(Journal of Nutrition. 2001;131:186S-190S.)
© 2001 The American Society for Nutritional Sciences
Supplement
Speaker Abstracts
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Nutritional Modulation of Human Carcinogenesis. John Potter. Fred Hutchinson Cancer Research Center,
Seattle, WA.
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The role of nutrition in cancer etiology is complex and multifaceted.
When animal experiments provided most of the information we had on
mechanisms of carcinogenesis, food was seen as a potential source of
initiating agents as well as promoters. Many of the judgments about the
role of nutrients and food-related exposures were made around
mutagenic capacity. Subsequently, it has become clear that nutrition
has a much wider role. Specific dietary compounds, such as heterocyclic
amines, aflatoxins, polycyclic aromatic hydrocarbons and
N-nitroso compounds—found in cooked, cured and spoiled food
and alcoholic drinks—may act as early-stage agents. The induction
of specific metabolizing enzymes by bioactive compounds—found
particularly in vegetables and fruits—may increase the likelihood of
detoxifying and excreting these DNA damaging compounds, largely, but
not always, reducing risk. Later in the cancer process, the growth of
an initiated clone is central. Obesity frequently increases the
likelihood of growth promotion probably via hormones and growth
factors. Physical activity tends to act in the reverse direction. Late
in the process, DNA damage is again a key issue. Folate—particularly
from vegetables—may reduce uracil misincorporation, DNA breakage and
DNA hypomethylation. Other agents in plant foods may decrease oxidative
damage and increase apoptosis. In contrast, energy-dense, high-fat
diets may contribute to oxidative damage. Nutrition influences the
likelihood of carcinogenesis via the complex pattern of influences that
includes naturally occurring bioactive compounds, both beneficial and
deleterious; compounds that are the result of methods of cooking and
processing food; overall energy intake and physical activity.
Additionally, the interaction between these factors and inherent
susceptibility and protection—both genetic and acquired—will
influence the ultimate likelihood of cancer at various stages of life.
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Molecular Epidemiology of Dietary Fat in Relation to Breast
and Prostate Cancer. Alice S. Whittemore. Stanford University
School of Medicine, Stanford, CA.
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Exogenous exposures such as diet have been implicated in cancers of the
breast and prostate by the changing incidence in migrants and their
offspring. Dietary fat intakes have long been suspected as explanations
for these incidence changes. Some epidemiological data support roles
for intakes of total fat and of saturated fat in the etiology of
cancers of the breast and prostate. For example, among countries with
large variation in incidence of these cancers, strong positive
correlations are seen between incidence rates and estimated per capita
fat intakes. Also, Asian-American migrants to the United States
have rates intermediate between the high rates in U.S. whites and the
lower rates in Asia, which may reflect higher fat intakes after
migration. However, the evidence from analytical case-control and
cohort studies is less clear. Inherent limitations in all of these
studies indicate the need for new approaches, such as randomized
intervention trials. For breast cancer, within the next decade we will
learn the results of the Women’s Health Initiative. This is a trial in
which some 67,000 postmenopausal U.S. women were randomly assigned to
receive either a diet with less than 20% of energy from fat or their
usual diet and then were monitored for subsequent breast cancer
incidence. For prostate cancer, there are as yet no published results
of trials of dietary fat reduction to prevent either incidence or
progression of disease. Given the diverse effects of fatty acids on
cellular biology and chemistry, it seems likely that their
relationships to cancer risk are complex, involving the interplay of
fat with other dietary factors, such as antioxidant vitamins and
minerals, or with genetic factors that influence susceptibility.
Emerging research on possible molecular mechanisms underlying roles for
fatty acids in these cancers promises to motivate and buttress
epidemiological studies and clarify the effects of dietary fat
reduction in preventing these cancers.
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Role of Androgens and Nutritional Factors in Prostate
Development and Function. Donald J. Tindall, Charles Y.-F.
Young and Rachel Butler. Departments of Urology and
Biochemistry/Molecular Biology, Mayo Foundation, Rochester, MN.
