Field Of Liquid Biopsy Research Articles

Multidimensional fragmentomic profiling of cell-free DNA released from patient-derived organoids

BackgroundFragmentomics, the investigation of fragmentation patterns of cell-free DNA (cfDNA), has emerged as a promising strategy for the early detection of multiple cancers in the field of liquid biopsy. However, the clinical application of this approach has been hindered by a limited understanding of cfDNA biology. Furthermore, the prevalence of hematopoietic cell-derived cfDNA in plasma complicates the in vivo investigation of tissue-specific cfDNA other than that of hematopoietic origin. While conventional two-dimensional cell lines have contributed to research on cfDNA biology, their limited representation of in vivo tissue contexts underscores the need for more robust models. In this study, we propose three-dimensional organoids as a novel in vitro model for studying cfDNA biology, focusing on multifaceted fragmentomic analyses.ResultsWe established nine patient-derived organoid lines from normal lung airway, normal gastric, and gastric cancer tissues. We then extracted cfDNA from the culture medium of these organoids in both proliferative and apoptotic states. Using whole-genome sequencing data from cfDNA, we analyzed various fragmentomic features, including fragment size, footprints, end motifs, and repeat types at the end. The distribution of cfDNA fragment sizes in organoids, especially in apoptosis samples, was similar to that found in plasma, implying occupancy by mononucleosomes. The footprints determined by sequencing depth exhibited distinct patterns depending on fragment sizes, reflecting occupancy by a variety of DNA-binding proteins. Notably, we discovered that short fragments (< 118 bp) were exclusively enriched in the proliferative state and exhibited distinct fragmentomic profiles, characterized by 3 bp palindromic end motifs and specific repeats.ConclusionsIn conclusion, our results highlight the utility of in vitro organoid models as a valuable tool for studying cfDNA biology and its associated fragmentation patterns. This, in turn, will pave the way for further enhancements in noninvasive cancer detection methodologies based on fragmentomics.

Emerging histological and serological biomarkers in oral squamous cell carcinoma: Applications in diagnosis, prognosis evaluation and personalized therapeutics (Review).

Oral squamous cell carcinoma (OSCC) is the most common malignancy of the oral cavity and accounts for >90% of all oral cancers. Despite advances in diagnostic procedures and therapeutic interventions, overall survival has not improved significantly in recent decades, primarily due to late diagnosis, locoregional recurrence and treatment resistance. Identifying reliable biomarkers for early detection, prognosis evaluation and treatment response prediction is critical for improving clinical outcomes in patients with OSCC. In the present review, the prognostic and predictive utility of circulating biomarkers, such as circulating tumour cells, serological biomarkers and histological and genetic biomarkers, were explored in the context of OSCC. In addition, the potential role of immune checkpoints in the treatment of OSCC was highlighted and the rapidly evolving field of liquid biopsy and its potential to revolutionize diagnosis, prognosis evaluation and treatment were examined. The existing evidence for the clinical utility of these biomarkers was critically evaluated and the challenges and limitations associated with their introduction into routine clinical practice were addressed. In conclusion, the present review highlights the promising role of biomarkers in improving the current understanding of the pathogenesis of OSCC and offers potential avenues for improving patient care through personalized medicine approaches.

CfOmics: a cell-free multi-Omics database for diseases.

Liquid biopsy has emerged as a promising non-invasive approach for detecting, monitoring diseases, and predicting their recurrence. However, the effective utilization of liquid biopsy data to identify reliable biomarkers for various cancers and other diseases requires further exploration. Here, we present cfOmics, a web-accessible database (https://cfomics.ncRNAlab.org/) that integrates comprehensive multi-omics liquid biopsy data, including cfDNA, cfRNA based on next-generation sequencing, and proteome, metabolome based on mass-spectrometry data. As the first multi-omics database in the field, cfOmics encompasses a total of 17 distinct data types and 13 specimen variations across 69 disease conditions, with a collection of 11345 samples. Moreover, cfOmics includes reported potential biomarkers for reference. To facilitate effective analysis and visualization of multi-omics data, cfOmics offers powerful functionalities to its users. These functionalities include browsing, profile visualization, the Integrative Genomic Viewer, and correlation analysis, all centered around genes, microbes, or end-motifs. The primary objective of cfOmics is to assist researchers in the field of liquid biopsy by providing comprehensive multi-omics data. This enables them to explore cell-free data and extract profound insights that can significantly impact disease diagnosis, treatment monitoring, and management.

