Genomic Next-generation Sequencing Research Articles

Next generation sequencing and imprinting disorders: Current applications and future perspectives: Lessons from Silver-Russell syndrome

Imprinting Disorders are a group of rare diseases with overlapping phenotypes which are associated with similar molecular changes and affect imprinted chromosomal regions. Clinical features mainly occur prenatally or in childhood, but have a severe lifelong impact on health. Due to their clinical and molecular heterogeneity, the diagnosis of imprinting disorders is often challenging and requires testing of a broad spectrum of genomic variants and aberrant methylation of imprinted loci (epimutations). A significant number of patients suspicious for imprinting disorders remain without a molecular confirmation, and in these cases differential diagnoses have to be considered. In fact, in patients with clinical features suggestive for imprinting disorders, the precise identification of the molecular cause is relevant for both clinical management as well as for genetic counselling. Thus, a comprehensive testing approach has to be applied. Next generation sequencing (NGS) based studies show that this technique is a valuable tool to improve the diagnostic efficiency particularly in entities with broad differential diagnoses. Furthermore, the development of diverse NGS approaches allows new insights in the function of imprinted regions, their structures, interactions and regulation. Based on a large cohort of patients referred for routine Silver Russel syndrome testing, the appropriateness and limitations of first trial tests in imprinting disorders are demonstrated in this report, but the chances of genomic NGS approaches for diagnostics and research are elucidated as well. Finally, the significance of the precise molecular diagnosis for the personalized management of the patient, and genetic counselling of the family will be discussed.

The complete chloroplast genome of Rosa lucidissima, a critically endangered wild rose endemic to China

Rosa lucidissima is a critically endangered wild rose species endemic to China with both ornamental value and medicinal usefulness. The complete chloroplast genome sequence of R. lucidissima was generated by de novo assembly using whole genome next-generation sequencing to provide genomic data for further conservation genetics of this species and for phylogenetic analysis of genus Rosa. The complete chloroplast genome was 156,588 bp in length, including two inverted repeat (IR) regions (26,048 bp), a small single-copy (SSC) region (18,765 bp), and a large single-copy (LSC) region (85,713 bp). The chloroplast genome encoded 129 genes, including 84 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. The overall GC content of the whole genome was 37.25%. Phylogenetic analysis of 19 selected chloroplast genomes revealed that R. lucidissima was most closely related to R. chinensis var. spontanea.

Single-Cell Sequencing in Precision Medicine.

The application of next-generation sequencing in cancer genomics allowed for a better understanding of the genetics and pathogenesis of cancer. Single-cell genomics is a relatively new field that has enhanced our current knowledge of the genetic diversity of cells involved in the complex biological systems of cancer. Single-cell genomics is a rapidly developing field, and current technologies can assay a single cell's gene expression, DNA variation, epigenetic state, and nuclear structure. Statistical and computational methods are central tosingle-cell genomics and allows for extraction of meaningful information. The translational application of single-cell sequencing in precision cancer therapy has the potential to improve cancer diagnostics, prognostics, targeted therapy, early detection, and noninvasive monitoring. Furthermore, single-cell genomics will transform cancer research as even initial experiments have revolutionized our current understanding of gene regulation and disease.

Biobank Establishment and Sample Management in the Tohoku Medical Megabank Project.

The Tohoku Medical Megabank biobank (TMM biobank) is the first major population-based biobank established in Japan. The TMM biobank was established based on two population cohorts and is a reconstruction program from the Great East Japan Earthquake and Tsunami of 2011. The biobank stores more than 3.4 million tubes of biospecimens and associated health and analytic data obtained from approximately 150,000 TMM cohort participants between May 2013 and December 2018, and the TMM biobank currently shares high-quality specimens and data. Various biospecimens, including peripheral and cord blood mononuclear cells, buffy coat, plasma, serum, urine, breast milk and saliva have been collected in the TMM biobank. To minimize human error and maintain the quality of data and specimens, we have been utilizing laboratory information management system into various biobank procedures from registration to storage with various automation systems, such as liquid dispensing, DNA extraction and their storage. The biobank procedures for the quality management system (ISO 9001:2015) and information security management system (ISO 27001:2013) are certified by the International Organization for Standardization. The quality of our biobank samples fulfills the pre-analytical requirements for researchers conducting next-generation whole genome sequencing, DNA array analyses, proteomics, metabolomics, etc. We established analytical centers to conduct standard genomic and multiomic analyses in-house and share the generated data. Additionally, we generate thousands of Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines and proliferating T cells for functional studies. The TMM biobank serves as an indispensable infrastructure for academic, clinical and industrial research to actualize next-generation medicine in Japan.

