Recently, exome sequencing led to the identification of causal mutations in 16-31% of patients with intellectual disability (ID), leaving the underlying cause for many patients unidentified. In this ...context, the noncoding part of the human genome remains largely unexplored. For many long non-coding RNAs (lncRNAs) a crucial role in neurodevelopment and hence the human brain is anticipated. Here we aimed at identifying lncRNAs associated with neuronal development and ID. Therefore, we applied an integrated genomics approach, harnessing several public epigenetic datasets. We found that the presence of neuron-specific H3K4me3 confers the highest specificity for genes involved in neurodevelopment and ID. Based on the presence of this feature and GWAS hits for CNS disorders, we identified 53 candidate lncRNA genes. Extensive expression profiling on human brain samples and other tissues, followed by Gene Set Enrichment Analysis indicates that at least 24 of these lncRNAs are indeed implicated in processes such as synaptic transmission, nervous system development and neurogenesis. The bidirectional or antisense overlapping orientation relative to multiple coding genes involved in neuronal processes supports these results. In conclusion, we identified several lncRNA genes putatively involved in neurodevelopment and CNS disorders, providing a resource for functional studies.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Shallow-depth sequencing of cell-free DNA, a cheap and standardized approach to obtain molecular information on tumors non-invasively, is insufficiently explored for lymphoma diagnosis and disease ...follow-up. This study collected 318 samples, including longitudinal liquid and paired solid biopsies, from a prospectively recruited cohort of 38 Hodgkin lymphoma (HL) and 85 aggressive B-cell non- HL patients, represented by 81 diffuse large B-cell lymphoma (DLBCL) cases. Following sequencing, copy number alterations and viral read fractions were derived and analyzed. At diagnosis, liquid biopsies showed detectable copy number alterations in 84.2% of HL (88.6% for classical HL) and 74.1% of DLBCL patients. Copy number profiles between liquid-solid pairs were highly concordant within DLBCL (r=0.815±0.043); and, compared to tissue, HL liquid biopsies had abnormalities with higher amplitudes (P=.010), implying that tumor DNA is more abundant in plasma. Additionally, 39.5% of HL and 13.6% of DLBCL cases had a significantly elevated number of plasmatic Epstein-Barr virus DNA fragments, achieving a sensitivity of 100% compared to current standard. Longitudinal analysis determined that, when detectable, copy number patterns were similar across (re)staging moments in refractory/relapsed patients. Moreover, the overall profile anomaly highly correlated with the total metabolic tumor volume (P.
Purpose
Providing additional insights on the efficacy of human nuclear transfer (NT). Here, and earlier, NT has been applied to minimize transmission risk of mitochondrial DNA (mtDNA) diseases. NT ...has also been proposed for treating infertility, but it is still unclear which infertility indications would benefit. In this work, we therefore additionally assess the applicability of NT to overcome failed fertilization.
Methods
Patient 1 carries a homoplasmic mtDNA mutation (m.11778G > A). Seventeen metaphase II (MII) oocytes underwent pre-implantation genetic testing (PGT), while five MII oocytes were used for spindle transfer (ST), and one in vitro matured (IVM) metaphase I oocyte underwent early pronuclear transfer (ePNT). Patients 2–3 experienced multiple failed intracytoplasmic sperm injection (ICSI) and ICSI-assisted oocyte activation (AOA) cycles. For these patients, the obtained MII oocytes underwent an additional ICSI-AOA cycle, while the IVM oocytes were subjected to ST.
Results
For patient 1, PGT-M confirmed mutation loads close to 100%. All ST-reconstructed oocytes fertilized and cleaved, of which one progressed to the blastocyst stage. The reconstructed ePNT-zygote reached the morula stage. These samples showed an average mtDNA carry-over rate of 2.9% ± 0.8%, confirming the feasibility of NT to reduce mtDNA transmission. For patient 2–3 displaying fertilization failure, ST resulted in, respectively, 4/5 and 6/6 fertilized oocytes, providing evidence, for the first time, that NT can enable successful fertilization in this patient population.
Conclusion
Our study showcases the repertoire of disorders for which NT can be beneficial, to overcome either mitochondrial disease transmission or failed fertilization after ICSI-AOA.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
To study the feasibility of in vitro maturation of ovarian tissue oocytes for fertility preservation in transgender men on testosterone treatment.
Cross-sectional study
University hospital
...Eighty-three transgender men enrolled from November 2015 to January 2019
In vitro maturation of cumulus-oocyte complexes (COCs) harvested at the time of gender confirmation surgery, and fertilization through intracytoplasmic sperm injection.
