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1.
Anticancer Res ; 40(10): 5577-5582, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32988881

RESUMO

BACKGROUND/AIM: Clusters of circulating tumor cells (CTCs) increase metastatic potential compared to single CTC. However, conventional technologies have been unable to generate an accurate analysis of single and cluster CTCs in the peripheral blood. We propose an effective strategy to detect and isolate both single and cluster CTCs using two size-selective microfilters. MATERIALS AND METHODS: Five ml of whole blood were collected from 10 patients with epidermal growth factor receptor mutation-positive non-small cell lung cancer. Single and cluster CTCs were identified using precision microfiltration membranes with two distinct pore sizes together with anti-EpCAM antibody labeling. RESULTS: Single and cluster CTCs were detected by simultaneously using two size-selective microfilters. The EGFR-L858R mutation was detected in the DNA from cells captured using both microfilters. CONCLUSION: Our method can be used to detect and isolate single and cluster CTCs in the whole blood and may facilitate the development of a liquid biopsy strategy.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/sangue , DNA Tumoral Circulante/sangue , Molécula de Adesão da Célula Epitelial/sangue , Células Neoplásicas Circulantes/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Pulmonar de Células não Pequenas/patologia , Separação Celular , Feminino , Humanos , Biópsia Líquida , Masculino , Pessoa de Meia-Idade , Mutação/genética , Células Neoplásicas Circulantes/patologia
2.
Nat Commun ; 11(1): 4489, 2020 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-32895384

RESUMO

We report a covalent chemistry-based hepatocellular carcinoma (HCC)-specific extracellular vesicle (EV) purification system for early detection of HCC by performing digital scoring on the purified EVs. Earlier detection of HCC creates more opportunities for curative therapeutic interventions. EVs are present in circulation at relatively early stages of disease, providing potential opportunities for HCC early detection. We develop an HCC EV purification system (i.e., EV Click Chips) by synergistically integrating covalent chemistry-mediated EV capture/release, multimarker antibody cocktails, nanostructured substrates, and microfluidic chaotic mixers. We then explore the translational potential of EV Click Chips using 158 plasma samples of HCC patients and control cohorts. The purified HCC EVs are subjected to reverse-transcription droplet digital PCR for quantification of 10 HCC-specific mRNA markers and computation of digital scoring. The HCC EV-derived molecular signatures exhibit great potential for noninvasive early detection of HCC from at-risk cirrhotic patients with an area under receiver operator characteristic curve of 0.93 (95% CI, 0.86 to 1.00; sensitivity = 94.4%, specificity = 88.5%).


Assuntos
Biomarcadores Tumorais/isolamento & purificação , Carcinoma Hepatocelular/diagnóstico , Detecção Precoce de Câncer/métodos , Vesículas Extracelulares/genética , Neoplasias Hepáticas/diagnóstico , Idoso , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Estudos de Casos e Controles , Química Click/instrumentação , Química Click/métodos , Química Computacional , Simulação por Computador , Diagnóstico Diferencial , Dimetilpolisiloxanos/química , Progressão da Doença , Detecção Precoce de Câncer/instrumentação , Feminino , Células Hep G2 , Humanos , Dispositivos Lab-On-A-Chip , Biópsia Líquida/instrumentação , Biópsia Líquida/métodos , Cirrose Hepática/sangue , Cirrose Hepática/patologia , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Masculino , Técnicas Analíticas Microfluídicas/instrumentação , Técnicas Analíticas Microfluídicas/métodos , Pessoa de Meia-Idade , Nanoestruturas/química , Nanofios/química , Estadiamento de Neoplasias , RNA Mensageiro/genética , RNA Mensageiro/isolamento & purificação , Curva ROC , Reação em Cadeia da Polimerase Via Transcriptase Reversa/instrumentação , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos
3.
J Immunother Cancer ; 8(2)2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32788235