Assembly of Ancient Mitochondrial Genomes Without a Closely Related Reference Sequence.

Recent methodological advances have transformed the field of ancient DNA (aDNA). Basic bioinformatics skills are becoming essential requirements to process and analyze the sheer amounts of data generated by current aDNA studies and in biomedical research in general. This chapter is intended as a practical guide to the assembly of ancient mitochondrial genomes, directly from genomic DNA-derived next-generation sequencing (NGS) data, specifically in the absence of closely related reference genomes. In a hands-on tutorial suitable for readers with little to no prior bioinformatics experience, we reconstruct the mitochondrial genome of a woolly mammoth deposited ~45,000years ago. We introduce key software tools and outline general strategies for mitogenome assembly, including the critical quality assessment of assembly results without a reference genome.

The Story of Helicobacter pylori: Depicting Human Migrations from the Phylogeography.

Helicobacter pylori is a spiral-shaped Gram-negative bacterium, which has infected more than half of the human population. Besides its colonisation capability, the genetic diversity of H. pylori is exceptionally well structured and belongs to several distinct genetic populations, depicting various prehistorical human migration events. The evolutionary relationship of H. pylori with its host had been started at least ~100,000years ago. In addition, the discovery of the ancient H. pylori genome from a European Copper Age glacier mummy, "The Iceman", gave the idea that the second out of Africa migration resulted in the recombinant population of hpEurope at least about 5300years ago. The advancement of next-generation genome sequencing discovered the prophage of H. pylori and could discriminate the big population of hpEurope into two different subpopulations. In addition, the implementation of the chromopainter/fineSTRUCTURE algorithm to the whole genome analysis of H. pylori provides a finer resolution population genetics of H. pylori; therefore it could also depict the recent migrations within the past 500years after colonial expansion. This discovery shows that the genetic recombination of H. pylori strains is far more dynamic compared to its human host, but still maintains the similarity to its host, suggesting that H. pylori is a handy tool to reconstruct the human migration both in the past and the recent.

Prospective, Real-time Metagenomic Sequencing During Norovirus Outbreak Reveals Discrete Transmission Clusters.

BackgroundNorovirus outbreaks in hospital settings are a common challenge for infection prevention teams. Given the high burden of norovirus in most communities, it can be difficult to distinguish between ongoing in-hospital transmission of the virus and new introductions from the community, and it is challenging to understand the long-term impacts of outbreak-associated viruses within medical systems using traditional epidemiological approaches alone.MethodsReal-time metagenomic sequencing during an ongoing norovirus outbreak associated with a retrospective cohort study.ResultsWe describe a hospital-associated norovirus outbreak that affected 13 patients over a 27-day period in a large, tertiary, pediatric hospital. The outbreak was chronologically associated with a spike in self-reported gastrointestinal symptoms among staff. Real-time metagenomic next-generation sequencing (mNGS) of norovirus genomes demonstrated that 10 chronologically overlapping, hospital-acquired norovirus cases were partitioned into 3 discrete transmission clusters. Sequencing data also revealed close genetic relationships between some hospital-acquired and some community-acquired cases. Finally, this data was used to demonstrate chronic viral shedding by an immunocompromised, hospital-acquired case patient. An analysis of serial samples from this patient provided novel insights into the evolution of norovirus within an immunocompromised host.ConclusionsThis study documents one of the first applications of real-time mNGS during a hospital-associated viral outbreak. Given its demonstrated ability to detect transmission patterns within outbreaks and elucidate the long-term impacts of outbreak-associated viral strains on patients and medical systems, mNGS constitutes a powerful resource to help infection control teams understand, prevent, and respond to viral outbreaks.

