Carfilzomib combined with suberanilohydroxamic acid (SAHA) synergistically promotes endoplasmic reticulum stress in non-small cell lung cancer cell lines

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Abstract

Purpose: The endoplasmic reticulum (ER) stress response is a therapeutic target for pharmacologic intervention in cancer cells. We hypothesized that combining carfilzomib (CFZ), a proteasome inhibitor, and vorinostat (SAHA), a histone deacetylase (HDAC) inhibitor, would synergistically activate ER stress in non-small cell lung cancer (NSCLC) cell lines, resulting in enhanced anti-tumor activity. Methods: Five NSCLC cell lines were treated with CFZ, SAHA, or the combination and cell proliferation measured using the MTT assay. Calcusyn software was utilized to determine the combination index as a measure of synergy. Cell viability and cytotoxicity were measured using trypan blue exclusion, CellTiter, and CytoTox assays. Western blot was used to measure markers of apoptosis, ER stress, and oxidative stress-related proteins. Reactive oxygen species (ROS) was measured using the fluorophore CM–H2DCFDA. Results: Synergistic activity was observed for all cell lines following 48 and 72 h of combined treatment. H520 and A549 cell lines were used to assess viability and apoptosis. In both cell lines, increased death and cleaved caspase-3 were observed following combination treatment as compared with single-agent treatments. Combination therapy was associated with upregulation of ER stress-regulated proteins including activating transcription factor 4, GRP78/BiP, and C/EBP homologous protein. Both cell lines also showed increased ROS and the oxidative stress-related protein, heat shock protein 70. Conclusion: Combining proteasome inhibition with HDAC inhibition enhances ER stress, which may contribute to the synergistic anticancer activity observed in NSCLC cell lines. Further preclinical and clinical studies of CFZ + SAHA in NSCLC are warranted.

Original languageEnglish (US)
Pages (from-to)549-560
Number of pages12
JournalJournal of Cancer Research and Clinical Oncology
Volume142
Issue number3
DOIs
StatePublished - Mar 1 2016

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Endoplasmic Reticulum Stress
Non-Small Cell Lung Carcinoma
Cell Line
Heat-Shock Proteins
Reactive Oxygen Species
Oxidative Stress
Activating Transcription Factor 4
Transcription Factor CHOP
Apoptosis
Proteasome Inhibitors
Histone Deacetylase Inhibitors
HSP70 Heat-Shock Proteins
Histone Deacetylases
Trypan Blue
Proteasome Endopeptidase Complex
Caspase 3
carfilzomib
vorinostat
Neoplasms
Cell Survival

Keywords

  • Carfilzomib
  • Non-small cell lung cancer
  • Proteasome inhibitor
  • SAHA

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

@article{ff0e4fa71fef40bd9eb5786aa4ff3b14,
title = "Carfilzomib combined with suberanilohydroxamic acid (SAHA) synergistically promotes endoplasmic reticulum stress in non-small cell lung cancer cell lines",
abstract = "Purpose: The endoplasmic reticulum (ER) stress response is a therapeutic target for pharmacologic intervention in cancer cells. We hypothesized that combining carfilzomib (CFZ), a proteasome inhibitor, and vorinostat (SAHA), a histone deacetylase (HDAC) inhibitor, would synergistically activate ER stress in non-small cell lung cancer (NSCLC) cell lines, resulting in enhanced anti-tumor activity. Methods: Five NSCLC cell lines were treated with CFZ, SAHA, or the combination and cell proliferation measured using the MTT assay. Calcusyn software was utilized to determine the combination index as a measure of synergy. Cell viability and cytotoxicity were measured using trypan blue exclusion, CellTiter, and CytoTox assays. Western blot was used to measure markers of apoptosis, ER stress, and oxidative stress-related proteins. Reactive oxygen species (ROS) was measured using the fluorophore CM–H2DCFDA. Results: Synergistic activity was observed for all cell lines following 48 and 72 h of combined treatment. H520 and A549 cell lines were used to assess viability and apoptosis. In both cell lines, increased death and cleaved caspase-3 were observed following combination treatment as compared with single-agent treatments. Combination therapy was associated with upregulation of ER stress-regulated proteins including activating transcription factor 4, GRP78/BiP, and C/EBP homologous protein. Both cell lines also showed increased ROS and the oxidative stress-related protein, heat shock protein 70. Conclusion: Combining proteasome inhibition with HDAC inhibition enhances ER stress, which may contribute to the synergistic anticancer activity observed in NSCLC cell lines. Further preclinical and clinical studies of CFZ + SAHA in NSCLC are warranted.",
keywords = "Carfilzomib, Non-small cell lung cancer, Proteasome inhibitor, SAHA",
author = "Hanke, {Neale T.} and Garland, {Linda L} and Baker, {Amanda F}",
year = "2016",
month = "3",
day = "1",
doi = "10.1007/s00432-015-2047-6",
language = "English (US)",
volume = "142",
pages = "549--560",
journal = "Journal of Cancer Research and Clinical Oncology",
issn = "0171-5216",
publisher = "Springer Verlag",
number = "3",

