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Transcript of Cancer Bio FINAL PAPER
Chemopreventive Effects of Lycopene and Other Various Carotenoids
Daniel Etter; A44073265
Dr. Bello-DeOcampo
Cancer Biology – ZOL 450
4/23/2015
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Table of Contents
Abstract…………………………………………………………………………………………………………3
Key Words……………………………………………………………………………………………………..3
Abbreviations………………………………………………………………………………………………...5
Objectives………………………………………………………………………………………………………6
Introduction…………………………………………………………………………………………………...7
Background……………………………...…………………………………………………………………….8
Reactive Oxygen Species and Oxidative Stress……………………………………….8
Antioxidants and Oxidative Stress………………………………………………………...9
Carotenoids…………………………………………………………………………………………10
Literature Review…………………………………………………………………………………………...11
-Carotene and -Carotene…………………………………………………………………..11
Lycopene…………………………………………………………….……………………………….15
-Cryptoxanthin………………………………………………………………………………......17
Lutein……………………….…………………………………………………………...…………….18
Discussion…………………………………………………………………………………………………..…...18
Dietary Risk Factors of Carcinogenesis………………………………………………….18
-Carotene and -Carotene…………………………………………………………………..20
Lycopene…………………………………………………………………………………………….22
-Cryptoxanthin………………………………………………………………………………….24
Lutein…………………………………………………………………………………………………24
Conclusions and Significance………………………………………………………………………….25
Figures………………………………………………………………………………………………………….27
References……………………………………………………………………………………………………..32
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Abstract
It is well documented that lifestyle factors such as an individual’s diet and smoking
are accountable for a large majority of cancers, so it is important to acknowledge this and
learn what you can do to improve your chances. Reactive oxygen species (ROS) are a group
of reactive molecules that are a byproduct of normal metabolism, but are commonly
overproduced in cells. ROS when overproduced have the ability to oxidize/damage
macromolecules such as deoxyribonucleic acid (DNA), lipids and proteins, which can
eventually lead to the development of cancer. They typical Western diet is high in refined
sugars and trans fatty acids, which employ multiple mechanisms that aid in the process of
carcinogenesis. Diets high in fruits and vegetables have been associated with a lower
incidence of cancer, and one group of molecules found in them that seem to possess
anticarcinogenic qualities are carotenoids. -Carotene showed to be approximately 10x
more potent of an inhibitor of proliferation than -carotene. Both and -carotene were
able to induce similar inhibition of invasion and migration, primarily through the decrease
in expression of matrix metalloproteinase 2/7/9 and the increase in the expression of the
tissue inhibitor of MMP (TIMP) 1 and 2. -Carotene has been shown to act as both a
procarcinogen and anticarcinogen, depending on whether or not the individual has a
history of smoking. Another carotenoid possessing potent in vitro anticarcinogenic
properties is lycopene, which studies have shown is able to inhibit proliferation/induce cell
cycle arrest, inhibit metastasis in a manner similar to and -carotene, induce the
antioxidant response element, induce apoptosis and improve cell-to-cell communication.
The carotenoids lutein and -cryptoxanthin have had much less research devoted to them,
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and although their consumption has been associated with lower cancer incidence future
research should take a closer examination into their specific mechanisms of action.
Key Words: Antioxidant, Carotenoid, Reactive Oxygen Species, Nuclear Factor E2-related
Factor 2 (Nrf-2), Growth Factor
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Abbreviations
ALU – Short interspersed element
AP-1 – Activator protein 1
ATBC - Alpha-Tocopherol, -Carotene Cancer Prevention Study
CARET - Beta Carotene and Retinol Efficacy Trial
CYP – Cytochrome P450
DNA – Deoxyribonucleic acid
DNMT – DNA methyltransferase
ERK – Extracellular signal-regulated kinase
FAK – Focal adhesion kinase
GCLC - Glutamate-cysteine ligase (catalytic subunit)
GCLM – Glutamate-cysteine ligase (modifier subunit)
GSR - Glutathione reductase
GSTs - Glutathione S-transferases
HDL – High-density lipoprotein
HO-1 - Heme oxygenase-1
ICAM – Intercellular adhesion molecule
JNK – c-Jun NH2-terminal kinase
LDH – Lactate dehydrogenase
LDL – Low-density lipoprotein
LINE-1 – Long interspersed element
MAPK – Mitogen activated protein kinase
mEH - Microsomal epoxide hydrolase 1
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MMP – Matrix metalloproteinase
NF-kB – Nuclear (transcription) factor kappa-B
NQO1 - NADPH:quinone oxidoreductase 1
Nrf-2 – Nuclear factor-E2 related factor 2
PAI – Plasminogen activator inhibitor protein
RAR - Retinoic acid receptor
RARE – Retinoic acid response element
RNA – Ribonucleic acid
ROS – Reactive oxygen species
TIMP – Tissue inhibitor of matrix metalloproteinase
UGT1A6 - UDP glucuronosyltransferase 1 family, polypeptide A6
uPA – Urokinase plasminogen activator
VCAM – Vascular cell adhesion molecule
Objectives
The objectives of the present review are to attempt to illustrate just how big of an
effect (positive or negative) an individual’s diet can have on the multistage process of
carcinogenesis. It will discuss multiple pro-carcinogenic factors in the typical Western diet
as well as numerous anti-carcinogenic agents and some of their associated mechanisms of
action, which may prove vital in halting/reversing the damage already done in many of our
bodies.
