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Phytosterols for Cancer Treatment
Josh Nooner, BS, CSCS
Phytochemicals
12/4/15
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Abstract
Cancer is a devastating disease that affects millions of people and remains one of the
world’s leading causes of death. In order to combat this disease, researchers are continuously
looking for ways to prevent and treat different forms of cancer. While a cure for cancer has yet to
be found, there has been promising evidence emerge for various treatment options that can halt
the progression of these cancers. One such option is through the use of phytosterols.
Phytosterols are plant-derived compounds that are similar in structure and function
to cholesterol. Phytosterols include both sterols and stanols, which differ only in a double bond
in the sterol ring. The most abundant sterols in plants and the human diet are sitosterol and
campesterol. Phytosterols have shown to reduce LDL cholesterol, improve lipid profiles, and
prevent cardiovascular disease. Additionally, the most recent research on phytosterols has shown
that they are also effective at treating different forms of cancer. Studies on phytosterols have
shown them to be effective at treating lung, stomach, breast, colon, and prostate cancers.
Phytosterols have shown to treat cancer by inducing apoptosis, reducing ROS levels,
preventing oxidative damage, increasing antioxidant enzymes, reducing blood cholesterol,
decreasing inflammatory cytokines, and decreasing angiogenesis to cancer cells. A dose of 300
mg/day has shown to exert these beneficial effects, however most Americans only consume 80
mg/day. Interestingly, this dose is also the dose needed to see beneficial changes in blood lipids
that will help to prevent cardiovascular disease. Phytosterols can be used safely by everyone
except individuals with sitosterolemia. Further research is needed to better understand some of
the long term effects, interactions with prescription drugs, and the exact mechanisms of
prevention. Phytosterols are a promising treatment option for many types of cancer and will be
used as a dietary intervention in a clinical setting in the future.
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Introduction
Phytosterols are a major group of phytochemicals that have been traditionally used to
prevent hypercholesterolemia and cardiovascular disease. In the 1990’s there was extensive
research being done on their effects on cholesterol and blood lipids. This came after the drastic
increase in the incidence of individuals with high cholesterol caused by the rise of fast food
chains. Research showed that phytosterols were effective at lowering blood cholesterol because
they are very similar in structure to cholesterol and lower the amount of cholesterol absorption.
After this discovery, phytosterols were added to many types of food in order to try and combat
the high rates of hypercholesterolemia. Margarine was the main food that these chemicals were
added to because they enter the enterocyte in micelles, therefore they absorb better when added
to fats. After the research died down regarding their effects on cardiovascular disease the
attention slowly turned towards cancer. The most recent studies on phytosterols are focused on
determining their effects on treating different types of cancer.
In the last 10-15 years there has been a large number of studies done in this area. All the
research is currently being done in animal models or in vitro. The only human studies that have
been conducted are epidemiological studies. The results found in the animal and cellular studies
still need to be confirmed in controlled clinical trials. Clinical trials in humans will be the next
step once we have a better understanding of the mechanisms involved. We do know some of the
mechanisms in which phytosterols exert their effects, but we need a deeper understanding of the
exact steps that are involved. Some of the pathways that we know about can be seen below in
figure 2 (Woyengo, 2009). We know that the main way that cancer progression is being
prevented is through the induction of apoptosis in cancerous cells. We also know that
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phytosterols increase caspase activity, reduced ROS, increase cellular antioxidant levels, reduce
blood cholesterol and prostaglandin synthesis, and decrease angiogenesis of cancer cells.
We have a basic understanding of the mechanisms involved as well as the dosage that we
need to see these benefits. The current studies on dosing recommend a range of 200-400 mg/day
of phytosterols with the average being around 300 mg/day. However, most Americans only
consume around 80 mg/day due to the low intake of vegetables in a Western diet. In order to see
these positive effects it is important that we keep the serum level of phytosterols at a relatively
constant concentration. To do this, we must eat a variety of phytosterol containing foods each
day.
