Santosh K. Katiyar, Ph.D. - University of Alabama at Birmingham

September 11, 2006; Botanicals Workshop
Animal models of skin carcinogenesis:
Green tea prevents skin cancer through
DNA-repair mechanism
Santosh K. Katiyar, Ph.D.
University of Alabama at Birmingham,
& Birmingham VA Medical Center, AL

• There are more than 3000 different diseases
or disorders of the skin, hair and nails, which
in an average year affect about 60 million
Americans.
• The combined annual cost to society of
medical care and lost wages from these
conditions is estimated to be in the billions.

• The skin is one of the largest organs in the
body, having enormous surface area ranging
from 1.5-2.0m 2 and making up about 16% of
body weight.
• Skin is constantly exposed to chemical agents
and other environmental pollutants. National
Institute of Occupational Safety and Health
(NIOSH) characterized skin diseases as one of
the most pervasive occupational health
problems in the Unites States. Data from the
Bureau of Labor Statistics indicate that skin
diseases attributed to work place exposures
accounts for more than 30% of all reported
occupational diseases.

Cancer Statistics
• The risk of cancer is a growing health problem
around the world particularly associated with the
constantly rise in life expectancy, changes in
environmental conditions, dietary habits and
lifestyle.
• According to current projections, one in five
Americans will develop at least one nonmelanoma
skin cancer during their life-time.
• Approximately 1.3 million new cases of skin
cancers are diagnosed each year in the United
States, which is equivalent to the incidence of
malignancies in all other organs combined.

Causes of Skin Cancer
• Following major factors contribute to the
skin cancer incidence:
– Heredity characters, e.g. Mutations like
addition, deletion, and substitution etc. in
genome.
– Environmental factors, e.g. UV radiation,
polycyclic aromatic hydrocarbon (PAH) or
toxic chemicals
– Dietary factors, e.g. diet containing toxic
ingredients, charcoal roasted meat etc.

Type of Skin Cancer/Carcinogenesis:
Animal Models
• Chemical carcinogenesis: Initiated by exposure to
chemical carcinogens like ubiquitous environmental
pollutant, benzo(a)pyrene [B(a)P] and/or 7,12-
dimethylbenz(a)anthracene (DMBA). Following
carcinogen exposure, tumor promoters may be
required to speed up the process of carcinogenesis.
– Tumor promoters
• TPA (12-O-tetradecanoylphorbol-13-acetate)
• Mezerein
• Benzoylperoxide (BPO)
• Photocarcinogenesis: Initiated and/or promoted by
ultraviolet (UV) radiation. UV is also a complete
carcinogen.

Multi-stage carcinogenesis model showing
initiation, promotion and progression stages
Sensitive to carcinogen (SENCAR) mouse model for chemical carcinogenesis.
SKH-1 hairless mouse model for photocarcinogenesis

Multi-stage carcinogenesis model leading to the development of
papillomas and subsequent progression to carcinomas.
Initiation
Promotion
Relative Tumor Yield
Papilloma
Carcinoma
+ +
_ _
++ + ++
+ ++ + +
++ +
_
+
_
_ _
Application of large dose of
DMBA
Application of small dose of
DMBA
Application of
TPA

UV Radiation and Facts
• Although several environmental, genetic and dietary
factors contribute to the development of skin cancers,
the most important is the chronic exposure to solar UV
radiation.
• Epidemiological, clinical and experimental biological
studies have shown that solar UV radiation is the major
etiologic agent in skin cancer development.
• UV radiation, particularly UVB (290-320 nm) spectrum,
acts as a tumor initiator, tumor promoter as well as a
complete carcinogen.
• UV irradiation to skin generates inflammatory responses,
oxidative stress, immune suppression and DNA damage
etc.. These factors contribute to various skin disorders
including high risk of skin cancer incidence.

Risk Factors
• Risk factors are well defined for nonmelanoma skin cancer.
Over-exposure to UV Radiation and fair skin type
susceptible to sunburns are the predominant risk factors.
Increasing frequency of exposure, age, and male gender also
contribute to increased risk.
• The incidence of skin cancers, especially SCCs, is also
increased among organ transplant recipients. Kidney and
heart transplant recipients showed a 66-fold increased risk
of SCC compared with the general population.
• A comprehensive study of 5,356 transplant recipients in
Sweden showed a 100-fold increased relative risk of
nonmelanoma skin cancer, almost exclusively in sunexposed
areas. The increased frequency of SCC in these
patients presumably attributable to long-term
immunosuppressive therapy, although non-immune
mechanisms may also play a role.

Dietary supplements may have the ability
to protect the skin from skin diseases.

