Publications by authors named "Nathan Treff"

106 Publications

Embryo Screening for Polygenic Disease Risk: Recent Advances and Ethical Considerations.

Genes (Basel) 2021 Jul 21;12(8). Epub 2021 Jul 21.

Department of Physics and Astronomy, Michigan State University, East Lansing, MI 48824, USA.

Machine learning methods applied to large genomic datasets (such as those used in GWAS) have led to the creation of polygenic risk scores (PRSs) that can be used identify individuals who are at highly elevated risk for important disease conditions, such as coronary artery disease (CAD), diabetes, hypertension, breast cancer, and many more. PRSs have been validated in large population groups across multiple continents and are under evaluation for widespread clinical use in adult health. It has been shown that PRSs can be used to identify which of two individuals is at a lower disease risk, even when these two individuals are siblings from a shared family environment. The relative risk reduction (RRR) from choosing an embryo with a lower PRS (with respect to one chosen at random) can be quantified by using these sibling results. New technology for precise embryo genotyping allows more sophisticated preimplantation ranking with better results than the current method of selection that is based on morphology. We review the advances described above and discuss related ethical considerations.
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http://dx.doi.org/10.3390/genes12081105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8393569PMC
July 2021

The "mosaic" embryo: misconceptions and misinterpretations in preimplantation genetic testing for aneuploidy.

Fertil Steril 2021 Jul 22. Epub 2021 Jul 22.

Genomic Prediction Inc., North Brunswick, New Jersey.

Preimplantation genetic testing for aneuploidy (PGT-A) remains one of the most controversial topics in reproductive medicine. With more than 40% of in vitro fertilization cycles in the United States reportedly involving PGT, both those in favor of and those opposed to PGT-A have significant interest in the efficacy of PGT-A. Ongoing issues include what patient population, if any, benefits from PGT-A, the true frequency of chromosomal mosaicism, whether embryonic aneuploidies self-correct, and how practitioners manage embryos designated as "mosaic." This review addresses several misconceptions and misinterpretations of data surrounding the genetic analysis and prediction of mosaicism in the preimplantation embryo.
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http://dx.doi.org/10.1016/j.fertnstert.2021.06.027DOI Listing
July 2021

Isn't it time to stop calling preimplantation embryos "mosaic"?

F S Rep 2020 Dec 4;1(3):164-165. Epub 2020 Nov 4.

Genomic Prediction, North Brunswick, New Jersey.

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http://dx.doi.org/10.1016/j.xfre.2020.10.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8244277PMC
December 2020

Editorial: Causes of Oocyte Aneuploidy and Infertility in Advanced Maternal Age and Emerging Therapeutic Approaches.

Front Endocrinol (Lausanne) 2021 23;12:652990. Epub 2021 Feb 23.

Genomic Prediction Inc., North Brunswick, NJ, United States.

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http://dx.doi.org/10.3389/fendo.2021.652990DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7940751PMC
February 2021

Allele-Specific Chromosome Removal after Cas9 Cleavage in Human Embryos.

Cell 2020 12 29;183(6):1650-1664.e15. Epub 2020 Oct 29.

Department of Pediatrics and Naomi Berrie Diabetes Center, Columbia University, New York, NY 10032, USA; Columbia University Stem Cell Initiative, New York, NY 10032, USA; Department of Obstetrics and Gynecology, Columbia University, New York, NY 10032, USA. Electronic address:

Correction of disease-causing mutations in human embryos holds the potential to reduce the burden of inherited genetic disorders and improve fertility treatments for couples with disease-causing mutations in lieu of embryo selection. Here, we evaluate repair outcomes of a Cas9-induced double-strand break (DSB) introduced on the paternal chromosome at the EYS locus, which carries a frameshift mutation causing blindness. We show that the most common repair outcome is microhomology-mediated end joining, which occurs during the first cell cycle in the zygote, leading to embryos with non-mosaic restoration of the reading frame. Notably, about half of the breaks remain unrepaired, resulting in an undetectable paternal allele and, after mitosis, loss of one or both chromosomal arms. Correspondingly, Cas9 off-target cleavage results in chromosomal losses and hemizygous indels because of cleavage of both alleles. These results demonstrate the ability to manipulate chromosome content and reveal significant challenges for mutation correction in human embryos.
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http://dx.doi.org/10.1016/j.cell.2020.10.025DOI Listing
December 2020

Preimplantation genetic testing for aneuploidy: A review of published blastocyst reanalysis concordance data.

Prenat Diagn 2021 04 4;41(5):545-553. Epub 2020 Oct 4.

Genomic Prediction, North Brunswick, New Jersey, USA.

Preimplantation genetic testing for aneuploidy (PGT-A) reduces miscarriage risk, increases the success of IVF, shortens time to pregnancy, and reduces multiple gestation rates without compromising outcomes. The progression of PGT-A has included common application of next-generation sequencing (NGS) from single nucleotide polymorphism microarray, quantitative real-time PCR, and array comparative hybridization platforms of analysis. Additional putative advances in PGT-A capability include classifying embryos as mosaic and predicting the presence of segmental imbalance. A critical component in the process of technical validation of these advancements involves evaluation of concordance between reanalysis results and initial testing results. While many independent studies have investigated the concordance of results obtained from the remaining embryo with the original PGT-A diagnosis, compilation and systematic analysis of published data has not been performed. Here, we review results from 26 primary research articles describing concordance in 1271 human blastocysts from 2260 pairwise comparisons. Results illustrate significantly higher discordance from PGT-A methods which utilize NGS and include prediction of mosaicism or segmental imbalance. These results suggest caution when considering new iterations PGT-A.
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http://dx.doi.org/10.1002/pd.5828DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8259107PMC
April 2021

Exome sequencing links CEP120 mutation to maternally derived aneuploid conception risk.

Hum Reprod 2020 09;35(9):2134-2148

Department of Genetics, Rutgers, The State University of New Jersey, Piscataway, NJ 08854, USA.

Study Question: What are the genetic factors that increase the risk of aneuploid egg production?

Summary Answer: A non-synonymous variant rs2303720 within centrosomal protein 120 (CEP120) disrupts female meiosis in vitro in mouse.

What Is Known Already: The production of aneuploid eggs, with an advanced maternal age as an established contributing factor, is the major cause of IVF failure, early miscarriage and developmental anomalies. The identity of maternal genetic variants contributing to egg aneuploidy irrespective of age is missing.

