Recurrent de novo missense variants across multiple histone H4 genes underlie a neurodevelopmental syndrome.

Chromatin is essentially an array of nucleosomes, each of which consists of the DNA double-stranded fiber wrapped around a histone octamer. This organization supports cellular processes such as DNA replication, DNA transcription, and DNA repair in all eukaryotes. Human histone H4 is encoded by fourteen canonical histone H4 genes, all differing at the nucleotide level but encoding an invariant protein. Here, we present a cohort of 29 subjects with de novo missense variants in six H4 genes (H4C3, H4C4, H4C5, H4C6, H4C9, and H4C11) identified by whole-exome sequencing and matchmaking. All individuals present with neurodevelopmental features of intellectual disability and motor and/or gross developmental delay, while non-neurological features are more variable. Ten amino acids are affected, six recurrently, and are all located within the H4 core or C-terminal tail. These variants cluster to specific regions of the core H4 globular domain, where protein-protein interactions occur with either other histone subunits or histone chaperones. Functional consequences of the identified variants were evaluated in zebrafish embryos, which displayed abnormal general development, defective head organs, and reduced body axis length, providing compelling evidence for the causality of the reported disorder(s). While multiple developmental syndromes have been linked to chromatin-associated factors, missense-bearing histone variants (e.g., H3 oncohistones) are only recently emerging as a major cause of pathogenicity. Our findings establish a broader involvement of H4 variants in developmental syndromes.

Longitudinal multi-omics of host-microbe dynamics in prediabetes.

Type 2 diabetes mellitus (T2D) is a growing health problem, but little is known about its early disease stages, its effects on biological processes or the transition to clinical T2D. To understand the earliest stages of T2D better, we obtained samples from 106 healthy individuals and individuals with prediabetes over approximately four years and performed deep profiling of transcriptomes, metabolomes, cytokines, and proteomes, as well as changes in the microbiome. This rich longitudinal data set revealed many insights: first, healthy profiles are distinct among individuals while displaying diverse patterns of intra- and/or inter-personal variability. Second, extensive host and microbial changes occur during respiratory viral infections and immunization, and immunization triggers potentially protective responses that are distinct from responses to respiratory viral infections. Moreover, during respiratory viral infections, insulin-resistant participants respond differently than insulin-sensitive participants. Third, global co-association analyses among the thousands of profiled molecules reveal specific host-microbe interactions that differ between insulin-resistant and insulin-sensitive individuals. Last, we identified early personal molecular signatures in one individual that preceded the onset of T2D, including the inflammation markers interleukin-1 receptor agonist (IL-1RA) and high-sensitivity C-reactive protein (CRP) paired with xenobiotic-induced immune signalling. Our study reveals insights into pathways and responses that differ between glucose-dysregulated and healthy individuals during health and disease and provides an open-access data resource to enable further research into healthy, prediabetic and T2D states.

Implementing evidence-based assertions of clinical actionability in the context of secondary findings: Updates from the ClinGen Actionability Working Group.

PURPOSE: The ClinGen Actionability Working Group (AWG) developed an evidence-based framework to generate actionability reports and scores of gene-condition pairs in the context of secondary findings from genome sequencing. Here we describe the expansion of the framework to include actionability assertions. METHODS: Initial development of the actionability rubric was based on previously scored adult gene-condition pairs and individual expert evaluation. Rubric refinement was iterative and based on evaluation, feedback, and discussion. The final rubric was pragmatically evaluated via integration into actionability assessments for 27 gene-condition pairs. RESULTS: The resulting rubric has a 4-point scale (limited, moderate, strong, and definitive) and uses the highest-scoring outcome-intervention pair of each gene-condition pair to generate a preliminary assertion. During AWG discussions, predefined criteria and factors guide discussion to produce a consensus assertion for a gene-condition pair, which may differ from the preliminary assertion. The AWG has retrospectively generated assertions for all previously scored gene-condition pairs and are prospectively asserting on gene-condition pairs under assessment, having completed over 170 adult and 188 pediatric gene-condition pairs. CONCLUSION: The AWG expanded its framework to provide actionability assertions to enhance the clinical value of their resources and increase their utility as decision aids regarding return of secondary findings.

Information-seeking preferences in diverse patients receiving a genetic testing result in the Clinical Sequencing Evidence-Generating Research (CSER) study.