RESUMO

To report a multi-institutional case series of patients with advanced microsatellite instability high (MSI-H) prostate adenocarcinoma identified with clinical cell-free DNA (cfDNA) next-generation sequencing (NGS) testing and treated with immune checkpoint inhibitors. Retrospective analysis of patients with metastatic castration-resistant prostate cancer (mCRPC) and MSI-H tumor detected by a commercially available cfDNA NGS assay Guardant360 (G360, Guardant Health) at eight different Academic Institutions in the USA, from September 2018 to April 2020. From a total of 14 MSI-H metastatic prostate cancer patients at participating centers, nine patients with mCRPC with 56% bone, 33% nodal, 11% liver and 11% soft-tissue metastases and a median PSA of 29.3 ng/dL, were treated with pembrolizumab after 2 lines of therapy for CRPC. The estimated median time on pembrolizumab was 9.9 (95% CI 1.0 to 18.8) months. Four patients (44%) achieved PSA50 after a median of 4 (3-12) weeks after treatment initiation including three patients with >99% PSA decline. Among the patients evaluable for radiographic response (n=5), the response rate was 60% with one complete response and two partial responses. Best response was observed after a median of 3.3 (1.4-7.6) months. At time of cut-off, four patients were still on pembrolizumab while four patients discontinued therapy due to progressive disease and one due to COVID-19 infection. Half of the patients with PSA50 had both MSI-H and pathogenic alterations in BRCA1 and BRCA2 in their G360 assays. The use of liquid biopsy to identify metastatic prostate cancer patients with MSI-H is feasible in clinical practice and may overcome some of the obstacles associated with prostate cancer tumor tissue testing. The robust activity of pembrolizumab in selected patients supports the generalized testing for MSI-H.


Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Instabilidade de Microssatélites , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais Humanizados/efeitos adversos , Antineoplásicos Imunológicos/efeitos adversos , Proteína BRCA1/genética , Proteína BRCA2/genética , Biomarcadores Tumorais/genética , DNA Tumoral Circulante/sangue , Infecções por Coronavirus , Humanos , Biópsia Líquida , Masculino , Pessoa de Meia-Idade , Mutação , Pandemias , Pneumonia Viral , Neoplasias da Próstata/patologia
5.
PLoS One ; 15(7): e0235611, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32634139

RESUMO

Liquid biopsy has become widely applied in clinical medicine along with the progress in innovative technologies, such as next generation sequencing, but the origin of circulating tumor DNA (ctDNA) has not yet been precisely established. We reported bimodal peaks of long fragment circulating free DNA (cfDNA) of 5 kb and short fragment cfDNA of 170 bp in patients with advanced lung cancer, and both contained ctDNA. In this paper, we demonstrate that the total amount of cfDNA is higher when patients with lung cancer have extrathoracic metastases, and the amount of long fragment cfDNA is significantly higher in those patients. To investigate the origin of long fragment cfDNA, conditioned media isolated from lung cancer cell lines was fractionated. Long fragment cfDNA was found concomitant with extracellular vesicles (EVs), but short fragment cfDNA was not observed in any fractions. However, in peripheral blood from a metastatic animal model both fragments were detected even with those same lung cancer cell lines. In human plasma samples, long fragment cfDNA was observed in the same fraction as that from conditioned media, and short fragment cfDNA existed in the supernatant after centrifugation at 100,000g. Concentration of ctDNA in the supernatant was two times higher than that in plasma isolated by the conventional procedure. Long fragment cfDNA associated with tumor progression might therefore be released into peripheral blood, and it is possible that the long fragment cfDNA escapes degradation by co-existing with EVs. Examination of the biological characteristics of long fragment cfDNA is a logical subject of further investigation.


Assuntos
DNA Tumoral Circulante/análise , Neoplasias Pulmonares/diagnóstico , Animais , Estudos de Casos e Controles , Linhagem Celular Tumoral , DNA Tumoral Circulante/química , Meios de Cultivo Condicionados/metabolismo , Modelos Animais de Doenças , Receptores ErbB/genética , Vesículas Extracelulares/genética , Vesículas Extracelulares/metabolismo , Frequência do Gene , Humanos , Biópsia Líquida , Pneumopatias/sangue , Pneumopatias/patologia , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Knockout , Mutação , Metástase Neoplásica , Transplante Heterólogo
6.
Rozhl Chir ; 99(4): 179-182, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32545981

