Circulating cell free DNA levels when considered in isolation have little clinical value in patients with pancreatic ductal adenocarcinoma compared to other chronic diseases

Abstract Pancreatic cancer is often detected late, when curative therapies are no longer possible. Here, we present non-invasive detection of pancreatic ductal adenocarcinoma (PDAC) by 5-hydroxymethylcytosine (5hmC) changes in circulating cell free DNA from a PDAC cohort (n = 64) in comparison with a non-cancer cohort (n = 243). Differential hydroxymethylation is found in thousands of genes, most significantly in genes related to pancreas development or function (GATA4, GATA6, PROX1, ONECUT1, MEIS2), and cancer pathogenesis (YAP1, TEAD1, PROX1, IGF1). cfDNA hydroxymethylome in PDAC cohort is differentially enriched for genes that are commonly de-regulated in PDAC tumors upon activation of KRAS and inactivation of TP53. Regularized regression models built using 5hmC densities in genes perform with AUC of 0.92 (discovery dataset, n = 79) and 0.92–0.94 (two independent test sets, n = 228). Furthermore, tissue-derived 5hmC features can be used to classify PDAC cfDNA (AUC = 0.88). These findings suggest that 5hmC changes enable classification of PDAC even during early stage disease.

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Tumor cell-free DNA copy number instability compared to CA19-9 as an early predictor of response to systemic therapy in pancreatic ductal adenocarcinoma (PDAC).

Journal of Clinical Oncology ◽

10.1200/jco.2017.35.15_suppl.e14524 ◽

2017 ◽

Vol 35 (15_suppl) ◽

pp. e14524-e14524

Author(s):

Madappa N. Kundranda ◽

Alexander Koenig ◽

Julia Beck ◽

Kirsten Bornemann-Kolatzki ◽

Jessica Coats ◽

...

Keyword(s):

Pancreatic Ductal Adenocarcinoma ◽

Real Time ◽

Systemic Therapy ◽

Copy Number ◽

Ductal Adenocarcinoma ◽

Malignant Neoplasms ◽

Cell Free Dna ◽

Free Dna ◽

Time Assessment

e14524 Background: Humoral tumor markers are used clinically for real-time assessment of therapeutic efficacy. In pancreatic ductal adenocarcinoma (PDAC) the predominant marker is CA19-9, which is not expressed by 10 to 30% of patients depending on race. We compared plasma cell-free DNA (cfDNA) copy number based assay with changes in serum CA19-9 levels and radiological responses to predict responses to systemic therapy. Methods: In a laboratory blinded, prospective multicenter pilot study, 40 non-resectable PDAC patients, treated with (m)FOLFIRINOX, CAPIRI, or gemcitabine +/- nab-paclitaxel) are currently enrolled. CA19-9 was determined in the local center’s laboratory. Tumor cfDNA was measured with a copy-number instability (CNI) scoring assay, determined by next generation sequencing in a centralized laboratory. The CNI score assesses the amount of cfDNA with somatic macro-alterations originating from malignant neoplasms. The difference of the values before commencing therapy (baseline) and prior to cycle 2 (either rising or falling) was calculated as a predictor of standardized radiological evaluation of chemotherapeutic efficacy. Results: 37 patients (3 drop-outs) had data for baseline and cycle 2, of which CA19-9 was elevated and evaluable in 29 patients. The direction from baseline to cycle 2 of CA19-9 and CNI scores were in agreement in 18/29 patients. 9 of 11 cases with discordant CNI score and CA19-9 had treatment response data, and CNI correlated with 7/9 (78%); in contrast 7/9 had rising CA19-9, when response was stable disease or better (22% concordance). In the 27 patients with available imaging, CNI predicted better (n = 18) than CA19-9 (n = 10) (p = 0.03 Fisher’s exact). Conclusions: This comparative study on cfDNA versus CA19-9 suggest that cfDNA CNI quantitation is a potentially more reliable blood based marker for early real-time assessment of efficacy in systemic PDAC therapy than CA19-9, compared to standard of care imaging. The better prediction after the first cycle might be due to the very short in vivo half-life of cfDNA ( < 1hr) compared to about one week for CA19-9. These results justify a larger prospective validation trial.

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Detection of early stage pancreatic cancer using 5-hydroxymethylcytosine signatures in circulating cell free DNA

10.1101/422675 ◽

2018 ◽

Cited By ~ 1

Author(s):

Francois Collin ◽

Yuhong Ning ◽

Tierney Phillips ◽

Erin McCarthy ◽

Aaron Scott ◽

...

Keyword(s):

