133P - Heparinase enables reliable quantification of circulating tumor DNA from heparin plasma samples by droplet digital PCR

Date 11 September 2017
Event ESMO 2017 Congress
Session Poster display session
Topics Cancer in Adolescents
Biomarkers
Hepatobiliary Cancers
Translational Research
Presenter David Sefrioui
Citation Annals of Oncology (2017) 28 (suppl_5): v22-v42. 10.1093/annonc/mdx363
Authors D. Sefrioui1, L. Beaussire2, F. Clatot3, A. Perdrix4, P. Michel5, F. Di Fiore5, N. Sarafan-Vasseur2
  • 1Gastroenterology, CHU Hôpitaux de Rouen-Charles Nicolle, 76031 - Rouen/FR
  • 2Institute For Biomedical Research And Innovation, Inserm U1079, University of Rouen, 76000 - rouen/FR
  • 3Department Of Medical Oncology, Henri Becquerel Center, 76000 - Rouen/FR
  • 4Department Of Biopathology, Henri Becquerel Center, 76000 - Rouen/FR
  • 5Department Of Oncology, CHU Charles Nicolle, 76031 - Rouen/FR

Abstract

Background

Circulating tumor DNA has been highlighted as a potential “liquid biopsy”, which can be used to identify prognostic and predictive alterations in oncology. Heparin is often used as plasma anticoagulant source for tumor marker analysis such as CA19.9 or CA15.3 but corresponds also to an inhibitory of PCR not enabling ctDNA detection. We aimed to evaluate the impact of heparinase addition on heparinized plasma samples to recovery the possibility of ctDNA analysis on samples initially dedicated for tumor markers analysis.

Methods

Plasma samples were collected in heparinized (n = 194) and EDTA (n = 8) tubes from hormone receptor-positive metastatic breast cancer (HR+MBC) patients resistant to aromatase inhibitor (AI) treatment (n = 144) and from newly diagnosed pancreatic adenocarcinoma (PA) patients (n = 50). ESR1 and KRAS mutations were used as targets for ctDNA detection in HR+MBC and PA patients, respectively. ctDNA was detected by droplet digital PCR after an amplification step either without or with heparinase (H- and H+ respectively). PCR efficiency and ctDNA detection rate were compared between H- and H+ subgroups as well as with EDTA subgroup.

Results

Heparinase addition improved significantly PCR efficiency for 91/144 HR+MBC and 26/50 PA patients enabling ctDNA detection in 22/91 (24%) and 13/26 (50%) of these patients. Moreover, heparinase condition did not quantitatively and qualitatively alter the ctDNA detection for patients without heparin inhibition of PCR and comparable results for ctDNA detection were obtained between H+ and EDTA subgroups.

Conclusions

Heparinase addition allows removing the heparin inhibition on cfDNA amplification and to detect and quantify accurately ctDNA levels by dPCR in the heparinized plasma samples.

Clinical trial identification

Legal entity responsible for the study

Rouen University Hospital

Funding

None

Disclosure

All authors have declared no conflicts of interest.