Mortality and morbidity associated with oral squamous cell carcinoma (OSCC) remain unacceptably high with disfiguring treatment options and a death rate of 1 per hour in the United States. The approval of cituximab for advanced OSCC has been the only new treatment for these patients since the 1970s, although it has not significantly increased overall survival. To address the paucity of effective new therapies, we undertook a high-throughput screen to discover small molecules and natural products that could induce endoplasmic reticulum (ER) stress and enforce a terminal unfolded protein response (UPR) in OSCC. The terpenoid cantharidin (CNT), previously used to treat various malignancies in culture-specific medical practices for over 2,000 y, emerged as a hit. CNT and its analog, cantharidic acid, potently induced protein and gene expression profiles consistent with the activation of ER stress, the UPR, and apoptosis in OSCC cells. Murine embryonic fibroblasts null for the UPR-associated transcription factors Atf4 or Chop were significantly protected from CNT, implicating a key role for the UPR in the death response. These data validate that our high-throughput screen can identify novel modulators of UPR signaling and that such compounds might provide a new therapeutic approach to treating patients with OSCC.
CrewJPFuggleSBicknellRCranstonDWde BenedettiAHarrisAL. 2000. Eukaryotic initiation factor-4E in superficial and muscle invasive bladder cancer and its correlation with vascular endothelial growth factor expression and tumour progression. Br J Cancer. 82(1):161–166.
2.
DeFattaRJNathanCODe BenedettiA. 2000. Antisense RNA to eIF4E suppresses oncogenic properties of a head and neck squamous cell carcinoma cell line. Laryngoscope. 110(6):928–933.
3.
DengLDongJWangW. 2013. Exploiting protein phosphatase inhibitors based on cantharidin analogues for cancer drug discovery. Mini Rev Med Chem. 13(8):1166–1176.
4.
EfferthTDaveyMOlbrichARuckerGGebhartEDaveyR. 2002. Activity of drugs from traditional Chinese medicine toward sensitive and MDR1- or MRP1-overexpressing multidrug-resistant human CCRF-CEM leukemia cells. Blood Cells Mol Dis. 28(2):160–168.
5.
EfferthTRauhRKahlSTomicicMBöchzeltHTomeMEBriehlMMBauerRKainaB. 2005. Molecular modes of action of cantharidin in tumor cells. Biochem Pharmacol. 69(5):811–818.
6.
FranklinSPhoTAbreoFWNassarRDe BenedettiAStuckerFJNathanCO. 1999. Detection of the proto-oncogene eIF4E in larynx and hypopharynx cancers. Arch Otolaryngol Head Neck Surg. 125(2):177–182.
7.
FribleyAMCruzPGMillerJRCallaghanMUCaiPNarulaNNeubigRRShowalterHDLarsenSDKirchhoffPD. 2011. Complementary cell-based high-throughput screens identify novel modulators of the unfolded protein response. J Biomol Screen. 16(8):825–835.
8.
FribleyAMEvenchikBZengQParkBKGuanJYZhangHHaleTJSoengasMSKaufmanRJWangCY. 2006. Proteasome inhibitor PS-341 induces apoptosis in cisplatin-resistant squamous cell carcinoma cells by induction of Noxa. J Biol Chem. 281(42):31440–31447.
9.
FribleyAMMillerJRBrownellALGarshottDMZengQReistTENarulaNCaiPXiYCallaghanMU. 2015. Celastrol induces unfolded protein response-dependent cell death in head and neck cancer. Exp Cell Res. 330(2):412–422.
10.
FribleyAZengQWangCY. 2004. Proteasome inhibitor PS-341 induces apoptosis through induction of endoplasmic reticulum stress-reactive oxygen species in head and neck squamous cell carcinoma cells. Mol Cell Biol. 24(22):9695–9704.
11.
HanJBackSHHurJLinYHGildersleeveRShanJYuanCLKrokowskiDWangSHatzoglouM. 2013. ER-stress-induced transcriptional regulation increases protein synthesis leading to cell death. Nat Cell Biol. 15(5):481–490.
12.
HonkanenRE. 1993. Cantharidin, another natural toxin that inhibits the activity of serine/threonine protein phosphatases types 1 and 2A. FEBS Lett. 330(3):283–286.
13.
KacarNTasliLKorkmazSErginSErdoganBS. 2012. Cantharidin-podophylotoxin-salicylic acid versus cryotherapy in the treatment of plantar warts: a randomized prospective study. J Eur Acad Dermatol Venereol. 26(7):889–893.
14.
KadiogluOKermaniNSKelterGSchumacherUFiebigHHGretenHJEfferthT. 2014. Pharmacogenomics of cantharidin in tumor cells. Biochem Pharmacol. 87(3):399–409.
15.
KawadaMAmemiyaMIshizukaMTakeuchiT. 1999. Cytostatin, an inhibitor of cell adhesion to extracellular matrix, selectively inhibits protein phosphatase 2A. Biochim Biophys Acta. 1452(2):209–217.
16.
KerekatteVSmileyKHuBSmithAGelderFDe BenedettiA. 1995. The proto-oncogene/translation factor eIF4E: a survey of its expression in breast carcinomas. Int J Cancer. 64(1):27–31.
17.
