Nutrition and Gene Regulation

Molecular Targets for Bioactive Food Components1

J. A. Milner2
Nutritional Sciences Research Group, Division of Cancer Prevention, National Cancer Institute,
Department of Health and Human Services, Rockville, MD 20852

ABSTRACT Mounting evidence points to dietary habits as an important determinant of cancer risk and tumor
behavior. Although the linkages with diet are intriguing, the literature is also laden with inconsistencies. The reasons
for these inconsistencies are likely multi-factorial, but probably reflect variations in the ability of bioactive constit-
uents to reach or affect critical molecular targets. Fluctuations in the foods consumed not only influence the intake
of particular bioactive components, but may alter metabolism and potentially influence the sites of action of both
essential and nonessential nutrients. Genetic polymorphisms are increasingly recognized as another factor that can
alter the response to dietary components (nutritional transcriptomic effect) by influencing the absorption, metab-
olism, or sites of action. Likewise, variation in DNA methylation patterns and other epigenetic events that influence
overall gene expression can be influenced by dietary intakes. Furthermore, variation in the ability of food
components to increase or depress gene expression (nutrigenomic effect) may account for some of the observed

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inconsistencies in the response to dietary change. Because a host of food components are recognized to influence
phosphorylation and other posttranslational events, it is also likely that these and other proteomic modifications
account for at least part of the response and variation that is reported in the literature. Collectively, it is clear that
bioactive food components can influence a number of key molecular events that are involved in health and disease
resistance. As the era of molecular nutrition unfolds, a greater understanding of how these foods and components
influence cancer will surely arise. Such information will be critical in the development of effective tailored strategies
for reducing cancer burden. Just as important, however, is that as this information unfolds it is utilized within a
responsible bioethical framework. J. Nutr. 134: 2492S–2498S, 2004.

KEY WORDS: ● bioactive ● nutrigenomics ● nutrigenetics ● proteomics ● metabolomics

During the past several decades, enormous progress has been
made in developing national nutrition policies and research agen-
das that are fundamental to dealing with a host of problems faced
by segments of the population. Despite the many successes, nu-
trition-related health issues remain commonplace and account
for many of the leading causes of death of Americans. According
to the USDA, about $71 billion could be saved per year if
healthier diets were consumed because of reduced medical costs,
increased productivity, and extended lives (1). Since this esti-
mate only accounts for conditions such as coronary heart disease,
stroke, cancer, and diabetes—and not other diet-related dis-
1
Presented at the 6th Postgraduate Course on Nutrition entitled “Nutrition and
Gene Regulation” Symposium at Harvard Medical School, Boston, MA, March 13–
14, 2003. This symposium was supported by Conrad Taff Nutrition Educational Fund,
ConAgra Foods, GlaxoSmithKline Consumer Healthcare, McNeil Nutritionals, Nestle
Nutrition Institute, The Peanut Institute, Procter & Gamble Company Nutrition Sci-
ence Institute, Ross Products Division–Abbott Laboratories, and Slim Fast Foods
Company. The proceedings of this symposium are published as a supplement to The
Journal of Nutrition. Guest editors for the supplement publication were: W. Allan
Walker, Harvard Medical School, George Blackburn, Harvard Medical School, Ed-
ward Giovanucci, Harvard School of Public Health, Boston, MA, and Ian Sanderson,
University of London, London, UK.
2
To whom correspondence should be addressed.
E-mail: milnerj@mail.nih.gov.
3
Abbreviations used: ALL, acute lymphocytic leukemia; ATBC Study, Alpha-
Tocopherol, Beta-Carotene Cancer Prevention Study; methyleneTHF, 5,10-meth-
ylenetetrahydrofolate; methylTHF, 5-methyltetrahydrofolate; MTHFR, methyl-
enetetrahydrofolate reductase.
eases—the savings could be far greater. Obesity alone has been
estimated to cost $117 billion and osteoporosis another $14
billion per year in medical expenses. Thus, the appropriate use of
diet can have profound benefits on both personal health and
financial well-being.
The diet and cancer link
Some of the earliest descriptions of the symptoms of cancer
appeared in ancient Chinese and Arabic medical writings (2).
One of the first significant attempts to determine the geo-
graphical distribution of cancer and possible causes and link-
ages appeared in the early 19th century in Europe. Findings
from their questionnaire that were published in 1806 by the
Society for Investigating the Nature and Cure of Cancer noted
“with regard to cancer, it is not only necessary to observe the
effects of climate and local situation but to extend our views to
different employments, as those in various metals and manu-
factures; in mines and collieries; in the army and navy; in
those who lead sedentary or active lives; in the married or
single; in the different sexes, and many other circumstances”
(3). It is disheartening that some of the same issues continue
to be debated almost 200 y after their synopsis was published.
Regardless, evidence from a variety of sources points to dietary
habits as a significant environmental factor in the overall
cancer process. While optimizing the intake of specific foods

0022-3166/04 $8.00 © 2004 American Society for Nutritional Sciences.

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 MOLECULAR TARGETS FOR FOOD COMPONENTS
and/or their bioactive components seems a prudent, noninva-
sive, and cost-effective strategy for reducing cancer burden,
this is far from a simple process. The complexity of the diet
coupled with the heterogeneity of the cancer process makes it
a daunting task to establish true interrelationships. Neverthe-
less success is key to enormous societal and personal satisfac-
tion.
Bioactive food components
While epidemiological and preclinical evidence continues to
highlight dietary components ranging from macro- to micro-
constituents as likely modifiers of cancer processes, there are also
alarming inconsistencies in the literature (4– 6). Evidence al-
ready exists that not only can the magnitude of the response vary
across studies, but even the direction of the response can shift.