We have a number of dietary options in which we can get our daily phytosterol intake
from. The foods that have the highest concentration of phytosterols are vegetable oils and
avocados which contain 95 mg of beta-sitosterol/200 calories. Phytosterols are also found in high
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amounts in fortified foods such as margarine which has 77mg/200 calories. Other sources of
phytosterols include nuts, seeds, dark chocolate, and eggs. Pistachios, hazelnuts, macadamias,
pecans, and walnuts are all good sources which range from 77mg/200 calories down to
20mg/200 calories respectively. The easiest way to get phytosterols in the diet is to cook with
vegetable oils and use healthy alternative fortified margarines for cooking. It is very important
that we eat these types of foods daily due to the low bioavailability of phytosterols.
Current studies show that phytosterols have a range of absorption from 0.5 – 8%. The
majority of the research shows that around 5% of phytosterols are absorbed. The reason for such
low bioavailability is due to a number of mechanisms. First, there is a low absorption of
phytosterols from the intestinal lumen into the enterocyte. Secondly, there is poor esterification
of phytosterols once they enter the enterocyte. This causes the phytosterols that do happen to get
absorbed to be kicked right back out into the intestinal lumen. Lastly, there is a high rate of
biliary excretion of phytosterols once they reach the liver. Take these three factors combined and
we have a very low concentration of phytosterols in the blood. This is why it is important to
consume these foods daily.
Consuming these foods daily is also safe for almost every population. A daily intake of
300-400 mg/day of phytosterols has shown to have no side effects except for in one very specific
population. There is a very rare autosomal recessive inherited lipid metabolic disorder that can
result in sitosterolemia. This disease is a result of a mutation in both copies of the ABCG5 or
ABCG8 gene. This disease renders the ABC transport proteins ineffective and results in elevated
blood sterols, atherosclerosis, and xanthomas. If these individuals consume a diet high in
phytosterols it can result in pockets of fatty deposits on the surface of the skin. These individuals
should avoid high phytosterol intake and be put on a low fat diet.
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Major Research Findings
I will know give a detailed discussion of my major research findings. As previously
stated, all the current research in this area is being done in animal model and cell studies.
Therefore, the majority of the articles I will cover are from these models. I will discuss 4 animal
studies, 3 cell studies, and 1 human epidemiological study.
Animal Studies
My first study is titled, Phytosterols inhibit the tumor growth and lipoprotein
oxidizability induced by a high-fat diet in mice with inherited breast cancer (Llaverias, 2013).
This study was published in 2013 in the Journal of Nutritional Biochemistry. The authors took 4-
week-old female PyMT Tg mice and randomized them into 2 groups. One group consumed a 2%
phytosterol supplement added to their normal powdered food, and the second group consumed
their normal powdered food without the phytosterol supplementation. Each of these groups was
then further subdivided into two more groups, the first being placed on a low fat low cholesterol
diet (6.2% fat, no cholesterol, energy density 3.1 kcal/g, calories from protein, fat and
carbohydrate, 24%, 18% and 58%, respectively), and the second being placed on a high fat high
cholesterol diet (21.2% fat, 0.2% cholesterol, energy density 4.5 kcal/g, calories from protein, fat
and carbohydrate, 15.2%, 42% and 42.7%, respectively). Phytosterols were composed of 20%
campesterol, 22% stigmasterol, and 41% β-sitosterol. Mice were maintained on the diets until
euthanized (at 4, 8 or 13 weeks of age).
The results from this study showed a decrease in total tumor weight in the high fat high
cholesterol diet when supplemented with phytosterols. There was also a significant reduction in
total lesion area in the high fat high cholesterol group that was supplemented with phytosterols.
Lastly, the results showed that cyclin D1, a gene that promotes breast cancer, was decreased
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using a western blot analysis. These are very positive results when considering their application
to human health and disease. These results indicate that dietary phytosterol supplementation
delayed tumor onset and progression in the setting of a typical Western diet. Interestingly, the
protection against cancer provided by phytosterols occurred at the same dose that reduced blood
cholesterol and cardiovascular risk. Here we can see that individuals with breast cancer can see
positive reductions in tumor promoter genes and tumor size with phytosterol supplementation.
My second study is titled, In vitro and in vivo (SCID mice) effects of phytosterols on the
growth and dissemination of human prostate cancer PC-3 cells (Awad, 2001). For the in vitro
portion of the study the authors supplemented PC-3 cells with either cholesterol, campesterol, or
beta-sitosterol. The authors then tested tumor invasiveness and cell migration. For the in vivo
portion of the study the authors fed mice for two weeks ad libitum either a cholesterol
supplemented diet or a phytosterol supplemented diet. Tumor cells were then injected into the
mice and their growth was monitored for 8 weeks.
