Endometrial Liquid Biopsy Provides a miRNA Roadmap of the Secretory Phase of the Human Endometrium

2019 ◽  
Vol 105 (3) ◽  
pp. 877-889 ◽  
Author(s):  
Alessia Grasso ◽  
Roser Navarro ◽  
Nuria Balaguer ◽  
Inmaculada Moreno ◽  
Pilar Alama ◽  
...  
Keyword(s):  
Functional Analysis ◽  
Ribonucleic Acid ◽  
Liquid Biopsy ◽  
Bioinformatic Analysis ◽  
Endometrial Receptivity ◽  
Mirna Signature ◽  
Secretory Phase ◽  
Messenger Ribonucleic Acid ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

Abstract Context Endometrial liquid biopsy (ELB) is a minimally invasive alternative for research and diagnosis in endometrial biology. Objective We sought to establish an endometrial micro ribonucleic acid (miRNA) roadmap based on ELB during the secretory phase of the menstrual cycle in both natural and hormonal replacement therapy (HRT) cycles. Design Human ELB samples (n = 58) were obtained from healthy ovum donors undergoing a natural and an HRT cycle consecutively. miRNA profiles were identified using next-generation sequencing (NGS). For functional analysis, messenger ribonucleic acid targets were chosen among those reported in the endometrial receptivity analysis. Results The human endometrial secretory phase is characterized by a dynamic miRNA secretion pattern that varies from the prereceptive to the receptive stages. No differences in miRNA profiles were found among natural versus HRT cycles in the same women, reinforcing the similarities in functional and clinical outcomes in natural versus medicated cycles. Bioinformatic analysis revealed 62 validated interactions and 81 predicted interactions of miRNAs differentially expressed in the HRT cycle. Annotation of these genes linked them to 51 different pathways involved in endometrial receptivity. Conclusion This NGS-based study describes the miRNA signature in human ELB during the secretory phase of natural and HRT cycles. A consistent endometrial miRNA signature was observed in the acquisition of endometrial receptivity. Interestingly, no significant differences in miRNA expression were found in natural versus HRT cycles reinforcing the functional clinical similarities between both approaches.

scholarly journals Technical Evaluation of Commercial Mutation Analysis Platforms and Reference Materials for Liquid Biopsy Profiling

Cancers ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 1588 ◽  
Author(s):  
Sabrina Weber ◽  
Benjamin Spiegl ◽  
Samantha O. Perakis ◽  
Christine M. Ulz ◽  
Peter M. Abuja ◽  
...  
Keyword(s):  
Mutation Analysis ◽  
Reference Materials ◽  
Test Performance ◽  
Liquid Biopsy ◽  
Attractive Alternative ◽  
Routine Practice ◽  
Patients With Cancer ◽  
Technical Evaluation ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

Molecular profiling from liquid biopsy, in particular cell-free DNA (cfDNA), represents an attractive alternative to tissue biopsies for the detection of actionable targets and tumor monitoring. In addition to PCR-based assays, Next Generation Sequencing (NGS)-based cfDNA assays are now commercially available and are being increasingly adopted in clinical practice. However, the validity of these products as well as the clinical utility of cfDNA in the management of patients with cancer has yet to be proven. Within framework of the Innovative Medicines Initiative (IMI) program CANCER-ID we evaluated the use of commercially available reference materials designed for ctDNA testing and cfDNA derived from Diagnostic Leukaphereses (DLA) for inter- and intra-assay as well as intra- and inter-laboratory comparisons. In three experimental setups, a broad range of assays including ddPCR, MassARRAY and various NGS-based assays were tested. We demonstrate that both reference materials with predetermined VAFs and DLA samples are extremely useful for the performance assessment of mutation analysis platforms. Moreover, our data indicate a substantial variability of NGS assays with respect to sensitivity and specificity highlighting the importance of extensive validation of the test performance before offering these tests in clinical routine practice.

