新的肿瘤分子标记物--肿瘤相关循环DNA

肿瘤患者外周血循环DNA的存在及相关改变为肿瘤的诊断和监测提供了新的靶分子,血浆／血清作为容易获得的检测对象比传统的癌组织活检具有更大的使用弹性和临床应用价值,循环DNA作为新的肿瘤标志物的研究在不久的未来将成为科学研究的热点。     

循环肿瘤DNA突变检测方法研究进展

循环肿瘤DNA(Circulating Tumor DNA,ctDNA)含有肿瘤的遗传信息,与肿瘤组织具有高度的一致性,可代替肿瘤组织用于肿瘤的早期诊断,预后监测和药物疗效监测,是一种具有良好临床应用前景的液体活检标记物。但血液中的ctDNA片段化程度高,含量稀少,且与野生型DNA混合存在(约1%甚至更低),并会随着患者肿瘤分期等情况动态变化,造成了ctDNA检测困难,需要高灵敏度高特异性的突变检测方法,才能够从大量的野生型DNA中检测出微量突变型ctDNA。目前,灵敏度和特异性能够满足ctDNA检测需求的方法主要有扩增阻滞突变系统PCR(Amplification Refractory Mutation System PCR,ARMS-PCR)、钳制PCR(Clamping-PCR)、数字化PCR(Digital-PCR)、西格诺公司的质谱分辨技术(Sequenom UltraseekTM)和高通量测序技术。本文对这些方法的原理、特点、最新进展和应用前景进行了综述,为研究人员选择合适的ctDNA检测方法提供理论依据。     

循环肿瘤细胞在女性实体器官肿瘤中的研究进展

循环肿瘤细胞(circulating tumor cells,CTCs)指的是从实体的肿瘤或转移的病灶进入外周血液循环的恶性肿瘤细胞。自发现以来,随着其检验技术日趋成熟,循环肿瘤细胞(CTCs)日渐成为肿瘤学炙手可热的研究对象。因为它将通过外周血的检验来实现监测肿瘤的发生、发展、转移、复发等情况,相对于肿瘤实体活检,"液体活检"不仅让患者易于接受,更有利于医务工作者监测病情变化。本文综述了循环肿瘤细胞(CTCs)的检测方法并综述了循环肿瘤细胞在女性实体肿瘤--乳腺癌、卵巢癌、宫颈癌、子宫内膜癌中的研究进展。其中着重介绍了其在早期乳腺癌及复发转移性乳腺癌中的重大意义以及在评价治疗效果中的分子学特征。实践表明,循环肿瘤细胞(CTCs)与HE-4、CA125的联合应用在评估卵巢癌化疗敏感性中也具有重要的临床意义。     

循环肿瘤DNA检测技术的研究进展

循环肿瘤DNA(circulating tumor DNA,ct DNA)检测,即所谓的"液体活检"。与传统的组织活检技术相比,"液体活检"能够更加全面地反映肿瘤的特征。随着ct DNA研究的逐渐深入,其检测技术的灵敏度和特异性都得到了很大的提升,而其中最具代表性的检测技术有数字PCR(digital PCR,d PCR)技术、流式技术的磁珠乳液扩增方法(bead,emulsion,amplification and magnetic,BEAMing)、标记扩增深度测序(tagged-amplicon deep sequencing,TAm-Seq)等,这些技术的发展有望开启精准医疗的新时代。     

扩增子测序技术应用于乳腺癌循环游离DNA无创性检测的研究

目的:应用扩增子测序方法检测乳腺癌循环游离DNA肿瘤相关突变。方法:军事医学科学院附属医院的10例HER2阳性晚期乳腺癌患者血样入组,应用扩增子测序技术检测血细胞和血浆中的循环游离DNA(cfDNA),筛选出肿瘤相关基因突变,即循环肿瘤DNA(ctDNA)。结果:检测10例晚期乳腺癌患者外周循环血50个基因,共检出11个突变基因,其中PIK3CA、ERBB4的阳性检出率均达到100%,AKT、TP53的阳性检出率为90%。结论:检测覆盖度及深度均良好,结果提示11个基因的热点突变区域出现单核苷酸的多态性基因突变,突变基因与PI3K-AKT-m TOR通路、Ras-Raf-MEK-ERK通路明显相关。扩增子测序技术和筛选方法可以用于乳腺癌ctDNA的无创性检测。     

