Discordance rates between day 3 and day 6 chromosome results and the predictive value of time-lapse morphokinetics

NGS Results for Donor Cycles aged 18 to 30 years old from IVF Centers with Ten or More Cycles

No. Cycles per Center (a-i)

Avg Donor Age

Avg No. Biopsy per Patient

No Amplification (%)a

Degraded DNA (%)a

Euploid (%)b

Aneuploid(%)b

Mosaic (%)b

a-16 b-23 c-12 d-41 e-14 f-19 g-18 h-36 i-13 p

24.00
2.31 25.27
2.35 22.92
1.44 25.50
2.44 24.00
2.04 25.42
3.02 25.41
3.10 26.14
2.58 25.31
2.36 0.005

7.06
2.86 6.55
3.53 7.75
3.14 7.30
3.83 10.43
4.11 9.21
5.48 8.53
4.52 8.03
4.74 5.31
2.56 0.055

0.88
4.17 0.63
1.78 4.30
10.76 1.03
5.45 0.68
4.45 1.14
3.33 0.00
0.00 0.35
3.33 0.00
0.00 0.351

2.65
5.46 0.69
4.26 2.15
3.73 2.40
4.86 1.37
10.69 4.00
10.44 2.76
11.04 1.38
7.84 2.90
11.09 0.987

44.95
28.11 52.08
27.70 43.68
20.17 56.29
25.07 50.35
21.62 38.82
20.88 47.92
24.64 45.49
23.29 37.31
22.20 0.169

32.11
26.90 18.06
18.98 39.08
19.18 26.22
22.86 17.48
10.69 27.65
19.82 18.06
21.02 26.74
17.13 14.93
12.18 0.018

22.94
13.58 29.86
26.63 17.24
12.20 17.48
18.73 32.17
19.79 33.53
15.07 34.03
21.67 27.78
26.31 47.76
21.26 <0.001

a.Percent expressed as total over total number biopsied b.Percent expressed as total over total embryos with diagnosis given

RESULTS: A total of 268 cycles from 38 different IVF centers resulted from the query. A total of 192 cycles from 9 different IVF centers were included in the analysis. The average age of patients was 25.15
2.60 years old (range 19-30), with a cumulative total of 1492 blastocysts biopsied. The overall euploid rate was 48.94%
24.61,with an aneuploid rate of 24.65%
20.76 and mosaic rate of 26.41%
25.0. When the 9 centers were compared as a group, there was a significant difference in mosaic rate (p<0.001) and aneuploid rate (p0.018), but not in the euploid rate (p 0.166). Linear regression on all 268 cycles showed no association between age and rate of euploidy (R¼0.023), aneuploidy (R¼0.056) or mosaicism (R¼0.054); indicating differences in donor ages amongst centers has no affect. There was no significant correlation between the number of blastocysts for biopsy and the percent euploid(R¼0.122), aneuploid(R¼0.035) or mosaic (R ¼0.099). CONCLUSIONS: The difference in mosaic and aneuploid rateswith NGS amongst donor cycles within different IVF institutions shows that there may be a significant practice or laboratory component affecting outcomes. The lack of association of mosaicism and aneuploidy in this population further indicates environmental factors may play an important role.

CONCLUSIONS: In our study, early TLM parameters did not predict blastulation or euploid status on day 3 or day 6 using CC2 and CC3 data. Discordant results were as high as 58% between day 3 and 6. This supports the theory that abnormal blastomeres likely undergo selective apoptosis and repair mechanisms during embryo growth, making day 3 CCS inaccurate. It is also possible that limited DNA material present in single cells is prone to chromosome dropout during the whole genome amplification process, leading to false positives (aneuploids). Discordant NGS Results

Embryo 1 2 3 4 5 6 7

NGS Results Day 3 46, XY, +6, +18 44, XY, -1, -11 42, XX, -4, -8 48, XX, +1, +5 44, XO, -5 48, XY, +5, +10 50, XY, +5, +11, +21

