Selective inhibition of CDK7 reveals high-confidence targets and new models for TFIIH function in transcription.

CDK7 associates with the 10-subunit TFIIH complex and regulates transcription by phosphorylating the C-terminal domain (CTD) of RNA polymerase II (RNAPII). Few additional CDK7 substrates are known. Here, using the covalent inhibitor SY-351 and quantitative phosphoproteomics, we identified CDK7 kinase substrates in human cells. Among hundreds of high-confidence targets, the vast majority are unique to CDK7 (i.e., distinct from other transcription-associated kinases), with a subset that suggest novel cellular functions. Transcription-associated factors were predominant CDK7 substrates, including SF3B1, U2AF2, and other splicing components. Accordingly, widespread and diverse splicing defects, such as alternative exon inclusion and intron retention, were characterized in CDK7-inhibited cells. Combined with biochemical assays, we establish that CDK7 directly activates other transcription-associated kinases CDK9, CDK12, and CDK13, invoking a "master regulator" role in transcription. We further demonstrate that TFIIH restricts CDK7 kinase function to the RNAPII CTD, whereas other substrates (e.g., SPT5 and SF3B1) are phosphorylated by the three-subunit CDK-activating kinase (CAK; CCNH, MAT1, and CDK7). These results suggest new models for CDK7 function in transcription and implicate CAK dissociation from TFIIH as essential for kinase activation. This straightforward regulatory strategy ensures CDK7 activation is spatially and temporally linked to transcription, and may apply toward other transcription-associated kinases.

Exome sequencing in multiplex families with left-sided cardiac defects has high yield for disease gene discovery.

Congenital heart disease (CHD) is a common group of birth defects with a strong genetic contribution to their etiology, but historically the diagnostic yield from exome studies of isolated CHD has been low. Pleiotropy, variable expressivity, and the difficulty of accurately phenotyping newborns contribute to this problem. We hypothesized that performing exome sequencing on selected individuals in families with multiple members affected by left-sided CHD, then filtering variants by population frequency, in silico predictive algorithms, and phenotypic annotations from publicly available databases would increase this yield and generate a list of candidate disease-causing variants that would show a high validation rate. In eight of the nineteen families in our study (42%), we established a well-known gene/phenotype link for a candidate variant or performed confirmation of a candidate variant's effect on protein function, including variants in genes not previously described or firmly established as disease genes in the body of CHD literature: BMP10, CASZ1, ROCK1 and SMYD1. Two plausible variants in different genes were found to segregate in the same family in two instances suggesting oligogenic inheritance. These results highlight the need for functional validation and demonstrate that in the era of next-generation sequencing, multiplex families with isolated CHD can still bring high yield to the discovery of novel disease genes.

Cost-effectiveness of 4CMenB vaccination against gonorrhea: importance of dosing schedule, vaccine sentiment, targeting strategy, and duration of protection.

BACKGROUND: Observational evidence suggests the 4CMenB meningococcal vaccine may partially protect against gonorrhea, with one dose being two-thirds as protective as two. We examined the cost-effectiveness of vaccinating men-who-have-sex-with-men (MSM) in England, with one- or two-dose primary vaccination. METHODS: Integrated transmission-dynamic health-economic modeling explored the effects of targeting strategy, first- and second-dose uptake levels, and duration of vaccine protection, using observational estimates of vaccine protection. RESULTS: Vaccination with one or two primary doses is always cost-saving, irrespective of uptake, although vaccine sentiment is an important determinant of impact and cost-effectiveness. The most impactful and cost-effective targeting is offering "Vaccination-according-to-Risk" (VaR), to all patients with gonorrhea plus those reporting high numbers of sexual partners. If VaR is not feasible to implement then the more-restrictive strategy of "Vaccination-on-Diagnosis" (VoD) with gonorrhea is cost-effective, but much less impactful. Under conservative assumptions, VaR(2-dose) saves £7.62M(95%CrI:1.15-17.52) and gains 81.41(28.67-164.23) QALYs over 10 years; VoD(2-dose) saves £3.40M(0.48-7.71) and gains 41.26(17.52-78.25) QALYs versus no vaccination. Optimistic versus pessimistic vaccine-sentiment assumptions increase net benefits by ∼30%(VoD) or ∼60%(VaR). CONCLUSIONS: At UK costs, targeted 4CMenB vaccination of MSM gains QALYs and is cost-saving at any uptake level. Promoting uptake maximizes benefits and is an important role for behavioral science.

SNVstory: inferring genetic ancestry from genome sequencing data.

