Showing papers in "Molecular Diagnosis & Therapy in 2021"
TL;DR: An ensemble method, based on a deep autoencoder and random forest classifier, to predict potential circRNA–disease associations that outperforms previous methods with high degree of robustness and is compared with state-of-the-art classifiers and related methods.
Abstract: Circular RNAs (circRNA) are endogenous non-coding RNA molecules with a stable circular conformation. Growing evidence from recent experiments reveals that dysregulations and abnormal expressions of circRNAs are correlated with complex diseases. Therefore, identifying the causal circRNAs behind diseases is invaluable in explaining the disease pathogenesis. Since biological experiments are difficult, slow-progressing, and prohibitively expensive, computational approaches are necessary for identifying the relationships between circRNAs and diseases. We propose an ensemble method called AE-RF, based on a deep autoencoder and random forest classifier, to predict potential circRNA–disease associations. The method first integrates circRNA and disease similarities to construct features. The integrated features are sent to the deep autoencoder, to extract hidden biological patterns. With the extracted deep features, the random forest classifier is trained for association prediction. AE-RF achieved AUC scores of 0.9486 and 0.9522, in fivefold and tenfold cross-validation experiments, respectively. We conducted case studies on the top-most predicted results and three common human cancers. We compared the method with state-of-the-art classifiers and related methods. The experimental results and case studies demonstrate the prediction power of the model, and it outperforms previous methods with high degree of robustness. Training the classifier with the unique features retrieved by the autoencoder enhanced the model’s predictive performance. The top predicted circRNAs are promising candidates for further biological tests.
25 citations
TL;DR: Piflufolastat F 18 (PYLARIFY®) is an 18F-labeled diagnostic imaging agent that has been developed by Progenics Pharmaceuticals Inc., a Lantheus company, for positron emission tomography (PET) that targets prostatespecific membrane antigen (PSMA) as discussed by the authors.
Abstract: Piflufolastat F 18 (PYLARIFY®) is an 18F-labelled diagnostic imaging agent that has been developed by Progenics Pharmaceuticals Inc., a Lantheus company, for positron emission tomography (PET) that targets prostate-specific membrane antigen (PSMA). Piflufolastat F 18 was approved in the USA on 27 May 2021 for PET of PSMA positive lesions in men with prostate cancer with suspected metastasis who are candidates for initial definitive therapy or with suspected recurrence based on elevated serum prostate specific antigen (PSA) level. This article summarizes the milestones in the development of piflufolastat F 18 leading to this approval as a radioactive diagnostic agent in prostate cancer.
24 citations
TL;DR: In this paper, a meta-analysis of variable epidermal growth factor receptor (EGFR) gene mutations in non-small cell lung cancer (NSCLC) is conducted to provide a worldwide overview of EGFR mutation and submutation prevalence.
Abstract: Background Identification of variable epidermal growth factor receptor (EGFR) gene mutations in non-small cell lung cancer (NSCLC) is important for the selection of appropriate targeted therapies. This meta-analysis was conducted to provide a worldwide overview of EGFR mutation and submutation (specifically exon 19 deletions, exon 21 L858R substitutions, and others) prevalence, and identify important covariates that influence EGFR mutation status in patients with advanced NSCLC to address this clinical data gap. Methods Embase® and MEDLINE® in Ovid were searched for studies published between 2004 and 2019 with cohorts of ≥ 50 adults with EGFR mutations, focusing on stage III/IV NSCLC (≤ 20% of patients with stage I/II NSCLC). Linear mixed-effects models were fitted to EGFR mutation endpoints using logistic transformation (logit), assuming a binomial distribution. The model included terms for an intercept reflecting European studies and further additive terms for other continents. EGFR submutations examined were exon 19 deletions, exon 21 L858R substitutions, and others. Results Of 3969 abstracts screened, 57 studies were included in the overall EGFR mutation analysis and 74 were included in the submutation analysis relative to the overall EGFR mutation population (Europe, n = 12; Asia, n = 51; North America, n = 5; Central America, n = 1; South America, n = 1; Oceania, n = 1; Global, n = 3). The final overall EGFR mutations model estimated Asian and European prevalence of 49.1% and 12.8%, respectively, and included an additive covariate for the proportion of male patients in a study. There were no significant covariates in the submutation analyses. Most submutations were actionable: exon 19 deletions (49.2% [Asia]; 48.4% [Europe]); exon 21 L858R substitutions (41.1% [Asia]; 29.9% [Europe]). Conclusions Although EGFR mutation prevalence was higher in Asian than Western countries, data support worldwide testing for EGFR overall and submutations to inform appropriate targeted treatment decisions.
24 citations
TL;DR: A review of liquid biopsy methods for tumor genotyping can be found in this paper, where the authors provide a general overview of the major applications of liquid-biopsy in blood, including its utility for molecular response and minimal residual disease monitoring.