Circulating tumour DNA testing in metastatic breast cancer: Integration with tissue testing.

Breast cancer biomarker profiling predominantly relies on tissue testing (surgical and/or biopsy samples). However, the field of liquid biopsy, particularly the analysis of circulating tumour DNA (ctDNA), has witnessed remarkable progress and continues to evolve rapidly. The incorporation of ctDNA-based testing into clinical practice is creating new opportunities for patients with metastatic breast cancer (MBC). ctDNA offers advantages over conventional tissue analyses, as it reflects tumour heterogeneity and enables multiple serial biopsies in a minimally invasive manner. Thus, it serves as a valuable complement to standard tumour tissues and, in certain instances, may even present a potential alternative approach. In the context of MBC, ctDNA testing proves highly informative in the detection of disease progression, monitoring treatment response, assessing actionable biomarkers, and identifying mechanisms of resistance. Nevertheless, ctDNA does exhibit inherent limitations, including its generally low abundance, necessitating timely blood samplings and rigorous management of the pre-analytical phase. The development of highly sensitive assays and robust bioinformatic tools has paved the way for reliable ctDNA analyses. The time has now come to establish how ctDNA and tissue analyses can be effectively integrated into the diagnostic workflow of MBC to provide patients with the most comprehensive and accurate profiling. In this manuscript, we comprehensively analyse the current methodologies employed in ctDNA analysis and explore the potential benefits arising from the integration of tissue and ctDNA testing for patients diagnosed with MBC.

Improved Characterization of Circulating Tumor Cells and Cancer-Associated Fibroblasts in One-Tube Assay in Breast Cancer Patients Using Imaging Flow Cytometry.

Circulating tumor cells (CTCs) and circulating cancer-associated fibroblasts (cCAFs) have been individually considered strong indicators of cancer progression. However, technical limitations have prevented their simultaneous analysis in the context of CTC phenotypes different from epithelial. This study aimed to analyze CTCs and cCAFs simultaneously in the peripheral blood of 210 breast cancer patients using DAPI/pan-keratin (K)/vimentin (V)/alpha-SMA/CD29/CD45/CD31 immunofluorescent staining and novel technology-imaging flow cytometry (imFC). Single and clustered CTCs of different sizes and phenotypes (i.e., epithelial phenotype K+/V- and epithelial-mesenchymal transition (EMT)-related CTCs, such as K+/V+, K-/V+, and K-/V-) were detected in 27.6% of the samples and correlated with metastases. EMT-related CTCs interacted more frequently with normal cells and tended to occur in patients with tumors progressing during therapy, while cCAFs coincided with CTCs (mainly K+/V- and K-/V-) in seven (3.3%) patients and seemed to correlate with the presence of metastases, particularly visceral ones. This study emphasizes the advantages of imFC in the field of liquid biopsy and highlights the importance of multimarker-based analysis of different subpopulations and phenotypes of cancer progression-related cells, i.e., CTCs and cCAFs. The co-detection of CTCs and cCAFs might improve the identification of patients at higher risk of progression and their monitoring during therapy.

Current status of molecular diagnostic approaches using liquid biopsy

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal cancers, and developing an efficient and reliable approach for its early-stage diagnosis is urgently needed. Precancerous lesions of PDAC, such as pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasms (IPMN), arise through multiple steps of driver gene alterations in KRAS, TP53, CDKN2A, SMAD4, or GNAS. Hallmark mutations play a role in tumor initiation and progression, and their detection in bodily fluids is crucial for diagnosis. Recently, liquid biopsy has gained attention as an approach to complement pathological diagnosis, and in addition to mutation signatures in cell-free DNA, cell-free RNA, and extracellular vesicles have been investigated as potential diagnostic and prognostic markers. Integrating such molecular information to revise the diagnostic criteria for pancreatic cancer can enable a better understanding of the pathogenesis underlying inter-patient heterogeneity, such as sensitivity to chemotherapy and disease outcomes. This review discusses the current diagnostic approaches and clinical applications of genetic analysis in pancreatic cancer and diagnostic attempts by liquid biopsy and molecular analyses using pancreatic juice, duodenal fluid, and blood samples. Emerging knowledge in the rapidly advancing liquid biopsy field is promising for molecular profiling and diagnosing pancreatic diseases with significant diversity.

