Genomic Screening Research Articles

Growth in perceived clinical genetics competency among primary care providers participating in genomic population health screening.

Limited competency in genetics among primary care providers (PCPs) is a barrier to use of genetic information in healthcare. Formal genetics lessons require time and interest, and knowledge wanes. We hypothesized another path to competency: participation in our PCP-centered adult clinical genomic population health screening program. We asked participating Family Medicine PCPs about their perceptions of growth in their genetics competency. An anonymous, voluntary, cross-sectional survey was developed and distributed to PCPs offering the screening. Results were compiled after 3weeks. PCPs rated several program resources for value and provided open-ended feedback. Seventy-five percent of respondents agreed that genetics is important to their practice. Eighty-seven percent felt that their knowledge of clinical genetics topics had grown. Eighty-seven percent perceived increased confidence in offering genetic testing and in discussing common genetic results with patients. Respondents gained appreciation for the scope of clinical utility that genetic information offers patients. Each education resource rated at least 3.75 out of 5 for contributing to genetics knowledge. The case-specific Genomic Medicine Action Plan rated highest in educational value, 4.5 out of 5. Most responding PCPs offering genomic population health screening perceived growth in their genetic competency and found hands-on, case-based resources most useful.

Genomic medicine in neonatal care: progress and challenges.

During the neonatal period, many genetic disorders present and contribute to neonatal morbidity and mortality. Genomic medicine-the use of genomic information in clinical care- has the potential to significantly reduce morbidity and mortality in the neonatal period and improve outcomes for this population. Diagnostic genomic testing for symptomatic newborns, especially rapid testing, has been shown to be feasible and have diagnostic and clinical utility, particularly in the short-term. Ongoing studies are assessing the feasibility and utility, including personal utility, of implementation in diverse populations. Genomic screening for asymptomatic newborns has also been studied, and the acceptability and feasibility of such an approach remains an active area of investigation. Emerging precision therapies, with examples even at the "n-of-1" level, highlight the promise of precision diagnostics to lead to early intervention and improve outcomes. To sustainably implement genomic medicine in neonatal care in an ethical, effective, and equitable manner, we need to ensure access to genetics and genomics knowledge, access to genomic tests, which is currently limited by payors, feasible processes for ordering these tests, and access to follow up in the clinical and research realms. Future studies will provide further insight into enablers and barriers to optimize implementation strategies.

Evaluation of Malignant Hyperthermia Features in Patients with Pathogenic or Likely Pathogenic RYR1 Variants Disclosed through a Population Genomic Screening Program.

Malignant hyperthermia (MH) susceptibility is a heritable musculoskeletal disorder that can present as a potentially fatal hypermetabolic response to triggering anesthesia agents. Genomic screening for variants in MH-associated genes RYR1 and CACNA1S provides an opportunity to prevent morbidity and mortality. There are limited outcomes data from disclosing variants in RYR1, the most common MH susceptibility gene, in unselected populations. The authors sought to identify the rate of MH features or fulminant episodes after triggering agent exposure in an unselected population undergoing genomic screening including actionable RYR1 variants. The MyCode Community Health Initiative by Geisinger (USA) is an electronic health record-linked biobank that discloses pathogenic and likely pathogenic variants in clinically actionable genes to patient-participants. Available electronic anesthesia and ambulatory records for participants with actionable RYR1 results returned through December 2020 were evaluated for pertinent findings via double-coded chart reviews and reconciliation. Descriptive statistics for observed phenotypes were calculated. One hundred fifty-two participants had an actionable RYR1 variant disclosed during the study period. None had previous documented genetic testing for MH susceptibility; one had previous contracture testing diagnosing MH susceptibility. Sixty-eight participants (44.7%) had anesthesia records documenting triggering agent exposure during at least one procedure. None received dantrolene treatment or had documented muscle rigidity, myoglobinuria, hyperkalemia, elevated creatine kinase, severe myalgia, or tea-colored urine. Of 120 possibly MH-related findings (postoperative intensive care unit admissions, hyperthermia, arterial blood gas evaluation, hypercapnia, or tachycardia), 112 (93.3%) were deemed unlikely to be MH events; 8 (6.7%) had insufficient records to determine etiology. Results demonstrate a low frequency of classic intraanesthetic hypermetabolic phenotypes in an unselected population with actionable RYR1 variants. Further research on the actionability of screening for MH susceptibility in unselected populations, including economic impact, predictors of MH episodes, and expanded clinical phenotypes, is necessary.