RESUMO

INTRODUCTION: Colorectal cancer (CRC) is the third most common malignant disease worldwide. The stage of the disease at the time of diagnosis and the capture of an early recurrence have a direct impact on long-term survival. Existing control screening methods often do not reflect real-time metastatic disease. In patients with detectable circulating tumor DNA (ctDNA), liquid biopsy can be an effective monitoring tool. CASE REPORT: In 2012, we performed sigmoid resection in a 57 years old patient for advanced CRC. The follow-up assessments included: blood samples for CA 19-9 and CEA, endoscopy and imaging methods. We also sampled peripheral blood to determine the level of ctDNA. Its value corresponded to the development of the disease throughout the period. Twice it outperformed imaging methods. CEA showed some degree of unreliability, especially after prolonged illness. CA 19-9 was in the normal range at all times. CONCLUSION: Circulating tumor DNA is an effective tool in the diagnosis of recurrent metastatic CRC. In patients with detectable ctDNA, its level correlates with the tumoral mass in real time. It has a predictive value in monitoring the treatment response. Its implementation in the follow-up of patients with CRC may have an impact on the choice of treatment strategy and consequently on patient survival.


Assuntos
DNA Tumoral Circulante/genética , Neoplasias Colorretais , Biomarcadores Tumorais/genética , Humanos , Biópsia Líquida , Pessoa de Meia-Idade , Recidiva Local de Neoplasia
7.
Anticancer Res ; 40(6): 3435-3444, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32487642

RESUMO

BACKGROUND/AIM: Although it has been suggested that circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) might be used in a complementary manner in lung cancer diagnosis, limited confirmatory data are available. In this prospective study, we evaluated the diagnostic performance of each assay separately and in combination. PATIENTS AND METHODS: From March 2018 to January 2019, patients with suspected primary lung cancer, who underwent routine lung cancer work-up and peripheral blood sampling, were prospectively enrolled in the study. Epithelial cell adhesion molecule and cytokeratin served as markers of CTCs. In terms of ctDNA analysis, single-nucleotide variants were evaluated via next-generation sequencing. RESULTS: We analyzed 111 patients, including 99 with primary lung cancer and 12 with benign pulmonary disease. The median number of CTCs in 10 ml of blood was 3. The most frequently detected single nucleotide variants of ctDNA were TP53, CDKN2A, and EGFR. The diagnostic sensitivity of conventional tumor marker (combination of carcinoembryonic antigen/CYFRA 21-1/neuron-specific enolase) was 66.7%, while those of the ctDNA and CTC assays were 72.7% and 65.7%, respectively. The sensitivity of the CTC/ctDNA combination (95.0%) was significantly greater than those of the CTC (p<0.001), ctDNA (p<0.001), or conventional tumor marker (p<0.001) alone. Subgroup analysis revealed that the sensitivity of the combination assay was greater than those of the CTC or ctDNA assays alone, regardless of tumor stage or histopathology type. CONCLUSION: The CTC/ctDNA combination assay enhanced the sensitivity of primary lung cancer diagnosis. The combination assay strategy may be clinically useful and could enhance the early detection of lung cancer (ClinicalTrials.gov number: NCT03479099).


Assuntos
Biomarcadores Tumorais , DNA Tumoral Circulante , DNA de Neoplasias , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/etiologia , Células Neoplásicas Circulantes/patologia , Idoso , Idoso de 80 Anos ou mais , DNA de Neoplasias/sangue , Suscetibilidade a Doenças , Feminino , Humanos , Biópsia Líquida , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Polimorfismo de Nucleotídeo Único
8.
Anticancer Res ; 40(6): 3477-3484, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32487647

RESUMO

BACKGROUND/AIM: The Paris System (TPS) has recently been proposed as a method to standardize urinary cytology reporting. In this study, we evaluated the impact of implementing TPS compared to the traditional reporting system. PATIENTS AND METHODS: In total, 299 urine samples were reclassified according to TPS. We examined correlations between cytological and histological diagnoses, and calculated probabilities for detecting high-grade urothelial carcinoma (HGUC). RESULTS: TPS resulted in a decrease in the proportion of cases diagnosed as atypical urothelial cell (AUC) (43% to 31%). Among the AUC cases, the proportion of histologically confirmed HGUC cases rose (75% to 80%), as did the proportion of low-grade urothelial neoplasms (57% to 71%). All probabilities for detecting HGUC significantly increased using TPS. CONCLUSION: TPS improved the diagnostic yield of urinary cytology. The implementation of TPS is expected to be a major step towards standardizing urinary cytology reporting and providing clear information to clinicians.