Abstract: In the era of personalized medicine and targeted therapies for the management of patients with cancer, ultrasensitive detection methods for tumor genotyping, such as next-generation sequencing or droplet digital polymerase chain reaction (ddPCR), play a significant role. In the search for less invasive strategies for diagnosis, prognosis and disease monitoring, the number of publications regarding liquid biopsy approaches using ddPCR has increased substantially in recent years. There is a long list of malignancies in which ddPCR provides a reliable and accurate tool for detection of nucleic acid-based markers derived from cell-free DNA, cell-free RNA, circulating tumor cells, extracellular vesicles or exosomes when isolated from whole blood, plasma and serum, helping to anticipate tumor relapse or unveil intratumor heterogeneity and clonal evolution in response to treatment. This updated review describes recent developments in ddPCR platforms and provides a general overview about the major applications of liquid biopsy in blood, including its utility for molecular response and minimal residual disease monitoring in hematological malignancies or the therapeutic management of patients with colorectal or lung cancer, particularly for the selection and monitoring of treatment with tyrosine kinase inhibitors. Although plasma is the main source of genetic material for tumor genomic profiling, liquid biopsy by ddPCR is being investigated in a wide variety of biologic fluids, such as cerebrospinal fluid, urine, stool, ocular fluids, sputum, saliva, bronchoalveolar lavage, pleural effusion, mucin, peritoneal fluid, fine needle aspirate, bile or pancreatic juice. The present review focuses on these "alternative" sources of genetic material and their analysis by ddPCR in different kinds of cancers.
24 citations
TL;DR: In this article, a review summarises the latest emerging therapies that may become mainstream treatment in the coming years and highlights the current and developing therapeutic landscape in novel pharmacological treatment, dietary supplementation, exercise training, device use, mitochondrial donation, tissue replacement gene therapy, hypoxic therapy and mitochondrial base editing.
Abstract: Primary mitochondrial disease (PMD) is a group of complex genetic disorders that arise due to pathogenic variants in nuclear or mitochondrial genomes. Although PMD is one of the most prevalent inborn errors of metabolism, it often exhibits marked phenotypic variation and can therefore be difficult to recognise. Current treatment for PMD revolves around supportive and preventive approaches, with few disease-specific therapies available. However, over the last decade there has been considerable progress in our understanding of both the genetics and pathophysiology of PMD. This has resulted in the development of a plethora of new pharmacological and non-pharmacological therapies at varying stages of development. Many of these therapies are currently undergoing clinical trials. This review summarises the latest emerging therapies that may become mainstream treatment in the coming years. It is distinct from other recent reviews in the field by comprehensively addressing both pharmacological non-pharmacological therapy from both a bench and a bedside perspective. We highlight the current and developing therapeutic landscape in novel pharmacological treatment, dietary supplementation, exercise training, device use, mitochondrial donation, tissue replacement gene therapy, hypoxic therapy and mitochondrial base editing.
23 citations
TL;DR: In this paper, the authors discuss the possible role played by the immune response in predicting this risk, along with the most common clinical and pathological factors and molecular tools that have been developed and are already in use.
Abstract: Hormone-receptor positive (HR+) breast cancer (BC) (including the luminal A and the luminal B subtypes) is the most common type of tumor in women diagnosed with early-stage BC (EBC). It represents a highly heterogeneous subgroup that is characterized by different risks of relapse. The aim of this review is to discuss the possible role played by the immune response in predicting this risk, along with the most common clinical and pathological factors and molecular tools that have been developed and are already in use. As opposed to what has previously been observed in the most aggressive human epidermal growth factor receptor 2 (HER2)-positive and triple-negative breast cancer (TNBC) subtypes, a high proportion of tumor-infiltrating lymphocytes (TILs)—reflecting a spontaneous and pre-existing immune response to the tumor—has been linked to a worse prognosis in HR+ EBC. This work provides some immune biological rationale explaining these findings and provides the basics to understand the principal clinical trials that are testing immunotherapy in HR+ (luminal) BC.
19 citations
TL;DR: In this paper, the origins and rationale for seeking to improve the mutation-based detection of circulating cell-free DNA (ccfDNA) by using ccfDNA size profiling are reviewed.
Abstract: Circulating cell-free DNA (ccfDNA) has emerged as a promising diagnostic tool in oncology. Identification of tumour-derived ccfDNA (i.e. circulating tumour DNA [ctDNA]) provides non-invasive access to a malignancy’s molecular landscape to diagnose, inform therapeutic strategies, and monitor treatment efficacy. Current applications of ccfDNA to detect somatic mutations, however, have been largely constrained to tumour-informed searches and identification of common mutations because of the interaction between ctDNA signal and next-generation sequencing (NGS) noise. Specifically, the low allele frequency of ctDNA associated with non-metastatic and early-stage lesions may be indistinguishable from artifacts that accrue during sample preparation and NGS. Thus, using ccfDNA to achieve non-invasive and personalized molecular profiling to optimize individual patient care is a highly sought goal that remains limited in clinical practice. There is growing evidence, however, that further advances in the field of ccfDNA diagnostics may be achieved by improving detection of somatic mutations through leveraging the inherently shorter fragment lengths of ctDNA compared to non-neoplastic ccfDNA. Here, the origins and rationale for seeking to improve the mutation-based detection of ctDNA by using ccfDNA size profiling are reviewed. Subsequently, in vitro and in silico methods to enrich for a target ccfDNA fragment length are detailed to identify current practices and provide perspective into the potential of using ccfDNA size profiling to impact clinical applications in oncology.
18 citations
TL;DR: In this article, the advantages and challenges of clinical and laboratory-initiated exome re-analysis are reviewed and a model for the future that could potentially maximize the clinical utility by integrating information from electronic medical records and knowledge databases into routine clinical workflows is proposed.
Abstract: Novel gene-disease discoveries, rapid advancements in technology, and improved bioinformatics tools all have the potential to yield additional molecular diagnoses through the reanalysis of exome sequencing data. Collaborations between clinical laboratories, ordering physicians, and researchers are also driving factors that can contribute to these new insights. Automation in ongoing natural history collection, evolving phenotype updates, advancements in processing next-generation sequencing data, and up-to-date variant-gene-disease databases are increasingly needed for systematic exome reanalysis. Here, we review some of the advantages and challenges for clinician-initiated and laboratory-initiated exome reanalysis, and we propose a model for the future that could potentially maximize the clinical utility of exome reanalysis by integrating information from electronic medical records and knowledge databases into routine clinical workflows.