BACKGROUND: Genetic ancestry, inferred from genomic data, is a quantifiable biological parameter. While much of the human genome is identical across populations, it is estimated that as much as 0.4% of the genome can differ due to ancestry. This variation is primarily characterized by single nucleotide variants (SNVs), which are often unique to specific genetic populations. Knowledge of a patient's genetic ancestry can inform clinical decisions, from genetic testing and health screenings to medication dosages, based on ancestral disease predispositions. Nevertheless, the current reliance on self-reported ancestry can introduce subjectivity and exacerbate health disparities. While genomic sequencing data enables objective determination of a patient's genetic ancestry, existing approaches are limited to ancestry inference at the continental level. RESULTS: To address this challenge, and create an objective, measurable metric of genetic ancestry we present SNVstory, a method built upon three independent machine learning models for accurately inferring the sub-continental ancestry of individuals. We also introduce a novel method for simulating individual samples from aggregate allele frequencies from known populations. SNVstory includes a feature-importance scheme, unique among open-source ancestral tools, which allows the user to track the ancestral signal broadcast by a given gene or locus. We successfully evaluated SNVstory using a clinical exome sequencing dataset, comparing self-reported ethnicity and race to our inferred genetic ancestry, and demonstrate the capability of the algorithm to estimate ancestry from 36 different populations with high accuracy. CONCLUSIONS: SNVstory represents a significant advance in methods to assign genetic ancestry, opening the door to ancestry-informed care. SNVstory, an open-source model, is packaged as a Docker container for enhanced reliability and interoperability. It can be accessed from https://github.com/nch-igm/snvstory .

Full-length isoform concatenation sequencing to resolve cancer transcriptome complexity.

BACKGROUND: Cancers exhibit complex transcriptomes with aberrant splicing that induces isoform-level differential expression compared to non-diseased tissues. Transcriptomic profiling using short-read sequencing has utility in providing a cost-effective approach for evaluating isoform expression, although short-read assembly displays limitations in the accurate inference of full-length transcripts. Long-read RNA sequencing (Iso-Seq), using the Pacific Biosciences (PacBio) platform, can overcome such limitations by providing full-length isoform sequence resolution which requires no read assembly and represents native expressed transcripts. A constraint of the Iso-Seq protocol is due to fewer reads output per instrument run, which, as an example, can consequently affect the detection of lowly expressed transcripts. To address these deficiencies, we developed a concatenation workflow, PacBio Full-Length Isoform Concatemer Sequencing (PB_FLIC-Seq), designed to increase the number of unique, sequenced PacBio long-reads thereby improving overall detection of unique isoforms. In addition, we anticipate that the increase in read depth will help improve the detection of moderate to low-level expressed isoforms. RESULTS: In sequencing a commercial reference (Spike-In RNA Variants; SIRV) with known isoform complexity we demonstrated a 3.4-fold increase in read output per run and improved SIRV recall when using the PB_FLIC-Seq method compared to the same samples processed with the Iso-Seq protocol. We applied this protocol to a translational cancer case, also demonstrating the utility of the PB_FLIC-Seq method for identifying differential full-length isoform expression in a pediatric diffuse midline glioma compared to its adjacent non-malignant tissue. Our data analysis revealed increased expression of extracellular matrix (ECM) genes within the tumor sample, including an isoform of the Secreted Protein Acidic and Cysteine Rich (SPARC) gene that was expressed 11,676-fold higher than in the adjacent non-malignant tissue. Finally, by using the PB_FLIC-Seq method, we detected several cancer-specific novel isoforms. CONCLUSION: This work describes a concatenation-based methodology for increasing the number of sequenced full-length isoform reads on the PacBio platform, yielding improved discovery of expressed isoforms. We applied this workflow to profile the transcriptome of a pediatric diffuse midline glioma and adjacent non-malignant tissue. Our findings of cancer-specific novel isoform expression further highlight the importance of long-read sequencing for characterization of complex tumor transcriptomes.

Novel pathogenic GATA6 variant associated with congenital heart disease, diabetes mellitus and necrotizing enterocolitis.

