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©The Author(s) 2018. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Feb 21, 2018; 24(7): 810-818
Published online Feb 21, 2018. doi: 10.3748/wjg.v24.i7.810
Cryopreservation for delayed circulating tumor cell isolation is a valid strategy for prognostic association of circulating tumor cells in gastroesophageal cancer
Daniel Brungs, David Lynch, Alison WS Luk, Elahe Minaei, Marie Ranson, Morteza Aghmesheh, Kara L Vine, Martin Carolan, Mouhannad Jaber, Paul de Souza, Therese M Becker
Daniel Brungs, Elahe Minaei, Marie Ranson, Morteza Aghmesheh, Kara L Vine, Martin Carolan, Illawarra Health and Medical Research Institute, University of Wollongong, Wollongong 2500, Australia
Daniel Brungs, Elahe Minaei, Marie Ranson, Kara L Vine, School of Biological Sciences, University of Wollongong, Wollongong 2500, Australia
Daniel Brungs, Morteza Aghmesheh, Martin Carolan, Mouhannad Jaber, Illawarra Cancer Centre, Wollongong Hospital, Wollongong 2500, Australia
Daniel Brungs, David Lynch, Alison WS Luk, Elahe Minaei, Marie Ranson, Morteza Aghmesheh, Kara L Vine, Martin Carolan, Mouhannad Jaber, Paul de Souza, Therese M Becker, CONCERT-Translational Cancer Research Centre, New South Wales 2000, Australia
David Lynch, Paul de Souza, Therese M Becker, Centre for Circulating Tumor Cell Diagnostics and Research, Ingham Institute for Applied Medical Research, Liverpool Hospital, Sydney 2170, Australia
Paul de Souza, Therese M Becker, School of Medicine, University of Western Sydney, Sydney 2170, Australia
Paul de Souza, Therese M Becker, South Western Medical School, University of New South Wales, Sydney 2170, Australia
Institutional review board statement: This study was approved by South Western Sydney Local Health District Human Research Ethics Committee (Project Number 15/072). A written informed consent was obtained from each participant before sample collection.
Conflict-of-interest statement: All authors have no conflicts of interest to declare.
Data sharing statement: No additional data are available.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Daniel Brungs, BSc, MBBS, Doctor, Illawarra Cancer Care Centre, Wollongong Hospital, 348 Crown St, Wollongong NSW 2500, Australia. daniel.brungs@health.nsw.gov.au
Telephone: +61-2-42225200 Fax: +61-2-42225243
Received: November 6, 2017
Peer-review started: November 7, 2017
First decision: November 30, 2017
Revised: December 11, 2017
Accepted: December 19, 2017
Article in press: December 19, 2017
Published online: February 21, 2018
AIM
To demonstrate the feasibility of cryopreservation of peripheral blood mononuclear cells (PBMCs) for prognostic circulating tumor cell (CTC) detection in gastroesophageal cancer.
METHODS
Using 7.5 mL blood samples collected in EDTA tubes from patients with gastroesopheagal adenocarcinoma, CTCs were isolated by epithelial cell adhesion molecule based immunomagnetic capture using the IsoFlux platform. Paired specimens taken during the same blood draw (n = 15) were used to compare number of CTCs isolated from fresh and cryopreserved PBMCs. Blood samples were processed within 24 h to recover the PBMC fraction, with PBMCs used for fresh analysis immediately processed for CTC isolation. Cryopreservation of PBMCs lasted from 2 wk to 25.2 mo (median 14.6 mo). CTCs isolated from pre-treatment cryopreserved PBMCs (n = 43) were examined for associations with clinicopathological variables and survival outcomes.
RESULTS
While there was a significant trend to a decrease in CTC numbers associated with cryopreserved specimens (mean number of CTCs 34.4 vs 51.5, P = 0.04), this was predominately in samples with a total CTC count of > 50, with low CTC count samples less affected (P = 0.06). There was no significant association between the duration of cryopreservation and number of CTCs. In cryopreserved PBMCs from patient samples prior to treatment, a high CTC count (> 17) was associated with poorer overall survival (OS) (n = 43, HR = 4.4, 95%CI: 1.7-11.7, P = 0.0013). In multivariate analysis, after controlling for sex, age, stage, ECOG performance status, and primary tumor location, a high CTC count remained significantly associated with a poorer OS (HR = 3.7, 95%CI: 1.2-12.4, P = 0.03).
CONCLUSION
PBMC cryopreservation for delayed CTC isolation is a valid strategy to assist with sample collection, transporting and processing.
Core tip: This study demonstrates a novel and robust protocol for the cryopreservation and thawing of patient blood samples, demonstrating reliable circulating tumor cell isolation and characterisation after the long term storage of patient samples. Using the largest patient cohort reported to date, we validated our method by confirming the independent prognostic association of circulating tumor cell (CTC) enumeration from cryopreserved peripheral blood mononuclear cells. Cryopreservation may assist with the wider incorporation of CTC collection and analysis in biobanking, retrospective studies, and large international clinical trials, by facilitating specimen storage, bulk transporting, and batch processing.