NGS Results Day 6 46, XY 46, XO, +1 47, XX, +10 46, XX 46, XX 46, XY 46, XY

P-137 Tuesday, October 18, 2016 DISCORDANCE RATES BETWEEN DAY 3 AND DAY 6 CHROMOSOME RESULTS AND THE PREDICTIVE VALUE OF TIME-LAPSE MORPHOKINETICS. J. R. Ho,a N. Arrach,b W. Salem,a S. A. Ingles,a K. Bendikson,a K. Chung,a R. Paulson,a A. Ahmady.a aUniversity of Southern California, Los Angeles, CA; bGenetics, Progenesis, La Jolla, CA. OBJECTIVE: Time-lapse morphokinetics (TLM) may provide a noninvasive method for selecting higher quality embryos for transfer. Some studies suggest early kinetic parameters may give insight to embryos with better developmental competence. However, there are mixed data regarding the ability of TLM to predict blastulation and chromosomal status. In this study, we investigate the relationship of TLM with blastulation and ploidy status. Comprehensive chromosomal screening (CCS) was performed on day 3 and day 6 on the same embryo, allowing us to also evaluate discordance rates between results. DESIGN: Prospective study of previously cryopreserved human embryos. MATERIALS AND METHODS: 25 cryopreserved zygote embryos were thawed and cultured in the ESCO Miri-Time Lapse incubator. On day 3, each embryo underwent blastomere biopsy, and on day 6, all embryos regardless of progression were sent for CCS. CCS analysis was performed by next generation sequencing (NGS). Kruskall Wallis was used to analyze the relationship between TLM parameters, blastulation, and ploidy status. For TLM, we included time at which embryos reached each developmental stage. We included second and third cell cycle duration (CC2 and CC3), as well as time between 3-cell to 4-cell (S2), and 5-cell to 8-cell (S3) stages in the analysis. RESULTS: 22 out of the 25 embryos survived the thaw process. 11 embryos (50%) grew to full blastocyst stage. Contrary to previous studies, timing of the second and third cycle (CC2 and CC3), were not predictive of blastulation or euploid status. There were no significant differences in TLM medians between those that formed full blastocysts vs those that arrested early. Other TLM parameters did not predict ploidy status. 14 out of 22 (63%) of NGS results on day 6 were euploid. Of 12 euploid embryos, corresponding day 3 results were discordant in 7 (58%) and showed aneuploidy. Table 1 displays discordant results.

FERTILITY & STERILITYÒ

References: 1. Wong C, Loewke K, Bossert N, et al. Non-invasive imaging of human embryos before embryonic genome activation predicts development to the blastocyst stage. Nature Biotech 2010 28(10):1114-1121. Supported by: Progenesis, Inc. (La Jolla, CA). P-138 Tuesday, October 18, 2016 KARYOMAPPING: THE GOLD STANDARD FOR PREIMPLANR. Prates,b TATION GENETIC DIAGNOSIS. K. M. Doyle,a C. Stimach,a F. Licciardi,c A. Dokras,d S. H. Chen,e T. K. McWilliams,a C. A. Benadiva,f J. Kitchen,a M. Konstantinidis.b aGenesis Genetics, Plymouth, MI; bReprogenetics, Livingston, NJ; cOBGYN, New York University Langone Medical Center, New York, NY; dObstetrics and Gynecology, Reproductive Endocrinology, Philadelphia, PA; eGynecology and Obstetrics, IRMS at Saint Barnabas, Livingston, NJ; fREI, University of Connecticut, Farmington, CT. OBJECTIVE: Preimplantation Genetic Diagnosis (PGD) has helped couples avoid inherited genetic diseases for over 25 years. Karyomapping can streamline this process with a high resolution and rapid result test design. This study is an evaluation of the Single Nucleotide Polymorphism (SNP)based karyomapping platform as the best standard for preimplantation genetic diagnosis of single gene disorders, using an analysis of over 9,000 trophectoderm biopsies and 304 diverse genetic conditions. DESIGN: Retrospective, multi-faceted analysis of embryo biopsies submitted for single-gene diagnosis on the new karyomapping test platform. MATERIALS AND METHODS: Familial DNA from either buccal swabs or blood samples was analyzed along with amplified embryo DNA using the Illumina Karyomapping assay. The data was analyzed with BlueFuse Multi software to detect affected familial haplotypes and determine disease status of embryos. When requested, aneuploidy screening was performed in parallel using either Comparative Genomic Hybridization microarray (aCGH) or Next Generation Sequencing.

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