BACKGROUND: Pathogenic GATA6 variants have been associated with congenital heart disease (CHD) and a spectrum of extracardiac abnormalities, including pancreatic agenesis, congenital diaphragmatic hernia, and developmental delay. However, the comprehensive genotype-phenotype correlation of pathogenic GATA6 variation in humans remains to be fully understood. METHODS: Exome sequencing was performed in a family where four members had CHD. In vitro functional analysis of the GATA6 variant was performed using immunofluorescence, western blot, and dual-luciferase reporter assay. RESULTS: A novel, heterozygous missense variant in GATA6 (c.1403 G > A; p.Cys468Tyr) segregated with affected members in a family with CHD, including three with persistent truncus arteriosus. In addition, one member had childhood onset diabetes mellitus (DM), and another had necrotizing enterocolitis (NEC) with intestinal perforation. The p.Cys468Tyr variant was located in the c-terminal zinc finger domain encoded by exon 4. The mutant protein demonstrated an abnormal nuclear localization pattern with protein aggregation and decreased transcriptional activity. CONCLUSIONS: We report a novel, familial GATA6 likely pathogenic variant associated with CHD, DM, and NEC with intestinal perforation. These findings expand the phenotypic spectrum of pathologic GATA6 variation to include intestinal abnormalities. IMPACT: Exome sequencing identified a novel heterozygous GATA6 variant (p.Cys468Tyr) that segregated in a family with CHD including persistent truncus arteriosus, atrial septal defects and bicuspid aortic valve. Additionally, affected members displayed extracardiac findings including childhood-onset diabetes mellitus, and uniquely, necrotizing enterocolitis with intestinal perforation in the first four days of life. In vitro functional assays demonstrated that GATA6 p.Cys468Tyr variant leads to cellular localization defects and decreased transactivation activity. This work supports the importance of GATA6 as a causative gene for CHD and expands the phenotypic spectrum of pathogenic GATA6 variation, highlighting neonatal intestinal perforation as a novel extracardiac phenotype.

The perceptual timescape: Perceptual history on the sub-second scale.

There is a high-capacity store of brief time span (∼1000 ms) which information enters from perceptual processing, often called iconic memory or sensory memory. It is proposed that a main function of this store is to hold recent perceptual information in a temporally segregated representation, named the perceptual timescape. The perceptual timescape is a continually active representation of change and continuity over time that endows the perceived present with a perceived history. This is accomplished primarily by two kinds of time marking information: time distance information, which marks all items of information in the perceptual timescape according to how far in the past they occurred, and ordinal temporal information, which organises items of information in terms of their temporal order. Added to that is information about connectivity of perceptual objects over time. These kinds of information connect individual items over a brief span of time so as to represent change, persistence, and continuity over time. It is argued that there is a one-way street of information flow from perceptual processing either to the perceived present or directly into the perceptual timescape, and thence to working memory. Consistent with that, the information structure of the perceptual timescape supports postdictive reinterpretations of recent perceptual information. Temporal integration on a time scale of hundreds of milliseconds takes place in perceptual processing and does not draw on information in the perceptual timescape, which is concerned with temporal segregation, not integration.

Impact of interventions to reduce nosocomial transmission of SARS-CoV-2 in English NHS Trusts: a computational modelling study.

BACKGROUND: Prior to September 2021, 55,000-90,000 hospital inpatients in England were identified as having a potentially nosocomial SARS-CoV-2 infection. This includes cases that were likely missed due to pauci- or asymptomatic infection. Further, high numbers of healthcare workers (HCWs) are thought to have been infected, and there is evidence that some of these cases may also have been nosocomially linked, with both HCW to HCW and patient to HCW transmission being reported. From the start of the SARS-CoV-2 pandemic interventions in hospitals such as testing patients on admission and universal mask wearing were introduced to stop spread within and between patient and HCW populations, the effectiveness of which are largely unknown. MATERIALS/METHODS: Using an individual-based model of within-hospital transmission, we estimated the contribution of individual interventions (together and in combination) to the effectiveness of the overall package of interventions implemented in English hospitals during the COVID-19 pandemic. A panel of experts in infection prevention and control informed intervention choice and helped ensure the model reflected implementation in practice. Model parameters and associated uncertainty were derived using national and local data, literature review and formal elicitation of expert opinion. We simulated scenarios to explore how many nosocomial infections might have been seen in patients and HCWs if interventions had not been implemented. We simulated the time period from March-2020 to July-2022 encompassing different strains and multiple doses of vaccination. RESULTS: Modelling results suggest that in a scenario without inpatient testing, infection prevention and control measures, and reductions in occupancy and visitors, the number of patients developing a nosocomial SARS-CoV-2 infection could have been twice as high over the course of the pandemic, and over 600,000 HCWs could have been infected in the first wave alone. Isolation of symptomatic HCWs and universal masking by HCWs were the most effective interventions for preventing infections in both patient and HCW populations. Model findings suggest that collectively the interventions introduced over the SARS-CoV-2 pandemic in England averted 400,000 (240,000 - 500,000) infections in inpatients and 410,000 (370,000 - 450,000) HCW infections. CONCLUSIONS: Interventions to reduce the spread of nosocomial infections have varying impact, but the package of interventions implemented in England significantly reduced nosocomial transmission to both patients and HCWs over the SARS-CoV-2 pandemic.