Botanical Supplements
• Epicatechins or polyphenols from green
tea (Camellia sinensis)
• Grape seed proanthocyanidins(Vitis
vinifera)
• Silymarin from the seeds of milk thistle
(Silybum marianum)

Topical application of (-)-epigallocatechin-3-
gallate (EGCG) from green tea prevents
photocarcinogenesis in mice.
NORMAL UVB-Alone EGCG+UVB
Neoplasia, 5: 555-565, 2003

• EGCG prevents UVB-induced immune
suppression, and UVB-induced immune
suppression is considered as a risk factor for the
development of non-melanoma skin cancer.
• We have also shown that inhibition of UVBinduced
immunosuppression in mice by EGCG is
associated with the increased levels of IL-12.
Carcinogenesis, 20: 2117-2124, 1999.
• IL-12 has anti-tumor activity and DNA repair
ability. Therefore,

Hypothesis
• We hypothesize that chemopreventive
effect of EGCG against photocarcinogenesis
is mediated through IL-12.

EGCG inhibits UVB-induced tumor incidence in
WT mice but not in IL-12 KO mice
% Mice with tumors
100
80
60
40
20
0
Wild-Types
UVB Alone
EGCG+UVB
40%
(p

EGCG inhibits UVB-induced tumor multiplicity
in WT mice but not in IL-12 KO mice
No. of tumors/group
80
60
40
20
0
Wild-Types
UVB Alone
EGCG+UVB
20 25 30 35
Weeks
62%
p

EGCG inhibits tumor growth greater
in WT than in IL-12 KO mice
Tumor vol./tumor
±SD (mm 3 )
200
150
100
50
0
Wild-Types
81%
UVB EGCG+UV
200
150
100
50
0
IL-12 KO
39%
UVB EGCG+UV

EGCG inhibits tumor growth greater
in WT than in IL-12 KO mice
Tumor vol./tumor
±SD (mm 3 )
200
150
100
50
0
Wild-Types
81%
UVB EGCG+UV
200
150
100
50
0
IL-12 KO
39%
UVB EGCG+UV

EGCG reduces UV-induced CPDs rapidly in
WT mice but not in IL-12 KO mice
WT
IL-12 KO
Normal
Control
EGCG
Control
EGCG
UVB-24 h
UVB-½ h

EGCG reduces or repairs UV-induced CPDs
rapidly in WT mice than in IL-12 KO mice
% CPD + Cells
100
80
60
40
20
0
WT
* *
½ h 24 h 48 h ½ h 24 h 48 h
UVB
EGCG+UVB
*P

EGCG reduces or repairs UV-induced CPDs
rapidly in WT mice than in IL-12 KO mice
UVB
EGCG
Normal
Wild-Types IL-12 KO
+ + + +
- + - +
½h
24 h

UV-induced Sunburn Cells
• The formation of sunburn cells after UV
irradiation is primarily a consequence of DNA
damage.
• We therefore investigated the possibility
whether EGCG has the ability to stimulate
DNA repair in sunburn cells and that it is
mediated through IL-12?

EGCG repairs UVB-induced sunburn cells
rapidly in wild-types than IL-12 knock out mice
Percent sunburn cells
1
0.8
0.6
0.4
0.2
0
Wild-types
UVB
EGCG + UVB
*
*
*
0 10 24 48
Hrs. after UVB exposure
*P

EGCG Repairs UVB-induced CPDs in XPA-
Proficient Cells but not in XPA-Deficient Cells
XPAproficient
XPAdeficient
Control
UVB
EGCG +
UVB
24 hr post UV irradiation
% CPD+ cells
100
80
60
40
20
0
100
80
60
40
20
0
UVB
p

EGCG repairs UVB-induced CPDs in XPAproficient
cells but not in XPA-deficient cells
EGCG
UVB
XPAdeficient
XPAproficient
Dot-blot Analysis
- + - +
- - + +
24 hr post UVB irradiation

Conclusion
• EGCG from green tea has the ability to
enhance the repair of UVB-induced DNA
damage through induction of IL-12, which may
lead to prevention of photocarcinogenesis.
• Although it is possible that not all cells with
UVB-induced DNA damage may be repaired
or eliminated using EGCG, the reduction in
UVB-induced DNA damage by EGCG should
be beneficial.

Financial Support
• National Cancer Institute (NCI)
• National Institute of Environmental Health
Sciences (NIEHS).
• NCCAM/NIH
• Cancer Research & Prevention Foundation
• Veterans Administration
• UAB/Purdue Botanical Center
• UAB Skin Diseases Research Center
• Mitsui Norin Co. Ltd., Japan

Working Support
• Anshu Mittal, Ph.D.
• Praveen Vayalil, Ph.D.
• Manjeshwar Baliga, Ph.D.
• Sudheer Mantena, Ph.D.
• Som D. Sharma, Ph.D.
• Syed M. Meeran, Ph.D.
• Suchitra Katiyar, MPH
• Suhail Akhtar, Ph.D.

Thanks for
kind attention !