INTERNATIONAL JOURNAL OF ONCOLOGY 41: 1513-1519, 2012
Abstract. Quinazolinone compounds have been shown to
have antitumor activity in many human cancer cell lines. In
the present study, we investigated the anti-metastatic activity
of MJ-33 (2-(3-ethoxyphenyl)-6-pyrrolidinylquinazolinone),
a novel quinazolinone derivate, and the signaling pathway
of MJ-33 in human prostate cells. MJ-33 exhibited a growth
inhibitory effect on DU145, LNCaP and PC-3 cells by MTT
assay. DU145 cells showed greater sensitivity to the growth
inhibition of MJ-33 than that of LNCaP and PC-3 cells. MJ-33
also had an inhibitory effect on the invasion, migration and
adhesion of DU145 cells using Boyden chamber transwell
assays, wound-healing and adhesion assay. In addition, MJ-33
inhibited cell metastasis through the reduction of matrix metal-
loproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9)
and urokinase-type plasminogen activator (u-PA) enzyme
activities and protein levels by gelatin zymography assay and
western blot analysis, respectively. MJ-33 reduced the protein
levels of p-JNK, p-p38, p-ERK, p-AKT and nuclear NF-κB
(p65), c-fos and c-Jun protein levels by western blotting. Using
electrophoretic mobility-shift assay (EMSA), we demonstrated
that MJ-33 blocked the activation of transcription factor AP-1
(activator protein-1) and NF-κB, which led to the inhibition of
MMP-2 and MMP-9 expression. Collectively, our data showed
that MJ-33 decreased protein levels of MAPKs (mitogen-
activated protein kinases), AKT, AP-1 and NF-κB, resulting in
the inhibition of matrix metalloproteinases. Downregulation
of MMP-2 and MMP-9 reduces the invasion, migration and
adhesion activities of DU145 cells. MJ-33 may be a promising
agent against prostate cancer metastasis.
Prostate cancer is one of the leading causes of mortality in
men (1) and has a highly variable natural history. Prostate
cancer may be present as an indolent and silent entity
throughout a man's life and then grow rapidly following
metastasis to the lymph nodes and bones with a median life
expectancy of 24-36 months (2). Therefore, prostate cancer
is largely asymptomatic until metastases are present and it
is largely a disease of the elderly. It is reasonable to propose
that agents which inhibit metastasis could have great thera-
Quinazoline derivatives are known for multiple effects,
such as anti-malarial, anti-inflammatory (3), anti-bacterial,
and antitumor activities (4). In recent years, we have designed
and synthesized a series of quinazoline derivatives as new
anti-mitotic agents (5,6). Our previous study showed that
synthesized 6-pyrrolidinyl-4-quinazolinone derivative MJ-29
inhibited tubulin polymerization through binding to β-tubulin
at the colchicine-binding site and acted as an anti-mitotic
agent (5). Furthermore, we demonstrated that 6-fluoro-
exhibits anti-metastatic effects in human osteosarcoma U-2 OS
cells through targeting the insulin-like growth factor-I receptor
(IGF-IR) (6). In the present study, we investigated the effects of
MJ-33 on invasion, migration and adhesion in DU145 human
prostate cancer cells. MJ-33 inhibited migration and invasion
through downregulation of matrix metalloproteinases (MMPs)
and urokinase-type plasminogen activator (u-PA) through the
AP-1 and NF-κB signaling pathways.