消化道肿瘤的循环血长链非编码RNA及其检测新技术

长链非编码RNA(lncRNA)是一类转录本长度大于200个核苷酸的非编码RNA分子，广泛参与细胞增殖、分化、凋亡、染色质重塑等重要生命过程。大量研究证实，lncRNA可以在血液循环中稳定存在，其表达水平与消化道肿瘤的发生、发展和转归密切相关。本文简要回顾了近年来消化道肿瘤循环血lncRNA的最新研究进展，着重讨论了循环血lncRNA在消化道肿瘤诊断和预后评估中的临床应用价值及其检测新技术。     

液体活检对食管癌诊疗和预后的价值

食管癌是一种全球性的恶性肿瘤,由于其进展迅速并缺乏早期发现和有效预后的生物标志物,造成了患者较高的死亡率和较差的预后情况。近些年来,在精准肿瘤诊疗的大背景下,液体活检作为一种新兴的非侵入性检测方法,在食管癌疾病进展中可以实现动态监测,逐渐在临床上引起关注。液体活检通过从体液中获取肿瘤相关的生物标志物,如循环肿瘤DNA、循环肿瘤细胞、外泌体内容物等,来评估肿瘤的存在、特征和预后等。对食管癌患者进行及时有效的预后评估有助于改善其临床结局,故该文将对液体活检在食管癌的诊疗和预后中的科学研究及临床应用现状进行详细阐述,并指出目前液体活检中存在的挑战及其未来发展方向,期望能为食管癌的早期和超早期诊断、疗效动态监测、预后评估、个体化精准治疗决策的制定提供依据。     

非编码RNA在肿瘤液体活检中的研究进展

液体活检是一种基于分析体液样本中的生物标志物来诊断疾病、监测疾病进展和评估疗效的非侵入性的新型微创诊断技术。液体活检主要包括探查和检测循环肿瘤细胞、ctDNA、非编码RNAs和细胞外囊泡等。非编码RNA之前被认为是一类无功能的RNA,但近年来研究表明它们在肿瘤的发病机制中扮演着重要的角色。它们构成了基因调控的一个重要层面,其表达水平在多种癌种中呈现失调势态,这提示了它们作为肿瘤生物学标志物的临床应用潜力。飞速发展的高通量测序技术使得在泛癌领域建立非编码RNA表达谱的分子表征成为可能。该文系统阐述了非编码RNA作为非侵入性肿瘤标志物的研究进展,评估了其作为肿瘤生物标志物的适用性,同时总结了近期的检测技术突破对肿瘤分子标志物发展的影响。     

基于物理过滤与原位杂交技术的骨肉瘤循环肿瘤细胞检测及临床意义

目的:探寻一种有效地从骨肉瘤患者外周血中富集并鉴定循环肿瘤细胞的方法。方法:利用基于物理过滤与原位杂交结合的技术对骨肉瘤患者外周血循环肿瘤细胞分离并鉴定。采用直径8μm纳米滤膜截留外周血中体积较大的白细胞及肿瘤细胞,利用多重RNA原位杂交技术检测CD45、EpCAM、CK8、CK18、CK19、vimentin及twist基因表达,并根据结果对滤膜截留下的细胞进行鉴定并分型。结果:本研究所使用的基于物理过滤与原位杂交技术的循环肿瘤细胞检测方法可以高效地从骨肉瘤患者外周血中富集骨肉瘤循环肿瘤细胞,该方法富集细胞的效率超过90%。15例健康志愿者中1例志愿者检测结果阳性。20例纳入研究的骨肉瘤患者中19例患者外周血中检测出CTC,CTC计数范围为0-20。肿瘤转移患者外周血CTC计数为11.33±5.88,肿瘤未转移患者外周血CTC计数为4.36±2.98,差异具有统计学意义(P=0.0022)。肿瘤转移患者外周血间质型CTC比例高于肿瘤未转移患者(P=0.0031)。结论:利用基于物理过滤与原位杂交结合的技术可以有效地检测骨肉瘤患者外周血循环肿瘤细胞。CTC检测结果可以作为辅助判断肿瘤转移情况的辅助指标。     

单细胞基因组测序技术新进展及其在生物医学中的应用

随着单细胞基因组测序技术的建立与发展,对细胞基因组特征的分析进入了单细胞水平。单细胞的基因组分辨率不但使研究人员能够在单细胞尺度上分析肿瘤细胞的异质性,也使得传统上难以检测的稀有细胞的基因组研究成为可能。这些稀有细胞往往具有重要的生物学意义或临床价值,如癌症患者血液中循环肿瘤细胞(circulatingtumorcell,CTC)的基因组检测或三代试管婴儿植入前胚胎细胞的遗传缺陷诊断与筛查(preimplantation genetic diagnosis/screening, PGD/PGS)。本文总结了近年来发展的各种单细胞基因组扩增技术及其优缺点,并介绍了单细胞基因组测序技术在肿瘤生物学和临床检测中的应用,以期为单细胞基因组测序技术在临床检测中应用开发提供参考。     

