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BMC Cancer
Published in: BMC Cancer 1/2018

Open Access 01-12-2018 | Research article

Inhibition of autophagy sensitizes cancer cells to Photofrin-based photodynamic therapy

Authors: Antoni Domagala, Joanna Stachura, Magdalena Gabrysiak, Angelika Muchowicz, Radoslaw Zagozdzon, Jakub Golab, Malgorzata Firczuk

Published in: BMC Cancer | Issue 1/2018

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Abstract

Background

Accumulating evidence suggest that autophagy plays a pivotal role in various anticancer therapies, including photodynamic therapy (PDT), acting as a pro-death or pro-survival mechanism in a context-dependent manner. Therefore, we aimed to determine the role of autophagy in Photofrin-based PDT.

Methods

In vitro cytotoxic/cytostatic effects of PDT were evaluated with crystal violet cell viability assay. Autophagy induction was analyzed by immunoblotting and immunofluorescence using anti-LC3 antibody. Autophagy was inhibited by shRNA-mediated ATG5 knockdown or CRISPR/Cas9-mediated ATG5 knockout. Apoptosis was assessed by flow cytometry analysis of propidium iodide and anexin V-positive cells as well as by detection of cleaved PARP and caspase 3 proteins using immunoblotting. Protein carbonylation was evaluated by the 2,4-dinitrophenylhydrazine (DNPH) method.

Results

Photofrin-PDT leads to robust autophagy induction in two cancer cell lines, Hela and MCF-7. shRNA-mediated knockdown of ATG5 only partially blocks autophagic response and only marginally affects the sensitivity of Hela and MCF-7 cells to PDT. ATG5 knockout in HeLa cell line utilizing CRISPR/Cas9 genome editing results in increased PDT-mediated cytotoxicity, which is accompanied by an enhanced apoptotic response and increased accumulation of carbonylated proteins.

Conclusions

Altogether, these observations imply that autophagy contributes to Photofrin-PDT resistance by enabling clearance of carbonylated and other damaged proteins. Therefore, autophagy inhibition may serve as a strategy to improve PDT efficacy.
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Metadata
Title
Inhibition of autophagy sensitizes cancer cells to Photofrin-based photodynamic therapy
Authors
Antoni Domagala
Joanna Stachura
Magdalena Gabrysiak
Angelika Muchowicz
Radoslaw Zagozdzon
Jakub Golab
Malgorzata Firczuk
Publication date
01-12-2018
Publisher
BioMed Central
Published in
BMC Cancer / Issue 1/2018
Electronic ISSN: 1471-2407
DOI
https://doi.org/10.1186/s12885-018-4126-y

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