Cerebral organoids containing an AUTS2 missense variant model microcephaly.

Variants in the AUTS2 gene are associated with a broad spectrum of neurological conditions characterized by intellectual disability, microcephaly, and congenital brain malformations. Here, we use a human cerebral organoid model to investigate the pathophysiology of a heterozygous de novo missense AUTS2 variant identified in a patient with multiple neurological impairments including primary microcephaly and profound intellectual disability. Proband cerebral organoids exhibit reduced growth, deficits in neural progenitor cell (NPC) proliferation and disrupted NPC polarity within ventricular zone-like regions compared to control cerebral organoids. We used CRISPR-Cas9-mediated gene editing to correct this variant and demonstrate rescue of impaired organoid growth and NPC proliferative deficits. Single-cell RNA sequencing revealed a marked reduction of G1/S transition gene expression and alterations in WNT-ß-catenin signalling within proband NPCs, uncovering a novel role for AUTS2 in NPCs during human cortical development. Collectively, these results underscore the value of cerebral organoids to investigate molecular mechanisms underlying AUTS2 syndrome.

Infantile metastatic ependymoma with a novel molecular profile and favorable outcome to intensive chemotherapy without irradiation: Case-based review.

Ependymal tumors are the third most common brain tumor under 14 years old. Even though metastatic disease is a rare event, it affects mostly young children and carries an adverse prognosis. The factors associated with dissemination and the best treatment approach have not yet been established and there is limited published data on how to manage metastatic disease, especially in patients under 3 years of age. We provide a review of the literature on clinical characteristics and radiation-sparing treatments for metastatic ependymoma in children under 3 years of age treated. The majority (73%) of the identified cases were above 12 months old and had the PF as the primary site at diagnosis. Chemotherapy-based approaches, in different regimens, were used with radiation reserved for progression or relapse. The prognosis varied among the studies, with an average of 50%-58% overall survival. This study also describes the case of a 7-month-old boy with metastatic posterior fossa (PF) ependymoma, for whom we identified a novel SPECC1L-RAF1 gene fusion using a patient-centric comprehensive molecular profiling protocol. The patient was successfully treated with intensive induction chemotherapy followed by high-dose chemotherapy and autologous hematopoietic progenitor cell rescue (AuHSCR). Currently, the patient is in continuous remission 5 years after his diagnosis, without radiation therapy. The understanding of the available therapeutic approaches may assist physicians in their management of such patients. This report also opens the perspective of newly identified molecular alterations in metastatic ependymomas that might drive more chemo-sensitive tumors.

Bayesian reconstruction of household transmissions to infer the serial interval of COVID-19 by variants of concern: analysis from a prospective community cohort study (Virus Watch).

BACKGROUND: The serial interval is a key epidemiological measure that quantifies the time between an infector's and an infectee's onset of symptoms. This measure helps investigate epidemiological links between cases, and is an important parameter in transmission models used to estimate transmissibility and inform control strategies. The emergence of multiple variants of concern (VOC) during the SARS-CoV-2 pandemic has led to uncertainties about potential changes in the serial interval of COVID-19. We estimated the household serial interval of multiple VOC using data collected by the Virus Watch study. This online, prospective, community cohort study followed-up entire households in England and Wales since mid-June 2020. METHODS: This analysis included 5842 symptomatic individuals with confirmed SARS-CoV-2 infection among 2579 households from Sept 1, 2020, to Aug 10, 2022. SARS-CoV-2 variant designation was based upon national surveillance data of variant prevalence by date and geographical region. We used a Bayesian framework to infer who infected whom by exploring all transmission trees compatible with the observed dates of symptoms, given assumptions on the incubation period and generation time distributions using the R package outbreaker2. FINDINGS: We characterised the serial interval of COVID-19 by VOC. The mean serial interval was shortest for omicron BA5 (2·02 days; 95% credible interval [CrI] 1·26-2·84) and longest for alpha (3·37 days; 2·52-4·04). The mean serial interval before alpha (wild-type) was 2·29 days (95% CrI 1·39-2·94), 3·11 days (2·28-3·90) for delta, 2·72 days (2·01-3·47) for omicron BA1, and 2·67 days (1·90-3·46) for omicron BA2. We estimated that 17% (95% CrI 5-26) of serial interval values are negative across all variants. INTERPRETATION: Most methods estimating the reproduction number from incidence time series do not allow for a negative serial interval by construction. Further research is needed to extend these methods and assess biases introduced by not accounting for negative serial intervals. To our knowledge, this study is the first to use a Bayesian framework to estimate the serial interval of all major SARS-CoV-2 VOC from thousands of confirmed household cases. FUNDING: UK Medical Research Council and Wellcome Trust.