Antitumor effects of the novel quinazolinone MJ-33: Inhibition
of metastasis through the MAPK, AKT, NF-κB and AP-1
signaling pathways in DU145 human prostate cancer cells
MANN-JEN HOUR1*, SHIH-CHANG TSAI3*, HSI-CHIN WU7,2, MENG-WEI LIN3, JING-GUNG CHUNG3,
JIN-BIN WU1,5,6, JO-HUA CHIANG8, MINORU TSUZUKI9,6 and JAI-SING YANG2,4
Schools of 1Pharmacy, and 2Medicine, China Medical University, Taichung 404; Departments of 3Biological
Science and Technology, and 4Pharmacology, China Medical University, Taichung 404; 5Graduate Institute
of Pharmaceutical Chemistry, China Medical University, Taichung 404; 6Tsuzuki Institute for Traditional
Medicine, China Medical University, Taichung 404; 7Department of Urology, China Medical University
Hospital, Taichung 404; 8Department of Life Sciences, National Chung Hsing University, Taichung 402, Taiwan,
R.O.C.; 9Department of Biochemistry, Nihon Pharmaceutical University, Saitama 362-0806, Japan
Received April 24, 2012; Accepted June 20, 2012
Correspondence to: Dr Jai-Sing Yang, Department of Pharmacology,
China Medical University, No. 91 Hsueh-Shih Road, Taichung 40402,
Key words: quinazolinone MJ-33, invasion, migration and adhesion,
MAPK, AKT, NF-κB and AP-1 signaling pathways, human prostate
cancer DU145 cells
INTERNATIONAL JOURNAL OF ONCOLOGY 41: 1513-1519, 2012
1. Jemal A, Siegel R, Ward E, Murray T, Xu J and Thun MJ: Cancer
statistics, 2007. CA Cancer J Clin 57: 43-66, 2007.
2. Petrylak DP: The current role of chemotherapy in metastatic
hormone-refractory prostate cancer. Urology 65: 3-8, 2005.
3. Chandrika PM, Yakaiah T, Rao AR, et al: Synthesis of novel
4,6-disubstituted quinazoline derivatives, their anti-inflamma-
tory and anti-cancer activity (cytotoxic) against U937 leukemia
cell lines. Eur J Med Chem 43: 846-852, 2008.
4. Chen Z, Huang X, Yang H, et al: Anti-tumor effects of B-2, a
novel 2,3-disubstituted 8-arylamino-3H-imidazo[4,5-g]quinazo-
line derivative, on the human lung adenocarcinoma A549 cell
line in vitro and in vivo. Chem Biol Interact 189: 90-99, 2011.
5. Yang JS, Hour MJ, Huang WW, Lin KL, Kuo SC and Chung JG:
MJ-29 inhibits tubulin polymerization, induces mitotic arrest, and
triggers apoptosis via cyclin-dependent kinase 1-mediated Bcl-2
phosphorylation in human leukemia U937 cells. J Pharmacol
Exp Ther 334: 477-488, 2010.
6. Chen KT, Hour MJ, Tsai SC, et al: The novel synthesized
(LJJ-10) compound exhibits anti-metastatic effects in human
osteosarcoma U-2 OS cells through targeting insulin-like growth
factor-I receptor. Int J Oncol 39: 611-619, 2011.
7. Liu KC, Huang AC, Wu PP, et al: Gallic acid suppresses the
migration and invasion of PC-3 human prostate cancer cells
via inhibition of matrix metalloproteinase-2 and -9 signaling
pathways. Oncol Rep 26: 177-184, 2011.
8. Ma CY, Ji WT, Chueh FS, et al: Butein inhibits the migration
and invasion of SK-HEP-1 human hepatocarcinoma cells through
suppressing the ERK, JNK, p38, and uPA signaling multiple
pathways. J Agric Food Chem 59: 9032-9038, 2011.
9. Lan YH, Wu YC, Wu KW, et al: Death receptor 5-mediated
TNFR family signaling pathways modulate gamma-humulene-
induced apoptosis in human colorectal cancer HT29 cells. Oncol
Rep 25: 419-424, 2011.
10. Huang WW, Ko SW, Tsai HY, et al: Cantharidin induces G2/M
phase arrest and apoptosis in human colorectal cancer colo 205
cells through inhibition of CDK1 activity and caspase-dependent
signaling pathways. Int J Oncol 38: 1067-1073, 2011.
11. Yu FS, Yang JS, Yu CS, et al: Safrole induces apoptosis in human
oral cancer HSC-3 cells. J Dent Res 90: 168-174, 2011.
