolled into the study at 12 months after randomization | 12 months after randomization | 
| Clinical pregnancy | Having at least 1 gestational sac on ultrasound at 5 weeks' gestation | At 7 weeks after embryo placement] | 
| Cumulative clinical pregnancy at 12 months | After 12 months, most patients doing IVF have finished all their frozen embryos; therefore, we consider this time point for analyzing the cumulative clinical pregnancy rate. Cumulative Clinical pregnancy is total case clinical pregnancy per total patient to be enrolled into the study at 12 months after randomization | 12 months after randomization | 
| Ongoing pregnancy | Having at least 1 gestational sac on ultrasound at 12 weeks' gestation with heart beat activity | At 12 weeks after embryo placement | 
| Cumulative ongoing pregnancy at 12 months | After 12 months, most patients doing IVF have finished all their frozen embryos; therefore, we consider this time point for analyzing the cumulative ongoing pregnancy rate. Cumulative ongoing pregnancy is total case ongoing pregnancy per total patient to be enrolled into the study at 12 months after randomization | 12 months after randomization | 
| Ectopic pregnancy | A pregnancy outside the uterine cavity, diagnosed by ultrasound, surgical visualisation, or histopathology | At 12 weeks of gestation | 
| Cumulative ectopic pregnancy at 12 months | After 12 months, most patients doing IVF have finished all their frozen embryos; therefore, we consider this time point for analyzing the cumulative ectopic pregnancy rate. Cumulative ectopic pregnancy is total case ectopic pregnancy per total patient to be enrolled into the study at 12 months after randomization | 12 months after randomization | 
| Miscarriage | The spontaneous loss of an intra-uterine pregnancy prior to 12 completed weeks of gestational age | At 12 weeks of gestation | 
| Cumulative miscarriage | After 12 months, most patients doing IVF have finished all their frozen embryos; therefore, we consider this time point for analyzing the cumulative miscarriage rate. Cumulative miscarriage is total case miscarriage per total patient to be enrolled into the study at 12 months after randomization | 12 months after randomization | 
| Multiple pregnancy | ≥1 gestational sac at early pregnancy ultrasound | At 6 to 8 weeks' gestation | 
| Cumulative multiple pregnancy | After 12 months, most patients doing IVF have finished all their frozen embryos; therefore, we consider this time point for analyzing the cumulative multiple pregnancy rate. Cumulative multiple pregnancy is total case multiple pregnancy per total patient to be enrolled into the study at 12 months after randomization | 12 months after randomization | 
| Vanishing twins | Vanishing twins is defined as a pregnancy with two or more gestational sacs or positive heart beats at 7 weeks of gestation, but only one at 12 weeks' gestation. | At 12 weeks' gestation | 
| Cumulative vanishing twins | After 12 months, most patients doing IVF have finished all their frozen embryos; therefore, we consider this time point for analyzing the cumulative vanishing twins rate. Cumulative vanishing twins is total case vanishing twins per total patient to be enrolled into the study at 12 months after randomization | 12 months after randomization | 
| Multiple delivery | Birth of more than one baby beyond 24 weeks | At 24 weeks' gestation | 
| Cumulative multiple delivery | After 12 months, most patients doing IVF have finished all their frozen embryos; therefore, we consider this time point for analyzing the cumulative multiple delivery rate. Cumulative multiple delivery is total case multiple delivery per total patient to be enrolled into the study at 12 months after randomization | 12 months after randomization | 
| Hypertensive disorders of pregnancy | Pregnancy-induced hypertension, pre-eclampsia and eclampsia | At 20 weeks of gestation or beyond after the completion of the first transfer | 
| Gestational diabetes mellitus | using a 75g oral glucose tolerance test | At 24 to 28 weeks of gestation | 
| Cumulative live birth rate | Cumulative live birth rate at 12 months after the randomization. | At 24 weeks of gestation | 
| Birth weight | Weight of singletons and twins | At the time of delivery | 
| Low birth weight | Weight < 2500 gm at birth | At birth | 
| Major congenital abnormalities | Structural, functional, and genetic anomalies, that occur during pregnancy, and identified antenatally, at birth, or later in life, and require surgical repair of a defect, or are visually evident, or are life-threatening, or cause death. Any congenital anomaly will be included as followed definition of congenital abnormalities in Surveillance of Congenital Anomalies by Division of Birth Defects and Developmental Disabilities, NCBDDD, Centers for Disease Control and Prevention (2020). | At birth | 
| High birth weight | Weight >4000 gm at birth | At birth | 
| Very low birth weight | Weight < 1500 gm at birth | At birth | 
| Large for gestational age | defined as birth weight >90th centile for gestation, based on standardised ethnicity based charts | At birth | 
| small for gestational age | defined as less than 10th centile for gestational age at delivery based on standardised ethnicity based charts | At birth | 
| Very high birth weight | Weight >4500 gm at birth | At birth | 
| Antepartum haemorrhage | including placenta previa, placenta accreta and unexplained | At birth | 
| Perinatal mortality | the death of a fetus or infant from 24 weeks of gestation to the end of the neonatal period of 4 weeks after birth. | 24 weeks of gestation to the end of the neonatal period of 4 weeks after birth | 
| Admission to NICU | The admittance of the newborn to NICU | At birth | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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POSEIDON, in vitro fertilization, live birth, low prognosis, AOA