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Introduction
Cancer is one of the most unfortunate realities in life, but a reality nonetheless.
Therefore, we must accept it for what it is and do what we can for those who fall victim to
its processes. To put the problem into perspective, there will be an estimated 1,658,370
new cancer diagnoses in the year 2015 in the United States alone (American Cancer Society
Facts and Figures, 2015). Fortunately, there are certain lifestyle choices an individual can
make that can aid in reducing the risk of developing cancer in one’s lifetime.
It is estimated that only about 5-10% of cancers are of genetic origin, or inherited
from the individual’s parents. This means that approximately 90-95% of cancers can be
attributed to lifestyle/environmental factors (Anand et al., 2008). Such factors include
tobacco consumption (smoked and smokeless), alcohol consumption, sun exposure,
environmental pollutants, infections, stress, physical inactivity, obesity and diet. Since my
generation was in elementary school we have heard of the dangers of tobacco, in particular
its carcinogenic capabilities, so many of us have steered clear. Unfortunately, this same
level of education was not given to us regarding the impact that our diet has on disease
prevention.
The typical Western diet is characterized by its caloric excess and its nutritional
deficits, due in part to the overconsumption of low cost, calorie dense food options. This
problem is especially prevalent among people in the younger demographic, with 57
percent of individuals 18-29 reporting that they consume fast food at least weekly (Gallup
Annual Consumption Poll, 2013). The problem with fast food, and countless other
foodstuffs in the American diet, includes but is not limited to the overabundance of
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saturated fats/hydrogenated oils and refined sugars. Implications of these excesses will be
examined further in the discussion section. In addition to the overconsumption of fast food
items, 37.7 percent of American adults reported consuming fruit less than one time per day
while 22.6 percent reported consuming vegetables less than one time per day (CDC, 2013).
This shortage of fruit and vegetable consumption is an important contributing factor
to carcinogenesis. Diets high in fruits and vegetables have been consistently associated
with a lower incidence of cancer and this is due in part to the fact that fruits and vegetables
are some of the best sources of antioxidants in our diets (Gonzalez, 2006). It is these
antioxidants that protect our body from oxidative damage by scavenging free radicals and
quenching reactive oxygen species (ROS), which have been linked to development of cancer
(Ahsan and Waris, 2006). Some such compounds found within fruits and vegetables that
possess such chemopreventive capabilities are a class of compounds known as carotenoids,
and will be the focus of this review of research.
Background
Reactive Oxygen Species and Oxidative Stress
Reactive oxygen species (ROS) are a group of chemically reactive molecules that
contain oxygen, such as oxygen ions and peroxides. ROS are usually produced through
normal metabolic pathways, however they can also be produced in response to certain
dietary and lifestyle choices. Some dietary factors that can cause an increase in ROS
production include excesses in either dietary sugar (Yu et al., 2011) or trans fats (Bryk et
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al., 2011). Additionally, a lifestyle factor such as smoking is well known to increase the
production of ROS (Lin et al., 2012).
When under normal homeostatic conditions, ROS are produced but are kept under
strict regulation where they play an important role in multiple cell signaling pathways such
as proliferation and apoptosis (Ahsan and Waris, 2006). Problems arise when regulatory
mechanisms of the cell dysfunction and ROS production is increased. These elevated levels
of ROS can then induce oxidative stress though oxidation of the individual’s
deoxyribonucleic acid (DNA), lipids or proteins. Such oxidative damage may result in
multiple DNA modifications, however in regards to cancer these mutations are typically
substitutions in the G-C base pairings, such as that seen in the most common mutation in
the p53 suppressor gene where there is a GT conversion (Ahsan and Waris, 2006).
Mutations in the DNA then lead to altered gene/protein expression, which is ultimately the
source of the observed and sustained phenotypic changes resulting in carcinogenesis.
Signal transduction pathways important in cell growth, such as activator protein 1 (AP-1)
and nuclear transcription factor kappa B (NF-kB), are among the common signaling
pathways induced by ROS (Valko et al., 2006).