Anomalies in the review process and interpretation of the evidence in the NICE guideline for chronic fatigue syndrome and myalgic encephalomyelitis.

Chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME) is a disabling long-term condition of unknown cause. The National Institute for Health and Care Excellence (NICE) published a guideline in 2021 that highlighted the seriousness of the condition, but also recommended that graded exercise therapy (GET) should not be used and cognitive-behavioural therapy should only be used to manage symptoms and reduce distress, not to aid recovery. This U-turn in recommendations from the previous 2007 guideline is controversial.We suggest that the controversy stems from anomalies in both processing and interpretation of the evidence by the NICE committee. The committee: (1) created a new definition of CFS/ME, which 'downgraded' the certainty of trial evidence; (2) omitted data from standard trial end points used to assess efficacy; (3) discounted trial data when assessing treatment harm in favour of lower quality surveys and qualitative studies; (4) minimised the importance of fatigue as an outcome; (5) did not use accepted practices to synthesise trial evidence adequately using GRADE (Grading of Recommendations, Assessment, Development and Evaluations trial evidence); (6) interpreted GET as mandating fixed increments of change when trials defined it as collaborative, negotiated and symptom dependent; (7) deviated from NICE recommendations of rehabilitation for related conditions, such as chronic primary pain and (8) recommended an energy management approach in the absence of supportive research evidence.We conclude that the dissonance between this and the previous guideline was the result of deviating from usual scientific standards of the NICE process. The consequences of this are that patients may be denied helpful treatments and therefore risk persistent ill health and disability.

Does Rectoanal Intussusception Limit Improvements in Clinical Outcome and Quality of Life After Sacral Nerve Stimulation for Fecal Incontinence?