Assuntos
Citodiagnóstico/métodos , Biópsia Líquida/métodos , Urinálise/métodos , Neoplasias Urológicas/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Biópsia , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Gradação de Tumores , Guias de Prática Clínica como Assunto , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Neoplasias Urológicas/urina
9.
Eur Urol Focus ; 6(5): 1086-1096, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32540268

RESUMO

CONTEXT: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic that erupted in December 2019 has affected more than a million people from over 200 countries, claiming over 70 000 lives (by April 7, 2020). As the viral infection is driven by increased angiotensin-converting enzyme-2 (ACE2) expression, with the kidney exhibiting the highest expression, it is crucial to gain insights into the mechanisms underlying renal cell carcinoma (RCC) and coronavirus disease 2019 (COVID-19). OBJECTIVE: This study considers up-to-date information on the biological determinants shared by COVID-19 and renal disease, and aims to provide evidence-based recommendations for the clinical management of RCC patients with COVID-19. EVIDENCE ACQUISITION: A literature search was performed using all sources (MEDLINE, EMBASE, ScienceDirect, Cochrane Libraries, and Web of Science). As of March 31, 2020, the Center for Disease Control reported that of the adults hospitalized for COVID-19 with underlying conditions in the USA, 74.8% had chronic renal disease. EVIDENCE SYNTHESIS: Evidence is discussed from epidemiological studies on SARS-CoV-2 pandemic and molecular studies on the role of kidney in facilitating routes for SARS-CoV-2 entry, leading to increased virulence of SARS-CoV-2 and clinical manifestation of symptoms in RCC. CONCLUSIONS: This analysis will advance our understanding of (1) the molecular signatures shared by RCC and COVID-19 and (2) the clinical implications of overlapping signaling pathways in the therapeutic management of RCC and COVID-19 patients. PATIENT SUMMARY: Amid the coronavirus disease 2019 (COVID-19) pandemic, patients diagnosed with renal cell carcinoma and infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may receive complimentary treatment modalities to enhance therapeutic response.


Assuntos
Betacoronavirus/metabolismo , Carcinoma de Células Renais/metabolismo , Infecções por Coronavirus/metabolismo , Neoplasias Renais/metabolismo , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/metabolismo , Insuficiência Renal Crônica/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Lesão Renal Aguda/epidemiologia , Lesão Renal Aguda/metabolismo , Lesão Renal Aguda/terapia , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Anticorpos Neutralizantes/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Antivirais/uso terapêutico , Carcinoma de Células Renais/epidemiologia , Comorbidade , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/fisiopatologia , Antagonistas dos Receptores de Endotelina/uso terapêutico , Hospitalização , Humanos , Ipilimumab/uso terapêutico , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/epidemiologia , Biópsia Líquida , Nivolumabe/uso terapêutico , Pandemias , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/epidemiologia , Pneumonia Viral/fisiopatologia , Inibidores de Proteínas Quinases/uso terapêutico , Diálise Renal , Insuficiência Renal Crônica/epidemiologia , Insuficiência Renal Crônica/terapia , Serina Endopeptidases/metabolismo , Índice de Gravidade de Doença , Sunitinibe/uso terapêutico
10.
PLoS One ; 15(6): e0234182, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32492056