Liquid biopsy – An advance diagnostic tool for tracking of tumor cells

An eminent tool is required for easy detection of disease aggressiveness, identification, and prognosis of treatment. This shows the importance of a non-invasive procedure called liquid biopsy. Liquid biopsy is a liquid biomarker that identify the tumor markers from human body fluids. The small amount of tissue obtained by needle biopsies may not represent the most aggressive subclones and some remote sites cannot be reached out by needles. Here is a review that will describe and compare and contrast sources such as circulating tumor cells, cell-free DNA, exosomes role in the liquid biopsy field.

Monte Carlo simulation of micro-x-ray fluorescence (μ-XRF) based detection of immunomagnetically labeled tumor cells

Objective. Circulating tumor cells (CTCs) carry crucial information related to the spreading and proliferation of tumors, especially at early stages of the disease. Despite the huge clinical potential held by CTCs in cancer therapy, capture and detection of these cells from the patient's peripheral blood system is rather challenging since CTCs are extremely rare cells. The objective of this paper is, based on Monte Carlo simulations, to propose the detection of immunomagnetically labelled tumor cells by micro-x-ray fluorescence (μ-XRF). Approach. The simulations were carried out with the Monte Carlo N-Particle, version 6.2, (MCNP6.2) code. The model simulates 20 μm cancer cell lines and 10 μm CTCs tagged with Fe3O4@SiO2 spherical nanoparticles of diameters 25 nm, 60 nm and 110 nm. A 17.5 keV monochromatic, micro-focused x-ray beam of diameter 15 μm, impinges on cancer cells immersed in a phosphate-buffered saline solution. The simulations also include a polymeric sample holder and a silicon drift detector with a beryllium window and silver collimator. Main results. The results show the dependence of the signal intensity (Fe Kα line) on cell and nanoparticle sizes. Samples containing two and three CTCs were also simulated in particular geometrical configurations. It is presented how the inter-cell distances and cell positions relative to the incident x-ray beam affect the signal. In addition, within the parameters used in the simulations, μ-XRF method provides a minimum detection limit of 9.4 pg of Fe, which corresponds to detecting a single 10 μm CTC labeled with 110 nm Fe3O4@SiO2 nanoparticles at 6.3% binding. Significance. The μ-XRF based method proposed in this paper for detecting CTCs, combined with immunomagnetic nanoparticles (NPs), has the potential to be innovative in the field of liquid biopsy.

Abstract 6602: Refining liquid biopsy: generating more information from cell free DNA

