Cellular and molecular mechanisms breaking immune tolerance in inborn errors of immunity

In addition to susceptibility to infections, conventional primary immunodeficiency disorders (PIDs) and inborn errors of immunity (IEI) can cause immune dysregulation, manifesting as lymphoproliferative and/or autoimmune disease. Autoimmunity can be the prominent phenotype of PIDs and commonly includes cytopenias and rheumatological diseases, such as arthritis, systemic lupus erythematosus (SLE), and Sjogren’s syndrome (SjS). Recent advances in understanding the genetic basis of systemic autoimmune diseases and PIDs suggest an at least partially shared genetic background and therefore common pathogenic mechanisms. Here, we explore the interconnected pathogenic pathways of autoimmunity and primary immunodeficiency, highlighting the mechanisms breaking the different layers of immune tolerance to self-antigens in selected IEI.     

Diagnosing,discarding, or de-VUSsing: A practical guide to (un)targeted metabolomics as variant-transcending functional tests

PurposeFor patients with inherited metabolic disorders (IMDs), any diagnostic delay should be avoided because early initiation of personalized treatment could prevent irreversible health damage. To improve diagnostic interpretation of genetic data, gene function tests can be valuable assets. For IMDs, variant-transcending functional tests are readily available through (un)targeted metabolomics assays. To support the application of metabolomics for this purpose, we developed a gene-based guide to select functional tests to either confirm or exclude an IMD diagnosis.MethodsUsing information from a diagnostic IMD exome panel, Kyoto Encyclopedia of Genes and Genomes, and Inborn Errors of Metabolism Knowledgebase, we compiled a guide for metabolomics-based gene function tests. From our practical experience with this guide, we retrospectively selected illustrative cases for whom combined metabolomic/genomic testing improved diagnostic success and evaluated the effect hereof on clinical management.ResultsThe guide contains 2047 metabolism-associated genes for which a validated or putative variant-transcending gene function test is available. We present 16 patients for whom metabolomic testing either confirmed or ruled out the presence of a second pathogenic variant, validated or ruled out pathogenicity of variants of uncertain significance, or identified a diagnosis initially missed by genetic analysis.ConclusionMetabolomics-based gene function tests provide additional value in the diagnostic trajectory of patients with suspected IMD by enhancing and accelerating diagnostic success.     

Next-generation sequencing technologies: An overview

Since the days of Sanger sequencing, next-generation sequencing technologies have significantly evolved to provide increased data output, efficiencies, and applications. These next generations of technologies can be categorized based on read length. This review provides an overview of these technologies as two paradigms: short-read, or “second-generation,” technologies, and long-read, or “third-generation,” technologies. Herein, short-read sequencing approaches are represented by the most prevalent technologies, Illumina and Ion Torrent, and long-read sequencing approaches are represented by Pacific Biosciences and Oxford Nanopore technologies. All technologies are reviewed along with reported advantages and disadvantages. Until recently, short-read sequencing was thought to provide high accuracy limited by read-length, while long-read technologies afforded much longer read-lengths at the expense of accuracy. Emerging developments for third-generation technologies hold promise for the next wave of sequencing evolution, with the co-existence of longer read lengths and high accuracy.     

A single‐center pilot study in Malaysia on the clinical utility of whole‐exome sequencing for inborn errors of immunity

Primary immunodeficiency diseases refer to inborn errors of immunity (IEI) that affect the normal development and function of the immune system. The phenotypical and genetic heterogeneity of IEI have made their diagnosis challenging. Hence, whole‐exome sequencing (WES) was employed in this pilot study to identify the genetic etiology of 30 pediatric patients clinically diagnosed with IEI. The potential causative variants identified by WES were validated using Sanger sequencing. Genetic diagnosis was attained in 46.7% (14 of 30) of the patients and categorized into autoinflammatory disorders (n = 3), diseases of immune dysregulation (n = 3), defects in intrinsic and innate immunity (n = 3), predominantly antibody deficiencies (n = 2), combined immunodeficiencies with associated and syndromic features (n = 2) and immunodeficiencies affecting cellular and humoral immunity (n = 1). Of the 15 genetic variants identified, two were novel variants. Genetic findings differed from the provisional clinical diagnoses in seven cases (50.0%). This study showed that WES enhances the capacity to diagnose IEI, allowing more patients to receive appropriate therapy and disease management.     

