Time-Lapse Incubation for Embryo Culture - Morphokinetics and Environmental Stability

Infertility Clinical Trial

Official title:

Time-Lapse Incubation for Embryo Culture - The Impact of Morphokinetics and Environmental Stability on Reproductive Outcomes

Clinical Trial Details — Status: Recruiting

Administrative data

• NCT number: NCT02657811
• Other study ID #: 0169-15-SZMC
• Status: Recruiting
• Phase: N/A
• First received: January 5, 2016
• Last updated: February 19, 2017
• Start date: October 2016
• Est. completion date: August 2018

Study information

• Verified date: February 2017
• Source: Shaare Zedek Medical Center
• Contact: Ruth Ronn, M.D. C.M.
• Phone: 972-2-655-5111
• Email: RuthRonn[@]szmc.org.il
• Is FDA regulated: No
• Study type: Interventional

Clinical Trial Summary

Embryo culture and selection has been a continuous challenge in evolution since the birth of In Vitro Fertilization (IVF). Traditionally, embryo quality and its presumed suitability for transfer were assessed based on morphologic features. However, the consensus as to the optimal time points for embryo assessment and as to 'preferable' characteristics have been challenging. Alongside this has been the challenge of achieving balance between multiple points of assessment, yet stabilizing the embryo environment for growth. In standard incubation, each new morphological assessment of embryos in culture theoretically creates an additional disruption to culture.

Most recently, time-lapse incubators (TLI) have been introduced as a novel embryo culture system attempting to limit culture disturbances. These incubators have been integrated with digital imaging, allowing for a substantial limitation in embryo handling and environmental disturbances. They have also introduced new morphokinetic parameters to embryo assessment and to optimizing selection of embryos. Thus far, a limited number of studies have examined the clinical outcomes and value of time lapse monitoring systems versus the more ubiquitous incubators (e.g. multichamber) for reproductive outcomes. In particular, the isolated value of morphokinetics in embryo assessment and of this new stable culture environment in TLI are still in question.

The objectives of this study are to prospectively assess and compare fertility outcomes when embryos are cultured in the TLI system versus more traditional bench incubators (BI). We will specifically assess the added value of the closed and isolated TLI compared to BI on reproductive outcomes, as well as the value of morphokinetic grading in IVF.

Description:

Research Objectives:

We are now reaching a new age with an opportunity to advance to theoretically better culture environments and improvements on measures of embryo assessment.

The objectives of this research study will be to evaluate and compare the reproductive outcomes in isolating one of two new interventions introduced by the Time-Lapse Incubator (TLI) system. The first will be an assessment of the TLI environment compared to the standard Bench Incubator (BI) environment. The second will be an assessment of the added morphokinetic grading of embryos compared to the traditional morphologic grading alone.

Although the primary objective of this study will focus on clinical pregnancy rates and fresh embryo transfers, further research using embryos frozen from this study will be conducted to also evaluate cumulative pregnancy rates per oocyte pick-up (OPU) cycle in the future.

Methods The current study will be a prospective randomized and double-blinded study using three patient arms. The first arm will include patients randomized to embryo culture in a tri-gas bench incubator (Miri, Benchtop Multi-room incubator). These embryos will undergo multiple evaluations using light microscopy and traditional morphologic assessment according to accepted criteria. The second arm will include patients randomized to embryo culture in a time-lapse incubator (Miri TL, Time-Lapse Incubator). These embryos will remain in the TLI and undergo both morphologic and morphokinetic evaluation and grading according to a multivariable scoring model. They will not be removed from incubation for the duration of culture. The third arm will include patients also randomized to embryo culture in a time-lapse incubator (Miri TL, Time-Lapse Incubator). These embryos will remain in the TLI and undergo only traditional morphologic assessment according to accepted criteria with no additional imaging. They will also not be removed from incubation for the duration of culture. The time points and evaluated parameters will be identical to those in arm 1 of the study.

Patients will be assessed for suitability, inclusion and exclusion criteria by the physician and nursing team prior to initiation of an IVF/ICSI cycle at our centre. Once the patient is deemed eligible, a member of the care team will discuss details of the study with the patient.

Approved study subjects will undergo standard controlled ovarian hyperstimulation (COH). Protocols and their corresponding medications will be decided upon at the discretion of the treating physician. These may include long gonadotropin-releasing hormone (GnRH) agonist protocol and GnRH antagonist protocol. Follicular aspiration will be performed in the IVF unit via transvaginal needle aspiration. Endometrial preparation and luteal phase support will be recommended as per our departmental protocol. The number of embryos for transfer will be defined prior to cycle initiation, according to The Israel Society of Obstetrics & Gynecology.