Antioxidants and Oxidative Stress
As the name might suggest, an antioxidant’s activity is to inhibit the damage done to
cellular structure and/or function via ROS. The body has multiple naturally occurring
enzymes that possess antioxidant capabilities such as NADPH, but due to the poor dietary
habits of many Westerners this balance between oxidants formed by normal aerobic
respiration and antioxidants is weighted in favor of oxidants (Ahsan and Waris, 2006). This
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imbalance is what eventually leads to the oxidative damage to various cellular
macromolecules as illustrated in Figure 1. Because many of us are constantly consuming
foods that are seen to increase levels of ROS in the body, it is important that we also
consume adequate amounts of foods high in antioxidants, such as fruits and vegetables.
One such group of antioxidants is known as carotenoids.
Carotenoids
Carotenoids are a group of more than 600 naturally occurring, fat-soluble pigments
that give plants their bright coloration (Higdon, 2004). They are a group of molecules called
tetraterpenoids, which are hydrocarbon chains consisting of 40 carbon atoms. There are
two main classifications of carotenoids; xanthophylls are those containing oxygen in their
otherwise hydrocarbon structure and carotenes are those that are pure hydrocarbon
molecules.
Xanthophylls (and carotenes) are not produced endogenously by humans and must
therefore be consumed in the diet by eating fruits and vegetables. Xanthophylls serve an
important role in protecting plants from high-intensity light-induced oxidative stress. They
include the molecules -cryptoxanthin and lutein, and function to absorb high blue
wavelengths, thus protecting the chloroplast from over excitation and subsequent
production of ROS through the action of the xanthophyll cycle (Latowski et al., 2011). With
sufficient dietary intake, humans preferentially sequester lutein and zeaxanthin in the
macula lutea of the retina, where it too protects the cells from ROS damage. Additionally,
recent research has elicited the positive effects of lutein and zeaxanthin on not only vision,
but on cognition as well (Johnson, E.J. 2014).
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Carotenes are carotenoids that possess purely hydrocarbon structures. These
molecules, such as -carotene, -carotene and Lycopene, function during photosynthesis
by transmitting its absorbed light energy to chlorophyll as well as absorbing the energy
from singlet oxygen. Two of the main dietary carotenes, -carotene and -carotene, are
considered provitamin A carotenoids, meaning that the body is able to metabolize these
molecules into vitamin A or its metabolites (Tanaka et al., 2012). It is believed that this
provitamin A activity is responsible for some, if not most, of the anticarcinogenic effects of
these provitamin A carotenoids.
Literature Review
-Carotene and -Carotene
-Carotene is one of the major dietary sources of vitamin A and some foods
containing the highest concentrations of this carotenoid include carrots, sweet potatoes
and winter squash. This carotenoid has been shown to exhibit significant antiproliferative
effects in vitro on the human neuroblastoma cell line GOTO in both a dose and time-
dependent manner (Murakoshi et al., 1989). The N-myc protein is highly expressed and
necessary for fetal development due to its ability to stimulate cell growth and proliferation,
but in cancer the gene amplification of this proto-oncogene results in activation and an
over-expression of the protein by 5-1000 times that of normal (DeOcampo, 2015).
Densitometry autoradiographs were used to determine the expression of N-myc messenger
ribonucleic acid (RNA) and -carotene was shown to significantly suppress the expression
of this messenger RNA in a concentration and time dependent manner, roughly ten times
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more effectively than did -carotene. In an additional experiment varying concentrations of
-carotene and -carotene were added to cultures containing the GOTO cell line and the
results showed that both possessed antiproliferative properties, with -carotene being
effective at a ten times lesser concentration (Murakoshi et al., 1989). -Carotene’s effect on
cell cycle progression was then determined by measuring the relative amounts of DNA
within the cell via flow cytometry of the control and the cells treated with -carotene.
Results indicated the cells treated with -carotene to be accumulating in the G0-G1 phase of
the cell cycle (Murakoshi et al., 1989). Additional in-vivo mice trials have shown that -
carotene is approximately 10x more potent of an inhibitor of liver, lung and skin
carcinogenesis than -carotene (Murakoshi et al., 1992).
Recent in-vitro and in-vivo studies have sought to indicate whether or not -
carotene (and -carotene) may possess any antimetastatic properties in the highly invasive
Lewis lung carcinoma (LLC). Cell culture containing the LLC cells were treated with -
carotene, -carotene and control and data showed that (at similar concentrations) and -
carotene induced comparable inhibition of invasion and migration of the LCC cells (Liu et
al., 2015). There was no effect on proliferation, however. Their effect (if any) on matrix
metalloproteinase (MMP) 9, MMP 2 and urokinase plasminogen activator (uPA) was then
examined in culture. and -carotene each significantly reduced the activity of MMP 9/2,
but only -carotene was shown to reduce the activity of uPA (Liu et al., 2015).