scholarly journals Molecular Profiling of Malignant Pleural Effusions with Next Generation Sequencing (NGS): Evidence that Supports Its Role in Cancer Management

2020 ◽  
Vol 10 (4) ◽  
pp. 206
Author(s):  
Georgia Ι. Grigoriadou ◽  
Stepan M. Esagian ◽  
Han Suk Ryu ◽  
Ilias P. Nikas
Keyword(s):  
Next Generation Sequencing ◽  
Cancer Patients ◽  
Advanced Cancer ◽  
Liquid Biopsy ◽  
Molecular Profiling ◽  
Pleural Effusions ◽  
Advanced Cancer Patients ◽  
Cancer Management ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

Malignant pleural effusions (MPEs) often develop in advanced cancer patients and confer significant morbidity and mortality. In this review, we evaluated whether molecular profiling of MPEs with next generation sequencing (NGS) could have a role in cancer management, focusing on lung cancer. We reviewed and compared the diagnostic performance of pleural fluid liquid biopsy with other types of samples. When applied in MPEs, NGS may have comparable performance with corresponding tissue biopsies, yield higher DNA amount, and detect more genetic aberrations than blood-derived liquid biopsies. NGS in MPEs may also be preferable to plasma liquid biopsy in advanced cancer patients with a MPE and a paucicellular or difficult to obtain tissue/fine-needle aspiration biopsy. Of interest, post-centrifuge supernatant NGS may exhibit superior results compared to cell pellet, cell block or other materials. NGS in MPEs can also guide clinicians in tailoring established therapies and identifying therapy resistance. Evidence is still premature regarding the role of NGS in MPEs from patients with cancers other than lung. We concluded that MPE processing could provide useful prognostic and theranostic information, besides its diagnostic role.

scholarly journals Next Generation Sequencing Identifies Five Novel Mutations in Lebanese Patients with Bardet–Biedl and Usher Syndromes

Genes ◽  
2019 ◽  
Vol 10 (12) ◽  
pp. 1047 ◽  
Author(s):  
Lama Jaffal ◽  
Wissam H Joumaa ◽  
Alexandre Assi ◽  
Charles Helou ◽  
George Cherfan ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Usher Syndrome ◽  
Bioinformatic Analysis ◽  
Next Generation ◽  
Novel Mutations ◽  
Pathogenic Variants ◽  
Whole Exome ◽  
Targeted Ngs ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

Aim: To identify disease-causing mutations in four Lebanese families: three families with Bardet–Biedl and one family with Usher syndrome (BBS and USH respectively), using next generation sequencing (NGS). Methods: We applied targeted NGS in two families and whole exome sequencing (WES) in two other families. Pathogenicity of candidate mutations was evaluated according to frequency, conservation, in silico prediction tools, segregation with disease, and compatibility with inheritance pattern. The presence of pathogenic variants was confirmed via Sanger sequencing followed by segregation analysis. Results: Most likely disease-causing mutations were identified in all included patients. In BBS patients, we found (M1): c.2258A > T, p. (Glu753Val) in BBS9, (M2): c.68T > C; p. (Leu23Pro) in ARL6, (M3): c.265_266delTT; p. (Leu89Valfs*11) and (M4): c.880T > G; p. (Tyr294Asp) in BBS12. A previously known variant (M5): c.551A > G; p. (Asp184Ser) was also detected in BBS5. In the USH patient, we found (M6): c.188A > C, p. (Tyr63Ser) in CLRN1. M2, M3, M4, and M6 were novel. All of the candidate mutations were shown to be likely disease-causing through our bioinformatic analysis. They also segregated with the corresponding phenotype in available family members. Conclusion: This study expanded the mutational spectrum and showed the genetic diversity of BBS and USH. It also spotlighted the efficiency of NGS techniques in revealing mutations underlying clinically and genetically heterogeneous disorders.

scholarly journals Monitoring of RAS mutant clones in plasma of patients with RAS mutant metastatic colorectal cancer

