pubs.acs.org/JAFC Published on Web 11/17/2010
©2010 American Chemical Society
J. Agric. Food Chem. 2010, 58, 12999–13005 12999
Antitumor Activity of Capsaicin on Human Colon Cancer Cells
in Vitro and Colo 205 Tumor Xenografts in Vivo
HSU-FENG LU,†,‡,§YUAN-LIANG CHEN,§JAI-SING YANG,#YI-YUAN YANG,^
JIA-YOU LIU,†SHU-CHUN HSU,XKUANG-CHI LAI,[,4AND JING-GUNG CHUNG*,b,1
Ecology, Fu-Jen Catholic University, Taipei 242, Taiwan,§Department of Medical Technology, Yuanpei
University, Hsinchu 306, Taiwan,#Department of Pharmacology, China Medical University,
Taichung 404, Taiwan,^Department of Biotechnology, Taipei Medical University, Taipei 110, Taiwan,
XSchool of Nutrition, China Medical University, Taichung 404, Taiwan,[School of Medicine, China
Medical University, Taichung 404, Taiwan,4Department of Surgery, China Medical University Beigang
Hospital, Yunlin 651, Taiwan,bDepartment of Biological Science and Technology, China Medical
University, Taichung 404, Taiwan, and1Department of Biotechnology, Asia University,
Wufeng, Taichung 413, Taiwan
Capsaicin was reported to inhibit cancer cell growth. The aim of this study was to evaluate the
antitumor potential of capsaicin by studying antitumor activity in vitro as well as in vivo. The in vitro
studies are to examine the effects of capsaicin on human colon cancer colo 205 cells after exposure
to capsaicin. The results showed that capsaicin induced cytotoxic effects in a time- and dose-
dependent manner and increased reactive oxygen species (ROS) and Ca2þbut decreased the level
of mitochondrial membrane potential (ΔΨm) in colo 205 cells. Data from Western blotting analysis
indicated that the levels of Fas, cytochrome c, and caspases were increased, leading to cell
apoptosis. Capsaicin decreased the levels of anti-apoptotic proteins such as Bcl-2 and increased the
levels of pro-apoptotic proteins such as Bax. Capsaicin-induced apoptosis in colo 205 cells was also
done through the activations of caspase-8, -9 and -3. In vivo studies in immunodeficient nu/nu mice
bearing colo 205 tumor xenografts showed that capsaicin effectively inhibited tumor growth. The
potent in vitro and in vivo antitumor activities of capsaicin suggest that capsaicin might be developed
for the treatment of human colon cancer.
KEYWORDS: Capsaicin; human colon cancer colo 205 cells; apoptosis; Fas/FasL; caspases activation
Cancer is the leading cause of death all over the world. One in
every four deaths in the United States is from cancer. The
incidence rate of colorectal cancer is 850,000/per year, and it
causes 500,000 deaths in every part of the world (1). For males
and females, the colon/rectum is the third leading site among all
primary sites in Taiwan. The poor prognostic outcome of colo-
rectal cancer is due to its resistance to current therapies, main-
taining it as the leading cause of cancer-related death. Successful
treatment with chemotherapeutic agents is largely dependent on
their ability to trigger cell death in tumor cells. Therefore, novel
inducers of apoptosis provide a new therapeutic approach for
anticancer design. Several previous studies demonstrated that
certain phytochemicals present in medicinal herbs exert antitu-
morigenic activity by inducing apoptosis in cancer cells.