Study Design, Size, Duration: Patients undergoing fertility treatment (n = 166) were deidentified and selected for whole-exome sequencing.

Participants/materials, Setting, Methods: Patients self-identified their ethnic groups and their ages ranged from 22 to 49 years old. The study was performed using genomes from White, non-Hispanic patients divided into controls (97) and cases (69) according to the number of aneuploid blastocysts derived during each IVF procedure. Following a gene prioritization strategy, a mouse oocyte system was used to validate the functional significance of the discovered associated genetic variants.

Main Results And The Role Of Chance: Patients producing a high proportion of aneuploid blastocysts (considered aneuploid if they missed any of the 40 chromatids or had extra copies) were found to carry a higher mutational burden in genes functioning in cytoskeleton and microtubule pathways. Validation of the functional significance of a non-synonymous variant rs2303720 within Cep120 on mouse oocyte meiotic maturation revealed that ectopic expression of CEP120:p.Arg947His caused decreased spindle microtubule nucleation efficiency and increased incidence of aneuploidy.

Limitations, Reasons For Caution: Functional validation was performed using the mouse oocyte system. Because spindle building pathways differ between mouse and human oocytes, the defects we observed upon ectopic expression of the Cep120 variant may alter mouse oocyte meiosis differently than human oocyte meiosis. Further studies using knock-in 'humanized' mouse models and in human oocytes will be needed to translate our findings to human system. Possible functional differences of the variant between ethnic groups also need to be investigated.

Wider Implications Of The Findings: Variants in centrosomal genes appear to be important contributors to the risk of maternal aneuploidy. Functional validation of these variants will eventually allow prescreening to select patients that have better chances to benefit from preimplantation genetic testing.

Study Funding/competing Interest(s): This study was funded through R01-HD091331 to K.S. and J.X. and EMD Serono Grant for Fertility Innovation to N.R.T. N.R.T. is a shareholder and an employee of Genomic Prediction.

Trial Registration Number: N/A.
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http://dx.doi.org/10.1093/humrep/deaa148DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7828473PMC
September 2020

Preimplantation Genetic Testing for Polygenic Disease Relative Risk Reduction: Evaluation of Genomic Index Performance in 11,883 Adult Sibling Pairs.

Genes (Basel) 2020 06 12;11(6). Epub 2020 Jun 12.

Genomic Prediction Inc., 675 US Highway One, North Brunswick, NJ 08902, USA.

Preimplantation genetic testing for polygenic disease risk (PGT-P) represents a new tool to aid in embryo selection. Previous studies demonstrated the ability to obtain necessary genotypes in the embryo with accuracy equivalent to in adults. When applied to select adult siblings with known type I diabetes status, a reduction in disease incidence of 45-72% compared to random selection was achieved. This study extends analysis to 11,883 sibling pairs to evaluate clinical utility of embryo selection with PGT-P. Results demonstrate simultaneous relative risk reduction of all diseases tested in parallel, which included diabetes, cancer, and heart disease, and indicate applicability beyond patients with a known family history of disease.
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http://dx.doi.org/10.3390/genes11060648DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7349610PMC
June 2020

PREIMPLANTATION GENETIC TESTING: Preimplantation genetic testing for polygenic disease risk.

Reproduction 2020 Nov;160(5):A13-A17

Genomic Prediction Inc., North Brunswick, New Jersey, USA.

Since its introduction to clinical practice, preimplantation genetic testing (PGT) has become a standard of care for couples at risk of having children with monogenic disease and for chromosomal aneuploidy to improve outcomes for patients with infertility. The primary objective of PGT is to reduce the risk of miscarriage and genetic disease and to improve the success of infertility treatment with the delivery of a healthy child. Until recently, the application of PGT to more common but complex polygenic disease was not possible, as the genetic contribution to polygenic disease has been difficult to determine, and the concept of embryo selection across multiple genetic loci has been difficult to comprehend. Several achievements, including the ability to obtain accurate, genome-wide genotypes of the human embryo and the development of population-level biobanks, have now made PGT for polygenic disease risk applicable in clinical practice. With the rapid advances in embryonic polygenic risk scoring, diverse considerations beyond technical capability have been introduced.
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http://dx.doi.org/10.1530/REP-20-0071DOI Listing
November 2020

Utility and First Clinical Application of Screening Embryos for Polygenic Disease Risk Reduction.

Front Endocrinol (Lausanne) 2019 4;10:845. Epub 2019 Dec 4.

Genomic Prediction Inc., North Brunswick, NJ, United States.

For over 2 decades preimplantation genetic testing (PGT) has been in clinical use to reduce the risk of miscarriage and genetic disease in patients with advanced maternal age and risk of transmitting disease. Recently developed methods of genome-wide genotyping and machine learning algorithms now offer the ability to genotype embryos for polygenic disease risk with accuracy equivalent to adults. In addition, contemporary studies on adults indicate the ability to predict polygenic disorders with risk equivalent to monogenic disorders. Existing biobanks provide opportunities to model the clinical utility of polygenic disease risk reduction among sibling adults. Here, we provide a mathematical model for the use of embryo screening to reduce the risk of type 1 diabetes. Results indicate a 45-72% reduced risk with blinded genetic selection of one sibling. The first clinical case of polygenic risk scoring in human preimplantation embryos from patients with a family history of complex disease is reported. In addition to these data, several common and accepted practices place PGT for polygenic disease risk in the applicable context of contemporary reproductive medicine. In addition, prediction of risk for PCOS, endometriosis, and aneuploidy are of particular interest and relevance to patients with infertility and represent an important focus of future research on polygenic risk scoring in embryos.
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http://dx.doi.org/10.3389/fendo.2019.00845DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6915076PMC
December 2019

Validation of concurrent preimplantation genetic testing for polygenic and monogenic disorders, structural rearrangements, and whole and segmental chromosome aneuploidy with a single universal platform.

Eur J Med Genet 2019 Aug 23;62(8):103647. Epub 2019 Apr 23.

Genomic Prediction Inc., 675 US Highway One, North Brunswick, NJ, 08902, USA.

