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Trial registered on ANZCTR


Registration number
ACTRN12618001064291
Ethics application status
Approved
Date submitted
15/06/2018
Date registered
26/06/2018
Date last updated
14/01/2019
Date data sharing statement initially provided
14/01/2019
Type of registration
Prospectively registered

Titles & IDs
Public title
A trial for the clinical effectiveness of a Non-invasive Preimplantation Genetic Screening (PGS) method for determining the ploidy status of embryos of patients undergoing In Vitro Fertilisation (IVF) treatment that are not suitable for standard biopsy
Scientific title
NEST4E Non-invasive Preimplantation Genetic Screening (PGS) testing for IVF patient embryos that are not suitable for standard biopsy to determine the proportion of embryos that are unable to be screened using standard PGS that can be conclusively screened for aneuploidy using NEST4E by analysing cell-free DNA in the embryo culture media.
Secondary ID [1] 295192 0
Nil
Universal Trial Number (UTN)
U1111-1215-5976
Trial acronym
NEST4E not for biopsy
Linked study record
ACTRN12617000500358

Health condition
Health condition(s) or problem(s) studied:
Infertility 308328 0
Embryo genetics 308329 0
Condition category
Condition code
Reproductive Health and Childbirth 307329 307329 0 0
Fertility including in vitro fertilisation

Intervention/exposure
Study type
Interventional
Description of intervention(s) / exposure
The purpose of this study is to implement a clinic trial for a non-invasive Preimplantation Genetic Screening (PGS) method for human embryos that are not suitable for standard biopsy and PGS screening. Only embryos that are not suitable for biopsy and can not undergo standard PGS screening will be used for this trail, All embryos that are suitable for biopsy will undergo standard PGS screening. Embryos that are suitable for biopsy are Grade 1 and 2 expanded blastocysts. Embryos that are suitable for freezing but not for biopsy are Grade 3 non-expanded blastocysts ie. early blastocysts.

Patients treatment will not be altered in any way and will proceed as per standard IVF cycle, including medications, oocyte retrieval, insemination and fertilization and embryo culture. Patient embryos will be cultured as per standard protocols; G1/G2 Plus media (Vitrolife) overlayed with oil (Ovoil, Vitrolife) in 6% CO2, 5% O2, 37 degrees.
On day 6, embryos will be assessed for morphology and graded as per current clinical protocols by clinical staff not involved in the trial. All embryos suitable for biopsy (Grade 1 and 2 - expanded blastocysts) will undergo standard biopsy and PGS treatment. For all embryos that are suitable for freezing but not for biopsy (Grade 3 - non-expanded blastocysts ie. early blastocysts), spent culture medium will be sampled and assessed for aneuploidy as described below.

A 4 micro litre drop of the spent culture medium will be sampled using PCR grade filtered tips, under laminar flow, and placed into PCR grade 0.6 ml tubes. DNA from the culture media sample will be amplified by WGA kit and Next Generation Sequencing using standard Illumina protocols.

• Euploid embryos will be made available for transfer. If a pregnancy results from the transfer of the embryo, patients will be offered a NIPT (Non-Invasive Prenatal Testing) first trimester prenatal screening as per standard patient care.
• Embryos assessed as aneuploid will have a confirmatory assessment using standard protocols.

The main risk of this non-invasive technology is that a false positive will be obtained, i.e. indicating an embryo is aneuploid and the confirmatory biopsy screens the embryo as euploid. The screening is not 100% accurate and there remains a risk that the result could be incorrect. However, the thaw and confirmatory PGS biopsy assessment of embryos screened as aneuploid will minimise the risk of discarding a normal embryo. These embryos will be thawed, cultured on and biopsied as per standard protocols, It is described as a 'modified confirmatory biopsy' because embryos that are screened using biopsy and standard PGS are not ever thawed and biopsied a second time to confirm these initial screening results.