The effect of and -carotene on the expression of the tissue inhibitor of MMP
(TIMP) -1, TIMP-2 and plasminogen activator inhibitor (PAI)-1 proteins was examined as
well. Data showed that both molecules markedly increased expression of all three proteins
in LLC cells in culture when at similar concentrations (Liu et al., 2015). The ability of -
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carotene to alter the expression of integrin -1 and subsequent integrin -1 mediated
signaling molecules, such as those in the focal adhesion kinase (FAK) and mitogen activated
protein kinase (MAPK) families, was then examined. Results showed that -1 integrin
expression was markedly reduced (Liu et al., 2015). The effects of -carotene on metastasis
in LLC mice are summarized in Figure 2. Data also showed that activation of multiple -1
integrin mediated downstream signaling molecules was also significantly reduced
following -carotene treatment. The phosphorylation status of extracellular signal-
regulated kinase (ERK)-1, ERK-2, FAK, c-Jun NH2-terminal kinase (JNK)-1, JNK-2, and p38
(of the MAPK family) were all significantly reduced (Liu et al., 2015). However, data elicited
that although altered phosphorylation caused a change in activation, the protein expression
of these molecules remained constant (Liu et al., 2015).
In-vivo mouse experiments on LLC mice studied the effects of an only -carotene
treatment, only taxol treatment, and a combination treatment on body weight, primary
tumor growth and lung metastasis. Data showed that -carotene treatment, taxol
treatment, and a combination treatment did not alter body weight of the LLC-bearing mice
(Liu et al., 2015). In addition, -carotene treatment alone did not alter primary tumor
growth, whereas taxol (and a combination) treatment was shown to significantly inhibit
the growth of the primary tumor (Liu et al., 2015). Data showed that both -carotene
treatment and taxol treatment both significantly reduced lung metastasis, with taxol being
the more potent inhibitor. Additionally, results indicated that combination treatment with
-carotene and taxol together was more successful than either of the treatments alone (Liu
et al., 2015).
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Similar in-vivo LLC mice studies examined the effects -carotene, taxol, and
combination treatment on the expression of integrin -1, TIMP-1/2, and PAI-1 in addition
to the phosphorylation of FAK. Treatment of LLC mice with -carotene and taxol alone
markedly reduced integrin -1 expression, while only taxol treatment inhibited the
phosphorylation of FAK. Combination treatment did not enhance these effects (Liu et al.,
2015). The results indicated that both -carotene and taxol significantly increased the
expression of TIMP-1 and PAI-1, but only taxol increased the expression of TIMP-2.
Combination treatment proved to enhance these effects (Liu et al., 2015).
Studies on -carotene seem to suggest that under normal circumstances it acts as an
antioxidant and enhances immune function, but in smokers and other high-risk individuals
it can actually act as a pro oxidant and aid in the process of carcinogenesis. Two large
intervention studies, the Alpha-Tocopherol, -Carotene Cancer Prevention Study (ATBC)
and the Beta Carotene and Retinol Efficacy Trial (CARET), showed a strong correlation
between high-dose -carotene supplementation and lung cancer incidence when compared
to the control (ATBC Study Group, 1994), (Omenn et al., 1994). Recent in-vitro analysis of
rat microsomal membranes studied the procarcinogenic effects of -carotene in the
“presence” of cigarette smoke (tar in this case) and -tocopherol (form of vitamin E). Data
showed that at low (nonphysiological) O2 pressures, -carotene acted as an antioxidant and
reduced the amount of lipid peroxidation caused by the tar in the presence and absence of
-tocopherol (Palozza et al., 2006). At physiological O2 pressure, -carotene significantly
increased the lipid peroxidation induced by the tar, and this effect was reduced in the
presence of -tocopherol (Palozza et al., 2006). In addition, the effect of varying O2
pressure on -carotene consumption and oxidation was examined and data showed that
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both consumption and oxidation of -carotene was significantly increased during an
increase in air pressure (Palozza et al., 2006). Another study that may provide insight into
-carotene’s role as a co-carcinogen looked into its effects on microsomal cytochrome p450
(CYP)-linked microoxygenase activity and ROS production in lung, liver, kidney and
intestinal tissues. Data showed that not only were a multitude of CYP-induced enzymes
increased, but the associated increase in ROS production was substantial (up to a 33-fold
increase in liver tissue) (Paolini et al., 2001).
