<title>1. INTRODUCTION</title> <p>Since the discovery of the first PPAR by Issemann and
Green in 1990 [##REF##2129546##1##], the role of this fascinating class of nuclear receptors in
normal physiology and pathophysiology has become progressively more important.
The potential biological activities attributed to PPARs have been expanding
ever since they were identified as potential mediators
of the hypolipidemic effect of fibrates in humans and as participants in
peroxisome proliferation and hepatocarcinogenicity in rodents. So far, PPARs
have been implicated in diverse processes including lipid and carbohydrate metabolism,
energy expenditure, immune and inflammatory processes, vascular homeostasis,
tissue remodeling, and cell differentiation and proliferation in normal and
neoplastic tissues [##UREF##0##2##–##REF##16511592##8##].</p> <p>It is
evident, but too often ignored, that important interrelationships must exist among
PPARs, mitochondria, and cancer. Regardless of the precise nature of these
interconnections, PPARs undoubtedly have a significant energetic, plastic, and
signaling role in the pathophysiology of cancer cells purely by virtue of their
central role in lipid metabolism. This role necessarily involves interaction with mitochondria.
Mitochondria are not only the main site of lipid oxidative metabolism, but they
are also the cellular powerhouses that coordinate cellular metabolism and serve
as the origin of important anabolic fluxes and signal transduction pathways [##REF##15304251##9##, ##REF##17475665##10##].</p> <p>Thus, the role of mitochondria in
cancer is under a critical re-evaluation, particularly in light of the
so-called Warburg effect: most cancer cells exhibit increased aerobic
glycolysis, and use this metabolic pathway for generation of ATP as a main
source of their energy supply. Too often this does not justify the complex
metabolic alterations present in different types of neoplasia [##REF##16728625##11##–##REF##16982728##13##].</p> <p>Similarly,
the interrelationships between PPARs and cancer are not entirely clear. Some
studies show that PPARs have antineoplastic and/or cancer differentiating
activities, while others show that they have important carcinogenic properties
[##REF##15192441##6##, ##REF##17308085##14##–##UREF##3##20##].</p> <p>These
reports highlight the multifaceted role of PPARs in neoplastic cells. Hence,
the roles of PPAR in normal physiology and pathophysiology should be clarified,
since this may benefit our understanding of how cancer occurs and how it can be
treated.</p> <p>Discussion
of the interrelationships among PPARs, mitochondria, and cancer should first
involve careful evaluation of some misleading factors that have contributed to
confusion about PPAR biology.</p>
<title>4. CONCLUSION</title> <p>Many molecular mechanisms have been
proposed to explain how PPARs, directly and/or indirectly, may induce cancer
cell cycle arrest and induction or cancer cell differentiation or
dedifferentiation. In spite of this, the molecular interrelationships between
the mechanisms of functional modulation of PPAR and these important cellular
phenotypic changes are still debated. It is clear that the various molecular
modifications observed in different studies (decrease in cyclin D1, inhibition
of IkB, induction of TSC22, NF-kB, GADD153, PTEN, etc.) may depend on the
particular cell and cell functional status and that a potential master signal
should be investigated.</p> <p>Here, we
have briefly described the molecular link between PPARs and cancer from a
mitochondrial point of view. In our opinion, the most important factor linking
cancer to PPARs is represented by their “synthetic ligands,” which are
characterized by other important and debated extrareceptor activities.