The results of this study showed that the tumor cell density increased in the cholesterol
supplemented group, but both the campesterol and beta-sitosterol supplemented groups greatly
reduced the tumor cell density. Additionally, the cholesterol supplemented group significantly
increased tumor cell migration, whereas the campesterol and beta-sitosterol supplemented groups
both significantly reduced tumor cell migration. Lastly, total tumor are was significantly reduced
at 6, 7, and 8 weeks post tumor inoculation in the mice fed the phytosterol diet compared to the
high cholesterol diet. In summary, cell density and cell migration were significantly reduced in
the phytosterol supplemented cells, and total tumor area at 8 weeks was significantly lower in the
phytosterol fed mice. If these results can be replicated in human studies, phytosterol
supplementation will become a common dietary intervention to treat human prostate cancers.
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My third animal study is titled, Protective effect of plant sterols against chemically
induced colon tumors in rats (Raicht, 1980). The methods for this study included 4 experimental
groups. The first group is a control chow and intracolonic 0.9% NaCI solution (N=10), the
second group is a control chow plus B-sitosterol (0.2%) and intracolonic 0.9% NaCI solution
(N=10), the third group is a control chow and intracolonic MNU3 (N=71), the fourth group is a
control chow plus B-sitosterol (0.2%) and intracolonic MNU (N=48). There was a 28 week
intervention period and at week 28 the colon was opened, and the number of tumors was
recorded.
The main results from this study showed that there was a decrease in colonic tumor
formation when the plant sterol B-sitosterol was added to the diet of rats. Group 3, the control
group with added carcinogen had 38 rats with tumors. However, group 4, the group with
carcinogen and phytosterol supplementation only had 16 rats with tumors. This is significant
because it shows that the addition of phytosterols to the diet of mice causes a reduction in tumor
formation. If we apply this research to humans we would hopefully see a similar reduction in
cancer rates. Human clinical trials are required to confirm this effect in humans.
My final animal study is titled, β-sitosterol, β-sitosterol glucoside, and a mixture of β-
sitosterol and β-sitosterol glucoside modulate the growth of estrogen-responsive breast cancer
cells in vitro and in ovariectomized athymic mice (Ju, 2004). The researchers injected estrogen
pellets and MCF-7 cells into ovariectomized female mice. The mice were then split into four
experimental groups. A negative control group, a BSS (Beta – Sitosterol) group, a BSSG (Beta –
Sitosterol glucoside) group, and a MC (BSS:BSSG = 99:1) group. The estrogenic and
antiestrogenic effects of dietary phytosterols on tumor growth were then measured after an 18
week intervention period.
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The results showed that tumor surface area was significantly reduced in all three of the
phytosterol groups compared to the negative control group. Additionally, all three phytosterol
groups significantly reduced the plasma estrogen levels in the mice with induced breast cancer.
The results of this study are very useful in furthering our understanding of phytosterols.
According to the data, it doesn’t matter whether the supplemented phytosterols are BSS, BSSG,
or MC. We will see the same effect when using any of the three. We can take this knowledge and
apply it to a therapeutic diet to treat breast cancer in humans.
In Vitro Studies
My first cellular study is titled, Chemopreventive potential of β-sitosterol in experimental
colon cancer model-an in vitro and in vivo study (Baskar, 2010). For the in vitro portion of the
study β-sitosterol was isolated from A. curassavica leaves. Its ability to induce apoptosis was
then determined by its in vitro antiradical activity and cytotoxic studies using human colon
adenocarcinoma cell lines. For the in vivo portion, the methodology involved injecting 1,2-
dimethylhydrazine (DMH, 20 mg/kg b.w.) into male Wistar rats. Rats were then supplemented
with β-sitosterol in 3 different concentrations which were 5 mg/kg bw, 10 mg/kg bw, and 20
mg/kg bw.
The study had a 16 week experimental period.