Abstract Liquid biopsy for profiling of cell free DNA (cfDNA) in blood holds huge promise to transform how we experience and manage cancer by early detection and identification of residual disease and subtype. However, a standard blood draw yields an average of only 10 ng of cfDNA, of which DNA derived from the tumor is a small minority. Therefore, we are faced with a dilemma when utilizing the limited sample to obtain maximum information. Genetic sequencing provides information on actionable somatic mutations but detection of a few loci in a minority of the sample is challenging. Modified cytosine profiles of cancer are differential from non-cancer at many more loci and so provide a stronger signal. Moreover, they can be used to distinguish tissue-of-origin of the tumor. However, methods such as bisulfite sequencing, EM-seq and TAPS sacrifice genetic information (namely C-&amp;gt;T mutations, which are the most common mutation in cancer) to measure modified cytosine. Genetic and modified cytosine data together have been shown to be more powerful for the detection of early cancer than either alone. Here we present a technology, which sequences at base resolution the complete genetic sequence integrated with modified cytosine from low nanogram amounts of cfDNA. It consists of (i) a single pre-sequencing workflow, which creates a copy of the original DNA and performs enzymatic base conversions which discriminate genetic and epigenetic states and (ii) post-sequencing data processing which resolves the resultant sequencing data to an information-rich 16-state code and derives genetic variants integrated with modified cytosine levels, within easy-to-use software. It is, in principle, compatible with any sequencing methodology and is shown here optimized for the Illumina fleet. In cfDNA we show accuracy of modC measurement is higher than that of bisulfite sequencing and EM-seq; and the accuracy of genetic sequencing is higher than that of Illumina alone. Accuracy of measurement is extremely important for liquid biopsy where tumor DNA is in the minority and observed in a small number of reads. We demonstrate the impact of varying base calling error rate on limit of detection for rare alleles in cfDNA samples. Further we show derivation of cfDNA fragment characteristics and that this is produced in combination with genetic and modC information. This further increases the signal available from cfDNA in only one workflow. We suggest this method will help advance the field of liquid biopsy towards its promise. Citation Format: Fabio Puddu, Casper K. Lumby, Nick Harding, David J. Morley, Jamie Scotcher, Robert Crawford, Jens Füllgrabe, Walraj S. Gosal, Shirong Yu, Daniel Brudzewsky, Jane Haywood, Andrada Tomoni, Philippa Burns, Joanna D. Holbrook, Paidi Creed. Refining liquid biopsy: generating more information from cell free DNA [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6602.

Abstract 780: Digital PCR-based EGFR T790M mutation detection from human blood samples collected in PAXgene Blood ccfDNA Tubes

Abstract Introduction: Circulating cell-free DNA (ccfDNA) is used as an analyte in liquid biopsy-based cancer research with promising applications in the fields of cancer screening, therapeutic decision making and monitoring and minimal residual disease control. However, despite the multitude of future applications, challenges regarding the sensitivity of analytical tests remain, thereby limiting the routine use of ccfDNA within the above mentioned fields. In this study, we investigated the compatibility of the PAXgene® Blood ccfDNA Tube and the QIAcuity® Digital PCR System as an optimal “sample to insight” workflow with regard to the detection of EGFR T790M mutations spiked into apparently healthy human donor blood specimens. Methods: Blood from 22 apparently healthy consented donors was collected into PAXgene Blood ccfDNA Tubes* (PreAnalytiX). Within 2 h of blood draw, blood samples were either spiked with an EGFR-Multiplex 5% AF cfDNA standard (SensID) containing an EGFR T790M mutation or left unspiked as a control. Tubes were incubated for 3 d at room temperature to simulate transport over a weekend and plasma was generated by double centrifugation. Plasma pools were generated from spiked and unspiked samples and ccfDNA was isolated with the QIAamp® Circulating Nucleic Acid Kit (QIAGEN). CcfDNA quantity was measured by Qubit® dsDNA HS Assay (Thermo Fisher Scientific) and fragment size distribution was determined by TapeStation® Cell-free DNA ScreenTape® (Agilent Technologies). EGFR T790M mutation detection was performed using the dPCR Mutation Assay EGFR 6240 Human (QIAGEN) with a 26K 24-well Nanoplate (QIAGEN) on the QIAcuity Digital PCR System (QIAGEN). Data analysis was conducted using the QIAcuity Software Suite (QIsAGEN). Results: CcfDNA isolated from PAXgene Blood ccfDNA Tubes had a fragment size distribution profile with distinct mononucleosomal peaks at ~180 bps without high-molecular weight DNA, indicating successful nucleated cell stabilization over the simulated transport duration and extraction of high-quality ccfDNA. Digital PCR analysis of extracted ccfDNA revealed positive detection of EGFR T790M mutations in spiked blood samples at a relative mutation fraction of approximately 2% in the investigated samples. Conclusion: PAXgene Blood ccfDNA Tubes enabled high-quality ccfDNA isolation by stabilizing nucleated cells during simulated transport condition. Spiked EGFR T790M mutations were successfully detected by dPCR following sample storage. This proof of principle study shows the combined usability of both systems for mutation detection in human blood specimens, highlighting the potential of both technologies for a broader application in the liquid biopsy field to detect oncogenic driver as well as tumor suppressor gene mutations in a multitude of tumor types. *The PAXgene Blood ccfDNA Tube is for Research Use Only in the US. Not for use in diagnostic procedures. Citation Format: Christian Neander, Andrea Huxhold, Maike Schoenborn, Thorsten Voss. Digital PCR-based EGFR T790M mutation detection from human blood samples collected in PAXgene Blood ccfDNA Tubes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 780.