Patient randomization will occur after Human Chorionic Gonadotropin (hCG) administration has already been ensured, prior to Intracytoplasmic Sperm Injection (ICSI). The randomization procedure will be accomplished using computer-generated randomization.

After follicular aspiration and transfer to the laboratory, the follicular fluid will be examined for presence of oocytes. Oocytes will then be scored for maturity and any morphologically abnormal features. ICSI will then be performed using fresh sperm from the corresponding partner.

Embryos will then be placed in culture media. The embryos within culture media will then be placed in either the Miri Benchtop or Miri Timelapse incubator according to their corresponding group of randomization. Note that the environment of the Miri benchtop and Miri TL incubators will be considered identical. oxygen concentration and carbon dioxide concentrations set at 5% and 5.5% respectively.

Embryos in arm 1 of the study (Miri Benchtop incubator) will be morphologically assessed by light microscopy at pre-defined time points and according to accepted criteria by one of the trained laboratory embryologists. The time points for evaluation will be at 16 - 20 hours post-ICSI (for normal fertilization), at 44 - 48 hours (day 2) post-ICSI, and then at 64 - 72 hours (day 3) post-ICSI. Evaluated parameters will include cell number, cell size, cell symmetry, and percent fragmentation. Any gross abnormalities will be noted. Embryos will then be graded on day 3 according to these evaluated parameters and transferred according to preferential grading. If embryo transfer has been predetermined for day 5, additional assessment will take place at approximately 116 hours.

Embryos in arm 2 of the study (Miri TL incubator) will be morphologically and morphokinetically assessed by the same laboratory embryologists using digital images generated by the incubator's integrated time-lapse imaging system. Assessment and scoring will be performed as per our scoring classification system. Morphological screening of embryos will initially be performed in order to discard or exclude those clearly not viable for transfer. Morphokinetic parameters will then be used in order to rank remaining embryos score categories from a maximum of 4.0 to a minimum of -2.0, in order of hypothesized decreasing implantation potential.

Embryos in arm 3 of the study (Miri TL incubator) will be morphologically assessed by the embryologists using digital images generated by the incubator's integrated time-lapse imaging system. As noted above, the time points and evaluated parameters will be identical to those in arm 1 of the study. Decisions on embryos for transfer will also be identical.

For each patient, embryos will be selected for transfer based on their morphologic scoring alone (Arm 1 and 3) or by the morphokinetic decision tree scoring (Arm 2). The number of embryos for transfer will have been pre-determined (as noted above).

Those embryos not selected and deemed appropriate for future transfer will undergo cryopreservation by vitrification.

Blinding of the current study will be ensured at multiple points. The gynecologists performing oocyte retrieval and embryo transfer will be blinded to the predefined and randomized patient group. Patients will be unaware of their group of randomization. Statisticians will also be blinded to the group of randomization in calculating pregnancy outcomes. It will unfortunately not be possible to ensure blinding of those embryologists performing the morphologic and morphokinetic assessments.

Sample Size calculation:

The pregnancy rate in our IVF Unit in the sub-group of patients with demographic and clinical characteristics similar to those included in the study, is about 40%. Assuming that in the TLI group pregnancy rate will increase by 10%, the sample size required per group is 124 patients per arm, with an alpha risk of 5% and a power of 80%.

Statistics:

The statistical analysis will be done by intention to treat.

Recruitment information / eligibility

• Status: Recruiting
• Enrollment: 372
• Est. completion date: August 2018
• Est. primary completion date: December 2017
• Accepts healthy volunteers: No
• Gender: Female
• Age group: 18 Years to 35 Years

Eligibility

Inclusion Criteria:

• Patients requiring assisted reproductive technologies for one or more of the following reasons:

• Male factor infertility

• Unexplained infertility

• Mechanical factor infertility

• Ovulatory infertility

• Patients undergoing fertility treatment at Shaare Zedek Medical Centre alone.

• Patients attempting pregnancy with autologous gametes.

• Patients receiving embryo transfers according to the Israel Society of Obstetrics and Gynecology guideline on number of embryos for transfer during in vitro fertilization.

• Patients undergoing their first or second ICSI cycle (cumulative to all other institutions involved in prior treatment) since their previous pregnancy.

• BMI criteria: >18 and <30 kg/m2

Exclusion Criteria:

• An infertility diagnosis that includes the following:

• Severe male factor infertility requiring testicular aspiration, testicular biopsy for sperm retrieval, or less than 1000 sperm per ejaculate.