18 citations
TL;DR: In this article, the authors evaluated the analytical performance of the newly developed rapid molecular assays GENECUBE® HQ SARS-CoV-2 (SARS CoV2) and GENE® FLU A/B (A/B) for detecting severe acute respiratory syndrome coronavirus 2.
Abstract: Molecular tests are the mainstay of detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, their accessibility can be limited by the long examination time and inability to evaluate multiple samples at once. This study evaluated the analytical performance of the newly developed rapid molecular assays GENECUBE® HQ SARS-CoV-2 and GENECUBE® FLU A/B. This prospective study was conducted between 14 December 2020 and 9 January 2021 at a polymerase chain reaction (PCR) center. Samples were collected from the nasopharynx with flocked swabs. Molecular tests were performed with the GENECUBE® system and reference reverse transcription (RT)-PCR, and the results of the two assays were compared. Among 1065 samples, 81 (7.6%) were positive for SARS-CoV-2 on the reference RT-PCR. Three showed discordance between GENECUBE® HQ SARS-CoV-2 and the reference RT-PCR; the total, positive, and negative samples of concordance for the two assays were 99.7%, 100%, and 99.7%, respectively. All discordant cases were positive with GENECUBE® HQ SARS-CoV-2 and negative with the reference RT-PCR. SARS-CoV-2 was detected in all three samples using another molecular assay for SARS-CoV-2. For GENECUBE® FLU A/B, the total, positive, and negative samples of concordance for the two assays were 99.5%, 100%, and 99.1%. The GENECUBE® HQ SARS-CoV-2 and GENECUBE® FLU A/B demonstrated sufficient analytical performance to detect SARS-CoV-2 and influenza virus A/B.
17 citations
TL;DR: A review of bi-and tri-specific NK/tumor engagers that are emerging as a new generation of immunotherapeutics can be found in this paper, where the authors explore the bi- and tri-Specific NK/Tumor Engagers (Bi-Teng et al. 2017).
Abstract: Natural killer (NK) cell-mediated cancer immunotherapy has grown significantly over the past two decades. More recently, multi-specific engagers have been developed as cancer therapeutics to effectively arm endogenous NK cells to more potently induce specific cytolytic responses against tumor targets. This review explores the bi- and tri-specific NK/tumor engagers that are emerging as a new generation of immunotherapeutics. These molecules vary in configuration, but they typically have small molecular weights and domains that engage specific tumor antigens and NK cell-activating receptors such as CD16, NKp30, NKp46, and NKG2D. They have demonstrated compelling potential in boosting NK cell cytotoxicity against specific tumor targets. This highly adaptable off-the-shelf platform, which in some formats also integrates cytokines, is poised to revolutionize targeted NK cell immunotherapy, either as a monotherapy or in combination with other effective anti-cancer therapies.
14 citations
TL;DR: In this article, the authors consider potential methods for preventing metastasis based on targeting molecular and cellular features of metastatic CTCs, including CTC clusters, and emphasize current knowledge gaps in CTC biology to develop highly effective therapeutics and strategies for metastasis suppression.
Abstract: Metastasis is the main cause of cancer death. Metastatic foci are derived from tumor cells that detach from the primary tumor and then enter the circulation. Circulating tumor cells (CTCs) are generally associated with a high probability of distant metastasis and a negative prognosis. Most CTCs die in the bloodstream, and only a few cells form metastases. Such metastatic CTCs have a stem-like and hybrid epithelial-mesenchymal phenotype, can avoid immune surveillance, and show increased therapy resistance. Targeting metastatic CTCs and their progenitors in primary tumors and their descendants, particularly disseminated tumor cells, represents an attractive strategy for metastasis prevention. However, current therapeutic strategies mainly target the primary tumor and only indirectly affect metastasis-initiating cells. Here, we consider potential methods for preventing metastasis based on targeting molecular and cellular features of metastatic CTCs, including CTC clusters. Also, we emphasize current knowledge gaps in CTC biology that should be addressed to develop highly effective therapeutics and strategies for metastasis suppression.
TL;DR: In this article, the authors discuss the pivotal role of IDH mutations in gliomagenesis and the complex interactions between the genomic and epigenetic landscapes, providing an overview of how, in the last decade, therapeutic approaches for low-grade gliomas have evolved.
Abstract: Diffuse low-grade gliomas account for approximately 20% of all primary brain tumors, they arise from glial cells and show infiltrative growth without histological features of malignancy. Mutations of the IDH1 and IDH2 genes constitute a reliable molecular signature of low-grade gliomas and are the earliest driver mutations occurring during gliomagenesis, representing a relevant biomarker with diagnostic, prognostic, and predictive value. IDH mutations induce a neomorphic enzyme that converts α-ketoglutarate to the oncometabolite D-2-hydroxyglutarate, which leads to widespread effects on cellular epigenetics and metabolism. Currently, there are no approved molecularly targeted therapies and the standard treatment for low-grade gliomas consists of radiation therapy and chemotherapy, with rising concern about treatment-related toxicities. Targeting D-2-hydroxyglutarate is considered a novel attractive therapeutic approach for low-grade gliomas and the insights from clinical trials suggest that mutant-selective IDH inhibitors are the ideal candidates, with a favorable benefit/risk ratio. A pivotal question is whether blocking IDH neomorphic activity may activate alternative oncogenetic pathways, inducing acquired resistance to IDH inhibitors. Based on this rationale, combination therapies to enhance the antitumor activity of IDH inhibitors and approaches aimed at exploiting, rather than inhibiting, the metabolism of IDH-mutant cancer cells, such as poly (adenosine 5'-diphosphate-ribose) polymerase inhibitors, are emerging from preclinical research and clinical trials. In this review, we discuss the pivotal role of IDH mutations in gliomagenesis and the complex interactions between the genomic and epigenetic landscapes, providing an overview of how, in the last decade, therapeutic approaches for low-grade gliomas have evolved.
