Clinical whole-exome sequencing for the diagnosis of rare disorders with congenital anomalies and/or intellectual disability: substantial interest of prospective annual reanalysis

PurposeCongenital anomalies and intellectual disability (CA/ID) are a major diagnostic challenge in medical genetics—50% of patients still have no molecular diagnosis after a long and stressful diagnostic “odyssey.” Solo clinical whole-exome sequencing (WES) was applied in our genetics center to improve diagnosis in patients with CA/ID.MethodsThis retrospective study examined 416 consecutive tests performed over 3 years to demonstrate the effectiveness of periodically reanalyzing WES data. The raw data from each nonpositive test was reanalyzed at 12 months with the most recent pipeline and in the light of new data in the literature. The results of the reanalysis for patients enrolled in the third year are not yet available.ResultsOf the 416 patients included, data for 156 without a diagnosis were reanalyzed. We obtained 24 (15.4%) additional diagnoses: 12 through the usual diagnostic process (7 new publications, 4 initially misclassified, and 1 copy-number variant), and 12 through translational research by international data sharing. The final yield of positive results was 27.9% through a strict diagnostic approach, and 2.9% through an additional research strategy.ConclusionThis article highlights the effectiveness of periodically combining diagnostic reinterpretation of clinical WES data with translational research involving data sharing for candidate genes.     

Update on cytology samples for the use of molecular pathology and other ancillary tests in the move towards next-generation sequencing

The use of molecular pathology is becoming more evident every year, with 92% of pathologists stating that they use molecular pathology on a regular basis when reporting diagnostic work. This growth has been led not just through service development and improvement, but also with the introduction of new treatments offering patients significant improvement and overall better survival from cancer. These new treatments tend to require a targeted genomic test to identify specific cohorts of patients that will benefit. This has led to a reform of genomic and molecular pathology testing across the four nations of the United Kingdom, with the introduction of new regional testing hubs. These centres mainly use next generation sequencing platforms, to test for both the inherited and somatic mutations which drive disease. Next generation sequencing offers high-throughput cost effective sequencing of DNA, either via sequencing; individual genes, groups of genes, coding DNA known as Exome sequencing and whole genome sequencing to analysis every base pair in the patients DNA. This technology comes with new ways of working within the laboratory and relays heavily on the data analysis via bioinformatic analysis to; analyse, manipulate, quality control and interpret the vast about of data from these platforms. With the regionalization of testing to the genomic hubs, and with the increased movement of diagnostic cytopathology towards near patient sampling, this opens an opportunity for cytopathology departments to work in partnership with the new genetic testing hubs, to make sure that both parties can avail off the superior benefits offered by cytology samples; safe procurement, formalin free, high diagnostic value, rapid, minimally invasive and a high DNA yield. Cytopathology can also offer a rapid report based on sample suitability for molecular testing based on cellular content, but this to will change with the possible presence of cell-free DNA (cfDNA) which has been reported to be present in abundance in serous fluid supernatants. This supernatant that is normally discarded in the preparation of samples may hold vast diagnostic potential in the move to NGS testing.     

Cancer fatigue: a neurobiological review for psychiatrists

BACKGROUND: Fatigue, both during and subsequent to treatment, is a ubiquitous and debilitating phenomenon for patients undergoing treatment for cancer of all types. OBJECTIVE: This review will focus on understanding the complex neurobiological mechanisms underlying the phenomenon of cancer-related fatigue (CRF) and their relevance for various treatment routes. METHOD: The review will describe the prevalence of CRF, differential diagnosis, consideration of neurobiological mechanisms, and routes for intervention. DISCUSSION: The review will provide suggestions for future study and research, and a future article will describe behavioral and educational strategies psychiatrists may use in alleviating cancer fatigue.     