PURPOSE: Accurate and understandable information after genetic testing is critical for patients, family members, and professionals alike. METHODS: As part of a cross-site study from the Clinical Sequencing Evidence-Generating Research consortium, we investigated the information-seeking practices among patients and family members at 5 to 7 months after genetic testing results disclosure, assessing the perceived utility of a variety of information sources, such as family and friends, health care providers, support groups, and the internet. RESULTS: We found that individuals placed a high value on information obtained from genetics professionals and health care workers, independent of genetic testing result case classifications as positive, inconclusive, or negative. The internet was also highly utilized and ranked. Study participants rated some information sources as more useful for positive results compared with inconclusive or negative outcomes, emphasizing that it may be difficult to identify helpful information for individuals receiving an uncertain or negative result. There were few data from non-English speakers, highlighting the need to develop strategies to reach this population. CONCLUSION: Our study emphasizes the need for clinicians to provide accurate and comprehensible information to individuals from diverse populations after genetic testing.

Candidate variants in TUB are associated with familial tremor.

Essential tremor (ET) is the most common adult-onset movement disorder. In the present study, we performed whole exome sequencing of a large ET-affected family (10 affected and 6 un-affected family members) and identified a TUB p.V431I variant (rs75594955) segregating in a manner consistent with autosomal-dominant inheritance. Subsequent targeted re-sequencing of TUB in 820 unrelated individuals with sporadic ET and 630 controls revealed significant enrichment of rare nonsynonymous TUB variants (e.g. rs75594955: p.V431I, rs1241709665: p.Ile20Phe, rs55648406: p.Arg49Gln) in the ET cohort (SKAT-O test p-value = 6.20e-08). TUB encodes a transcription factor predominantly expressed in neuronal cells and has been previously implicated in obesity. ChIP-seq analyses of the TUB transcription factor across different regions of the mouse brain revealed that TUB regulates the pathways responsible for neurotransmitter production as well thyroid hormone signaling. Together, these results support the association of rare variants in TUB with ET.

Predicting genes from phenotypes using human phenotype ontology (HPO) terms.

The interpretation of genomic variants following whole exome sequencing (WES) can be aided using human phenotype ontology (HPO) terms to standardize clinical features and predict causative genes. We performed WES on 453 patients diagnosed prior to 18 years of age and identified 114 pathogenic (P) or likely pathogenic (LP) variants in 112 patients. We utilized PhenoDB to extract HPO terms from provider notes and then used Phen2Gene to generate a gene score and gene ranking from each list of HPO terms. We assigned Phen2Gene gene rankings to 6 rank classes, with class 1 covering raw gene rankings of 1 to 10 and class 2 covering rankings from 11 to 50 out of a total of 17,126 possible gene rankings. Phen2Gene ranked causative genes into rank class 1 or 2 in 27.7% of cases and the genes in rank class 1 were all associated with well-characterized phenotypes. We found significant associations between the gene score and the number of years, since the gene was first published, the number of HPO terms with an hierarchical depth greater or equal to 11, and the number of Online Mendelian Inheritance in Man terms associated with the phenotype and gene. We conclude that genes associated with recognizable phenotypes and terms deep in the HPO hierarchy have the best chance of producing a high gene score and ranking in class 1 to 2 using Phen2Gene software with HPO terms. Clinicians and laboratory staff should consider these results when HPO terms are employed to prioritize candidate genes.

Integration of stakeholder engagement from development to dissemination in genomic medicine research: Approaches and outcomes from the CSER Consortium.

PURPOSE: There is a critical need for genomic medicine research that reflects and benefits socioeconomically and ancestrally diverse populations. However, disparities in research populations persist, highlighting that traditional study designs and materials may be insufficient or inaccessible to all groups. New approaches can be gained through collaborations with patient/community stakeholders. Although some benefits of stakeholder engagement are recognized, routine incorporation into the design and implementation of genomics research has yet to be realized. METHODS: The National Institutes of Health-funded Clinical Sequencing Evidence-Generating Research (CSER) consortium required stakeholder engagement as a dedicated project component. Each CSER project planned and carried out stakeholder engagement activities with differing goals and expected outcomes. Examples were curated from each project to highlight engagement strategies and outcomes throughout the research lifecycle from development through dissemination. RESULTS: Projects tailored strategies to individual study needs, logistical constraints, and other challenges. Lessons learned include starting early with engagement efforts across project stakeholder groups and planned flexibility to enable adaptations throughout the project lifecycle. CONCLUSION: Each CSER project used more than 1 approach to engage with relevant stakeholders, resulting in numerous adaptations and tremendous value added throughout the full research lifecycle. Incorporation of community stakeholder insight improves the outcomes and relevance of genomic medicine research.