Preimplantation genetic testing (PGT) has been successfully applied to reduce the risk of miscarriage, improve IVF success rates, and prevent inheritance of monogenic disease and unbalanced translocations. The present study provides the first method capable of simultaneous testing of aneuploidy (PGT-A), structural rearrangements (PGT-SR), and monogenic (PGT-M) disorders using a single platform. Using positive controls to establish performance characteristics, accuracies of 97 to >99% for each type of testing were observed. In addition, this study expands PGT to include predicting the risk of polygenic disorders (PGT-P) for the first time. Performance was established for two common diseases, hypothyroidism and type 1 diabetes, based upon availability of positive control samples from commercially available repositories. Data from the UK Biobank, eMERGE, and T1DBASE were used to establish and validate SNP-based predictors of each disease (7,311 SNPs for hypothyroidism and 82 for type 1 diabetes). Area under the curve of disease status prediction from genotypes alone were 0.71 for hypothyroidism and 0.68 for type 1 diabetes. The availability of expanded PGT to evaluate the risk of polygenic disorders in the preimplantation embryo has the potential to lower the prevalence of common genetic disease in humans.
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http://dx.doi.org/10.1016/j.ejmg.2019.04.004DOI Listing
August 2019

Investigating the impact of the timing of blastulation on implantation: management of embryo-endometrial synchrony improves outcomes.

Hum Reprod Open 2018 20;2018(4):hoy022. Epub 2018 Dec 20.

Division of Reproductive Endocrinology, Department of Obstetrics and Gynecology, Thomas Jefferson University, Philadelphia, PA, USA.

Study Question: Do embryos with delayed blastulation have inferior reproductive potential or poorer outcomes due in part to embryo and endometrial synchrony?

Summary Answer: Diminished outcomes in embryos with delayed blastulation undergoing fresh embryo transfer (ET) may be attributed to a loss of embryonic-endometrial synchrony.

What Is Known Already: Embryos that blastulate slowly have lower sustained implantation rates (SIR) than those which blastulate normally on Day 5 (D5). Traditionally this has been attributed to reduced embryo quality; however, dyssynchrony with the endometrium is also a possibility and has not been fully described. This convenient cohort composed of groups that resulted from a practice wide change in laboratory protocol allows for evaluation of embryo and endometrial synchrony as it related to blastocyst expansion.

Study Design Size Duration: A retrospective cohort analysis was carried out from January 2009 to February 2013. Three cohorts were identified: D5 ET, D6 ET and frozen ET that comprised 822 patients, 763 patients and 718 patients, respectively. Each of these cohorts had slowly blastulating and normally blastulating embryos identified.

Participants/materials Setting Methods: The study setting was academic affiliated private practice. All first fresh or cryopreserved ETs from 2010 to 2013 were studied. Non-biopsied embryos were classified into two groups on D5: slowly blastulating (Morula-Gardner 1) or normally blastulating (Gardner 2-6). Only single ETs or transfer of two embryos within the same classification group were included. Outcomes were compared between classification groups in embryos undergoing transfer on D5, D6, or after cryopreservation. This assesses the impact of transfer timing in fresh cycles as well as isolating a pure embryonic factor in frozen ET cycles. Sustained implantation was defined as heart beat detection at discharge sonogram at 8 weeks gestation. SIR was defined as the number of embryos transferred meeting criteria for sustained implantation divided by the total number of embryos transferred.

Main Results And The Role Of Chance: In total, 3391 embryos were transferred to 1966 patients. On D5, SIRs were significantly lower with slowly blastulating embryos (44% versus 64% in women <35 years of age ( < 0.001) and 18% versus 56% in women ≥35 years of age ( < 0.001)). Fresh D6 ETs also had significantly lower SIRs for embryos that were slowly blastulating on D5 (52% versus 63% in <35 years of age ( < 0.05) and 32% versus 48% in ≥35 years of age ( < 0.005)) despite continued development to full blastocysts and being morphologically equivalent at the time of ET, suggesting dyssynchrony. However, when slowly blastulating embryos underwent vitrification and then ET, they had SIRs which were equivalent to their normally blastulating counterparts (57% versus 60% in <35 years of age ( = 0.5) and 37% versus 42% in ≥35 years of age ( = 0.3)). An intraclass correlation and a generalized estimating equation corrected for patient age was performed which confirmed these findings. The normalization in cryopreserved ETs indicates that dyssynchrony may be a major adverse factor limiting outcomes with late blastulating embryos in fresh cycles.

Limitations Reasons For Caution: This is a retrospective study comprising cohorts from a convenient sample chosen due to a uniform change in embryology laboratory protocol regarding ET day, however, this was done independent of the management of embryo and endometrial synchrony. Although strict ultrasound and serum progesterone criteria were utilized to make endometrial receptivity uniform, pathologic states of pre-receptive and post-receptive endometrium cannot be ruled out.

Wider Implications Of The Findings: The data surrounding embryo and endometrial synchrony should be considered in patients undergoing IVF and attention to the variations in blastulation rates can be applied to any patient undergoing extended embryo culture.

Study Funding/competing Interests: None.
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http://dx.doi.org/10.1093/hropen/hoy022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6396639PMC
December 2018

Integrated RNA-seq and ChIP-seq analysis reveals a feed-forward loop regulating H3K9ac and key labor drivers in human placenta.

Placenta 2019 01 10;76:40-50. Epub 2019 Jan 10.

Department of Obstetrics, Gynecology, and Reproductive Sciences, Division of Maternal-Fetal Medicine, Rutgers Robert Wood Johnson Medical School, New Brunswick, NJ, 08901, USA. Electronic address:

Background: Chromatin alterations are important mediators of gene expression changes. We have recently shown that activated non-canonical NF-κB signaling (RelB/p52) recruits histone acetyltransferase CBP and deacetylase HDAC1 to selectively acetylate H3K9 (H3K9ac) to induce expression of corticotropin-releasing hormone (CRH) and prostaglandin-endoperoxide synthase-2 (PTGS2) in the human placenta. Both of these genes play a role in initiating parturition in human pregnancy.

Methods: We performed chromatin immunoprecipitation followed by gene sequencing (ChIP-seq) in primary term human cytotrophoblast (CTB) with use of antibodies to RelB, CBP, HDAC1 and H3K9ac. We further associated these chromatin alterations with gene expression changes from mid-trimester to term in CTB by RNA sequencing (RNA-seq).

Results: We detected a genome-wide differential gene enrichment between mid-trimester and term human placenta. Pathway analysis identified that cytokine-cytokine receptor interaction, NF-κB, and TNF are the leading pathways enriched in term placenta and associated with these chromatin alterations.

Discussions: Our analysis has provided the first-time characterization of the key players of human placental origin with molecular changes resulting from chromatin modifications, which could drive human labor.
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http://dx.doi.org/10.1016/j.placenta.2019.01.010DOI Listing
January 2019

Molecular Testing for Preimplantation Genetic Diagnosis of Single Gene Disorders.

