**1. Introduction**

Liquid biopsy represents a spectrum of technologies for the molecular analysis of blood and other biological liquids (e.g. urine, saliva or others) in order to detect cell-free/ tumour nucleic acids, exosomes, microRNAs, tumour-educated platelets and circulating or free-floating tumour or foetal cells, depending on the clinical context. In contrast to classic tumour markers, e.g. prostate-specific antigen (PSA) or carcinoembryonic antigen (CEA), liquid biopsy is distinguished by high specificity as it provides genomic, proteomic and cellular characteristics of the disease. The anticipated outstanding reliability of these tests has been reflected in the term itself: "liquid biopsy" is expected to be at least as informative as tissue biopsy, used as the gold standard in certain diagnostic fields, especially in oncology.

The advantages of liquid biopsy include non-invasive approach which is patient-friendly, associated with remarkably low possibility of complications and technically feasible even in patients who are in serious general status or affected by tumour or metastases that are not easily accessible by conventional tissue biopsy. The clinically simple application allows the repeated use of liquid biopsy resulting in real-time follow-up for the disease course. The testing is fast and exact as definite molecular markers are sought for. Generally, liquid biopsy is a much awaited tool to overcome the limitations set by tumour heterogeneity upon conventional tissue biopsy representing only a small part of the whole tumour [1]. However, some technological modifications of liquid biopsy in certain patients can be subjected to the same restrictions regarding heterogeneity. A characteristic example would be the assessment of circulating tumour cells (CTCs) in patients with low burden of malignant cells in the blood. For instance, the diagnostic threshold of

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

two CTCs per 7.5 mL of blood ensured high sensitivity and specificity for gastric cancer diagnosis, reaching 85.3 and 90.3%, respectively [2]. Thus, the diagnostic value of liquid biopsy in such setting has proven to be high, but the few malignant cells per sample cannot represent the full scope of cancer heterogeneity, by the number being inferior to tissue biopsy. Similarly, to characterize the tumour exactly, the circulating tumour DNA must represent at least 10% of the whole blood burden of cell-free DNA—a threshold that is not always reached [1].

Liquid biopsies are increasingly applied in the evaluation of oncological patients (**Figure 1**) due to the previously listed benefits over conventional tissue biopsies. However, this approach is not limited to the diagnostics of malignant tumours [3, 4].

In addition to the increasing use in oncology, liquid biopsy can be used in obstetrics and gynaecology to evaluate the molecular characteristics of foetus [3, 4]; in transplantology for the early detection of graft rejection; in haemodialysis and critical care medicine to identify tissue damage; and in rheumatology, e.g. to assess systemic lupus erythematosus [4].

Currently, liquid biopsy is not a routine laboratory test in clinical practice [5], and even its clinical efficacy has been seriously questioned by the American Society of Clinical Oncology and the College of American Pathologists [6], but certain tests are approved for use or reaching the clinical life (e.g. *EGFR* gene mutation testing, approved by the Food and Drug Administration (FDA) on 2016; tumour mutation burden in blood, confirmed in 2018 as an effective tool to predict the efficacy of immunological checkpoint inhibitors), and the related field is rapidly expanding [7, 8]. The development is seen as the progress in technologies, software and quality control systems as well as diagnosis-based research to substantiate many promising clinical applications (**Figure 2**).

**2. Circulating tumour cells and related issues**

**Figure 2.** The potential applications of liquid biopsy in oncology.

Invasion is a typical feature of malignant tumours. An epithelial malignancy, namely, carcinoma, starts its invasive growth from intraepithelial carcinoma in situ, destroys the basement membrane, infiltrates connective tissues and reaches smaller or larger blood and lymphatic vessels, entering the circulation. Thus, at some point of cancer pathogenesis, malignant cells appear in the bloodstream. This is a crucial step of carcinogenesis, leading to the metastatic spread—the hallmark of malignant process. Circulating tumour cells (CTCs) have been described by Thomas Ashworth as long ago as in 1869 [9]. Similarly, circulating foetal cells have been identified in peripheral maternal blood [3]. However, major technological advances were necessary to develop reliable tests to identify the circulating non-haematological cells. CTCs are generally recognised as non-leukocytic nucleated cells in the bloodstream. They can be recognised by their physical properties including large size, mechanical plasticity and dielectric mobility. Physical filters, density gradient, dielectric, microfluidic or photoacoustic methods are used for physical separation. Immunophenotype by expression of different antigens also can be applied to identify CTCs; the relevant methods include cytometric highthroughput imaging and immunomagnetic and adhesion-based separation as well as nega-

Introductory Chapter: Liquid Biopsy — A Promising Technology of the Future

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tive depletion of leukocytes and CTC identification by tumour markers [5].

**Figure 1.** Role of liquid biopsy in the stepwise evaluation of cancer patient.

**Figure 2.** The potential applications of liquid biopsy in oncology.

two CTCs per 7.5 mL of blood ensured high sensitivity and specificity for gastric cancer diagnosis, reaching 85.3 and 90.3%, respectively [2]. Thus, the diagnostic value of liquid biopsy in such setting has proven to be high, but the few malignant cells per sample cannot represent the full scope of cancer heterogeneity, by the number being inferior to tissue biopsy. Similarly, to characterize the tumour exactly, the circulating tumour DNA must represent at least 10% of the whole blood burden of cell-free DNA—a threshold that is not

Liquid biopsies are increasingly applied in the evaluation of oncological patients (**Figure 1**) due to the previously listed benefits over conventional tissue biopsies. However, this approach

In addition to the increasing use in oncology, liquid biopsy can be used in obstetrics and gynaecology to evaluate the molecular characteristics of foetus [3, 4]; in transplantology for the early detection of graft rejection; in haemodialysis and critical care medicine to identify

Currently, liquid biopsy is not a routine laboratory test in clinical practice [5], and even its clinical efficacy has been seriously questioned by the American Society of Clinical Oncology and the College of American Pathologists [6], but certain tests are approved for use or reaching the clinical life (e.g. *EGFR* gene mutation testing, approved by the Food and Drug Administration (FDA) on 2016; tumour mutation burden in blood, confirmed in 2018 as an effective tool to predict the efficacy of immunological checkpoint inhibitors), and the related field is rapidly expanding [7, 8]. The development is seen as the progress in technologies, software and quality control systems as well as diagnosis-based research to substantiate many

tissue damage; and in rheumatology, e.g. to assess systemic lupus erythematosus [4].

is not limited to the diagnostics of malignant tumours [3, 4].

promising clinical applications (**Figure 2**).

**Figure 1.** Role of liquid biopsy in the stepwise evaluation of cancer patient.

always reached [1].

4 Liquid Biopsy
