Departments of Cell Biology and Medicine, Vanderbilt University Medical Center and Veterans Affairs Medical Center, Nashville, Tennessee 37232
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ABSTRACT |
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Colorectal cancer (CRC) represents
a significant cause of morbidity and mortality worldwide. Recently,
ligands for the nuclear hormone receptor peroxisome
proliferator-activated receptor (PPAR
) have exhibited promise in
the treatment of CRC. For example, activation of PPAR
reduces the
proliferation of cultured CRC cells grown in vitro or in vivo using the
nude mouse xenograft model of tumor growth. Furthermore, agonists of
the receptor also reduce the development of preneoplastic lesions in a
model of carcinogen-induced CRC in rats. However, ligands for the
receptor paradoxically enhance intestinal adenoma formation in another murine model of intestinal polyposis, the APCMin
mice. These disparate results may be due to the inherent limitations of
the APCMin mouse as a model for humans with CRC.
Finally, genetic studies identifying loss of function mutations of
PPAR
in human CRC specimens strongly suggest a tumor suppressive
role for the receptor during the development of CRC.
nuclear hormone receptor; intestinal epithelial cell differentiation
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ARTICLE |
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COLORECTAL CANCER (CRC) represents a
significant cause of morbidity and mortality worldwide. Despite its
prevalence, current therapies for the disease remain unsatisfactory and
largely ineffectual, particularly in patients with advanced disease.
Recently, ligands for the nuclear hormone receptor peroxisome
proliferator-activated receptor (PPAR
) have proven effective in
preclincal models of CRC. These findings have direct clinical
relevance, because synthetic activators of PPAR
are currently being
used for the treatment of non-insulin-dependent diabetes mellitus and
do not appear to be associated with significant toxicity. However, the excitement of these findings is tempered by the fact that ligands for
PPAR
are reported to enhance intestinal adenoma formation in a mouse
model of intestinal polyposis. Thus recent studies on PPAR
in the
colon have produced a controversial question that has yet to be
completely answered: does activation of PPAR
prevent or enhance CRC growth?
PPAR forms functional heterodimers with members of the retinoid X
receptor family of nuclear receptors (for review, see Ref. 27). Putative endogenous ligands for the receptor include
both polyunsaturated fatty acids and the eicosanoids
15-deoxy
12,14-PGJ2 , 13-hydroxyoctadecadienoic acid, and 15-hydroxyeicosatetraenoic acid,
but their respective roles in PPAR
signaling in vivo remain unclear.
High-affinity synthetic ligands that selectively activate PPAR
include the thiazolidinedione (TZD) family of drugs. Early studies on
PPAR
established its role as a dominant regulator of adipocyte
differentiation (for review, see Ref. 15). Part of the
cellular response of preadipogenic cell lines to PPAR
activators
includes growth arrest characterized by cell cycle withdrawal
(1). Not surprisingly, activation of PPAR
was also shown to inhibit the growth of liposarcoma cell lines in vitro (25), a result that has now been confirmed in clinical
trials in patients with the disease (6). Because many cell
types express at least low levels of PPAR
, the ability of the
receptor to regulate differentiation and cell growth pathways in
nonadipogenic cell lineages has been examined. Activating ligands of
PPAR
have been found to inhibit the growth of virtually all cell
types tested, including epithelial cells derived from the breast
(12), prostate (13), stomach
(23), and lung (4).