Methods Mol Biol 2019 ;1885:61-71

Thomas Jefferson University, Basking Ridge, NJ, USA.

Preimplantation genetic testing has evolved tremendously from the early days of FISH detection for a select few chromosome aneuploidies to now combining the detection of all whole chromosome imbalances in conjunction with single gene disorder testing for inherited diseases. As universal carrier screening and exome or genome studies become more commonplace, more and more families are becoming interested in reducing the risk of having a child with a severe disease using preimplantation genetic testing. We describe here the use of quantitative PCR (qPCR) for the custom construction of single gene disorder testing plans for families, the validation of the probes designed, and the protocol for diagnosing an embryo biopsy. qPCR has been shown to have the lowest risk of failed amplification and allele dropout and thus the lowest risk of a misdiagnosis, while also currently providing the fastest protocol to allow for rapid turnaround of results.
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http://dx.doi.org/10.1007/978-1-4939-8889-1_4DOI Listing
June 2019

The pros and cons of preimplantation genetic testing for aneuploidy: clinical and laboratory perspectives.

Fertil Steril 2018 08;110(3):353-361

Ronald O. Perelman and Claudia Cohen Center for Reproductive Medicine, Weill Cornell Medical College, New York, New York.

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http://dx.doi.org/10.1016/j.fertnstert.2018.06.002DOI Listing
August 2018

Cumulus cell transcriptome profiling is not predictive of live birth after in vitro fertilization: a paired analysis of euploid sibling blastocysts.

Fertil Steril 2018 03 7;109(3):460-466.e2. Epub 2018 Feb 7.

Reproductive Medicine Associates of New Jersey, Basking Ridge, New Jersey; Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Sidney Kimmel Medical College of Thomas Jefferson University, Philadelphia, Pennsylvania.

Objective: To compare the transcriptome of cumulus cells associated with a euploid embryo that resulted in live birth with that of a sibling euploid embryo without sustained implantation.

Design: Paired analysis.

Setting: Academic institution.

Patient(s): Couples undergoing in vitro fertilization (IVF)/intracytoplasmic sperm injection with preimplantation genetic screening with female age ≤42 years and normal ovarian reserve.

Intervention(s): Transcriptome profiling of cumulus cells from sibling oocytes for correlation with live birth after euploid blastocyst transfer. Embryos were individually cultured to facilitate association with clinical outcomes. The cumulus cell transcriptome from the embryo resulting in live birth was compared with that of its sibling embryo without sustained implantation to investigate potential biomarkers that may aid in embryo selection.

Main Outcome Measure(s): Differential gene expression in cumulus cells associated with a euploid embryo resulting in live birth and its sibling euploid embryo without sustained implantation using next-generation RNA sequencing (RNAseq).

Result(s): Cumulus cell RNAseq of 34 samples (from 17 patients) generated an average of 10.4 ± 4 × 10 reads per sample. A total of 132 differentially expressed genes between sibling embryos that resulted in a live birth and those that did not were identified (P<.05). However, after correcting for multiple testing none of the genes remained significantly differentially expressed (false discovery rate <.05).

Conclusion(s): The RNAseq profiles were similar between cumulus cells associated with a euploid embryo resulting in live birth and its sibling embryo that did not sustain implantation. The cumulus cell transcriptome is not predictive of live birth within an individual patient's cohort of euploid embryos.
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http://dx.doi.org/10.1016/j.fertnstert.2017.11.002DOI Listing
March 2018

Validation of a targeted next generation sequencing-based comprehensive chromosome screening platform for detection of triploidy in human blastocysts.

Reprod Biomed Online 2018 Apr 2;36(4):388-395. Epub 2018 Jan 2.

Rutgers, The State University of New Jersey, Department of Genetics, Piscataway, NJ, USA.

Triploidy accounts for ~2% of natural pregnancies and 15% of cytogenetically abnormal miscarriages. This study aimed to validate triploidy detection in human blastocysts, its frequency and parental origin using genotyping data generated in parallel with chromosome copy number analysis by a targeted next generation sequencing (tNGS)-based comprehensive chromosome screening platform. Phase 1: diploid and triploid control samples were blinded, sequenced by tNGS and karyotype predictions compared for accuracy. Phase 2: tNGS was used to calculate the frequency of triploidy in 18,791 human blastocysts from trophectoderm (TE) biopsies. Phase 3: parental origin of the inherited extra alleles was evaluated by sequencing parental gDNA to validate triploidy predictions from Phase 2. All karyotypes and ploidy in controls from Phase 1 were correctly predicted by two independent methods. A blastocyst triploidy frequency of 0.474% (89/18,791) was observed in Phase 2 of the study. Finally, five suspected triploid blastocysts with parental DNA available were confirmed to be triploid and of maternal origin. tNGS provides higher sequencing depth in contrast to other contemporary NGS platforms, allowing for accurate single nucleotide polymorphism calling and accurate detection of triploidy in TE biopsies. Triploidy in intracytoplasmic sperm injection-derived blastocysts is rare and mostly of maternal origin.
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http://dx.doi.org/10.1016/j.rbmo.2017.12.015DOI Listing
April 2018

Mitochondrial DNA content is associated with ploidy status, maternal age, and oocyte maturation methods in mouse blastocysts.

J Assist Reprod Genet 2017 Dec 24;34(12):1587-1594. Epub 2017 Oct 24.

Rutgers, the State University of New Jersey, New Brunswick, NJ, 08901, USA.

Purpose: It was reported that mitochondrial DNA (mtDNA) was significantly increased in aneuploid human embryos compared to euploid embryos and was also associated with maternal age. In this study, we further established the mouse model of mtDNA quantitation in reproductive samples based on whole-genome amplification (WGA) and next-generation sequencing (NGS).

Methods: WGA followed by NGS-based mtDNA quantitation was first performed on 6 single- and 100-cell samples from a tumor-derived mouse cell line, which was exposed to ethidium bromide to reduce mtDNA content. The relative mtDNA content was normalized to nuclear DNA. This method was then applied to mouse reproductive samples, including 40 pairs of oocytes and polar bodies from 8 CD-1 female mice of advanced reproductive age and 171 blastocysts derived via in vitro maturation (IVM) or in vivo maturation (IVO) from young (6-9 weeks) and reproductively aged (13.5 months) female CF-1 mice.

