Activation of the Unfolded Protein
Response Contributes toward the
Antitumor Activity of Vorinostat1
Soumen Kahali*,†, Bhaswati Sarcar*,†, Bin Fang‡,
Eli S. Williams§, John M. Koomen‡,¶, Philip J. Tofilon§
and Prakash Chinnaiyan*,†
*Radiation Oncology, H. Lee Moffitt Cancer Center, Tampa,
FL, USA;†Experimental Therapeutics, H. Lee Moffitt Cancer
Center, Tampa, FL, USA;‡Proteomics, H. Lee Moffitt Cancer
Center, Tampa, FL, USA;§Drug Discovery, H. Lee Moffitt
Cancer Center, Tampa, FL, USA;¶Molecular Oncology,
H. Lee Moffitt Cancer Center, Tampa, FL, USA
Histone deacetylase (HDAC) inhibitors represent an emerging class of anticancer agents progressing through clinical
trials. Although their primary target is thought to involve acetylation of core histones, several nonhistone substrates
have been identified, including heat shock protein (HSP) 90, which may contribute towards their antitumor activity.
Glucose-regulated protein 78 (GRP78) is a member of the HSP family of molecular chaperones and plays a central role
in regulating the unfolded protein response (UPR). Emerging data suggest that GRP78 is critical in cellular adaptation
and survival associated with oncogenesis and may serve as a cancer-specific therapeutic target. On the basis of
sharedhomology withHSP family proteins,wesoughttodetermine whether GRP78could serve asa moleculartarget
of the HDAC inhibitor vorinostat. Vorinostat treatment led to GRP78 acetylation, dissociation, and subsequent activa-
tion of its client protein double-stranded RNA-activated protein-like endoplasmic reticulum kinase (PERK). Investiga-
tions in a panel of cancer cell lines identified that UPR activation after vorinostat exposure is specific to certain lines.
Mass spectrometry performed on immunoprecipitated GRP78 identified lysine-585 as a specific vorinostat-
induced acetylation site of GRP78. Downstream activation of the UPR was confirmed, including eukaryotic initiat-
ing factor 2α phosphorylation and increase in ATF4 and C/EBP homologous protein expression. To determine the
biologic relevance of UPR activation after vorinostat, RNA interference of PERK was performed, demonstrating
significantly decreased sensitivity to vorinostat-induced cytotoxicity. Collectively, these findings indicate that
its antitumor activity.
Neoplasia (2010) 12, 80–86
Although cancer has traditionally been considered a disease originating
from genetic alterationsresultingin functional loss of tumor-suppressor
genes or gain of oncogenes, epigenetic modifications, or modulating
gene expression through mechanisms other than changes in the under-
lying DNA sequence have emerged as a contributing factor toward
inanopen configuration,allowing accessibility forspecifictranscription
factors and/or the general transcription machinery . The opposing
Abbreviations: ATF, activated transcription factor; CHOP, C/EBP homologous protein;
eIF, eukaryotic initiation factor; GRP78, glucose-regulated protein 78; HSP, heat shock
endoplasmic reticulum kinase; UPR, unfolded protein response
Address all correspondence to: Prakash Chinnaiyan, MD, H. Lee Moffitt Cancer Center
and Research Institute, SRB3, 12902 Magnolia Dr, Tampa, FL 33612.
1This work was supported in part by the Miles for Moffitt Research Award (P.C.). The
Moffitt Proteomics Facility is supported by the US Army Medical Research and Material
Command under Award No. DAMD17-02-2-0051 for a National Functional Genomics
Center, the National Cancer Institute under Award No. P30-CA076292 as a Cancer
Center Support Grant, and the Moffitt Foundation.
Received 18 August 2009; Revised 8 October 2009; Accepted 8 October 2009
Copyright © 2010 Neoplasia Press, Inc. All rights reserved 1522-8002/10/$25.00
Volume 12 Number 1 January 2010pp. 80–86
understood at present [16,17]. In our studies, vorinostat induced the
expression of CHOP, which is a well-described mediator governing
UPR activation through RNA interference of PERK leads to a signifi-
pathwayplaysinits antitumor activity. However, PERK represents only
one of the three central mediators of UPR activation, which also in-
cludes ATF6 and inositol-requiring gene 1. Further study involving
the influence of vorinostat on these pathways may provide additional
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Vorinostat-induced Activation of the Unfolded Protein Response Kahali et al.Neoplasia Vol. 12, No. 1, 2010