Patients will have any embryo screened as euploid via standard PGS or via NEST4E non-invasive screening available for transfer. The decision on which embryo to transfer and how many to transfer is outside the scope of the trial. Standard patient care practices will be followed and this decision will be are made by the patients treating clinician in consultation with senior embryologists not involved in the trial. Patients typically have a single embryos transfer however double embryo transfers are requested by patients for various reasons but are only approved by their treating clinician upon informed consent once the of the risks involved have been given.

For all stages/steps of the trial, patient treatment and embryology will be performed by those involved in standard patient care, ie. nurses, clincians, emrbyologyists. The only part of the trial that will involve someone outside of this scope is the informed consenting to be part of the trial will be performed by Dr Francesca Bell who is a research associate at Repromed. She will also be reposnible for the collection of culture media for embryos that will be screened non-invasively. The WGA DNA amplification, sequencing and screening of embryos will be formed by a qualified member of the genetics team at Repromed who
is responsible for standard PGS patient embryo care.
Intervention code [1] 301531 0
Early detection / Screening
Comparator / control treatment
No control group
Control group
Uncontrolled

Outcomes
Primary outcome [1] 306291 0
The proportion of embryos that are unable to be screened using standard biopsy and PGS that can be conclusively screened for aneuploidy using NEST4E.
Timepoint [1] 306291 0
When a conclusive result is obtained from NEST4E screening (sequencing of amplified DNA from the embryo culture media) for embryos that can not be biopsied.
Secondary outcome [1] 348093 0
For euploid embryos screened using NEST4E and transferred resulting in pregnancy - percentage concordance of media sample to 1st trimester screening.
Timepoint [1] 348093 0
Following non-invasive NIPT based first trimester screening which can be performed as early as 10 weeks.
Secondary outcome [2] 348094 0
For aneuploid embryos screened using NEST4E- percentage of embryos with concordance to confirmatory biopsy and standard PGS screening following thawing and a an extended culture protocol.
Timepoint [2] 348094 0
Next generation sequencing results of biopsied cells can be obtained with 2 weeks post trophectoderm biopsy
Secondary outcome [3] 348095 0
Clinical pregnancy rates of transferred embryos
Timepoint [3] 348095 0
Determination of a positive pregnancy test occurs 12-14 days post embryo transfer via detection of HCG levels from a maternal blood test. Confirmation of a viable pregnancy is performed at 6-8 weeks of pregnancy via ulstrasound to detect fetal heart beat Live birth outcomes will be collected 2-4 weeks post the due date as per standard clinical protocol.
Secondary outcome [4] 348096 0
Live birth rates of embryos screened as euploid using the non-invasive NEST4E test
Timepoint [4] 348096 0
Live birth outcomes will be collected as per standard Repromed patient care. This information is generally collected in writing from the treating obstetrician/doctor, or alternatively directly from the patient with a follow up phone call from the clinic nurses several weeks post due date. It is a regulatory requirement that gestation length, birthweight, sex, method of delivery and details of any maternal or neonatal complications are reported to ANZARD (Australian & New Zealand Assisted Reproduction Database).

Eligibility
Key inclusion criteria
1. Any patient for whom PGS is considered as a treatment option will be eligible
2. Patients that have blastocysts’ that are not suitable for standard biopsy and PGS.

Under the current Ethical guidelines on the use of assisted reproductive technology in clinical practice and research (2017), only patients that have already consented to the use of PGT to select against a genetic condition, disease or abnormality will be approached in this study. This is to recognise that the decision to have genetic testing requires serious ethical consideration and is to prevent patients consenting to this trial to access this technology without having considered the full ethical implications.
Minimum age
18 Years
Maximum age
53 Years
Gender
Both males and females
Can healthy volunteers participate?
No
Key exclusion criteria
1. Patients involved in the Donor program (oocyte and sperm)
2. Patients with a known infectious disease (HIV, HEP B, HEP C)
3. Patients using Preimplantation Genetic Diagnosis (PGD) for single genes or translocations.

Study design
Purpose of the study
Diagnosis
Allocation to intervention
Non-randomised trial
Procedure for enrolling a subject and allocating the treatment (allocation concealment procedures)
Allocation is not concealed
Methods used to generate the sequence in which subjects will be randomised (sequence generation)
non-randomised trial
Masking / blinding
Open (masking not used)
Who is / are masked / blinded?