Specifically, these agents can induce oxidative stress, which has an ambiguous
role in cancer, leading it to act as a double-edged sword.</p> <p>In this sense, mitochondria plays a critical role as
one of the most important organelles for generating reactive species. The
metabolic stress and energetic failure that result from fibrate- and
thiazolidinedione-induced mitochondrial impairment may also play an important
part in cancer regression, especially in cells that require active and complete
anabolic pathways to sustain cancer growth, an aspect that has not always been
completely or correctly evaluated.</p> <p>Considering their
physicochemical properties discussed above, it is worthy of note that tumor regression induced by PPAR
ligands may be a useful approach for the treatment of
neoplasias of the central nervous system, which are classically difficult to
treat with conventional chemotherapy. Specifically, interesting results have
already been obtained in terms of decrease of cell proliferation, apoptosis induction, and
expression of markers typical of a more differentiated phenotype in
glioblastoma and astrocytoma cell lines [##REF##17949236##84##–##REF##14988421##90##], in primary cultures of human
glioblastoma cells derived from surgical specimens [##UREF##14##91##], and above all, in patients
with high-grade gliomas (glioblastoma or anaplastic glioma) [##REF##16094629##92##].</p> <p>Moreover, given that
pharmacological modulation of PPAR in cancer cells typically arrests the cell
cycle in the G0/G1 phase, combination therapy with a PPAR agonist and an antimitotic antitumor
agent deserves careful consideration.</p> <p>Furthermore,
it may be useful to distinguish between real differentiating agents, which show
low cytotoxicity indices relative to their differentiation activity
(thiazolidinediones, fibrates, retinoids) and spurious differentiating agents,
which show low differentiating activity and high cytotoxicity indices (old and
new HDAC inhibitors) [##REF##14998325##93##, ##REF##16892355##94##].</p> <p>A mitochondrial approach to analysis of the
molecular link between PPARs and cancer certainly adds new levels of complexity
to the already complicated picture. However, an optimal definition of all
molecular mechanisms relating PPARs, mitochondria, and cancer may be
fundamental to our understanding of the real therapeutic index of
pharmacological modulation of these nuclear receptors. This is important not
only in cancer, but also in the other diseases in which PPARs play a
significant role, including atherosclerosis, hyperlipoproteinemias, metabolic
syndrome, diabetes mellitus, and obesity. Moreover, a complete understanding of
the pharmacotoxicological profile of these agents may reduce the incidence of
dangerous side effects that have already dramatically afflicted patients
treated with PPAR ligands [##REF##15039476##37##–##REF##17848652##40##].</p>
<p>Recommended by Dipak Panigrahy</p> <p>Before the discovery of peroxisome proliferator activated receptors (PPARs), it was well known that certain drugs considered as classical PPAR-alpha agonists induced hepatocarcinoma or peroxisome proliferation in rodents. These drugs were derivatives of fibric acid, and they included clofibrate, bezafibrate, and fenofibrate. However, such toxicity has never been observed in human patients treated with these hypolipidemic drugs. Thiazolidinediones are a new class of PPAR activators showing greater specificity for the <italic>γ</italic> isoform of PPARs. These drugs are used as insulin sensitizers in the treatment of type II diabetes. In addition, they have been shown to induce cell differentiation or apoptosis in various experimental models of cancer. PPAR-<italic>α</italic> ligands have also been shown to induce cancer cell differentiation and, paradoxically, PPAR-<italic>γ</italic> drug activators have been reported to act as carcinogens. The confusing picture that emerges from these data is further complicated by the series of intriguing side effects observed following administration of pharmacological PPAR ligands (rhabdomyolysis, liver and heart toxicity, anemia, leucopenia). These side effects cannot be easily explained by simple interactions between the drug and nuclear receptors. Rather, these side effects seem to indicate that the ligands have biological activity independent of the nuclear receptors. Considering the emerging role of mitochondria in cancer and the potential metabolic connections between this organelle and PPAR physiology, characterization of the reciprocal influences is fundamental not only for a better understanding of cancer biology, but also for more defined pharmacotoxicological profiles of drugs that modulate PPARs.</p>
<title>2. PPARs IN PATHOPHYSIOLOGY AND
MISLEADING FACTORS</title> <title>2.1. Synthetic ligands</title> <p>Initially,
the physiological ligands of PPARs were unknown and PPARs were classified as
“orphan receptors.” Their function was studied using synthetic ligands of PPAR-<italic>α</italic>, the first PPAR discovered. These
synthetic ligands were a heterogeneous class of molecules ranging from
trichloroacetic acid to plasticizers such as di-2-ethylhexyl phthalate (DEHP) and mono-2-ethylhexyl phthalate (MEHP) [##REF##2129546##1##]. The
structural heterogeneity of the ligands seems to reflect the conformation of
the PPAR ligand binding domain (LBD), which forms a large, Y-shaped hydrophobic
pocket with relatively low ligand specificity [##UREF##0##2##, ##REF##17132851##7##]. For PPAR-<italic>α</italic>, the ligands used most often in
experiments were fibrates such as clofibric acid, bezafibrate, and gemfibrozil.
