scholarly journals The Use of Microfluidic Technology for Cancer Applications and Liquid Biopsy

Micromachines ◽  
2018 ◽  
Vol 9 (8) ◽  
pp. 397 ◽  
Author(s):  
Arutha Kulasinghe ◽  
Hanjie Wu ◽  
Chamindie Punyadeera ◽  
Majid Warkiani
Keyword(s):  
Tumor Markers ◽  
Liquid Biopsy ◽  
Point Of Care ◽  
High Sensitivity ◽  
Circulating Tumor Dna ◽  
Diagnostic Tools ◽  
Tumor Biomarkers ◽  
Blood Draw ◽  
Tumor Biopsy ◽  
Microfluidic Technology

There is growing awareness for the need of early diagnostic tools to aid in point-of-care testing in cancer. Tumor biopsy remains the conventional means in which to sample a tumor and often presents with challenges and associated risks. Therefore, alternative sources of tumor biomarkers is needed. Liquid biopsy has gained attention due to its non-invasive sampling of tumor tissue and ability to serially assess disease via a simple blood draw over the course of treatment. Among the leading technologies developing liquid biopsy solutions, microfluidics has recently come to the fore. Microfluidic platforms offer cellular separation and analysis platforms that allow for high throughout, high sensitivity and specificity, low sample volumes and reagent costs and precise liquid controlling capabilities. These characteristics make microfluidic technology a promising tool in separating and analyzing circulating tumor biomarkers for diagnosis, prognosis and monitoring. In this review, the characteristics of three kinds of circulating tumor markers will be described in the context of cancer, circulating tumor cells (CTCs), exosomes, and circulating tumor DNA (ctDNA). The review will focus on how the introduction of microfluidic technologies has improved the separation and analysis of these circulating tumor markers.

scholarly journals Electroanalytical Biosensors for Circulating Tumor DNA Detection—A Brief Review

2021 ◽  
Author(s):  
Zhijia Peng ◽  
Xiaogang Lin ◽  
Weiqi Nian ◽  
Xiaodong Zheng ◽  
Jayne Wu
Keyword(s):  
Real Time ◽  
Point Of Care ◽  
Low Cost ◽  
High Sensitivity ◽  
High Specificity ◽  
Circulating Tumor Dna ◽  
Minimal Invasive ◽  
Diagnosis And Treatment ◽  
Detection Technology ◽  
Tumor Dna

Early diagnosis and treatment have always been highly desired in the fight against cancer, and detection of circulating tumor DNA (ctDNA) has recently been touted as highly promising for early cancer screening. Consequently, the detection of ctDNA in liquid biopsy gains much attention in the field of tumor diagnosis and treatment, which has also attracted research interest from the industry. However, traditional gene detection technology is difficult to achieve low cost, real-time and portable measurement of ctDNA. Electroanalytical biosensors have many unique advantages such as high sensitivity, high specificity, low cost and good portability. Therefore, this review aims to discuss the latest development of biosensors for minimal-invasive, rapid, and real-time ctDNA detection. Various ctDNA sensors are reviewed with respect to their choices of receptor probes, detection strategies and figures of merit. Aiming at the portable, real-time and non-destructive characteristics of biosensors, we analyze their development in the Internet of Things, point-of-care testing, big data and big health.

scholarly journals Direct-RT-qPCR Detection of SARS-CoV-2 without RNA Extraction as Part of a COVID-19 Testing Strategy: From Sample to Result in One Hour

Diagnostics ◽  
2020 ◽  
Vol 10 (8) ◽  
pp. 605 ◽  
Author(s):  
Eva Kriegova ◽  
Regina Fillerova ◽  
Petr Kvapil
Keyword(s):  
High Throughput Screening ◽  
High Speed ◽  
Limit Of Detection ◽  
Point Of Care ◽  
Rna Extraction ◽  
Rna Isolation ◽  
High Sensitivity ◽  
Ease Of Use ◽  
Diagnostic Tools ◽  
Heat Inactivation