• Untreated hydrosalpinx

• Persistently thin endometrial lining or endometrial factor

• Severe Endometriosis

• Low ovarian reserve (=8 antral follicle count (AFC) follicles measuring 2 to 10 mm in diameter on day 2-4 of menstrual cycle, or day 3 follicle stimulating hormone (FSH) =10 milli-International unit (mIU) /mL)

• High risk for Ovarian Hyperstimulation Syndrome (Estradiol level >13 000 pmol/L or >20 follicles =16 mm diameter during controlled ovarian hyper stimulation, =20 oocytes collected).

• Patients obtaining GnRH agonist for final follicular maturation

• Patients requiring preimplantation genetic diagnosis (PGD)

• Patients who require freezing of all oocytes or embryos

• More than 2 previous cycles of IVF since previous pregnancy

• Current smokers

Related Conditions & MeSH terms

• Genetic Diseases
• Infertility

Intervention

Device:
• Miri, Benchtop Multi-room incubator
Bench Incubation - Morphologic Assessment These embryos will undergo multiple evaluations using light microscopy and traditional morphologic assessment.
• Miri TL, Time-Lapse incubator
Time Lapse Incubation - Morphologic/Morphokinetic Assessment These embryos will remain in the TLI and undergo both morphologic and morphokinetic evaluation and grading according to a hierarchical multivariable model. They will not be removed from incubation for the duration of culture.
• Miri TL, Time-Lapse incubator
Time Lapse Incubation - Morphologic Assessment These embryos will remain in the TLI and undergo only traditional morphologic assessment according to accepted criteria with no additional imaging. They will also not be removed from incubation for the duration of culture. The time points and evaluated parameters will be identical to those in arm 1 of the study.

Locations

Country	Name	City	State
Israel	Shaare Zedek Medical Center	Jerusalem

Sponsors (1)

Lead Sponsor	Collaborator
Shaare Zedek Medical Center

Country where clinical trial is conducted

Israel, 

References & Publications (26)

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Armstrong S, Arroll N, Cree LM, Jordan V, Farquhar C. Time-lapse systems for embryo incubation and assessment in assisted reproduction. Cochrane Database Syst Rev. 2015 Feb 27;(2):CD011320. doi: 10.1002/14651858.CD011320.pub2. Review. — View Citation

Azzarello A, Hoest T, Mikkelsen AL. The impact of pronuclei morphology and dynamicity on live birth outcome after time-lapse culture. Hum Reprod. 2012 Sep;27(9):2649-57. doi: 10.1093/humrep/des210. — View Citation

Basile N, Morbeck D, García-Velasco J, Bronet F, Meseguer M. Type of culture media does not affect embryo kinetics: a time-lapse analysis of sibling oocytes. Hum Reprod. 2013 Mar;28(3):634-41. doi: 10.1093/humrep/des462. — View Citation

Ciray HN, Aksoy T, Goktas C, Ozturk B, Bahceci M. Time-lapse evaluation of human embryo development in single versus sequential culture media--a sibling oocyte study. J Assist Reprod Genet. 2012 Sep;29(9):891-900. doi: 10.1007/s10815-012-9818-7. — View Citation

Conaghan J, Chen AA, Willman SP, Ivani K, Chenette PE, Boostanfar R, Baker VL, Adamson GD, Abusief ME, Gvakharia M, Loewke KE, Shen S. Improving embryo selection using a computer-automated time-lapse image analysis test plus day 3 morphology: results from a prospective multicenter trial. Fertil Steril. 2013 Aug;100(2):412-9.e5. doi: 10.1016/j.fertnstert.2013.04.021. — View Citation

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Dal Canto M, Coticchio G, Mignini Renzini M, De Ponti E, Novara PV, Brambillasca F, Comi R, Fadini R. Cleavage kinetics analysis of human embryos predicts development to blastocyst and implantation. Reprod Biomed Online. 2012 Nov;25(5):474-80. doi: 10.1016/j.rbmo.2012.07.016. — View Citation

Fujiwara M, Takahashi K, Izuno M, Duan YR, Kazono M, Kimura F, Noda Y. Effect of micro-environment maintenance on embryo culture after in-vitro fertilization: comparison of top-load mini incubator and conventional front-load incubator. J Assist Reprod Genet. 2007 Jan;24(1):5-9. — View Citation

Goodman LR, Goldberg J, Falcone T, Austin C, Desai N. Does the addition of time-lapse morphokinetics in the selection of embryos for transfer improve pregnancy rates? A randomized controlled trial. Fertil Steril. 2016 Feb;105(2):275-85.e10. doi: 10.1016/j.fertnstert.2015.10.013. — View Citation

