Methodological development and biological observations of cell free DNA with a simple direct fluorescent assay in colorectal cancer

Boysen, Anders; Sørensen, Brita Singers; Lefèvre, Anna Cecilie; Abrantes, Rafaela; Johansen, Julia S.; Jensen, Benny Vittrup; Schou, Jesper Sølver; Larsen, Finn Ole; Nielsen, Dorte; Taflin, Helena; Gustavson, Bengt; Wettergren, Yvonne; Ree, Anne Hansen; Dueland, Svein; Pallisgaard, Niels; Spindler, Karen Lise Garm

doi:10.1016/j.cca.2018.09.029

Cited by 12 publications

(14 citation statements)

References 15 publications

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“…For comparison, blood samples were available from 94 fully anonymized presumed healthy individuals from the Lolland-Falster Health Study (NCT02482896), as previously described (13). All healthy donors gave written informed consent and the study was approved by Region Zealand's Ethical Committee on Health Research (approval no.…”

Section: Methodsmentioning

confidence: 99%

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Total cell‑free DNA measurement in metastatic colorectal cancer with a fast and easy direct fluorescent assay

et al. 2022

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Treatment for metastatic colorectal cancer (mCRC) is focused on prolonging survival and maintaining quality of life. It is important to establish prognostic and predictive markers to avoid extended, ineffective treatment. The aim of the present study was, by a novel approach, to analyze the association between cell-free (cf)DNA levels and outcome in patients receiving systemic therapy for incurable mCRC. The study was a prospective non-interventional biomarker study for patients receiving standard of systemic treatment for mCRC. Patients with mCRC, who, according to standard guidelines, were considered for treatment with EGFR inhibitors, were included. The cfDNA levels in consecutive plasma samples were measured by a direct fluorescence assay. The study included 47 patients. Blood samples were available at baseline (n=47); prior to the third treatment cycle (n=31); the first (n=33), second (n=22) and third response evaluation during treatment (n=17); and at progression (n=22). The disease control rate was 42 and 91% in patients with high (≥75th percentile of baseline cfDNA levels) and low cfDNA levels (<75th percentile of baseline cfDNA levels), respectively (P<0.001). Median progression-free survival (PFS) was 3.8 and 8.5 months in patients with high and low cfDNA levels, respectively (hazard ratio=3.03, 95% CI 1.46-6.29, P<0.01). Median overall survival (OS) was 5.0 and 26.6 months in patients with high and low cfDNA levels, respectively (hazard ratio=3.48, 95% CI 1.44-8.44, P<0.01). In the multivariate analysis, baseline cfDNA levels remained a significant predictor of PFS and OS. In conclusion, cfDNA is a promising prognostic tool in the personalized treatment of mCRC. cfDNA levels were estimated by a simple, rapid and inexpensive method (OPTIPAL II: ClinicalTrials.gov identifier no. NCT03750175; registered November 21, 2018).

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Section: Methodsmentioning

confidence: 99%

“…Direct fluorescent assay (DFA), first described by Goldshtein et al (12) in 2009 and refined by Boysen et al (13) in 2018, is able to quantify cfDNA in plasma or serum. This method is simple, rapid and inexpensive.…”

Section: Introductionmentioning

confidence: 99%

Total cell‑free DNA measurement in metastatic colorectal cancer with a fast and easy direct fluorescent assay

et al. 2022

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“…The clinical data and outcome of the study has been presented separately (13), thus only translational aspects are addressed here. For comparison, we had blood samples available from 94 healthy individuals from a Danish biobank, as previously described (14). After HAI treatment patients underwent standard of care surveillance with regular Computed Tomography (CT) scans.…”

Section: Methodsmentioning

confidence: 99%

“…Blood samples for translational use were drawn at baseline prior to the administration of the first intrahepatic dose of oxaliplatin and at a fixed schedule during follow up. Analysis of cfDNA was done using a direct fluorescent assay for cfDNA analysis, not requiring any prior processing, as preliminary reported by Douvdevani and colleagues (15,16) and further modified by our group, as previously published (14).…”