These inconsistencies may reflect the cumulative effects of nu-
merous variables that influence the effective quantity of the
bioactive component occurring at the target site or that influence
its ability to bring about a metabolic or phenotypic change.
Figure 1 illustrates the multiple steps where bioactive food com-
ponents can interact with genes and their expression in altering
phenotypes. Undeniably genes can influence absorption, metab-
olism, or transport of a bioactive food component (nutritional
transcriptomic effect) or its site of action and thus influence the
overall response to the diet. Likewise, bioactive food components
can alter the genetic expression of a host of cellular events
(nutritional transcriptomic effect) and thus influence cancer out-
comes. It has long been recognized that nutrients can modify
proteins once formed through a variety of processes including
phosphorylation. Thus, it is likely that nutritional proteomics is
in some way key to the changes in tumor incidence or behavior
that is observed in model systems following modification of the
diet or as suggested by epidemiological and limited clinical stud-
ies. Finally, if the effective concentration of the bioactive com-
ponent never reaches target tissue or is masked by other inter-
mediates, then metabolomics becomes a limiting factor.
Collectively, it is clear the response to bioactive food components
is highly dependent on a number of cellular events and regulatory
processes that attempt to maintain homeostasis and/or survival.
The complexity of the diet is illustrated by the number of
essential and nonessential food components that have been
proposed to modify one or more steps in the cancer process
(7). Table 1 provides an incomplete list of some of the
numerous food components that may be important in modi-
fying cancer risk and tumor behavior. Collectively, evidence
FIGURE 1 Influence of bioactive food components on genetic,
epigenetic, and proteomic events. A host of bioactive food components
are known to influence one or more stages of this process.
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exists that both essential and nonessential dietary components
can alter the cancer process. These dietary components are not
limited to plants, since zoochemicals arising from the con-
sumption of animal products can provide compounds like
conjugated linoleic acid and n-3 fatty acids that may also
influence cancer. Likewise, compounds arising from mush-
rooms (fungochemicals) have been proposed to have antican-
cer properties (8). Finally, it is conceivable that microorgan-
isms residing in the gastrointestinal tract may play a key role
by forming compounds (bacteriochemicals) that may increase
or decrease cancer (9,10).
The magnitude of the problem of identifying which dietary
component is most important in increasing or decreasing risk
is evident by the literally thousands of compounds consumed
through the diet each day. Likewise, the dearth of information
about some components limits the ability to unravel which
bioactive components are most important. For example,
whereas it is estimated that humans may be exposed to ⬎5000
flavonoids, only a few have been examined for their anticar-
cinogenic effects. Examination of classes of food components
as suggested in Table 1 may assist in expediting this process.
Nevertheless, this will be a complicated process because it is
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likely that multiple steps in the cancer process are being
modified simultaneously, including sites such as drug detoxifi-
cation, DNA repair, cell proliferation, apoptosis, differentia-
tion, and angiogenesis (Fig. 2). Overgeneralizations may also
lead to false impressions about what are key facts influencing
the cancer process. Clearly, absolute intake, duration of expo-
sure, and speciation are fundamental to the ability of any of
these bioactive food components to influence the cancer pro-
cess, at least in model systems (11,12). Although there is good
reason to believe that similar responses will occur in humans,
the literature is less plentiful with data. To move this area
forward additional attention is needed to determine the crit-
ical intake and duration required to bring about a desired
physiological change.
It is already evident that considerable variation exists in the
amount of bioactive components that occur in a particular
food. For example, the selenium content of foods is known to
be quite variable, likely because of the variation in the content
of the soil. This variation in soil content may also manifest
itself in other food items (such as animal products) that are
consumed. Recent evidence demonstrates the content of se-
lenium in beef, which is a significant source of selenium in the
North America, can range more than 2-fold depending on
where the cattle were raised (13). The content of organic
components can also vary significantly within and among
plants. ␣-tocopherol in broccoli has been reported to vary
from 0.46 to 4.25 mg/100 g fresh weight, with a mean of 1.62
(14). Assessing the intake of some constituents is particularly
daunting because of significant limitations in available com-
positional databases. Even a single database may be inadequate
because the content of food component in plants can depend
not only on preparation techniques, but also on various envi-
ronmental factors, including soil conditions, climate, season,
horticultural practices, and the type and age of the plant. Such
variations in food content can lead to wide fluxes in exposures
at potential target sites for cancer prevention. The variability
in intake of polyphenols arising from tea consumption illus-
trates the complexity of trying to examine a dietary factor as a
modifier of cancer risk. Although the daily mean intake of
polyphenols from tea for 116 individuals was estimated to be
80.8 mg, the range varied from 3 to 588 (15). Relatively few
studies have examined reproducibility of intake for specific
food items using the food frequency questionnaires. Using 2
different questionnaires for tea the correlation for black tea
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TABLE 1
Partial list of bioactive food components that may influence cancer risk and tumor behavior

Class Bioactive component Dietary source

Vitamins Vitamin D Dairy products

Folic acid Vegetables
Vitamin A Vegetables
Vitamin E (-tocopherol) Vegetable oils
Ascorbic acid Vegetables, fruits
Minerals Calcium Dairy products, vegetables
Selenium Cereal grains, meat, fish
Zinc Meat, vegetables
Carotenoids Lycopene Tomatoes
Lutein Dark green vegetables
␤-Carotene Orange-yellow vegetables
Flavonoids Genistein Soybeans, soy products
Resveratrol Grapes, red wine
Quercetin Vegetables, fruits
Tangeretin Citrus fruits
Catechins Grapes
(-)-Epigallocatechin-3-gallate Green tea
Organosulfur Allyl sulfur Allium vegetables
Diallyl sulfide Garlic
Isothiocyanates Allyl isothiocyanate Cabbage

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Benzyl isothiocyanate Garden cress
Sulforaphane Broccoli
Indoles Indole-3-carbinol Cruciferous vegetables
Monoterpenes D-Limonene Citrus fruit oils
D-Carvone Caraway seed oil
Phenolic acids Curcumin Turmeric, curry, mustard
Caffeic acid Fruits, coffee beans, soybeans
Ferulic acid Fruits, soybeans
Chlorogenic acid Fruits, coffee beans, soybeans
Chlorophyll Chlorophyll Green vegetables
Chlorophyllin Green vegetables

Adapted from Huang, M.-T., Osawa, T., Ho, C.-T., & Rosen, R. T. (1994) Food Phytochemicals for Cancer Prevention I: Fruits and Vegetables.