Extracellular urinary microRNAs as non-invasive biomarkers of endometrial and ovarian cancer.

Gynecological cancers account for a large number of cancer-related deaths in women. Endometrial cancer is the most prevalent, while ovarian cancer is the deadliest gynecological cancer worldwide. To overcome the clinical need for easy and rapid testing, there is a growing interest in cancer detection in non-invasive modalities. With a growing field of liquid biopsy, urine became interesting source of cancer biomarkers. The aim of this manuscript is to provide an overview on the origin, analysis and the clinical significance of urine microRNAs in gynecological cancers, with afocus on ovarian and endometrial cancer.MicroRNAs, a class of small non-coding nucleic acids, are emerging as a non-invasive biomarkers due to the feasibility and the extreme stability in body fluids. Specific miRNA expression signatures have been previously identified in ovarian and endometrial cancer. The aim of this manuscript is to provide an overview on the origin, analysis and the clinical significance of urine microRNAs in gynecological cancers, with the focus on ovarian and endometrial cancer. CONCLUSION: The advantages and limitations of urine microRNA utility and technologies are discussed. Previously detected microRNA from urine of the patients are summarized to evaluate their potential as non-invasive clinical biomarkers in gynecological oncology.

Research on liquid biopsy for cancer: A bibliometric analysis

Background: In recent years, liquid biopsy has shown great potential for improving cancer diagnosis and treatment. This study aimed to explore the trends and prospects in liquid biopsy for cancer from a bibliometric perspective.Methods: Reviews and articles on liquid biopsy and cancer were collected from the Web of Science Core Collection (WoSCC). Key bibliometric characteristics were analyzed using CiteSpace. Co-occurrence analysis of keywords and co-citation analysis of references was performed.Results: A total of 6331 publications from 11 years of scientific research were retrieved. Ninety-five countries and 7004 institutions in liquid biopsy and cancer contributed. The United States (US) and China published the most articles. The institution with the most published articles was the University of Texas MD Anderson Cancer Center. The most published journals were Cancer and Frontiers in Oncology. “Bettegowda (2014)" was the most cited reference with the highest burst strength in the last decade. Cluster analysis revealed that the recent hot topics were “circulating tumor cells,” “cancer,” and “exosomes."Conclusions: This bibliometric analysis maps the basic knowledge structure of the field of liquid biopsy for cancer. The field is entering a phase of rapid development. The hot spots identified in this study deserve further investigation.

Toward Personalized Nanomedicine: The Critical Evaluation of Micro and Nanodevices for Cancer Biomarker Analysis in Liquid Biopsy.

Liquid biopsy for the analysis of circulating cancer biomarkers (CBs) is a major advancement toward the early detection of cancer. In comparison to tissue biopsy techniques, liquid biopsy is relatively painless, offering multiple sampling opportunities across easily accessible bodily fluids such as blood, urine, and saliva. Liquid biopsy is also relatively inexpensive and simple, avoiding the requirement for specialized laboratory equipment or trained medical staff. Major advances in the field of liquid biopsy are attributed largely to developments in nanotechnology and microfabrication that enables the creation of highly precise chip-based platforms. These devices can overcome detection limitations of an individual biomarker by detecting multiple markers simultaneously on the same chip, or by featuring integrated and combined target separation techniques. In this review, the major advances in the field of portable and semi-portable micro, nano, and multiplexed platforms for CB detection for the early diagnosis of cancer are highlighted. A comparative discussion is also provided, noting merits and drawbacks of the platforms, especially in terms of portability. Finally, key challenges toward device portability and possible solutions, as well as discussing the future direction of the field are highlighted.