The results showed a dose dependent increase in induction of apoptotic cells by B-
sitosterol using both flow cytometry and fluorescent staining. Furthermore, there was a
significant reduction in ACF lesions with increased B-sitosterol supplementation, and there also
was a dose dependent decrease in B-Catenin (cell adhesion) and PCNA (DNA Replication)
genes using western blot analysis. These results show us that there is an increase in apoptosis of
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these colon cancer cells, a very beneficial and much wanted effect for delaying cancer
progression. The results also show us that the phytosterols prevent further lesions from
developing along with eliminating the current ones through apoptosis. Lastly, these results show
us just how these effects take place, through the decrease in activation of cell adhesion and
replication genes. If the same effect can be demonstrated in human colon cancer cells this would
become a great way to treat colon cancer in humans in the future and could improve treatment
options for this disease.
My second cell study is titled, Cholesterol and phytosterols differentially regulate the
expression of caveolin 1 and a downstream prostate cell growth-suppressor gene (Ifere, 2010).
This study used PC-3 and DU145 cells treated with either cholesterol or phytosterols for 72
hours. Necrosis and cell growth were measured, induction of cell growth-suppressor gene
expression was evaluated, and apoptosis was examined.
Western blots showed increases in pro-apoptotic genes and decreases in anti-apoptotic
genes. NDRG1 gene (tumor suppressor gene) increased with campsiterol, Caveolin 1 gene
(tumor suppressor gene) increased with campsiterol, BCL-xl gene and BCL-2 gene both
decreased with campsiterol, and Bcl-Sx and P53 genes both increased with campsiterol. The
opposite effect was seen in the cholesterol supplemented group. The modulation of these genes
creates an anti-cancer effect induced by phytosterols. Modification at the genetic level by
phytosterols is showing to be one of the main mechanisms in which they exert their beneficial
effects. If we can use phytosterols to modify the genes of human prostate cells we can effectively
reduce the progression of prostate cancer in humans.
My final cell study is titled, β-Sitosterol inhibits cell growth and induces apoptosis in
SGC-7901 human stomach cancer cells (Zhao, 2009). The authors used SGC-7901 human
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stomach cancer cells for this study. The cells were treated with different concentrations of β-
sitosterol. Proliferation, cytotoxicity, and apoptosis were examined using various assays and
western blotting.
The data from this study showed that there is a dose dependent decrease in proliferation
of stomach cancer cells at both 1, 3, and 5 days as seen in figure 2 below. This figure is showing
the inhibition rate of cancer cells at different concentrations of b-sitosterol.
Figure 2 - Inhibition Rate of stomach cancer cells
This study also looked at the morphological changes to the stomach cells and found that there
was a decrease in the size of the cells with increasing concentrations of phytosterols.
Additionally, the authors found that B-sitosterol increased expression of pro-apoptotic genes, and
decreased expression of anti-apoptotic genes. Western blotting showed that Pro-Caspase 3 and
Bax genes were increased while Bcl-2 genes were decreased. This information reinforces the
results from the previous study in showing that phytosterols are effective at modulating genes
that code for apoptosis. If we can learn how to modify these genes in humans through nutrition
we can create a pro-apoptotic cellular environment and effectively cause cell death to occur in
cancerous human stomach cells.
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Human Studies
The one human study that I will discuss is titled, Phytosterols and risk of lung cancer: a
case-control study in Uruguay (Mendilaharsu, 1998). This is an epidemiological study that was
completed using the data from a case-control study conducted during the period of 1993-1996.
All patients with newly diagnosed primary lung cancer diagnosed in the four major hospitals of
Montevideo were included in this study. There was a 96% response rate that included 463 cases
with lung cancer and 465 hospitalized controls. Detailed medical, lifestyle, and diet
questionnaires were completed by all participants.
The most important findings from this study were that there was a significantly lower risk
of developing lung cancer in the highest quintiles of phytosterol consumption. The odds ratios
were determined and showed that there was a trend for increased odds of getting lung cancer the
lower the phytosterol intake went. This was also seen when the results were broken down by
type of phytosterol (stigmasterol, campesterol, b-sitosterol) and when broken down by type of
cancer cell (squamous cell, small cell, adenocarcinoma). Because this study was conducted on
humans it is very applicable to disease prevention. This study shows us that those who ate more
phytosterols in their diets had significantly lower risks of developing lung cancer. Therefore, it
would be wise to introduce phytosterols into the diets of individuals at risk for developing lung
cancer in order to prevent lung cancer from developing.