O10 Universal genomic newborn screening for early, treatable, and severe conditions- including 33 genes of NMD: Baby Detect

O10 Universal genomic newborn screening for early, treatable, and severe conditions- including 33 genes of NMD: Baby Detect

1276P Genomic screening by multi-gene assay for perioperative targeted therapy in patients with early-stage non-small cell lung cancer: LC-SCRUM-Advantage/MRD

1276P Genomic screening by multi-gene assay for perioperative targeted therapy in patients with early-stage non-small cell lung cancer: LC-SCRUM-Advantage/MRD

Next-generation sequencing-based newborn screening initiatives in Europe: an overview

Aim: This article describes results from a survey targeting healthcare professionals (HCPs) leading newborn screening (NBS) initiatives in Europe. The survey was developed within the framework of a dedicated working group set up by the International Rare Diseases Research Consortium (IRDiRC) to gather collective efforts relating to NBS. The objectives of the survey were to gain a better understanding of approaches being tested for the expansion of NBS and to raise awareness of the significant momentum across Europe to evaluate novel technologies for use in future NBS programs. Methods: A web-based survey including 57 questions was developed to gather information about genomic newborn screening initiatives in Europe that are using next-generation sequencing (NGS) as a first-tier test. Responses were analyzed qualitatively, and aggregated results are presented herein. The identity of some initiatives is not presented to preserve confidentiality. Results: The findings of the survey indicated that most initiatives are in the planning stage and have not yet started. Although all 14 studies are heterogeneous in design, there is broad consensus that NGS approaches to NBS will, in the short term, be implemented in parallel with current screening programs. The results of this survey can be used to inform the design of studies still in the early planning stages. Conclusion: Here, we provide an overview of NGS-based initiatives in Europe. Importantly, the initiatives described herein will generate evidence to evaluate the utility and feasibility of NGS approaches to NBS, thereby shortening the pathway to responsible implementation of NGS in NBS and informing future research efforts.

Could federated data analysis be the catalyst accelerating the introduction of newborn genome screening for the detection of genetic disease?

Data federation intermediated through trusted research environments can help accelerate the adoption and utilization of newborn genome screening worldwide. Data federation will protect individual datasets from unauthorized security breaches, allow analysis in situ , and bypass the need for cumbersome data sharing agreements between parties. Finally, data federation could accelerate the adoption of new therapies for rare genetic diseases with the use of synthetic clinical trials.

Practical Characterization Strategies for Comparison, Qualification, and Selection of Cell Viability Detection Methods for Cellular Therapeutic Product Development and Manufacturing.

Cellular therapy development and manufacturing has focused on providing novel therapeutic cell-based products for various diseases. The International Organization for Standardization (ISO) has provided guidance on critical quality attributes (CQAs) that shall be considered when testing and releasing cellular therapeutic products. Cell count and viability measurements are two of the CQAs that are determined during development, manufacturing, testing, and product release. The ISO Cell Counting Standard Part 1 and 2 addressed the needs for improving the quality of cell counting results. However, there is currently no guidance on the qualification and selection of a fit-for-purpose cell viability detection method. In this work, we present strategies for the characterization and comparison of AO/PI and AO/DAPI staining methods using the heat-killed (HK) and low temperature/nutrient-deprived (LT/ND) cell death models to evaluate the comparability of cell viability measurements and identify potential causes of differences. We compared the AO/PI and AO/DAPI staining methods using HK and LT/ND-generated dead cells, investigated the staining time effects on cell viability measurements, and determined their viability linearity with different mixtures of live and dead cells. Furthermore, we validated AO/PI and AO/DAPI cell viability measurement with a long-term cell proliferation assay. Finally, we demonstrate a practical example of cell viability measurement comparison using AO/PI and AO/DAPI on antibiotic-selected transduced Jurkat and THP-1 cells to select a fit-for-purpose method for functional genomics screening. The proposed strategies may potentially enable scientists to properly characterize, compare, and select cell viability detection methods that are critical for cellular therapeutic product development and manufacturing.

Applying the R = MC2 implementation science heuristic to assess the impact of readiness on reach and implementation of a population-wide genomic screening program.