Enzymatic Methods for Mutation Detection in Cancer Samples and Liquid Biopsies.

Low-level tumor somatic DNA mutations in tissue and liquid biopsies obtained from cancer patients can have profound implications for development of metastasis, prognosis, choice of treatment, follow-up, or early cancer detection. Unless detected, such low-frequency DNA alterations can misinform patient management decisions or become missed opportunities for personalized medicine. Next-generation sequencing technologies and digital-PCR can resolve low-level mutations but require access to specialized instrumentation, time, and resources. Enzymatic-based approaches to detection of low-level mutations provide a simple, straightforward, and affordable alternative to enrich and detect such alterations and is broadly available to low-resource laboratory settings. This review summarizes the traditional uses of enzymatic mutation detection and describes the latest exciting developments, potential, and applications with specific reference to the field of liquid biopsy in cancer.

A label-free fluorescent sensor for rapid and sensitive detection of ctDNA based on fluorescent PDA nanoparticles.

Technological advances in the detection of circulating tumor DNA (ctDNA) have made new options available for diagnosis, classification, biological studies, and treatment selection. However, effective and practical methods for analyzing this emerging class of biomarkers are still lacking. In this work, a fluorescent biosensor was designed for the label-free detection of ctDNA (EGFR 19 del for non-small cell lung cancer, NSCLC). The biosensor was based on the fact that MnO2 nanosheets (MnO2 NSs) have stronger affinity towards single-stranded DNA (ssDNA), as compared with double-stranded DNA (dsDNA). As a high-performance nanoenzyme, MnO2 NSs could oxidize dopamine (DA) into fluorescent polydopamine nanoparticles (FL-PDA NPs), which could be used as a fluorescence signal. The probe ssDNA could be adsorbed on the surface of MnO2 NSs through π-π stacking, and the active site would be masked, causing a lower fluorescence. After the targets were recognized by probe ssDNA to form dsDNA, its affinity for MnO2 NSs decreased and the active site recovered, causing a restored fluorescence. It was verified that Mn ions, •OH radicals and electron transfer were the important factors in the catalytic oxidation of DA. Under the optimal experimental conditions, this biosensor exhibited a detection limit of 380 pM and a linear range of 25-125 nM, providing reliable readout in a short time (45 min). This sensor exhibited outstanding specificity, stability and reproducibility. In addition, this sensor was applied to the detection of ctDNA in serum samples and cell lysates. It is demonstrated that FL-PDA NPs can be used as a fluorescence signal for easy, rapid and label-free detection of ctDNA without any other amplification strategies, and the proposed strategy has great potential for biomarker detection in the field of liquid biopsy.

Multiomic Approaches for Cancer Biomarker Discovery in Liquid Biopsies: Advances and Challenges.

Cancer is a complex and heterogeneous disease that poses a significant threat to global health. Early diagnosis and treatment are critical for improving patient outcomes, and the use of liquid biopsies has emerged as a promising approach for cancer detection and monitoring. Traditionally, cancer diagnosis has relied on invasive tissue biopsies, the collection of which can prove challenging for patients and the results of which may not always provide accurate results due to tumor heterogeneity. Liquid biopsies have gained increasing attention as they provide a non-invasive and accessible source of cancer biomarkers, which can be used to diagnose cancer, monitor treatment response, and detect relapse. The integration of -omics technologies, such as proteomics, genomics, and metabolomics, has further enhanced the capabilities of liquid biopsies by introducing precision oncology and enabling the tailoring of treatment for individual patients based on their unique tumor biology. In this review, we will discuss the challenges and advances in the field of cancer liquid biopsies and the integration of -omics technologies for different types of liquid biopsies, including blood, tear, urine, sweat, saliva, and cerebrospinal fluid.

Current concepts of anti-EGFR targeting in metastatic colorectal cancer.

