Impact of DNA repair and reactive oxygen species levels on radioresistance in pancreatic cancer

Lily Nguyen; Sophie Dobiasch; Günter Schneider; Roland M Schmid; Omid Azimzadeh; Kristiyan Kanev; Dominik Buschmann; Michael W Pfaffl; Stefan Bartzsch; Thomas E Schmid; Daniela Schilling; Stephanie E Combs

doi:10.1016/j.radonc.2021.03.038

Impact of DNA repair and reactive oxygen species levels on radioresistance in pancreatic cancer

Radiother Oncol. 2021 Jun:159:265-276. doi: 10.1016/j.radonc.2021.03.038. Epub 2021 Apr 9.

Authors

Affiliations

¹ Institute of Radiation Medicine (IRM), Department of Radiation Sciences (DRS), Helmholtz Zentrum München, Neuherberg, Germany; Department of Radiation Oncology, School of Medicine, Klinikum rechts der Isar, Technical University of Munich (TUM), Germany.
² Institute of Radiation Medicine (IRM), Department of Radiation Sciences (DRS), Helmholtz Zentrum München, Neuherberg, Germany; Department of Radiation Oncology, School of Medicine, Klinikum rechts der Isar, Technical University of Munich (TUM), Germany; Deutsches Konsortium für Translationale Krebsforschung (DKTK), Partner Site Munich, Munich, Germany.
³ Department of Medicine II, School of Medicine, Klinikum rechts der Isar, Technical University of Munich (TUM), Germany; Deutsches Krebsforschungszentrum (DKFZ) and German Cancer Consortium (DKTK), Heidelberg, Germany.
⁴ Department of Medicine II, School of Medicine, Klinikum rechts der Isar, Technical University of Munich (TUM), Germany.
⁵ Institute of Radiation Biology (ISB), Department of Radiation Sciences (DRS), Helmholtz Zentrum München, Neuherberg, Germany.
⁶ Division of Animal Physiology and Immunology, TUM School of Life Sciences Weihenstephan, Technical University of Munich (TUM), Freising, Germany.
⁷ Institute of Radiation Medicine (IRM), Department of Radiation Sciences (DRS), Helmholtz Zentrum München, Neuherberg, Germany; Department of Radiation Oncology, School of Medicine, Klinikum rechts der Isar, Technical University of Munich (TUM), Germany; Deutsches Konsortium für Translationale Krebsforschung (DKTK), Partner Site Munich, Munich, Germany. Electronic address: stephanie.combs@tum.de.

PMID: 33839203
DOI: 10.1016/j.radonc.2021.03.038

Abstract

Purpose: Radioresistance in pancreatic cancer patients remains a critical obstacle to overcome. Understanding the molecular mechanisms underlying radioresistance may achieve better response to radiotherapy and thereby improving the poor treatment outcome. The aim of the present study was to elucidate the mechanisms leading to radioresistance by detailed characterization of isogenic radioresistant and radiosensitive cell lines.

Methods: The human pancreatic cancer cell lines, Panc-1 and MIA PaCa-2 were repeatedly exposed to radiation to generate radioresistant (RR) isogenic cell lines. The surviving cells were expanded, and their radiosensitivity was measured using colony formation assay. Tumor growth delay after irradiation was determined in a mouse pancreatic cancer xenograft model. Gene and protein expression were analyzed using RNA sequencing and Western blot, respectively. Cell cycle distribution and apoptosis (Caspase 3/7) were measured by FACS analysis. Reactive oxygen species generation and DNA damage were analyzed by detection of CM-H₂DCFDA and γH2AX staining, respectively. Transwell chamber assays were used to investigate cell migration and invasion.

Results: The acquired radioresistance of RR cell lines was demonstrated in vitro and validated in vivo. Ingenuity pathway analysis of RNA sequencing data predicted activation of cell viability in both RR cell lines. RR cancer cell lines demonstrated greater DNA repair efficiency and lower basal and radiation-induced reactive oxygen species levels. Migration and invasion were differentially affected in RR cell lines.

Conclusions: Our data indicate that repeated exposure to irradiation increases the expression of genes involved in cell viability and thereby leads to radioresistance. Mechanistically, increased DNA repair capacity and reduced oxidative stress might contribute to the radioresistant phenotype.

Keywords: Invasion; Migration; Pancreatic cancer; RNA sequencing; Radioresistance.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Apoptosis
Cell Line, Tumor
DNA Repair
Gene Expression Regulation, Neoplastic*
Humans
Mice
Pancreatic Neoplasms* / genetics
Pancreatic Neoplasms* / radiotherapy
Radiation Tolerance / genetics
Reactive Oxygen Species

Substances

Reactive Oxygen Species