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The development and function of the prostate depend on the metabolic
conversion of testosterone to 5
-dihydrotestosterone, which binds to
the nuclear androgen receptor (AR). This binding induces dimerization
of the AR and interaction with androgen response elements on specific
genes that are critical for proliferation and differentiation of
prostate cells. The AR gene itself is tightly regulated by androgens as
well as by factors that induce cAMP, AP1 and NF-
B. Additionally,
suppressor elements within the AR gene may modulate its transcriptional
activity. A number of other prostate-specific genes are highly
dependent on androgens for their expression. Both prostate-specific
antigen and human kallikrein-2 are androgen dependent and serve as
markers of androgen action within prostate cells. Moreover, the
expressions of these genes are modulated by several nutritional
factors, such as vitamins D and A, via their respective nuclear
receptors. Another nuclear receptor, the peroxisome
proliferator-activated receptor (PPAR), binds to and is activated
by the prostanoid 15d-PGJ2, a
metabolite of a long-chain fatty acid. Because some essential fatty
acids (EFA) have been implicated in the development and progression of
advanced prostate cancer, we investigated the effect of
15d-PGJ2 on three human prostate
cancer cell lines: LNCaP, DU145 and PC-3. We found that
15d-PGJ2 induced cell death in all
three cell lines. Mitochondrial transmembrane potentials were
significantly reduced by 15d-PGJ2
treatment. However, cytoplasmic changes were indicative of type 2
(autophagic) nonapoptotic programmed cell death. These data demonstrate
that PPAR-
is a negative regulator of prostate cell growth.
Furthermore, they suggest that some fatty acids may play a role in
controlling the death of prostate cells.
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Effects of Estrogenic Exposures on Prostate Development.
Gail S. Prins. Departments of Urology and Physiology & Biophysics,
University of Illinois at Chicago, Chicago, IL.
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The developing prostate gland is markedly influenced by the hormonal
milieu, most notably androgens, which dictate its growth and
differentiation. In addition, the developing prostate is particularly
sensitive to estrogenic exposures. During prostate morphogenesis,
elevated levels of endogenous estrogens (maternal or excess local
production) or exogenous estrogens (diethylstilbestrol or, potentially,
environmental estrogens) have been shown to induce permanent
disturbances in prostate growth and predispose to precancerous lesions,
a process referred to as developmental estrogenization or estrogen
imprinting. The rodent prostate has evolved as a useful model to study
early estrogenic exposure because the gland develops postnatally. Using
the rat prostate as our model, we have shown that estradiol
administration on days 1–5 of life results in a permanent reduction in
prostatic growth and activational response to androgens during
adulthood, an effect mediated in part through a reduction in androgen
receptor expression. When neonatally estrogenized rats age, prostatic
hyperplasia and dysplasia are prominent, and PIN-3 lesions are observed
when these animals are given exogenous testosterone. Structural and
functional epithelial cytodifferentiation during development is
perturbed or, for some endpoints, permanently blocked by neonatal
estrogens, as determined by markers for basal and luminal cytokeratins
and secretory proteins (prostate-binding protein, urokinase, 26-kDa
protease). Recent evidence indicates that estrogen-induced changes
in the expression of E-cadherin and connexins 32 and 43 in
epithelial cells of the adult prostate may result in impaired
cell-cell adhesion and defective cell-cell communication, which
may be one of the key mechanisms through which changes towards a
dysplastic state are mediated. During the first 5 d of life,
estrogen receptor (ER) is present in the proximal mesenchymal cells
whereas ERß mRNA is expressed at low levels in the epithelium. Thus
both cell types may be targets of estrogen action during development.