RESUMO

The development of noninvasive approaches for brain tumor diagnosis and monitoring continues to be a major medical challenge. Although blood-based liquid biopsy has received considerable attention in various cancers, limited progress has been made for brain tumors, at least partly due to the hindrance of tumor biomarker release into the peripheral circulation by the blood-brain barrier. Focused ultrasound (FUS) combined with microbubbles induced BBB disruption has been established as a promising technique for noninvasive and localized brain drug delivery. Building on this established technique, we propose to develop FUS-enabled liquid biopsy technique (FUS-LBx) to enhance the release of brain tumor biomarkers (e.g., DNA, RNA, and proteins) into the circulation. The objective of this study was to demonstrate that FUS-LBx could sufficiently increase plasma levels of brain tumor biomarkers without causing hemorrhage in the brain. Mice with orthotopic implantation of enhanced green fluorescent protein (eGFP)-transfected murine glioma cells were treated using magnetic resonance (MR)-guided FUS system in the presence of systemically injected microbubbles at three peak negative pressure levels (0.59, 1.29, and 1.58 MPa). Plasma eGFP mRNA levels were quantified with the quantitative polymerase chain reaction (qPCR). Contrast-enhanced MR images were acquired before and after the FUS sonication. FUS at 0.59 MPa resulted in an increased plasma eGFP mRNA level, comparable to those at higher acoustic pressures (1.29 MPa and 1.58 MPa). Microhemorrhage density associated with FUS at 0.59 MPa was significantly lower than that at higher acoustic pressures and not significantly different from the control group. MRI analysis revealed that post-sonication intratumoral and peritumoral hyperenhancement had strong correlations with the level of FUS-induced biomarker release and the extent of hemorrhage. This study suggests that FUS-LBx could be a safe and effective brain-tumor biomarker release technique, and MRI could be used to develop image-guided FUS-LBx.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Ultrassonografia de Intervenção/métodos , Animais , Biomarcadores Tumorais/sangue , Barreira Hematoencefálica , Neoplasias Encefálicas/diagnóstico por imagem , Linhagem Celular Tumoral , Meios de Contraste , Feminino , Glioblastoma/diagnóstico por imagem , Proteínas de Fluorescência Verde/sangue , Proteínas de Fluorescência Verde/genética , Hemorragias Intracranianas/etiologia , Hemorragias Intracranianas/patologia , Biópsia Líquida/métodos , Imagem por Ressonância Magnética , Camundongos , Ultrassonografia de Intervenção/efeitos adversos
11.
PLoS One ; 15(6): e0234302, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32525942

RESUMO

Circulating tumor DNA (ctDNA)-based molecular profiling is rapidly gaining traction in clinical practice of advanced cancer patients with multi-gene next-generation sequencing (NGS) panels. However, clinical outcomes remain poorly described and deserve further validation with personalized treatment of patients with genomic alterations detected in plasma ctDNA. Here, we describe the outcomes, disease control rate (DCR) at 3 months and progression-free survival (PFS) in oncogenic-addicted advanced NSCLC patients with actionable alterations identified in plasma by ctDNA liquid biopsy assay, InVisionFirst®-Lung. A pooled retrospective analysis was completed of 81 advanced NSCLC patients with all classes of alterations predicting response to current FDA approved drugs: sensitizing common EGFR mutations (78%, n = 63) with T790M (73%, 46/63), ALK / ROS1 gene fusions (17%, n = 14) and BRAF V600E mutations (5%, n = 4). Actionable driver alterations detected in liquid biopsy were confirmed by prior tissue genomic profiling in all patients, and all patients received personalized treatment. Of 82 patients treated with matched targeted therapies, 10% were at first-line, 41% at second-line, and 49% beyond second-line. Acquired T790M at TKI relapse was detected in 73% (46/63) of patients, and all prospective patients (34/46) initiated osimertinib treatment based on ctDNA results. The 3-month DCR was 86% in 81 evaluable patients. The median PFS was of 14.8 months (12.1-22.9m). Baseline ctDNA allelic fraction of genomic driver did not correlate with the response rate of personalized treatment (p = 0.29). ctDNA molecular profiling is an accurate and reliable tool for the detection of clinically relevant molecular alterations in advanced NSCLC patients. Clinical outcomes with targeted therapies endorse the use of liquid biopsy by amplicon-based NGS ctDNA analysis in first line and relapse testing for advanced NSCLC patients.