Hashimoto S, Kato N, Saeki K, Morimoto Y. Selection of high-potential embryos by culture in poly(dimethylsiloxane) microwells and time-lapse imaging. Fertil Steril. 2012 Feb;97(2):332-7. doi: 10.1016/j.fertnstert.2011.11.042. — View Citation

Hlinka D, Kalatová B, Uhrinová I, Dolinská S, Rutarová J, Rezácová J, Lazarovská S, Dudáš M. Time-lapse cleavage rating predicts human embryo viability. Physiol Res. 2012;61(5):513-25. — View Citation

Kirkegaard K, Ahlström A, Ingerslev HJ, Hardarson T. Choosing the best embryo by time lapse versus standard morphology. Fertil Steril. 2015 Feb;103(2):323-32. doi: 10.1016/j.fertnstert.2014.11.003. Review. — View Citation

Lemmen JG, Agerholm I, Ziebe S. Kinetic markers of human embryo quality using time-lapse recordings of IVF/ICSI-fertilized oocytes. Reprod Biomed Online. 2008 Sep;17(3):385-91. — View Citation

Meseguer M, Herrero J, Tejera A, Hilligsøe KM, Ramsing NB, Remohí J. The use of morphokinetics as a predictor of embryo implantation. Hum Reprod. 2011 Oct;26(10):2658-71. doi: 10.1093/humrep/der256. — View Citation

Meseguer M, Rubio I, Cruz M, Basile N, Marcos J, Requena A. Embryo incubation and selection in a time-lapse monitoring system improves pregnancy outcome compared with a standard incubator: a retrospective cohort study. Fertil Steril. 2012 Dec;98(6):1481-9.e10. doi: 10.1016/j.fertnstert.2012.08.016. — View Citation

Morbeck DE, Krisher RL, Herrick JR, Baumann NA, Matern D, Moyer T. Composition of commercial media used for human embryo culture. Fertil Steril. 2014 Sep;102(3):759-766.e9. doi: 10.1016/j.fertnstert.2014.05.043. — View Citation

Practice Committee of American Society for Reproductive Medicine.; Practice Committee of Society for Assisted Reproductive Technology.. Criteria for number of embryos to transfer: a committee opinion. Fertil Steril. 2013 Jan;99(1):44-6. doi: 10.1016/j.fertnstert.2012.09.038. — View Citation

Rubio I, Galán A, Larreategui Z, Ayerdi F, Bellver J, Herrero J, Meseguer M. Clinical validation of embryo culture and selection by morphokinetic analysis: a randomized, controlled trial of the EmbryoScope. Fertil Steril. 2014 Nov;102(5):1287-1294.e5. doi: 10.1016/j.fertnstert.2014.07.738. — View Citation

Rubio I, Kuhlmann R, Agerholm I, Kirk J, Herrero J, Escribá MJ, Bellver J, Meseguer M. Limited implantation success of direct-cleaved human zygotes: a time-lapse study. Fertil Steril. 2012 Dec;98(6):1458-63. doi: 10.1016/j.fertnstert.2012.07.1135. — View Citation

Swain JE. Decisions for the IVF laboratory: comparative analysis of embryo culture incubators. Reprod Biomed Online. 2014 May;28(5):535-47. doi: 10.1016/j.rbmo.2014.01.004. Review. — View Citation

VerMilyea MD, Tan L, Anthony JT, Conaghan J, Ivani K, Gvakharia M, Boostanfar R, Baker VL, Suraj V, Chen AA, Mainigi M, Coutifaris C, Shen S. Computer-automated time-lapse analysis results correlate with embryo implantation and clinical pregnancy: a blinded, multi-centre study. Reprod Biomed Online. 2014 Dec;29(6):729-36. doi: 10.1016/j.rbmo.2014.09.005. — View Citation

Wong CC, Loewke KE, Bossert NL, Behr B, De Jonge CJ, Baer TM, Reijo Pera RA. Non-invasive imaging of human embryos before embryonic genome activation predicts development to the blastocyst stage. Nat Biotechnol. 2010 Oct;28(10):1115-21. doi: 10.1038/nbt.1686. — View Citation

Yang Z, Zhang J, Salem SA, Liu X, Kuang Y, Salem RD, Liu J. Selection of competent blastocysts for transfer by combining time-lapse monitoring and array CGH testing for patients undergoing preimplantation genetic screening: a prospective study with sibling oocytes. BMC Med Genomics. 2014 Jun 22;7:38. doi: 10.1186/1755-8794-7-38. — View Citation