In the colon, levels of PPAR mRNA are nearly equivalent to that
found in adipocytes (7), with the highest levels of
receptor expression observed in the postmitotic, differentiated
epithelial cells facing the lumen (11). Consistent with
this expression pattern, exposure of cultured human CRC cells to
PPAR
agonists induces growth inhibition associated with a delay in
the G1 phase of the cell cycle and an increase in several
markers of cellular differentiation (3, 8, 17). However,
whether these antineoplastic, prodifferentiation effects of PPAR
ligands in the colon operate in vivo is not clear. Three animal models
that have been used to test this hypothesis, including adenomatous
polyposis coliMin [(APC)Min] mice,
azoxymethane (AOM)-treated rats, and nude mice with tumor cell
xenografts. The antineoplastic effects of PPAR
have been confirmed
in the latter two models. Rats treated with the chemical carcinogen AOM
develop preneoplastic colonic lesions termed aberrant crypt foci (ACF),
which later progress into carcinomas. Tanaka et al. (24)
demonstrated that administration of the PPAR
ligand troglitazone
significantly reduces the number of ACF lesions. Importantly, it will
be necessary to carry out these studies further to determine whether
these effects are extended to a reduction in the number of
adenocarcinomas in the colon. Additionally, administration of
troglitazone also significantly reduces the tumor volume of a CRC cell
xenografted onto the flanks of athymic mice (17).
In contrast to the above findings, PPAR ligands have been reported
to enhance polyp number in the APCMin mouse model of CRC.
These mice harbor a nonsense mutation in the tumor suppressor gene
APC (20); loss of function mutations in this
gene are responsible for the hereditary polyposis syndrome familial
adenomatous polyposis (FAP) and are also thought to be one of the major
genetic initiating events for a large percentage of sporadic CRCs
(14). Two different groups have reported that PPAR
ligands slightly increase colon polyps in APCMin mice
(increasing on average from a mean of 1 colonic polyp in control mice
to 3 colonic polyps in PPAR
ligand-treated mice) (10,
16). No effects on either polyp number or size were seen in the
small intestine. It has been proposed that the increase in colonic
polyps seen with PPAR
ligand treatment is consistent with the
increase in polyp burden observed when these mice are placed on a
high-fat diet. Because fatty acids can potentially act as endogenous
activators of PPAR
, the receptor may serve as a molecular link
between dietary fat and colorectal carcinogenesis. However,
there are important differences between these two types of treatment,
because, unlike the case with PPAR
ligand treatment, APCMin mice placed on a high-fat diet have significant
increases in polyp number in both the small and large intestines
(26). It should be emphasized that PPAR
ligands did not
induce polyp formation in wild-type mice, implying the potential need
for a predisposed genetic susceptibility in order for PPAR
ligands
to induce this effect. Additionally, extensive carcinogenic testing of
several TZD derivatives that are potent PPAR
ligands have not
demonstrated any protumorigenic effects of these compounds (T. M. Willson, personal communication). Finally, both of these studies relied exclusively on the TZD class of PPAR
agonists, and the results observed could be due to effects of TZD compounds unrelated to PPAR
agonism. Novel and potent non-TZD PPAR
agonists have been developed,
and one of these, GW7845, was shown to be a potent suppressor of
tumorigenesis in a rodent model of carcinogen-induced breast carcinoma
(21). It will be important to determine the effects on
polyposis in the APCMin mice of such non-TZD PPAR
ligands.
How can these results be reconciled with the antineoplastic effects of
the same compounds observed both in cultured CRC cells and other
preclinical models of tumor growth? And, just how relevant are these
findings to human disease? One explanation may be that treatment in
these particular mice with an agent that accelerates colon epithelial
cell differentiation results in abnormal and excessive proliferation of
the intestinal stem cell compartment; such a chronic stimulus may
paradoxically result in the selection of stem cells with loss of normal
growth control mechanisms. Additionally, in the APCMin
mice, polyp formation is much more pronounced in the small intestine (ranging from 70-100 polyps/mouse) compared with the colon
(~0-2 polyps/mouse). Thus one problematic issue is that PPAR
agonists only affected polyp formation in the colon, where the polyp
numbers are so small that limited statistical confidence can be
achieved. Finally, the APCMin model is clearly imperfect,
because in humans with FAP, the majority of intestinal polyps is found
in the large intestine. Furthermore, the adenomatous polyps in these
animals rarely progress into invasive carcinomas. Recently, Shibata et
al. (19) created a novel mouse model of FAP using a
conditional gene-targeting strategy that specifically deleted exon 14 in both alleles of APC in the colorectal epithelium, resulting in the
rapid development of colonic, but not small intestinal, adenomas and
adenocarcinomas. It will be of major interest to determine whether
PPAR
activators also enhance intestinal adenoma formation in this
variant model of the APCMin mouse.
