IMR Press / JMCM / Volume 3 / Issue 2 / DOI: 10.31083/j.jmcm.2020.02.730
Open Access Original Research
Pleiotropic activities of nitric oxide-releasing doxorubicin on P-glycoprotein/ABCB1
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1 Department of Oncology, University of Torino, 10126 Torino, Italy
J. Mol. Clin. Med. 2020, 3(2), 37–46; https://doi.org/10.31083/j.jmcm.2020.02.730
Submitted: 10 May 2020 | Accepted: 8 June 2020 | Published: 20 June 2020
Copyright: © 2020 Costamagna et al. Published by IMR press.
This is an open access article under the CC BY 4.0 license (https://creativecommons.org/licenses/by/4.0/).
Abstract

Doxorubicin is one of the first-line chemotherapeutic drugs for osteosarcoma, but the rate of success is below 60% of patients. The main cause of this low success is the presence of P-glycoprotein (P-gp/ABCB1) that effluxes the drug, limiting the intracellular accumulation and toxicity of Doxorubicin. P-gp also inhibits immunogenic cell death promoted by Doxorubicin. Nitric oxide-releasing Doxorubicin is a synthetic anthracycline effective against P-gp-positive osteosarcoma cells. It is not known how it impacts on P-gp expression and immunogenic cell death induction. To address this point, we treated human Doxorubicin-sensitive osteosarcoma U-2OS cells and their resistant variants with increasing amount of P-gp, with Dox and Nitric oxide-releasing Doxorubicin. While Doxorubicin was cytotoxic only in U-2OS cells, Nitric oxide-releasing Doxorubicin maintained its cytotoxic properties in all the resistant variants. Nitric oxide-releasing Doxorubicin elicited a strong nitrosative stress in whole cell extracts, endoplasmic reticulum and plasma membrane. P-gp was nitrated in all these compartments. The nitration caused protein ubiquitination and lower catalytic efficacy. The removal of P-gp from cell surface upon Nitric oxide-releasing Doxorubicin treatment disrupted its interaction with calreticulin, an immunogenic cell death-inducer that is inhibited by P-gp. Drug resistant cells treated with Nitric oxide-releasing Doxorubicin exposed calreticulin, were phagocytized by dendritic cells and expanded anti-tumor CD8+ T-lymphocytes. The efficacy of Nitric oxide-releasing Doxorubicin was validated in Dox-resistant osteosarcoma xenografts and was higher in immune-competent humanized mice than in immune-deficient mice, confirming that part of Nitric oxide-releasing Doxorubicin efficacy relies on the restoration of immunogenic cell death. Nitric oxide-releasing Doxorubicin was a pleiotropic anthracycline reducing activity and expression of P-gp, and restoring immunogenic cell death. It can be an innovative drug against P-gp-expressing/ Doxorubicin-resistant osteosarcomas.

Keywords
Osteosarcoma
doxorubicin
nitric oxide
P-glycoprotein
calreticulin
immunogenic cell death
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