Population-wide genomic screening for genes that have high penetrance and clinical actionability enhances the opportunity to identify individuals at risk for developing hereditary conditions. Organizational readiness has been shown to influence the likelihood of successful implementation of complex initiatives such as the integration of population-wide genomic screening in clinical settings. We use the organizational readiness heuristic R = MC2 to better understand three factors that influence readiness for implementation of In Our DNA SC, a population-wide genomic screening program: motivation to implement, general capacity of an organization, and innovation-specific capacities. We then assessed the influence of these readiness factors on implementation outcomes of reach (measured through enrollment rate) and implementation (measured through the number of DNA samples collected). Data were collected pre-implementation and captured during the three-month pilot phase of the In Our DNA SC program. We collected administrative data from the electronic health record and quantitatively captured elements of readiness through surveys distributed to provider champions and clinical administrative champions at the 10 sites implementing the population-wide genomic screening program. We facilitated innovation-specific capacity through training offered at each site, as well as technical assistance through weekly meetings with other implementing sites, and resources available to all staff. Forty percent of provider champions attended training and 80% of administrative champions attended training. An average of 3.7 additional staff were trained at each implementing site. Satisfaction with training positively influenced reach (β = 0.0121, p = 0.0271) but did not impact implementation. Provider engagement (innovation capabilities) was associated with reach (β = 0.0020, p = 0.0251) and clinical administrator engagement was associated with sample collection rate (β = 0.2599, β = 0.038). Readiness to change is considered one of the most important factors in understanding the potential opportunity for implementation. We found that motivation to adopt a population-wide genomic screening program positively impacted the program's reach. The type of champion influenced discrete outcomes, with provider champions positively impacting reach and administrative champions influencing implementation (assessed through sample collection rate). As genomics continues to be integrated into clinical practice, it will be important to understand the contextual factors that influence readiness for implementation and design support throughout the life-course of implementation to ensure the success of large-scale, complex initiatives.

Unequal global implementation of genomic newborn screening.

Unequal global implementation of genomic newborn screening.

Genome sequencing-based transcriptomic analysis reveals novel genes in Peucedanum praeruptorum

BackgroundPeucedanum praeruptorum Dunn, a traditional Chinese herbal medicine, contains coumarin and volatile oil components that have clinical application value. However, early bolting often occurs in the medicinal materials of Apiaceae plants. The rhizomes of the medicinal parts are gradually lignified after bolting, resulting in a sharp decrease in the content of coumarins. At present, the link between coumarin biosynthesis and early bolting in P. praeruptorum has not been elucidated.ResultsCombining the genome sequencing and the previous transcriptome sequencing results, we reanalyzed the differential transcripts of P. praeruptorum before and after bolting. A total of 62,088 new transcripts were identified, of which 31,500 were unknown transcripts. Functional classification and annotation showed that many genes were involved in the regulation of transcription, defense response, and carbohydrate metabolic processes. The main domains are the pentatricopeptide repeat, protein kinase, RNA recognition motif, leucine-rich repeat, and ankyrin repeat domains, indicating their pivotal roles in protein modification and signal transduction. Gene structure analysis showed that skipped exon (SE) was the most dominant alternative splicing, followed by the alternative 3’ splice site (A3SS) and the alternative 5’ splice site (A5SS). Functional enrichment of differentially expressed genes showed that these differentially expressed genes mainly include transmembrane transporters, channel proteins, DNA-binding proteins, polysaccharide-binding proteins, etc. In addition, genes involved in peroxisome, hexose phosphate pathway, phosphatidylinositol signaling system, and inositol phosphate metabolism pathway were greatly enriched. A protein-protein interaction network analysis discoverd 1,457 pairs of proteins that interact with each other. The expression levels of six UbiA genes, three UGT genes, and four OMT genes were higher during the bolting stage. This observation suggests their potential involvement in the catalytic processes of prenylation, glycosylation, and methylation of coumarins, respectively. A total of 100 peroxidase (PRX) genes were identified being involved in lignin polymerization, but only nine PRX genes were highly expressed at the bolting stage. It is worth noting that 73 autophagy-related genes (ATGs) were first identified from the KEGG pathway-enriched genes. Some ATGs, such as BHQH00009837, BHQH00013830, and novel8944, had higher expression levels after bolting.ConclusionsComparative transcriptome analysis and large-scale genome screening provide guidance and new opinions for the identification of bolting-related genes in P. praeruptorum.