After neonatal estrogen exposure, E|$$ . R expression is
up-regulated in periductal stromal cells along the length of the
ducts, which allows for amplification of estrogenic effects in those
cells specifically. In contrast, ERß is not autoregulated in
epithelial cells in response to estrogen treatment. More conclusive
studies with E|$$ . R knockout mice indicate that stromal cell
E|$$ . R is the dominant ER mediating developmental estrogenization
of the prostate. Downstream mediators of estrogen action include
transforming growth factor (TGF) ß1, which is
expressed by differentiating smooth muscle cells. Our data suggests
that transient expression of TGFß1 may be
involved in driving prostate epithelial cells into a differentiation
pathway during a critical developmental window. Exposure to high levels
of estrogens interrupts this signaling pathway at several sites, which
ultimately results in epithelial differentiation blockade. It is
hypothesized that the accumulation of undifferentiated basal cells or
of epithelial cells possessing differentiation defects in response to
early estrogenic exposure is an early event that predisposes the
prostate gland toward precancerous lesions as the animals age.
[Supported by National Institutes of Health DK-40890 and Environmental
Protection Agency R826299.]
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Estrogen Receptor Knockout Mice. Molecular and Endocrine
Phenotypes. Kenneth S. Korach. Laboratory of
Reproductive and Developmental Toxicology, National Institute of
Environmental Health Sciences, National Institutes of Health, Research
Triangle Park, NC.
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Estrogen receptors (ERs) are thought to play a crucial role in
development, reproduction and normal physiology. Using gene targeting
techniques, we have produced lines of transgenic mice homozygous for
the disrupted ER gene (ERKO) and ERß genes (ßERKO). Western
blot analysis showed no detectable ER protein in AERKO uteri.
Ribonuclease protection assay analysis has detected comparable levels
of ER
mRNA in tissues of ERKO mice, suggesting that
ERß expression is not dependent on ER. ERKO mice were totally
unresponsive to uterotropic assays with estradiol, hydroxytamoxifen and
diethylstilbestrol. Further support came from the failure of treatment
with estrogen or epidermal growth factor (EGF) to induce DNA synthesis
in ERKO uteri, even though EGF signaling was shown to be intact by
stimulation of c-fos, a gene regulatory response.
Progesterone receptor mRNA was detected in ERKO mice but not
stimulated by estrogen in the uterus, mammary gland and ovary,
indicating an estrogen-dependent and estrogen-independent gene
regulation. ERKO females are infertile and have hypoplastic uteri
and hyperemic ovaries with no corpora lutea. The ERKO ovarian
phenotype occurs developmentally by age 30 d and continues to
become more overt. Ovarian gonadotropin receptor levels, serum estrogen
and luteinizing hormone are elevated compared with wild type (WT)
females. Analysis of the mammary glands of adult ERKO females showed
a primitive ductal rudiment rather than the fully developed ductal tree
seen in WT or ßERKO mice. ERKO males are also infertile, with
atrophy of the testes and seminiferous tubule dysmorphogenesis
resulting in decreased spermatogenesis and inactive sperm. Sperm
transplantation studies of ERKO males rescues the infertility
phenotype. Males have reduced bone density and some alterations in
cardiovascular function. Phenotypic differences were seen in sex
behavior and aggressive behavior in both ERKO males and females
compared with the patterns in WT mice. Initial findings in ßERKO
females assessed by continuous breeding studies and superovulation are
arrested folliculogenesis and subfertility. In contrast to the ERKO
males, the ßERKO males are fertile and have normal sexual behavior.
Recent development of a viable double ER /ß knockout shows a
unique ovarian phenotype of transdifferentiation of granulosa to
Sertoli cells. Further characterization of the mice and comparison of
the individual and double ER gene knockout phenotypes will be required
to more fully understand the physiological consequences of
ER-mediated actions and the specific roles of the two different
forms of ER in estrogen hormone responsiveness.
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Diet Intervention in the Prevention of Cancer. Cheryl L. Rock. Department of Family and Preventive
Medicine, and Cancer Prevention and Control Program, University of
California, San Diego.