2022 ◽  
Author(s):  
A. Fernández Montes ◽  
E. Élez ◽  
A. Vivancos ◽  
N. Martínez ◽  
P. González ◽  
...  
Keyword(s):  
Liquid Biopsy ◽  
Line Treatment ◽  
First Line ◽  
Liquid Biopsies ◽  
Therapeutic Implications ◽  
Next Generation Sequencing Ngs ◽  
Solid Liquid ◽  
Mutant Genes ◽  
Generation Sequencing ◽  
First Line Treatment

Abstract Purpose Some patients with histologically confirmed primary mCRC and mutated RAS reported undetectable RAS mutant clones in plasma after receiving anti-VEGF treatment. The aim was to prospectively assess it with its potential therapeutic implications. Methods RAS mutant genes in solid biopsy (before first-line treatment: FOLFOX/CAPOX + bevacizumab) were compared in liquid biopsy (before second-line treatment: panitumumab + FOLFIRI), using Idylla™ system. Discordant results between solid/liquid biopsies were assessed by the next-generation sequencing (NGS) test (solid/liquid biopsies). Results Twenty-three patients were assessed (seven had RAS mutant discrepancies between solid/liquid biopsies). The NGS test confirmed that 3/23 (13%) patients had undetectable RAS mutant clones in liquid biopsy and 3/23 (13%) presented discrepancies in solid biopsy (Idylla™ system vs. NGS test). Conclusion Thirteen percentage of patients had undetectable RAS mutant clones in liquid biopsy after first-line treatment. However, some discrepancies between solid and liquid biopsies have been observed. These results suggest a need to improve accuracy of RAS analyses, especially in solid biopsies.

scholarly journals Generic Protocols for the Analytical Validation of Next-Generation Sequencing-Based ctDNA Assays: A Joint Consensus Recommendation of the BloodPAC’s Analytical Variables Working Group

2020 ◽  
Vol 66 (9) ◽  
pp. 1156-1166 ◽  
Author(s):  
James H Godsey ◽  
Angela Silvestro ◽  
J Carl Barrett ◽  
Kelli Bramlett ◽  
Darya Chudova ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Liquid Biopsy ◽  
Working Group ◽  
Circulating Tumor Dna ◽  
Next Generation ◽  
Analytical Validation ◽  
Consensus Recommendation ◽  
Next Generation Sequencing Ngs ◽  
Development And Validation ◽  
Generation Sequencing

Abstract Liquid biopsy, particularly the analysis of circulating tumor DNA (ctDNA), has demonstrated considerable promise for numerous clinical intended uses. Successful validation and commercialization of novel ctDNA tests have the potential to improve the outcomes of patients with cancer. The goal of the Blood Profiling Atlas Consortium (BloodPAC) is to accelerate the development and validation of liquid biopsy assays that will be introduced into the clinic. To accomplish this goal, the BloodPAC conducts research in the following areas: Data Collection and Analysis within the BloodPAC Data Commons; Preanalytical Variables; Analytical Variables; Patient Context Variables; and Reimbursement. In this document, the BloodPAC’s Analytical Variables Working Group (AV WG) attempts to define a set of generic analytical validation protocols tailored for ctDNA-based Next-Generation Sequencing (NGS) assays. Analytical validation of ctDNA assays poses several unique challenges that primarily arise from the fact that very few tumor-derived DNA molecules may be present in circulation relative to the amount of nontumor-derived cell-free DNA (cfDNA). These challenges include the exquisite level of sensitivity and specificity needed to detect ctDNA, the potential for false negatives in detecting these rare molecules, and the increased reliance on contrived samples to attain sufficient ctDNA for analytical validation. By addressing these unique challenges, the BloodPAC hopes to expedite sponsors’ presubmission discussions with the Food and Drug Administration (FDA) with the protocols presented herein. By sharing best practices with the broader community, this work may also save the time and capacity of FDA reviewers through increased efficiency.

scholarly journals The most common technologies and tools for functional genome analysis