Lycopene
Lycopene is a non-provitamin A carotenoid that is found in high concentrations in
guava, watermelon and tomatoes and has been the subject of much recent research. Many
of lycopene’s biological functions are due to its ability to activate the antioxidant response
element, which is a series of cellular oxidative stress defense enzymes (Solis et al., 2013).
The effects of lycopene’s metabolite apo-10’-lipopenoic acid on nuclear factor-E2 related
factor 2 (Nrf-2) and subsequent antioxidant enzyme expression was examined in the
human bronchial epithelial cell line BEAS-2B. Data showed that not only did apo-10’-
lipopenoic acid cause nuclear translocation of Nrf-2, but it also induced an increase in the
messenger RNA of multiple phase II antioxidant/detoxification enzymes including heme
oxygenase-1 (HO-1), NADPH:quinone oxidoreductase 1 (NQO1), glutathione S-transferases
(GSTs), glutathione reductase (GSR), the catalytic and modifier subunits of glutamate-
cysteine ligase (GCLC and GCLM, respectively), microsomal epoxide hydrolase 1 (mEH)
and Uridine diphosphate (UDP) glucuronosyltransferase 1 family, polypeptide A6
(UGT1A6) (Lian and Wang, 2008). Additionally, treatment with apo-10’-lipopenoic acid
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resulted in the increase of the endogenous antioxidant glutathione and a significant
decrease in intracellular ROS production (Lian and Wang, 2008). Apo-10’-lipopenoic acid
was also found to decrease H2O2-induced oxidative damage as noted by a reduction of
lactate dehydrogenase (LDH) release (Lian and Wang, 2008).
Multiple in vitro and in vivo studies on lycopene have illustrated its effects on
multiple growth factors and their related signaling pathways, which are summarized in
Figure 5. For example, lycopene has been shown to decrease levels of circulating insulin-
like growth factor 1 (IGF-1) and vascular endothelial growth factor (VEGF), while also
inhibiting the autophosphorylation and activation of platelet derived growth factor (PDGF)
(Solis et al., 2013). Studies have also elicited the fact that lycopene has the ability to induce
apoptosis and/or cell cycle arrest, inhibit metastasis, and enhance cell-cell communication
(Solis et al., 2013). In vivo studies showed lycopene’s association with a reduction in the
average number of tumors per mouse in both lung and liver cancer, seen in Table 1
(Nishino et al., 2002).
Lycopene’s effects on the androgen-sensitive LNCaP prostate cell line and androgen-
independent PC3 prostate cell line have also been investigated. In the androgen sensitive
LNCaP cell line data showed an induction of cell cycle arrest at the G1/S phase transition as
well as an induction of apoptosis (Ivanov et al., 2007). The methylation status of the GSTP-1
promoter region, as well as the expression DNA methyltransferases (DNMT), was
unaffected following lycopene treatment in the LNCaP cell line (Fu et al., 2014). However,
LNCaP cells did experience a reduction in methylation of the long interspersed element
(LINE-1) and short interspersed element (ALU) (Fu et al., 2014). The androgen-
independent PC3 cell line also saw an induction of cell cycle arrest at the G1/S phase
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transition and although it did not experience an induction of apoptosis, it did experience a
reactivation of glutathione-S-transferase- (GSTP-1) (Ivanov et al., 2007). Additionally,
data showed that lycopene treatment of the PC3 cell line experienced a significant increase
in methylation of the GSTP-1 promoter and subsequent GSTP-1 expression (Fu et al., 2014).
Data also showed a reduction of PC3 cells’ DNMT 3A protein expression (Fu et al., 2014).
-Cryptoxanthin
-Cryptoxanthin is a provitamin A carotenoid and falls into the xanthophyll
category. Common sources include red peppers and citrus fruits. The correlation of serum
-cryptoxanthin levels and lung cancer incidence was examined in a cohort study of
Chinese men, since they have a relatively high dietary intake of citrus. Data showed that
high serum -cryptoxanthin levels were associated with a significant reduction in lung
cancer risk (Yuan et al., 2003). The effects of -cryptoxanthin on the non-small cell lung
cancer cell lines A549 and BEAS-2B have also been examined in vitro. Data indicated -
cryptoxanthin exposure resulted in a significant reduction in the total number of cells due
to the induction of cell cycle arrest at the G1/S phase transition (Lian et al., 2006). Data also
indicated that -cryptoxanthin increased the expression of messenger RNA and protein
levels of the retinoic acid receptor (RAR), which correlated with an increase in the RAR-
mediated response element (RARE) (Lian et al., 2006). In vivo trials on F334 rats have
studied the effects of -cryptoxanthin on colon cancer incidence, and found that -
cryptoxanthin reduced the incidence of N-methylnitrosourea induced colon cancer
(Narisawa et al., 1999).