Next-generation sequencing-based clinical sequencing: toward precision medicine in solid tumors.

Numerous technical and functional advances in next-generation sequencing (NGS) have led to the adoption of this technique in conventional clinical practice. Recently, large-scale genomic research and NGS technological innovation have revealed many more details of somatic and germline mutations in solid tumors. This development is allowing for the classification of tumor type sub-categories based on genetic alterations in solid tumors, and based on this information, new drugs and targeted therapies are being administered to patients. This has largely been facilitated by gene panel testing, which allows for a better understanding of the genetic basis for an individual's response to therapy. NGS-based comprehensive gene panel testing is a clinically useful approach to investigate genomic mechanisms, including therapy-related signaling pathways, microsatellite instability, hypermutated phenotypes, and tumor mutation burden. In this review, we describe the concept of precision medicine in solid tumors using NGS-based comprehensive gene panel testing, as well as the importance of quality control of tissue sample handling in routine NGS-based genomic testing, and we discuss issues for the future adoption of this technique in Japan.

Congenital X-Linked Myelodysplasia with Tetraploidy Is Associated with De Novo Germline C-Terminal Mutation of SEPT6, a Septin Filament Protein

Private germline mutations affecting hematopoiesis can cause progressive myelodysplasia and thus constitute pre-leukemic states. These can remain undetected, progressively transform and reveal themselves as infant leukemias, which can be linked to translocations involving the mixed lineage-leukemia (MLL) gene. Septin proteins play key roles in mammalian cell division and cytokinesis and are found as fusion partners of MLL in infant and early childhood acute myeloid leukemia (AML). We identified and describe a human germline disorder of septins in a newborn with myelodysplasia who required early hematopoietic stem cell transplant (HSCT) to prevent progressive disease.Materials & Methods: A Caucasian newborn male with no birth defects/malformations or suspicious family history presented with severe progressive neutropenia and was found to have bone marrow (BM) dysplasia with tetraploidy of myeloid progenitors. The patient developed unfavorable clonal aberrations (trisomy 7,8,9) and increased tetraploidy. Due to his progressive cytopenias and concern about risk of leukemic transformation, he underwent an allogeneic busulfan-cyclophosphamide/ATG conditioned DQ-mismatched unrelated HSCT at age 1 yo. He is currently 8 years post-HSCT with normal trilineage hematopoiesis (full donor chimerism), no graft versus host disease or any other non-hematological phenotype. To investigate the genetic etiology of this unique phenotype, we performed family trio germline exome/whole genome next-generation sequencing (NGS), and somatic pre-HSCT BM NGS for the index case. An established algorithm filtered for significant candidates following a de novo germline model. Immunohistochemical (IHC) staining of the pre- and post-HSCT BM biopsies for the candidate protein was performed. To validate the germline origin of the candidate mutation, we generated patient and control skin fibroblasts and induced pluripotent stem cells (iPSCs) that underwent fidelity testing by murine injection teratoma assays and 16-marker immunofluorescence (IF) staining. We then studied hematopoietic progenitor cells (HPCs) derived from iPSC-embryoid bodies (EB) in methylcellulose assays. To further determine the pathogenic nature of the mutation, we generated CRISPR/Cas9 knock-out of the human erythroleukemic cell line (TF-1) and studies these cells by cytomorphology, DNA and cell cycle assays. In-silico protein analysis of the candidate mutation and its effects on septin filament formation was performed.Results: Family trio and disease-tissue NGS identified a novel, germline C-terminal mutation in SEPT6, which was acquired de novo in the patient, and was not found in any database of common polymorphisms. IHC of pre-HSCT patient BM showed reduced Septin-6 staining in megakaryocyte and granulocyte precursors compared to post-HSCT and controls. Patient-derived iPSCs carried the mutation, were cytogenetically normal and bona-fide pluripotent by IF and teratoma assays. EB-derived HPCs from these cells recapitulated the patient's phenotype as they differentially failed to produce granulocyte vs erythroid colonies (fold-reductions CFU-M:8, CFU-G:36, CFU-GM:46, BFU-E:6, see Figure). Despite multiple approaches, SEPT6 CRISPR/Cas9 knock-out/in of the patient's mutation was not tolerated in iPSCs and human myeloid (granulo-/myelocytic) cell lines (HL-60, Molm-13, K562), and only tolerated in erythroid TF-1 cells. Analysis of SEPT6 knock-out TF-1 single-clone lines revealed a propensity to poly-nuclearity/lobation, as observed in the patient's BM. SEPT6 knock-in of C-terminal mutations caused cell death, consistent with existing literature. In silico protein analysis (incl. previously published crystallographic data) suggests that the mutation a) most likely modulates the key role of the coiled-coil SEPT6 domain in septin filament stabilization/bundling/bending, and thus deleteriously impacts cytokinesis, and b) perturbs the equilibrium of splice variants, possibly conferring tissue specificity.Conclusions: Mutation of the C-terminus of human SEPT6 causes aberrant cytokinesis in HPCs leading to a severe congenital neutropenia with tetraploidy and progressive myelodysplasia and cytogenetic aberrations. This report implicates a human germline disorder of SEPT6, and further investigations are required to elucidate the role septins in normal and disordered myelopoiesis. DisclosuresWilliams:Bluebird Bio: Research Funding.