TL;DR: In this paper, the authors propose to divide patients into subgroups of asthma pheno-/endotypes based on combined clinical and cellular biological characteristics and link them to targeted treatments.
Abstract: Asthma is a heterogeneous condition, but firm identification of heterogeneity-focused treatments is still lacking. Dividing patients into subgroups of asthma pheno-/endotypes based on combined clinical and cellular biological characteristics and linking them to targeted treatments could be a potentially useful approach to personalize therapy for better outcomes. Nonetheless, there are still many problems related to the identification and validation of asthma phenotypes and endotypes. Alternatively, a precision-medicine strategy for the management of patients with airways disease that is free from the traditional diagnostic labels and based on identifying "treatable traits" in each patient might be preferable. However, it would represent a quite unsophisticated approach because the definition of a treatable trait is too imprecise. In fact, there is still no understanding of the mechanisms underlying treatable traits that allow directing any targeted therapies against any particular treatable trait. Fortunately, in-depth identification of underlying molecular pathways to guide targeted treatment in individual patients is in progress thanks to the improvement in big data management obtained from ‘-omic’ sciences that is greatly increasing knowledge concerning asthma.
TL;DR: The relatively high percentage of significantly downregulated off-target genes for which one mismatch is present corroborates the requirement for intense bioinformatic screens and stringent specificity criteria to design antisense oligonucleotides with only minimal sequence complementarity to any non-target sequence.
Abstract: The field of antisense oligonucleotide therapeutics is rapidly growing and in addition to small molecules and therapeutic antibodies, oligonucleotide-based gene expression modifiers have been developed as fully accepted therapeutics. Antisense oligonucleotides are designed to modify gene expression of their specific target genes. However, as their effect relies on Watson–Crick base pairing, they could also bind to other unintended complementary RNAs showing sufficient sequence homology, which in turn could lead to off-target effects. It is assumed that these off-target effects depend on the degree of complementarity between the antisense oligonucleotides and off-target sequences. Aim of this study was the investigation of the effects of antisense oligonucleotides on the expression of potential off-targets having a defined number of mismatches to the oligonucleotide sequence. We extend recent studies by investigating the off-target profile of two 17-mer antisense oligonucleotides in two distinct human cell lines by a whole-transcriptome study using RNA sequencing. The relatively high percentage of significantly downregulated off-target genes for which one mismatch is present corroborates the requirement for intense bioinformatic screens and stringent specificity criteria to design antisense oligonucleotides with only minimal sequence complementarity to any non-target sequence. Avoiding suppression of off-target genes by a thorough bioinformatics screen should strongly reduce the risk for toxicities caused by antisense oligonucleotide-mediated off-target RNA suppression and finally result in safer antisense oligonucleotide-based therapeutics.
TL;DR: In this article, the authors discuss the development and clinical implementation of commercial liquid biopsy platforms for circulating tumor DNA minimal residual disease detection of solid tumors and highlight the technological features that enable highly sensitive detection of tumor-derived genomic alterations.
Abstract: The detection of circulating tumor DNA via liquid biopsy has become an important diagnostic test for patients with cancer. While certain commercial liquid biopsy platforms designed to detect circulating tumor DNA have been approved to guide clinical decisions in advanced solid tumors, the clinical utility of these assays for detecting minimal residual disease after curative-intent treatment of nonmetastatic disease is currently limited. Predicting disease response and relapse has considerable potential for increasing the effective implementation of neoadjuvant and adjuvant therapies. As a result, many companies are rapidly investing in the development of liquid biopsy platforms to detect circulating tumor DNA in the minimal residual disease setting. In this review, we discuss the development and clinical implementation of commercial liquid biopsy platforms for circulating tumor DNA minimal residual disease detection of solid tumors. Here, we aim to highlight the technological features that enable highly sensitive detection of tumor-derived genomic alterations, the factors that differentiate these commercial platforms, and the ongoing trials that seek to increase clinical implementation of liquid biopsies using circulating tumor DNA-based minimal residual disease detection.
TL;DR: In this article, the authors presented the vital diagnostic function that could be performed by circulating exosomes of neuronal origin in identifying the subtype of stroke, its severity, and the recovery stages.