Assuntos
Carcinogênese/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Genômica , Sequenciamento de Nucleotídeos em Larga Escala , Neoplasias Pulmonares/genética , Mutação , Idoso , Carcinoma Pulmonar de Células não Pequenas/patologia , Estudos de Viabilidade , Feminino , Humanos , Estimativa de Kaplan-Meier , Biópsia Líquida , Neoplasias Pulmonares/patologia , Masculino , Estadiamento de Neoplasias
12.
Adv Exp Med Biol ; 1257: 95-105, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32483734

RESUMO

Liquid biopsies encompass a number of new technologies designed to derive tumor data through the minimally invasive sampling of an accessible body fluid. These technologies remain early in their clinical development, and applications for patients with osteosarcoma are actively under investigation. In this chapter, we outline the current state of liquid biopsy technologies as they apply to cancer generally and osteosarcoma specifically, focusing on assays that detect and profile circulating tumor DNA (ctDNA), microRNAs (miRNA), and circulating tumor cells (CTCs). At present, ctDNA assays are the most mature, with multiple assays demonstrating the feasibility of detecting and quantifying ctDNA from blood samples of patients with osteosarcoma. Initial studies show that ctDNA can be detected in the majority of patients with osteosarcoma and that the detection and level of ctDNA correlates with a worse prognosis. Profiling of ctDNA can also identify specific somatic events that may have prognostic relevance, such as 8q gain in osteosarcoma. miRNAs are stable RNAs that regulate gene expression and are known to be dysregulated in cancer, and patterns of miRNA expression have been evaluated in multiple studies of patients with osteosarcoma. While studies have identified differential expression of many miRNAs in osteosarcomas compared to healthy controls, a consensus set of prognostic miRNAs has yet to be definitively validated. Recent studies have also demonstrated the feasibility of capturing CTCs in patients with osteosarcoma. The development of assays that quantify and profile CTCs for use as prognostic biomarkers or tools for biologic discovery is still in development. However, CTC technology holds incredible promise given the potential to perform multi-omic approaches in single cancer cells to understand osteosarcoma heterogeneity and tumor evolution. The next step required to move liquid biopsy technologies closer to helping patients will be wide-scale collection of patient samples from large prospective studies.


Assuntos
Neoplasias Ósseas , DNA Tumoral Circulante , Biópsia Líquida , Osteossarcoma , Biomarcadores Tumorais/sangue , Neoplasias Ósseas/terapia , DNA Tumoral Circulante/sangue , Humanos , Biópsia Líquida/normas , Osteossarcoma/terapia , Estudos Prospectivos
13.
PLoS One ; 15(6): e0231854, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32479545

RESUMO

UltraPrep is an open-source, two-step method for purification of cell-free DNA that entails extraction of total DNA followed by size-selective enrichment of the smaller fragments that are characteristic of DNA originating from fragmentation between nucleosome. The advantages of the two related protocols that are described are that they can easily accommodate a wide range of sample input volumes, they rely on simple, magnetic bead-based technology, the yields of cfDNA are directly comparable to the most popular methods for cfDNA purification, and they dramatically reduce the cost of cfDNA isolation relative to currently available commercial methods. We provide a framework for physical and molecular quality analysis of purified cfDNA and demonstrate that the cfDNA generated by UltraPrep meets or exceeds the quality metrics of the most commonly used procedure. In addition, our method removes high molecular weight genomic DNA (hmwgDNA) that can interfere with downstream assay results, thereby addressing one of the primary concerns for preanalytical collection of blood samples.


Assuntos
Ácidos Nucleicos Livres/isolamento & purificação , Extração em Fase Sólida/métodos , Ácidos Nucleicos Livres/sangue , Humanos , Biópsia Líquida , Magnetismo , Nucleossomos/genética , Dióxido de Silício/química , Extração em Fase Sólida/economia
15.
Transl Res ; 223: 40-60, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32413499

RESUMO

Improvement of breast cancer (BC) patient's outcome is directly related to early detection. However, there is still a lack of reliable biomarkers for diagnosis, prognosis and, treatment follow up in BC, leading researchers to study the potential of liquid biopsy based on circulating microRNAs (c-miRNAs). These c-miRNAs can be cell-free or associated with extracellular vesicles (EVs), and have great advantages such as stability in biofluids, noninvasive accessibility compared to current techniques (core-biopsy and surgery), and expression associated with pathogenic conditions. Recently, a new promising field of EV-derived miRNAs (EV-miRNAs) as cancer biomarkers has emerged, receiving special attention due to their selective vesicle sorting which makes them accurate for disease detection. In this review, we discuss new findings about c-miRNA and their potential as biomarkers for BC diagnosis, prognosis, and therapy. Additionally, we address the impact of limitations associated with the standardization of analysis techniques and methods on the implementation of these biomarkers in the clinical setting.


