Basic Study
Copyright ©The Author(s) 2018. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Sep 7, 2018; 24(33): 3749-3759
Published online Sep 7, 2018. doi: 10.3748/wjg.v24.i33.3749
Establishment, functional and genetic characterization of a colon derived large cell neuroendocrine carcinoma cell line
Michael Gock, Christina S Mullins, Christine Harnack, Friedrich Prall, Robert Ramer, Anja Göder, Oliver H Krämer, Ernst Klar, Michael Linnebacher
Michael Gock, Ernst Klar, Department of General Surgery, University of Rostock, Rostock 18055, Germany
Christina S Mullins, Christine Harnack, Michael Linnebacher, Department of General Surgery, Section of Molecular Oncology and Immunotherapy, University of Rostock, Rostock 18055, Germany
Friedrich Prall, Institute of Pathology, University of Rostock, Rostock 18055, Germany
Robert Ramer, Institute of Pharmacology, University of Rostock, Rostock 18055, Germany
Anja Göder, Oliver H Krämer, Institute of Toxicology, University Medical Center Mainz, Mainz 55131, Germany
Author contributions: Linnebacher M and Klar E designed research; Mullins CS, Harnack C, Prall F, Ramer R and Göder A performed research; Gock M, Krämer OH and Linnebacher M analyzed data; Gock M, Mullins CS, Krämer OH and Linnebacher M wrote the paper.
Institutional review board statement: This study was reviewed and approved by Ethikkommission an der Medizinischen Fakultät der Universität Rostock, St.-Georg-Str. 108, Rostock 18055, Germany.
Informed consent statement: All procedures were approved by Ethikkommission an der Medizinischen Fakultät der Universität Rostock (reference number II HV 43/2004) in accordance with generally accepted guidelines for the use of human material.
Conflict-of-interest statement: To the best of our knowledge, no conflict of interest exists.
Data sharing statement: No additional data are available.
ARRIVE guidelines statement: The ARRIVE Guidelines have been adopted.
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: Michael Linnebacher, PhD, Postdoc, Research Scientist, Department of General Surgery, Section of Molecular Oncology and Immunotherapy, University of Rostock, Schillingallee 35, Rostock 18055, Germany. michael.linnebacher@med.uni-rostock.de
Telephone: +49-381-4946043 Fax: +49-381-4946002
Received: February 28, 2018
Peer-review started: March 1, 2018
First decision: April 18, 2018
Revised: June 14, 2018
Accepted: June 16, 2018
Article in press: June 16, 2018
Published online: September 7, 2018
ARTICLE HIGHLIGHTS
Research background

Only a few gastrointestinal neuroendocrine tumor lines have been published in the last decades. A major problem is the very heterogeneous pathological terminology when trying to classify these cell lines according to the lately revised WHO classification with regard to their original tumors. Particularly, reports of poorly differentiated neuroendocrine carcinoma (NEC) are very scarce mainly due to their low incidence.

Research motivation

Gaining insights in the biology of large cell NEC is essential for the identification of potentially therapeutic molecular targets of this highly malignant neoplasia. Individual tumor models deliver exceptional tools for further research of these objectives. However, well characterized and low passage NEC models are still rare.

Research objectives

Main objective of the study was the establishing and profound characterization of an patient derived ultra-low passage NEC cell line and corresponding patient-derived xenograft (PDX) model that allows drug response testing and prediction.

Research methods

Cell line establishment could be realized from direct in vitro culturing of colonic NEC tissue. In addition, a PDX model could be established from frozen tumor samples. Profound analysis of morphological features, invasive and migratory behavior as well as expression of neuroendocrine markers was done. Detailed phenotypic analysis was performed by microscopy and multicolor flow cytometry. Chromosomal aberrations were mapped by array comparative genomic hybridization and DNA profiling was analyzed by DNA fingerprinting. At last drug responsiveness was evaluated and the sensitivity against chemotherapeutic agents assessed.

Research results

The cell line displayed characteristic morphological and molecular features of large cell NEC with KI-67 > 50%. In vitro and in vivo experiments demonstrated that the cell line retained their malignant properties. Molecular-pathological analysis revealed a CpG island promoter methylation positive cell line with microsatellite instability being absent. The KRAS gene was not mutated whereas a BRAF V600E mutation was detected. A high sensitivity to such drugs as etoposide, cisplatin and 5-FU could be observed with a more resistant phenotype to rapamycin.

Research conclusions

Taken together, this study describes the development and basic characterization of powerful matched in vitro and in vivo patient-derived models not only to perform basic research to better understand the biology of NECs, but also to establish novel therapeutic options.

Research perspectives

This descriptive study exemplifies the methodology and characterization of a large cell NEC cell line directly from original patients’ tumor material. This will help to improve the ability for personalizing tumor therapy in the near future.