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Much of the evidence linking dietary factors to cancer risk is based on
observational studies, and the interpretation of these data has
considerable limitations. However, results from cell culture studies
support the biologic plausibility that many dietary constituents,
particularly those provided by plant foods (i.e., vegetables, fruit,
whole grains), may reduce the risk and possibly improve the prognosis
after the diagnosis of cancer. To date the overwhelming majority of
clinical trials testing the effect of nutritional factors on cancer
risk have involved single nutrients or combinations of vitamin and
mineral supplements. Results from these studies, with a few exceptions,
do not support a single-nutrient or reductionist focus. Diet
intervention studies that involve whole foods and overall dietary
patterns are currently underway, although reliance on self-report
dietary data in these studies is problematic. Thus, biological
indicators of diet and cellular activities are crucial in the
evaluation of the effect of these nutritional interventions in addition
to the examination and consideration of diet-gene interactions.
Diet-gene interactions likely contribute considerably to the
observed variations in cancer risk in response to exposures to
nutritional factors. Examples of diet intervention studies with a focus
on biomarkers are the Women’s Healthy Eating and Living (WHEL) Study
and the Cancer Prevention for Women (CAPRE) Study. The WHEL Study is
testing whether a high-vegetable, low-fat diet can reduce risk for
breast cancer recurrence. The CAPRE Study is testing whether a diet
high in vegetables and fruit can increase the regression rate of
cervical dysplasia. Biological indicators of diet in these studies
demonstrate that intakes of vegetables and fruits can be substantially
increased in these target populations of women.
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A Role for Vitamin D Receptor Agonists in the Treatment of
Prostate Cancer. Nancy Weigel. Department of Molecular
and Cellular Biology, Baylor College of Medicine, Houston, TX.
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Epidemiological studies show an inverse correlation between exposure to
sunlight—the chief source of vitamin D for Americans—and prostate
cancer mortality. Vitamin D is subsequently metabolized to its active
form, 1,25-dihydroxyvitamin D (calcitriol), which acts through the
vitamin D receptor (VDR), a member of the nuclear receptor family of
transcription factors. We and others have found that calcitriol
inhibits the growth of LNCaP and to a lesser extent PC3, prostate
cancer cells, in vitro. However, the concentration required to achieve
this would cause hypercalcemia in vivo. The development of less
calcemic VDR agonists raises the possibility that these may be of use
clinically in the treatment of prostate cancer. We have tested two VDR
agonists—a calcitriol analogue, EB1089, from Leo Pharmaceutical
Products and a novel nonsecosteroid VDR agonist from Ligand
Pharmaceuticals—in an LNCaP nude mouse tumor model. Both compounds
inhibit tumor growth without inducing hypercalcemia or causing weight
loss. In vitro calcitriol treatment causes both
G0/G1 accumulation and
induces apoptosis with concomitant down-regulation of Bcl-2 and
BclXL in LNCaP cells. PC-3 cells, although growth
inhibited, exhibit neither of these changes. Our studies show that in
these cells, calcitriol induces transforming growth factor-ß
(TGF-ß) production, which then inhibits the growth of the cells. In
contrast, LNCaP cells are resistant to TGF-ß treatment. Collectively,
these studies show that calcitriol induces pleiotropic responses and
that analogs of calcitriol may be useful alone or in combination with
other compounds in the treatment of prostate cancer.
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Human Selenium Binding Protein in Prostate Cancer. Arthur J. Sytkowski. Beth Israel Deaconess Medical Center, Boston, MA.
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A deficiency in the essential trace element selenium has long been
associated with increased cancer risk, including an increased incidence
of liver, lung, colon and prostate cancer. A recent clinical trail of
dietary selenium supplementation resulted in a reduction in total
cancer mortality, total cancer incidence and incidences of prostate,
lung and colorectal cancer. Even more recently, a marked inverse
correlation between dietary selenium and the risk of advanced prostate
cancer was reported. However, despite the strong association of
selenium deficiency with malignancy, the precise mechanism of this
effect is not understood, although several hypotheses have been
proposed. In experimental rodent in vivo systems, dietary selenium
supplementation inhibits tumorigenesis. It has been proposed that this
cancer-protective effect of selenium in rodents may be mediated by
one or more selenium-binding proteins. In an effort to identify
genes whose expression in human prostate cancer is correlated either
with an aggressive or an indolent phenotype, we have discovered that
the human homologue of one of these selenium-binding proteins,
designated hSP56, is expressed by the slowly growing,
androgen-sensitive human prostate cancer cell line LNCaP but not by
the more rapidly-growing, androgen-insensitive line PC-3.