Zhang JQ, Li XL, Peng Y, Guo X, Heng BC, Tong GQ. Reduction in exposure of human embryos outside the incubator enhances embryo quality and blastulation rate. Reprod Biomed Online. 2010 Apr;20(4):510-5. doi: 10.1016/j.rbmo.2009.12.027. — View Citation

* Note: There are 26 references in all — Click here to view all references

Outcome

Type	Measure	Description	Time frame	Safety issue
Primary	Ongoing pregnancy rate	The number of women with a viable pregnancy (including gestational sac and fetal heart beat) at 12 weeks gestation divided by the number of women having an embryo transfer.	12 weeks gestation
Secondary	Embryo morphology - Number of pronuclei present following fertilization	Number of pronuclei present in the embryo 1 day following fertilization. Morphologic assessment parameter used to grade embryo quality.	Day 1 post-fertilization
Secondary	Embryo morphology - Number of cells present	Number of cells present in the embryo 2 and 3 days following fertilization. Morphologic assessment parameter used to grade embryo quality.	Day 2 and Day 3 post-fertilization
Secondary	Embryo morphology - % fragmentation	Percent fragmentation in the embryo 2 and 3 days following fertilization. Morphologic assessment parameter used to grade embryo quality.	Day 2 and Day 3 post-fertilization
Secondary	Embryo morphology - Presence of multi nucleation	Presence of multi-nucleation in the embryo on day 2 following fertilization. Morphologic assessment parameter used to grade embryo quality.	Day 2 post-fertilization
Secondary	Embryo morphology - Symmetry	Symmetry in the embryo 2 and 3 days following fertilization. Morphologic assessment parameter used to grade embryo quality.	Day 2 and Day 3 post-fertilization
Secondary	Embryo morphology - Stage of development at day 5	Stage of development in the embryo 5 days following fertilization. Characterized as: Early blastocyst, Blastocyst, Expanded, Hatched/Hatching, or other. Morphologic assessment parameter used to grade embryo quality.	Day 5 post-fertilization
Secondary	Embryo morphology - Inner Cell Mass (ICM) grade	ICM grade in the embryo 5 days following fertilization. Morphologic assessment parameter used to grade embryo quality.	Day 5 post-fertilization
Secondary	Embryo morphology - Trophectoderm grade	Trophectoderm grade in the embryo 5 days following fertilization. Morphologic assessment parameter used to grade embryo quality.	Day 5 post-fertilization
Secondary	Embryo morphokinetics - Abrupt division	Presence or absence of abrupt division of the embryo from one to three or more cells: a morphokinetic assessment parameter used to grade embryo quality.	Day 0 to Day 3 post-fertilization
Secondary	Embryo morphokinetics - Time of cleavage to five-blastomere embryo	Presence or absence of optimal time range for cleavage to five-blastomere embryo (48.8-56.6 hours): a morphokinetic assessment parameter used to grade embryo quality.	Day 0 to Day 3 post-fertilization
Secondary	Embryo morphokinetics - Time from two to four-blastomere embryo	Presence or absence of optimal time duration of division from 2 to 3 cells and subsequent division to 4 cells (0-0.76 hours): a morphokinetic assessment parameter used to grade embryo quality.	Day 1 to Day 3 post-fertilization
Secondary	Embryo morphokinetics - Time from two to three-blastomere embryo	Presence or absence of optimal time duration of division from two-cell to three-cell embryo (0-11.9 hours): a morphokinetic assessment parameter used to grade embryo quality.	Day 1 to Day 3 post-fertilization
Secondary	Oocyte characteristics - Polar body presence	Polar body presence of oocyte on day of oocyte retrieval	Oocyte retrieval until fertilization - less than 1 day
Secondary	Biochemical pregnancy rate	The number of women with a positive ß-hCG approximately 14 days after embryo transfer divided by the number of women having an embryo transfer.	14 days after embryo transfer
Secondary	Cumulative ongoing pregnancy rate	Cumulative number of women with a viable pregnancy at 12 weeks (where pregnancy was obtained with fresh and vitrified embryos from the same ovarian stimulation cycle) divided by the number of women having undergone an oocyte pickup cycle.	One year from oocyte pickup
Secondary	Live birth rate	The percentage of all Oocyte Pick Up (OPU) cycles that lead to live birth Approximate time frame will be time of one ovarian stimulation cycle plus term pregnancy	One year and forty weeks
Secondary	Spontaneous abortion rate	Loss of an embryo or fetus prior to 20 weeks gestation or weighing less than 500 grams.	First 20 weeks of gestation

External Links

•   Practice Guideline #201- Number of embryos for transfer during in vitro fertilization. Israel Society of Obstetrics and Gynecology. 2010 May 23;:1-1