Abstract: Stroke is one of the major causes of morbidity and mortality globally, with devastating effects. It is diagnosed mainly by clinical assessment and brain imaging; however, it is challenging to discriminate stroke from similar conditions with parallel presentations. While brain imaging provides detection of stroke infarcts, it does not provide useful information on the biology and prognosis of the underlying disease process. The complex pathophysiology of stroke infarcts is a barrier in developing sensitive diagnostic tools, which consequently has a detrimental effect on development of treatment regimens. Early diagnosis of stroke is vital for better management, but currently there is no diagnostic blood-based biomarker. The cargo of exosomes can give an insight into the physiological or pathophysiological status of the cell. Exosomes have gained great interest as a means of intercellular communication and recently have been explored as a potential biomarker tool. Circulating exosomes in the blood result from of a contribution from all tissues. The sub-population of exosomes released from brain cells circulating in body fluids are known as neuronal exosomes. This overview presents the vital diagnostic function that could be performed by circulating exosomes of neuronal origin in identifying the subtype of stroke, its severity, and the recovery stages. A number of potential biomarkers that are obtained from circulating exosomes have showed promising potential to function as stroke biomarkers; however, further work is needed to characterize the neuronal exosomes and its payload and to determine the pathways it uses in the complex pathophysiology of stroke. The identification is a subset of exosomal biomarkers that are specific to stroke will enhance the early detection and prognosis of the disease.
TL;DR: In this article, a review aimed to briefly describe genomic, transcriptomic, proteomic, and metabolomic profiles of human seminal plasma in an attempt to introduce potential candidate non-invasive biomarkers for sperm retrieval success in men with non-obstructive azoospermia.
Abstract: Non-obstructive azoospermia (NOA) is a severe form of male factor infertility resulting from the impairment of sperm production. Surgical sperm retrieval followed by intracytoplasmic sperm injection (ICSI) is the only alternative for NOA patients to have their own genetic children. Nevertheless, due to an approximately 50% chance of success, harvesting sperm from these patients remains challenging. Thus, discovering noninvasive biomarkers, which are able to reliably predict the probability of sperm acquisition, not only can eliminate the risk of surgery but also can lower the costs of NOA diagnosis and treatment. Seminal plasma is the non-cellular and liquid portion of the ejaculate that consists of the secretions originating from testes and male accessory glands. In past years, a wide range of biomolecules including DNAs, RNAs, proteins, and metabolic intermediates have been identified by omics techniques in human seminal plasma. The current review aimed to briefly describe genomic, transcriptomic, proteomic, and metabolomic profiles of human seminal plasma in an attempt to introduce potential candidate noninvasive biomarkers for sperm-retrieval success in men with NOA.
TL;DR: This systematic review supports the use of structural and functional biomarkers in early AMD and intermediate AMD, which are more reproducible and less invasive than the other classes of biomarkers described.
Abstract: The purpose of the review was to identify structural, functional, blood-based, and other types of biomarkers for early, intermediate, and late nonexudative stages of age-related macular degeneration (AMD) and summarize the relevant data for proof-of-concept clinical trials. AMD is a leading cause of blindness in the aging population, yet no treatments exist for its most common nonexudative form. There are limited data on the diagnosis and progression of nonexudative AMD compared to neovascular AMD. Our objective was to provide a comprehensive, systematic review of recently published biomarkers (molecular, structural, and functional) for early AMD, intermediate AMD, and geographic atrophy and to evaluate the relevance of these biomarkers for use in future clinical trials. A literature search of PubMed, ScienceDirect, EMBASE, and Web of Science from January 1, 1996 to November 30, 2020 and a patent search were conducted. Search terms included “early AMD,” “dry AMD,” “intermediate AMD,” “biomarkers for nonexudative AMD,” “fundus autofluorescence patterns,” “color fundus photography,” “dark adaptation,” and “microperimetry.” Articles were assessed for bias and quality with the Mixed-Methods Appraisal Tool. A total of 94 articles were included (61,842 individuals). Spectral-domain optical coherence tomography was superior at highlighting detailed structural changes in earlier stages of AMD. Fundus autofluorescence patterns were found to be most important in estimating progression of geographic atrophy. Delayed rod intercept time on dark adaptation was the most widely recommended surrogate functional endpoint for early AMD, while retinal sensitivity on microperimetry was most relevant for intermediate AMD. Combinational studies accounting for various patient characteristics and machine/deep-learning approaches were best suited for assessing individualized risk of AMD onset and progression. This systematic review supports the use of structural and functional biomarkers in early AMD and intermediate AMD, which are more reproducible and less invasive than the other classes of biomarkers described. The use of deep learning and combinational algorithms will gain increasing importance in future clinical trials of nonexudative AMD.
TL;DR: In this paper, the performance of droplet digital polymerase chain reaction (ddPCR) was evaluated to detect SARS-CoV-2 and compared it to the conventional RT-qPCR.
Abstract: Since the initial coronavirus disease outbreak in late 2019 (COVID-19), reverse-transcription real-time polymerase chain reaction (RT-qPCR) has become the gold standard test to detect severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2). However, a more sensitive and accurate diagnostic tool was required. Therefore, droplet digital polymerase chain reaction (ddPCR) was suggested as an alternative method. Here, we evaluated the performance of ddPCR to detect SARS-CoV-2 and compared it to the performance of RT-qPCR. The analytical performances, including limit of blank and limit of detection, were established using positive and negative SARS-CoV-2 reference materials. A total of 366 RNA extracts (173 positive and 193 negative by RT-qPCR) were collected from four institutions and tested with a Bio-Rad SARS-CoV-2 ddPCR kit that detects the SARS-CoV-2 genome using primers for N1 and N2. Limit of blank was set at 0, and the limits of detection of N1 and N2 were 1.99 copies/μL and 5.18 copies/μL, respectively. Linearity was evaluated using serial dilution samples, which demonstrated good results (R2: 0.999, linear range: 5.88–6825.25 copies/μL for N1 and R2: 0.999, 5.53–5855.47 copies/μL for N2). The results of ddPCR and RT-qPCR revealed substantial agreement (Cohen’s kappa: 0.639, p < 0.01). The 63 samples with positive ddPCR but negative RT-qPCR showed low copy numbers, and 55% of them had COVID-19-related symptoms. Droplet digital polymerase chain reaction demonstrated excellent sensitivity for SARS-Cov-2 detection and consistently agreed with the results from conventional RT-qPCR. Furthermore, ddPCR provided quantitative data that can be used to monitor changes in the viral load of patients with COVID-19.