1281. Emergence of a B/F1 HIV Recombinant in the Philippines: A Potentially New Circulating Recombinant Form

BackgroundThe Philippines has one of the fastest growing HIV epidemics globally. This was accompanied by a switch from subtype B to CRF01_AE. With a large population of returning overseas workers, new subtypes are being introduced regularly. Because diagnosis of HIV in the Philippines is usually late, superinfections can occur and may give rise to new circulating recombinant forms (CRFs). We propose a new CRF from the Philippines.MethodsFollowing institutional board approval, treatment-naive patients from two HIV treatment hubs (San Lazaro Hospital and the Philippine General Hospital) were recruited. Blood samples underwent Sanger sequencing of the PR and RT regions and next-generation sequencing (NGS) of near-full length genomes. Sequences were analyzed for recombination using the online tool jumping profile Hidden Markov Model (http://jphmm.gobics.de/).Results247 samples underwent Sanger sequencing of the PR and RT regions of the pol gene. Phylogenetic analysis indicated a clustering of four of the samples. Further analysis showed all four samples had the same breakpoints at nucleotides 2875, 2996, and 3001 (HXB2 numbering). All four patients were male, MSM, with a mean age of 28 years old (24–32), and >10 sexual partners. Mean CD4 count was 464 cells/µL and median viral load was 2.67 × 104 copies/mL. Two patients had sex with foreigners. To get a better overall view of subtype composition, we performed NGS using Illumina HiSeq. NGS showed the majority of the genome to be subtype F1 with segments of subtype B inserted in the pol, vpu, and env genes. A blast analysis of the consensus sequence showed 8,932 out of 8,943 nucleotides (99%) matched a 1999 sample from Argentina. Phylogenetic analysis of these samples show clustering of the four B/F1 recombinants with some South American sequences. No drug resistance mutations were identified.ConclusionMutation and recombination contribute to the extensive genetic diversity of HIV. Understanding this is important in choosing treatment regimens, developing future vaccines, and pursuing epidemiological investigations. The emergence of a new CRF in the Philippines underlies the importance of conducting routine surveillance for new HIV recombinant forms.Figure 1.New CRF genome showing subtype components.Disclosures All authors: No reported disclosures.

Insect Transcription Factors: A Landscape of Their Structures and Biological Functions in Drosophila and beyond.

