Ignite Creation Date:
2024-05-06 @ 1:02 PM
Last Modification Date:
2024-10-26 @ 1:08 PM
Study NCT ID:
NCT03915054
Status:
COMPLETED
Last Update Posted:
2021-06-16
First Post:
2019-04-03
Brief Title:
Amphiregulin Versus Non-Amphiregulin Supplementation to Maturation Culturing Medium in IVM
Sponsor:
Mỹ Đức Hospital
Organization:
Mỹ Đức Hospital
Study Overview
Official Title:
Effect of the CAPA Culture Step on Meiotic and Developmental Competence of Human Oocytes After Using Two Previously Used Meiotic Maturation Triggers in a SIBLING Oocyte Study Design
Status:
COMPLETED
Status Verified Date:
2021-06
Last Known Status:
None
Delayed Posting:
No
If Stopped, Why?:
Not Stopped
Has Expanded Access:
False
If Expanded Access, NCT#:
N/A
Has Expanded Access, NCT# Status:
N/A
Acronym:
None
Brief Summary:
Clinical use of IVM was pioneered in the nineties but has not yet become a realistic option for wide-scale practice for several reasons Fundamentally despite recent progress in improving the implantation and the pregnancy rates using in-vitro matured oocytes results of IVM remain lower than treatment cycles utilizing conventional ART To improve the outcome of IVM cycles this study focuses on improving in-vitro culture conditions
In-vitro maturation IVM of human oocytes obtained from minimally stimulated or unstimulated ovaries offers a more patient friendly treatment option than the conventional Assisted Reproductive Technology ART treatment with controlled ovarian hyperstimulation COH Typically IVM will be offered to women with polycystic ovaries PCOPCOS or to patients with an excellent ovarian reserve ie a high antral follicle count IVM treatment is characterized by minimal administration of FSH or hMG and NO hCG trigger The IVM approach is less disruptive to patients daily life through the reduced need for hormonal and ultrasound monitoring avoids a range of minor and major complications such as ovarian hyperstimulation syndrome and aims to reduce the total cost of infertility treatment for the patient and for the health care budget
Human oocytes retrieved from small antral follicles are able to resume meiosis by undergoing germinal vesicle breakdown and extrusion of the first polar body if oocytes have reached meiotic competence These oocytes can be fertilized although only a proportion less than 50 of them can develop further into viable embryos It has been hypothesized that failure of embryonic development may at least in part be due to an immature oocyte cytoplasm A novel human in vitro maturation IVM culture system named CAPACITATION-IVM is being investigated hereafter named CAPA using 1 natural compounds known to influence cAMP levels within the cumulus-oocyte-complex and 2 compounds that are crucial for the oocyte-cumulus cross-talk Keeping cyclic AMP high after retrieval in the GV oocyte prevents the occurrence of nuclear maturation enabling increased communication between the oocyte and the cumulus cells This allows for the improvement in the synchronization of nuclear and cytoplasmic maturation processes in the oocyte to the benefit of embryo quality
In-vitro maturation IVM of human oocytes obtained from minimally stimulated or unstimulated ovaries offers a more patient friendly treatment option than the conventional Assisted Reproductive Technology ART treatment with controlled ovarian hyperstimulation COH Typically IVM will be offered to women with polycystic ovaries PCOPCOS or to patients with an excellent ovarian reserve ie a high antral follicle count IVM treatment is characterized by minimal administration of FSH or hMG and NO hCG trigger The IVM approach is less disruptive to patients daily life through the reduced need for hormonal and ultrasound monitoring avoids a range of minor and major complications such as ovarian hyperstimulation syndrome and aims to reduce the total cost of infertility treatment for the patient and for the health care budget