Perspectives and preferences regarding genomic secondary findings in underrepresented prenatal and pediatric populations: A mixed-methods approach.

PURPOSE: Patients undergoing clinical exome sequencing (ES) are routinely offered the option to receive secondary findings (SF). However, little is known about the views of individuals from underrepresented minority pediatric or prenatal populations regarding SF. METHODS: We explored the preferences for receiving hypothetical categories of SF (H-SF) and reasons for accepting or declining actual SF through surveying (n = 149) and/or interviewing (n = 47) 190 families undergoing pediatric or prenatal ES. RESULTS: Underrepresented minorities made up 75% of the probands. In total, 150 families (79%) accepted SF as part of their child/fetus's ES. Most families (63%) wanted all categories of H-SF. Those who declined SF as part of ES were less likely to want H-SF across all categories. Interview findings indicate that some families did not recall their SF decision. Preparing for the future was a major motivator for accepting SF, and concerns about privacy, discrimination, and psychological effect drove decliners. CONCLUSION: A notable subset of families (37%) did not want at least 1 category of H-SF, suggesting more hesitancy about receiving all available results than previously reported. The lack of recollection of SF decisions suggests a need for alternative communication approaches. Results highlight the importance of the inclusion of diverse populations in genomic research.

Preference for secondary findings in prenatal and pediatric exome sequencing.

OBJECTIVE: We aimed to determine the frequency of accepting secondary findings in families undergoing exome sequencing in prenatal and pediatric settings. METHODS: This was a secondary analysis of prospectively enrolled patients undergoing trio exome sequencing for congenital anomalies or developmental disorders in prenatal and pediatric settings, in which families were offered receiving secondary findings (initially assessed in the proband and, if identified, then in the parents). The primary outcome was frequency of accepting secondary findings. Secondary outcomes included frequency of acceptance in prenatal versus pediatric settings, and sociodemographic differences between those who accepted versus declined secondary findings. RESULTS: There were 682 families included in the cohort (289 prenatal and 393 pediatric). Overall, 84% (576/682) of families accepted secondary findings: 86.2% (249/289) of families undergoing prenatal versus 83.2% (327/393) pediatric (p = 0.30) testing. Secondary findings were identified in 2.6% (15/576) of cases, with no difference between prenatal and pediatric settings. There were no differences in sociodemographics between families that accepted versus declined secondary findings. CONCLUSION: The majority of families undergoing exome sequencing accepted secondary findings; this did not differ in prenatal versus pediatric settings. This highlights the need for guidance surrounding the offer of secondary findings in the prenatal setting.

New cases that expand the genotypic and phenotypic spectrum of Congenital NAD Deficiency Disorder.

Nicotinamide adenine dinucleotide (NAD) is an essential coenzyme involved in over 400 cellular reactions. During embryogenesis, mammals synthesize NAD de novo from dietary l -tryptophan via the kynurenine pathway. Biallelic, inactivating variants in three genes encoding enzymes of this biosynthesis pathway (KYNU, HAAO, and NADSYN1) disrupt NAD synthesis and have been identified in patients with multiple malformations of the heart, kidney, vertebrae, and limbs; these patients have Congenital NAD Deficiency Disorder HAAO and four families with biallelic variants in KYNU. These patients present similarly with multiple malformations of the heart, kidney, vertebrae, and limbs, of variable severity. We show that each variant identified in these patients results in loss-of-function, revealed by a significant reduction in NAD levels via yeast genetic complementation assays. For the first time, missense mutations are identified as a cause of malformation and shown to disrupt enzyme function. These missense and frameshift variants cause moderate to severe NAD deficiency in yeast, analogous to insufficient synthesized NAD in patients. We hereby expand the genotypic and corresponding phenotypic spectrum of Congenital NAD Deficiency Disorder.

A review and definition of 'usual care' in genetic counseling trials to standardize use in research.

