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OBJECTIVE: To evaluate the technical accuracy, inheritance, and pathogenicity of small copy number variants (CNVs) detected by a targeted next-generation sequencing-based preimplantation genetic testing for aneuploidy (PGT-A) platform. DESIGN: Retrospective observational study performed between 2020 and 2022. SETTING: Clinic. PATIENT(S): A total of 12,157 patients who underwent clinical PGT-A performed by targeted next-generation sequencing for whole chromosome and large segmental aneuploidies. INTERVENTION(S): An incidental finding was reported when a CNV gain/loss of at least 3 consecutive amplicons appeared in at least 2 embryos from the same in vitro fertilization cycle. MAIN OUTCOME MEASURE(S): The primary outcome measures were the specificity, incidence, inheritance, and pathogenicity of small CNVs detected by the PGT-A platform. Accuracy of the PGT-A platform CNV calls was assessed via concordance with the CNV calls (size and genomic location) on chromosomal microarray of the gamete provider(s). Parental origin of the CNV and pathogenicity classifications were also reported. RESULT(S): An incidental finding that met reporting criteria was identified in 75 (0.62%; 95% confidence interval, 0.5%-0.8%) of 12,157 unique PGT-A patients. Chromosomal microarray follow-up was requested for all cases, and results were received for 1 or both members of 65 reproductive couples. In all cases, 1 of the gamete providers was confirmed to have the CNV identified in the embryos (100.0%, N = 65/65; 95% confidence interval, 94.5-100). The identified CNV was of maternal origin in 34 cases (52.3%) and of paternal origin in 31 cases (47.7%). A significant correlation was identified between PGT-A-predicted CNV sizes and chromosomal microarray detected sizes (r = 0.81) and genomic coordinates on parental deoxyribonucleic acid. Twenty-six (40%) of the CNVs were classified as benign/likely benign, 30 (46.2%) as a variant of uncertain significance, and 9 (13.8%) as pathogenic/likely pathogenic. CONCLUSION(S): Certain PGT-A platforms may enable the detection of inherited, small CNVs with extremely high specificity without prior knowledge of parental status. Most CNVs in this data set were confirmed to be benign/likely benign or a variant of uncertain significance. Pathogenic/likely pathogenic CNVs associated with a broad range of phenotypic features may also be detected, although a reliable negative predictive value for small CNVs with current PGT-A technologies is unknown because of the many technical challenges.
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During IVF treatments, normal fertilization is generally evidenced by the appearance of two pronuclei, one arising from the oocyte and the other from the male gamete. Embryos derived from zygotes with a pronuclei number other than two are assumed to possess a ploidy abnormality and their transfer is usually avoided owing to increased risk of implantation failure, miscarriage, and molar pregnancies. Nonetheless, the inclusion of genotyping data in preimplantation genetic testing has revealed that a normal diploid configuration is possible in embryos deriving from zygotes with an abnormal pronuclei number such as tripronuclear and one pronucleus. Here, we present a one-of-a-kind transfer of a tetrapronuclear-derived embryo that was discovered to be diploid and negative for other whole chromosome or segmental aneuploidies during preimplantation genetic testing using a targeted next-generation sequencing approach. The transfer resulted in the live birth of a healthy infant who is now 4 years old and has no apparent health or developmental impairments.
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Nacimiento Vivo , Diagnóstico Preimplantación , Embarazo , Humanos , Femenino , Masculino , Fertilización In Vitro/métodos , Diagnóstico Preimplantación/métodos , Transferencia de Embrión , Pruebas Genéticas/métodos , Aneuploidia , BlastocistoRESUMEN
Segmental aneuploidies (SAs) are structural imbalances, namely, gains or losses, involving a chromosomal segment. Most preimplantation genetic testing platforms can detect segmental imbalances greater than 5-10 Mb, either full or mosaic; however, questions remain about clinical significance. An in-depth review was performed to determine the accuracy, frequency, and types of SAs detected in preimplantation embryos. A comprehensive search of the literature revealed an incidence of approximately 8.15% in preimplantation embryos, compared with a prevalence of 3.55% in prenatal diagnosis samples. Several studies have used rebiopsy analysis to validate the accuracy and reproducibility of such findings in blastocyst-stage embryos. A comparison of these studies yielded a mean confirmation rate of SAs slightly higher than 30%. This result could be attributed to their mitotic origin as well as to the technical limitations of preimplantation genetic testing. In addition, the few available studies in which embryos with a segmental finding were transferred in utero are analyzed to discuss the reproductive competence of such embryos. Except for 1 study, all outcomes were described for segmental embryos in a mosaic state. As a result, there is still insufficient evidence to provide accurate information about the effect of segmental imbalances on embryonic reproductive competence and to determine gestational and newborn risks.