Transcription factors (TFs) play essential roles in the transcriptional regulation of functional genes, and are involved in diverse physiological processes in living organisms. The fruit fly Drosophila melanogaster, a simple and easily manipulated organismal model, has been extensively applied to study the biological functions of TFs and their related transcriptional regulation mechanisms. It is noteworthy that with the development of genetic tools such as CRISPR/Cas9 and the next-generation genome sequencing techniques in recent years, identification and dissection the complex genetic regulatory networks of TFs have also made great progress in other insects beyond Drosophila. However, unfortunately, there is no comprehensive review that systematically summarizes the structures and biological functions of TFs in both model and non-model insects. Here, we spend extensive effort in collecting vast related studies, and attempt to provide an impartial overview of the progress of the structure and biological functions of current documented TFs in insects, as well as the classical and emerging research methods for studying their regulatory functions. Consequently, considering the importance of versatile TFs in orchestrating diverse insect physiological processes, this review will assist a growing number of entomologists to interrogate this understudied field, and to propel the progress of their contributions to pest control and even human health.

Author Correction: A comparison of tools for the simulation of genomic next-generation sequencing data.

The originally published article contained errors in Fig. 1 (Decision tree for the selection of a suitable NGS genomic simulator), whereby the labels 'Variants' and 'No Variants' had been switched. The correct figure is presented in this notice.

Long-read sequencing identified intronic repeat expansions in SAMD12 from Chinese pedigrees affected with familial cortical myoclonic tremor with epilepsy.

The locus for familial cortical myoclonic tremor with epilepsy (FCMTE) has long been mapped to 8q24 in linkage studies, but the causative mutations remain unclear. Recently, expansions of intronic TTTCA and TTTTA repeat motifs within SAMD12 were found to be involved in the pathogenesis of FCMTE in Japanese pedigrees. We aim to identify the causative mutations of FCMTE in Chinese pedigrees. We performed genetic linkage analysis by microsatellite markers in a five-generation Chinese pedigree with 55 members. We also used array-comparative genomic hybridisation (CGH) and next-generation sequencing (NGS) technologies (whole-exome sequencing, capture region deep sequencing and whole-genome sequencing) to identify the causative mutations in the disease locus. Recently, we used low-coverage (~10×) long-read genome sequencing (LRS) on the PacBio Sequel and Oxford Nanopore platforms to identify the causative mutations, and used repeat-primed PCR for validation of the repeat expansions. Linkage analysis mapped the disease locus to 8q23.3-24.23. Array-CGH and NGS failed to identify causative mutations in this locus. LRS identified the intronic TTTCA and TTTTA repeat expansions in SAMD12 as the causative mutations, thus corroborating the recently published results in Japanese pedigrees. We identified the pentanucleotide repeat expansion in SAMD12 as the causative mutation in Chinese FCMTE pedigrees. Our study also suggested that LRS is an effective tool for molecular diagnosis of genetic disorders, especially for neurological diseases that cannot be positively diagnosed by conventional clinical microarray and NGS technologies.

Use DNA fingerprinting to identify and confirm triploid embryos in whole genome next generation sequencing (NGS) preimplantation genetic screen (PGS)

