Two different microarray technologies for preimplantation genetic diagnosis (PGD) and screening (PGS), due to reciprocal translocation imbalances, demonstrate equivalent clinical pregnancy rates

Next-generation sequencing: challenges in reproductive genetics

Is There a Relationship Between Early Embryo Cleavage and Genetic Abnormalities? Lessons From Preimplantation Genetic Diagnosis (PGD)

Current features of preimplantation genetic diagnosis

Deep impact: sequencing embryo biopsy specimens at increasing depth

Preimplantation genetic diagnosis (PGD) is gradually widely used in prevention of gene diseases and chromosomal abnormalities. Much improvement has been achieved in biopsy technique and molecular diagnosis. Blastocyst biopsy can increase diagnostic accuracy and reduce allele dropout. It is cost-effective and currently plays an important role. Whole genome amplification permits subsequent individual detection of multiple gene loci and screening all 23 pairs of chromosomes. For PGD of chromosomal translocation, fluorescence in-situ hybridization (FISH) is traditionally used, but with technical difficulty. Array comparative genomic hybridization (CGH) can detect translocation and 23 pairs of chromosomes that may replace FISH. Single nucleotide polymorphisms array with haplotyping can further distinguish between normal chromosomes and balanced translocation. PGD may shorten time to conceive and reduce miscarriage for patients with chromosomal translocation. PGD has a potential value for mitochondrial diseases. Preimplantation genetic haplotyping has been applied for unknown mutation sites of single gene disease. Preimplantation genetic screening (PGS) using limited FISH probes in the cleavage-stage embryo did not increase live birth rates for patients with advanced maternal age, unexplained recurrent abortions, and repeated implantation failure. Polar body and blastocyst biopsy may circumvent the problem of mosaicism. PGS using blastocyst biopsy and array CGH is encouraging and merit further studies. Cryopreservation of biopsied blastocysts instead of fresh transfer permits sufficient time for transportation and genetic analysis. Cryopreservation of embryos may avoid ovarian hyperstimulation syndrome and possible suboptimal endometrium.

Diagnóstico genético pré‐implantacional e sua aplicação na reprodução humana assistida

Preimplantation Genetic Diagnosis (PGD) is currently an alternative for couples with high risk of pregnancies with genetic anomalies; it offers the possibility of avoiding the need to terminate affected pregnancies, since it allows the selection of unaffected embryos for transfer. PGD for inherited disorders has become extremely accurate (99.5%), and may currently be performed for any single gene disorders in which mutation is identified. PGD has been performed for more than 100 different conditions resulting in the birth of at least 1000 healthy children free of genetic disorder. PGD is presently also used together with preimplantation HLA typing for treatment of affected sibling with genetic and acquired disorders requiring HLA matched stem cell transplantation. This is not only to allow couples to have an unaffected child but also to select a potential donor progeny for stem cell transplantation. In Turkey, thalassemia is the most commonly seen genetic disorder the rate of thalassemia carriers is about 3–4% in Turkey. The majority of our PGD cases are thalassemia carriers. They do not only require thalassemia mutation analysis but also HLA typing for their affected child. In this study PGD results of 236 Turkish couples with or without HLA typing will be presented and discussed. A full diagnosis was achieved in 91.0% of the biopsied samples. In Group I, 17.8% of the analyzed embryos were found to be HLA compatible. HLA compatible and disease free embryos were 12.9% of all diagnosed embryos. In group II, 17.2% of embryos were found to be HLA matched and 71.4% HLA non-matched. The majority of our HLA typing combined with PGD cases were β-Thalassemia carriers (87.9%). The mutations analyzed have high heterogeneity, the most frequent mutation was IVS-I-110 G-A and comprised 46.2% of all mutations. To date, 70 healthy and HLA compatible children have been born. Twenty-five sick children have already been cured with cord blood cell and/or bone marrow transplantation. Twenty-one children are waiting for their newborn siblings to gain sufficient weight and maturity for the donation of stem cells. The successful transplantations have been performed for the following indications: β- Thalassemia (n = 19), Wiskott Aldrich syndrome (n = 2), Glanzmann Disease (n = 1), X-Adrenoleukodystrophy (n=1) and acute myeloid leukemia (n = 1) and Diamond Blackfan anemia (n = 1). This data presents one of the world’s largest experiences on preimplantation HLA typing, and the outcome of stem cell transplantation is the largest number available from one center. Our results indicate HLA typing with or without mutation analysis is a promising and effective therapeutic tool for curation of an affected sibling.