Section: Methodsmentioning

confidence: 99%

Prognostic and predictive value of circulating DNA for hepatic arterial infusion of chemotherapy for patients with colorectal cancer liver metastases

et al. 2020

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Hepatic arterial infusion (HAI) of chemotherapy is an experimental treatment option for patients with colorectal cancer liver metastases (CRCLM). The current study aimed to investigate the predictive and prognostic value of cell free DNA (cfDNA) in patients with CRCLM receiving HAI with oxaliplatin and systemic capecitabine. Plasma samples from 62 patients were investigated who were included into a single arm phase II study investigating HAI treatment for patients with CRCLM. The clinical outcome of the trial has been presented previously. In brief, treatment consisted of intrahepatic infusion of oxaliplatin 100 mg/m 2 every second week with concomitant oral capecitabine 3,500 mg/m 2 every second week for up to 12 cycles. Blood samples were drawn at baseline and follow-up and plasma was analyzed for cell free DNA using a direct fluorescent assay. The baseline level of plasma cfDNA was 0.92 ng/µl (95% CI 0.84-1.00). Patients with a baseline value of cfDNA above the 75th quartile had a median overall survival of 2.4 years (95% CI 0.7-2.8), compared with 3.9 years (95% CI 2.8-5.9) for patients below the 75th quartile (P=0.02). The baseline level of cfDNA was significantly lower (0.91 ng/µl, 95% CI 0.76-0.98) in patients who achieved an objective response compared to non-responders (1.79 ng/µl; 95% CI 0.99-2.57; P=0.02). The current study demonstrated a possible prognostic and predictive value of cfDNA for patients with CRCLM undergoing HAI with oxaliplatin and concomitant capecitabine.

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“…Afterwards, many other studies have also evaluated the concordance between tissue and plasma KRAS mutations obtaining elevated specificity and sensitivity rates [ 102 , 110 ]. Even though many different manuscripts have observed better correlations between cfDNA concentrations and the tumor mutation burden in mCRC [ 107 , 110 , 131 ], the cfDNA analysis could be also performed in patients at earlier cancer stages [ 102 , 110 ].…”

Section: Colon and Rectal Cancersmentioning

confidence: 99%

Clinical Utility of Liquid Biopsy-Based Actionable Mutations Detected via ddPCR

et al. 2021

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Cancer is one of the leading causes of death worldwide and remains a major public health challenge. The introduction of more sensitive and powerful technologies has permitted the appearance of new tumor-specific molecular aberrations with a significant cancer management improvement. Therefore, molecular pathology profiling has become fundamental not only to guide tumor diagnosis and prognosis but also to assist with therapeutic decisions in daily practice. Although tumor biopsies continue to be mandatory in cancer diagnosis and classification, several studies have demonstrated that liquid biopsies could be used as a potential tool for the detection of cancer-specific biomarkers. One of the main advantages is that circulating free DNA (cfDNA) provides information about intra-tumoral heterogeneity, reflecting dynamic changes in tumor burden. This minimally invasive tool has become an accurate and reliable instrument for monitoring cancer genetics. However, implementing liquid biopsies across the clinical practice is still ongoing. The main challenge is to detect genomic alterations at low allele fractions. Droplet digital PCR (ddPCR) is a powerful approach that can overcome this issue due to its high sensitivity and specificity. Here we explore the real-world clinical utility of the liquid biopsy ddPCR assays in the most diagnosed cancer subtypes.

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Methodological development and biological observations of cell free DNA with a simple direct fluorescent assay in colorectal cancer

Cited by 12 publications

References 15 publications

Total cell‑free DNA measurement in metastatic colorectal cancer with a fast and easy direct fluorescent assay

Total cell‑free DNA measurement in metastatic colorectal cancer with a fast and easy direct fluorescent assay

Prognostic and predictive value of circulating DNA for hepatic arterial infusion of chemotherapy for patients with colorectal cancer liver metastases

Clinical Utility of Liquid Biopsy-Based Actionable Mutations Detected via ddPCR

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