American Chemical Society, Washington, DC.

consumption was 0.68. Nevertheless, complicating this issue is
the uncertainty about the health benefits that might be attrib-
uted to intermittent vs. sustained intake of a given amount of
a particular food or its component(s).
FIGURE 2 Bioactive food components are known to influence
multiple biological processes. Determining which is most instrumental
in bringing about a phenotypic change is critical to the future of nutrition
and health because it will assist in determining who may benefit and
who may be placed at risk by intervention strategies.
Some of the most compelling evidence that diet can influ-
ence the cancer process comes from a rather recent nutritional
intervention study with selenium-yeast (16). Although spec-
ulation about the propensity of selenium to serve as an anti-
cancer agent has been expressed for ⬎30 y (17), this inter-
vention study provided some of the most intriguing evidence
for a biological response, although admittedly the protective
effects only surfaced in secondary analysis. Nevertheless, these
findings are largely supportive by population studies and ran-
domized clinical trials, suggesting an inverse association exists
between increased selenium intake and a reduction in esoph-
ageal, gastric, lung, prostate, and colorectal cancers. Preclini-
cal studies with animal models supply additional evidence that
an inverse association exists between the dietary intake of
selenium and carcinogenesis (18). Although the majority of
selenium’s role in health promotion has focused on its anti-
oxidant activity, it has several diverse biologic effects includ-
ing an ability to suppress cell proliferation, enhance immune
response, alter the metabolism of carcinogens, and induce
apoptosis. New genomic and proteomic technologies offer
exciting opportunities to examine each of these events simul-
taneously and ultimately to determine which is most instru-
mental in bringing about a change in the incidence or behav-
ior of a tumor.
New technologies and nutritional genomics
DNA microarrays (19,20) and serial analysis of gene ex-
pression (SAGE) (21) are being increasingly employed to
 MOLECULAR TARGETS FOR FOOD COMPONENTS
identify multi-site gene-food component interactions that
were merely dreams just a few years ago. Today, the expression
of literally thousands of genes can now be monitored simulta-
neously between normal and abnormal tissue and used to
identify and characterize the response to prevention and ther-
apeutic interventions. As the understanding about how bio-
active food components modify genetic events becomes
clearer, the current conundrum surrounding appropriate di-
etary recommendations should evolve into rational and per-
sonalized intervention strategies to improve health and mini-
mize the risk of disease.
The term nutritional genomics has been used to describe
work at the interface of plant biochemistry, genomics, and
human nutrition. The term was coined by DellaPenna (22) to
enhance the understanding of reactions and interactions at the
molecular or genomic level. As this area continues to unfold,
it is becoming increasingly apparent that a host of genomic
interrelationships with the diet exist which encompass the
broad topic of nutrigenomics. Genetic changes arising as a
result of single point mutations, rearrangements, or copy num-
ber involving either deletions or additions can likely influence
the response to various dietary components. Unfortunately,
relatively few causal diet and chronic disease associations can
be articulated with any degree of certainty. It is noteworthy
that important exceptions may include the relationship be-
tween dietary fat and coronary heart disease and the interre-
lationship between calcium consumption and fracture risk. In
both cases a biomarker, i.e., plasma lipoproteins and bone
mineral density, respectively, was identified that was respon-
sive to dietary intervention. Unfortunately, few if any surro-
gate or intermediate biomarkers have been validated for can-
cer. The absolute response to diseased conditions is becoming
increasingly recognized to depend on a number of gene poly-
morphisms, and thus may contribute to variation in response
and possibly account for the dearth of validated cancer
biomarkers. For example, those that are linked to dietary
habits appear to have a pivotal role in lipoprotein metabolism
and cardiovascular disease including apoproteins (apo) E, B,
A-IV, and C-III, LDL receptor, microsomal transfer protein
(MTP), fatty acid-binding protein (FABP), cholesteryl ester
transfer protein (CETP), lipoprotein lipase, and hepatic lipase
(23). Likewise, evidence exists that vitamin D receptor poly-
morphisms may influence the loss of bone mass resulting from
other dietary variables including caffeine consumption (24).
The health benefits attributed to soluble fiber also appear to be
linked with genetic backgrounds because data from Hegele et
al. (25) suggest that polymorphism occurring with the angio-
tensinogen gene can influence the response in blood pressure
brought about by these carbohydrates.