In vitro maturation, fertilization, and blastulation rates; comparison of morphokinetics with vitrified-warmed oocytes; and analysis of the genetic profiles of embryos. Secondary outcomes: association between serum hormone levels; COCs’ morphologic characteristics, and vitrification rate.
All participants were on testosterone treatment for a median of 83 (64Quartile 1; 113.2Quartile 2) weeks. A total of 1,903 COCs (mean per participant, 23 ± 15.8) were collected. The in vitro maturation rate was 23.8%, vitrification rate was 21.5%, and survival rate after warming was 72.6% (n = 151). Intracytoplasmic sperm injection was performed in 139 oocytes. The rate of normal fertilized oocytes was 34.5%, and 25 (52.1%) embryos reached day 3. One blastocyst was achieved on day 5. Aberrant cleavage patterns and early embryo arrest were observed in 22 (45.8%) and 44 (91.7%) zygotes, respectively. Compared with vitrified-warmed donor oocytes, a delay was observed in pronuclei disappearance, t2 (time to reach 2 cell stage) timings, and CC1 (the duration of the 1st cell cycle) and SS3 (synchronization of cleavage pattern (calculated as t8-t5) time intervals. A normal genetic pattern was seen in 42% embryos. The proportion of vitrified oocytes was negatively associated with progesterone (odds ratio, 0.76) and positively associated with antimüllerian hormone serum levels (odds ratio, 1.23). The highest vitrification rate was achieved by the morphologic characteristic 344 at day 0 and by 433 at day 2.
Ovarian tissue oocytes matured in vitro show low developmental capacity in transgender men, when collected under testosterone treatment.
Baja eficiencia de los ovocitos madurados in vitro provenientes de complejos de Cumulus recuperados durante la preparación de tejido ovárico en el momento de la cirugía de reafirmación de género y bajo tratamiento con testosterona para preservación de fertilidad en hombres transgénero.
Estudiar la viabilidad de la maduración in vitro de ovocitos de tejido ovárico para preservación de fertilidad en varones transgénero en tratamiento con testosterona.
Estudio de muestra transversal.
Hospital universitario.
Se incluyeron 83 varones transgénero entre Noviembre de 2015 y Enero de 2019.
Maduración in vitro de complejos cúmulo-ovocitos (COCs) recuperados en el momento de la cirugía de confirmación de género y fertilización mediante inyección intracitoplasmática de espermatozoides.
Tasas de maduración in vitro, fertilización y blastulación; comparación de morfokinética con ovocitos vitrificados y desvitrificados y análisis del perfil genético de los embriones. Resultados secundarios: Relación entre los niveles hormonales séricos, características morfokinéticas y tasa de vitrificación.
Todos los participantes habían estado en tratamiento con testosterona durante una media de 83 semanas (64Cuartil 1; 113.2Cuartil 2). Se recuperaron un total de 1,903 COCs (23 ± 15.8 de media por participante). La tasa de maduración in vitro fue del 23.8%, la tasa de vitrificación fue de 21.5% y la tasa de supervivencia a la descongelación fue del 72.6% (n=151). Se realizó inyección intracitoplasmática de espermatozoides en 130 ovocitos. La tasa de ovocitos con fertilización normal fue del 34.5% y 25 (52.1%) de los embriones llegaron a día 3. Se logró un blastocisto en día 5. Se observaron patrones de división aberrante y bloqueo embrionario precoz en 22 (45.8%) y 44 (91.7%) de los cigotos respectivamente. Comparados con los ovocitos de donante vitrificados-desvitrificados, se observó un retraso en la desaparición de los pronúcleos, t2 (tiempo para alcanzar estadio de 2 células) y CC1 (la duración del primer ciclo celular) y SS3 (sincronización del patrón de tiempo de los intervalos de división) calculado como t8-t5. Se observó un patrón genético normal en el 42% de los embriones. La proporción de ovocitos vitrificados se relacionó negativamente con los niveles de progesterona (odds ratio 0.76) y positivamente con los niveles séricos de hormona antimulleriana (odds ratio 1.23). La tasa más alta de vitrificación se logró con características morfológicas 344 en el día 0 y con 433 en día 2.
Los ovocitos de tejido ovárico madurados in vitro muestran baja capacidad de desarrollo en varones transgénero cuando se recuperan bajo tratamiento con testosterona.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Noninvasive prenatal screening (NIPS) using cell-free DNA has transformed prenatal care. Belgium was the first country to implement and fully reimburse NIPS as a first-tier screening test offered to ...all pregnant women. A consortium consisting of all Belgian genetic centers report the outcome of two years genome-wide NIPS implementation.
