SMAD4 Gene Mutation Renders Pancreatic Cancer Resistance to Radiotherapy through Promotion of Autophagy

Understanding the mechanism of radioresistance could help develop strategies to improve therapeutic response of patients with PDAC. The gene is frequently mutated in pancreatic cancer. In this study, we investigated the role of deficiency in pancreatic cancer cells' response to radiotherapy. We...

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Bibliographic Details
Published in:Clinical cancer research 2018-07, Vol.24 (13), p.3176-3185
Main Authors: Wang, Feng, Xia, Xiaojun, Yang, Chunying, Shen, Jianliang, Mai, Junhua, Kim, Han-Cheon, Kirui, Dickson, Kang, Ya'an, Fleming, Jason B, Koay, Eugene J, Mitra, Sankar, Ferrari, Mauro, Shen, Haifa
Format: Article
Language:English
Subjects:
Acetylcysteine
Animals
Antineoplastic Agents - pharmacology
Apoptosis - genetics
Apoptosis - radiation effects
Assaying
Autophagy
Autophagy - genetics
Autophagy - radiation effects
Biomarkers, Tumor
Cancer
Cell Line, Tumor
Cell Proliferation
Cell Survival - genetics
Cell Survival - radiation effects
Chloroquine
Depletion
Disease Models, Animal
DNA Damage
Experimental design
Flow cytometry
Gene Knockdown Techniques
Genomic Instability
Humans
Immunohistochemistry
Inhibitors
Mice
Mutation
Pancreatic cancer
Pancreatic Neoplasms - genetics
Pancreatic Neoplasms - metabolism
Pancreatic Neoplasms - radiotherapy
Phagocytosis
Point mutation
Proteins
Radiation
Radiation effects
Radiation therapy
Radiation Tolerance - genetics
Radioresistance
Radiosensitivity
Reactive oxygen species
Reactive Oxygen Species - metabolism
siRNA
Smad4 protein
Smad4 Protein - genetics
Smad4 Protein - metabolism
Survival
Tumors
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Summary:Understanding the mechanism of radioresistance could help develop strategies to improve therapeutic response of patients with PDAC. The gene is frequently mutated in pancreatic cancer. In this study, we investigated the role of deficiency in pancreatic cancer cells' response to radiotherapy. We downregulated SMAD4 expression with siRNA or shRNA and overexpressed SMAD4 in mutant pancreatic cancer cells followed by clonogenic survival assay to evaluate their effects on cell radioresistance. To study the mechanism of radioresistance, the effects of loss on reactive oxygen species (ROS) and autophagy were determined by flow cytometry and immunoblot analysis, respectively. Furthermore, we measured radioresistance by clonogenic survival assay after treatment with autophagy inhibitor (Chloroquine) and ROS inhibitor (N-acetyl-l-cysteine) in -depleted pancreatic cancer cells. Finally, the effects of on radioresistance were also confirmed in an orthotopic tumor model derived from -depleted Panc-1 cells. -depleted pancreatic cancer cells were more resistant to radiotherapy based on clonogenic survival assay. Overexpression of wild-type SMAD4 in -mutant cells rescued their radiosensitivity. Radioresistance mediated by depletion was associated with persistently higher levels of ROS and radiation-induced autophagy. Finally, depletion induced radioresistance in Panc-1-derived orthotopic tumor model ( = 0.038). More interestingly, we observed that the protein level of SMAD4 is inversely correlated with autophagy in orthotopic tumor tissue samples. Our results demonstrate that defective is responsible for radioresistance in pancreatic cancer through induction of ROS and increased level of radiation-induced autophagy. .
ISSN:1078-0432
1557-3265
DOI:10.1158/1078-0432.ccr-17-3435