Liquid biopsy: A new avenue in pathology

Liquid biopsy is a relatively new entity. This non‐invasive technique provides real‐time information about a tumour. The liquid biopsy contains circulating tumour cells, cell‐free DNA and exosomes. The main indications for liquid biopsy include early diagnosis, screening, detection of minimal residual disease, designing personalised treatment and predicting biological behaviour of the tumour. In this review, we discuss various aspects of liquid biopsy and compare it with conventional biopsy.     

Current and Future Perspectives of Cell-Free DNA in Liquid Biopsy

A liquid biopsy is a minimally invasive or non-invasive method to analyze a range of tumor material in blood or other body fluids, including circulating tumor cells (CTCs), cell-free DNA (cfDNA), messenger RNA (mRNA), microRNA (miRNA), and exosomes, which is a very promising technology. Among these cancer biomarkers, plasma cfDNA is the most widely used in clinical practice. Compared with a tissue biopsy of traditional cancer diagnosis, in assessing tumor heterogeneity, a liquid biopsy is more reliable because all tumor sites release cfDNA into the blood. Therefore, a cfDNA liquid biopsy is less invasive and comprehensive. Moreover, the development of next-generation sequencing technology makes cfDNA sequencing more sensitive than a tissue biopsy, with higher clinical applicability and wider application. In this publication, we aim to review the latest perspectives of cfDNA liquid biopsy clinical significance and application in cancer diagnosis, treatment, and prognosis. We introduce the sequencing techniques and challenges of cfDNA detection, analysis, and clinical applications, and discuss future research directions.     

Liquid biopsy for early diagnosis of non-small cell lung carcinoma: recent research and detection technologies

Liquid biopsy, an innovative method for early diagnosis of cancer, has changed the traditional method of diagnosing lung cancer and is considered a feasible auxiliary diagnostic tool. To date, various reports emphasize the need for non-small cell lung carcinoma (NSCLC), both with higher incidence and mortality and less effective treatments; thus, emphasizing the need for early detection of NSCLC for improved patient outcomes. Invasive tissue biopsy is a common diagnostic tool that is usually extracted from the primary tumor to indicate molecular composition. In comparison, liquid biopsy taken from body fluid reflects extensive malignant features nonexistent in primary tumors. Owing to new detection technologies, liquid biopsy reduces the need for invasive treatments and enhances the accuracy and specificity of early detection of cancer in clinical settings. This review summarizes some latest research on the diagnosis of early-stage NSCLC via liquid biopsy, including circulating DNA, circulating tumor cells, exosomes, and tumor-educated platelets, as well as their detection technologies, such as fluorescence in-situ hybridization-based, polymerase chain reaction-based, next-generation sequencing-based, Chip-based, and microfluidic methods. Additionally, we outline the existing challenges and possible solutions for liquid-biopsy biomarkers. Our study mainly highlights the merits of liquid biopsy as a promising biomarker for non-invasive detection in the future, particularly for the early detection of NSCLC, thereby benefitting human health.     

Current and emerging applications of liquid biopsy in pan-cancer

Cancer morbidity and mortality are growing rapidly worldwide and it is urgent to develop a convenient and effective method that can identify cancer patients at an early stage and predict treatment outcomes. As a minimally invasive and reproducible tool, liquid biopsy (LB) offers the opportunity to detect, analyze and monitor cancer in any body fluids including blood, complementing the limitations of tissue biopsy. In liquid biopsy, circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) are the two most common biomarkers, displaying great potential in the clinical application of pan-cancer. In this review, we expound the samples, targets, and newest techniques in liquid biopsy and summarize current clinical applications in several specific cancers. Besides, we put forward a bright prospect for further exploring the emerging application of liquid biopsy in the field of pan-cancer precision medicine.     

DNA methylation signatures in circulating cell-free DNA as biomarkers for the early detection of cancer

Detecting cell-free DNA(cfDNA) or circulating tumor DNA(ctDNA) in plasma or serum could serve as a "liquid biopsy", which would be useful for numerous diagnostic applications. cfDNA methylation detection is one of the most promising approaches for cancer risk assessment. Here, we reviewed the literature related to the use of serum or plasma circulating cell-free DNA for cancer diagnosis in the early stage and their power as future biomarkers.     