Results: Exposure to ethidium bromide for 3 and 6 days decreased mtDNA levels in both the single- and 100-cell samples as expected. Results demonstrated that the first polar body contained an average of 0.9% of mtDNA relative to oocytes. Compared to the cells in blastocysts, oocytes contained about 180 times as much mtDNA per cell. mtDNA levels were compared among blastocysts from reproductively young and old female mice that had either been produced by IVM or IVO. Cells in blastocysts from younger mice contained significantly lower amounts of mtDNA compared to aged mice (P < 0.0001). Cells in blastocysts produced via IVO had higher mtDNA content than IVM-derived blastocysts (P = 0.0001). Cells in aneuploid blastocysts were found to have significantly higher (1.74-fold) levels of mtDNA compared to euploid blastocysts (P = 0.0006).

Conclusion: A reliable method for assessing mtDNA content in mouse gametes and embryos was established. Relative mtDNA levels were elevated in aneuploid embryos relative to euploid embryos, were higher in blastocysts from reproductively old mice relative to young mice, and were lower in embryos derived from IVM compared to IVO.
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http://dx.doi.org/10.1007/s10815-017-1070-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5714829PMC
December 2017

Abnormally fertilized oocytes can result in healthy live births: improved genetic technologies for preimplantation genetic testing can be used to rescue viable embryos in in vitro fertilization cycles.

Fertil Steril 2017 12 15;108(6):1007-1015.e3. Epub 2017 Sep 15.

Genera, Centers for Reproductive Medicine, Clinica Valle Giulia, Rome, Italy; Genetyx, Molecular Genetics Laboratory, Marostica, Italy.

Objective: To test whether abnormally fertilized oocyte (AFO)-derived blastocysts are diploid and can be rescued for clinical use.

Design: Longitudinal-cohort study from January 2015 to September 2016 involving IVF cycles with preimplantation genetic testing for aneuploidy (PGT-A). Ploidy assessment was incorporated whenever a blastocyst from a monopronuclear (1PN) or tripronuclear zygote (2PN + 1 smaller PN; 2.1 PN) was obtained.

Setting: Private IVF clinics and genetics laboratories.

Patient(s): A total of 556 women undergoing 719 PGT-A cycles.

Intervention(s): Conventional chromosome analysis was performed on trophectoderm biopsies by quantitative polymerase chain reaction. For AFO-derived blastocysts, ploidy assessment was performed on the same biopsy with the use of allele ratios for hetorozygous SNPs analyzed by means of next-generation sequencing (1:1 = diploid; 2:1 = triploid; loss of heterozygosity = haploid). Balanced-diploid 1PN- and 2.1PN-derived blastocysts were transferred in the absence of normally fertilized transferable embryos.

Main Outcome Measure(s): Ploidy constitution and clinical value of AFO-derived blastocysts in IVF PGT-A cycles.

Result(s): Of the 5,026 metaphase II oocytes injected, 5.2% and 0.7% showed 1PN and 2.1PN, respectively. AFOs showed compromised embryo development (P<.01). Twenty-seven AFO-derived blastocysts were analyzed for ploidy constitution. The 1PN-derived blastocysts were mostly diploid (n = 9/13; 69.2%), a few were haploid (n = 3/13; 23.1%), and one was triploid (n = 1/13; 7.7%). The 2.1PN-derived blastocysts were also mostly diploid (n = 12/14; 85.7%), and the remainder were triploid. Twenty-six PGT-A cycles resulted in one or more AFO-derived blastocysts (n = 26/719; 3.6%). Overall, eight additional balanced-diploid transferable embryos were obtained from AFOs. In three cycles, the only balanced-diploid blastocyst produced was from an AFO (n = 3/719; 0.4%). Three AFO-derived live births were achieved: one from a 1PN zygote and two from 2.1PN zygotes.

Conclusion(s): Enhanced PGT-A technologies incorporating reliable ploidy assessment provide an effective tool to rescue AFO-derived blastocysts for clinical use.
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http://dx.doi.org/10.1016/j.fertnstert.2017.08.004DOI Listing
December 2017

Evaluation of comprehensive chromosome screening platforms for the detection of mosaic segmental aneuploidy.

J Assist Reprod Genet 2017 Aug 2;34(8):975-981. Epub 2017 Jun 2.

Reproductive Medicine Associates of New Jersey, 140 Allen Rd, Basking Ridge, NJ, 07920, USA.

Purpose: A subset of preimplantation embryos identified as euploid may in fact possess both whole and sub-chromosomal mosaicism, raising concerns regarding the predictive value of current comprehensive chromosome screening (CCS) methods utilizing a single biopsy. Current CCS methods may be capable of detecting sub-chromosomal mosaicism in a trophectoderm biopsy by examining intermediate levels of segmental aneuploidy within a biopsy. This study evaluates the sensitivity and specificity of segmental aneuploidy detection by three commercially available CCS platforms utilizing a cell line mixture model of segmental mosaicism in a six-cell trophectoderm biopsy.

Methods: Two cell lines with known karyotypes were obtained and mixed together at specific ratios of six total cells (0:6, 1:5, 2:4, 3:3, 4:2, 5:1, and 6:0). A female cell line containing a 16.2 Mb deletion on chromosome 5 and a male cell line containing a 25.5 Mb deletion on chromosome 4 were used to create mixtures at each level. Six replicates of each mixture were prepared, randomized, and blinded for analysis by one of the three CCS platforms (SNP-array, VeriSeq NGS, or NexCCS). Sensitivity and specificity of segmental aneuploidy at each level of mosaicism was determined and compared between each platform. Additionally, an alternative VeriSeq NGS analysis method utilizing previously published criteria was evaluated.

Results: Examination of the default settings of each platform revealed that the sensitivity was significantly different between NexCCS and SNP up to 50% mosaicism, custom VeriSeq, and SNP-array up to 66% mosaicism, and between NexCCS and custom VeriSeq up to 50% mosaicism. However, no statistical difference was observed in mixtures with >50% mosaicism with any platform. No comparison was made between default VeriSeq, as it does not report segmental imbalances. Furthermore, while the use of previously published criteria for VeriSeq NGS significantly increased sensitivity at low levels of mosaicism, a significant decrease in specificity was observed (66% false positive prediction of segmental aneuploidy).