Increased copy number of the target gene squalene monooxygenase as the main resistance mechanism to terbinafine in Leishmania infantum

We use here two genomic screens in an attempt to understand the mode of action and resistance mechanism of terbinafine, an antifungal contemplated as a potential drug against the parasite Leishmania. One screen consisted in in vitro drug evolution where 5 independent mutants were selected step-by-step for terbinafine resistance. Sequencing of the genome of the 5 mutants revealed no single nucleotide polymorphisms related to the resistance phenotype. However, the ERG1 gene was found amplified as part of a linear amplicon, and transfection of ERG1 fully recapitulated the terbinafine resistance phenotype of the mutants. The second screen, Cos-seq, consisted in selecting a gene overexpression library with terbinafine followed by the sequencing of the enriched cosmids. This screen identified two cosmids derived from loci on chromosomes 13 and 29 encoding the squalene monooxygenase (ERG1) and the C8 sterol isomerase (ERG2), respectively. Transfection of the ERG1-cosmid, but not the ERG2-cosmid, produced resistance to terbinafine. Our screens suggest that ERG1 is the main, if not only, target for terbinafine in Leishmania and amplification of its gene is the main resistance mechanism.

Combination of Genomic Landsscape and 3D Culture Functional Assays Bridges Sarcoma Phenotype to Target and Immunotherapy.

Genomic-based precision medicine has not only improved tumour therapy but has also shown its weaknesses. Genomic profiling and mutation analysis have identified alterations that play a major role in sarcoma pathogenesis and evolution. However, they have not been sufficient in predicting tumour vulnerability and advancing treatment. The relative rarity of sarcomas and the genetic heterogeneity between subtypes also stand in the way of gaining statistically significant results from clinical trials. Personalized three-dimensional tumour models that reflect the specific histologic subtype are emerging as functional assays to test anticancer drugs, complementing genomic screening. Here, we provide an overview of current target therapy for sarcomas and discuss functional assays based on 3D models that, by recapitulating the molecular pathways and tumour microenvironment, may predict patient response to treatments. This approach opens new avenues to improve precision medicine when genomic and pathway alterations are not sufficient to guide the choice of the most promising treatment. Furthermore, we discuss the aspects of the 3D culture assays that need to be improved, such as the standardisation of growth conditions and the definition of in vitro responses that can be used as a cut-off for clinical implementation.

Principles of Genetic Counseling in Inherited Heart Conditions.

Cardiac genetic counseling is the process of helping individuals adapt to a personal diagnosis or family history of an inherited heart condition. The process is shown to benefit patients and includes specialized skills, such as counseling children and interpreting complex genetic results. Emerging areas include: evolving service delivery models for caring for patients and communicating risk to relatives, new areas of need including postmortem molecular autopsy, and new populations of individuals found to carry a likely pathogenic/pathogenic cardiac variant identified through genomic screening. This article provides an overview of the cardiac genetic counseling process and evolving areas in the field.

A small, polyphyletic group of Firmicutes synthesizes trimethylamine froml-carnitine.

Gut microbiota-derived trimethylamine (TMA) is associated with cardiometabolic disorders and exemplifies a microbial involvement in the etiology of emerging, noncommunicable diseases, the leading causes of death worldwide. Three biochemical pathways taking dietary compounds as intake have been described with distinct taxa involved that are all present at low relative abundances. A recently discovered pathway is now considered to be the main route for TMA synthesis from l-carnitine involving γ-butyrobetaine as an intermediate product. By comprehensive (meta) genomic screening of publicly available data, namely, genomes of the UHGG catalog (n > 200,000) and 10 metagenomic (transcriptomic) data sets, we revealed bacteria synthesizing TMA via this pathway and specified their ecophysiology. Results will contribute to stratification of individuals based on their gut microbiota's potential to synthesize TMA and might aid in the development of strategies restricting TMA formation.