Translational extracts active biologically in vitro obtained from eukaryotic monolayer cells: a versatile method for viral RNA studies

Preservation of enzymatic activities in biological samples, especially after freeze/thawing, is a crucial requirement in virological research. Theoretically, this preservation can be achieved with the presence of cryopreservative agents. In contrast to tedious methods, it was found that this might be readily achieved by using well-defined conditions, including sucrose in the samples. Hence, the generation of a translational extract obtained from eukaryotic cells that have grown as monolayers is described below. This versatile method could be used advantageously for the in vitro translation of messenger RNAs, added exogenously, including viral mRNAs. The translational extract can be prepared freshly on a daily basis, or more conveniently it can be frozen and thawed subsequently for further use, without loss of activity. It can replace the Krebs ascites fluid and the commercial rabbit reticulocyte lysate. The procedure employed for the preservation of the biological activity of the translational extract can be extended to various other biological samples.     

Cargos and genes: insights into vesicular transport from inherited human disease

Many cellular functions depend on the correct delivery of proteins to specific intracellular destinations. Mutations that alter protein structure and disrupt trafficking of the protein (the "cargo") occur in many genetic disorders. In addition, an increasing number of disorders have been linked to mutations in the genes encoding components of the vesicular transport machinery responsible for normal protein trafficking. We review the clinical phenotypes and molecular pathology of such inherited "protein-trafficking disorders", which provide seminal insights into the molecular mechanisms of protein trafficking. Further characterisation of this expanding group of disorders will provide a basis for developing new diagnostic techniques and treatment strategies and offer insights into the molecular pathology of common multifactorial diseases that have been linked to disordered trafficking mechanisms.     

Development of Alzheimer-disease neuroimaging-biomarkers using mouse models with amyloid-precursor protein-transgene expression

There are important recent developments in Alzheimer's disease (AD) translational research, especially with respect to the imaging of amyloid pathology in vivo using MRI and PET technologies. Here we exploit the most widely used transgenic mouse models of amyloid pathology in order to relate the imaging findings to our knowledge about the histopathological phenotype of these models. The development of new diagnostic criteria of AD necessitates the use of biological markers to diagnose AD even in the absence of overt dementia or early symptomatic mild cognitive impairment. The validity of the diagnosis will depend on the availability of an in vivo marker to reflect underlying neurobiological changes of AD. Transgenic models with essential features of AD pathology and mechanisms provide a test setting for the development and evaluation of new biological imaging markers. Among the best established imaging markers of amyloid pathology in transgenic animals are high-field MRI of brain atrophy, proton spectroscopy of neurochemical changes, high-field MRI of amyloid plaque load, and in vivo plaque imaging using radio-labelled ligands with PET. We discuss the implications of the findings as well as the methodological limitations and the specific requirements of these technologies. We furthermore outline future directions of transgene-imaging research. Transgene imaging is an emerging area of translational research that implies strong multi- and interdisciplinary collaborations. It will become ever more valuable with the introduction of new diagnostic standards and novel treatment approaches which will require valid and reliable biological markers to improve the diagnosis and early treatment of AD patients.     

Molecular genetics and potential gene therapy

Contemporary molecular techniques including gene cloning, DNA sequencing, and gene transfer permit precise and comprehensive analysis of genetic disorders. For example, the molecular basis of hemoglobin synthesis disorders (the thalassemias) can now be ascribed to more than 30 different specific mutations. These affect virtually all aspects of gene expression. The more recent capacity to reintroduce cloned genes into mammalian cells in a functional form has raised the prospect of gene therapy, that is, the replacement of an abnormal gene with its normal counterpart or merely the introduction of a normal gene into a cell containing a defective copy. Genetic correction of enzyme-deficiency disorders whose effects are manifest in bone-marrow-derived cells seems most likely to be amenable to "somatic" (as opposed to germ line) gene therapy. Treatment of severe combined immunodeficiency due to adenosine deaminase (ADA) deficiency may be a suitable model for this approach. This report will review the molecular genetics of ADA, the methods by which ADA gene sequences may be transferred into various cells, and goals for current and future research.     

High-quality biobanking for personalized precision medicine: BioSpecimen Sciences at the helm

Optimal procurement and deep molecular (“omic”) characterization of human biospecimens are the key elements of success for developing truly personalized precision medicine of the future. We review key elements of developing and implementing a high-quality, cutting-edge BioSpecimen Sciences Program. Using our recently established academic programme as a specific example, we focus on quality of biospecimen practice covering topics including research consent of patients, logistics of ultra-rapid biobanking, computational and database aspects, quality control measures, proficiency testing, and optimal pathophysiome-preservation for data-rich omic interrogation of biospecimens. We emphasize that Pathology as the central medical speciality of personalized precision medicine will be the key driver for high-quality BioSpecimen Sciences in the modern healthcare setting.     