Interestingly, hSP56 expression was markedly increased after
growing LNCaP cells in androgen-depleted medium, suggesting that
androgen may down-regulate hSP56 expression
simultaneously with its growth-promoting effect. We confirmed this
by showing that the addition of androgen to the cells greatly reduced
hSP56 transcript levels. We also quantified steady state
hSP56 mRNA levels in normal human tissues and in several
human neoplastic tissues and cells. Pronounced differences were found
among the tissues and cancer cell lines examined. Among normal tissues,
high levels were observed in liver, lung, colon, prostate, kidney and
pancreas and moderate levels were found in spleen, ovary and heart. In
thymus, testis, peripheral blood leukocyte and brain the expression was
barely detectable. In contrast, hSP56 expression was
detectable in only 4 of 11 neoplastic cell lines examined. Currently
ongoing gain-of-function and loss-of-function studies should help to
elucidate the role of hSP56 in the prostate cancer
phenotype.
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Vitamin A and Its Receptors in Breast Cancer. Xiao-kun Zhang, Bingzhen Lin, Hui Li, Siva Kumar Kolluri, Xihua Cao, Guo-quan Chen and Feng Lin. Burnham Institute, La Jolla, CA.
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Vitamin A and its natural and synthetic analogs, retinoids, regulate a
broad range of biological activities largely through two types of
nuclear receptors—the retinoic acid receptors (RARs) and retinoid X
receptors (RXRs). These receptors are members of the
steroid-thyroid-retinoid hormone receptor superfamily. Epidemiological,
in vitro and animal studies also indicate that vitamin A derivatives
are promising anti–breast cancer agents. The anti-cancer effects
of vitamin A are mainly mediated by one of the vitamin A receptors, the
RARß, which induce growth inhibition and apoptosis and inhibit
activities of oncogenes c-jun and c-fos (Lin et al. 2000
, Liu et al. 1996
, Wu et al. 1997
, Wu et al. 1997
).
Unfortunately, RARß expression is often lost in
estrogen-independent breast cancer cells. This has contributed
significantly to the loss of anti-cancer activities of vitamin A in
these hormone-refractory cancer cells, a major obstacle in vitamin
A treatment. Expression of RARß is mainly mediated by a retinoic
acid–responsive element (ßRARE) in the RARß promoter, which binds
to RAR/RXR heterodimers. Recent studies from our laboratory demonstrate
that lack of RARß expression in estrogen-independent breast
cancer cells is due to loss of orphan receptor COUP-TF (Lin et al. 2000
). COUP-TF, through its binding to a DR-8 element present in
the RARß promoter, acts as an accessory protein for RAR to recruit
its coactivator CBP (CREB-binding protein) and induces transcriptional
activity of the RARß promoter in a retinoic acid–and
RAR-dependent manner (Lin et al. 2000
). These findings provide a
molecular mechanism for down-regulation of RARß in
estrogen-independent breast cancer cells and possibilities of
restoring vitamin A activity in these cells. In searching for
alternative approaches to induce RARß expression in
estrogen-independent breast cancer cells, we observed that a group
of specific vitamin A derivatives that bind selectively to RXR strongly
induce RARß expression in estrogen-independent breast cancer
cells (Wu et al. 1997
). These compounds activate RARß promoter
activity through activation of RXR/nur77 heterodimers that also bind to
ßRARE. Recently, we also studied another group of vitamin A
derivatives that are potent inducers of apoptosis of
estrogen-dependent and -independent breast cancer cells. These
derivatives induce apoptosis independently of retinoid receptor (Li et al. 1998
). Their proapoptotic effects require expression of orphan
receptor TR3 (also called nur77 or NGFI-B) (Li et al. 2000
). TR3, in
response to the compounds, translocates from the nucleus to the
cytoplasm, where it targets mitochondria and induces cytochrome c
release and apoptosis. Our results demonstrate a novel mechanism by
which a nuclear transcriptional factor translocates to mitochondria to
initiate apoptosis and also indicate that these new vitamin A
derivatives may represent a novel class of compounds suitable for
treatment of estrogen-independent breast cancer cells.