Pancreatic Cancer ◽

Early Stage ◽

Training Data ◽

Ductal Adenocarcinoma ◽

Data Sets ◽

Cell Free Dna ◽

Non Invasive ◽

Pancreatic Cancers ◽

Free Dna ◽

Circulating Cell Free Dna

AbstractPancreatic cancers are typically diagnosed at late stage where disease prognosis is poor as exemplified by a 5-year survival rate of 8.2%. Earlier diagnosis would be beneficial by enabling surgical resection or earlier application of therapeutic regimens. We investigated the detection of pancreatic ductal adenocarcinoma (PDAC) in a non-invasive manner by interrogating changes in 5-hydroxymethylation cytosine status (5hmC) of circulating cell free DNA in the plasma of a PDAC cohort (n=51) in comparison with a non-cancer cohort (n=41). We found that 5hmC sites are enriched in a disease and stage specific manner in exons, 3’UTRs and transcription termination sites. Our data show that 5hmC density is reduced in promoters and histone H3K4me3-associated sites with progressive disease suggesting increased transcriptional activity. 5hmC density is differentially represented in thousands of genes, and a stringently filtered set of the most significant genes points to biology related to pancreas (GATA4, GATA6, PROX1, ONECUT1) and/or cancer development (YAP1, TEAD1, PROX1, ONECUT1, ONECUT2, IGF1 and IGF2). Regularized regression models were built using 5hmC densities in statistically filtered genes or a comprehensive set of highly variable 5hmC counts in genes and performed with an AUC = 0.94-0.96 on training data. We were able to test the ability to classify PDAC and non-cancer samples with the Elastic net and Lasso models on two external pancreatic cancer 5hmC data sets and found validation performance to be AUC = 0.74-0.97. The findings suggest that 5hmC changes enable classification of PDAC patients with high fidelity and are worthy of further investigation on larger cohorts of patient samples.

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Circulating Cell-Free DNA-Based Liquid Biopsy Markers for the Non-Invasive Prognosis and Monitoring of Metastatic Pancreatic Cancer

Cancers ◽

10.3390/cancers12071754 ◽

2020 ◽

Vol 12 (7) ◽

pp. 1754 ◽

Cited By ~ 1

Author(s):

Marta Toledano-Fonseca ◽

M. Teresa Cano ◽

Elizabeth Inga ◽

Rosa Rodríguez-Alonso ◽

M. Auxiliadora Gómez-España ◽

...

Keyword(s):

Liquid Biopsy ◽

Hepatic Metastases ◽

Progression Free Survival ◽

The Body ◽

Ductal Adenocarcinoma ◽

Cell Free Dna ◽

Ras Mutation ◽

Non Invasive ◽

Free Dna ◽

Circulating Cell Free Dna

Liquid biopsy may assist in the management of cancer patients, which can be particularly applicable in pancreatic ductal adenocarcinoma (PDAC). In this study, we investigated the utility of circulating cell-free DNA (cfDNA)-based markers as prognostic tools in metastatic PDAC. Plasma was obtained from 61 metastatic PDAC patients, and cfDNA levels and fragmentation were determined. BEAMing technique was used for quantitative determination of RAS mutation allele fraction (MAF) in cfDNA. We found that the prognosis was more accurately predicted by RAS mutation detection in plasma than in tissue. RAS mutation status in plasma was a strong independent prognostic factor for both overall survival (OS) and progression-free survival (PFS). Moreover, RAS MAF in cfDNA was also an independent risk factor for poor OS, and was strongly associated with primary tumours in the body/tail of the pancreas and liver metastases. Higher cfDNA levels and fragmentation were also associated with poorer OS and shorter PFS, body/tail tumors, and hepatic metastases, whereas cfDNA fragmentation positively correlated with RAS MAF. Remarkably, the combination of CA19-9 with MAF, cfDNA levels and fragmentation improved the prognostic stratification of patients. Furthermore, dynamics of RAS MAF better correlated with patients’ outcome than standard CA19-9 marker. In conclusion, our study supports the use of cfDNA-based liquid biopsy markers as clinical tools for the non-invasive prognosis and monitoring of metastatic PDAC patients.

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Evaluation of cell-free DNA as a biomarker for pancreatic malignancies

The International Journal of Biological Markers ◽

10.5301/jbm.5000088 ◽

2015 ◽

Vol 30 (1) ◽

pp. 136-141 ◽

Cited By ~ 25

Author(s):

Katarzyna Sikora ◽

Chiara Bedin ◽

Caterina Vicentini ◽

Giorgio Malpeli ◽

Edoardo D'Angelo ◽

...

Keyword(s):

Pancreatic Ductal Adenocarcinoma ◽

Pancreatic Neuroendocrine Tumor ◽

Diagnostic Value ◽

Ductal Adenocarcinoma ◽

Cell Free Dna ◽

Total Load ◽

Free Dna ◽

Spearman's Rho ◽

Diagnostic Potential ◽

Spearman’S Rho

Background Currently, no reliable blood-based assay for early detection of pancreatic ductal adenocarcinoma (PDAC) is available. Cell-free DNA (cfDNA) quantitation in patients’ plasma has been recently applied in monitoring several cancer types. This study evaluates the diagnostic potential of cfDNA in PDAC patients. Methods Plasma cfDNA levels and integrity ratio were assayed using quantitative real-time PCR of Alu-repeat amplicons in patients with pancreatic ductal adenocarcinoma (n=50), pancreatic neuroendocrine tumor (n=23), and chronic pancreatitis (n=20), as well as in healthy volunteers without evidence of pancreatic disease (n=23). Results The total load of cfDNA, obtained by Alu83 quantitation, was the highest in PDAC patients than in any of the other patient groups (Welch t test; p<0.001) and was an average predictor of PDAC disease (AUC=0.664; CI, 0.56-0.77). A nonlinear association between Alu83 levels and subjects’ age was detected (Spearman's rho=0.35; p<0.001) in the overall population, as well as within the PDAC patients’ group (Spearman's rho=0.47; p<0.001). Necrosis-derived cfDNA fragments, quantitated with the Alu244 amplicon, were barely detectable in any of the samples and, in that respect, comparable between the different subject groups. CfDNA integrity estimation (Alu244/Alu83 ratio) was significantly affected by the limited detectability of plasma Alu244 levels. Conclusion The lack of detectable levels of necrosis-derived cfDNA in pancreatic pathologies considerably affects the clinical use of such biomarker in PDAC patients. Different methods of analysis should be applied in the evaluation of the cfDNA diagnostic value in pancreas pathology.

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