Assuntos
Biomarcadores Tumorais/sangue , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/patologia , MicroRNA Circulante/metabolismo , Vesículas Extracelulares/metabolismo , Biópsia Líquida , Neoplasias da Mama/genética , Feminino , Humanos , Prognóstico
16.
J Cancer Res Clin Oncol ; 146(8): 2051-2066, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32462295

RESUMO

PURPOSE: To explore whether targeted next generation sequencing (NGS) of liquid biopsy in advanced non-small cell lung cancer (NSCLC) could potentially overcome the innate problems that arise with standard tissue biopsy, like intratumoral heterogeneity and the inability to obtain adequate samples for analysis. METHODS: The Scopus, Cochrane Library, and MEDLINE (via PubMed) databases were searched for studies with matched tissue and liquid biopsies from advanced NSCLC patients, analyzed with targeted NGS. The number of mutations detected in tissue biopsy only, liquid biopsy only, or both was assessed and the positive percent agreement (PPA) of the two methods was calculated for every clinically relevant gene. RESULTS: A total of 644 unique relevant articles were retrieved and data were extracted from 38 studies fulfilling the inclusion criteria. The sample size was composed of 2000 mutations tested in matched tissue and liquid biopsies derived from 1141 patients. No studies analyzed circulating tumor cells. The calculated PPA rates were 53.6% (45/84) for ALK, 53.9% (14/26) for BRAF, 56.5% (13/23) for ERBB2, 67.8% (428/631) for EGFR, 64.2% (122/190) for KRAS, 58.6% (17/29) for MET, 54.6% (12/22) for RET, and 53.3% (8/15) for ROS1. We additionally recorded data for 65 genes that are not recommended by current guidelines for mutational testing. An extra category containing results of unspecified genes was added, with a PPA rate of 55.7% (122/219). CONCLUSION: Despite many advantages, liquid biopsy might be unable to fully substitute its tissue counterpart in detecting clinically relevant mutations in advanced NSCLC patients. However, it may serve as a helpful tool when making therapeutic decisions. More studies are needed to evaluate its role in everyday clinical practice.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Biópsia/métodos , Carcinoma Pulmonar de Células não Pequenas/sangue , DNA Tumoral Circulante/genética , Análise Mutacional de DNA , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Biópsia Líquida/métodos , Neoplasias Pulmonares/sangue , Células Neoplásicas Circulantes/patologia
17.
J Chromatogr A ; 1624: 461148, 2020 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-32376029

RESUMO

The variation patterns of transcription factors (TFs) provide direct information for the states of cell populations, which is of significance for biomedical research and clinical diagnostics. Herein, we show that through multi-channeled isothermal amplification, it is feasible to connect DNA-based signal transduction with chromatography for multiplexed detection of TFs. The described system is referred to as "PAC" which includes three major steps: (i) Protection, which uses DNA-modified magnetic beads to capture TFs and converts the capturing event into triggering signal; (ii) Amplification, which receives the triggering signal and generate DNA reporters through multi-channeled extension and nicking of oligonucleotides; and (iii) Chromatography, which separates and detects the DNA reporters in liquid chromatography. The quantitative detection of five essential TFs includes p50, p53, AP-1, MITF, and c-Myc is realized in a multiplexed manner, with the lowest detection limit of 0.5 pM. PAC can also provide effective means to measure the above five TFs in real samples, including cultured cells, xenograft tumors, and blood-based liquid biopsy. This study not only established a solution for multiplexed measurement of TFs for molecular diagnostics, but also paved avenue for bridging the gap between DNA nanotechnology and chromatography.