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Diagnóstico Preimplantación , Femenino , Humanos , Recién Nacido , Embarazo , Aneuploidia , Blastocisto/patología , Pruebas Genéticas , Reproducibilidad de los ResultadosRESUMEN
Preimplantation genetic testing for aneuploidy (PGT-A) is used as a frequent add-on for in-vitro fertilization (IVF) to improve clinical outcomes. The purpose is to select a euploid embryo following chromosomal testing on embryo biopsies. The current practice includes comprehensive chromosome screening (CCS) technology applied on trophectoderm (TE) biopsies. Despite its widespread use, PGT-A remains a controversial topic mainly because all of the RCTs comprised only good prognosis patients with 2 or more blastocysts available; hence the results are not generalizable to all groups of patients. Furthermore, with the introduction of the highly-sensitive platforms into clinical practice (i.e. next-generation sequencing [NGS]), a result consistent with intermediate copy number surfaced and is termed "Mosaic," consistent with a mixture of euploid and aneuploid cells within the biopsy sample. The optimal disposition and management of embryos with mosaic results is still an open question, as many 'mosaics' generated healthy live births with no identifiable congenital anomalies. The present article provides a complete and comprehensive up-to-date review on PGT-A. It discusses in detail the findings of all the published RCTs on PGT-A with CCS, comments on the subject of "mosaicism" and its current management, and describes the latest technique of non-invasive PGT-A.
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Diagnóstico Preimplantación , Embarazo , Femenino , Humanos , Diagnóstico Preimplantación/métodos , Pruebas Genéticas/métodos , Aneuploidia , Blastocisto/patología , MosaicismoRESUMEN
PURPOSE: Carrier screening (CS) is a term used to describe a genetic test performed on individuals without family history of genetic disorders, to investigate the carrier status for pathogenic variants associated with multiple recessive conditions. The advent of next-generation sequencing enabled simultaneous CS for an increasing number of conditions; however, a consensus on which diseases to include in gene panels and how to best develop the provision of CS is far to be reached. Therefore, the provision of CS is jeopardized and inconsistent and requires solving several important issues. METHODS: In 2020, the Italian Society of Human Genetics (SIGU) established a working group composed of clinical and laboratory geneticists from public and private fields to elaborate a document to define indications and best practice of CS provision for couples planning a pregnancy. RESULTS: Hereby, we present the outcome of the Italian working group's activity and compare it with previously published international recommendations (American College of Medical Genetics and Genomics (ACMG), American College of Obstetricians and Gynecologists (ACOG), and Royal Australian and New Zealand College of Obstetricians and Gynaecologists (RANZCOG)). We determine a core message on genetic counseling and nine main subject categories to explore, spanning from goals and execution to technical scientific, ethical, and socio-economic topics. Moreover, a level of agreement on the most critical points is discussed using a 5-point agreement scale, demonstrating a high level of consensus among the four societies. CONCLUSIONS: This document is intended to provide genetic and healthcare professionals involved in human reproduction with guidance regarding the clinical implementation of CS.
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Asesoramiento Genético , Pruebas Genéticas , Embarazo , Femenino , Humanos , Australia , Personal de Salud , ReproducciónRESUMEN
The most important factor associated with oocytes' developmental competence has been widely identified as the presence of chromosomal abnormalities. However, growing application of genome-wide sequencing (GS) in population diagnostics has enabled the identification of multifactorial genetic predispositions to sub-lethal pathologies, including those affecting IVF outcomes and reproductive fitness. Indeed, GS analysis in families with history of isolated infertility has recently led to the discovery of new genes and variants involved in specific human infertility endophenotypes that impact the availability and the functionality of female gametes by altering unique mechanisms necessary for oocyte maturation and early embryo development. Ongoing advancements in analytical and bioinformatic pipelines for the study of the genetic determinants of oocyte competence may provide the biological evidence required not only for improving the diagnosis of isolated female infertility but also for the development of novel preventive and therapeutic approaches for reproductive failure. Here, we provide an updated discussion and review of the progresses made in preconception genomic medicine in the identification of genetic factors associated with oocyte availability, function, and competence.
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Infertilidad , Oocitos , Desarrollo Embrionario , Femenino , Genómica , Humanos , Oogénesis/genéticaRESUMEN
Prostate cancer (PCa) is a slow-growing tumor representing one of the major causes of all new cancer cases and cancer mortality in men worldwide. Although screening methods for PCa have substantially improved, the outcome for patients with advanced PCa remains poor. The elucidation of the molecular mechanism that drives the progression from a slow-growing, organ-confined tumor to a highly invasive and castration-resistant PCa (CRPC) is therefore important. We have already proved the diagnostic potential of indoleamine-2,3-dioxygenase (IDO) when detected in urine of individuals at risk of developing PCa. The aim of this study was to implement IDO as a prognostic marker for PCa patients undergoing surgical treatment. We have thus conducted an observational study by collecting 100 urine samples from patients undergoing radical prostatectomy as first treatment of choice. To test the integrity of our investigation, scale dilution cells of an established PC3 cell line were added to urine of healthy donors and used for gene expression analysis by a TaqMan assay on the catalytic part of IDO mRNA. Our data show that the quantification of IDO mRNA in urine of patients has a very promising ability to identify patients at high risk of cancer advancement, as defined by Gleason score. Our goal is to lay the groundwork to develop a superior test for PCa. The data generated are thus necessary (i) to strengthen the IDO-based diagnostic/prognostic test and (ii) to provide patients and clinicians with an affordable and easy screening test.