The performance for the common trisomies and for secondary findings was evaluated based on 153,575 genome-wide NIP tests. Furthermore, the evolution of the number of invasive tests and the incidence of Down syndrome live births was registered.
Trisomies 21, 18, and 13 were detected in respectively 0.32%, 0.07%, and 0.06% of cases, with overall positive predictive values (PPVs) of 92.4%, 84.6%, and 43.9%. Rare autosomal trisomies and fetal segmental imbalances were detected in respectively 0.23% and 0.07% of cases with PPVs of 4.1% and 47%. The number of invasive obstetric procedures decreased by 52%. The number of trisomy 21 live births dropped to 0.04%.
Expanding the scope of NIPS beyond trisomy 21 fetal screening allows the implementation of personalized genomic medicine for the obstetric population. This genome-wide NIPS approach has been embedded successfully in prenatal genetic care in Belgium and might serve as a framework for other countries offering NIPS.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Chromothripsis represents a novel phenomenon in the structural variation landscape of cancer genomes. Here, we analyze the genomes of ten patients with congenital disease who were preselected to ...carry complex chromosomal rearrangements with more than two breakpoints. The rearrangements displayed unanticipated complexity resembling chromothripsis. We find that eight of them contain hallmarks of multiple clustered double-stranded DNA breaks (DSBs) on one or more chromosomes. In addition, nucleotide resolution analysis of 98 breakpoint junctions indicates that break repair involves nonhomologous or microhomology-mediated end joining. We observed that these eight rearrangements are balanced or contain sporadic deletions ranging in size between a few hundred base pairs and several megabases. The two remaining complex rearrangements did not display signs of DSBs and contain duplications, indicative of rearrangement processes involving template switching. Our work provides detailed insight into the characteristics of chromothripsis and supports a role for clustered DSBs driving some constitutional chromothripsis rearrangements.
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► Constitutional complex chromosomal rearrangements display unanticipated complexity resembling chromothripsis ► Some chromothripsis rearrangements involve clustered double-stranded DNA breaks ► There exist distinct classes of chromothripsis rearrangements
Complex genomic rearrangements are associated with birth defects and cancer development. Kloosterman, Cuppen, and colleagues use whole-genome sequencing analysis to unravel the precise structure of ten complex genomic rearrangements in patients with congenital malformations. Eight of the rearrangements are caused by chromosome shattering and nonhomologous DNA repair, while two rearrangements involve replicative repair processes. These rearrangements represent two different instances of chromothripsis. The authors show that chromosome shattering and repair may frequently underlie complex genomic rearrangements causing developmental defects.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Objective To establish the value of array comparative genomic hybridization (CGH) for preimplantation genetic diagnosis (PGD) in embryos of translocation carriers in combination with vitrification ...and frozen embryo transfer in nonstimulated cycles. Design Retrospective data analysis study. Setting Academic centers for reproductive medicine and genetics. Patient(s) Thirty-four couples undergoing PGD for chromosomal rearrangements from October 2013 to December 2015. Intervention(s) Trophectoderm biopsy at day 5 or day 6 of embryo development and subsequently whole genome amplification and array CGH were performed. Main Outcome Measure(s) This approach revealed a high occurrence of aneuploidies and structural rearrangements unrelated to the parental rearrangement. Nevertheless, we observed a benefit in pregnancy rates of these couples. Result(s) We detected chromosomal abnormalities in 133/207 embryos (64.2% of successfully amplified), and 74 showed a normal microarray profile (35.7%). In 48 of the 133 abnormal embryos (36.1%), an unbalanced rearrangement originating from the parental translocation was identified. Interestingly, 34.6% of the abnormal embryos (46/133) harbored chromosome rearrangements that were not directly linked to the parental translocation in question. We also detected a combination of unbalanced parental-derived rearrangements and aneuploidies in 27 of the 133 abnormal embryos (20.3%). Conclusion(s) The use of trophectoderm biopsy at the blastocyst stage is less detrimental to the survival of the embryo and leads to a more reliable estimate of the genomic content of the embryo than cleavage-stage biopsy. In this small cohort PGD study, we describe the successful implementation of array CGH analysis of blastocysts in patients with a chromosomal rearrangement to identify euploid embryos for transfer.