is consumed, there is a very low incidence of gastrointestinal
cancers, including colorectal and stomach cancers, compared to
the rest of Asia, including Japan and China. Capsaicin (trans-8-
methyl-N-vanillyl-6-nonenamide) is widely used as a food addi-
tive and as an analgesic agent. In addition to its well-known role
in nociception, it is mediated by vanilloid receptor 1 specifically
expressed in dorsal root ganglion neurons. Capsaicin has also
been considered as a potential anticancer agent as it inhibits cell
proliferation and induces apoptosis in various types of cancer
cells (2,3). Capsaicin is a major pungent ingredient of red pepper
and isthe activecomponent ofchili peppers,a plant belonging to
the genus Capsicum. Natural capsaicin directly inhibits the
growth of leukemic cells, and this was reported in several clinical
studies conducted in Japan and China. Some studies described
their early experimental findings as showing that daily consump-
tion of hot peppers may actually prevent certain types of cancer,
although these studies used pure capsaicin directly injected into
isolated diseased cells in a laboratory setting (4).
Morre et al. reported that capsaicin acted against certain
carcinogens and mutagens and, especially, was able to kill
prostate cancer cells by causing them to undergo apoptosis (5).
Morre et al. found that capsaicin preferentially repressed the
growth of some transformed human and mouse cells (5,6). Hail
*Address correspondence to this author at the Department of
Biological Science and Technology, China Medical University, No. 91,
Hsueh-Shih Road, Taichung 404, Taiwan (phone þ886 4 2205 3366, ext
2161; fax þ886 4 2205 3764; e-mail firstname.lastname@example.org).
Article J. Agric. Food Chem., Vol. 58, No. 24, 201013005
mic reticulum; DCFH-DA, 20,70-dichlorofluorescin diacetate;
ROS, reactive oxygen species; PI, propidium iodide.
(1) Benson, A.B., 3rd. Epidemiology, diseaseprogression,andeconom-
ic burden of colorectal cancer. J. Manag. Care Pharm. 2007, 13 (6
Suppl. C), S5-S18.
(2) Vatanasapt, V.; Martin, N.; Sriplung, H.; Chindavijak, K.; Sontipong,
S.; Sriamporn, H.; Parkin, D. M.; Ferlay, J. Cancer incidence in
Thailand, 1988-1991. Cancer Epidemiol. Biomarkers Prev. 1995, 4 (5),
(3) Vatanasapt, V.; Sriamporn, S.; Vatanasapt, P. Cancer control in
Thailand. Jpn. J. Clin. Oncol. 2002, 32 (Suppl.), S82-S91.
(4) Ito, K.; Nakazato, T.; Yamato, K.; Miyakawa, Y.; Yamada, T.;
Hozumi, N.; Segawa, K.; Ikeda, Y.; Kizaki, M. Induction of apoptosis
in leukemic cells by homovanillic acid derivative, capsaicin, through
by reactive oxygen species. Cancer Res. 2004, 64 (3), 1071-1078.
(5) Morre, D. J.; Chueh, P. J.; Morre, D. M. Capsaicin inhibits
preferentially the NADH oxidase and growth of transformed cells
in culture. Proc. Natl. Acad. Sci. U.S.A. 1995, 92 (6), 1831-1835.
(6) Morre, D. J.; Sun, E.; Geilen, C.; Wu, L. Y.; de Cabo, R.;
Krasagakis, K.; Orfanos, C. E.; Morre, D. M. Capsaicin inhibits
plasma membraneNADH oxidase and growth of human and mouse
melanoma lines. Eur. J. Cancer 1996, 32A (11), 1995-2003.
(7) Hail, N., Jr.; Lotan, R. Examining the role of mitochondrial
respiration in vanilloid-induced apoptosis. J. Natl. Cancer Inst.
2002, 94 (17), 1281-1292.
(8) Macho, A.; Calzado, M. A.; Munoz-Blanco, J.; Gomez-Diaz, C.;
Gajate, C.; Mollinedo, F.; Navas, P.; Munoz, E. Selective induction
of apoptosis by capsaicin in transformed cells: the role of reactive
oxygen species and calcium. Cell Death Differ. 1999, 6 (2), 155-165.