TL;DR: In this paper, a detailed molecular investigation of a cohort of 4185 patients with referrals for inherited cardiac diseases was performed using a next-generation sequencing (NGS) workflow based on a panel of 105 genes involved in sudden cardiac death.
Abstract: Molecular diagnosis in inherited cardiac diseases is challenging because of the significant genetic and clinical heterogeneity. We present a detailed molecular investigation of a cohort of 4185 patients with referrals for inherited cardiac diseases. Patients suffering from cardiomyopathies (3235 probands), arrhythmia syndromes (760 probands), or unexplained sudden cardiac arrest (190 cases) were analyzed using a next-generation sequencing (NGS) workflow based on a panel of 105 genes involved in sudden cardiac death. (Likely) pathogenic variations were identified for approximately 30% of the cohort. Pathogenic copy number variations (CNVs) were detected in approximately 3.1% of patients for whom a (likely) pathogenic variation were identified. A (likely) pathogenic variation was also detected for 21.1% of patients who died from sudden cardiac death. Unexpected variants, including incidental findings, were present for 28 cases. Pathogenic variations were mainly observed in genes with definitive evidence of disease causation. Our study, which comprises over than 4000 probands, is one of most important cohorts reported in inherited cardiac diseases. The global mutation detection rate would be significantly increased by determining the putative pathogenicity of the large number of variants of uncertain significance. Identification of "unexpected" variants also showed the clinical utility of genetic testing in inherited cardiac diseases as they can redirect clinical management and medical resources toward a meaningful precision medicine. In cases with negative result, a WGS approach could be considered, but would probably have a limited impact on mutation detection rate as (likely) pathogenic variations were essentially clustered in genes with strong evidence of disease causation.
TL;DR: This review provides an illustrative overview of the current research trends using CRISPR/Cas technology and highlights the latest developments in CRISpr/Cas technique includingCRISPR imaging, discovery of novel CRISP systems, and applications in altering the genome, epigenome or RNA in different organisms.
Abstract: The emergence of the versatile gene-editing technology using programmable sequence-specific endonuclease system (CRISPR-Cas9) has instigated a major upheaval in biomedical research. In a brief span of time, CRISPR/Cas has been adopted by research labs around the globe because of its potential for significant progress and applicability in terms of efficiency, versatility and simplicity. It is a breakthrough technique for systematic genetic engineering, genome labelling, epigenetic and transcriptional modulation, and multiplexed gene editing, amongst others. This review provides an illustrative overview of the current research trends using CRISPR/Cas technology. We highlight the latest developments in CRISPR/Cas technique including CRISPR imaging, discovery of novel CRISPR systems, and applications in altering the genome, epigenome or RNA in different organisms. Finally, we address the potential challenges of this technique for its future use. Development of new CRISPR/Cas systems.
TL;DR: In this paper, the role of next-generation sequencing (NGS) in the work-up of infectious, neoplastic, neuroimmunological, and neurodegenerative CNS diseases was evaluated.
Abstract: Cerebrospinal fluid (CSF) is a clear and paucicellular fluid that circulates within the ventricular system and the subarachnoid space of the central nervous system (CNS), and diverse CNS disorders can impact its composition, volume, and flow. As conventional CSF testing suffers from suboptimal sensitivity, this review aimed to evaluate the role of next-generation sequencing (NGS) in the work-up of infectious, neoplastic, neuroimmunological, and neurodegenerative CNS diseases. Metagenomic NGS showed improved sensitivity-compared to traditional methods-to detect bacterial, viral, parasitic, and fungal infections, while the overall performance was maximized in some studies when all diagnostic modalities were used. In patients with primary CNS cancer, NGS findings in the CSF were largely concordant with the molecular signatures derived from tissue-based molecular analysis; of interest, additional mutations were identified in the CSF in some glioma studies, reflecting intratumoral heterogeneity. In patients with metastasis to the CNS, NGS facilitated diagnosis, prognosis, therapeutic management, and monitoring, exhibiting higher sensitivity than neuroimaging, cytology, and plasma-based molecular analysis. Although evidence is still rudimentary, NGS could enhance the diagnosis and pathogenetic understanding of multiple sclerosis in addition to Alzheimer and Parkinson disease. To conclude, NGS has shown potential to aid the research, facilitate the diagnostic approach, and improve the management outcomes of all the aforementioned CNS diseases. However, to establish its role in clinical practice, the clinical validity and utility of each NGS protocol should be determined. Lastly, as most evidence has been derived from small and retrospective studies, results from randomized control trials could be of significant value.
TL;DR: The role of salivary exosomes in mediating the reciprocal interplay between glucose metabolism and human papillomavirus (HPV)-driven oropharyngeal cancer (OPC) remains unelucidated.