Vineis et al. (26) reviewed how metabolic polymorphisms
likely influenced the interrelationship between diet and can-
cer. The prevalence of these polymorphisms is increasingly
being recognized as a significant variable that may influence
the interpretation of results of otherwise solidly-designed stud-
ies. Linkages between polymorphisms in genes associated with
drug metabolism and dietary habits are becoming more appar-
ent (27,28) and are not only important for explaining alter-
ations in the initiation phase of carcinogenesis, but also inter-
actions with chemotherapeutic agents. A low prevalence of
polymorphisms in genes coding for activation (phase I) en-
zymes CYP1A1 (0.07) and CYP2E1 (0.02) was observed in a
sample of subjects involved with the Alpha-Tocopherol, Beta-
Carotene Cancer Prevention Study (ATBC Study), while
there was also a high prevalence in genes coding for detoxifi-
cation (phase II) enzymes GSTM1 (0.40) and NQO1 (0.20)
(29). It is not clear what impact these gene profiles had on the
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overall outcome of the study. Interestingly, 7 of 10 members of
this sample carried the VDR-Taq1 polymorphism (t) associ-
ated with lower risk for prostate cancer, which may account in
part for lower cancer rates in Finland when compared with the
United States (29). Further, in a nested case-control study
within the ATBC Study, glutathione peroxidase 1 (hGPX1),
a selenium-dependent enzyme involved in detoxification of
hydrogen peroxide, was found to have a polymorphism exhib-
iting a proline to leucine replacement at codon 198. This
polymorphism conferred a relative risk for lung cancer risk of
1.8 for heterozygotes and 2.3 for homozygous variants com-
pared with homozygote wild types (30). More recently, poly-
morphism in glutathione peroxidase has been linked to an
increased risk of breast cancer (31).
Several genetic commonplace polymorphisms can influence
folate homeostasis and thus may assist in explaining the link-
age between this B-vitamin and cancer (32,33). Polymor-
phisms of the methylenetetrahydrofolate reductase (MTHFR)
gene code for the protein that converts 5,10-methylenetetra-
hydrofolate (methyleneTHF), the major form of intracellular
folate, to 5-methyltetrahydrofolate (methylTHF), the major
form of circulating folate in plasma. MethyleneTHF, a cofactor
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for methylating deoxyuridylate to produce thymidylate, is the
rate-limiting nucleotide in DNA synthesis. Depressed
MTHFR activity increases the availability of methyleneTHF,
thereby reducing the chances of inadequate thymidylate and
misincorporation of uracil into DNA. The reduction in uracil
incorporation into DNA reduces chromosome instability and
potentially a reduced risk of cancer (32). Epidemiologic studies
revealed that when folate intake is adequate, colorectal cancer
risk is reduced by ⬎50% in individuals with the MTHFR
677TT genotype compared with the MTHFR 677CC geno-
type, and risk of adult acute lymphocytic leukemia (ALL) is
reduced by 77% (34). When folate intake was inadequate,
however, the genetic profile became less important. Recent
clinical data demonstrated that genomic DNA hypomethyla-
tion was greater in leukocytes of individuals with the MTHFR
677TT genotype compared to those with the MTHFR 677CC
genotype and was inversely correlated with folate status (35).
Several studies suggested that reducing MTHFR activity
may limit the availability of methyl groups for synthesis of
S-adenosylmethionine (SAM) and increase DNA hypomethy-
lation, and that when folate status is low this mechanism may
negate the beneficial effect of the MTHFR 677TT polymor-
phism (35). MTHFR 1298 A-C polymorphism may also have
an influence on adult ALL (34).
The diversity of molecular targets that can be influenced by
food components highlights the complexity and breadth of
interactions. Figure 2 and 3 reveal that the effects of a bio-
active food component may be a manifestation of nuclear and
cytoplasmic events that regulate the abundance and/or activity
of specific proteins involving several cancer processes. Fluctu-
ations in these proteins can lead not only to changes in overall
cellular metabolism, but also can markedly influence the pro-
portion of cells that are dividing, undergoing apoptosis, or
differentiating. Characterizing which of these molecular
events is most important in bringing about a phenotypic effect
is the primary thrust of an RFA that has been released by NCI
during the last year. This RFA (CA 03– 001), entitled “Col-
laborations on Nutritional Modulation of Genetic Pathways
Leading to Cancer,” will provide financial support to assist in
building teams to address fundamental molecular issues about
the mechanism(s) by which bioactive food components influ-
ence the cancer process. It is anticipated that this RFA will be
re-released soon.
2496S SUPPLEMENT
FIGURE 3 Model diagram for ex-
amining the interrelationship between
bioactive food components and events
involved with the cancer process. Mod-
ified figure from the Cancer Genome
Anatomy Project produced by BioCarta
Inc. and the article published by Smelzer
and Kim (67).
Nutrigenomics
Rather compelling evidence that a variety of nutrients
interact with specific molecular targets (36 –39). Invariably,
studies reveal that a host of messages are modified by the
presence or absence of a food component. Our understanding
of the cellular events that allow for these collective changes
remains obscure. However, as presented in this symposium
common events such as mRNA stability and polymorphisms
may help explain the multitude of effects that are observed
(40). While understanding the basis for the multiple nutri-
genomic effects is important, it is also fundamental to under-
stand that several processes are influenced simultaneously.
Thus, it is likely that the clustering of gene expression changes
and then expressing these in terms of a ratio with some other
process may be most informative. Such an approach was used
with some success in examining shifts in cell proliferation and
apoptosis (41– 43). Additional characterization of these mes-
sage profiles will be needed to examine for commonality and
disparities in response among various nutrients so that poten-
tial synergies and antagonisms can be identified. Figure 3
provides an example for the way that multiple bioactive food
components may influence one or more steps in a process.
Such interrelationships will need to be examined to under-
stand the influence of the entire diet or possibly the food
matrix on biological responses. Knowledge about genes and
food components influencing key cellular processes will surely
be fundamental for determining who will and will not respond
to intervention strategies.