Due to the lack of protective immunity in the general population and the absence of effective antivirals and vaccines, the Coronavirus disease 2019 (COVID-19) pandemic continues in some countries, with local epicentres emerging in others. Due to the great demand for effective COVID-19 testing programmes to control the spread of the disease, we have suggested such a testing programme that includes a rapid RT-qPCR approach without RNA extraction. The Direct-One-Step-RT-qPCR (DIOS-RT-qPCR) assay detects severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in less than one hour while maintaining the high sensitivity and specificity required of diagnostic tools. This optimised protocol allows for the direct use of swab transfer media (14 μL) without the need for RNA extraction, achieving comparable sensitivity to the standard method that requires the time-consuming and costly step of RNA isolation. The limit of detection for DIOS-RT-qPCR was lower than seven copies/reaction, which translates to 550 virus copies/mL of swab. The speed, ease of use and low price of this assay make it suitable for high-throughput screening programmes. The use of fast enzymes allows RT-qPCR to be performed under standard laboratory conditions within one hour, making it a potential point-of-care solution on high-speed cycling instruments. This protocol also implements the heat inactivation of SARS-CoV-2 (75 °C for 10 min), which renders samples non-infectious, enabling testing in BSL-2 facilities. Moreover, we discuss the critical steps involved in developing tests for the rapid detection of COVID-19. Implementing rapid, easy, cost-effective methods can help control the worldwide spread of the COVID-19 infection.

Non-invasive detection of acquired resistance to FGFR inhibition in patients with cholangiocarcinoma harboring FGFR2 alterations.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. 4096-4096 ◽  
Author(s):  
Anna M. Varghese ◽  
Juber Ahamad A Patel ◽  
Yelena Yuriy Janjigian ◽  
Fanli Meng ◽  
S Duygu Selcuklu ◽  
...  
Keyword(s):  
Targeted Therapy ◽  
Acquired Resistance ◽  
High Sensitivity ◽  
Circulating Tumor Dna ◽  
Resistance Mechanisms ◽  
Tumor Evolution ◽  
Background Error ◽  
Tumor Biopsy ◽  
Invasive Approach ◽  
Non Invasive

4096 Background: FGFR2 alterations are present in 14% of cholangiocarcinomas (CCA) and are promising targets of investigational FGFR-directed therapies. Cell-free DNA profiling has emerged as a non-invasive approach to monitor disease and longitudinally characterize tumor evolution. We describe the use of circulating tumor DNA (ctDNA) among patients (pts) with FGFR2-altered CCA receiving FGFR-targeted therapy in the identification of acquired FGFR2 mutations (mut) at resistance. Methods: Serial blood samples were collected from 8 pts with FGFR-altered CCA for ctDNA isolation and next generation sequencing. Plasma ctDNA collected at baseline and resistance to FGFR-targeted therapy were sequenced using a custom ultra-deep coverage cfDNA panel, MSK-ACCESS, incorporating dual index primers and unique molecular barcodes to enable background error suppression and high-sensitivity mut detection. The assay was enhanced to include all protein-coding exons and relevant introns of FGFR2. In 5/8 pts, genomic profiling of an initial tumor biopsy was performed. Results: 8 pts with FGFR2-altered CCA (7 gene fusions, 1 amplification) were treated with FGFR-targeted therapies. 7/8 pts exhibited stable disease or partial response. 19 total acquired mut in FGFR2 were detected at resistance in 5/8 pts (between 1-9 unique mut identified in each sample). All mut were located in the kinase domain. Conclusions: Acquired mut in FGFR2 are seen in pts who have developed resistance to targeted therapy. CtDNA can be used to identify these mut at the time of acquired resistance. The multitude of FGFR2 mut observed within individual pts suggest heterogeneity and evolutionary convergence of resistance mechanisms. Our results illustrate the utility of ctDNA as a less invasive way to monitor for signs of resistance and to identify other potential targetable alterations. [Table: see text]

The results of a study of the concordance of RAS status in liquid and invasive biopsies in patients with metastatic colorectal cancer in the Republic of Kazakhstan.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e16017-e16017
Author(s):  
Kaldigul Smagulova ◽  
Dilyara Kaidarova ◽  
Yelena Ukolova ◽  
Madina Orazgalieva ◽  
Anel Kurmankulova
Keyword(s):  
Colorectal Cancer ◽  
Liquid Biopsy ◽  
Point Mutations ◽  
Diagnostic System ◽  
Circulating Tumor Dna ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Wild Type ◽  
Kras Gene ◽  
Tumor Biopsy