Understanding cancer drug resistance with Sleeping Beauty functional genomic screens: Application to MAPK inhibition in cutaneous melanoma

Combined BRAF and MEK inhibition is an effective treatment for BRAF-mutant cutaneous melanoma. However, most patients progress on this treatment due to drug resistance. Here, we applied the Sleeping Beauty transposon system to understand how melanoma evades MAPK inhibition. We found that the specific drug resistance mechanisms differed across melanomas in our genetic screens of five cutaneous melanoma cell lines. While drivers that reactivated MAPK were highly conserved, many others were cell-line specific. One such driver, VAV1, activated a de-differentiated transcriptional program like that of hyperactive RAC1, RAC1P29S. To target this mechanism, we showed that an inhibitor of SRC, saracatinib, blunts the VAV1-induced transcriptional reprogramming. Overall, we highlighted the importance of accounting for melanoma heterogeneity in treating cutaneous melanoma with MAPK inhibitors. Moreover, we demonstrated the utility of the Sleeping Beauty transposon system in understanding cancer drug resistance.

Targeted and Logical Discovery of Piperazic Acid-Bearing Natural Products Based on Genomic and Spectroscopic Signatures.

A targeted and logical discovery method was devised for natural products containing piperazic acid (Piz), which is biosynthesized from ornithine by l-ornithine N-hydroxylase (KtzI) and N-N bond formation enzyme (KtzT). Genomic signature-based screening of a bacterial DNA library (2020 strains) using polymerase chain reaction (PCR) primers targeting ktzT identified 62 strains (3.1%). The PCR amplicons of KtzT-encoding genes were phylogenetically analyzed to classify the 23 clades into two monophyletic groups, I and II. Cultivating hit strains in media supplemented with 15NH4Cl and applying 1H-15N heteronuclear multiple bond correlation (HMBC) along with 1H-15N heteronuclear single quantum coherence (HSQC) and 1H-15N HSQC-total correlation spectroscopy (HSQC-TOCSY) NMR experiments detected the spectroscopic signatures of Piz and modified Piz. Chemical investigation of the hit strains prioritized by genomic and spectroscopic signatures led to the identification of a new azinothricin congener, polyoxyperuin B seco acid (1), previously reported chloptosin (2) in group I, depsidomycin D (3) incorporating two dehydropiperazic acids (Dpz), and lenziamides A and B (4 and 5), structurally novel 31-membered cyclic decapeptides in group II. By consolidating the phylogenetic and chemical analyses, clade-structure relationships were elucidated for 19 of the 23 clades. Lenziamide A (4) inhibited STAT3 activation and induced G2/M cell cycle arrest, apoptotic cell death, and tumor growth suppression in human colorectal cancer cells. Moreover, lenziamide A (4) resensitized 5-fluorouracil (5-FU) activity in both in vitro cell cultures and the in vivo 5-FU-resistant tumor xenograft mouse model. This work demonstrates that the genomic and spectroscopic signature-based searches provide an efficient and general strategy for new bioactive natural products containing specific structural motifs.

Exploring biomarkers for autophagy-mediated macrophage pyroptosis in atherosclerosis.

This study tried to investigate the macrophage autophagy-related pyroptosis in atherosclerosis. The gene expression omnibus (GEO) dataset of GSE100927 was used for differentially expressed genes (DEG) screening, gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG), CIBERSORT, weighted correlation network analysis (WGCNA), receiver operating characteristic (ROC), gene set enrichment analysis (GSEA), and correlation analysis, and GSE159677 was used for single-cell analysis, all conducted in R software. Protein-protein interaction (PPI) was constructed in STRING and analyzed in Cytoscape. Transcription factors, drugs, and tissue co-expression network were explored in NetworkAnalyst. A total of 110 autophagy-related DEG (DEATG) were identified, and GO/KEGG revealed the top items enriched in autophagy, phagosome and lysosome. CIBERSORT showed 11 cell types were markedly differentially expressed (p < .05). WGCNA found the turquoise and yellow module were positively correlated with macrophage M0 (corr = 0.5, P = 6e-6) and M2 (corr = 0.54, P = 1e-6), respectively. Then 35 immune-related DEATG were identified, and functional analysis showed immune effector process, interleukin-6 and myeloid cell activation were enriched besides autophagy. PPI and MCC algorithm identified 6 hub genes in regulating macrophage-related autophagy, and ROC indicated high prediction value (area under curve = 0.961). GSEA enriched 6 common pathways associated with autophagy and atherosclerosis pathogenesis, and immune correlation suggested these hub genes were correlated with macrophages M0/M1, monocytes and T cells. Then venn plot found 3 central genes in mediating macrophage autophagy-associated pyroptosis in atherosclerosis, and single-cell analysis demonstrated cell distribution, then validated in THPA human samples. Our data discovered hub genes responsible for macrophage autophagy-mediated pyroptosis in atherosclerosis, and functional analysis with immune cell distribution evidenced their high phenotype-trait prediction value.