The descriptor 'usual care' refers to standard or routine care. Yet, no formal definition exists. The need to define what constitutes usual care arises in clinical research. Often one arm in a trial represents usual care in comparison with a novel intervention. Accordingly, usual care in genetic counseling research appears predominantly in randomized controlled trials. Recent standards for reporting genetic counseling research call for standardization, but do not address usual care. We (1) inventoried all seven studies in the Clinical Sequencing Evidence-Generating Consortium (CSER) about how genetic counseling was conceptualized, conducted, and whether a usual care arm was involved; (2) conducted a review of published randomized control trials in genetic counseling, comparing how researchers describe usual care groups; and (3) reviewed existing professionally endorsed definitions and practice descriptions of genetic counseling. We found wide variation in the content and delivery of usual care. Descriptions frequently detailed the content of usual care, most often noting assessment of genetic risk factors, collecting family histories, and offering testing. A minority included addressing psychological concerns or the risks versus benefits of testing. Descriptions of how care was delivered were vague except for mode and type of clinician, which varied. This significant variation, beyond differences expected among subspecialties, reduces the validity and generalizability of genetic counseling research. Ideally, research reflects clinical practice so that evidence generated can be used to improve clinical outcomes. To address this objective, we propose a definition of usual care in genetic counseling research that merges common elements from the National Society of Genetic Counselors' practice definition, the Reciprocal Engagement Model, and the Accreditation Council for Genetic Counselors' practice-based competencies. Promoting consistent execution of usual care in the design of genetic counseling trials can lead to more consistency in representing clinical care and facilitate the generation of evidence to improve it.

The expanding spectrum of NFIB-associated phenotypes in a diverse patient population-A report of two new patients.

NFIB (Nuclear Factor I B) haploinsufficiency has recently been identified as a cause of intellectual disability and macrocephaly. Here we describe two patients with pathogenic variants in NFIB. The first is a 6-year-old Latino male with developmental delays, mild hypotonia, facial anomalies, and brain magnetic resonance imaging findings comprising mild thinning of the corpus callosum, with more marked thinning of the splenium and blunting of the rostrum and cavum septum pellucidum. Exome sequencing identified a previously described de novo variant in NFIB, c.265C>T, predicting p.Arg89Ter. The second is a 5-year-old Latino male with developmental delays, hypotonia, dysmorphic features, a preauricular tag and pit, a small ventricular septal defect, and brain magnetic resonance imaging findings including a dysmorphic corpus callosum and a small posterior fossa. A single nucleotide polymorphism microarray identified a 92 kb interstitial deletion at 9p23 including several exons of NFIB and no other known genes. Our two patients add to the knowledge of this rare condition through our addition of new brain MRI findings and dysmorphic features. Additionally, these are the first known Latino patients to be described with NFIB haploinsufficiency, expanding our understanding of the associated facial features in diverse populations. Further data are needed to determine genotype-phenotype relationships for NFIB.

A novel truncating variant in ring finger protein 113A (RNF113A) confirms the association of this gene with X-linked trichothiodystrophy.

We describe an 11-year old boy with severe global developmental delays, failure to thrive and growth retardation, refractory seizures with recurrent status epilepticus, hypogammaglobulinemia, hypergonadotropic hypogonadism, and duodenal strictures. He had facial and skin findings compatible with trichothiodystrophy, including sparse and brittle hair, thin eyebrows, and dry skin. Exome sequencing showed a hemizygous, truncating variant in RNF113A, c.903_910delGCAGACCA, predicting p.(Gln302fs*12), that was inherited from his mother. Although his clinical features overlap closely with features described in the two previously reported male first cousins with RNF113A loss of function mutations, the duodenal strictures seen in this patient have not been reported. Interestingly, the patient's mother had short stature and 100% skewed X-inactivation as seen in other obligate female carriers. A second male with developmental delays, microcephaly, seizures, ambiguous genitalia, and facial anomalies that included sparse and brittle hair, thin eyebrows and dry skin was recently reported to have c.897_898delTG, predicting p.(Cys299*) in RNF113A and we provide additional clinical details for this patient. This report further supports deleterious variants in RNF113A as a cause of a novel trichothiodystrophy syndrome.

Digital Health: Tracking Physiomes and Activity Using Wearable Biosensors Reveals Useful Health-Related Information.