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Diet and Mammographic Densities. N. F.
Boyd. Ontario Cancer Institute Toronto, Canada.
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We are conducting a long-term randomized controlled trial to find
out whether intervention with a low-fat, high-carbohydrate diet
reduces breast cancer risk. Subjects with extensive mammographic
densities, a risk factor for breast cancer, were randomly assigned to
an intervention group and given intensive individual counseling aimed
at reducing total dietary fat to 15% of energy or to a control group
that received general advice about diet but were not counseled to
change their fat intake. Mammograms from 817 subjects taken at baseline
and compared with those taken 2 y later by using a quantitative
image analysis system and without knowledge of the dietary group of the
subjects or of the sequence in which pairs of images had been taken.
After 2 y the total area of the breast was reduced by an average
of 246.2 mm2 (2.5%) in the intervention group
compared with an average of 9.2 mm2 (<0.1%) in
controls (P = 0.01). The area of density was reduced by
376.6 mm2 (6%) in the intervention group
compared with an average of 153.1 mm2 (2.5%) in
the control group (P = 0.02). The effect of the
intervention on breast area was no longer statistically significant
after weight change, menopausal status and age at trial entry were
taken into account (P = 0.10). The effect of the
intervention on area of density remained statistically significant
after controlling for weight loss, age at entry and menopausal status
(P = 0.05).
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Conjugated Linoleic Acid Acts Directly on the Mammary
Epithelium and Indirectly through the Mammary Stroma to Prevent Breast
Cancer. M. M. Ip, D. Zangani, P. Masso-Welch, C.
Ip, S. Shoemaker, D. Liu and B. Lisafeld. Roswell Park Cancer
Institute, Buffalo, NY.
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Conjugated linoleic acid (CLA), a trace fatty acid found in meat and
dairy products, inhibits rat mammary carcinogenesis at low levels in
the diet. To determine possible mechanisms by which CLA exerts its
effects, the following studies were carried out. CLA (
40%
c9, t11- and 40% t10,
c12-octadecadienoic acids) inhibited growth of rat mammary
epithelial cells in three-dimensional primary culture with both an
inhibition of DNA synthesis and a stimulation of apoptosis contributing
to this effect. CLA also inhibited the growth of initiated rat mammary
epithelial cells as well as rat and mouse mammary tumor cell lines; in
these studies, t10, c12-CLA was considerably more
potent than c9, t11-CLA. In addition to this
direct action on epithelial cells, we also found that CLA may act
indirectly, by altering the microenvironment in which tumors develop.
In these studies we examined the effect of CLA on a multipotent stromal
cell population that is present in the rat mammary gland and is able to
acquire a fibroblastic, adipocytic or endothelial phenotype, depending
on culture conditions (Zangani et al. Differentiation 64: 91–101, 1999). We hypothesized that CLA might direct mammary stromal
cell differentiation towards a particular pathway. In these
experiments, t10, c12-CLA was found to be a
potent adipogenic factor, stimulating adipocyte differentiation even in
the absence of exogenous hormonal supplementation; c9,
t11-CLA was less effective. Conversely, CLA inhibited the
formation of capillary-like structures formed by the mammary
stromal cells when cultured on a reconstituted basement membrane. This
suggests that one mechanism by which CLA exerts its preventive action
on mammary carcinogenesis may be through an inhibition of angiogenesis.