2017 ◽  
Vol 24 (1) ◽  
pp. 1-11 ◽  
Author(s):  
Evelina Gasperskaja ◽  
Vaidutis Kučinskas
Keyword(s):  
Functional Analysis ◽  
Human Genome ◽  
Genome Analysis ◽  
Bioinformatic Analysis ◽  
Genome Wide ◽  
Polymerase Chain ◽  
Scientific Results ◽  
The Right ◽  
Generation Sequencing ◽  
Functional Genome

Since the  sequence of the  human genome is complete, the  main issue is how to understand the information written in the DNA sequence. Despite numerous genome-wide studies that have already been performed, the challenge to determine the function of genes, gene products, and also their interaction is still open. As changes in the human genome are highly likely to cause pathological conditions, functional analysis is vitally important for human health. For many years there have been a  variety of technologies and tools used in functional genome analysis. However, only in the  past decade there has been rapid revolutionizing progress and improvement in high-throughput methods, which are ranging from traditional real-time polymerase chain reaction to more complex systems, such as next-generation sequencing or mass spectrometry. Furthermore, not only laboratory investigation, but also accurate bioinformatic analysis is required for reliable scientific results. These methods give an opportunity for accurate and comprehensive functional analysis that involves various fields of studies: genomics, epigenomics, proteomics, and interactomics. This is essential for filling the  gaps in the  knowledge about dynamic biological processes at both cellular and organismal level. However, each method has both advantages and limitations that should be taken into account before choosing the right method for particular research in order to ensure successful study. For this reason, the present review paper aims to describe the most frequent and widely-used methods for the comprehensive functional analysis.

scholarly journals A DNA methylation-based liquid biopsy for triple-negative breast cancer

2021 ◽  
Vol 5 (1) ◽  
Author(s):  
Katrina Cristall ◽  
Francois-Clement Bidard ◽  
Jean-Yves Pierga ◽  
Michael J. Rauh ◽  
Tatiana Popova ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Triple Negative Breast Cancer ◽  
Liquid Biopsy ◽  
Blood Test ◽  
Triple Negative ◽  
Superior Performance ◽  
Serum Samples ◽  
Clonal Hematopoiesis ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

AbstractHere, we present a next-generation sequencing (NGS) methylation-based blood test called methylation DETEction of Circulating Tumour DNA (mDETECT) designed for the optimal detection and monitoring of metastatic triple-negative breast cancer (TNBC). Based on a highly multiplexed targeted sequencing approach, this assay incorporates features that offer superior performance and included 53 amplicons from 47 regions. Analysis of a previously characterised cohort of women with metastatic TNBC with limited quantities of plasma (<2 ml) produced an AUC of 0.92 for detection of a tumour with a sensitivity of 76% for a specificity of 100%. mDETECTTNBC was quantitative and showed superior performance to an NGS TP53 mutation-based test carried out on the same patients and to the conventional CA15-3 biomarker. mDETECT also functioned well in serum samples from metastatic TNBC patients where it produced an AUC of 0.97 for detection of a tumour with a sensitivity of 93% for a specificity of 100%. An assay for BRCA1 promoter methylation was also incorporated into the mDETECT assay and functioned well but its clinical significance is currently unclear. Clonal Hematopoiesis of Indeterminate Potential was investigated as a source of background in control subjects but was not seen to be significant, though a link to adiposity may be relevant. The mDETECTTNBC assay is a liquid biopsy able to quantitatively detect all TNBC cancers and has the potential to improve the management of patients with this disease.