e16017 Background: Liquid biopsy is increasingly of interest as an alternative to invasive biopsy of solid tumors for predicting, making decisions about the treatment and monitoring of the disease. Particular preference is given to liquid biopsy in cases where it is not possible to obtain a sufficient amount of material or material of poor quality with a tumor biopsy. However, in order to find out the clinical significance of circulating tumor DNA (ctDNA), it is important to first establish the sensitivity of the method using tumor-plasma consistency studies. Methods: We selected 38 patients with a confirmed diagnosis of colorectal cancer (CRC), in whom was established the progression of the disease. All patients underwent two diagnostic methods, which we divided conditionally into 2 groups: A – invasive biopsy from available metastatic foci; to detect mutations in exons 2, 3, and 4 the KRAS gene used a reagent kit to detect 18 point mutations in codons 12,13,61,117,146 and a reagent kit to identify 10 mutations of the NRAS gene in codons 12,13,61,146 (Entrogen) on a Rotor-Gene 6000 Amplifier; B - liquid biopsy, to determine 21 mutations in the codons 12,13,59,61,117,146 of the KRAS gene, 18 mutations in the codons of 12,13,59,61,117,146 of the NRAS gene, was used Idylla automated molecular diagnostic system. Results: Of the 38 studied samples of group A: 22 (57.9%) had wild type KRAS, 16 (42.1%) were mutated, in group B: wild type had 25 (65.8%) patients, mutated - 13 (34, 2%). Thus, we see a discrepancy in the results in 3 (7.9%) of 38 cases. Conclusions: The results indicate a high degree of sensitivity (92.1%) of liquid biopsy as a diagnostic method, but confirmation of concordance with traditional tissue biopsy requires further in-depth study of this issue in a larger sample of patients.

scholarly journals Multiplex Enrichment and Detection of Rare KRAS Mutations in Liquid Biopsy Samples using Digital Droplet Pre-Amplification

2018 ◽  
Author(s):  
Erica D. Pratt ◽  
Robert W. Cowan ◽  
Sara L. Manning ◽  
Edmund Qiao ◽  
Heather Cameron ◽  
...  
Keyword(s):  
Single Molecule ◽  
Liquid Biopsy ◽  
High Sensitivity ◽  
Circulating Tumor Dna ◽  
Tumor Burden ◽  
Allelic Frequency ◽  
Cancer Surveillance ◽  
Ductal Adenocarcinoma ◽  
Kras Mutations ◽  
Oncology Research

AbstractOncology research is increasingly incorporating molecular detection of circulating tumor DNA (ctDNA) as a tool for cancer surveillance and early detection. However, non-invasive monitoring of conditions with low tumor burden remains challenging, as the diagnostic sensitivity of most ctDNA assays is inversely correlated with total DNA concentration and ctDNA abundance. Here we present the Multiplex Enrichment using Droplet Pre-Amplification (MED-Amp) method, which com-bines single-molecule emulsification and short-round PCR preamplification with digital droplet PCR (ddPCR) detection of mutant DNA template. The MED-Amp assay increased mutant signal by over 50-fold with minimal distortion in allelic frequency. We demonstrate detection of as few as 3 mutant copies in wild-type DNA concentrations ranging from 5 to 50ng. The MED-Amp assay successfully detected KRAS mutant ctDNA in 86% plasma samples obtained from patients with metastatic pancreatic ductal adenocarcinoma. This assay for high-sensitivity rare variant detection is appropriate for liquid biopsy samples, or other limited clinical biospecimens

Liquid Biopsy and Lung Cancer

2018 ◽  
Vol 63 (6) ◽  
pp. 489-496 ◽  
Author(s):  
Pasquale Pisapia ◽  
Umberto Malapelle ◽  
Giancarlo Troncone
Keyword(s):  
Lung Cancer ◽  
Liquid Biopsy ◽  
False Negative ◽  
High Sensitivity ◽  
Circulating Tumor Dna ◽  
Sample Collection ◽  
Negative Results ◽  
Major Disadvantage ◽  
Analytical Phase ◽  
Nsclc Patients

The identification of non-small cell lung cancer (NSCLC) patients potentially responsive to targeted therapies relies on a number of relevant biomarkers, including EGFR, ALK, ROS-1, and PD-L1. Biomarker identification is most commonly based on surgical sample collection. However, when tissues are difficult to reach or when multiple analyses are necessary to monitor tumor progression and treatment response, liquid biopsy is a valid noninvasive alternative. This analysis, which is preferentially performed on circulating tumor DNA (ctDNA) extracted from plasma samples, has the major advantage of reducing the inherent risks and discomfort of tissue biopsy. However, a major disadvantage is that it yields only a low number of ctDNA targets. Thus, to avoid false-positive and false-negative results, it is important to adopt and validate technologies with high sensitivity and specificity in the pre-analytical phase of sampling. This review succinctly addresses the principal methodologies for analyzing plasma-derived ctDNA in NSCLC patients.

scholarly journals ctDNA Detection in Microfluidic Platform: A Promising Biomarker for Personalized Cancer Chemotherapy