}

TY - JOUR

T1 - Carfilzomib combined with suberanilohydroxamic acid (SAHA) synergistically promotes endoplasmic reticulum stress in non-small cell lung cancer cell lines

AU - Hanke, Neale T.

AU - Garland, Linda L

AU - Baker, Amanda F

PY - 2016/3/1

Y1 - 2016/3/1

N2 - Purpose: The endoplasmic reticulum (ER) stress response is a therapeutic target for pharmacologic intervention in cancer cells. We hypothesized that combining carfilzomib (CFZ), a proteasome inhibitor, and vorinostat (SAHA), a histone deacetylase (HDAC) inhibitor, would synergistically activate ER stress in non-small cell lung cancer (NSCLC) cell lines, resulting in enhanced anti-tumor activity. Methods: Five NSCLC cell lines were treated with CFZ, SAHA, or the combination and cell proliferation measured using the MTT assay. Calcusyn software was utilized to determine the combination index as a measure of synergy. Cell viability and cytotoxicity were measured using trypan blue exclusion, CellTiter, and CytoTox assays. Western blot was used to measure markers of apoptosis, ER stress, and oxidative stress-related proteins. Reactive oxygen species (ROS) was measured using the fluorophore CM–H2DCFDA. Results: Synergistic activity was observed for all cell lines following 48 and 72 h of combined treatment. H520 and A549 cell lines were used to assess viability and apoptosis. In both cell lines, increased death and cleaved caspase-3 were observed following combination treatment as compared with single-agent treatments. Combination therapy was associated with upregulation of ER stress-regulated proteins including activating transcription factor 4, GRP78/BiP, and C/EBP homologous protein. Both cell lines also showed increased ROS and the oxidative stress-related protein, heat shock protein 70. Conclusion: Combining proteasome inhibition with HDAC inhibition enhances ER stress, which may contribute to the synergistic anticancer activity observed in NSCLC cell lines. Further preclinical and clinical studies of CFZ + SAHA in NSCLC are warranted.

AB - Purpose: The endoplasmic reticulum (ER) stress response is a therapeutic target for pharmacologic intervention in cancer cells. We hypothesized that combining carfilzomib (CFZ), a proteasome inhibitor, and vorinostat (SAHA), a histone deacetylase (HDAC) inhibitor, would synergistically activate ER stress in non-small cell lung cancer (NSCLC) cell lines, resulting in enhanced anti-tumor activity. Methods: Five NSCLC cell lines were treated with CFZ, SAHA, or the combination and cell proliferation measured using the MTT assay. Calcusyn software was utilized to determine the combination index as a measure of synergy. Cell viability and cytotoxicity were measured using trypan blue exclusion, CellTiter, and CytoTox assays. Western blot was used to measure markers of apoptosis, ER stress, and oxidative stress-related proteins. Reactive oxygen species (ROS) was measured using the fluorophore CM–H2DCFDA. Results: Synergistic activity was observed for all cell lines following 48 and 72 h of combined treatment. H520 and A549 cell lines were used to assess viability and apoptosis. In both cell lines, increased death and cleaved caspase-3 were observed following combination treatment as compared with single-agent treatments. Combination therapy was associated with upregulation of ER stress-regulated proteins including activating transcription factor 4, GRP78/BiP, and C/EBP homologous protein. Both cell lines also showed increased ROS and the oxidative stress-related protein, heat shock protein 70. Conclusion: Combining proteasome inhibition with HDAC inhibition enhances ER stress, which may contribute to the synergistic anticancer activity observed in NSCLC cell lines. Further preclinical and clinical studies of CFZ + SAHA in NSCLC are warranted.

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KW - Proteasome inhibitor

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U2 - 10.1007/s00432-015-2047-6

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