Conclusion: These results demonstrate the potential of NGS-based detection methods to detect segmental mosaicism within a biopsy. However, these data also demonstrate that a balance between sensitivity and specificity should be more carefully considered. These results emphasize the importance of vigorous preclinical evaluation of new testing criteria prior to clinical implementation providing a point of departure for further algorithm development and improved detection of mosaicism within preimplantation embryos.
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http://dx.doi.org/10.1007/s10815-017-0924-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5533675PMC
August 2017

Advances in Preimplantation Genetic Testing for Monogenic Disease and Aneuploidy.

Annu Rev Genomics Hum Genet 2017 08 12;18:189-200. Epub 2017 May 12.

Foundation for Embryonic Competence, Basking Ridge, New Jersey 07920.

Genetic testing of preimplantation embryos promises to prevent monogenic disease in children born to at-risk couples, the transfer of unbalanced embryos to patients carrying a balanced translocation, and the use of aneuploid embryos created during in vitro fertilization. Technologies have evolved from fluorescence in situ hybridization to next-generation-sequencing-based aneuploidy screening and allow for simultaneous testing of multiple genetic abnormalities in a single biopsy. The field has also shifted away from polar body or blastomere biopsy and toward trophectoderm biopsy as the new standard. This review describes the multitude of available platforms and methodologies used in contemporary preimplantation genetic testing.
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http://dx.doi.org/10.1146/annurev-genom-091416-035508DOI Listing
August 2017

Comprehensive chromosome screening and gene expression analysis from the same biopsy in human preimplantation embryos.

Mol Hum Reprod 2017 05;23(5):330-338

Reproductive Medicine Associates of New Jersey, 140 Allen Road, Basking Ridge, NJ 07920, USA.

Study Question: Can simultaneous comprehensive chromosome screening (CCS) and gene expression analysis be performed on the same biopsy of preimplantation human embryos?

Summary Answer: For the first time, CCS and reliable gene expression analysis have been performed on the same human preimplantation embryo biopsy.

What Is Known Already: A single trophectoderm (TE) biopsy is routinely used for many IVF programs offering CCS for selection of only chromosomally normal embryos for transfer. Although the gene expression profiling of human preimplantation embryos has been described, to date no protocol allows for simultaneous CCS and gene expression profiling from a single TE biopsy.

Study Design, Size And Duration: This is a proof of concept and validation study structured in two phases. In Phase 1, cell lines were subjected to a novel protocol for combined CCS and gene expression analysis so as to validate the accuracy and reliability of the proposed protocol. In Phase 2, 20 donated human blastocysts were biopsied and processed with the proposed protocol in order to obtain an accurate CCS result and characterize their gene expression profiles using the same starting material.

Participants/materials, Setting And Method: A novel protocol coupling quantitative real-time PCR-based CCS and gene expression analysis using RT-PCR was designed for this study. Phase 1: six-cell aliquots of well-characterized fibroblast cell lines (GM00323, 46,XY and GM04435, 48,XY,+16,+21) were subjected to the proposed protocol. CCS results were compared with the known karyotypes for consistency, and gene expression levels were compared with levels of purified RNA from same cell lines for validation of reliable gene expression profiling. Phase 2: four biopsies were performed on 20 frozen human blastocysts previously diagnosed as trisomy 21 (10 embryos) and monosomy 21 (10 embryos) by CCS. All samples were processed with the proposed protocol and re-evaluated for concordance with the original CCS result. Their gene expression profiles were characterized and differential gene expression among embryos and early embryonic cell lineages was also evaluated.

Main Results And The Role Of Chance: CCS results from cell lines showed 100% consistency with their known karyotypes. ΔΔCt values of differential gene expression of four selected target genes from the cell lines GM4435 and GM0323 were comparable between six-cell aliquots and purified RNA (Collagen type I alpha-1 (COL1A1), P = 0.54; Fibroblast growth factor-5 (FGF5), P = 0.11; Laminin subunit beta-1 (LAMB1), P = 1.00 and Atlastin-1 (ATL1), P = 0.23). With respect to human blastocysts, 92% consistency was reported after comparing embryonic CCS results with previous diagnosis. A total of 30 genes from a human stem cell pluripotency panel were selected to evaluate gene expression in human embryos. Correlation coefficients of expression profiles from biopsies of the same embryo (r = 0.96 ± 0.03 (standard deviation), n = 45) were significantly higher than when biopsies from unrelated embryos were evaluated (r = 0.93 ± 0.03, n = 945) (P < 0.0001). Growth differentiation factor 3 (GDF3) was found to be significantly up-regulated in the inner cell mass (ICM), whereas Caudal type homebox protein-2 (CDX2), Laminin subunit alpha-1 (LAMA1) and DNA methyltransferase 3-beta (DNMT3B) showed down-regulation in ICM compared with TE. Trisomy 21 embryos showed significant up-regulation of markers of cell differentiation (Cadherin-5 (CDH5) and Laminin subunit gamma-1 (LAMC1)), whereas monosomy 21 blastocysts showed higher expression of genes reported to be expressed in undifferentiated cells (Gamma-Aminobutyric Acid Type-A Receptor Beta3 Subunit (GABRB3) and GDF3).

Large Scale Data: N/A.

Limitations, Reasons For Caution: Gene expression profiles of chromosomally normal embryos were not assessed due to restrictive access to euploid embryos for research. Nonetheless, the profile of blastocysts with single aneuploidies was characterized and compared. Only 30 target genes were analyzed for gene expression in this study. Increasing the number of target genes will provide a more comprehensive transcriptomic signature and reveal potential pathways paramount for embryonic competence and correct development.

Wider Implications Of The Findings: This is the first time that CCS and gene expression analysis have been performed on the same human preimplantation embryo biopsy. Further optimization of this protocol with other CCS platforms and inclusion of more target genes will provide innumerable research and clinical applications, such as discovery of biomarkers for embryonic reproductive potential and characterization of the transcriptomic signatures of embryos, potentially allowing for further embryo selection prior to embryo transfer and therefore improving outcomes.

Study Funding And Competing Interests: This study was funded by the Foundation for Embryonic Competence, Basking Ridge, NJ, USA. No conflicts of interests declared.
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http://dx.doi.org/10.1093/molehr/gax014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5420574PMC
May 2017

Identification and characterization of Aurora kinase B and C variants associated with maternal aneuploidy.

Mol Hum Reprod 2017 06;23(6):406-416

Department of Genetics, Rutgers, The State University of New Jersey, 145 Bevier Rd. Piscataway, NJ 08854, USA.

Study Question: Are single nucleotide variants (SNVs) in Aurora kinases B and C (AURKB, AURKC) associated with risk of aneuploid conception?