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Lutein
Lutein is considered a non-provitamin A carotenoid of the xanthophyll class and is
found in high concentrations in kale, spinach and egg yolks. Recent research has studied
the effects in vivo of lutein treatment during the promoting stage of carcinogenesis. Data
showed that in both mouse lung and skin cancer lutein, when administered during the
same period as the tumor promoter, significantly reduced the average number of tumors
shown by the mice (Nishino et al., 2002). Additionally, lutein was shown in vivo to inhibit
the formation of aberrant crypt foci in the rat colon (Nishino et al., 2002).
Discussion
Dietary Risk Factors of Carcinogenesis
It is no longer a secret that improper dietary habits are one of the biggest risk
factors of carcinogenesis, with an estimated 30-35 percent of cancers being linked to an
individual’s diet (Anand et al., 2008). This statistic is comparable to that of tobacco, which
is known to be littered with thousands of chemicals. Of these chemicals, at least 60 of them
are known carcinogens (DeOcampo, 2015). There are many aspects of the modern Western
diet that can be considered a risk factor of carcinogenesis. Two of the more common risk
factors include excesses of refined sugar and partially hydrogenated oils/trans fats.
The problem associated with excess refined sugar is multifold. One side of the
equation is that increased carbohydrate intake is typically seen as one of the primary
causes of obesity, a well-known risk factor of carcinogenesis (Anand et al., 2008).
Intuitively, such an overconsumption of refined sugars can commonly result in chronic
18
elevated blood glucose levels, or hyperglycemia. Prolonged hyperglycemic conditions may
result in multiple modified intracellular signaling pathways. Signaling pathways relevant to
carcinogenesis that may be modified following high glucose exposure include those
involved with cell-to-cell adhesion molecule gene expression, inflammatory gene
expression, growth factor production and ROS production/oxidative stress (Popov, D.,
2010). Such alterations to signaling pathways induces phenotypic changes characterizing
malignancy.
Throughout recent years much attention has been brought regarding the health
risks associated with consuming trans fat. Trans fatty acids can be difficult to avoid even
today because they are widely used in the production of fried foods like chips and French
fries (partially hydrogenated vegetable oils used in frying), baked goods (many shortenings
use these oils), and margarines. Trans fatty acids simultaneously increases the “bad
cholesterol” known as low-density lipoprotein (LDL) and decrease the “good” cholesterol
known as high-density lipoprotein (HDL). Initially shown to have implications on
cardiovascular health by aiding in the development of heart disease, recent studies have
suggested that trans fat consumption is also positively correlated with an increase risk of
colon cancer (Slattery et al., 2001) and breast cancer (Breastcancer.org, 2015).
Nearly 90 percent of all cancers fall into the carcinoma category, or cancers that
arise from mutations in epithelial cells (DeOcampo, 2015). One such theory on the origin of
cancer that supports this statistic is the chronic inflammation/irritation theory. Trans fatty
acids have been shown to induce an inflammatory response in human aortic endothelial
cells that is mediated by NF-B activation (Bryk et al., 2011). During the inflammatory
response endothelial cells activate NADPH oxidase, which results in an increase in
19
intracellular ROS production. These ROS then function as second messengers, activating a
multitude of protein kinases and transcription factors, including NF-B (Bryk et al., 2011).
Once activated, NF-B translocates into the nucleus, binding to the promoter regions of the
inflammatory intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion
molecule-1 (VCAM-1) genes, inducing an upregulation of the expression of these cellular
adhesion molecules on the cell surface as illustrated in Figure 4 (Bryk et al., 2011). This
upregulation of surface proteins ICAM-1 and VCAM-1 is important in the process of
carcinogenesis because CAMs bind to or are associated with the cytoskeleton, linking
molecules, second messenger systems, growth factor receptors, oncogenes products and
transcription products (DeOcampo, 2015).
-Carotene and -Carotene
The -carotene and -carotene molecules are important dietary sources of
antioxidants, due in part to their provitamin A metabolic activities. Adequate intake of
these and other carotenoids are important as they function to inhibit the carcinogenic
effects of certain dietary elements, such as excess sugar consumption/uptake and trans
fatty acids consumption. The N-myc protein has the ability to push cell growth and
proliferation, and it appeared that the maximum inhibition of growth and proliferation,
marked by the relative amount of cells in the G1/G0 phase of the cell cycle, occurred while
the expression of N-myc messenger RNA was maximally suppressed. Additionally, N-myc
messenger RNA recovery was associated with a decreased number of cells in the G1/G0
phase and an increase in the number of cells in the S phase indicating that cell cycle arrest
may be mediated through the repression of the N-myc gene (Murakoshi et al., 1989).