Transgenic and knockout models provide evidence that genes
can markedly influence the response to dietary components (43–
45). Likewise, the use of transgenic and knockout models can
provide valuable clues about the site(s) of action of specific
bioactive food components (36,43,46,47). Without question,
more information is needed to examine how genetic profiles
influence the ability of specific dietary components to bring about
phenotypic changes. Understanding the dynamic interactions
between food components and molecular targets is the basis of a
recent RFA that focuses on prostate cancer (RFA 03– 003). It is
believed that enhanced emphasis on molecular targets for pros-
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tate cancer will energize the nutrition community to explore
molecular sites of actions in other tissues.
Microarray technology that allows parallel analysis of ex-
pression patterns of a large number of genes in a single exper-
iment has created an exciting new frontier for the nutrition
and health care community. Unfortunately, far too frequently
conclusions are being drawn on the basis of limited observa-
tions. It is becoming increasingly apparent that cellular vari-
ation in gene expression profiles requires that multiple chips be
examined for each experimental condition. Studies by Wang
et al. (48) suggest a minimum of 5 cDNA chips may be needed
to detect a doubling or reduction in half of the expression of
a gene. As the desire to detect even small changes in gene
expression grows, it will likely be necessary to examine even
greater numbers of samples. The vast amount of gene expres-
sion data that will be generated from these multiple samples
requires a robust database system that will facilitate efficient
data storage, retrieval, secure access, data dissemination, and
integrated data analyses. It will also be necessary to examine if
commonality in response across tissues is occurring. A recent
NIH Announcement (CA 03– 027) is aimed at supporting
research to determine if bioactive food components alter gene
expression across various tissues and if there is consistency in
response among animal models and what occurs in humans.
Proteomics
Techniques are being developed and refined to assist in the
identification of the proteome, defined as all of the proteins
present in a particular cell at a particular moment (49). The
importance of these types of investigations stems from the fact
that gene expression does not always correlate with protein ex-
pression and the influence of food components may be either
translational or posttranslational, rather than at the transcrip-
tional level (50,51). There are a host of examples about how
specific dietary components can alter protein phosphorylation
and glycosylation (52–54). Some of these changes may occur
because of several processes including oxidative stress (55,56). In
some cases this may be through shifts in the quantity or expres-
 MOLECULAR TARGETS FOR FOOD COMPONENTS
sion of specific kinases and/or phosphatases. Thus, proteomic-
based studies, although technically challenging, complement
genomic studies and are essential in any comprehensive research
strategy aimed at examining the molecular processes and factors
involved in modulating disease risk. The use of current proteomic
tools will not only advance the field of nutritional sciences, but
will make the discipline more valued for its involvement in
disease prevention including cancer.
Metabolomics
The response to a bioactive food component cannot be
considered to occur in isolation but must be considered in the
context of the entire diet (57). Although interactions occur-
ring among nutrients remain an area poorly examined, there
are a few examples that illustrate potential negative and pos-
itive interactions. For example, ascorbic acid can reduce the
ability of selenium to retard a chemically induced tumor,
presumably by reducing it to elemental selenium (58). How-
ever, other constituents of the diet such as allyl sulfur com-
pounds found in processed garlic can enhance the anticancer
potential of selenium (59). Studies aimed at examining inter-
actions may simultaneously provide important insights into
site of action of various food components. Verification of the
importance of such changes in terms of phenotypic outcome
will be key to the usefulness of these or other types of predic-
tive models. One approach to examining interactions among
nutrients may be through metabolomic research. Metabolom-
ics involves the systematic estimation of metabolomes, i.e., the
characterization of all metabolites and small molecular weight
compounds occurring in an organism. While methods for
metabolomics are in the early stages of development, they
nonetheless offer exciting opportunities for the determination
of relationships between exposures and phenotypic outcomes
and possible interactions among nutrients. Various approaches
have been suggested and many analyses and visualizations are
currently being explored to obtain an operational view of
metabolomics (60 – 62).
As with other techniques, how to deal with the reams of
data are of paramount importance (63). Several methods,
including principal component analysis and clustering, are
being examined to analyze metabolomic data. Metabolomic
subsets, such as the lipid and amino acid metabolome, are
already suggesting that some very useful information can be
obtained (64,65).
Future research directions
Nutritional sciences remain an exciting and relevant research
area. The success of this discipline depends on dedicated scientists
who utilize the rapidly developing technologies to identify mo-
lecular sites of action of bioactive food components and how
these sites relate to disease prevention. Without a doubt, identi-
fying the mechanisms will be challenging, especially considering
the multiple sites of interactions among individual food compo-
nents, and the influence of genetic susceptibilities. Nevertheless,
innovative strategies should identify components of this mecha-
nistic puzzle so that a greater insight into the relationship be-
tween diet and health can be established. Resolving conflicting
and inconsistent data among studies will surely be one of the most
important aspects of basic research on the mechanisms of action
of bioactive food components.
Many research questions relevant to nutrition and cancer
remain to be addressed. For instance, how do interactions
among individual food components influence genetic expres-
sions occurring within a cell? Can the enhanced antiprolifera-
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tive effects of combining bioactive food components found in
vitro (66) be verified in vivo? Furthermore, is there common-
ality in response to bioactive components across tissues? How
do age and gender influence the response to food components
on molecular targets? How can interindividual variance in
nutrient metabolism best be taken into account? Can biomar-
kers that indicate nutrient status be identified and validated?
How might exfoliated cells be used as predictors of the re-
sponse to dietary intervention strategies? Does inhibition of a
cancer-related pathway by the interaction of a nutrient with a
specific molecular target cause compensatory activity to be
initiated through other pathways, possibly negating any ben-
eficial effect of the nutrient? If nutrients interact with cancer-
related molecular targets through several mechanisms, how
can the targets/mechanisms most important for reducing can-
cer risk be identified? Can nutrient-modulated biomarkers that
can serve as surrogate endpoint biomarkers for clinical disease
be identified and validated? Will currently used study designs
and analytical techniques be adequate in molecular-based nu-
trition and cancer research?