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Samla Gauri ◽  
Mohd R. Ahmad
Keyword(s):  
Cancer Chemotherapy ◽  
Liquid Biopsy ◽  
Treatment Resistance ◽  
Circulating Tumor Cell ◽  
Circulating Tumor Dna ◽  
Microfluidic System ◽  
Tumor Biopsy ◽  
Ctdna Analysis ◽  
Personalized Cancer

Early detection and characterization of circulating tumor DNA (ctDNA) can reveal mint of comprehensive biological insights from indicating the presence of tumor, identifying mutational changes of malignant cells, and allowing precision or targeted therapy together with monitoring disease progression, treatment resistance, and relapse of the disease. Apart from these, one of the greatest axiomatic implications of ctDNA detection is that it provides a new shed of light as noninvasive liquid biopsy as a replaceable procedure of surgical tumor biopsy. Despite the tremendous potential of ctDNA in cancer research, there remains a paucity of quantitative study on ctDNA detection and analysis. The majority of previously published microfluidic-based studies have focused on circulating tumor cell (CTC) detection and have failed to address the potential of ctDNA. The studies on microfluidic ctDNA detection are not consistent might be due to the complexity of ctDNA isolation as they present in low concentration in blood plasma. Researchers need to leverage the ability of microfluidic system for ctDNA analysis so that the significant enigma about cancer can be resolved effectively. This study, therefore, highlights the importance of ctDNA as cancer biomarker for liquid biopsy and provides an overview of the current laboratory as well as microfluidic techniques for ctDNA detection. This paper also attempts to show the emergence of new strands of microfluidic ctDNA detection and analysis for personalized cancer chemotherapy.

Validation of the denaturing capillary electrophoresis (DCE) assay for non-invasive liquid biopsy in lung carcinoma: A study of concordance with the cobas EGFR mutation test v2.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. e23040-e23040
Author(s):  
Marek Minarik ◽  
Barbora Belsanova ◽  
Ondrej Fiala ◽  
Milos Pesek ◽  
Lucie Benesova
Keyword(s):  
Capillary Electrophoresis ◽  
Liquid Biopsy ◽  
Egfr Mutation ◽  
Low Cost ◽  
Stage Iv ◽  
High Sensitivity ◽  
Circulating Tumor Dna ◽  
Plasma Samples ◽  
Iso 15189 ◽  
Lung Cancer Patients

e23040 Background: The detection of circulating tumor DNA (ctDNA), referred to as “liquid biopsy” is becoming a preferred option for some patients with lung adenocarcinomas. The key for a suitable technology is high sensitivity enabling to detect small fractions of mutated alleles, the method has to be solid against false-positives. The aim of this work was to compare our method based on capillary electrophoresis under denaturing conditions (DCE) to FDA and CE IVD approved commercial kit - cobas EGFR Mutation Test v2. Methods: A total of 44 plasma samples were acquired from patients with clinicaly confirmed stage IV lung adenocarcinoma. There were samples from patients prior to receiving therapy (with a known tissue EGFR genotype) as well as several ones showing progression on a previously administered antiEGFR-therapy (with no tissue info available at the time of plasma sampling). DNA was isolated from multiple 2 mL aliquots of each plasma. The first aliquot was run on cobas z480 real-time instrument using unmodified manufacturer's protocol. Another 2ml aliquot was run by DCE assay (accredited method under ISO 15189:2012), which is based on a PCR followed by a fragment analysis on an ABI capillary analyzer (Sanger sequencer). Results: The mutual concordance was 84.1% (37/44). cobas assay identified 11 EGFR-positive plasma samples (6 x Ex19del, 3 x L858R, 1x T790M combined with Ex19del and 1x T790M combined with L858R). DCE identified 8 EGFR-positive samples (6 x Ex19del, 1 x L858R, 1xT790M combined with L858R). DCE missed 5 of cobas-positive samples (2x Ex19del, 2x L858R and 1x T790M) while in turn identifying 2x Ex19del plasma samples that were concordant with tissue, but missed by cobas. Overall concordance with tissue was 73.3% (22/30) for cobas, 70.0% (21/30) for DCE assay and 80.0% (24/30) for both combined. Conclusions: DCE approach is reliable for low levels of mutations in plasma of lung cancer patients and comparable (and complementary) to cobas. Its advantage is simplicity, low cost and a universal, straighforward adaptability to virtually any mutation at any site allowing to detect oncogenic as well as tumor supressor mutations in ctDNA. Supported by grant 17-30748A.

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