A new wave of portable biosensors allows frequent measurement of health-related physiology. We investigated the use of these devices to monitor human physiological changes during various activities and their role in managing health and diagnosing and analyzing disease. By recording over 250,000 daily measurements for up to 43 individuals, we found personalized circadian differences in physiological parameters, replicating previous physiological findings. Interestingly, we found striking changes in particular environments, such as airline flights (decreased peripheral capillary oxygen saturation [SpO2] and increased radiation exposure). These events are associated with physiological macro-phenotypes such as fatigue, providing a strong association between reduced pressure/oxygen and fatigue on high-altitude flights. Importantly, we combined biosensor information with frequent medical measurements and made two important observations: First, wearable devices were useful in identification of early signs of Lyme disease and inflammatory responses; we used this information to develop a personalized, activity-based normalization framework to identify abnormal physiological signals from longitudinal data for facile disease detection. Second, wearables distinguish physiological differences between insulin-sensitive and -resistant individuals. Overall, these results indicate that portable biosensors provide useful information for monitoring personal activities and physiology and are likely to play an important role in managing health and enabling affordable health care access to groups traditionally limited by socioeconomic class or remote geography.

Developmental and epileptic encephalopathy in two siblings with a novel, homozygous missense variant in SCN1B.

Developmental and epileptic encephalopathies are genetic disorders in which both the developmental disability and the frequent epileptic activity are the effect of a specific gene variant. While heterozygous variants in SCN1B have been described in families with generalized epilepsy with febrile seizures plus, Type 1, only three cases of homozygous, missense variants in SCN1B have been reported in association with autosomal recessive inheritance of a severe developmental and epileptic encephalopathy. We present two siblings who are homozygous for a novel, missense variant in SCN1B, c.265C>T, predicting p.Arg89Cys. The proband is an 11-year-old female with infantile-onset, fever-induced, intractable generalized tonic-clonic seizures, myoclonic seizures, and developmental slowing and autism spectrum disorder occurring later in the course of the disease. Her 4-year-old brother had a similar epilepsy phenotype, but still displays normal development. This variant has not been previously reported in the homozygous state in control databases. The variant was predicted to be damaging and occurred in the vicinity of other epileptic encephalopathy-associated missense variants that are biallelic and located in the extracellular immunoglobulin loop domain of the protein, which mediates interaction of the beta-1 subunit with cellular adhesion molecules. Our report is the first set of siblings with homozygosity for the p.Arg89Cys variant in SCN1B and further implicates biallelic mutations in this gene as a cause of epileptic encephalopathy mimicking Dravet syndrome. Interestingly, the phenotype we observed was milder compared to that previously described in patients with recessive SCN1B mutations.

Much ado about nothing: A qualitative study of the experiences of an average-risk population receiving results of exome sequencing.

The increasing availability of exome sequencing to the general ("healthy") population raises questions about the implications of genomic testing for individuals without suspected Mendelian diseases. Little is known about this population's motivations for undergoing exome sequencing, their expectations, reactions, and perceptions of utility. In order to address these questions, we conducted in-depth semi-structured interviews with 12 participants recruited from a longitudinal multi-omics profiling study that included exome sequencing. Participants were interviewed after receiving exome results, which included Mendelian disease-associated pathogenic and likely pathogenic variants, pharmacogenetic variants, and risk assessments for multifactorial diseases such as type 2 diabetes. The primary motivation driving participation in exome sequencing was personal curiosity. While they reported feeling validation and relief, participants were frequently underwhelmed by the results and described having expected more from exome sequencing. All participants reported discussing the results with at least some family, friends, and healthcare providers. Participants' recollection of the results returned to them was sometimes incorrect or incomplete, in many cases aligning with their perceptions of their health risks when entering the study. These results underscore the need for different genetic counseling approaches for generally healthy patients undergoing exome sequencing, in particular the need to provide anticipatory guidance to moderate participants' expectations. They also provide a preview of potential challenges clinicians may face as genomic sequencing continues to scale-up in the general population despite a lack of full understanding of its impact.

Developing a genomics rotation: Practical training around variant interpretation for genetic counseling students.