Abstract: Increasing evidence supports the notion that human papillomavirus (HPV) DNA integration onto the human genome can influence and alter the molecular cargo in the exosomes derived from head and neck cancer cells. However, the molecular cargo of salivary exosomes derived from HPV-driven oropharyngeal cancer (HPV-driven OPC) remains unelucidated. Salivary exosomes morphology and molecular characterizations were examined using the nanoparticle tracking (NTA), western blot analysis, transmission electron microscopy (TEM) and mass spectrometry analysis. We report that HPV16 DNA was detected (80%) in isolated salivary exosomes of HPV-driven OPC patients. Importantly, we demonstrate elevated protein levels of six main glycolytic enzymes [i.e., aldolase (ALDOA), glyceraldehye-3-phosphate dehydrogenase (GAPDH), lactate dehydrogenase A/B (LDHA and LDHB), phosphoglycerate kinase 1 (PGK1) and pyruvate kinase M1/2 (PKM)] in isolated salivary exosomes of HPV-driven OPC patients, suggesting a novel mechanism underlying the potential role of salivary exosomes in mediating the reciprocal interplay between glucose metabolism and HPV-driven OPC. Our data demonstrate the potential diagnostic value of HPV16 DNA and glycolytic enzymes in salivary exosomes in discriminating healthy controls from HPV-driven OPC patients, thereby opening new avenues in the future for clinical translation studies.
TL;DR: This article summarizes the milestones in the development of elivaldogene autotemcel leading to this first approval for the treatment of early CALD in patients < 18 years of age, with an ABCD1 genetic mutation.
Abstract: Elivaldogene autotemcel (SKYSONA™, eli-cel; Lenti-D™ gene therapy) is a gene therapy that has been developed by bluebird bio for the treatment of cerebral adrenoleukodystrophy (CALD), a rare, X-linked genetic disease that mainly affects the nervous system and adrenal glands. In July 2021, elivaldogene autotemcel received its first approval (in the EU) for the treatment of for the treatment of early CALD in patients < 18 years of age, with an ABCD1 genetic mutation, and for whom a human leukocyte antigen (HLA)-matched sibling haematopoietic stem cell (HSC) donor is not available. This article summarizes the milestones in the development of elivaldogene autotemcel leading to this first approval.
TL;DR: Oncolytic viruses act as double-edged swords against the tumors through direct cytolysis of cancer cells and the induction of antitumor immunity in colorectal cancer.
Abstract: Colorectal cancer is the third most common neoplasm in the world and the third leading cause of cancer-related deaths in the USA. A safer and more effective therapeutic intervention against this malignant carcinoma is called for given the limitations and toxicities associated with the currently available treatment modalities. Immuno-oncolytic or oncolytic virotherapy, the use of viruses to selectively or preferentially kill cancer cells, has emerged as a potential anticancer treatment modality. Oncolytic viruses act as double-edged swords against the tumors through the direct cytolysis of cancer cells and the induction of antitumor immunity. A number of such viruses have been tested against colorectal cancer, in both preclinical and clinical settings, and many have produced promising results. Oncolytic virotherapy has also shown synergistic antitumor efficacy in combination with conventional treatment regimens. In this review, we describe the status of this therapeutic approach against colorectal cancer at both preclinical and clinical levels. Successes with and the challenges of using oncolytic viruses, both as monotherapy and in combination therapy, are also highlighted.
TL;DR: Wang et al. as discussed by the authors compared the performance of RT-PCR and NGS in the detection of echinoderm microtubule-associated protein-like 4 (EML4)-ALK fusion in Chinese patients with NSCLC.
Abstract: Precise detection of anaplastic lymphoma kinase (ALK) rearrangement guides the application of ALK-targeted tyrosine kinase inhibitors (ALK-TKIs) in patients with non-small-cell lung cancer (NSCLC). Next-generation sequencing (NGS) has been widely used in clinics, but DNA-based NGS used to detect fusion genes has delivered false-negative results. However, fusion genes can be successfully detected at the transcription level and with higher sensitivity using RNA-based reverse transcription polymerase chain reaction (RT-PCR). This study compared the performance of RT-PCR and NGS in the detection of echinoderm microtubule-associated protein-like 4 (EML4)-ALK fusion in Chinese patients with NSCLC. Formalin-fixed paraffin-embedded tissues from 153 patients who were pathologically diagnosed as having NSCLC were collected from November 2017 to October 2019. Both DNA/RNA-based NGS and RNA-based RT-PCR were used to detect EML4-ALK fusion. For samples with discordant ALK status results, fluorescence in situ hybridization (FISH) or Sanger sequencing was used to further confirm the ALK status. In total, 124 samples were successfully analyzed using both NGS and RT-PCR. For 118 samples, results were consistent between NGS and RT-PCR, with 25 reported as ALK fusion positive and 93 as ALK fusion negative, achieving a concordance rate of 95.16%. Among the six samples with disconcordant results, five were positive using RT-PCR but negative using NGS, and one was positive using NGS but negative using RT-PCR. Four of six cases with disconcordant results (three RT-PCR positive and one NGS positive) were successfully validated using either FISH or Sanger sequencing. Compared with NGS, RT-PCR appears to be a reliable method of detecting EML4-ALK fusion in patients with NSCLC.
TL;DR: The restoration of Immunological imbalance in SAA patients after omalizumab is surely intriguing and represents a glimpse of light in the comprehension of immunological effects of treatment.