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Genomic disorders are often caused by recurrent copy number variations (CNVs), with nonallelic homologous recombination (NAHR) as the underlying mechanism. Recently, several microhomology-mediated ...repair mechanisms--such as microhomology-mediated end-joining (MMEJ), fork stalling and template switching (FoSTeS), microhomology-mediated break-induced replication (MMBIR), serial replication slippage (SRS), and break-induced SRS (BISRS)--were described in the etiology of non-recurrent CNVs in human disease. In addition, their formation may be stimulated by genomic architectural features. It is, however, largely unexplored to what extent these mechanisms contribute to rare, locus-specific pathogenic CNVs. Here, fine-mapping of 42 microdeletions of the FOXL2 locus, encompassing FOXL2 (32) or its regulatory domain (10), serves as a model for rare, locus-specific CNVs implicated in genetic disease. These deletions lead to blepharophimosis syndrome (BPES), a developmental condition affecting the eyelids and the ovary. For breakpoint mapping we used targeted array-based comparative genomic hybridization (aCGH), quantitative PCR (qPCR), long-range PCR, and Sanger sequencing of the junction products. Microhomology, ranging from 1 bp to 66 bp, was found in 91.7% of 24 characterized breakpoint junctions, being significantly enriched in comparison with a random control sample. Our results show that microhomology-mediated repair mechanisms underlie at least 50% of these microdeletions. Moreover, genomic architectural features, like sequence motifs, non-B DNA conformations, and repetitive elements, were found in all breakpoint regions. In conclusion, the majority of these microdeletions result from microhomology-mediated mechanisms like MMEJ, FoSTeS, MMBIR, SRS, or BISRS. Moreover, we hypothesize that the genomic architecture might drive their formation by increasing the susceptibility for DNA breakage or promote replication fork stalling. Finally, our locus-centered study, elucidating the etiology of a large set of rare microdeletions involved in a monogenic disorder, can serve as a model for other clustered, non-recurrent microdeletions in genetic disease.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Accurate lung cancer classification is crucial to guide therapeutic decisions. However, histological subtyping by pathologists requires tumor tissue-a necessity that is often intrinsically associated ...with procedural difficulties. The analysis of circulating tumor DNA present in minimal-invasive blood samples, referred to as liquid biopsies, could therefore emerge as an attractive alternative.
Concerning adenocarcinoma, squamous cell carcinoma, and small cell carcinoma, our proof of concept study investigates the potential of liquid biopsy-derived copy number alterations, derived from single-end shallow whole-genome sequencing (coverage 0.1-0.5×), across 51 advanced stage lung cancer patients.
Genomic abnormality testing reveals anomalies in 86.3% of the liquid biopsies (16/20 for adenocarcinoma, 13/16 for squamous cell, and 15/15 for small cell carcinoma). We demonstrate that copy number profiles from formalin-fixed paraffin-embedded tumor biopsies are well represented by their liquid equivalent. This is especially valid within the small cell carcinoma group, where paired profiles have an average Pearson correlation of 0.86 (95% CI 0.79-0.93). A predictive model trained with public data, derived from 843 tissue biopsies, shows that liquid biopsies exhibit multiple deviations that reflect histological classification. Most notably, distinguishing small from non-small cell lung cancer is characterized by an area under the curve of 0.98 during receiver operating characteristic analysis. Additionally, we investigated how deeper paired-end sequencing, which will eventually become feasible for routine diagnosis, empowers tumor read enrichment by insert size filtering: for all of the 29 resequenced liquid biopsies, the tumor fraction could be increased in silico, thereby "rescuing" three out of five cases with previously undetectable alterations.
Copy number profiling of cell-free DNA enables histological classification. Since shallow whole-genome sequencing is inexpensive and often fully operational at routine molecular laboratories, this finding has current diagnostic potential, especially for patients with lesions that are difficult to reach.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Structural genomic variations play an important role in human disease and phenotypic diversity. With the rise of high-throughput sequencing tools, mate-pair/paired-end/single-read sequencing has ...become an important technique for the detection and exploration of structural variation. Several analysis tools exist to handle different parts and aspects of such sequencing based structural variation analyses pipelines. A comprehensive analysis platform to handle all steps, from processing the sequencing data, to the discovery and visualization of structural variants, is missing. The ViVar platform is built to handle the discovery of structural variants, from Depth Of Coverage analysis, aberrant read pair clustering to split read analysis. ViVar provides you with powerful visualization options, enables easy reporting of results and better usability and data management. The platform facilitates the processing, analysis and visualization, of structural variation based on massive parallel sequencing data, enabling the rapid identification of disease loci or genes. ViVar allows you to scale your analysis with your work load over multiple (cloud) servers, has user access control to keep your data safe and is easy expandable as analysis techniques advance. URL: https://www.cmgg.be/vivar/
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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