BACKGROUND: Sacral nerve stimulation is a treatment option for severe, medically refractory fecal incontinence, although its use in patients with anatomic abnormalities remains controversial. OBJECTIVE: This study aimed to determine whether patients with rectoanal intussusception achieve similar benefits from device implantation to patients without rectoanal intussusception. DESIGN: Retrospective review of a prospectively maintained database. Demographics and clinical data were collected for each patient, including preoperative pelvic floor testing. Defecographies were reanalyzed in a blinded manner. Preoperative rectoanal intussusception was determined on the basis of the Oxford system (grade III-IV vs not; grade V excluded). SETTINGS: Academic-affiliated pelvic health center. PATIENTS: All patients undergoing sacral nerve stimulation for fecal incontinence between July 2011 and July 2019. MAIN OUTCOME MEASURES: Cleveland Clinic Florida Incontinence/Wexner Scores, Fecal Incontinence Severity Indices, and Fecal Incontinence Quality of Life Indices at 1 year. RESULTS: One hundred sixty-nine patients underwent sacral nerve stimulation for fecal incontinence during the study period. The average age was 60.3 years and 91% were female. Forty-six patients (27.2%) had concomitant rectoanal intussusception (38 patients [22.5%] grade III and 8 patients [4.7%] grade IV). Before surgery, patients reported an average of 10.8 accidents per week and a Wexner score of 15.7, with no difference between patients with and without rectoanal intussusception ( p = 0.22 and 0.95). At 1 year after surgery, the average Wexner score was 9.5. There was no difference in postoperative Wexner scores (10.4 vs 9.2, p = 0.23) or improvement over time between patients with and without rectoanal intussusception (-6.7 vs -5.7, p = 0.40). Similarly, there was no difference in quality of life or frequency of incontinence to liquid or solid stool. LIMITATIONS: Single-institution, moderate sample size, incomplete survey response. CONCLUSIONS: Concomitant rectoanal intussusception does not appear to affect clinical outcomes or quality of life after sacral nerve stimulation for fecal incontinence. Appropriate patients with fecal incontinence and rectoanal intussusception can be considered for sacral nerve stimulation placement. See Video Abstract at http://links.lww.com/DCR/C192 . LA INTUSUSCEPCIN RECTOANAL LIMITA LAS MEJORAS EN EL RESULTADO CLNICO Y LA CALIDAD DE VIDA DESPUS DE LA NEUROESTIMULACION SACRA PARA LA INCONTINENCIA FECAL: ANTECEDENTES:La neuroestimulación sacra es una opción de tratamiento para la incontinencia fecal grave refractaria al tratamiento médico, aunque su uso en pacientes con anomalías anatómicas sigue siendo controvertido.OBJETIVO:Determinar si los pacientes con intususcepción rectoanal logran beneficios similares de la implantación del dispositivo a los pacientes sin intususcepción rectoanal.DISEÑO:Revisión retrospectiva de una base de datos mantenida prospectivamente. Se recopilaron datos demográficos y clínicos de cada paciente, incluidas las pruebas preoperatorias del piso pélvico. Las defecografías se volvieron a analizar de forma ciega. La intususcepción rectoanal preoperatoria se determinó según el sistema de Oxford (grado III-IV vs. no; grado V excluido).ESCENARIO:Centro académico de salud pélvica.PACIENTES:Todos los pacientes sometidos a neuroestimulación sacra por incontinencia fecal entre julio de 2011 y julio de 2019.PRINCIPALES MEDIDAS DE RESULTADO:Cleveland Clinic Florida Incontinence/Wexner Scores, Índices de gravedad de la incontinencia fecal, Índices de calidad de vida de la incontinencia fecal al año.RESULTADOS:169 pacientes se sometieron a neuroestimulación sacra por incontinencia fecal durante el período de estudio. La edad promedio fue de 60.3 años y el 91% eran mujeres. Cuarenta y seis pacientes (27.2%) tenían intususcepción rectoanal concomitante (38 [22.5%] grado III y 8 [4.7%] grado IV). Antes de la cirugía, los pacientes informaron un promedio de 10.8 accidentes por semana y una puntuación de Wexner de 15.7 sin diferencia entre pacientes con y sin intususcepción rectoanal (p = 0.22 y 0.95). Un año después de la cirugía, la puntuación promedio de Wexner fue de 9.5. No hubo diferencia en las puntuaciones de Wexner posoperatorias (10.4 frente a 9.2, p = 0.23) o mejoría con el tiempo entre los pacientes con y sin intususcepción rectoanal (-6.7 frente a -5.7, p = 0.40). De manera similar, no hubo diferencia en la calidad de vida o la frecuencia de incontinencia de heces líquidas o sólidas.LIMITACIONES:Institución única, tamaño de muestra moderado, respuesta de encuesta incompleta.CONCLUSIÓN:La intususcepción rectoanal concomitante no parece afectar los resultados clínicos o la calidad de vida después de la neuroestimulación sacra para la incontinencia fecal. Los pacientes apropiados con incontinencia fecal e intususcepción rectoanal pueden ser considerados para la neuroestimulación sacra. Consulte Video Resumen en http://links.lww.com/DCR/C192(Traducción-Dr. Jorge Silva Velazco ).

Novel frameshift variant in MYL2 reveals molecular differences between dominant and recessive forms of hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is characterized by thickening of the ventricular muscle without dilation and is often associated with dominant pathogenic variants in cardiac sarcomeric protein genes. Here, we report a family with two infants diagnosed with infantile-onset HCM and mitral valve dysplasia that led to death before one year of age. Using exome sequencing, we discovered that one of the affected children had a homozygous frameshift variant in Myosin light chain 2 (MYL2:NM_000432.3:c.431_432delCT: p.Pro144Argfs*57;MYL2-fs), which alters the last 20 amino acids of the protein and is predicted to impact the most C-terminal of the three EF-hand domains in MYL2. The parents are unaffected heterozygous carriers of the variant and the variant is absent in control cohorts from gnomAD. The absence of the phenotype in carriers and the infantile presentation of severe HCM is in contrast to HCM associated with dominant MYL2 variants. Immunohistochemical analysis of the ventricular muscle of the deceased patient with the MYL2-fs variant showed a marked reduction of MYL2 expression compared to an unaffected control. In vitro overexpression studies further indicate that the MYL2-fs variant is actively degraded. In contrast, an HCM-associated missense variant (MYL2:p.Gly162Arg) and three other MYL2 stop-gain variants (p.E22*, p.K62*, p.E97*) that result in loss of the EF domains are stably expressed but show impaired localization. The degradation of the MYL2-fs can be rescued by inhibiting the cell's proteasome function supporting a post-translational effect of the variant. In vivo rescue experiments with a Drosophila MYL2-homolog (Mlc2) knockdown model indicate that neither the MYL2-fs nor the MYL2:p.Gly162Arg variant supports normal cardiac function. The tools that we have generated provide a rapid screening platform for functional assessment of variants of unknown significance in MYL2. Our study supports an autosomal recessive model of inheritance for MYL2 loss-of-function variants in infantile HCM and highlights the variant-specific molecular differences found in MYL2-associated cardiomyopathy.