12. Lu CC, Yang JS, Huang AC, et al: Chrysophanol induces
necrosis through the production of ROS and alteration of ATP
levels in J5 human liver cancer cells. Mol Nutr Food Res 54:
13. Chiang JH, Yang JS, Ma CY, et al: Danthron, an anthraquinone
derivative, induces DNA damage and caspase cascades-mediated
apoptosis in SNU-1 human gastric cancer cells through mito-
chondrial permeability transition pores and Bax-triggered
pathways. Chem Res Toxicol 24: 20-29, 2011.
14. Huang WW, Chiu YJ, Fan MJ, et al: Kaempferol induced
apoptosis via endoplasmic reticulum stress and mitochondria-
dependent pathway in human osteosarcoma U-2 OS cells. Mol
Nutr Food Res 54: 1585-1595, 2010.
15. Danthron. Rep Carcinog 12: 128-129, 2011.
16. Lo C, Lai TY, Yang JS, et al: Gallic acid inhibits the migration
and invasion of A375.S2 human melanoma cells through the
inhibition of matrix metalloproteinase-2 and Ras. Melanoma Res
21: 267-273, 2011.
17. Kuo TC, Yang JS, Lin MW, et al: Emodin has cytotoxic and
protective effects in rat C6 glioma cells: roles of Mdr1a and
nuclear factor kappaB in cell survival. J Pharmacol Exp Ther
330: 736-744, 2009.
18. Wang SW, Pan SL, Huang YC, et al: CHM-1, a novel synthetic
quinolone with potent and selective antimitotic antitumor activity
against human hepatocellular carcinoma in vitro and in vivo. Mol
Cancer Ther 7: 350-360, 2008.
19. Hsu SC, Yang JS, Kuo CL, et al: Novel quinolone CHM-1
induces apoptosis and inhibits metastasis in a human osterogenic
sarcoma cell line. J Orthop Res 12: 1637-1644, 2009.
20. Gondi CS and Rao JS: Therapeutic potential of siRNA-mediated
targeting of urokinase plasminogen activator, its receptor, and
matrix metalloproteinases. Methods Mol Biol 487: 267-281,
21. Chen PN, Hsieh YS, Chiou HL and Chu SC: Silibinin inhibits
cell invasion through inactivation of both PI3K-Akt and MAPK
signaling pathways. Chem Biol Interact 156: 141-150, 2005.
22. Westermarck J and Kahari VM: Regulation of matrix metallo-
proteinase expression in tumor invasion. FASEB J 13: 781-792,
23. Aguirre Ghiso JA, Alonso DF, Farias EF, Gomez DE and de Kier
Joffe EB: Deregulation of the signaling pathways control-
ling urokinase production. Its relationship with the invasive
phenotype. Eur J Biochem 263: 295-304, 1999.
24. Peng PL, Hsieh YS, Wang CJ, Hsu JL and Chou FP: Inhibitory
effect of berberine on the invasion of human lung cancer cells via
decreased productions of urokinase-plasminogen activator and
matrix metalloproteinase-2. Toxicol Appl Pharmacol 214: 8-15,
25. Deryugina EI and Quigley JP: Matrix metalloproteinases and
tumor metastasis. Cancer Metastasis Rev 25: 9-34, 2006.
26. Lin YT, Yang JS, Lin HJ, et al: Baicalein induces apoptosis in
SCC-4 human tongue cancer cells via a Ca2+-dependent mito-
chondrial pathway. In Vivo 21: 1053-1058, 2007.
27. Chuang JY, Huang YF, Lu HF, et al: Coumarin induces cell
cycle arrest and apoptosis in human cervical cancer HeLa cells
through a mitochondria- and caspase-3 dependent mechanism
and NF-kappaB down-regulation. In Vivo 21: 1003-1009, 2007.
28. Huang YT, Hwang JJ, Lee LT, et al: Inhibitory effects of a lutein-
izing hormone-releasing hormone agonist on basal and epidermal
growth factor-induced cell proliferation and metastasis-associ-
ated properties in human epidermoid carcinoma A431 cells. Int J
Cancer 99: 505-513, 2002.