新生儿遗传代谢病的筛查诊断及治疗进展

遗传代谢病在新生儿早期症状大多无特异性,由于累及的部位和病情轻重差异大所以极易造成误诊.孕妇在产前应对胎儿进行遗传代谢疾病的筛查诊断,降低新生儿遗传性代谢病出生缺陷率.新生儿科医生应综合评价筛查结果及时准确地做出临床诊断,以便 对遗传代谢疾病危象期患儿采取补救措施,使他们得到进一步评估及特殊治疗.该文就新生儿遗传代谢病主要的筛查诊断及治疗方法作一综述.     

Next-generation sequencing and clinical histocompatibility testing

Histocompatibility testing is essential for donor identification and risk assessment in solid organ and hematopoietic stem cell transplant. Additionally, it is useful for identifying donor specific alleles for monitoring donor specific antibodies in post-transplant patients. Next-generation sequence (NGS) based human leukocyte antigen (HLA) typing has improved many aspects of histocompatibility testing in hematopoietic stem cell and solid organ transplant. HLA disease association testing and research has also benefited from the advent of NGS technologies. In this review we discuss the current impact and future applications of NGS typing on clinical histocompatibility testing for transplant and non-transplant purposes.     

How to: Diagnose inborn errors of intrinsic and innate immunity to viral,bacterial, mycobacterial,and fungal infections

BackgroundInborn errors of intrinsic and innate immunity constitute the focus of a growing research field that investigates the molecular mechanisms underlying susceptibility to infections previously not considered part of the spectrum of inborn errors of immunity. These so-called nonconventional inborn errors of immunity often occur as infections caused by a narrow spectrum of microorganisms in otherwise healthy subjects.ObjectivesThis review aimed to provide a framework for identifying and evaluating patients with viral, bacterial, mycobacterial, and fungal infection needing further assessment for inborn errors of intrinsic and innate immunity.SourcesA literature search was performed using PubMed, from inception until 1 May 2022. The search included the following keywords: “inborn errors of immunity”; “inborn errors of innate immunity”; “primary immune deficiency”; “primary immunodeficiency”; “infections”; “infectious susceptibility”; “virus”; “pyogenic bacteria”; “mycobacteria”; “fungi”. All article types were considered.ContentWe review the definition of what can be considered an inborn error of immunity and how the definition changed over the last ～25 years. We further provide criteria to rule out secondary immunodeficiencies, identify patients needing further clinical and laboratory immunological assessment, and suspect and diagnose an inborn error of intrinsic and innate immunity. These steps are proposed as part of an algorithm.ImplicationsPatients with unexplained life-threatening infections, including otherwise healthy subjects, should be systematically screened for known inborn errors of immunity. The early diagnosis can prevent recurrence of life-threatening infections in the patients and reduce the total burden of infectious diseases.     

新生儿重症监护病房遗传代谢病分析

目的明确新生儿重症监护病房（NICU）遗传代谢病患者临床特点以及对其应采取的措施。方法留取从2008年11月1日到2009年10月31日,29例遗传代谢病高危新生儿的干血滤纸片,经含氨基酸、酰基肉碱内标的甲醇萃取,盐酸正丁醇衍生后,进行液相串联质谱分析。结果29例高危儿中检测出8例（27.6%）。甲基丙二酸血症患者3例,酪氨酸血症患者2例,枫糖尿病患者1例,甲基丙二酸血症合并酪氨酸血症患者2例。4例患者需要机械通气,辅助呼吸,6例患者出现严重代谢性酸中毒。住院周期的中位值为3天（0-7天）,4例（50%）患者死亡。结论遗传代谢病在某些NICUs还是比较常见的,而且入院前多没有明确诊断。入院患者多需要侵入性（如机械通气）辅助治疗,住院周期较短,病死率较高。这些病人在诊断和治疗方面给临床医生提出了挑战。     

Next-generation sequencing characterization of HLA in multi-generation families of Kuwaiti descent

The frequency of HLA genes in one population may not accurately represent frequencies in other populations. In this study, we characterized extended human leukocyte antigen (HLA) haplotypes in several families of Kuwaiti descent by high-resolution typing using next-generation technology. A total 81 members (including patients and related donors) from 21 families were enrolled. No haplotypes were shared among multiple families. Of 77 haplotypes identified, 23 were not listed in the HaploStats database. Two haplotypes were most common in African Americans, six in Asian Pacific Islanders, three in Caucasians, three in Hispanics, and three in Native Americans. The remaining identified haplotypes were not among the most common 200 HLA haplotypes in any of the five major populations. This cohort had 202 (19%) unique alleles, including 20 rare alleles, 16 very rare alleles, and 2 novel ones. Furthermore, no frequency data were available for 30% (23/77) of the observed haplotypes, and 6% (3/49) of B?～?C blocks identified were not available in the HaploStats database. Kuwaiti individuals carry unique HLA haplotypes that are not shared by the majority of individuals historically reported to the US National Marrow Donor Program registry.     

