TRAIL-Based High Throughput Screening Reveals a Link between TRAIL-Mediated Apoptosis and Glutathione Reductase, a Key Component of Oxidative Stress Response.

Authors

Dmitri Rozanov
Anton Cheltsov
Eduard Sergienko
Stefan Vasile
Vladislav Golubkov
Alexander E Aleshin
Trevor Levin
Elie Traer
Byron Hann
Julia Freimuth
Nikita Alexeev, George Washington University
Max A Alekseyev
Sergey P Budko
Hans Peter Bächinger
Paul Spellman

Document Type

Journal Article

Publication Date

1-1-2015

Journal

PLoS One

Volume

10

Issue

6

DOI

10.1371/journal.pone.0129566

Keywords

Animals; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Dose-Response Relationship, Drug; Doxorubicin; Drug Discovery; Glutathione; Glutathione Reductase; High-Throughput Screening Assays; Humans; Mice; Oxidative Stress; Reactive Oxygen Species; Small Molecule Libraries; TNF-Related Apoptosis-Inducing Ligand

Abstract

A high throughput screen for compounds that induce TRAIL-mediated apoptosis identified ML100 as an active chemical probe, which potentiated TRAIL activity in prostate carcinoma PPC-1 and melanoma MDA-MB-435 cells. Follow-up in silico modeling and profiling in cell-based assays allowed us to identify NSC130362, pharmacophore analog of ML100 that induced 65-95% cytotoxicity in cancer cells and did not affect the viability of human primary hepatocytes. In agreement with the activation of the apoptotic pathway, both ML100 and NSC130362 synergistically with TRAIL induced caspase-3/7 activity in MDA-MB-435 cells. Subsequent affinity chromatography and inhibition studies convincingly demonstrated that glutathione reductase (GSR), a key component of the oxidative stress response, is a target of NSC130362. In accordance with the role of GSR in the TRAIL pathway, GSR gene silencing potentiated TRAIL activity in MDA-MB-435 cells but not in human hepatocytes. Inhibition of GSR activity resulted in the induction of oxidative stress, as was evidenced by an increase in intracellular reactive oxygen species (ROS) and peroxidation of mitochondrial membrane after NSC130362 treatment in MDA-MB-435 cells but not in human hepatocytes. The antioxidant reduced glutathione (GSH) fully protected MDA-MB-435 cells from cell lysis induced by NSC130362 and TRAIL, thereby further confirming the interplay between GSR and TRAIL. As a consequence of activation of oxidative stress, combined treatment of different oxidative stress inducers and NSC130362 promoted cell death in a variety of cancer cells but not in hepatocytes in cell-based assays and in in vivo, in a mouse tumor xenograft model.

Comments

Reproduced with permission of PLoS ONE.

Creative Commons License

This work is licensed under a Creative Commons Attribution 4.0 License.

APA Citation

Rozanov, D., Cheltsov, A., Sergienko, E., Vasile, S., Golubkov, V., Aleshin, A., Levin, T., Traer, E., Hann, B., Freimuth, J., Alexeev, N., Alekseyev, M., Budko, S., Bächinger, H., & Spellman, P. (2015). TRAIL-Based High Throughput Screening Reveals a Link between TRAIL-Mediated Apoptosis and Glutathione Reductase, a Key Component of Oxidative Stress Response.. PLoS One, 10 (6). http://dx.doi.org/10.1371/journal.pone.0129566

Peer Reviewed

1

Open Access

1