20
The efficacy of -carotene as an anticarcinogenic agent has been in question,
especially when talking about high-risk individuals like smokers. One reason smokers may
experience adverse effects of -carotene supplementation is because many of their cells,
especially those of the lungs, have already experienced one or more hits (mutations) to
their DNA, due to the high carcinogenicity of tobacco and the high concentration of ROS in
cigarette smoke. Oxidation of certain biomolecules such as DNA, lipids or proteins can lead
to the development of multiple chronic diseases, including cancer (Rao and Rao, 2007).
With regards to lipids, -carotene reduces lipid peroxidation at low O2 pressure, however
at physiological O2 pressure -carotene causes a significant increase in lipid peroxidation,
which may have carcinogenic implications (Palozza et al., 2006). Also having important
carcinogenic implications was -carotene’s induction of CYP and associated enzymes with
the subsequent significant overproduction of ROS.
-Carotene seemed to exhibit a more potent antimetastatic effect than that of -
carotene. Proteins of the MMP family function in the breakdown of extracellular matrix,
and are believed to play a role in malignancy/metastasis (Liu et al., 2015). Alternatively,
TIMP proteins function by inhibiting these MMP proteins. Although both reduced the
expression of MMP-9 and MMP-2 while also increasing expression of TIMP-1, TIMP-2 and
PAI-1, only -carotene reduced the expression of uPA, which aids in the activation of
plasminogen and formation of active MMPs (Liu et al., 2015). Additionally, -carotene was
shown to markedly reduce the expression of integrin -1 (and associated downstream
kinases), which has important carcinogenic implications because integrins serve as the
primary cell surface receptor for extracellular matrix proteins, such as MMPs, (DeOcampo,
2015).
21
Lycopene
Lycopene has been shown to be the most potent in vitro antioxidant of all the
studied carotenoid molecules. Many of lycopene/lycopene metabolites’ protective effects
are thought to be mediated through its antioxidant activity, which is the result of the
upregulation of Nrf-2. Nrf-2 has important implications in carcinogenesis because it
mediates the activation of the antioxidant response element when dissociation from the
inhibitory Keap-1 protein and translocation to the nucleus occurs (Lian and Wang, 2008).
This antioxidant response element increases the expression of a multitude of antioxidant
enzymes, which can control ROS overproduction.
Multiple growth factor signaling pathways associated with carcinogenic activity
have also been shown to be inhibited by lycopene exposure. IGF-1 signaling, important in
proliferation and apoptosis, was reduced by lycopene’s ability to increase levels of
circulating IGF binding proteins 1 and 2 (Solis et al., 2013). Lycopene also decreases levels
of circulating VEGF, which upon binding to its receptor can induce proliferation, migration,
and angiogenesis among other things (Solis et al., 2013). A third growth factor that is
inhibited by lycopene is PDGF. The PDGF is activated after undergoing dimerization and
becoming a hetero- or homodimer, and it is this process that lycopene inhibits thus
inhibiting activation and downstream signaling (Solis et al., 2013).
In the body lycopene is preferentially sequestered in certain tissues, such as breast
tissue and tissue of the prostate gland, which may prove to be important in future
treatment options. Lycopene seems to have different effects on the androgen sensitive and
androgen-independent prostate cancer cell lines, LNCaP and PC3 cell lines, respectively.
Lycopene was able to induce cell cycle arrest in both cell lines, via downregulation of the
22
IGF-1 receptor and consequent signaling pathways (Ivanov et al., 2007). The tumor
suppressor retinoblastoma protein (Rb) therefore remains bound to E2F, avoiding
phosphorylation and cell cycle progression. The tumor suppressor p53 is upregulated as
well as the cyclin dependent kinase (Cdk) inhibitors p21 and p27, further suppressing
proliferation (Ivanov et al., 2007). Apoptosis was induced only in the androgen-sensitive
LNCaP cell line and was mediated by the upregulation of pro-apoptotic proteins such as
Bax and a downregulation of anti-apoptotic proteins such as Bcl-2, BclXL and survivin,
which suggests the androgen receptor’s part in activating the genes that code these
proteins (Ivanov et al., 2007).