Summary Answer: Two SNVs were found in patients with extreme aneuploid concepti rates with respect to their age; one variant, AURKC p.I79V, is benign, while another, AURKB p.L39P, is a potential gain-of-function mutant with increased efficiency in promoting chromosome alignment.

What Is Known Already: Maternal age does not always predict aneuploidy risk, and rare gene variants can be drivers of disease. The AURKB and AURKC regulate chromosome segregation, and are associated with reproductive impairments in mouse and human.

Study Design, Size, Duration: An extreme phenotype sample selection scheme was performed for variant discovery. Ninety-six DNA samples were from young patients with higher than average embryonic aneuploidy rates and an additional 96 DNA samples were from older patients with lower than average aneuploidy rates.

Participants/materials, Setting, Methods: Using the192 DNA samples, the coding regions of AURKB and AURKC were sequenced using next generation sequencing. To assess biological significance, we expressed complementary RNA encoding the human variants in mouse oocytes. Assays such as determining subcellular localization and assessing catalytic activity were performed to determine alterations in protein function during meiosis.

Main Results And The Role Of Chance: Ten SNVs were identified using three independent variant-calling methods. Two of the SNVs (AURKB p.L39P and AURKC p.I79V) were non-synonymous and identified by at least two variant-identification methods. The variant encoding AURKC p.I79V, identified in a young woman with a higher than average rate of aneuploid embryos, showed wild-type localization pattern and catalytic activity. On the other hand, the variant encoding AURKB p.L39P, identified in an older woman with lower than average rates of aneuploid embryos, increased the protein's ability to regulate alignment of chromosomes at the metaphase plate. These experiments were repeated three independent times using 2-3 mice for each trial.

Large Scale Data: N/A.

Limitations, Reasons For Caution: Biological significance of the human variants was assessed in an in vitro mouse oocyte model where the variants are over-expressed. Therefore, the human protein may not function identically to the mouse homolog, or the same in mouse oocytes as in human oocytes. Furthermore, supraphysiological expression levels may not accurately reflect endogenous activity. Moreover, the evaluated variants were identified in one patient each, and no trial linking the SNV to pregnancy outcomes was conducted. Finally, the patient aneuploidy rates were established by performing comprehensive chromosome screening in blastocysts, and because of the link between female gamete aneuploidy giving rise to aneuploid embryos, we evaluate the role of the variants in Meiosis I. However, it is possible that the chromosome segregation mistake arose during Meiosis II or in mitosis in the preimplantation embryo. Their implications in human female meiosis and aneuploidy risk remain to be determined.

Wider Implications Of The Findings: The data provide evidence that gene variants exist in reproductively younger or advanced aged women that are predictive of the risk of producing aneuploid concepti in humans. Furthermore, a single amino acid in the N-terminus of AURKB is a gain-of-function mutant that could be protective of euploidy.

Study Funding/competing Interests: This work was supported by a Research Grant from the American Society of Reproductive Medicine and support from the Charles and Johanna Busch Memorial Fund at Rutgers, the State University of NJ to K.S. and the Foundation for Embryonic Competence, Inc to N.T. The authors declare no conflicts of interest.
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http://dx.doi.org/10.1093/molehr/gax018DOI Listing
June 2017

Levels of trophectoderm mitochondrial DNA do not predict the reproductive potential of sibling embryos.

Hum Reprod 2017 04;32(4):954-962

Reproductive Medicine Associates of New Jersey, Basking Ridge, NJ, USA.

Study Question: What is the predictive value of trophectoderm mitochondrial DNA (mtDNA) quantity for blastocyst reproductive potential?

Summary Answer: This study demonstrates that, within a given cohort, mtDNA quantitation does not distinguish between embryos that implant and embryos that do not implant after double embryo transfer (DET).

What Is Already Known: An association between implantation failure and increased quantities of mtDNA has been observed in two studies but not in a third.

Study Design, Size And Duration: A total of 187 patients (nine who received donor oocytes) with DET of one male and one female euploid blastocyst were included in this retrospective study, with 69 singleton deliveries providing the primary dataset to evaluate the predictive value of mtDNA for reproductive potential between January 2010 and July 2016.

Participants/materials, Setting And Method: MtDNA was quantified in cell lines to validate the quantitative PCR assay on limited quantities of starting material and then applied to 374 blastocyst biopsies. Pregnancies resulting in a singleton outcome were analyzed and newborn gender was utilized as a means to identify the implanted embryo. MtDNA quantity was then compared between implanted and non-implanted embryos in order to define the predictive value of mtDNA content for reproductive potential in this subset of patients.

Main Results And The Role Of Chance: An initial comparison of mtDNA levels between all successful and unsuccessful embryos revealed no significant differences. In order to control for patient-specific variables, gender was subsequently used to identify the implanted embryo in DETs resulting in a singleton (n = 69). No systematic difference in relative mtDNA quantity was detected between implanted and non-implanted embryos.

Limitations, Reasons For Caution: This study was conducted at a single center and did not evaluate the entire cohort of embryos from each patient to evaluate cohort specific variation in mtDNA quantity. Although the largest of its kind so far, the sample size of DETs leading to a singleton was relatively small.

Wider Implications Of The Findings: These data highlight the importance of control over patient-specific variables when evaluating candidate biomarkers of reproductive potential. All current available data suggest that mtDNA quantification needs further study before its clinical use to augment embryo selection.

Study Funding/competing Interests: The authors have no potential conflict of interest to declare. No external funding was obtained for this study.

Trial Registration Number: Not applicable.
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http://dx.doi.org/10.1093/humrep/dex034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5400072PMC
April 2017

Preimplantation embryonic mosaicism: origin, consequences and the reliability of comprehensive chromosome screening.

Curr Opin Obstet Gynecol 2017 Jun;29(3):168-174

aReproductive Medicine Associates of New Jersey, Basking Ridge, New Jersey bThomas Jefferson College of Biomedical Sciences, Thomas Jefferson University, Philadelphia, Pennsylvania, USA.

Purpose Of Review: Embryonic mosaicism represents an ongoing challenge for contemporary comprehensive chromosome screening platforms due to the unknown reproductive potential of mosaic embryos and technical difficulties of its detection from a single embryo biopsy.