Inborn errors of immunity underlying a susceptibility to pyogenic infections: from innate immune system deficiency to complex phenotypes

BackgroundPyogenic bacteria are associated with a wide range of clinical manifestations, ranging from common and relatively mild respiratory and cutaneous infections to life-threatening localized or systemic infections, such as sepsis and profound abscesses. Despite vaccination and the widespread use of effective antibiotic treatment, severe infection is still observed in a subset of affected patients.ObjectivesWe aim to summarize the available data regarding inborn errors of immunity that result in a high risk of severe pyogenic infections.SourcesCase series, as well as review and original articles on human genetic susceptibility to pyogenic infections were examined.ContentWe review host-associated factors resulting in inborn errors of immunity and leading to a susceptibility to pyogenic infections, including deficiency in major components of the immune system (e.g., neutrophils, complement, immunoglobulin, and spleen function) and novel monogenic disorders resulting in specific susceptibility to pyogenic infection. Specifically, innate immune system deficiency involving toll-like receptors and associated signaling typically predispose to a narrow spectrum of bacterial diseases in otherwise healthy people, making a diagnosis more difficult to suspect and confirm. More complex syndromes, such as hyper IgE syndrome, are associated with a high risk of pyogenic infections due to an impairment of the interleukin-6 or -17 signaling, demonstrating the pivotal role of these pathways in controlling bacterial infections.ImplicationsIn clinical practice, awareness of such conditions is essential, especially in the pediatric setting, to avoid a potentially fatal diagnostic delay, set the most proper and prompt treatment, and ensure prevention of severe complications.     

Usability study of clinical exome analysis software: Top lessons learned and recommendations

ObjectivesNew DNA sequencing technologies have revolutionized the search for genetic disruptions. Targeted sequencing of all protein coding regions of the genome, called exome analysis, is actively used in research-oriented genetics clinics, with the transition to exomes as a standard procedure underway. This transition is challenging; identification of potentially causal mutation(s) amongst ∼10⁶ variants requires specialized computation in combination with expert assessment. This study analyzes the usability of user interfaces for clinical exome analysis software. There are two study objectives: (1) To ascertain the key features of successful user interfaces for clinical exome analysis software based on the perspective of expert clinical geneticists, (2) To assess user-system interactions in order to reveal strengths and weaknesses of existing software, inform future design, and accelerate the clinical uptake of exome analysis.MethodsSurveys, interviews, and cognitive task analysis were performed for the assessment of two next-generation exome sequence analysis software packages. The subjects included ten clinical geneticists who interacted with the software packages using the “think aloud” method. Subjects’ interactions with the software were recorded in their clinical office within an urban research and teaching hospital. All major user interface events (from the user interactions with the packages) were time-stamped and annotated with coding categories to identify usability issues in order to characterize desired features and deficiencies in the user experience.ResultsWe detected 193 usability issues, the majority of which concern interface layout and navigation, and the resolution of reports. Our study highlights gaps in specific software features typical within exome analysis. The clinicians perform best when the flow of the system is structured into well-defined yet customizable layers for incorporation within the clinical workflow. The results highlight opportunities to dramatically accelerate clinician analysis and interpretation of patient genomic data.ConclusionWe present the first application of usability methods to evaluate software interfaces in the context of exome analysis. Our results highlight how the study of user responses can lead to identification of usability issues and challenges and reveal software reengineering opportunities for improving clinical next-generation sequencing analysis. While the evaluation focused on two distinctive software tools, the results are general and should inform active and future software development for genome analysis software. As large-scale genome analysis becomes increasingly common in healthcare, it is critical that efficient and effective software interfaces are provided to accelerate clinical adoption of the technology. Implications for improved design of such applications are discussed.     