Anti-EGFR targeting is one of the key strategies in the treatment of metastatic colorectal cancer (mCRC). For almost two decades oncologists have struggled to implement EGFR antibodies in the mCRC continuum of care. Both sidedness and RAS mutational status rank high among the predictive factors for the clinical efficacy of EGFR inhibitors. A prospective phase III trial has recently confirmed that anti-EGFR targeting confers an overall survival benefit only in left sided RAS-wildtype tumors when given in first line. It is a matter of discussion if more clinical benefit can be reached by considering putative primary resistance mechanisms (e.g., HER2, BRAF, PIK3CA, etc.) at this early stage of treatment. The value of this procedure in daily routine clinical utility has not yet been clearly delineated. Re-exposure to EGFR antibodies becomes increasingly crucial in the disease journey of mCRC. Yet re- induction or re-challenge strategies have been problematic as they relied on mathematical models that described the timely decay of EGFR antibody resistant clones. The advent of liquid biopsy and the implementation of more accurate next-generation sequencing (NGS) based high throughput methods allows for tracing of EGFR resistant clones in real time. These displays the spatiotemporal heterogeneity of metastatic disease compared to the former standard radiographic assessment and re-biopsy. These techniques may move EGFR inhibition in mCRC into the area of precision medicine in order to apply EGFR antibodies with the increase or decrease of EGFR resistant clones. This review critically discusses established concepts of tackling the EGFR pathway in mCRC and provides insight into the growing field of liquid biopsy guided personalized approaches of EGFR inhibition in mCRC.

BLOODPAC: Collaborating to chart a path towards blood-based screening for early cancer detection.

BLOODPAC: Collaborating to chart a path towards blood-based screening for early cancer detection.

Sensing red blood cell nano-mechanics: Toward a novel blood biomarker for Alzheimer's disease.

Red blood cells (RBCs) are characterized by a remarkable elasticity, which allows them to undergo very large deformation when passing through small vessels and capillaries. This extreme deformability is altered in various clinical conditions, suggesting that the analysis of red blood cell (RBC) mechanics has potential applications in the search for non-invasive and cost-effective blood biomarkers. Here, we provide a comparative study of the mechanical response of RBCs in patients with Alzheimer’s disease (AD) and healthy subjects. For this purpose, RBC viscoelastic response was investigated using atomic force microscopy (AFM) in the force spectroscopy mode. Two types of analyses were performed: (i) a conventional analysis of AFM force–distance (FD) curves, which allowed us to retrieve the apparent Young’s modulus, E; and (ii) a more in-depth analysis of time-dependent relaxation curves in the framework of the standard linear solid (SLS) model, which allowed us to estimate cell viscosity and elasticity, independently. Our data demonstrate that, while conventional analysis of AFM FD curves fails in distinguishing the two groups, the mechanical parameters obtained with the SLS model show a very good classification ability. The diagnostic performance of mechanical parameters was assessed using receiving operator characteristic (ROC) curves, showing very large areas under the curves (AUC) for selected biomarkers (AUC > 0.9). Taken all together, the data presented here demonstrate that RBC mechanics are significantly altered in AD, also highlighting the key role played by viscous forces. These RBC abnormalities in AD, which include both a modified elasticity and viscosity, could be considered a potential source of plasmatic biomarkers in the field of liquid biopsy to be used in combination with more established indicators of the pathology.

A light-activated magnetic bead strategy utilized in spatio-temporal controllable exosomes isolation.

Tumor-derived exosomes are considered as a key biomarker in the field of liquid biopsy. However, conventional separation techniques such as ultracentrifugation, co-precipitation and column chromatography cannot isolate samples with high throughput, while traditional immunomagnetic separation techniques, due to steric effect of magnetic beads, reducing sensitivity of exosomes optical detection. Herein, we provide a novel and simple nanoplatform for spatiotemporally controlling extraction and elution of exosomes via magnetic separation and light-activated cargo release. In this system, magnetic beads are co-modified by photoresponsive groups -nitrobenzyl group and aptamers that are compatible with CD63-a highly expressed exosomal surface-specific protein. Through exosomes extracted from cell model and nude mice xenograft tumor model morphological characterization and proteomic analysis, results showed that our novel magnetic bead system outperformed current ultracentrifugation in serum exosome extraction in terms of extraction time, yield, and proportion of populations with high CD63 expression. This strategy may be a powerful tool for exosome isolation in clinical liquid biopsies of cancer disease.