The answers to these and other questions may depend on
the development of effective collaborations between nutrition
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scientists with others because it is unlikely that single individ-
uals and/or institutions will have sufficient capabilities to
adequately address some of the overarching issues. Perhaps it is
important to recognize that developing and implementing this
new research paradigm in nutrition and cancer prevention will
take considerable time and patience. While the challenges will
be enormous the potential rewards in terms of both cancer
morbidity and mortality will be of equally great magnitude.
LITERATURE CITED
1. Frazao, E. (1999) High costs of poor eating patterns in the United
States. In: America’s Eating Habits: Changes and Consequences. USDA, Agri-
culture Information Bulletin No. 750 (Frazao, E., ed.), p. 5–32. Economic Research
Service, U.S. Department of Agriculture, Washington, DC.
2. Higginson, J. (1997) From geographical pathology to environmental
carcinogenesis: A historical reminiscence. Cancer Lett. 117: 133–142.
3. Society for Investigating the Nature and Cure of Cancer (1806) Edin-
burgh Med. Drug J. 2: 382–389.
4. Feskanich, D., Ziegler, R. G., Michaud, D. S., Giovannucci, E. L., Speizer,
F. E., Willett, W. & C. Colditz, G. A. (2000) Prospective study of fruit and
vegetable consumption and risk of lung cancer among men and women. J. Natl.
Cancer Inst. 92: 1812–1823.
5. Park, E. J. & Pezzuto, J. M. (2002) Botanicals in cancer chemopre-
vention. Cancer Metastasis Rev. 21: 231–255.
6. Olsen, A., Tjonneland, A., Thomsen, B. L., Loft, S., Stripp, C., Overvad, K.,
Moller, S. & Olsen, J. H. (2003) Fruits and vegetables intake differentially
affects estrogen receptor negative and positive breast cancer incidence rates. J.
Nutr. 133: 2342–2347.
7. Milner, J. A., McDonald, S. S., Anderson, D. E. & Greenwald, P. (2002)
Molecular targets for nutrients involved with cancer prevention. Nutr. Cancer 41:
1–16.
8. Wasser, S. P. (2002) Medicinal mushrooms as a source of antitumor
and immunomodulating polysaccharides. Appl. Microbiol. Biotechnol. 60: 258 –
274.
9. Guarner, F. & Malagelada, J. R. (2003) Gut flora in health and disease.
Lancet 361: 512–519.
10. Heerdt, B. G., Houston, M. A., Anthony, G. M. & Augenlicht, L. H. (1998)
Mitochondrial membrane potential (delta psi(mt)) in the coordination of p53-
independent proliferation and apoptosis pathways in human colonic carcinoma
cells. Cancer Res. 58: 2869 –2875.
11. Ip, C., Birringer, M., Block, E., Kotrebai, M., Tyson, J. F., Uden, P. C. &
Lisk, D. J. (2000) Chemical speciation influences comparative activity of
selenium-enriched garlic and yeast in mammary cancer prevention. J. Agric. Food
Chem. 48: 2062–2070.
12. Lamartiniere, C. A, Cotroneo, M. S., Fritz, W. A., Wang, J., Mentor-
Marcel, R. & Elgavish A. (2002) Genistein chemoprevention: timing and mech-
anisms of action in murine mammary and prostate. J. Nutr. 132: 552S–558S.
13. Hintze, K. J., Lardy, G. P., Marchello, M. J. & Finley, J. W. (2001) Areas
with high concentrations of selenium in the soil and forage produce beef with
enhanced concentrations of selenium. J. Agric. Food Chem. 49: 1062–1067.
14. Kurilich, A. C., Tsau, G. J., Brown, A., Howard, L., Klein, B. P, Jeffery,
E. H., Kushad, M., Wallig, M. A. & Juvik, J. (1999) Carotene, tocopherol, and
2498S
ascorbate contents in subspecies of Brassica oleracea. J. Agric. Food Chem. 47:
1576 –1581.
15. Hakim, I. A., Hartz, V., Harris, R. B., Balentine, D., Weisgerber, U. M.,
Graver, E., Whitacre, R.& Alberts, D. (2001) Reproducibility and relative validity
of a questionnaire to assess intake of black tea polyphenols in epidemiological
studies. Cancer Epidemiol. Biomarkers Prev. 10: 667– 678.
16. Clark, L. C., Combs, G. F., Jr., Turnbull, B. W., Slate, E. H., Chalker, D. K.,
Chow, J., Davis, L. S., Glover, R. A., Graham, G. F., Gross, E. G., Krongrad, A.,
Lesher, J. L., Jr., Park, H. K., Sanders, B. B., Jr., Smith, C. L. & Taylor, J. R.
(1996) Effects of selenium supplementation for cancer prevention in patients
with carcinoma of the skin. J. Am. Med. Assoc. 276: 1957–1963.
17. Schrauzer, G. N. (2000) Anticarcinogenic effects of selenium. Cell Mol.
Life Sci. 57: 1864 –1873.
18. Ip, C. (1998) Lessons from basic research in selenium and cancer
prevention. J. Nutr. 128: 1845–1854.
19. Moreno-Aliaga, M. J., Marti, A., Garcia-Foncillas, J. & Alfredo, M. J.
(2001) DNA hybridization arrays: a powerful technology for nutritional and
obesity research. Br. J. Nutr. 86: 119 –122.
20. Lobenhofer, E. K., Bushel, P. R., Afshari, C. A. & Hamadeh, H. K. (2001)
Progress in the application of DNA microarrays. Environ. Health Perspect. 109:
881– 891.
21. Polyak, K. & Riggins, G. J. (2001) Gene discovery using the serial
analysis of gene expression technique: implications for cancer research. J. Clin.
