Combined proteasome and histone deacetylase inhibition in non-small cell lung cancer.
OBJECTIVE
Inhibitors of histone deacetylases are potent inducers of cell-cycle arrest and apoptosis in certain malignancies. We have previously demonstrated that chemotherapy activates the antiapoptotic transcription factor nuclear factor kappa B in non-small cell lung cancer and fails to induce significant levels of apoptosis. We hypothesize that nuclear factor kappa B inhibition with the proteasome inhibitor bortezomib (formerly known as PS-341) will sensitize non-small cell lung cancer cells to histone deacetylase inhibitor-mediated apoptosis.


METHODS
Tumorigenic non-small cell lung cancer cells (A549, H358, and H460) were treated with bortezomib, followed by the histone deactylase inhibitor sodium butyrate. After treatment, nuclear factor kappa B transcriptional activity was measured by using a luciferase reporter assay and transcription of the nuclear factor kappa B-dependent gene IL8. Apoptosis was determined on the basis of caspase-3 activation and DNA fragmentation. Western blot analyses for the cell-cycle regulatory proteins p21 and p53 were performed, and cell-cycle alterations were determined by means of FACS analysis. Experiments were performed in triplicate, and statistical significance was determined by using unpaired t tests.


RESULTS
Butyrate increased nuclear factor kappa B transcriptional activity 4-fold relative to that seen in control cells (P =.05) in all non-small cell lung cancer cell lines. Treatment with bortezomib reduced butyrate-induced activation of nuclear factor kappa B to baseline levels. The proteins p21 and p53 were stabilized after treatment with bortezomib, correlating with a G(2)/M cell-cycle arrest. Treatment with butyrate alone resulted in minimal apoptosis, but combined histone deacetylase and proteasome inhibition increased apoptosis 3- to 4-fold (P =.02).


CONCLUSIONS
Combined molecular targeting of histone deacteylases and proteasomes synergistically induced apoptosis in non-small cell lung cancer. Pharmacologic nuclear factor kappa B suppression through proteasome inhibition, followed by treatment with histone deacetylase inhibitors, might represent a novel treatment strategy for patients with non-small cell lung cancer.
Denlinger
Chadrick E
CE
Department of Surgery, University of Virginia School of Medicine, Charlottesville, 22908, USA.
Keller
Michael D
MD
Mayo
Marty W
MW
Broad
R Michael
RM
Jones
David R
DR
D000971:Antineoplastic Combined Chemotherapy Protocols / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects; D001897:Boronic Acids / Q000008:administration & dosage; D000069286:Bortezomib; D002087:Butyrates / Q000008:administration & dosage; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000503:physiopathology; D002453:Cell Cycle / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003546:Cysteine Endopeptidases / Q000008:administration & dosage; D006655:Histone Deacetylases / Q000008:administration & dosage; D006801:Humans; D016209:Interleukin-8 / Q000235:genetics; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000503:physiopathology; D009097:Multienzyme Complexes / Q000008:administration & dosage; D016328:NF-kappa B / Q000187:drug effects / Q000378:metabolism; D011480:Protease Inhibitors / Q000008:administration & dosage; D046988:Proteasome Endopeptidase Complex; D011719:Pyrazines / Q000008:administration & dosage; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects; D016896:Treatment Outcome; D014407:Tumor Cells, Cultured
D001897:Boronic Acids; D002087:Butyrates; D016209:Interleukin-8; D009097:Multienzyme Complexes; D016328:NF-kappa B; D011480:Protease Inhibitors; D011719:Pyrazines; D000069286:Bortezomib; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D006655:Histone Deacetylases