Human oocytes retrieved from small antral follicles are able to resume meiosis by undergoing germinal vesicle breakdown and extrusion of the first polar body if oocytes have reached meiotic competence These oocytes can be fertilized although only a proportion less than 50 of them can develop further into viable embryos It has been hypothesized that failure of embryonic development may at least in part be due to an immature oocyte cytoplasm A novel human in vitro maturation IVM culture system named CAPACITATION-IVM is being investigated hereafter named CAPA using 1 natural compounds known to influence cAMP levels within the cumulus-oocyte-complex and 2 compounds that are crucial for the oocyte-cumulus cross-talk Keeping cyclic AMP high after retrieval in the GV oocyte prevents the occurrence of nuclear maturation enabling increased communication between the oocyte and the cumulus cells This allows for the improvement in the synchronization of nuclear and cytoplasmic maturation processes in the oocyte to the benefit of embryo quality
Detailed Description:
There are two types of patients can be distinguished in this study
Polycystic ovarian morphology normal cycle length up to 35 days The majority of subjects in this group have non-syndromic polycystic ovaries and do not have PCOS In these patients AMH is only moderately elevated and cyclical follicular development occurs under the influence of FSH
PCOS - Polycystic ovarian morphology oligomenorrhoea menstrual periods occurring at intervals of greater than 35 days with only four to nine periods in a year or amenorrhea In these patients with often strongly elevated AMH levels and concomitant hyperandrogenemia relative FSH resistance results in arrest of follicular growth at the small antral follicular state Spontaneous selection of a dominant follicle does not occur or occurs rarely A large proportion of follicles are atretic An important proportion of these patients have a BMI higher than 25 These patients generally receive the same standard gonadotropin stimulation regimen as the patients of the previous group but if the largest follicles have a diameter of less than 8 mm on the third day of stimulation patients receive one or two supplementary days of gonadotropin stimulation Moreover these patients generally take two-three weeks of OCP before the start of the IVM cycle and start stimulation approximately on day five after OCP withdrawal However since these patients do not have endogenous FSH-driven recruitment of follicles the start of stimulation is rather flexible
The use of OCP oral contraceptive pill before the IVM cycle is obligate OCP for example Microgynon 30 micrograms will be administered daily to clear the atretic follicles in the ovary OCP will be given for between 14 and 21 days this method allows for better programming the cycles and the workload in the embryology lab
Before the first IVM cycle Screening visit All subjects will undergo a pelvic ultrasound scan to evaluate suitability to undergo IVM treatment Patients will undergo a blood test for serology Hepatitis B HIV syphilis and baseline hormonal profiling LH FSH E2 progesterone AMH SHBG Testosterone Additional analysis of TSH thyroperoxidase antibodies prolactin Any concomitant medication taken in the last 3 months prior to the IVM attempt should be notified
First IVM treatment cycle
First clinic visit
All subjects will contact the study nurse on cycle day cd1 to initiate the first Capacitation culture cycle If cd1 is during the weekend subjects will contact the study nurse on Monday The subject will attend the clinic on day 1 2 or 3 of menstrual bleeding An ultrasound scan is performed to rule out the existence of ovarian cysts and a blood sample is taken for hormonal assessment HCG FSH LH E2 progesterone On the evening of cycle day 3 of the first clinic visit flexible Gonadotrophin stimulation will be started to enhance follicular development The first dose of HP-hMG 150 IU at 2 PM Second dose next day of HP-hMG 150 IU at 2 PM The subject will return for a pelvic ultrasound scan and a blood test FSH LH E2 progesterone after the two days of HP-hMG 150 IU stimulation ie on the morning of day 6
To patients with OCP bleeding On day one two three or four of the withdrawal bleeding that occurs after discontinuing the OCP the subject will attend the clinic to undergo a pelvic ultrasound scan and a blood sample is taken for hormonal assessment FSH LH E2 progesterone AMH total serum testosterone SHBG On the day of the first clinic visit flexible HP-hMG stimulation will be started to enhance follicular development at a first daily dose of HP-hMG at 8 PM The subject will return for a pelvic ultrasound scan and a blood test FSH LH E2 progesterone after two days of HP hMG stimulation