Oncol. 19: 2948 –2958.
22. DellaPenna, D. (1999) Nutritional genomics: manipulating plant micro-
nutrients to improve human health. Science 285: 375–379.
23. Vincent, S., Planells, R., Defoort, C., Bernard, M. C., Gerber, M., Prud-
homme, J., Vague, P. & Lairon, D. (2002) Genetic polymorphisms and lipopro-
tein responses to diets. Proc. Nutr. Soc. 61: 427– 434.
24. Rapuri, P. B., Gallagher, J. C., Kinyamu, H. K. & Ryschon, K. L. (2001)
Caffeine intake increases the rate of bone loss in elderly women and interacts with
vitamin D receptor genotypes. Am. J. Clin. Nutr. 74: 694 –700.
25. Hegele, R. A., Jugenberg, M., Connelly, P. W. & Jenkins, D.J.A. (1997)
Evidence for gene-diet interaction in the response of blood pressure to dietary
fibre, Nutr. Res. 17: 1229 –1238.
26. Vineis, P., Malats, N., Lang, M., d”Errico, A., Caporaso, N., Cuzick, J. &
Boffetta, P., eds. (1999) Metabolic polymorphisms and susceptibility to can-
cer. Lyon, France, International Agency for Research on Cancer.
27. Marchand, L. L., Wilkinson, G. R. & Wilkens, L. R. (1999) Genetic and
dietary predictors of CYP2E1 activity: a phenotyping study in Hawaii Japanese
using chlorzoxazone. Cancer Epidemiol. Biomark. Prev. 8: 495–500.
28. Eaton, D. L., Bammler, T. K. & Kelly, E. J. (2001) Interindividual
differences in response to chemoprotection against aflatoxin-induced hepatocar-
cinogenesis: implications for human biotransformation enzyme polymorphisms.
Adv. Exp. Med. Biol. 500: 559 –576.
29. Woodson, K., Ratnasinghe, D., Bhat, N. K., Stewart, C., Tangrea, J. A.,
Hartman, T. J., Stolzenberg-Solomon, R., Virtamo, J., Taylor, P. R. & Albanes, D.
(1999) Prevalence of disease-related DNA polymorphisms among participants
in a large cancer prevention trial. Eur. J. Cancer Prev. 8: 441– 447.
30. Ratnasinghe D., Tangrea, J. A., Andersen, M. R., Barrett, M. J., Virtamo,
J., Taylor, P. R. & Albanes, D. (2000) Glutathione peroxidase codon 198
polymorphism variant increases lung cancer risk. Cancer Res. 60: 6381– 6383.
31. Hu, Y. J. & Diamond, A. M. (2003) Role of glutathione peroxidase 1 in
breast cancer: loss of heterozygosity and allelic differences in the response to
selenium. Cancer Res. 63: 3347–3351.
32. Kim, Y.-I. (2000) Methylenetetrahydrofolate reductase polymor-
phisms, folate, and cancer risk: a paradigm of gene-nutrient interactions in
carcinogenesis. Nutr. Rev. 58: 205–209.
33. Rampersaud, G. C., Bailey, L. B. & Kauwell, G. P. (2002) Relationship
of folate to colorectal and cervical cancer: Review and recommendations for
practitioners. J. Am. Diet Assoc. 102: 1273–1282.
34. Skibola, C. F., Smith, M. T., Kane, E., Roman, E., Rollinson, S., Cart-
wright, R. A. & Morgan, G. (1999) Polymorphisms in the methylenetetrahydro-
folate reductase gene are associated with susceptibility to acute leukemia in
adults. Proc. Natl. Acad. Sci. U.S.A. 96: 12810 –12815.
35. Stern, L. L., Mason, J. B., Selhub, J. & Choi, S. W. (2000) Genomic
DNA hypomethylation, a characteristic of most cancers, is present in peripheral
leukocytes of individuals who are homozygous for the C677T polymorphism in the
methylenetetrahydrofolate reductase gene. Cancer Epidemiol. Biomark. Prev. 9:
849 – 853.
36. Yang, C. S., Maliakal, P. & Meng, X. (2002) Inhibition of carcinogenesis
by tea. Annu. Rev. Pharmacol. Toxicol. 42: 25–54.
37. Cheng, W. H., Quimby, F. W. & Lei, X. G. (2003) Impacts of glutathione
peroxidase-1 knockout on the protection by injected selenium against the pro-
oxidant-induced liver aponecrosis and signaling in selenium-deficient mice. Free
Radic. Biol. Med. 34: 918 –927.
38. Li, Y., Li, X. & Sarkar, F. H. (2003) Gene expression profiles of I3C- and
DIM-treated PC3 human prostate cancer cells determined by cDNA microarray
analysis. J. Nutr. 133: 1011–1019.
39. Li, Y. & Sarkar, F. H. (2002) Down-regulation of invasion and angio-
genesis-related genes identified by cDNA microarray analysis of PC3 prostate
cancer cells treated with genistein. Cancer Lett. 186: 157–164.
SUPPLEMENT
40. Cabelof, D. C., Yanamadala, S., Raffoul, J. J., Guo, Z., Soofi, A. &
Heydari, A. R. (2003) Caloric restriction promotes genomic stability by induc-
tion of base excision repair and reversal of its age-related decline. DNA Repair
(Amst.) 2: 295–307.
41. Beenken, S. W. & Bland, K. I. (2002) Biomarkers for breast cancer.
Minerva Chir. 57: 437– 448.
42. Creighton, C., Hanash, S. & Beer, D. (2003) Gene expression patterns
define pathways correlated with loss of differentiation in lung adenocarcinomas.
FEBS Lett. 540: 167–170.