(9) Macho, A.; Sancho, R.; Minassi, A.; Appendino, G.; Lawen, A.;
Munoz, E. Involvement of reactive oxygen species in capsaicinoid-
induced apoptosis in transformed cells. Free Radical Res. 2003, 37
Tsai, H. Y.; Chen, Y. F.; Chung, J. G. Curcumin-induced cell cycle
arrest and apoptosis in human acute promyelocytic leukemia HL-60
cells via MMP changes and caspase-3 activation. Anticancer Res.
2006, 26 (6B), 4361-4371.
H. J.; Chung, J. G. Crude extracts of Euchresta formosana radix
induce cytotoxicity and apoptosis in human hepatocellular carcino-
ma cell line (Hep3B). Anticancer Res. 2007, 27 (4B), 2415-2425.
W. C.; Chen, G. W. Curcumin-induced apoptosis of human colon
cancer colo 205 cells through the production of ROS, Ca2þand the
activation of caspase-3. Anticancer Res. 2006, 26 (6B), 4379-4389.
(13) Lu, H. F.; Wang, H. L.; Chuang, Y. Y.; Tang, Y. J.; Yang, J. S.; Ma,
Y. S.; Chiang, J. H.; Lu, C. C.; Yang, J. L.; Lai, T. Y.; Wu, C. C.;
Chung, J. G. Danthron induced apoptosis through mitochondria-
and caspase-3-dependent pathways in human brain glioblastoma
multiforms GBM 8401 cells. Neurochem. Res. 2010, 35 (3),390-398.
(14) Wu, P. P.; Kuo, S. C.; Huang, W. W.; Yang, J. S.; Lai, K. C.; Chen,
H. J.; Lin, K. L.; Chiu, Y. J.; Huang, L. J.; Chung, J. G. (-)-
Epigallocatechin gallate induced apoptosis in human adrenal cancer
NCI-H295 cells through caspase-dependent and caspase-indepen-
dent pathway. Anticancer Res. 2009, 29 (4), 1435-1442.
(15) Wen,Y. F.; Yang, J.S.;Kuo, S.C.;Hwang, C.S.;Chung, J.G.;Wu,
H. C.; Huang, W. W.; Jhan, J. H.; Lin, C. M.; Chen, H. J.
Investigation of anti-leukemia molecular mechanism of ITR-284, a
carboxamide analog, in leukemia cells and its effects in WEHI-3
leukemia mice. Biochem. Pharmacol. 2010, 79 (3), 389-398.
(16) Hsia, T. C.; Chung, J. G.; Lu, H. F.; Ho, H. C.; Yang, C. C.; Lu,
K. H.; Hung, C. F. The effect of paclitaxel on 2-aminofluorene-
DNA adducts formation and arylamine N-acetyltransferase activity
and gene expression in human lung tumor cells (A549). Food Chem.
Toxicol. 2002, 40 (5), 697-703.
(17) Ji, B. C.; Hsu, W. H.; Yang, J. S.; Hsia, T. C.; Lu, C. C.; Chiang,
J. H.; Yang, J. L.; Lin, C. H.; Lin, J. J.; Suen, L. J.; Gibson Wood,
W.; Chung, J. G. Gallic acid induces apoptosis via caspase-3 and
mitochondrion-dependent pathways in vitro and suppresses lung
xenograft tumor growth in vivo. J. Agric. Food Chem. 2009, 57 (16),
(18) Ho, Y. T.; Yang, J. S.; Lu, C. C.; Chiang, J. H.; Li, T. C.; Lin, J. J.;
Lai, K. C.; Liao, C. L.; Lin, J. G.; Chung, J. G. Berberine inhib-
its human tongue squamous carcinoma cancer tumor growth
in a murine xenograft model. Phytomedicine 2009, 16 (9), 887-
(19) Dethlefsen, L. A.; Prewitt, J. M.; Mendelsohn, M. L. Analysis of
tumor growth curves. J. Natl. Cancer Inst. 1968, 40 (2), 389-405.
