Targeted Sequencing of Plasma-Derived vs. Urinary cfDNA from Patients with Triple-Negative Breast Cancer

Herzog, Henrike; Dogan, Senol; Aktas, Bahriye; Nel, Ivonne

doi:10.3390/cancers14174101

Cited by 6 publications

(6 citation statements)

References 76 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Genomics-based analyses of urinary ctDNA have emphasized the somatic mutations linked to the primary tumors of BC patients, such as the PIK3CA and TP53 mutated genes, offering the non-invasive probing and real-time monitoring of BC relapse [ 164 ]. Based on cfDNA-seq, NF1 , CHEK2 , KMT2C, and PTEN were found to be the most frequently occurring mutated genes in the plasma and urine of BC patients [ 165 ].…”

Section: Breast Cancer Investigation In the Multi-omics Eramentioning

confidence: 99%

Omics-Based Investigations of Breast Cancer

Neagu,

Whitham,

Bruno

et al. 2023

Molecules

View full text Add to dashboard Cite

Breast cancer (BC) is characterized by an extensive genotypic and phenotypic heterogeneity. In-depth investigations into the molecular bases of BC phenotypes, carcinogenesis, progression, and metastasis are necessary for accurate diagnoses, prognoses, and therapy assessments in predictive, precision, and personalized oncology. This review discusses both classic as well as several novel omics fields that are involved or should be used in modern BC investigations, which may be integrated as a holistic term, onco-breastomics. Rapid and recent advances in molecular profiling strategies and analytical techniques based on high-throughput sequencing and mass spectrometry (MS) development have generated large-scale multi-omics datasets, mainly emerging from the three ”big omics”, based on the central dogma of molecular biology: genomics, transcriptomics, and proteomics. Metabolomics-based approaches also reflect the dynamic response of BC cells to genetic modifications. Interactomics promotes a holistic view in BC research by constructing and characterizing protein–protein interaction (PPI) networks that provide a novel hypothesis for the pathophysiological processes involved in BC progression and subtyping. The emergence of new omics- and epiomics-based multidimensional approaches provide opportunities to gain insights into BC heterogeneity and its underlying mechanisms. The three main epiomics fields (epigenomics, epitranscriptomics, and epiproteomics) are focused on the epigenetic DNA changes, RNAs modifications, and posttranslational modifications (PTMs) affecting protein functions for an in-depth understanding of cancer cell proliferation, migration, and invasion. Novel omics fields, such as epichaperomics or epimetabolomics, could investigate the modifications in the interactome induced by stressors and provide PPI changes, as well as in metabolites, as drivers of BC-causing phenotypes. Over the last years, several proteomics-derived omics, such as matrisomics, exosomics, secretomics, kinomics, phosphoproteomics, or immunomics, provided valuable data for a deep understanding of dysregulated pathways in BC cells and their tumor microenvironment (TME) or tumor immune microenvironment (TIMW). Most of these omics datasets are still assessed individually using distinct approches and do not generate the desired and expected global-integrative knowledge with applications in clinical diagnostics. However, several hyphenated omics approaches, such as proteo-genomics, proteo-transcriptomics, and phosphoproteomics-exosomics are useful for the identification of putative BC biomarkers and therapeutic targets. To develop non-invasive diagnostic tests and to discover new biomarkers for BC, classic and novel omics-based strategies allow for significant advances in blood/plasma-based omics. Salivaomics, urinomics, and milkomics appear as integrative omics that may develop a high potential for early and non-invasive diagnoses in BC. Thus, the analysis of the tumor circulome is considered a novel frontier in liquid biopsy. Omics-based investigations have applications in BC modeling, as well as accurate BC classification and subtype characterization. The future in omics-based investigations of BC may be also focused on multi-omics single-cell analyses.

show abstract

Section: Breast Cancer Investigation In the Multi-omics Eramentioning

confidence: 99%

Omics-Based Investigations of Breast Cancer

Neagu,

Whitham,

Bruno

et al. 2023

Molecules

View full text Add to dashboard Cite

show abstract

“…Herzong et al suggested urine as an alternative liquid biopsy fluid from which to extract ctDNAs for TNBC clinical analysis. They compared the performances of ctDNA collection and characterization in both urine and plasma samples from 15 presurgical TNBC patients [ 57 ]. The analysis was performed by amplification of a coding region of 93 genes, known to be related to breast cancer, and their sequencing by Illumina NGS (NextSeqv2.5High Output 300 bp cassette).…”

Section: Liquid Biopsy and Circulating Cell-free Nucleic Acids In Tnbcmentioning

confidence: 99%

“…Bioinformatical analysis has revealed 431 somatic breast cancer-related variants shared in both fluids. Among these, the most common pathogenic and probable pathogenic mutations are NF1, CHEK2, KMT2C, and PTEN, which are tumor suppressor genes commonly implicated in TNBC onset and progression [ 43 , 44 , 45 , 46 , 47 , 48 , 57 ]. These results proved that both plasma and urine-derived ctDNA from TNBC patients could be efficiently analyzed using a targeted sequencing approach.…”