Recent Findings: Mosaicism in preimplantation embryos is a product of mitotic errors arising primarily from anaphase lag and chromosome nondisjunction. To date, there is high variability among estimations of prevalence of mosaicism in blastocysts, the most recent ranging from 3.3 to 83%. It has been reported that alleged mosaic embryos can develop into healthy babies, although the proper study evaluating this question remains to be completed. Technical artefacts from comprehensive chromosome screening platforms may also hinder correct classification of embryos as genuine mosaics.

Summary: Although complex, embryonic mosaicism is a phenomenon that deserves further investigation. Many embryos classified as mosaic may have actual reproductive potential. The predictive value of intermediate chromosome copy number assignments for the remaining embryo and for ongoing reproductive potential needs more careful consideration. In addition, recent advancements in extended embryo culture raise the possibility of investigating whether preferential segregation, selective advantage of normal cells or surveillance of abnormal chromosome numbers occur at postimplantation stages.
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http://dx.doi.org/10.1097/GCO.0000000000000358DOI Listing
June 2017

Genomic instability during reprogramming by nuclear transfer is DNA replication dependent.

Nat Cell Biol 2017 04 6;19(4):282-291. Epub 2017 Mar 6.

Department of Pediatrics, Columbia University, New York 10032, USA.

Somatic cells can be reprogrammed to a pluripotent state by nuclear transfer into oocytes, yet developmental arrest often occurs. While incomplete transcriptional reprogramming is known to cause developmental failure, reprogramming also involves concurrent changes in cell cycle progression and nuclear structure. Here we study cellular reprogramming events in human and mouse nuclear transfer embryos prior to embryonic genome activation. We show that genetic instability marked by frequent chromosome segregation errors and DNA damage arise prior to, and independent of, transcriptional activity. These errors occur following transition through DNA replication and are repaired by BRCA1. In the absence of mitotic nuclear remodelling, DNA replication is delayed and errors are exacerbated in subsequent mitosis. These results demonstrate that independent of gene expression, cell-type-specific features of cell cycle progression constitute a barrier sufficient to prevent the transition from one cell type to another during reprogramming.
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http://dx.doi.org/10.1038/ncb3485DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5613662PMC
April 2017

Combination of uterine natural killer cell immunoglobulin receptor haplotype and trophoblastic HLA-C ligand influences the risk of pregnancy loss: a retrospective cohort analysis of direct embryo genotyping data from euploid transfers.

Fertil Steril 2017 Mar 6;107(3):677-683.e2. Epub 2017 Jan 6.

Reproductive Medicine Associates of New Jersey, Basking Ridge, New Jersey; Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania.

Objective: To compare maternal uterine natural killer cell immunoglobulin receptor (KIR) genotype and haplotype frequencies between patients whose euploid single-embryo transfer resulted in pregnancy loss and those that resulted in delivery and to determine if the risk of pregnancy loss was affected by the HLA-C genotype content in the embryo.

Design: Retrospective cohort.

Setting: Academic research center.

Patient(s): Autologous fresh IVF cycles resulting in positive serum β-hCG during 2009-2014.

Intervention(s): None.

Main Outcome Measure(s): 1) Relative risk of pregnancy loss according to maternal KIR genotypes and haplotypes. 2) Comparison of pregnancy loss rates within each KIR haplotype according to HLA-C ligand present in trophectoderm biopsy samples.

Result(s): A total of 668 euploid single-embryo transfers with stored maternal DNA and available preamplification DNA from prior trophectoderm biopsy samples were studied. KIR2DS1, KIR3DS1, and KIR2DS5 were more common in patients who experienced pregnancy loss. Carriers of KIR A haplotype exhibited a decreased risk of pregnancy loss compared with KIR B haplotype carriers. However, among KIR A haplotype carriers, the risk of loss was significantly influenced by whether the transferred embryo carried a C1 allele versus no C1 alleles.

Conclusion(s): KIR A haplotype carriers experienced fewer pregnancy losses than KIR B haplotype carriers after euploid single-embryo transfer. However, this risk was modified by HLA-C alleles present in the embryo. High-risk combinations (KIR A/homozygous C2 and KIR B/homozygous C1) resulted in a 51% increased risk of loss over all other combinations.
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http://dx.doi.org/10.1016/j.fertnstert.2016.12.004DOI Listing
March 2017

Reply: Detecting mosaicism in trophectoderm biopsies.

Hum Reprod 2017 03;32(3):714-715

Reproductive Medicine Associates of New Jersey, 140 Allen Road, Basking Ridge, NJ 07920, USA.

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http://dx.doi.org/10.1093/humrep/dew347DOI Listing
March 2017

High relative deoxyribonucleic acid content of trophectoderm biopsy adversely affects pregnancy outcomes.

Fertil Steril 2017 03 8;107(3):731-736.e1. Epub 2016 Dec 8.

Reproductive Medicine Associates of New Jersey, Basking Ridge, New Jersey; Sidney Kimmel College of Medicine, Thomas Jefferson University, Philadelphia, Pennsylvania.

Objective: To evaluate the association between relative DNA content of the trophectoderm biopsy and pregnancy outcomes.

Design: Retrospective cohort study.

Setting: Academic-affiliated private practice.

Patient(s): This study included patients undergoing their first single embryo transfer after trophectoderm biopsy and comprehensive chromosome screening (CCS) at a single center between January 2010 and February 2014.

Intervention(s): In phase 1 of the study, a standard curve was developed to estimate the relative DNA content of trophectoderm biopsies. Phase 2 of the study examined reproductive outcomes in patients undergoing single embryo transfer after trophectoderm biopsy and CCS. Samples were divided into quartiles according to their relative DNA content, and clinical outcomes were compared.

Main Outcome Measure(s): Chemical pregnancy rate, clinical implantation rate, ongoing pregnancy rate, live birth rate.

Result(s): The quartile of highest relative DNA content had a significantly lower live birth rate when compared with the other three quartiles (relative risk 0.84, 95% confidence interval 0.75-0.95). There was no difference between the quartiles regarding age, body mass index, ovarian response, or endometrial thickness. Among those patients who had a live birth, there was no difference in hCG levels, gestational age at delivery, or birth weight with respect to biopsy DNA content.

Conclusion(s): Trophectoderm biopsies with the highest relative DNA content are associated with lower live birth rates after single embryo transfer. Possible explanations for this phenomenon include diminished accuracy of the euploid diagnosis vs. a mechanical impact of the biopsy. Regardless of the cause, the outcomes emphasize the importance of obtaining appropriately sized trophectoderm biopsies for CCS.
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http://dx.doi.org/10.1016/j.fertnstert.2016.11.013DOI Listing
March 2017
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