Studies on fibrate binding to PPAR-<italic>α</italic> showed that these drugs caused a
hypotriglyceridemic effect by inducing transcription of several genes related
to oxidative metabolism of lipids, which occurs primarily in the mitochondria
[##UREF##0##2##–##REF##12746275##4##]. However, some discrepancies soon emerged. In fact, previous PPAR
research failed to provide thorough explanations of the drugs' other important
biological activities, such as peroxisome proliferation and
hepatocarcinogenicity in rodents or, most importantly, their side effects.
These side effects include angina crisis; elevation of serum aminotransferase,
which indicates liver damage; increases in serum creatine phosphokinase
concentrations, which can initiate myositis, myopathy and, in rare cases,
rhabdomyolysis; increases in serum creatinine concentration; and acute renal
failure in rare cases [##UREF##4##21##].</p> <p>Some of these biological activities had already led
investigators to propose that these ligands had effects independent of their
binding to PPAR. If true, neglecting these “extrareceptor” functions may
interfere with our understanding of PPAR pathophysiology. For example, studies
in the 1980s established that fibrates (clofibric acid, bezafibrate, and
gemfibrozil) can pass freely through red blood cell membranes and bind to human
hemoglobin at the level of the hydrophobic pocket of the alpha chain
interfaces. This binding lowers the oxygen affinity of hemoprotein more strongly
than does the natural allosteric effector 2,3-biphosphoglycerate [##UREF##5##22##], which
could, for example, lead to an angina crisis (by perturbation of
microcirculation in ischemic areas caused by abrupt changes in the level of
oxygen released from the blood) at the usual therapeutic plasmatic drug
concentrations [##REF##7756095##23##]. Around the same time, other researchers showed that the
agents acting as peroxisome proliferators hampered mitochondrial respiration,
with potentially significant clinical implications [##UREF##6##24##–##REF##10330687##26##]. Indeed, treatment
with fibrates was found to lead to some histological and biochemical features
characteristic of hepatic, muscular, and renal toxicities. This result led to the hypothesis that
disruption of the mitochondrial electron respiratory chain in conjunction with
other genetic or acquired predisposing factors may contribute to these toxic
effects independently of PPAR activation. Moreover, molecular analysis of the
interactions between human hemoglobin and fibrates indicated that particular
physicochemical aspects of the drug molecules, specifically their carboxylic
group and their significant hydrophobicity, might be responsible for their
biological activity. Interestingly, these physicochemical characteristics of
fibrates fit well with the milieu of mitochondria in general, namely the
difference in pH between the more alkaline matrix and the more acidic
intermembrane space, as well as with structural features of complex I (NADH
cytochrome c reductase) in the mitochondrial electron respiratory chain, which
is a large, hydrophobic protein component in the mitochondria [##REF##17475665##10##, ##REF##11870261##27##].</p> <p>All of these considerations led to studies that
examined how mitochondria were affected by fibrate administration, and whether
these effects might have clinical implications [##REF##14615970##18##, ##UREF##7##28##]. The results clearly
showed that fibrates could disrupt the mitochondrial electron respiratory chain
at the level of NADH cytochrome c reductase [##REF##14615970##18##]. This effect was even more
pronounced for ciglitazone, which was one of the first thiazolidinediones to be
synthesized. Thiazolidinediones are a class of molecules that are chemically
related to fibrates (##FIG##0##Figure 1##). Drug-induced mitochondrial dysfunction causes a
series of compensatory metabolic mechanisms, which, in addition to PPAR agonist