Abstract: Severe allergic asthma (SAA) is based on type 2 (T2-high) immune responses to allergens promoting type 2 T helper (Th2) cell cytokine responses and production of IgE antibodies. Omalizumab was the first biological drug licensed for clinical use in the management of IgE-mediated SAA. Despite emerging evidence supporting the prominent role of follicular T cells (Tfh), Breg and Treg subsets, in the development and progression of SAA, no data are available on the impact of omalizumab therapy. Ten SAA patients monitored at the Respiratory Diseases Unit of Siena University Hospital and ten healthy sex- and age-matched controls were enrolled in the study. Clinical and functional parameters were collected at baseline (T0) and after 6 months of therapy (T6). Cellular population analysis was determined through multicolour flow cytometry. SAA patients showed higher percentages of Th17.1, Tfh and Tfh2 while CD24hiCD27hi Breg cell, Treg and Tfr percentages were significantly lower than in controls. Higher percentages of Tfh2 in patients with nasal polyps than in those without and in controls were observed. At T6, significant decreases in Tfh and Tfh2 compared with T0 were observed. A slightly significant increase in Teffs was reported at T6 compared to T0. ΔIgE levels in serum were correlated with ΔCD19+CD24+CD27+ Breg cell percentages (r = − 0.86, p = 0.0022). Our data explored the changes in Tfh cells, Tregs and Bregs in severe asthma. The restoration of immunological imbalance in SAA patients after omalizumab is certainly intriguing and represents a glimpse into the comprehension of immunological effects of treatment.
TL;DR: In this article, the utility of miR-371a-3p levels as a tool for diagnosing testicular germ cell tumors was evaluated using a systematic review and meta-analysis.
Abstract: This study sought to assess the utility of miR-371a-3p levels as a tool for diagnosing testicular germ cell tumors. For this systematic review and meta-analysis we reviewed available published studies assessing the accuracy of miR-371a-3p as a tool for diagnosing testicular germ cell tumors. STATA 16.0 was used to calculate pooled sensitivity, specificity, negative likelihood ratio (NLR), positive likelihood ratio (PLR), and area under the curve (AUC) values. In total, six studies incorporating 1835 individuals were included in this analysis. Pooled results suggested that miR-371a-3p was able to differentiate between testicular germ cell tumors and non-testicular germ cell tumors or healthy individuals with a sensitivity of 0.90 [95% confidence interval (CI) 0.88–0.92], a specificity of 0.93 (95% CI 0.87–0.96), a PLR of 12.2 (95% CI 6.90–18.24), an NLR of 0.11 (95% CI 0.09–0.14) a diagnostic odds ratio (DOR) of 121.56 (95% CI 64.84–227.89), and an AUC of 0.94. MiR-371a-3p represents a viable biomarker associated with testicular germ cell tumors.
TL;DR: Type-specific HPV-DNA detection by ddPCR is highly specific but moderately sensitive for the determination of HPV status and showed clinical relevance, mainly when associated with p16-IHC status.
Abstract: High-risk human papillomavirus infection impacts staging and prognosis of oropharyngeal squamous cell carcinomas (OPSCCs). Determination of HPV status in tumor tissue by p16-immunohistochemistry (p16-IHC) can be challenging; therefore, complementary methodologies could be useful in a clinical setting. To test for accuracy and clinical relevance of HPV-DNA detection in formalin-fixed and paraffin-embedded (FFPE) tumor samples by droplet digital PCR (ddPCR). Fifty OPSCCs were tested for p16-IHC status followed by HPV-16/18 DNA detection/quantification in FFPE-recovered DNA using ddPCR. Accuracy for HPV status determination and association with patient information were also evaluated. 32.0% (16/50) of the cases were p16-IHC positive (p16 +), 42.0% (21/50) had detectable levels of HPV-16 DNA, and none were positive for HPV-18 DNA. A higher median viral load of HPV-16 DNA was observed in p16 + cases (p < 0.0001). Concordance between p16-IHC and HPV-16 DNA ranged from 78.0 to 86.0% and accuracy rates were between 78.0 and 86.0%. P16-IHC and HPV-16 DNA detection was associated with gender, smoking status, and tumor subsite, while only HPV-16 DNA was associated with cT stage. The combination of HPV positivity by p16-IHC and ddPCR showed higher overall survival rates in comparison with p16 + /HPV-DNA- and p16 − /HPV-DNA- results. Type-specific HPV-DNA detection by ddPCR is highly specific but moderately sensitive for the determination of HPV status and showed clinical relevance, mainly when associated with p16-IHC status. Results highlight the importance of performing HPV-DNA testing in combination with p16-IHC for proper identification of HPV-associated OPSCC and to improve clinical management of OPSCC patients.
TL;DR: The purpose of this review is to summarize the recent literature on epigenetics in most non-tumor dermatological pathologies, focusing on psoriasis, and to better understand the role of interactions between the environment and the genome in the physiopathogenesis of these diseases.
Abstract: Epigenetics is the study of the mechanisms that regulate gene expression without modifying DNA sequences. Knowledge of and evidence about how epigenetics plays a causative role in the pathogenesis of many skin diseases is increasing. Since the epigenetic changes present in tumor diseases have been thoroughly reviewed, we believe that knowledge of the new epigenetic findings in non-tumor immune-mediated dermatological diseases should be of interest to the general dermatologist. Hence, the purpose of this review is to summarize the recent literature on epigenetics in most non-tumor dermatological pathologies, focusing on psoriasis. Hyper- and hypomethylation of DNA methyltransferases and methyl-DNA binding domain proteins are the most common and studied methylation mechanisms. The acetylation and methylation of histones H3 and H4 are the most frequent and well-characterized histone modifications and may be associated with disease severity parameters and serve as therapeutic response markers. Many specific microRNAs dysregulated in non-tumor dermatological disease have been reviewed. Deepening the study of how epigenetic mechanisms influence non-tumor immune-mediated dermatological diseases might help us better understand the role of interactions between the environment and the genome in the physiopathogenesis of these diseases.