Whole genome next generation sequencing (NGS) for preimplantation genetic screen (PGS) has higher sensitivity and accuracy in detecting unbalanced translocation and partial aneuploid than targeted NGS. Whole genome NGS also detect some triploid, the most common polyploidy, if the sex chromosomes are unbalanced (69, XXY, and 69, XYY). Unlike targeted NGS, whole genome NGS unusually does not detect triploid 69, XXX, neither does it provide additional data to confirm triploid 69, XXY and 69, XYY identified in PGS. Here we tested using DNA fingerprinting in combination with whole genome NGS to identify and confirm sex chromosome balanced and unbalanced triploid in PGS. Feasibility and validation study. The feasibility is tested on of 5 known triploid cell lines and 5 known diploid cell lines. 24 fingerprinting loci were tested. Further validation and accuracy were tested on 16 blastocyst biopsy samples whole genome amplification product. All 16 samples had whole genome NGS for PGS and were potential triploid samples. 15 of them were identified by unbalanced sex chromosomes for PGS. 1 of them showed 46, XX for PGS but is suspected to be 69, XXX triploid because of the enlarged nuclei observed by the embryologist. DNA fingerprinting was performed on the 16 PGS samples to identify and confirm the triploid embryos. FISH was performed to confirm the DNA fingerprinting result. Homozygous or heterozygous loci (24 of 24) in DNA fingerprinting were detected for all 5 known diploid cells lines, confirming diploid. For the 5 known triploid cell lines, 5 to 10 out of the 24 DNA fingerprinting loci has 3 genotypes, confirming triploid. Among the 15 potential triploid samples identified by PGS, 13 showed triploid in DNA fingerprinting, with 3 to 11 loci having 3 genotypes; 2 showed diploid in DNA fingerprinting. The enlarged nuclei sample with 46, XX for PGS also showed triploid in DNA fingerprinting, with 7 out of 24 loci having 3 genotypes. The 16 embryos FISH result confirmed the DNA fingerprinting result, demonstrating DNA fingerprinting is accurate in identifying and confirming triploid in PGS sample. DNA fingerprinting test on whole genome amplified PGS samples is a simple and accurate method to examine triploid status of embryos. It can confirm triploid indicated by embryo morphology. Triploid identification in PGS based on unbalance sex chromosomes can be difficult and not reliable, especially when embryo DNA quality is compromised. In this situation DNA fingerprinting can identify and confirm the real triploid embryos. Used in combination with whole genome NGS, DNA fingerprinting allows patients to benefit from the advantages of the whole genome NGS, and at the same time, identifying the triploid embryos to reduce pregnancy loss.

Embryo quality and rate of development as predictive tools for embryo selection

This study was designed to examine the relationship between embryo quality, day of biopsy, and ploidy; with the goal of improving embryo selection criteria for frozen embryo transfer (FET). In this retrospective study, data were collected from 696 patients who underwent 904 autologous preimplantation genetic testing - aneuploidy (PGT-A) cycles between 2015 and 2017. Variables included for analysis were patient age, embryo morphology and day of trophectoderm biopsy. Only good and fair embryos based on SART criteria were considered for biopsy. A total of 3,502 embryos were biopsied and analyzed. Blastocysts from autologous oocytes were assessed for morphology prior to undergoing trophectoderm biopsy on day 5 or day 6. All embryos were then vitrified and ploidy assessed by a third party testing laboratory using array comparative genomic hybridization, next-generation sequencing, or single-nucleotide polymorphism microarray. Patients who utilized monogenic preimplantation genetic testing were not included in this study. Mosaic (0.14%) and inconclusive (1.04%) results were excluded from the data set. Statistical comparisons were made by Chi-Square analysis. The average age of autologous patients was 36.7. An average of 3.9 embryos were biopsied per cycle. Good quality embryos that were biopsied on day 5 showed the highest euploid rate of 63% (857/1370). Good quality embryos biopsied on day 6 had a euploid rate of 56% (588/1048). Fair quality embryos were significantly less likely to yield euploid results, with embryos biopsied on day 5 returning a euploid rate of 48% (187/393) and embryos biopsied on day 6 returning a euploid rate of 39% (271/691). When stratified by age, this trend held significance (p<.01). Establishing a hierarchy of embryo quality and rate of development can help guide embryo selection for frozen embryo transfers for patients who did not utilize PGT. These data suggest that in the absence of PGT, embryo quality is the single most important factor in selecting a euploid embryo for transfer, while rate of development to the blastocyst stage should be considered secondarily. Further studies will be necessary to identify other laboratory and clinical factors that contribute to the success of an FET cycle.Tabled 1STATISTICAL COMPARISONS WITHIN AGE GROUPS (*p<.01)Patient AgeDay 5 GOODDay 6 GOODDay 5 FAIRDay 6 FAIRTotal Embryos Tested<3575% (322/429)*72% (248/345)*58% (70/119)*58% (115/197)*109035-3766% (287/437)*60% (180/300)*53% (59/111)*41% (75/181)*102938-4055% (190/346)*44% (129/294)*41% (48/116)*31% (65/211)*967>4037% (58/158)*28% (31/109)*21% (10/47)*16% (16/102)*416Overall63% (857/1370)*56% (588/1048)*48% (187/393)*39% (271/691)*3502 Open table in a new tab