As mentioned earlier, the vast majority of the current literature is all being performed on
animal and cellular models. In order to further this type of research, it is very important that we
move to the next step and begin conducting human clinical trials. This will give us better
understanding of how phytosterols work in humans, and will allow us to test our hypothesis that
phytosterols will prevent and treat many types of human cancers.
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Summary
In summary, phytosterols are proving to be a promising treatment for many types of
cancers including prostate, colon, breast, lung, and stomach. However, most Americans do not
consume enough phytostersols, the average intake being 80 mg/day. In order to see the positive
effects that we know phytosterols can provide, we must find ways to get Americans to increase
their daily phytosterol intake to 300 mg/day. If health care professionals can begin to teach
people about the cancer prevention benefits of phytosterols I believe that we can begin to see
positive changes.
We have done a lot of research in this area using animal models and cell studies. We
know that the main mechanism phytosterols use is induction of apoptosis in cancerous cells.
Phytosterols have also shown to reduce ROS levels, prevent oxidative damage, increase
antioxidant enzymes, reduce blood cholesterol, decrease inflammatory cytokines, and decrease
angiogenesis. There are numerous positive effects that can be seen with phytosterol
supplementation. The cancer treatment effects of phytosterols are seen at the same concentration
that we see the cardiovascular protective effects. However, with bioavailability at only 5%, it is
crucial that we consume phytosterol containing foods on a regular basis. We must have a daily
intake in order to keep the serum concentration at a relatively constant level.
Once research has been conducted on humans in controlled clinical trials I believe that
we will begin to see phytosterol supplementation used as a common treatment for many types of
cancers. Phytosterols show very promising benefits in terms of both cardiovascular and cancer
treatment options and I believe that we will soon see them being used as a dietary intervention in
a clinical setting. As dietitians, it is our duty to be aware of emerging research being done in this
area and be able to use this information to educate and provide better patient outcomes.
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References
Woyengo, T. A., Ramprasath, V. R., & Jones, P. J. H. (2009). Anticancer effects of phytosterols. European Journal of Clinical Nutrition, 63(7), 813-820.
Llaverias, G., Escolà-Gil, J. C., Lerma, E., Julve, J., Pons, C., Cabré, A., ... & Blanco-Vaca, F. (2013). Phytosterols inhibit the tumor growth and lipoprotein oxidizability induced by a high-fat diet in mice with inherited breast cancer. The Journal of nutritional biochemistry, 24(1), 39-48.
Awad, A. B., Fink, C. S., Williams, H., & Kim, U. (2001). In vitro and in vivo (SCID mice) effects of phytosterols on the growth and dissemination of human prostate cancer PC-3 cells. European Journal of Cancer Prevention, 10(6), 507-513.
Raicht, R. F., Cohen, B. I., Fazzini, E. P., Sarwal, A. N., & Takahashi, M. (1980). Protective effect of plant sterols against chemically induced colon tumors in rats. Cancer Research, 40(2), 403-405.
Ju, Y. H., Clausen, L. M., Allred, K. F., Almada, A. L., & Helferich, W. G. (2004). β-sitosterol, β-sitosterol glucoside, and a mixture of β-sitosterol and β-sitosterol glucoside modulate the growth of estrogen-responsive breast cancer cells in vitro and in ovariectomized athymic mice. The journal of nutrition,134(5), 1145-1151.
Baskar, A. A., Ignacimuthu, S., Paulraj, G. M., & Al Numair, K. S. (2010). Chemopreventive potential of β-sitosterol in experimental colon cancer model-an in vitro and in vivo study. BMC complementary and alternative medicine,10(1), 24.
Ifere, G. O., Equan, A., Gordon, K., Nagappan, P., Igietseme, J. U., & Ananaba, G. A. (2010). Cholesterol and phytosterols differentially regulate the expression of caveolin 1 and a downstream prostate cell growth-suppressor gene. Cancer epidemiology, 34(4), 461-471.
Zhao, Y., Chang, S. K., Qu, G., Li, T., & Cui, H. (2009). β-Sitosterol inhibits cell growth and induces apoptosis in SGC-7901 human stomach cancer cells.Journal of agricultural and food chemistry, 57(12), 5211-5218.
Mendilaharsu, M., De Stefani, E., Deneo-Pellegrini, H., Carzoglio, J., & Ronco, A. (1998). Phytosterols and risk of lung cancer: a case-control study in Uruguay. Lung Cancer, 21(1), 37-45.