A systematic review of real-world applications of genome sequencing for newborn screening

Aim: With the costs of genomic sequencing falling quickly and an ever-increasing number of clinical laboratories equipped with new-generation sequencing machines, healthcare systems around the world are getting ready to enter the era of genomic newborn screening (NBS). However, the adoption of Genomic Sequencing (GS), encompassing whole-exome sequencing (WES) and whole-genome sequencing (WGS), in NBS programs raises a number of clinical, ethical, and legal questions as well as organizational and economic challenges. This systematic review is part of a feasibility study to assess the introduction of WGS for NBS in Lombardy region with the specific aim of gathering evidence from existing pilots in the field whose results have been published. Methods: Three different sources were identified for the selection of articles in order to obtain a various and unbiased set of publications. 33 articles were retained for analysis to answer the following questions: 1. Clinical: Does genomic sequencing demonstrate clinical utility in the context of NBS? What are the limitations of these kind of programs? 2. Societal: What are the social, ethical and psychological implications of using GS for NBS? 3. Governance: What are the legal, economic, and organizational challenges for GS-based NBS programs? Results: There is a general consensus in the literature on the key principles that should guide the adoption of GS in NBS, such as the inclusion of actionable genes only, the need for informed consent from the parents, the right of the newborn to an open future, which means the exclusion of late-onset diseases even when those are considered treatable. However, there are still several differences in how these principles are detailed and applied. Conclusion: Real-world evidence from a handful of pilot projects (namely BabySeq and NC-Nexus, both carried out in the USA) have been published recently; however, this evidence is not yet sufficient to put an end to the broad and animated debate on the use of GS for NBS. Ethical, legal, and social issues still constitute great challenges and major barriers to wide and uniform adoption of GS in NBS. On the clinical side, a number of issues remain unaddressed, such as the benefits and limitations of the different approaches (targeted sequencing, GS only versus GS+standard NBS), the genes/diseases to include and the frequency of incidental findings, identification of carrier status, and variants of uncertain significance (VUS). Further pilots and consultations with involved stakeholders will be necessary before GS-based NBS can be accepted and systematically implemented in national healthcare programs.

Exploring Genetic Markers: Mitochondrial DNA and Genomic Screening for Biodiversity and Production Traits in Donkeys.

Donkeys (Equus asinus) play a pivotal role as essential livestock in arid and semi-arid regions, serving various purposes such as transportation, agriculture, and milk production. Despite their significance, donkey breeding has often been overlooked in comparison to other livestock species, resulting in limited genetic improvement programs. Preserving donkey genetic resources within each country necessitates the establishment of breed conservation programs, focusing on managing genetic diversity among populations. In recent years, significant strides have been made in sequencing and analyzing complete mitochondrial DNA (mtDNA) molecules in donkeys. Notably, numerous studies have honed in on the mitochondrial D-loop region, renowned for its remarkable variability and higher substitution rate within the mtDNA genome, rendering it an effective genetic marker for assessing genetic diversity in donkeys. Furthermore, genetic markers at the RNA/DNA level have emerged as indispensable tools for enhancing production and reproduction traits in donkeys. Traditional animal breeding approaches based solely on phenotypic traits, such as milk yields, weight, and height, are influenced by both genetic and environmental factors. To overcome these challenges, genetic markers, such as polymorphisms, InDel, or entire gene sequences associated with desirable traits in animals, have achieved widespread usage in animal breeding practices. These markers have proven increasingly valuable for facilitating the selection of productive and reproductive traits in donkeys. This comprehensive review examines the cutting-edge research on mitochondrial DNA as a tool for assessing donkey biodiversity. Additionally, it highlights the role of genetic markers at the DNA/RNA level, enabling the informed selection of optimal production and reproductive traits in donkeys, thereby driving advancements in donkey genetic conservation and breeding programs.