Intervention assignment
Single group
Other design features
none
Phase
Not Applicable
Type of endpoint(s)
Efficacy
Statistical methods / analysis
Seventy-three patient couples will be recruited to participate in this phase 1 screening trial to assess the clinical implementation of our NEST4E non-invasive technology to screen embryos that are not suitable for standard biopsy and PGS for aneuploidy. The number of participants has been calculated using a sample size calculation for specificity assuming an 80% power, based on a 60% prevalence of aneuploidy for this cohort of patients.

Recruitment
Recruitment status
Recruiting
Date of first participant enrolment
Anticipated
Actual
Date of last participant enrolment
Anticipated
Actual
Date of last data collection
Anticipated
Actual
Sample size
Target
Accrual to date
Final
Recruitment in Australia
Recruitment state(s)
SA
Recruitment hospital [1] 11153 0
Repromed Day Surgery - Dulwich
Recruitment postcode(s) [1] 22978 0
5065 - Dulwich

Funding & Sponsors
Funding source category [1] 299780 0
Commercial sector/Industry
Name [1] 299780 0
Repromed
Address [1] 299780 0
180 Fullarton Road Dulwich SA 5065
Country [1] 299780 0
Australia
Primary sponsor type
Commercial sector/Industry
Name
Repromed
Address
180 Fullarton Road Dulwich SA 5065
Country
Australia
Secondary sponsor category [1] 299159 0
None
Name [1] 299159 0
Address [1] 299159 0
Country [1] 299159 0

Ethics approval
Ethics application status
Approved
Ethics committee name [1] 300669 0
Bellberry Human Research Ethics Committee E
Ethics committee address [1] 300669 0
129 Glen Osmond Road Eastwood South Australia 5063
Ethics committee country [1] 300669 0
Australia
Date submitted for ethics approval [1] 300669 0
14/03/2018
Approval date [1] 300669 0
08/06/2018
Ethics approval number [1] 300669 0
2018-02-082

Summary
Brief summary
The primary purpose of this trial is to evaluate the clinical implementation of our NEST4E non-invasive method for aneuploidy screening of embryos that are unable to be biopsied and screened using the standard PGS technology. Our hypothesis is that this non-invasive technology will be capable of providing a genetic screening result for embryos that we would not be able to provide a result for using standard methods currently used. Seventy three patients will be recruited by informed consent to participate in this trial. The spent culture media will be collected from these embryos and the cell-free DNA analysed to determine the ploidy status of the embryo.

Aim 1: For euploid embryos - determine the percentage concordance of media sample to 1st trimester screening. For aneuploid embryos- determine the percentage of embryos with concordance to confirmatory biopsy and standard PGS screening.
Aim 2: Determine clinical pregnancy rates and live birth outcomes of transferred embryos
Trial website
Trial related presentations / publications
Public notes

Contacts
Principal investigator
Name 84406 0
Prof Michelle Lane
Address 84406 0
Repromed 180 Fullarton Road Dulwich SA 5065
Country 84406 0
Australia
Phone 84406 0
+61 883338111
Fax 84406 0
Email 84406 0
mlane@repromed.com.au
Contact person for public queries
Name 84407 0
Dr Francesca Bell
Address 84407 0
Repromed 180 Fullarton Road Dulwich SA 5065
Country 84407 0
Australia
Phone 84407 0
+61 883338111
Fax 84407 0
Email 84407 0
fbell@repromed.com.au
Contact person for scientific queries
Name 84408 0
Dr Francesca Bell
Address 84408 0
Repromed 180 Fullarton Road Dulwich SA 5065
Country 84408 0
Australia
Phone 84408 0
+61 883338111
Fax 84408 0
Email 84408 0
fbell@repromed.com.au

Data sharing statement
Will individual participant data (IPD) for this trial be available (including data dictionaries)?
No
No/undecided IPD sharing reason/comment
the data for this trial will be published in peer reviewed publications
What supporting documents are/will be available?
No other documents available
Summary results
No Results