Second clinic visit at day 6 after 2 days of HP-hMG treatment
If the ultrasound scan shows the presence of a single dominant follicle larger than 12 mm with the other follicles all less than 10 mm and the oocyte retrieval OR will be scheduled two days later between 800 and 900 h It is important to keep the OR 42 - 46 hours after the last HP-hMG injection as a fixed timing If the ultrasound scan shows a maximal follicular diameter of less than 8 mm a final injection of HP-hMG will be administered at 2 PM and the oocyte retrieval OR will be scheduled two days later between 800 and 900 h It is important to keep the OR 42 - 46 hours after the last HP-hMG injection as a fixed timing
To patients with OCP bleeding If the ultrasound scan shows a maximal follicular diameter of approx 9 mm a final injection of HP-hMG will be administered at 2 PM and the oocyte retrieval OR will be scheduled two days later between 800 and 900 h under general anesthetics - the exact time of OR should be discussed with operating nurses and anesthetist It is important to keep the time of OR constant at 42 -46 hours after the last HP-hMG injection
Third visit - Oocyte retrieval Oocyte retrieval OR will be scheduled between 800 and 1000 h under general anesthetics On the day of OR
An ultrasound scan of both ovaries and of endometrial lining is recorded
A blood sample will be obtained for hormonal profiling LH FSH E2 progesterone
All cumulus-oocyte complexes COC retrieved from the patient are cultured in CAPA medium for 24 hours
Following CAPA culture half of oocytes will be divided randomly 5050 to the two maturation triggers medium
Group 1 Medicult IVM media AREGFSHHSAInsulinEstradiol AREG-TRIGGER group
Group 2 Medicult IVM media FSHGHhCGHSA CONTROL-TRIGGER group with no AREG added
In vitro oocyte maturation
Evaluate and register COCs for cumulus mass CM and cumulus contact between oocyte and cumulus cells Each time take photographs Discard fully denuded degenerated oocytes
Transfer half of the COCs 5-10 at a time from the Capacitation culture dish to the washing dish containing Maturation Medium 1 AREG-TRIGGER medium by using an Eppendorf micropipette and wash them thoroughly load pipette tips with 5µl max 10µl Then transfer COCs to the IVM dish with Maturation Medium 1 AREG-TRIGGER medium
Transfer the second half of the COCs from the Capacitation culture dish to the washing dish containing Maturation Medium 2 CONTROL-TRIGGER medium and wash them thoroughly load pipette tips with 5µl max 10µl Then transfer COCs to the IVM dish with Maturation Medium 2 CONTROL-TRIGGER medium
Keep COCs in pools of 10
Label the dish appropriately and culture COCs 30-32 hours in an incubator
Evaluation of maturation MII GVBD GV will be done at 30 hours Oocytes which have undergone GVBD but with no clear PB will be assessed at 32 hours Insemination will be performed using intra-cytoplasmic sperm injection 3-4 hours after oocyte retrieval or maturation check only matured oocytes will be inseminated
Fertilization check will be performed under an inverted microscope at 16-18 hours after insemination Embryo evaluation will be performed at 68 1 hours after fertilization using the Istanbul consensus Standard Embryo Vitrification protocol used Embryos will be vitrified per stimulation protocol obtained group 1 AREG-TRIGGER or group 2 - CONTROL-TRIGGER
Frozen embryo transfer The patient will be randomized to receive embryo randomly from AREG-TRIGGER or CONTROL-TRIGGER Where no embryos from the randomized group were available embryos from the other group were transferred
The first pregnancy test was performed 14 days after embryo transfer a positive pregnancy test was defined as serum beta hCG 5 mIUmL
Clinical pregnancy was defined as at least one gestational sac on ultrasound at 7 weeks gestation with the detection of heartbeat activity
Ongoing pregnancy was defined as pregnancy with a detectable heart rate at 12 weeks gestation after the completion of the first transfer
Live birth was defined as the birth of at least one newborn after 24 weeks gestation exhibiting any sign of life twins were a single count