Novel splice variants of the human kallikrein-related peptidases 11 (KLK11) and 12 (KLK12), unraveled by next-generation sequencing technology

2018 ◽  
Vol 399 (9) ◽  
pp. 1065-1071 ◽  
Author(s):  
Panagiotis G. Adamopoulos ◽  
Christos K. Kontos ◽  
Andreas Scorilas
Keyword(s):  
Next Generation Sequencing ◽  
Serine Proteases ◽  
Splice Variants ◽  
Bioinformatic Analysis ◽  
Next Generation ◽  
Next Generation Sequencing Technology ◽  
Normal Tissues ◽  
Rapid Amplification ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

AbstractTissue kallikrein, kallikrein-related peptidases (KLKs), and plasma kallikrein form the largest group of serine proteases in the human genome, sharing many structural and functional characteristics. In this study, we describe the molecular cloning of four novel splice variants of the humanKLK11andKLK12genes, discovered by combining 3′ rapid amplification of cDNA ends (3′ RACE), next-generation sequencing (NGS) technology, advanced bioinformatic analysis and Sanger sequencing. Expression analysis of these new transcripts in cell lines originating from 17 cancerous and two normal tissues revealed the expression pattern of each transcript. These novelKLK11andKLK12splice variants represent new potential cancer biomarkers.

scholarly journals NGS Screening for Identification of Novel Pexophagy-Related Mutation in Arabidopsis thaliana

Proceedings ◽  
2020 ◽  
Vol 76 (1) ◽  
pp. 7
Author(s):  
Katarzyna Sieńko ◽  
Kacper Żukowski ◽  
Kenji Yamada ◽  
Shino Goto-Yamada
Keyword(s):  
Arabidopsis Thaliana ◽  
Sequence Data ◽  
Bioinformatic Analysis ◽  
Mapping Method ◽  
Eukaryotic Cells ◽  
Causative Gene ◽  
Biochemical Pathways ◽  
Conventional Mapping ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

Peroxisomes are the type of organelles in eukaryotic cells that are involved in different biochemical pathways depending on the type of cell. We have isolated a number of peroxisome unusual positioning (peup) mutants, which display the accumulation of abnormal peroxisomes, and demonstrated that autophagy is involved in removing damaged organelles. These peup mutants also show defects of other autophagy-related processes, such as the recovery from dark-senescence, and also failed to induce vacuole-related vesicle formations during microautophagy under nutrient deprivations. The aim of this study was to identify the causative gene of the peup33 mutant using next-generation sequencing (NGS) as a tool. Identification of mutations with NGS will allow us to save time compared to the conventional mapping method. Here, we present the workflow of the experiment, the procedure of bioinformatic analysis and the software applied to the sequence data produced by NGS.

scholarly journals The Role of Liquid Biopsy in Hepatocellular Carcinoma Prognostication

Cancers ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 659 ◽  
Author(s):  
Ismail Labgaa ◽  
Augusto Villanueva ◽  
Olivier Dormond ◽  
Nicolas Demartines ◽  
Emmanuel Melloul
Keyword(s):  
Hepatocellular Carcinoma ◽  
Liquid Biopsy ◽  
Biological Fluids ◽  
Tissue Samples ◽  
Genomic Complexity ◽  
Minimally Invasive Access ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing ◽  
Related Mortality

Showing a steadily increasing cancer-related mortality, the epidemiological evolution of hepatocellular carcinoma (HCC) is concerning. Numerous strategies have attempted to prognosticate HCC but their performance is modest; this is partially due to the heterogeneous biology of this cancer. Current clinical guidelines endorse classifications and scores that use clinical variables, such as the Barcelona Clinic Liver Cancer (BCLC) classification. These algorithms are unlikely to fully recapitulate the genomic complexity of HCC. Integrating molecular readouts on a patient-basis, following a precision-medicine perspective, might be an option to refine prognostic systems. The limited access to HCC tissue samples is an important limitation to these approaches but it could be partially circumvented by using liquid biopsy. This concept consists of the molecular analysis of products derived from a solid tumor and released into biological fluids, mostly into the bloodstream. It offers an easy and minimally-invasive access to DNA, RNA, extracellular vesicles and cells that can be analyzed with next-generation sequencing (NGS) technologies. This review aims to investigate the potential contributions of liquid biopsy in HCC prognostication. The results identified prognostic values for each of the components of liquid biopsy, suggesting that this technology may help refine HCC prognostication.

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