(20) Di Giovanni, S.; Mirabella, M.; Papacci, M.; Odoardi, F.; Silvestri,
with cytochrome c oxidase deficiency in mitochondrial encephalo-
myopathies. Mol. Cell. Neurosci. 2001, 17 (4), 696-705.
(21) Lee, M. G.; Lee, K. T.; Chi, S. G.; Park, J. H. Costunolide induces
apoptosis by ROS-mediated mitochondrial permeability transi-
tion and cytochrome C release. Biol. Pharm. Bull. 2001, 24 (3),
(22) Chen, M.; Wang, J. Initiator caspases in apoptosis signaling path-
ways. Apoptosis 2002, 7 (4), 313-319.
(23) Budihardjo, I.; Oliver, H.; Lutter, M.; Luo, X.; Wang, X. Biochem-
ical pathways of caspase activation during apoptosis. Annu. Rev.
Cell. Dev. Biol. 1999, 15, 269-290.
(24) Rao, R. V.; Ellerby, H. M.; Bredesen, D. E. Coupling endoplasmic
(25) Salvesen, G. S.; Duckett, C. S. IAP proteins: blocking the road to
death’s door. Nat. Rev. Mol. Cell. Biol. 2002, 3 (6), 401-410.
(26) Earnshaw, W. C.; Martins, L. M.; Kaufmann, S. H. Mammalian
caspases: structure, activation, substrates, and functions during
apoptosis. Annu. Rev. Biochem. 1999, 68, 383-424.
(27) Strasser, A.; O’Connor, L.; Dixit, V. M. Apoptosis signaling. Annu.
Rev. Biochem. 2000, 69, 217-245.
(28) Rao, R. V.; Hermel, E.; Castro-Obregon, S.; del Rio, G.; Ellerby,
L. M.; Ellerby, H. M.; Bredesen, D. E. Coupling endoplasmic
reticulum stress to the cell death program. Mechanism of caspase
activation. J. Biol. Chem. 2001, 276 (36), 33869-33874.
(29) Antonsson, B. Bax and other pro-apoptotic Bcl-2 family “killer-
proteins” and their victim the mitochondrion. Cell Tissue Res. 2001,
306 (3), 347-361.
(30) Vaux, D. L.; Korsmeyer, S. J. Cell death in development. Cell 1999,
96 (2), 245-254.
(31) Meek, D. W. Multisite phosphorylation and the integration of stress
signals at p53. Cell Signal. 1998, 10 (3), 159-166.
(32) Ryan, K. M.; Phillips, A. C.; Vousden, K. H. Regulation and
function of the p53 tumor suppressor protein. Curr. Opin. Cell Biol.
2001, 13 (3), 332-337.
(33) Miyashita, T.; Reed, J. C. Tumor suppressor p53 is a direct
transcriptional activator of the human bax gene. Cell 1995, 80 (2),
(34) el-Deiry, W. S. p21/p53, cellular growth control and genomic
integrity. Curr. Top. Microbiol. Immunol. 1998, 227, 121-137.
(35) Mori, A.; Lehmann, S.; O’Kelly, J.; Kumagai, T.; Desmond, J. C.;
Pervan, M.; McBride, W. H.; Kizaki, M.; Koeffler, H. P. Capsaicin,
a component of red peppers, inhibits the growth of androgen-
independent, p53 mutant prostate cancer cells. Cancer Res. 2006,
66 (6), 3222-3229.
(36) Zhang, R.; Humphreys, I.; Sahu, R. P.; Shi, Y.; Srivastava, S. K. In
vitro and in vivo induction of apoptosis by capsaicin in pancreatic
cancer cells is mediated through ROS generation and mitochondrial
death pathway. Apoptosis 2008, 13 (12), 1465-1478.
Received for review August 27, 2010. Revised manuscript received
October 14, 2010. Accepted October 19, 2010. This work was
supported by Grant CMU98-ASIA-10 from China Medical University,