After a first unsuccessful IVM cycle The clinical and embryological data and results related to the first IVM cycle cumulative fresh and frozen embryo cycles will be discussed to establish a more patient-tailored approach for an eventual second IVM cycle The patient-tailored approach in the second IVM cycle might consist of modification of the management of the follicular phase
Polycystic ovarian morphology normal cycle length up to 35 days The majority of subjects in this group have non-syndromic polycystic ovaries and do not have PCOS In these patients AMH is only moderately elevated and cyclical follicular development occurs under the influence of FSH
PCOS - Polycystic ovarian morphology oligomenorrhoea menstrual periods occurring at intervals of greater than 35 days with only four to nine periods in a year or amenorrhea In these patients with often strongly elevated AMH levels and concomitant hyperandrogenemia relative FSH resistance results in arrest of follicular growth at the small antral follicular state Spontaneous selection of a dominant follicle does not occur or occurs rarely A large proportion of follicles are atretic An important proportion of these patients have a BMI higher than 25 These patients generally receive the same standard gonadotropin stimulation regimen as the patients of the previous group but if the largest follicles have a diameter of less than 8 mm on the third day of stimulation patients receive one or two supplementary days of gonadotropin stimulation Moreover these patients generally take two-three weeks of OCP before the start of the IVM cycle and start stimulation approximately on day five after OCP withdrawal However since these patients do not have endogenous FSH-driven recruitment of follicles the start of stimulation is rather flexible
The use of OCP oral contraceptive pill before the IVM cycle is obligate OCP for example Microgynon 30 micrograms will be administered daily to clear the atretic follicles in the ovary OCP will be given for between 14 and 21 days this method allows for better programming the cycles and the workload in the embryology lab
Before the first IVM cycle Screening visit All subjects will undergo a pelvic ultrasound scan to evaluate suitability to undergo IVM treatment Patients will undergo a blood test for serology Hepatitis B HIV syphilis and baseline hormonal profiling LH FSH E2 progesterone AMH SHBG Testosterone Additional analysis of TSH thyroperoxidase antibodies prolactin Any concomitant medication taken in the last 3 months prior to the IVM attempt should be notified
First IVM treatment cycle
First clinic visit
All subjects will contact the study nurse on cycle day cd1 to initiate the first Capacitation culture cycle If cd1 is during the weekend subjects will contact the study nurse on Monday The subject will attend the clinic on day 1 2 or 3 of menstrual bleeding An ultrasound scan is performed to rule out the existence of ovarian cysts and a blood sample is taken for hormonal assessment HCG FSH LH E2 progesterone On the evening of cycle day 3 of the first clinic visit flexible Gonadotrophin stimulation will be started to enhance follicular development The first dose of HP-hMG 150 IU at 2 PM Second dose next day of HP-hMG 150 IU at 2 PM The subject will return for a pelvic ultrasound scan and a blood test FSH LH E2 progesterone after the two days of HP-hMG 150 IU stimulation ie on the morning of day 6
To patients with OCP bleeding On day one two three or four of the withdrawal bleeding that occurs after discontinuing the OCP the subject will attend the clinic to undergo a pelvic ultrasound scan and a blood sample is taken for hormonal assessment FSH LH E2 progesterone AMH total serum testosterone SHBG On the day of the first clinic visit flexible HP-hMG stimulation will be started to enhance follicular development at a first daily dose of HP-hMG at 8 PM The subject will return for a pelvic ultrasound scan and a blood test FSH LH E2 progesterone after two days of HP hMG stimulation
Second clinic visit at day 6 after 2 days of HP-hMG treatment
If the ultrasound scan shows the presence of a single dominant follicle larger than 12 mm with the other follicles all less than 10 mm and the oocyte retrieval OR will be scheduled two days later between 800 and 900 h It is important to keep the OR 42 - 46 hours after the last HP-hMG injection as a fixed timing If the ultrasound scan shows a maximal follicular diameter of less than 8 mm a final injection of HP-hMG will be administered at 2 PM and the oocyte retrieval OR will be scheduled two days later between 800 and 900 h It is important to keep the OR 42 - 46 hours after the last HP-hMG injection as a fixed timing