Indeed, in preliminary studies in which angiogenesis was investigated
in vivo using a model of capillary invasion into growth
factor–impregnated matrigel, we found that functional angiogenesis was
inhibited in mice fed a diet containing 1% or 2% CLA. Taken together,
these observations suggest that CLA may prevent breast cancer both by a
direct effect on the mammary epithelium and a modification of the
differentiation program of mammary stromal cells.
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Fats, Lipoxygenases and Prostate Cancer. Charles
E. Myers and Jagat Ghosh. Cancer Center, University of Virginia
School of Medicine, Charlottesville, VA.
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Extensive epidemiology studies have linked a diet high in animal
products with an increased risk of prostate cancer. Arachidonic acid is
one fatty acid found in animal products but not in plant products. In
the late 1980s and early 1990s a series of articles reported that
arachidonic acid stimulated the growth of prostate cancer cells in
tissue culture. Furthermore, this growth stimulation was blocked by
drugs that inhibited all three lipoxygenases. At the time, it proved to
be impossible to identify the specific eicosanoid product that
stimulated cell growth. In previously published studies, we showed that
the proliferative response to arachidonic acid was blocked by
inhibitors of 5-lipoxygenase. When we examined the products of
5-lipoxygenase, the leukotrienes were inactive whereas
5-hydroxy-6,8,11,14-eicosatetraenoic acid (5-HETE), 5-HETE lactone and
5-oxoETE effectively stimulated proliferation. In the conduct of these
experiments, we noted that complete inhibition of 5-lipoxygenase caused
rapid onset of apoptosis. When we examined this process of cell death
in greater detail, we found that the apoptosis had a number of
interesting properties. First, mitochondrial permeability transition
occurred within 45 min, cell surface blebbing within 60 min and DNA
degradation by 120 min. Apoptosis was complete within 8 h.
Apoptosis was also independent of cell cycle and was not blocked by the
addition of testosterone or epidermal growth factor. We next proceeded
to examine how the presence of 5-HETE blocks apoptosis. Our results
indicate that this eicosanoid acts by blocking Fas-mediated cell
death. We were first able to document that prostate cancer cells
contain large amounts of preformed Fas and FAS-ligand (Fas-L), but
these proteins were sequestered intracellularly rather than being
arrayed on the cell surface were they could initiate death signaling.
Within 15 min after addition of the 5-lipoxygenase inhibitor, MK886,
Fas-L appeared at the cell surface, with Fas arriving later at 30
min. At 30 min the adaptor protein FADD also was found associated with
FAS. This was followed by recruitment of procaspase 8 to the death
signaling complex, activation of caspase 8 and, finally, activation of
caspase 3. These results show that inhibition of 5-HETE formation
causes rapid translocation of preformed Fas and Fas-L to the cell
surface followed by initiation of Fas-mediated death signaling.
Although the onset of apoptosis was temporally associated with
activation of Fas-signaling, this does not prove that Fas
activation is the cause of cell death. To establish this link, we next
treated the cells with antibodies to Fas and Fas-L. Unfortunately,
these antibodies were only able to partially block the onset of
apoptosis. There were a number of reasons why this might have happened.
First, other death pathways might have been activated. Second, the
antibodies we used may not compete effectively with Fas-L for Fas.
Third, inhibition of 5-HETE formation resulted in the formation of
macromolecular clusters of Fas that have been known to trigger cell
death independent of Fas-L. To clarify this issue, we next tested
the effect of a dominant negative FADD construct in which the portion
of the protein responsible for the recruitment and activation of
caspase 8 was replaced with green florescent protein. This dominant
negative FADD was able to completely block the onset of apoptosis
following inhibition of 5-lipoxygeanse. In summary, we have shown that
5-HETE supports the survival of prostate cancer cells through a novel
pathway that blocks translocation of Fas and Fas L to the cell surface.
Furthermore, this survival pathway functions independent of cell cycle,
P53 status or Akt phosphorylation.
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Inhibition of COX-2. A Novel Approach to Breast Cancer
Prevention. Andrew J. Dannenberg. Weill Medical College of Cornell
University and Strang Cancer Prevention Center, New York, NY.