With the wide adoption of next-generation sequencing (NGS)-based genetic tests, genetic counselors require increased familiarity with NGS technology, variant interpretation concepts, and variant assessment tools. The use of exome and genome sequencing in clinical care has expanded the reach and diversity of genetic testing. Regardless of the setting where genetic counselors are performing variant interpretation or reporting, most of them have learned these skills from colleagues, while on the job. Though traditional, lecture-based learning around these topics is important, there has been growing need for the inclusion of case-based, experiential training of genomics and variant interpretation for genetic counseling students, with the goal of creating a strong foundation in variant interpretation for new genetic counselors, regardless of what area of practice they enter. To address this need, we established a genomics and variant interpretation rotation for Stanford's genetic counseling training program. In response to changes in the genomics landscape, this has now evolved into three unique rotation experiences, each focused on variant interpretation in the context of various genomic settings, including clinical laboratory, research laboratory, and healthy genomic analysis studies. Here, we describe the goals and learning objectives that we have developed for these variant interpretation rotations, and illustrate how these concepts are applied in practice.

Evidence-based assessments of clinical actionability in the context of secondary findings: Updates from ClinGen's Actionability Working Group.

The use of genome-scale sequencing allows for identification of genetic findings beyond the original indication for testing (secondary findings). The ClinGen Actionability Working Group's (AWG) protocol for evidence synthesis and semi-quantitative metric scoring evaluates four domains of clinical actionability for potential secondary findings: severity and likelihood of the outcome, and effectiveness and nature of the intervention. As of February 2018, the AWG has scored 127 genes associated with 78 disorders (up-to-date topics/scores are available at www.clinicalgenome.org). Scores across these disorders were assessed to compare genes/disorders recommended for return as secondary findings by the American College of Medical Genetics and Genomics (ACMG) with those not currently recommended. Disorders recommended by the ACMG scored higher on outcome-related domains (severity and likelihood), but not on intervention-related domains (effectiveness and nature of the intervention). Current practices indicate that return of secondary findings will expand beyond those currently recommended by the ACMG. The ClinGen AWG evidence reports and summary scores are not intended as classifications of actionability, rather they provide a resource to aid decision makers as they determine best practices regarding secondary findings. The ClinGen AWG is working with the ACMG Secondary Findings Committee to update future iterations of their secondary findings list.

Association of AHSG with alopecia and mental retardation (APMR) syndrome.

Alopecia with mental retardation syndrome (APMR) is a very rare autosomal recessive condition that is associated with total or partial absence of hair from the scalp and other parts of the body as well as variable intellectual disability. Here we present whole-exome sequencing results of a large consanguineous family segregating APMR syndrome with seven affected family members. Our study revealed a novel predicted pathogenic, homozygous missense mutation in the AHSG (OMIM 138680) gene (AHSG: NM_001622:exon7:c.950G>A:p.Arg317His). The variant is predicted to affect a region of the protein required for protein processing and disrupts a phosphorylation motif. In addition, the altered protein migrates with an aberrant size relative to healthy individuals. Consistent with the phenotype, AHSG maps within APMR linkage region 1 (APMR 1) as reported before, and falls within runs of homozygosity (ROH). Previous families with APMR syndrome have been studied through linkage analyses and the linkage resolution did not allow pointing out to a single gene candidate. Our study is the first report to identify a homozygous missense mutation for APMR syndrome through whole-exome sequencing.

The Need to Standardize the Reanalysis of Genomic Sequencing Results: Findings from Interviews with Underserved Families in Genomic Research.

The reanalysis of genomic sequencing results has the potential to provide results that are of considerable medical and personal importance to recipients. Employing interviews with forty-seven predominantly medically underserved families and ethnographic observations we argue that there is pressing need to standardize the approach taken to reanalysis. Our findings highlight that study participants were unclear as to the likelihood of reanalysis happening, the process of initiating reanalysis, and whether they would receive revised results. Their reflections mirror the lack a specific focus upon reanalysis within consent and results sessions as observed in clinical settings. Mechanisms need to be put into place that standardize the approach to reanalysis in research and in clinical contexts. This would enable clinicians and genetic counsellors to communicate clearly with research participants with respect to potential for reanalysis of results and the process of reanalysis. We argue that that the role of reanalysis is too important to be referred to in an ad-hoc manner. Furthermore, the ad-hoc nature of the current process may increase health inequities given the likelihood that only those families who have the means to press for reanalysis are likely to receive it.