To patients with OCP bleeding If the ultrasound scan shows a maximal follicular diameter of approx 9 mm a final injection of HP-hMG will be administered at 2 PM and the oocyte retrieval OR will be scheduled two days later between 800 and 900 h under general anesthetics - the exact time of OR should be discussed with operating nurses and anesthetist It is important to keep the time of OR constant at 42 -46 hours after the last HP-hMG injection
Third visit - Oocyte retrieval Oocyte retrieval OR will be scheduled between 800 and 1000 h under general anesthetics On the day of OR
An ultrasound scan of both ovaries and of endometrial lining is recorded
A blood sample will be obtained for hormonal profiling LH FSH E2 progesterone
All cumulus-oocyte complexes COC retrieved from the patient are cultured in CAPA medium for 24 hours
Following CAPA culture half of oocytes will be divided randomly 5050 to the two maturation triggers medium
Group 1 Medicult IVM media AREGFSHHSAInsulinEstradiol AREG-TRIGGER group
Group 2 Medicult IVM media FSHGHhCGHSA CONTROL-TRIGGER group with no AREG added
In vitro oocyte maturation
Evaluate and register COCs for cumulus mass CM and cumulus contact between oocyte and cumulus cells Each time take photographs Discard fully denuded degenerated oocytes
Transfer half of the COCs 5-10 at a time from the Capacitation culture dish to the washing dish containing Maturation Medium 1 AREG-TRIGGER medium by using an Eppendorf micropipette and wash them thoroughly load pipette tips with 5µl max 10µl Then transfer COCs to the IVM dish with Maturation Medium 1 AREG-TRIGGER medium
Transfer the second half of the COCs from the Capacitation culture dish to the washing dish containing Maturation Medium 2 CONTROL-TRIGGER medium and wash them thoroughly load pipette tips with 5µl max 10µl Then transfer COCs to the IVM dish with Maturation Medium 2 CONTROL-TRIGGER medium
Keep COCs in pools of 10
Label the dish appropriately and culture COCs 30-32 hours in an incubator
Evaluation of maturation MII GVBD GV will be done at 30 hours Oocytes which have undergone GVBD but with no clear PB will be assessed at 32 hours Insemination will be performed using intra-cytoplasmic sperm injection 3-4 hours after oocyte retrieval or maturation check only matured oocytes will be inseminated
Fertilization check will be performed under an inverted microscope at 16-18 hours after insemination Embryo evaluation will be performed at 68 1 hours after fertilization using the Istanbul consensus Standard Embryo Vitrification protocol used Embryos will be vitrified per stimulation protocol obtained group 1 AREG-TRIGGER or group 2 - CONTROL-TRIGGER
Frozen embryo transfer The patient will be randomized to receive embryo randomly from AREG-TRIGGER or CONTROL-TRIGGER Where no embryos from the randomized group were available embryos from the other group were transferred
The first pregnancy test was performed 14 days after embryo transfer a positive pregnancy test was defined as serum beta hCG 5 mIUmL
Clinical pregnancy was defined as at least one gestational sac on ultrasound at 7 weeks gestation with the detection of heartbeat activity
Ongoing pregnancy was defined as pregnancy with a detectable heart rate at 12 weeks gestation after the completion of the first transfer
Live birth was defined as the birth of at least one newborn after 24 weeks gestation exhibiting any sign of life twins were a single count
After a first unsuccessful IVM cycle The clinical and embryological data and results related to the first IVM cycle cumulative fresh and frozen embryo cycles will be discussed to establish a more patient-tailored approach for an eventual second IVM cycle The patient-tailored approach in the second IVM cycle might consist of modification of the management of the follicular phase
Study Oversight
Has Oversight DMC:
None
Is a FDA Regulated Drug?:
False
Is a FDA Regulated Device?:
False
Is an Unapproved Device?:
None
Is a PPSD?:
None
Is a US Export?:
None
Is an FDA AA801 Violation?:
None