Section: Liquid Biopsy and Circulating Cell-free Nucleic Acids In Tnbcmentioning

confidence: 99%

An Overview of Circulating Cell-Free Nucleic Acids in Diagnosis and Prognosis of Triple-Negative Breast Cancer

Tierno

Grassi

Zanconati

et al. 2023

IJMS

View full text Add to dashboard Cite

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer due to its molecular heterogeneity and poor clinical outcomes. Analysis of circulating cell-free tumor nucleic acids (ctNAs) can improve our understanding of TNBC and provide efficient and non-invasive clinical biomarkers that may be representative of tumor heterogeneity. In this review, we summarize the potential of ctNAs to aid TNBC diagnosis and prognosis. For example, tumor fraction of circulating cell-free DNA (TFx) may be useful for molecular prognosis of TNBC: high TFx levels after neoadjuvant chemotherapy have been associated with shorter progression-free survival and relapse-free survival. Mutations and copy number variations of TP53 and PIK3CA/AKT genes in plasma may be important markers of TNBC onset, progression, metastasis, and for clinical follow-up. In contrast, the expression profile of circulating cell-free tumor non-coding RNAs (ctncRNAs) can be predictive of molecular subtypes of breast cancer and thus aid in the identification of TBNC. Finally, dysregulation of some circulating cell-free tumor miRNAs (miR17, miR19a, miR19b, miR25, miR93, miR105, miR199a) may have a predictive value for chemotherapy resistance. In conclusion, a growing number of efforts are highlighting the potential of ctNAs for future clinical applications in the diagnosis, prognosis, and follow-up of TNBC.

show abstract

“…CtDNA only represents a small part (0.1-10%) of the total cfDNA [10]. Still, cfDNA levels of cancer patients were reported to be increased compared to healthy individuals [11]. Increased cfDNA concentrations are not only cancer-specific but can also appear in other diseases or during pregnancy, physical exercise and physiological stress; thus, measuring the concentration alone is not sufficient.…”

Section: Introductionmentioning

confidence: 99%

“…In urine, the half-life time of cfDNA is shorter than in plasma due to high levels of nucleases like DNase I and DNase II, as well as contaminants present in urine [9,10]. In previous studies, we were able to show that targeted sequencing of ucfDNA from patients with breast cancer was feasible without using stabilizing buffers when samples were processed within 2-4 h after collection [11,21]. However, the cfDNA yield from 10 mL of urine appeared to be low.…”

Section: Introductionmentioning

confidence: 99%

The Challenge to Stabilize, Extract and Analyze Urinary Cell-Free DNA (ucfDNA) during Clinical Routine

Nel,

Münch,

Shamkeeva

et al. 2023

Diagnostics

Self Cite

View full text Add to dashboard Cite

Background: The “Liquid Biopsy” has become a powerful tool for cancer research during the last decade. Circulating cell-free DNA (cfDNA) that originates from tumors has emerged as one of the most promising analytes. In contrast to plasma-derived cfDNA, only a few studies have investigated urinary cfDNA. One reason might be rapid degradation and hence inadequate concentrations for downstream analysis. In this study, we examined the stability of cfDNA in urine using different methods of preservation under various storage conditions. Methodology: To mimic patient samples, a pool of healthy male and female urine donors was spiked with a synthetic cfDNA reference standard (fragment size 170 bp) containing the T790M mutation in the EGFR gene. Spiked samples were preserved with three different buffers and with no buffer over four different storage periods (0 h; 4 h; 12 h; 24 h) at room temperature vs. 4 °C. The preservatives used were Urinary Analyte Stabilizer (UAS, Novosanis, Wijnegem, Belgium), Urine Conditioning Buffer (UCB, Zymo, Freiburg, Germany) and a self-prepared buffer called “AlloU”. CfDNA was extracted using the QIAamp MinElute ccfDNA Mini Kit (Qiagen, Hilden, Germany). CfDNA concentration was measured using the Qubit™ 4 fluorometer (Thermo Fisher Scientific, Waltham, MA, USA). Droplet digital PCR (ddPCR) was used for detection and quantification of the T790M mutation. Results: Almost no spiked cfDNA was recoverable from samples with no preservation buffer and the T790M variant was not detectable in these samples. These findings indicate that cfDNA was degraded below the detection limit by urinary nucleases. Stabilizing buffers showed varying efficiency in preventing this degradation. The most effective stabilizing buffer under all storage conditions was the UAS, enabling adequate recovery of the T790M variant using ddPCR. Conclusion: From a technical point of view, stabilizing buffers and adequate storage conditions are a prerequisite for translation of urinary cfDNA diagnostics into clinical routine.

show abstract

Targeted Sequencing of Plasma-Derived vs. Urinary cfDNA from Patients with Triple-Negative Breast Cancer

Cited by 6 publications

References 76 publications

Omics-Based Investigations of Breast Cancer

Omics-Based Investigations of Breast Cancer

An Overview of Circulating Cell-Free Nucleic Acids in Diagnosis and Prognosis of Triple-Negative Breast Cancer

The Challenge to Stabilize, Extract and Analyze Urinary Cell-Free DNA (ucfDNA) during Clinical Routine

Contact Info

Product

Resources

About