Rate of development of euploid blastocyst affects clinical outcomes of frozen embryo transfer

Preimplantation genetic testing for aneuploidy (PGT-A) has been increasingly used to identify euploid embryos. The present study evaluates clinical outcomes of euploid frozen embryo transfers (FET) in 2016-2017, following trophectoderm (TE) biopsy (BX) on day 5 (D5), day 6 (D6), or day 7 (D7). Retrospective study. Trophectoderm biopsy was performed on embryos once they reached blastocyst (BL) stage on D5, D6, or D7, where the day of biopsy indicates the rate of development. We analyzed a total of 1889 FETs following D5, D6, or D7 TE BX. Comparative genomic hybridization array, SNP microarray, or Next-generation sequencing was used for PGT-A. Transfer was performed following vitrification and warming. Clinical outcomes of implantation rate (IR), ongoing pregnancy/delivered (OGD), and spontaneous abortion (SAb) were compared among different BX days per patient age groups, using Kruskal-Wallis testing for IR and Chi-squared analysis for OGD and SAb. Statistical significance was defined as P <0.05. For patients of all ages, all clinical outcomes (IR, OGD, and SAb) showed statistical significance correlated to the rate of embryo development. In patients under 35 years of age, OGD was significantly higher in D5 euploid embryos compared to D6 or D7. For patients 38 years of age and older, IR and OGD were statistically higher in D5 euploid embryos.Tabled 1Clinical Outcomes Post TE BX/FETD5 BX/FET#FETAll Ages: 684<35: 21535-37: 17538-40: 167>40: 127IR72.5%∗∗68.2%74.4%77.0%∗∗71.2%∗OGD+429 (63%)∗128 (60%)∗113 (65%)113 (68%)∗75 (59%)∗SAb (losses/CP;%)65 (10%)∗20 (9%)13 (7%)17 (10%)15 (12%)D6 BX/FET#FETAll Ages: 1113<35: 31635-37: 24838-40: 318>40: 231IR60.9%∗∗60.8%64.1%61.1%∗∗57.5%∗OGD+540 (49%)∗155 (49%)∗131 (53%)149 (47%)∗105 (45%)∗SAb (losses/CP;%)129 (12%)∗38 (12%)25 (10%)38 (12%)28 (12%)D7 BX/FET#FETAll Ages: 92<35: 2035-37: 1538-40: 41>40: 16IR43.2%∗∗45.5%68.8%31.7%∗∗43.8%∗OGD+32 (35%)∗8 (40%)∗8 (53%)11 (27%)∗5 (31%)∗SAb (losses/CP;%)8 (9%)∗2 (10%)2 (13%)2 (5%)2 (13%)*: P < 0.05 ; **: P < 0.01 ; + :Ongoing pregnancies were defined as those with fetal cardiac activity at or beyond 8 weeks of gestational age. Open table in a new tab *: P < 0.05 ; **: P < 0.01 ; + :Ongoing pregnancies were defined as those with fetal cardiac activity at or beyond 8 weeks of gestational age. Overall, D5 euploid BL showed improved clinical outcomes compared to D6 or D7, suggesting that delayed embryo development may have an adverse effect on clinical outcomes, despite the proven euploid status. Interestingly, for younger patients (<35), D5 euploid BL improved continuation of pregnancy (OGD), while other clinical outcomes were not significantly different among D5, D6, or D7. However, for older patients (≥38), implantation rates as well as ongoing pregnancy rates were improved for D5 embryos compared to D6 or D7. Even though D7 euploid BL result in much lower IR and OGD compared to D5 or D6, they still result in 35% OGD, suggesting that slow developing embryos can still be beneficial for the patients, and therefore should not be overlooked.

Taking up the torch

Taking up the torch

A New Era, New Strategies: Education and Communication Strategies to Manage Greater Access to Genomic Information.