Circulating nucleic acids as a new diagnostic tool

The discovery of circulating nucleic acids in the 1940s opened up new possibilities for the non-invasive detection, monitoring and screening of various human disorders. Several tumour markers that enable early cancer detection or tumour behaviour prediction have been detected in the plasma of cancer patients. Maternal plasma analysis can be used to detect certain fetal abnormalities, with the quantification of cell-free nucleic acids used to screen for several pregnancy-associated disorders. Some other applications are in transplant monitoring and graft rejection assessment, and in certain medical emergencies such as trauma and burn severity stratification. Many studies have yielded promising results in this field, but the techniques have yet to be applied in routine clinical practice. Large-scale studies using similar technologies and a broad spectrum of patients are still needed to verify the results of the various studies.     

Neueste technologische Entwicklungen für die Analyse von zirkulierender Tumor-DNA

The analysis of circulating tumor DNA (ctDNA) and circulating tumor cells, often known as liquid biopsy, is a rapidly developing field of medical research. Although it has taken decades since the discovery of cell-free DNA for it to be recognized as a suitable biomarker, the clinical benefit of ctDNA with regard to monitoring therapy response, the identification of resistance mechanisms, and novel emerging actionable targets, in addition to the detection of minimal residual disease, has recently been proven in numerous studies.Owing to the great variability of ctDNA in the circulation, together with the high degree of fragmentation, ctDNA is a challenging analyte. However, in recent years technological advances have contributed to a variety of routine applications of ctDNA analysis becoming a reality, given that a number of additional regulatory hurdles can be overcome.     

Tumour biomarkers—Tracing the molecular function and clinical implication

In recent years, with the increase in cancer mortality caused by metastasis, and with the development of individualized and precise medical treatment, early diagnosis with precision becomes the key to decrease the death rate. Since detecting tumour biomarkers in body fluids is the most non‐invasive way to identify the status of tumour development, it has been widely investigated for the usage in clinic. These biomarkers include different expression or mutation in microRNAs (miRNAs), circulating tumour DNAs (ctDNAs), proteins, exosomes and circulating tumour cells (CTCs). In the present article, we summarized and discussed some updated research on these biomarkers. We overviewed their biological functions and evaluated their multiple roles in human and small animal clinical treatment, including diagnosis of cancers, classification of cancers, prognostic and predictive values for therapy response, monitors for therapy efficacy, and anti‐cancer therapeutics. Biomarkers including different expression or mutation in miRNAs, ctDNAs, proteins, exosomes and CTCs provide more choice for early diagnosis of tumour detection at early stage before metastasis. Combination detection of these tumour biomarkers may provide higher accuracy at the lowest molecule combination number for tumour early detection. Moreover, tumour biomarkers can provide valuable suggestions for clinical anti‐cancer treatment and execute monitoring of treatment efficiency.     

Genetic profiling of cancer with circulating tumor DNA analysis

Circulating cell-free tumor DNA(ctDNA) in the blood is DNA released from apoptotic, circulating, and living tumor cells. ctDNA is about 140 nt in length and has a half-life of about 1.5 h. ctDNA analysis provides a noninvasive means to assess the genetic profile of cancer in real time. With the advent of molecular technologies, including digital PCR and massively parallel sequencing(MPS), ctDNA analysis has shown promise as a highly sensitive and specific alternative to conventional tissue biopsy in cancer detection, longitudinal monitoring, and precision therapy. This review provides an overview of the latest development in our understanding of the biologic characteristics, detection methodologies, and potential clinical implications of ctDNA, as well as the challenges in translating ctDNA analysis from the research arena to patient care.     

Liquid biopsy in pancreatic cancer – Current perspective and future outlook

Pancreatic cancer is a lethal condition with a rising incidence and often presents at an advanced stage, contributing to abysmal five-year survival rates. Unspecific symptoms and the current lack of biomarkers and screening tools hamper early diagnosis. New technologies for liquid biopsies and their respective evaluation in pancreatic cancer patients have emerged over recent years. The term liquid biopsy summarizes the sampling and analysis of circulating tumor cells (CTCs), small extracellular vesicles (sEVs), and tumor DNA (ctDNA) from body fluids. The major advantages of liquid biopsies rely on their minimal invasiveness and repeatability, allowing serial sampling for dynamic insights to aid diagnosis, particularly early detection, risk stratification, and precision medicine in pancreatic cancer. However, liquid biopsies have not yet developed into a new pillar for clinicians' routine armamentarium. Here, we summarize recent findings on the use of liquid biopsy in pancreatic cancer patients. We discuss current challenges and future perspectives of this potentially powerful alternative to conventional tissue biopsies.