A Novel Patient Selection Tool Is Highly Efficacious at Identifying Candidates for Outpatient Surgery When Applied to a Nonselected Cohort of Patients in a Community Hospital.

BACKGROUND: There is a paucity of validated selection tools to assess which patients can safely and predictably undergo same-day or 23-hour discharge in a community hospital. The purpose of this study was to assess the ability of our patient selection too to identify patients who are candidates for outpatient total joint arthroplasty (TJA) in a community hospital. METHODS: A retrospective review of 223 consecutive (unselected) primary TJAs was performed. The patient selection tool was retrospectively applied to this cohort to determine eligibility for outpatient arthroplasty. Utilizing length of stay and discharge disposition, we identified the proportion of patients discharged home within 23 hours. RESULTS: We found that 179 (80.1%) patients met eligibility criteria for short-stay TJA. Of the 223 patients in this study, 215 (96.4%) patients were discharged home; 17 (7.9%) were on the day of surgery, and 190 (88.3%) within 23 hours. Of the 179 eligible patients for short-stay discharge, 155 (86.6%) patients were discharged home within 23 hours. Overall, the sensitivity of the patient selection tool was 79%, the specificity was 92%, the positive predictive value was 87% and the negative predictive value was 96%. CONCLUSION: In this study, we found that more than 80% of patients undergoing TJA in a community hospital are eligible for short-stay arthroplasty with this selection tool. We found that this selection tool is safe and effective at predicting short-stay discharge. Further studies are needed to better ascertain the direct effects of these specific demographic traits on their effects on short-stay protocols.

Long-read whole genome sequencing reveals HOXD13 alterations in synpolydactyly.

Synpolydactyly 1, also called syndactyly type II (SDTY2), is a genetic limb malformation characterized by polydactyly with syndactyly involving the webbing of the third and fourth fingers, and the fourth and fifth toes. It is caused by heterozygous alterations in HOXD13 with incomplete penetrance and phenotypic variability. In our study, a five-generation family with an SPD phenotype was enrolled in our Rare Disease Genomics Protocol. A comprehensive examination of three generations using Illumina short-read whole-genome sequencing (WGS) did not identify any causative variants. Subsequent WGS using Pacific Biosciences (PacBio) long-read HiFi Circular Consensus Sequencing (CCS) revealed a heterozygous 27-bp duplication in the polyalanine tract of HOXD13. Sanger sequencing of all available family members confirmed that the variant segregates with affected individuals. Reanalysis of an unrelated family with a similar SPD phenotype uncovered a 21-bp (7-alanine) duplication in the same region of HOXD13. Although ExpansionHunter identified these events in most individuals in a retrospective analysis, low sequence coverage due to high GC content in the HOXD13 polyalanine tract makes detection of these events challenging. Our findings highlight the value of long-read WGS in elucidating the molecular etiology of congenital limb malformation disorders.

Evidence-Based Care for People with Chronic Fatigue Syndrome and Myalgic Encephalomyelitis.

Chronic fatigue syndrome (CFS), sometimes referred to as myalgic encephalomyelitis (ME) and often as CFS/ME, is an illness characterized by disabling fatigue and other symptoms, typically worsened by activity. The main evidence-based treatments are rehabilitative in nature and include specific types of cognitive behavior therapy (CBT) and graded exercise therapy (GET). In this article, we briefly review the evidence for their safety and effectiveness and propose that much of the controversy about them arises from misunderstandings about their nature and delivery. In particular, we emphasize that successful rehabilitation from CFS/ME does not indicate that the illness is not real. We recommend that rehabilitative treatment always be preceded by a thorough clinical assessment and delivered by appropriately trained therapists working in close collaboration with the patient. We conclude that properly applied rehabilitative treatments offer the best hope of safely improving fatigue and function for patients with CFS/ME. However, we also recognize the need for more research into the treatment of this neglected condition, especially for those most severely disabled by it.

Detection of brain somatic variation in epilepsy-associated developmental lesions.