As next-generation genomic sequencing, including whole-genome sequencing information, becomes more common in research, clinical, and public health contexts, there is a need for comprehensive communication strategies and education approaches to prepare patients and clinicians to manage this information and make informed decisions about its use, and nowhere is that imperative more pronounced than when genomic sequencing is applied to newborns. Unfortunately, in-person counseling is unlikely to be applicable or cost-effective when parents obtain genomic risk information directly via the Internet. As a rule, communication strategies should match how people are accessing health information. Today, many people can obtain health information in a variety of settings, including through direct-to-consumer services, via websites, and through other digital channels or settings. In response to these changes, new communication strategies need to be considered. Adopting a comprehensive communication model means understanding the multiple levels of influence experienced by parents and the clinicians who serve them. In addition, applying communication-science principles can help in addressing some key challenges to effectively communicating genomic information to parents.

PO-047 Etarget: a digital science solution to integrate clinical and genomic data for the manchester molecular tumour board (MTB)

IntroductionManchester Cancer Research Centre (MCRC) has established an MTB to facilitate precision medicine decision-making within the TARGET trial (Tumour chAracterisation to Guide Experimental Targeted Therapy). The MTB meets monthly to review clinical data and next generation sequencing (NGS) results from tumour tissue and circulating DNA (ctDNA) for patients being considered for early phase clinical trials. Initially the MTB relied on multiple paper reports. Here we present eTARGET, a digital solution developed by the digital Experimental Cancer Medicine Team (digitalECMT), which integrates clinical and genomic NGS data to facilitate decision-making for matching patients with clinical trials.Material and methodsThe digitalECMT explored data sources and existing reports to define end-user and data requirements. Following a successful prototype, a beta version was developed. Created in Microsoft Azure, a secure cloud computing platform, components included a storage account for data upload from three different sources, and a database for storing and integrating the data. The solution enabled automated extraction of individual pseudonymised clinical and genomic data. In addition, a web application to view the data was developed with clinical input.Results and discussionsThe beta version of eTARGET went online in October 2017 and has been utilised at 5 MTB meetings for 55 patient cases. This portal interface presents patient characteristics, treatment history and genomic data. The portal can be viewed remotely, across multiple locations, where all attendees see the same view. eTARGET has enabled the MTB to review individual patient data in a single portal, capture meeting outcomes in real-time and upload to the electronic patient record. Decisions regarding significant variants, trial matching or requirements for further analytical or translational analyses are captured.Conclusion eTARGET has shown that a digital solution can be implemented to overcome the challenge of integrating data from disparate sources in different organisations to create a single view of patient clinical and genomic data. We have shown the utility of eTARGET in a hospital setting to support decision-making for an MTB. The eTARGET project opens the possibility of wider MTB participation including cross centre collaboration. Next steps are to enhance the software to visualise the global molecular dataset and serial changes in NGS profiles on treatment.

Research Techniques Made Simple: Mass Spectrometry for Analysis of Proteins in Dermatological Research

Identifying previously unknown proteins or detecting the presence of known proteins in research samples is critical to many experiments conducted in life sciences, including dermatology. Sensitive protein detection can help elucidate new intervention targets and mechanisms of disease, such as in autoimmune blistering skin diseases, atopic eczema, or other conditions. Historically, peptides from highly purified single proteins were sequenced, with many limitations, by stepwise degradation from the N-terminus to the C-terminus with subsequent identification by UV absorbance spectroscopy of the released amino acids (i.e., Edman degradation). Recently, however, the availability of comprehensive protein databases from different species (derived from high-throughput next-generation sequencing of those organisms' genomes) and sophisticated bioinformatics analysis tools have facilitated the development and use of mass spectrometry for identification and global analysis of proteins, summarized as mass spectrometry-based proteomics. Mass spectrometry is an analytical technique measuring the mass (m)-to-charge (z) ratio of ionized biological molecules such as peptides. Proteins can be identified by correlating peptide-derived experimental mass spectrometry spectra with theoretical spectra predicted from protein databases. Here we briefly describe how this technique works, how it can be used for identification of proteins, and how this knowledge can be applied in elucidating human biology and disease.