Background: Preimplantation genetic diagnosis (PGD) is currently used for over 400 monogenic diseases. Some endocrine conditions that occur due to monogenic defects are either life threatening or can cause severe morbidities; thus, PGD may be an option to avoid the occurrence of such diseases. But a high cost and limited availability of licensed IVF centers can be a barrier for many patients who might otherwise benefit from PGD. Objective: To review the literature for endocrine monogenic disorders that have been prevented with PGD. Methods: An initial search in PubMed/Medline search was done to identify monogenic endocrine conditions using MeSH term “preimplantation diagnosis” and other searches using “preimplantation genetic diagnosis (PGD)”, plus “endocrine” or “monogenic” anywhere in the title or abstract. Studies were eligible for consideration if they included use of PGD for any known monogenic endocrine disease. Studies were omitted if the disease was non-endocrine or PGD was not used for embryo selection. Eleven articles (1999-2018) reported 15 cases using PGD for monogenic endocrine diseases. These were Congenital adrenal hyperplasia (n=2), X-linked adrenoleukodystrophy (n=2), Congenital lipoid adrenal hyperplasia (n=1), Persistent hyperinsulinemic hypoglycemia of infancy (n=4), Pseudohypoparathyroidism type 1a (n=1), Hypoparathyroidism-retardation-dysmorphism syndrome (n=2) and Multiple endocrine neoplasia types 1 (n=1) and 2a (n=2). Techniques used for PGD included PCR, whole genome amplification, FISH and more recently next generation sequencing. Clinical and outcome data of these cases were reviewed with respect to number of PGD cycles, successful pregnancy rates, live births and their genetic status. Results: Fifteen couples underwent 32 PGD cycles (ranging from 1-9 per couple), of which 17 cycles resulted in a pregnancy. Seven couples underwent a single PGD cycle. Four of these 7 couples had successful pregnancies each resulting in live births (all singletons), 1 couple was unable to achieve pregnancy, 1 chose to terminate the pregnancy as the karyotype detected a Turner fetus (45XO) and outcome data was not reported in one. The remaining 8 couples underwent multiple PGD cycles (2-9 per couple) and all had successful pregnancies in at least one cycle, resulting in 16 livebirths (6 singletons and 5 twins). Amongst the total live births (n=20), 60% were genetically unaffected for the tested monogenic disorder and 40% were carriers of the autosomal recessive gene mutation for which PGD was performed. Conclusion: Preimplantation genetic diagnosis may be a potential tool for preventing the inheritance of severe monogenic endocrine diseases in future generations. The current use of PGD in endocrine disorders is rare, but provides a promising option on a case by case basis provided the optimal resources are available.

Preimplantation genetic diagnosis (PGD) consists of diagnostic ­procedures detecting a genetic condition(s) in the oocyte or embryo produced by in vitro fertilization (IVF) prior to pregnancy. Chromosomal abnormalities and single gene disorders can be tested by PGD, which gives the parents the opportunity to choose unaffected embryos for transfer. Establishment of affected human embryonic stem cell (hESC) lines from affected preembryos with genetic disorders diagnosed by PGD provides a powerful research tool for exploring fundamental biological mechanisms of early stages of development. This, in turn, leads to the development of new approaches for diagnosing, treating, and preventing genetic disorders. Human embryonic stem cell (hESC) lines are pluripotent and can ­produce all types of cell lineages in the body. Considering the presence of genetic diversities and polymorphisms in populations, there is a need for large hESC line collections to provide the various genetic components for research purposes. There are several advantages and a uniqueness of hESC research. It cannot be done in animal models or with cell culture methods. Also, it has an invaluable place in pharmacogenomic testing and regenerative medicine applications as affected hESCs of some disorders are the only biological tools we have as disease models.