KokSHHongCYKuoMYLeeCHLeeJJLouIULeeMSHsiaoMLinSK. 2003. Comparisons of norcantharidin cytotoxic effects on oral cancer cells and normal buccal keratinocytes. Oral Oncol. 39(1):19–26.
18.
LiWChenZZongYGongFZhuYZhuYLvJZhangJXieLSunY. 2011. PP2A inhibitors induce apoptosis in pancreatic cancer cell line PANC-1 through persistent phosphorylation of IKKalpha and sustained activation of the NF-kappaB pathway. Cancer Lett. 304(2):117–127.
19.
LiWXieLChenZZhuYSunYMiaoYXuZHanX. 2010. Cantharidin, a potent and selective PP2A inhibitor, induces an oxidative stress-independent growth inhibition of pancreatic cancer cells through G2/M cell-cycle arrest and apoptosis. Cancer Sci. 101(5):1226–1233.
20.
LiXZhangKLiZ. 2011. Unfolded protein response in cancer: the physician’s perspective. J Hematol Oncol. 4:8.
21.
LiYMCasidaJE. 1992. Cantharidin-binding protein: identification as protein phosphatase 2A. Proc Natl Acad Sci U S A. 89(24):11867–11870.
22.
MaYHendershotLM. 2004. The role of the unfolded protein response in tumour development: friend or foe?Nat Rev Cancer. 4(12):966–977.
23.
NathanCOLiuLLiBDAbreoFWNandyIDe BenedettiA. 1997. Detection of the proto-oncogene eIF4E in surgical marginsmaypredict recurrence in head and neck cancer. Oncogene. 15(5):579–584.
24.
Puerto GalvisCEVargas MéndezLYKouznetsovVV. 2013. Cantharidin-based small molecules as potential therapeutic agents. Chem Biol Drug Des. 82(5):477–499.
25.
RauhRKahlSBoechzeltHBauerRKainaBEfferthT. 2007. Molecular biology of cantharidin in cancer cells. Chin Med. 2:8.
26.
SakoffJAAcklandSPBaldwinMLKeaneMAMcCluskeyA. 2002. Anticancer activity and protein phosphatase 1 and 2A inhibition of a new generation of cantharidin analogues. Invest New Drugs. 20(1):1–11.
27.
ScheunerDSongBMcEwenELiuCLaybuttRGillespiePSaundersTBonner-WeirSKaufmanRJ. 2001. Translational control is required for the unfolded protein response and in vivo glucose homeostasis. Mol Cell. 7(6):1165–1176.
28.
ShanHBCaiYCLiuYZengWNChenHXFanBTLiuXHXuZLWangBXianLJ. 2006. Cytotoxicity of cantharidin analogues targeting protein phosphatase 2A. Anticancer Drugs. 17(8):905–911.
SingletonDCHarrisAL. 2012. Targeting the ATF4 pathway in cancer therapy. Expert Opin Ther Targets. 16(12):1189–1202.
31.
SmithEMRitchieJMSummersgillKFHoffmanHTWangDHHaugenTHTurekLP. 2004. Human papillomavirus in oral exfoliated cells and risk of head and neck cancer. J Natl Cancer Inst. 96(6):449–455.
32.
TillJSMajmudarBN. 1981. Cantharidin poisoning. South Med J. 74(4):444–447.
33.
TsengIJSheuSYChenYTHuangCYLinCTLinPY. 2012. A simple procedure for preparation of N-thiazol, thiadiazol, pyridyl and sulfanylamidocantharidinimines analogues and evaluation of their cytotoxicities against human HL-60, MCF7, Neuro-2a and A549 carcinoma cell. Chem Pharm Bull (Tokyo). 60(11):1453–1457.
34.
VermaAKPrasadSB. 2013. Changes in glutathione, oxidative stress and mitochondrial membrane potential in apoptosis involving the anticancer activity of cantharidin isolated from redheaded blister beetles, Epicauta hirticornis. Anticancer Agents Med Chem. 13(7):1096–1114.
35.
WadaSUsamiIUmezawaYInoueHOhbaSSomenoTKawadaMIkedaD. 2010. Rubratoxin A specifically and potently inhibits protein phosphatase 2A and suppresses cancer metastasis. Cancer Sci. 101(3):743–750.
36.
WangGS. 1989. Medical uses of mylabris in ancient China and recent studies. J Ethnopharmacol. 26(2):147–162.
37.
WangQMora-JensenHWenigerMAPerez-GalanPWolfordCHaiTRonDChenWTrenkleWWiestnerA. 2009. ERAD inhibitors integrate ER stress with an epigenetic mechanism to activate BH3-only protein NOXA in cancer cells. Proc Natl Acad Sci U S A. 106(7):2200–2205.
38.
WangSKaufmanRJ. 2012. The impact of the unfolded protein response on human disease. J Cell Biol. 197(7):857–867.
39.
YiSNWassJVincentPIlandH. 1991. Inhibitory effect of norcantharidin on K562 human myeloid leukemia cells in vitro. Leuk Res. 15(10):883–886.
40.
ZhangZKelemenOvan SantenMAYeltonSMWendlandtAESviripaVMBollenMBeullensMUrlaubHLührmannR. 2011. Synthesis and characterization of pseudocantharidins, novel phosphatase modulators that promote the inclusion of exon 7 into the SMN (survival of motoneuron) pre-mRNA. J Biol Chem. 286(12):10126–10136.
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