骨与软组织肿瘤诊断方法的研究

骨与软组织肿瘤是骨科常见疾病,对患者的生活质量、生命健康造成严重威胁。其诊断方法是临床和基础研究的重点,尤其是伴随着分子生物学技术的进步,对骨与软组织肿瘤的发生机制的认识不断深入,诊断方法也在逐步改进,但病理学检测仍是该病诊断的“金标准”。本文将对骨与软组织肿瘤的诊断方法研究进展作一综述。     

Mastocytosis: current treatment concepts

An explosion of research on mastocytosis in the last decade has witnessed a greater understanding of the molecular basis of this heterogeneous group of disorders and the conclusion that similar disease phenotypes may indeed be the result of different underlying genotypes. Along with our growing knowledge base of mastocytosis, newer approaches of treating these disorders are becoming available, all under investigational use at this time. This short review highlights the state of the art of current treatment strategies for the different categories of mastocytosis. The future will undoubtedly witness an even greater array of therapeutic options, as we continue to learn more about this enigmatic disease.     

Advancing the management of obstructive airways diseases through translational research

Obstructive airways diseases (OAD) represent a huge burden of illness world‐wide, and in spite of the development of effective therapies, significant morbidity and mortality related to asthma and COPD still remains. Over the past decade, our understanding of OAD has improved vastly, and novel treatments have evolved. This evolution is the result of successful translational research, which has connected clinical presentations of OAD and underlying disease mechanisms, thereby enabling the development of targeted treatments. The next challenge of translational research will be to position these novel treatments for OAD for optimal clinical use. At the same time, there is great potential in these treatments providing even better insights into disease mechanisms in OAD by studying the effects of blocking individual immunological pathways. To optimize this potential, there is a need to ensure that translational aspects are added to randomized clinical trials, as well as real‐world studies, but also to use other trial designs such as platform studies, which allow for simultaneous assessment of different interventions. Furthermore, demonstrating clinical impact, that is research translation, is an increasingly important component of successful translational research. This review outlines concepts of translational research, exemplifying how translational research has moved management of obstructive airways diseases into the next century, with the introduction of targeted, individualized therapy. Furthermore, the review describes how these therapies may be used as research tools to further our understanding of disease mechanisms in OAD, through translational, mechanistic studies. We underline the current need for implementing basic immunological concepts into clinical care in order to optimize the use of novel targeted treatments and to further the clinical understanding of disease mechanisms. Finally, potential barriers to adoption of novel targeted therapies into routine practice and how these may be overcome are described.     

Development of Alzheimer-disease neuroimaging-biomarkers using mouse models with amyloid-precursor protein-transgene expression

There are important recent developments in Alzheimer's disease (AD) translational research, especially with respect to the imaging of amyloid pathology in vivo using MRI and PET technologies. Here we exploit the most widely used transgenic mouse models of amyloid pathology in order to relate the imaging findings to our knowledge about the histopathological phenotype of these models. The development of new diagnostic criteria of AD necessitates the use of biological markers to diagnose AD even in the absence of overt dementia or early symptomatic mild cognitive impairment. The validity of the diagnosis will depend on the availability of an in vivo marker to reflect underlying neurobiological changes of AD. Transgenic models with essential features of AD pathology and mechanisms provide a test setting for the development and evaluation of new biological imaging markers.Among the best established imaging markers of amyloid pathology in transgenic animals are high-field MRI of brain atrophy, proton spectroscopy of neurochemical changes, high-field MRI of amyloid plaque load, and in vivo plaque imaging using radio-labelled ligands with PET. We discuss the implications of the findings as well as the methodological limitations and the specific requirements of these technologies. We furthermore outline future directions of transgene-imaging research. Transgene imaging is an emerging area of translational research that implies strong multi- and interdisciplinary collaborations. It will become ever more valuable with the introduction of new diagnostic standards and novel treatment approaches which will require valid and reliable biological markers to improve the diagnosis and early treatment of AD patients.     