OBJECTIVE: Epilepsy-associated developmental lesions, including malformations of cortical development and low-grade developmental tumors, represent a major cause of drug-resistant seizures requiring surgical intervention in children. Brain-restricted somatic mosaicism has been implicated in the genetic etiology of these lesions; however, many contributory genes remain unidentified. METHODS: We enrolled 50 children who were undergoing epilepsy surgery into a translational research study. Resected tissue was divided for clinical neuropathologic evaluation and genomic analysis. We performed exome and RNA sequencing to identify somatic variation and we confirmed our findings using high-depth targeted DNA sequencing. RESULTS: We uncovered candidate disease-causing somatic variation affecting 28 patients (56%), as well as candidate germline variants affecting 4 patients (8%). In agreement with previous studies, we identified somatic variation affecting solute carrier family 35 member A2 (SLC35A2) and mechanistic target of rapamycin kinase (MTOR) pathway genes in patients with focal cortical dysplasia. Somatic gains of chromosome 1q were detected in 30% (3 of 10) of patients with Type I focal cortical dysplasia (FCD)s. Somatic variation in mitogen-activated protein kinase (MAPK) pathway genes (i.e., fibroblast growth factor receptor 1 [FGFR1], FGFR2, B-raf proto-oncogene, serine/threonine kinase [BRAF], and KRAS proto-oncogene, GTPase [KRAS]) was associated with low-grade epilepsy-associated developmental tumors. RNA sequencing enabled the detection of somatic structural variation that would have otherwise been missed, and which accounted for more than one-half of epilepsy-associated tumor diagnoses. Sampling across multiple anatomic regions revealed that somatic variant allele fractions vary widely within epileptogenic tissue. Finally, we identified putative disease-causing variants in genes not yet associated with focal cortical dysplasia. SIGNIFICANCE: These results further elucidate the genetic basis of structural brain abnormalities leading to focal epilepsy in children and point to new candidate disease genes.

Outcomes of Mechanical Thrombectomy Compared With Systemic Thrombolysis in Pulmonary Embolism: A Comprehensive Evaluation From the National Inpatient Sample Database.

BACKGROUND: Systemic thrombolysis (ST) may not be ideal for many patients with acute pulmonary embolism (PE) due to bleeding risk. In this analysis, we evaluated the safety and effectiveness of mechanical thrombectomy (MT) as an alternative to ST for acute PE. METHODS: Patients aged ≥18 years who underwent MT and/or ST for PE were identified from the National Inpatient Sample database from 2016 to 2017. Patients who underwent catheter-directed thrombolysis were excluded. We compared in-hospital outcomes of both groups in this retrospective study. RESULTS: Of 16 890 patients who received an intervention for acute PE, 1380 (8.2%) received MT and 15 510 (91.8%) received ST. There was no difference in age between both groups. In-hospital mortality was significantly lower in patients who received MT than that in those who received ST (11.9% vs 20.6%, odds ratio [OR]: 0.52, 95% confidence interval [CI]: 0.29-0.93, p=0.028). There was no statistically significant difference in terms of periprocedural bleeding, intracranial hemorrhage, and acute kidney injury between the 2 groups (p≥0.608 for all). Patients who received MT had a higher rate of respiratory complications (19.0% vs 11.6%, OR: 1.79, 95% CI: 1.06-3.03, p=0.030) and discharge to an outside facility (34.1% vs 19.2%, OR: 2.18, 95% CI: 1.41-3.37, p<0.001) than those who received ST. CONCLUSION: Mortality was significantly lower with MT than that with ST, but larger randomized studies are needed to validate this. The use of MT should be individualized on the basis of the patients' clinical presentation, risk profile, and local resources. CLINICAL IMPACT: In this study, we utilized the National Inpatient Sample database to study the in-hospital outcomes of pulmonary embolism patients who underwent mechanical thrombectomy compared to those who underwent systemic thrombolysis. We found that the patients who were diagnosed with pulmonary embolism and underwent mechanical thrombectomy had significantly lower mortality compared to those who were treated using systemic thrombolysis. This study was the first of its kind, utilizing the national inpatient sample database for evaluation of mechanical thrombectomy in comparison with the standard of care. These result would direct further randomized controlled trials for better evaluation of the utilization of mechanical thrombectomy in the correct clinical context. Furthermore, our study demonstrated comparable peri-operative complications between the mechanical thrombectomy group and the systemic thrombolysis group. These results would direct clinicians to consider mechanical thrombectomy if clinically indicated given the promising results.