Preimplantation Genetic Diagnosis (PGD) is the screening of embryos at the cleavage stage in order to select and transfer only the desired embryo. The main indications of PGD are monogenic disorders and aneuploidy. Additionally, PGD can be employed to distinguish a male from a female embryo or to choose a HLA-matched embryo to a child requiring bone marrow or cord blood transplant. The European Society of Human Reproduction and Embryology classifies PGD into two categories (Hum Reprod 24: 1786–810, 2009): high-risk PGD for patients at high risk of transmitting a genetic or chromosomal abnormality to their children, which includes single gene defects, namely autosomal recessive, autosomal dominant, and X-linked disorders, as well as chromosomal abnormalities (translocations, small deletions, etc.); and low-risk PGD (or PGS for Preimplantation Genetic Screening) for sex selection, and for infertile patients undergoing IVF with the aim of increasing the IVF pregnancy rates. Patients that fall into this category are those of advanced maternal age and repeated IVF failure and couples with normal karyotypes who have experienced repeated miscarriages.

Six-year experience of preimplantation genetic diagnosis (PGD) for single gene disorders and chromosomal abnormalities at CHA Institute

Preimplantation genetic diagnosis (PGD) is a technique used to test embryos created by in vitro fertilization for disorders caused by inherited mutations in single genes. Embryos free from the germline mutation identified in the couple are transferred into the uterus to allow implantation and pregnancy. PGD enables couples with inherited disorders a means of ensuring that their children are unaffected by the disease without the need of termination of affected pregnancies. PGD for inherited susceptibilities to cancer encompasses a diverse group of genes with variable degrees of penetrance and expressivity often making genotype/phenotype relationships hard to establish. This is one of the factors that makes PGD a preferred option to potential parents compared to prenatal diagnosis and termination for this group of disorders.

Preimplantation genetic diagnosis (PGD) relates exclusively to in vitro fertilisation techniques (IVF) that aim to prevent transmission of a serious genetic abnormality to the child. The genetic characteristics of the embryo created through IVF are analysed, and only the embryos free of the genetic abnormality are implanted in the womb. Performed worldwide since 1990, this technique has raised many legal and ethical debates due to the very wide variations of lawgiving between countries. This is shown by the report of the UNESCO IBC (2003), which described the techniques and the issues raised by preimplantation genetic diagnosis. In this article, the authors present the differences between prenatal diagnosis and preimplantation genetic diagnosis, the French legislation, then the range of legislation in Europe and finally the position of the European Court of Human Rights which sanctioned Italy and Latvia for refusing access to PGD.

Preimplantation genetic diagnosis and screening are widely accepted for chromosomal abnormality identification to avoid transferring embryos with genetic defects. Massively parallel sequencing (MPS) is a rapidly developing approach for genome analysis with increasing application in clinical practice. The purpose of this study was to use MPS for identification of aneuploidies and unbalanced chromosomal rearrangements after blastocyst biopsy. Trophectoderm (TE) samples of 38 blastocysts from 16 in vitro fertilization cycles were subjected to analysis. Low-coverage whole genome sequencing was performed using the Illumina HiSeq2000 platform with a novel algorithm purposely created for chromosomal analysis. The efficiency of this MPS approach was estimated by comparing results obtained by an Affymetrix single-nucleotide polymorphism (SNP) array. Whole genome amplification (WGA) products of TE cells were detected by MPS, with an average of 0.07× depth and 5.5% coverage of the human genome. Twenty-six embryos (68.4%) were detected as euploid, while six embryos (15.8%) contained uniform aneuploidies. Four of these (10.5%) were with solely unbalanced chromosomal rearrangements, whereas the remaining two embryos (5.3%) showed both aneuploidies and unbalanced rearrangements. Almost all these results were confirmed by the SNP array, with the exception of one sample, where different sizes of unbalanced rearrangements were detected, possibly due to chromosomal GC bias in array analysis. Our study demonstrated MPS could be applied to accurately detect embryonic chromosomal abnormality with a flexible and cost-effective strategy and higher potential accuracy.

Idiopathic recurrent miscarriage is caused mostly by aneuploid embryos