Photosensitivity in lupus erythematosus

Lupus erythematosus (LE) is an autoimmune disease which can be triggered by environmental factors such as solar irradiation. It has long been observed that especially ultraviolet (UV) exposure can induce and exacerbate skin lesions in patients with this disease. However, despite the frequency of photosensitivity in LE, the mechanisms by which UV irradiation activates autoimmune responses is only now becoming increasingly unfolded by advanced molecular and cellular biological investigations. Phototesting, according to a standardized protocol with UVA and UVB irradiation has proven to be a valid model to study photosensitivity in various subtypes of LE and to evaluate the underlying pathomechanisms of this disease. Detailed analysis of the molecular events that govern lesion formation in experimentally photoprovoced LE showed increased accumulation of apoptotic keratinocytes and impaired expression of the inducible nitric oxide synthase (iNOS). In the near future, gene expression profiling and proteomics will further increase our knowledge on the complexity of the "UV response" in LE. This review summarizes the current understanding of the clinical and molecular mechanisms that initiate photosensitivity in this disease.     

The eye as a model of ageing in translational research – Molecular,epigenetic and clinical aspects

The eye and visual system are valuable in many areas of translational research such as stem cell therapy, transplantation research and gene therapy. Changes in many ocular tissues can be measured directly, easily and objectively in vivo (e.g. lens transparency; retinal blood vessel calibre; corneal endothelial cell counts) and so the eye may also be a uniquely useful site as a model of ageing. This review details cellular, molecular and epigenetic mechanisms related to ageing within the eye, and describes ocular parameters that can be directly measured clinically and which might be of value in ageing research as the translational “window to the rest of the body”. The eye is likely to provide a valuable model for validating biomarkers of ageing at molecular, epigenetic, cellular and clinical levels. A research agenda to definitively establish the relationship between biomarkers of ageing and ocular parameters is proposed.     

Role of electrical activity in promoting neural repair

The nervous system communicates in a language of electrical activities. The motivation to replace function lost through injury or disease through electrical prostheses has gained traction through steady advances in basic and translational science addressing the interface between electrical prostheses and the nervous system. Recent experiments suggest that electrical activity, signaling through specific molecular pathways, promotes neuronal survival and regeneration. Such data suggests that electrical prostheses, in addition to replacing lost function, may slow underlying degenerative disease or induce regenerative response. Here we review these data with a focus on retinal neurons, and discuss current efforts to translate this effect of electrical activity into clinically applicable treatments.     

Diagnostic and theranostic microRNAs in the pathogenesis of atherosclerosis

MicroRNAs (miRNAs) are a group of small single strand and noncoding RNAs that regulate several physiological and molecular signalling pathways. Alterations of miRNA expression profiles may be involved with pathophysiological processes underlying the development of atherosclerosis and cardiovascular diseases, including changes in the functions of the endothelial cells and vascular smooth muscle cells, such as cell proliferation, migration and inflammation, which are involved in angiogenesis, macrophage function and foam cell formation. Thus, miRNAs can be considered to have a crucial role in the progression, modulation and regulation of every stage of atherosclerosis. Such potential biomarkers will enable us to predict therapeutic response and prognosis of cardiovascular diseases and adopt effective preclinical and clinical treatment strategies. In the present review article, the current data regarding the role of miRNAs in atherosclerosis were summarized and the potential miRNAs as prognostic, diagnostic and theranostic biomarkers in preclinical and clinical studies were further discussed. The highlights of this review are expected to present opportunities for future research of clinical therapeutic approaches in vascular diseases resulting from atherosclerosis with an emphasis on miRNAs.     

Allergic hypersensitivity to topical and systemic corticosteroids: a review

 Corticosteroids, which are potent anti-inflammatory and immunomodulator agents used in the treatment of various inflammatory diseases including allergic diseases, can in some cases produce immediate or delayed hypersensitivity reactions. This review summarizes the epidemiological and clinical characteristics of such reactions, including related diagnostic issues. It also presents a detailed analysis of the proposed immunological mechanisms including underlying cross-reactions.