43. Yang, W. C., Mathew, J., Velcich, A., Edelmann, W., Kucherlapati, R.,
Lipkin, M., Yang, K. & Augenlicht, L. H. (2001) Targeted inactivation of the
p21(WAF1/cip1) gene enhances Apc-initiated tumor formation and the tumor-
promoting activity of a Western-style high-risk diet by altering cell maturation in
the intestinal mucosal. Cancer Res. 61: 565–569.
44. Yang, X., Edgerton, S. M., Kosanke, S. D., Mason, T. L., Alvarez, K. M.,
Liu, N., Chatterton, R. T., Liu, B., Wang, Q., Kim, A., Murthy, S. & Thor, A. D.
(2003) Hormonal and dietary modulation of mammary carcinogenesis in mouse
mammary tumor virus-c-erbB-2 transgenic mice. Cancer Res. 63: 2425–2433.
45. Fong, L. Y., Ishii, H., Nguyen, V. T., Vecchione, A., Farber, J. L., Croce,
C. M. & Huebner, K. (2003) p53 deficiency accelerates induction and progres-
sion of esophageal and forestomach tumors in zinc-deficient mice. Cancer Res.
63: 186 –195.
46. Thornalley, P. J. (2002) Isothiocyanates: mechanism of cancer che-
mopreventive action. Anticancer Drugs 13: 331–338.
47. Gupta, S., Hastak, K., Ahmad, N., Lewin, J. S. & Mukhtar, H. (2001)
Inhibition of prostate carcinogenesis in TRAMP mice by oral infusion of green tea
polyphenols. Proc. Natl. Acad. Sci. U.S.A. 98: 10350 –10355.
48. Wang, T., Milner, M., Kim, Y. & Milner, J. (2003) Effects of Garlic
derived compound diallyl disulfide on prostate cancer cell LNCaP gene expres-
Downloaded from jn.nutrition.org by guest on May 4, 2014
sion: cDNA microarray expression profiling FASEB J. 17: A376.
49. Trayhurn, P. (2000) Proteomics and nutrition—a science for the first
decade of the new Millennium. Br. J. Nutr. 83: 1–2.
50. Banks, R. E., Dunn, M. J., Hochstrasser, D. F., Sanchez, J. C., Black-
stock, W., Pappin, D. J. & Selby, P. J. (2000) Proteomics: new perspectives,
new biomedical opportunities. Lancet 356: 1749 –1756.
51. Clarke, S. D. (2001) Polyunsaturated fatty acid regulation of gene
transcription: a molecular mechanism to improve the metabolic syndrome. J.
Nutr. 131: 1129 –1132.
52. Knowles, L. M. & Milner J. A. (2000) Diallyl disulfide inhibits p34(cdc2)
kinase activity through changes in complex formation and phosphorylation. Car-
cinogenesis 21: 1129 –1134.
53. Dees, C., Foster, J. S., Ahamed, S. & Wimalasena, J. (1997) Dietary
estrogens stimulate human breast cells to enter the cell cycle. Environ. Health
Perspect. 105 (Suppl 3): 633– 636.
54. Grancharov, K., Naydenova, Z., Lozeva, S. & Golovinsky, E. (2001)
Natural and synthetic inhibitors of UDP-glucuronosyltransferase. Pharmacol.
Ther. 89: 171–186.
55. Thompson, H. J., Heimendinger, J., Haegele, A., Sedlacek, S. M., Gillette,
C. & O’Neill, C., Wolfe, P., Conry, C. (1999) Effect of increased vegetable and
fruit consumption on markers of oxidative cellular damage. Carcinogenesis 20:
2261–2266.
56. Le Marchand, L. (2002) Cancer preventive effects of flavonoids–a
review. Biomed. Pharmacother. 56: 296 –301.
57. Milner, J. A. (2000) Functional foods: the US perspective. Am. J. Clin.
Nutr. 71: 1654S–1659S.
58. Ip, C. (1986) Interaction of vitamin C and selenium supplementation in
the modification of mammary carcinogenesis in rats. J. Natl. Cancer Inst. 77:
299 –303.
59. Milner, J. A (2001) A historical perspective on garlic and cancer. J.
Nutr. 131: 1027S–1031S.
60. German, J. B., Roberts, M. A., Fay, L. & Watkins, S. M. (2002) Metabo-
lomics and individual metabolic assessment: the next great challenge for nutri-
tion. J. Nutr. 132: 2486 –2487.
61. Hall, R., Beale, M., Fiehn, O., Hardy, N., Sumner, L. & Bino, R. (2002)
Plant metabolomics: the missing link in functional genomics strategies. Plant Cell
14: 1437–1440.
62. Reo, N. V. (2002) NMR-based metabolomics. Drug Chem. Toxicol. 25:
375–382.
63. Mendes, P. (2002) Emerging bioinformatics for the metabolome. Brief
Bioinform. 3: 134 –145.
64. Watkins, S. M., Reifsnyder, P. R., Pan, H. J., German, J. B. & Leiter, E. H.
(2002) Lipid metabolome-wide effects of the PPARgamma agonist rosiglitazone.
J. Lipid Res. 43: 1809 –1817.
65. Noguchi, Y., Sakai, R. & Kimura, T. (2003) Metabolomics and its
potential for assessment of adequacy and safety of amino acid intake. J. Nutr.
133 (Suppl 1): 2097S–2100S.
66. Sakamoto, K. (2000) Synergistic effects of thearubigin and genistein
on human prostate tumor cell (PC-3) growth via cell cycle arrest. Cancer Lett. 151:
103–109.
67. Smelzer, A. & Kim, Y. (2003) The effects of bioactive food components
on p53 pathway in cancer prevention. Nutrition Today (in press).