Lycopene’s inhibition of metastasis occurs in a dose dependent manner, seen in
Figure 4, by decreasing the expression of MMP-2/9, similarly to -carotene, as well MMP-7
(Solis et al., 2013). Lycopene and its metabolites also induce simultaneous upregulation of
TIMP-1/2, so this combined effect drastically reduces the cell’s ability to degrade the
surrounding extra-cellular matrix. Another antimetastatic property of lycopene was its
upregulation of the nm23-H1 gene. Increased expression of the nm23-H1 gene is
associated with a decrease in metastasis in liver, colon, breast, melanoma and gastric
carcinomas (Tee et al., 2006). This differs from thyroid carcinomas, neuroblastomas and
osteosarcomas, where an increase in nm23-H1 is actually associated with a more
aggressive/malignant phenotype, which may suggest tissue specific roles of nm23-H1 (Tee
et al., 2006). Lycopene treatment was also shown to decrease -catenin expression, an
important cellular adhesion and signal transduction molecule utilized during
carcinogenesis (Solis et al., 2013). Loss of cell-to-cell communication is believed to be a
factor in metastasis, so lycopene’s ability to increase cellular gap junction communication
23
via upregulation of connexin-43 (an important gap junction component) messenger RNA
may play a role in its antimetastatic properties (Solis et al., 2013).
-Cryptoxanthin
Citrus fruits are some of the best dietary sources of -cryptoxanthin, which is the
reason cohort study decided to use Chinese men near Shanghai (higher than average citrus
fruit consumption). Additionally, -cryptoxanthin supplemented rats showed a significant
reduction in the incidence of colon cancer. These preventative effects of -cryptoxanthin
may be mediated through its ability to induce cell cycle arrest by suppressing the
expression of cyclin D and cyclin E, which are necessary in cell cycle progression, and by
increasing the expression of the cyclin dependent kinase inhibitor p21 (Lian et al., 2006).
The increase in RAR messenger RNA and protein expression is significant because
retinoids, such as retinoic acid, are the biologically active form of vitamin A and therefore
increasing receptor expression should increase the efficacy of not only vitamin A, but of the
provitamin A molecules as well.
Lutein
In vivo studies on mice supplemented with lutein during the tumor-promoting
phase of the experiment suggest that its anticarcinogenic effects may occur during tumor
promotion, as evidenced by the reduction in the incidence of skin and lung tumors in the
lutein supplemented group. Lutein is preferentially sequestered in the human eye,
particularly in the macula, where it functions to absorb the retinal-damaging blue light.
24
Lutein is also the most abundant carotenoid in human brain tissue, where it is believed to
have antioxidant, anti-inflammatory and structural activities (Johnson, 2014).
Conclusions/Comments
This semester has given me a glimpse into just how complex the multistep process
of carcinogenesis really is. There are so many different mechanisms that interact that it can
be intimidating at times. This class was unique and I really enjoyed how it was structured
around in class lectures and individual research. The majority of classes I have taken over
the past five years have been primarily lectures with exams on memorized information, so
it was refreshing to have to research and integrate information on your own. It was also
interesting to see what topics students chose to present on, and the overall quality of the
majority of presentations was quite impressive.
The vast majority of cancers are attributed to lifestyle choices such as your diet and
smoking, which is why I was interested in our diet’s role in carcinogenesis. There are
numerous dietary factors that aid in cancer development, but two of particular concern in a
typical Western diet are excess refined sugars and the consumption of trans fatty acids. As
a country with millions of soda drinkers and fast food frequenters, it is important to know
the risks associated with such habits. Luckily, there are also some dietary choices you can
make that can inhibit the process/progression of carcinogenesis such as increasing your
intake of fruits and vegetables, which are rich in nutrients like antioxidants and
carotenoids.
With the exception of the aphid, animals do not possess the ability to produce
carotenoids endogenously, so we must make sure we have an adequate intake of fruits and
25
vegetables. Some carotenoids have proven to be potent in vitro and in vivo inhibitors of
several mechanisms and pathways associated with cancer development and progression.
These include mechanisms such as cell cycle arrest/inhibition of proliferation, inhibition of
various growth factor activity, induction of apoptosis, increased cell-to-cell communication
and the inhibition of metastasis. These effects seem to be intensified when working in
combination, so make sure to eat a variety of fruits and vegetables to capitalize on their
anticarcinogenic effects.
26
Figures
Figure 1: Simplified flowchart detailing the steps in disease progression after ROS
production and lack of repair. (Rao and Rao, 2007).
Figure 2: A summary of the antimetastatic effects of -carotene treated LLC cells in vitro.
(Liu et al., 2015).
27
Figure 3: Lycopene’s effects on cell invasion and migration occurs in a dose dependent
manner. (Huang et al., 2005).
28
Figure 4: The effects of trans fatty acid exposure on ICAM-1 and VCAM-1 expression in
endothelial cells. Graphs A and B are surface protein expression and graphs C and D are
messenger RNA expression. (Bryk et al., 2011).
29
Figure 5: Summary of the multiple growth factor signaling pathways effected by lycopene.
(Solis et al., 2013).
30
Table 1: Lycopene treatment on lung and liver carcinogenesis in mice, in vivo. (Nishino et
al., 2002).
31
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