Assuntos
Cromatografia Líquida , Técnicas de Amplificação de Ácido Nucleico , Fatores de Transcrição/análise , Animais , Células Cultivadas , DNA/química , Sondas de DNA , Humanos , Limite de Detecção , Biópsia Líquida , Camundongos , Nanoestruturas , Nanotecnologia , Oligonucleotídeos , Ensaios Antitumorais Modelo de Xenoenxerto
18.
PLoS One ; 15(5): e0232754, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32379795

RESUMO

Analyzing cell-free DNA (cfDNA) as a source of circulating tumor DNA is useful for diagnosing or monitoring patients with cancer. However, the concordance between cfDNA within liquid biopsy and genomic DNA (gDNA) within tumor tissue biopsy is still under debate. To evaluate the concordance in a clinical setting, we enrolled 54 patients with metastatic colorectal cancer and analyzed their plasma cfDNA, gDNA from peripheral blood mononuclear cells (PBMC), and gDNA from available matched tumor tissues using ultra-deep sequencing targeting 10 genes (38-kb size) recurrently mutated in colorectal cancer. We first established a highly reliable cut-off value using reference material. The sensitivity of detecting KRAS hotspot mutations in plasma was calculated as 100%, according to digital droplet PCR. We could selectively detect clinically important somatic alterations with a variant allele frequency as low as 0.18%. We next compared somatic mutations of the 10 genes between cfDNA and genomic DNA from tumor tissues and observed an overall 93% concordance rate between the two types of samples. Additionally, the concordance rate of patients with the time interval between liquid biopsy and tumor tissue biopsy within 6 months and no prior exposure to chemotherapy was much higher than those without. The patients with KRAS mutant fragments in plasma had poor prognosis than those without the mutant fragments (33 months vs. 63 months; p<0.05). Consequently, the profiling with our method could achieve highly concordant results and may facilitate the surveillance of the tumor status with liquid biopsy in CRC patients.


Assuntos
Ácidos Nucleicos Livres/genética , DNA Tumoral Circulante/genética , Neoplasias Colorretais/genética , Adulto , Idoso , Neoplasias Colorretais/patologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Biópsia Líquida , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/secundário , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/secundário , Masculino , Pessoa de Meia-Idade , Mutação , Metástase Neoplásica/genética , Proteínas Proto-Oncogênicas p21(ras)/genética
19.
Crit Rev Oncol Hematol ; 151: 102978, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32428812

RESUMO

Precision medicine was born with the development of new diagnostic techniques and targeted drugs, yielding better outcomes in cancer care. With the evolution and increasing sensitivity for detecting oncogenic drivers, liquid biopsies (LBs), specifically cell-free DNA (cfDNA) analysis, have been proposed as a minimally-invasive technique for genomic profiling. Ranging from sequencing techniques to PCR-based methods and other more complex strategies, this approach, currently applicable in some solid tumors with robust evidence, is showing promising opportunities in other cancers. However, difficulties in validating their clinical utility exist within limitation at different levels among several techniques, reporting of the results, lack of appropriate clinical trial designs, and unknown economic impact. One of the aims of the ISLB is to create recommendations to develop reliable and sustainable diagnostic, prognostic and predictive tools using LBs. This paper is addressing these objectives, helping the healthcare providers and scientific community to understand the potential of LB.


Assuntos
Ácidos Nucleicos Livres , Biópsia Líquida , Neoplasias/patologia , Medicina de Precisão/métodos , Biomarcadores Tumorais , Humanos , Sociedades Médicas
20.
Transl Res ; 222: 56-66, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32413498

RESUMO

Management of advanced prostate cancer remains complex, with substantial changes in treatment options emerging in recent years having implications for treatment selection and sequencing. Recognition of the importance of androgen signaling has led to life-prolonging treatments, as well as "liquid biopsy" techniques to guide these treatments in some settings. Therapies that target estrogen receptor signaling are efficacious but infrequently used options for treatment of castration-resistant prostate cancer. It is possible that nuances of estrogen receptor (ER) signaling, or selective modulation of ER signaling, might favorably influence outcomes in castration-resistant prostate cancer. Expression of ERs and their variants has been investigated in other cancers such as breast. Constitutively activating gene alterations can potentially lead to ER activation and subsequently promote cancer progression. The identification of these aberrations may help identify cancer phenotypes that are susceptible or resistant to therapies involved in ER signaling. This review outlines the current literature regarding ER signaling in prostate cancer, and provides background for exploration of potentially useful ER signaling biomarkers in advanced prostate cancer.


Assuntos
Carcinogênese/metabolismo , Neoplasias da Próstata/metabolismo , Receptores Estrogênicos/metabolismo , Transdução de Sinais , Humanos , Biópsia Líquida , Masculino , Processamento de RNA/genética , Receptores Estrogênicos/genética
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