It could be argued that the protumorigenic effects of PPAR ligands
are only seen in the background of genetic mutations in the
APC gene. Supporting this notion, CRC lesions in the AOM rat normally contain activating mutations in the
-catenin oncogene rather than loss-of-function mutations in APC (22). On the
other hand, PPAR
agonists do inhibit the growth in vitro of a broad spectrum of CRC cell lines, many of which harbor loss-of-function mutations in APC. However, these cell lines are derived from invasive adenocarcinomas that have evolved genetic and epigenetic perturbations in multiple pathways and thus represent imperfect models of early intestinal adenomas. In this regard, cell lines derived from a hybrid
of the Immorto and APCMin mice have previously been
isolated (5), and it would be of interest to determine
whether in these cells PPAR
activation also results in growth
inhibition and differentiation. Finally, because all of these studies
rely on compounds that may have biochemical targets independent of
PPAR
, it will also be of major interest to determine whether any
results seen with PPAR
ligands can be replicated in mice genetically
deficient in PPAR
[either PPAR
/+ mice, because homozygous
null PPAR
are embryonic lethal (2), or in mice with a
colon-targeted genetic ablation of PPAR
].
Recent studies on the genetic status of the PPAR locus in CRCs
appear to support the hypothesis that PPAR
has a tumor suppressive, rather than enhancing, role in the colon. Sarraf et al.
(18) reported that 8% of primary colorectal tumors
contained a loss-of-function point mutation in one allele of the
PPAR
gene. Four unique mutations in PPAR
were identified in the
study; one resulted in a truncated protein that lacked the entire
ligand binding domain, whereas the other three mutations caused defects
in the binding of either synthetic or natural ligands. In addition, a
chromosomal translocation was recently identified in a subset of human
thyroid follicular carcinomas that produces a fusion protein between
PAX8 and PPAR
; this oncoprotein is thought to promote thyroid
carcinoma formation in part by acting as a dominant-negative inhibitor
of wild-type PPAR
(9). Collectively, these findings
argue that PPAR
may be one of many factors that prevent the abnormal
growth and differentiation associated with malignancy.
In summary, cumulative evidence suggests that, as is the case with many
other cell types, activation of PPAR reduces the malignant potential
of CRC cells. Nevertheless, it remains to be determined whether the
protumorigenic effects of PPAR
activation seen in the
APCMin mice are simply an idiosyncratic result of one
particular model system or rather is a finding that can be generally
applied to humans. The controversy will likely be settled by the
results of ongoing experiments examining the biological role of PPAR
in the colon and in CRC as well as studies evaluating the incidence of
CRC in the cohort of humans chronically taking synthetic PPAR
ligands for diabetes.
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ACKNOWLEDGEMENTS |
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We thank the T. J. Martell Foundation and the National Colorectal Cancer Research Alliance for generous support.
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FOOTNOTES |
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This work was supported, in part, by the National Institutes of Health Grants RO1DK-47279 (to R. N. DuBois), P030 ES-00267-29 (to R. N. DuBois), and P01CA-77839 (to R. N. DuBois).
R. N. DuBois is a recipient of a Veteran's Affairs Research Merit Grant and is the Mina C. Wallace Professor of Cancer Prevention.
Address for reprint requests and other correspondence: R. N. DuBois, Dept. of Medicine/GI; MCN C-2104, Vanderbilt University Medical Center, 1161 21st Ave. South, Nashville, TN 37232-2279 (E-mail: raymond.dubois{at}mcmail.vanderbilt.edu).
10.1152/ajpgi.00486.2001
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