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There are two distinct isoforms of cyclooxygenase (COX) that catalyze
the formation of prostaglandins from arachidonic acid. These enzymes
have been designated COX-1 and COX-2. COX-1 is constitutively expressed
in most tissues whereas COX-2 results from an inducible
early-response gene. There is considerable evidence that COX-2 is
important in carcinogenesis. COX-2 is overexpressed in transformed
cells and in a variety of human malignancies including breast and colon
cancer. Knocking out the COX-2 gene markedly reduced the tumor
incidence in experimental models of intestinal and skin tumorigenesis.
Moreover, selective COX-2 inhibitors suppressed the formation of tumors
in numerous animal models including experimental breast cancer. A major
interest of this laboratory is developing strategies to suppress COX-2
gene expression as an approach to cancer prevention. Toward this end,
it is important to elucidate the signal transduction pathways that
regulate COX-2 gene expression and identify natural or synthetic
substances that block the induction of COX-2. We have shown that the
HER-2/neu oncogene and tumor promoting phorbol esters stimulate
AP-1-mediated induction of COX-2 transcription in human mammary
epithelial cells. We identified a variety of small molecules including
phenolic antioxidants, retinoids, and triterpenoids that inhibit
AP-1-mediated induction of COX-2. The identification of inhibitors of
COX-2 transcription may lead to novel strategies to prevent or treat
cancer.
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Tumor Angiogenesis: A Target for Dietary Interventions. Steven K. Clinton. Ohio State University, Arthur G. James
Cancer Hospital and Richard J. Solove Research Institute, Columbus, OH.
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Blood vessels supply the oxygen and nutrients that are crucial for
normal cell function and survival and also remove metabolic waste
products. Virtually all cells in normal tissues reside within 100 µm
of a capillary. The growth of new blood vessels—called
angiogenesis—is a tightly regulated, transitory and elegantly
orchestrated process during organogenesis, reproduction and tissue
repair after injury. In contrast, unrestricted angiogenesis is a
fundamental component of carcinogenesis contributing to local
progression and systemic metastases. Premalignant and early aberrant
proliferative lesions seem to initially lack the ability to stimulate
angiogenesis. However, a hallmark of progression to larger and more
aggressive neoplasia is the development of angiogenic ability.
Angiogenesis is regulated in tissues and tumors by counterbalancing
positive and negative signals involving several dozen soluble secreted
factors as well as cell-cell and cell-matrix interactions. The
ability to stimulate angiogenesis seems to be acquired at a discrete
point in tumor progression—termed the angiogenic switch—when the
proangiogenic process becomes dominant and normal vascular quiescence
is overcome. The mechanisms underlying the shift in the balance between
proangiogenic regulators and antiangiogenic regulators remain
incompletely understood and may differ among tumors of various etiology
and of different tissues. The scientific evidence for the importance of
angiogenesis in cancer is now very compelling. For example, rodent
studies demonstrate that tumors genetically engineered to overexpress
angiogenic growth factors have enhanced growth. Conversely, antibodies
to potent angiogenic growth factors, such as vascular endothelial
growth factor, impair neovascularization and the growth of tumors. The
more recent discovery and synthesis of several antiangiogenic growth
factors led to studies demonstrating dramatic tumor inhibition in mice.
Examination of several types of human cancer showed that those with
greater microvessel density—a biomarker of enhanced angiogenesis—have
a greater risk of metastatic spread and risk of death. It is apparent
that tumor angiogenesis offers an attractive target for diet and
nutritional interventions that may inhibit carcinogenesis or the growth
and progression of early cancers. Ongoing studies using in vitro and in
vivo models suggest that energy balance, several nutrients and a
variety of dietary components such as phytochemicals may influence the
balance between angiogenic and antiangiogenic regulators. Future
studies will further explore tumor angiogenesis as a novel target of
diet and nutritional interventions focusing on cancer prevention and as
a complement to therapeutic strategies.
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Nutritional Modulation of Human...
Molecular Epidemiology of...
