The Histone Deacetylase Inhibitor Suberoylanilide Hydroxamic Acid Induces Growth Inhibition and Enhances Gemcitabine-Induced Cell Deathin Pancreatic Cancer Nichole BoyerArnold, Nohea Arkus, Jason Gunn, and Murray Korc Abstract Purpose: Pancreatic cancer is an aggressive human malignancy that is generally refractory to chemotherapy. Histone deacetylase inhibitors are novel agents that modulate cell growth and survival. In this study, we sought to determine whether a relatively new histone deacetylase inhibitor, suberoylanilide hydroxamic acid (SAHA), inhibits pancreatic cancer cellgrowth. Experimental Design: The effects of SAHA on the growth of three pancreatic cancer cell lines (BxPC3, COLO-357, and PANC-1) were examined with respect to cell cycle progression, p21induction andlocalization, andinteractions with the nucleoside analogue gemcitabine. Results: SAHA induced a G 1 cell cycle arrest in BxPC-3 cells and COLO-357 cells but not in PANC-1cells.This arrest was dependent, in part, on induction of p21by SAHA, as p21was not induced in PANC-1cells, and knockdown of p21using small interfering RNA oligonucleotides nearly completely suppressed the effects of SAHA on cell cycle arrest in COLO-357 and partly attenuated the effects of SAHA in BxPC-3. COLO-357 and BxPC-3 cells, but not PANC-1cells, werealsosensitivetogemcitabine.Inthegemcitabine-resistantPANC-1cells,a48-hcotreatment with SAHA rendered the cells sensitive to the inhibitory and proapoptotic effects of gemcitabine. An additive effect on growth inhibition by SAHA and gemcitabine was observed in COLO-357 and BxPC-3 cells. Moreover, analysis of p21distribution in COLO-357 cells revealed that SAHA inducedthe cytoplasmiclocalizationofbothp21andphospho-p21. Conclusions: These data indicate that SAHA exerts proapoptotic effects in pancreatic cancer cells,inpart,byup-regulatingp21andsequesteringitinthecytoplasm,raisingthepossibilitythat SAHA may have therapeutic potentialin the treatment of pancreatic cancer. Pancreatic cancer is the fifth leading cause of cancer death in the United States. The disease presents late in life (mean age, >60 years) and the incidence virtually equals mortality where median survival time is 4 to 6 months at diagnosis and the 5-year survival rate is only 1% to 3% (1). The American Cancer Society estimates that in 2005, f32,180 people in the United States will be diagnosed with pancreatic cancer and f31,800 will die of the disease. Despite our advances in the understanding of the molecular biology of pancreatic cancer, the treatment options are still unsatisfactory and there is at present no curative treatment, hence the high death rates (2). Chemotherapy, antihormonal treatment (3), radiotherapy (4), and anti–pancreatic cancer–specific antibodies (3) have not led to a significant improvement. Moreover, due to the dis- seminated nature of the disease at presentation, surgical resection of pancreatic ductal adenocarcinoma is only carried out in 15% to 20% of patients (5). Pancreatic ductal adenocarcinoma is characterized by muta- tions and/or silencing of tumor suppressor genes, such as and p53 and Smad4 , the overexpression of mitogenic growth factors and their cognate high affinity tyrosine kinase re- ceptors,andmutationofK-ras (6).Therearealsodefectsincell cycle–regulating genes, such as the increased expression of cyclin D1 and mutation/silencing of p16 , which contribute to the inactivation of the retinoblastoma protein (7). These alter- ations contribute to the excessive growth of pancreatic tumors and to the resistance of pancreatic cancers to chemotherapeutic agents (8). Histone acetylation is a posttranslational modification of the nucleosomal histones that affects chromatin structure and modulatesgeneexpression.Theacetylationstatusofhistonesis modulated by histone acetyl transferases and histone deacety- lases (HDAC). Histone acetyl transferases are generally considered to be transcriptional activators because histone acetylation is associated with transcriptionally active chroma- tin, whereas HDACs are considered as transcriptional inhib- itors because histone deacetylation is associated with transcription repression (9, 10). Moreover, HDAC activity is increased in cancer cells and this increase has been shown to induce oncogenic transformation by modulating the function oftranscriptionfactors,suchasp53andnuclearfactor-nB(11), Human Cancer Biology Authors’Affiliation: DepartmentsofMedicine,andPharmacologyandToxicology, Dartmouth-Hitchcock Medical Center, Dartmouth Medical School, Hanover, New Hampshire Received4/17/06;revised8/27/06;accepted10/9/06. Grant support: U.S. Public Health Service grant CA-102687 (M. Korc). Thecostsofpublicationofthisarticleweredefrayedinpartbythepaymentofpage charges.This article must therefore be hereby marked advertisement in accordance with18 U.S.C. Section1734 solely toindicate this fact. Requests for reprints: Murray Korc, Department of Medicine, Dartmouth- Hitchcock Medical Center, Lebanon, NH 03756. Phone: 603-650-7936; Fax:603-650-6122;E-mail:Murray.Korc@Dartmouth.edu. F 2007AmericanAssociationforCancerResearch. doi:10.1158/1078-0432.CCR-06-0914 www.aacrjournals.org Clin Cancer Res 2007;13(1) January1, 2007 18 Research. on May 28, 2017. © 2007 American Association for Cancer clincancerres.aacrjournals.org Downloaded from