Mild inborn errors of metabolism in commonly used inbred mouse strains

Inbred mouse strains are a cornerstone of translational research but paradoxically many strains carry mild inborn errors of metabolism. For example, α-aminoadipic acidemia and branched-chain ketoacid dehydrogenase deficiency are known in C57BL/6J mice. Using RNA sequencing, we now reveal the causal variants in Dhtkd1 and Bckdhb, and the molecular mechanism underlying these metabolic defects. C57BL/6J mice have decreased Dhtkd1 mRNA expression due to a solitary long terminal repeat (LTR) in intron 4 of Dhtkd1. This LTR harbors an alternate splice donor site leading to a partial splicing defect and as a consequence decreased total and functional Dhtkd1 mRNA, decreased DHTKD1 protein and α-aminoadipic acidemia. Similarly, C57BL/6J mice have decreased Bckdhb mRNA expression due to an LTR retrotransposon in intron 1 of Bckdhb. This transposable element encodes an alternative exon 1 causing aberrant splicing, decreased total and functional Bckdhb mRNA and decreased BCKDHB protein. Using a targeted metabolomics screen, we also reveal elevated plasma C5-carnitine in 129 substrains. This biochemical phenotype resembles isovaleric acidemia and is caused by an exonic splice mutation in Ivd leading to partial skipping of exon 10 and IVD protein deficiency. In summary, this study identifies three causal variants underlying mild inborn errors of metabolism in commonly used inbred mouse strains.     

Cost-effectiveness frameworks for comparing genome and exome sequencing versus conventional diagnostic pathways: A scoping review and recommended methods

PurposeMethodological challenges have limited economic evaluations of genome sequencing (GS) and exome sequencing (ES). Our objective was to develop conceptual frameworks for model-based cost-effectiveness analyses (CEAs) of diagnostic GS/ES.MethodsWe conducted a scoping review of economic analyses to develop and iterate with experts a set of conceptual CEA frameworks for GS/ES for prenatal testing, early diagnosis in pediatrics, diagnosis of delayed-onset disorders in pediatrics, genetic testing in cancer, screening of newborns, and general population screening.ResultsReflecting on 57 studies meeting inclusion criteria, we recommend the following considerations for each clinical scenario. For prenatal testing, performing comparative analyses of costs of ES strategies and postpartum care, as well as genetic diagnoses and pregnancy outcomes. For early diagnosis in pediatrics, modeling quality-adjusted life years (QALYs) and costs over ≥20 years for rapid turnaround GS/ES. For hereditary cancer syndrome testing, modeling cumulative costs and QALYs for the individual tested and first/second/third-degree relatives. For tumor profiling, not restricting to treatment uptake or response and including QALYs and costs of downstream outcomes. For screening, modeling lifetime costs and QALYs and considering consequences of low penetrance and GS/ES reanalysis.ConclusionOur frameworks can guide the design of model-based CEAs and ultimately foster robust evidence for the economic value of GS/ES.     

Attitudes of the general population towards preconception expanded carrier screening for autosomal recessive disorders including inborn errors of metabolism

Background

A substantial number of severely debilitating and often ultimately fatal inborn errors of metabolism (IEMs) still lack an effective disease-modifying treatment. Informing couples before a pregnancy about an increased risk of having a child with an inherited disorder is now feasible by preconception expanded carrier screening (ECS). While knowledge about carrier status enhances reproductive autonomy, it may also result in ethical dilemmas. The purpose of this study was to assess the attitudes of the general Dutch population towards preconception ECS and to investigate which factors influence these attitudes.

新生儿遗传代谢病检测的进展

新生儿遗传代谢病筛查是世界上最大的筛查工程,近三十年来,随着筛查技术的发展,人们对新生儿遗传代谢病的认识、治疗及管理也在不断更新中.本文主要就新生儿遗传代谢病的历史、技术及最新的进展做一综述.     

Profiling of oxidative stress in patients with inborn errors of metabolism

Free radical formation resulting in oxidative stress is a hallmark of mitochondrial dysfunction. Indeed, oxidative stress has been demonstrated to be an underlying pathophysiologic process in various inborn errors of metabolism. Metabolic profiling of oxidative stress may provide a non-specific measure of disease activity that may further enable physicians to monitor disease. In the present study, we investigated two markers of oxidative damage in urinary samples from IEM subjects and controls: F-2 isoprostanes, a measure of lipid peroxidation and di-tyrosine, a measure of protein oxidation. We also determined urinary antioxidant activity in these samples. Subsets of IEM patients showed significantly higher levels of the damage markers isoprostanes and di-tyrosine. Of note, patients with cobalamin disorders (i.e., CblB and CblC) consistently had the highest levels of oxidative damage markers. Lower urine antioxidant capacity was seen in all subject categories, particularly cobalamin disorders and propionic acidemia. Longitudinal studies in subjects with MSUD showed good concordance between markers of oxidative damage and acute decompensation. Overall, quantifying oxidative stress offers a unique perspective to IEM. These measures may provide a means of addressing mitochondrial function in IEM and aid in the development of therapeutic targets and clinical monitoring in this diverse set of disorders.