Laboratory of Genetic Rare Diseases EA 7402 University of Montpellier |
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Update: 19-12-2016 |
Preimplantation Genetic Diagnosis for monogenic disorders | |||
The intented goal of Preimplantation Genetic Diagnosis (PGD) is to diagnose a specific genetic disease on single cells from embryos obtained through an in vitro fertilization (IVF) procedure, by selecting and transferring to the woman's uterus only embryos found to be unaffected, before a clinical pregnancy has been established. Genetic diagnosis is performed on one or two blastomeres biopsied from 3 days-old embryos, then embryos free of the disease under investigation are transferred to the mother on day 4 or 5. | |||
PGD cycle |
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PGD indications PGD indications are quite similar to those practiced in prenatal diagnosis, i.e serious hereditary disorders that may result in a therapeutic abortion following prenatal diagnosis of an affected fetus. According to european recommendations, couples requesting information about PND may also be counselled about PGD, limits and errors related to PGD, as well as the possibility to perform PND to confirm PGD results.
The PGD procedure is applied to monogenic disorders, that are classified into four main categories :
Risks and pitfalls
The study of specific DNA sequences in a single blastomere is a substantial challenge as many problems may compromise both the accuracy and reliability of the genetic diagnosis. For the detection of single gene defects, the prevalent method of diagnosis is PCR (polymerase chain reaction) allowing the exponential amplification of short DNA sequences, which are subsequently detected and analysed. Preclinical work-up Before clinical application, extensive preclinical trials have to be performed on several single cells isolated under an inverted microscope (lymphocytes or lymphoblasts), including the development of specific PCR protocols as well as techniques chosen to identify mutations and polymorphic markers. Otherwise, the aim of these preclinical experiments is to evaluate amplification efficiencies and ADO rates for all the DNA sequences to be studied in the procedure. For most of PGD indications, the development, optimization and validation procedures can take several weeks to several months, according to the genes studied and techniques used. Work-up results on single cells are considered satisfactory for clinical application when they fall within the limits set in the ESHRE PGD consortium's guidelines : amplification efficiencies may be above 90% and ADO rates may be lower than 10%. However, single cell amplification efficiencies and ADO rates can vary greatly with different cell types, especially in blastomeres from poor embryo quality that frequently exhibit inconclusive results. Available PGD indications for monogenic disorders in the CHU of Montpellier
In France, some PGD indications are only performed in our center (hemolytic disease, aniridia, molybdenum cofactor deficiency, vascular Ehlers-Danlos syndrome, Carvajal syndrome…). For other indications (such as cystic fibrosis and Duchenne muscular dystrophy), we have developed powerful protocols in order to accept in our PGD program most of the couples requesting PGD for these conditions. Finally, for other families, we have set up techniques enabling to distinguish healthy embryos from monosomic embryos (for the chromosome studied). Thus, the embryologist may select for transfer, among a large cohort of unaffected embryos, the embryo(s) displaying the best morphology and kinetics of development. Consequently, the rates of pregnancy are improved. |
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Anne GIRARDET (PhD, MCU-PH CHU Montpellier) Aliya ISHMUKHAMETOVA (PhD, PAA CHU Montpellier) Stéphanie PLAZA (PhD, Engineer CHU Montpellier) Sandie MEREUZE (PhD, Engineer CHU Montpellier) Victoria VIART (PhD, Engineer CHU Montpellier) Florielle SAGUET (TL CHU Montpellier) Garance VERRIERE (TL CHU Montpellier) |
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Girardet A, Ishmukhametova A, Viart V, Plaza S, Saguet F, Verriere G, Hamamah S, Coupier I, Haquet E, Anahory T, Willems M, Claustres M. Thirteen years' experience of 893 PGD cycles for monogenic disorders in a publicly funded, nationally regulated regional hospital service. Reprod Biomed Online. 2018 Feb;36(2):154-163. PMID: 29203382 |