activity, may be partially responsible for some of the pharmacological and
toxicological properties of this class of molecules. In fact, the resulting shut-down of
mitochondrial NADH oxidation drives cells to change their oxidative metabolism
in a way that is strictly correlated to the degree of complex I inhibition. Specifically,
upon treatment with fibrates, which are less potent inhibitors of complex I,
cells tend to use those components of the electron respiratory chain that
remain efficient (e.g., complex II). This leads the cell to use FADH<sub>2</sub> oxidation to obtain energy. In other words, compensatory mechanisms come into
play, which are probably sustained by glycerol catabolism viamitochondrial FAD-dependent glycerol-3-phosphate dehydrogenase or
by fatty acid <italic>β</italic>-oxidation viaelectron-transferring
flavoprotein (ETF). These changes have a significant hypotriglyceridemic effect
and a slight hypoglycemic effect. In contrast, using more potent complex I
inhibitors (i.e., thiazodilinediones) greatly reduces NADH dehydrogenase
activity, thus reducing the use of <italic>β</italic>-oxidation and increasing reliance on
glycolysis, resulting in a stronger hypoglycemic effect and a much weaker or
null hypotriglyceridemic effect (##FIG##1##Figure 2##) [##REF##14615970##18##].</p> <p>Based on
these findings, PPAR-<italic>α</italic> activation may be due, at least in part, to a shift in
the metabolic state: preferential use of lipids through glycerol catabolism via mitochondrial FAD-dependent
glycerol-3-phosphate dehydrogenase and fatty acid <italic>β</italic>-oxidation via ETF. In this way, as suggested by Kersten et al. [##REF##10839530##3##], fatty
acids would stimulate their own metabolism. Interestingly, this mechanism of
switching to lipid metabolism in response to fibrate-induced complex I
inhibition may explain the reported activation of genes in the cytochrome P450
IV family. This family of proteins is responsible for microsomal <italic>ω</italic>-oxidation of
long chain and very long chain fatty acids [##UREF##8##29##]. This pathogenic mechanism may
also provide a better explanation of the peroxisomal proliferation observed in
rodents given synthetic PPAR-<italic>α</italic> ligands. In fact, the lipid component of these
animals' diet contains a particularly high proportion of polyunsaturated fatty
acids [##UREF##9##30##]. Hence peroxisomal <italic>β</italic>-oxidation, which is normally more active in
rodents than in humans or other primates, may be further enhanced by inhibition
of mitochondrial NADH oxidation.</p> <p>Moreover,
the increase in free radical oxygen species resulting from stimulated
peroxisomal <italic>β</italic>-oxidation may
further increase the oxidative stress that results from complex I inhibition
[##REF##11864782##31##, ##REF##17056127##32##] and thereby contributes significantly to the observed carcinogenic
properties of PPAR ligands in rodents, particularly in the liver.</p> <p>From a
clinical point of view, thiazolidinediones, which are stronger inhibitors of
NADH cytochrome c reductase [##REF##14615970##18##, ##REF##15184076##33##–##REF##17499714##36##] than fibrates, strongly disrupt NADH
oxidation such that they may prevent the induction of <italic>β</italic>-oxidation, which could
otherwise serve as a compensatory energy source. This renders metabolism almost
exclusively dependent on glycolysis. Interestingly, damage to the pathways of
energy production, particularly in organs that are rich in mitochondria (e.g.,
liver, muscle, heart, and kidney), may explain (i) the prevalent hypoglycemic
activity of <italic>γ</italic>-ligands, despite minor or absent hypolipidemic effects, (ii) the
weight gain due mainly to water retention typically observed in patients
treated with PPAR-<italic>γ</italic> ligands, (iii) the differentiation of adipose cells, and
(iv) the dramatic cardiac
and hepatic toxicities often observed following administration of
thiazolidinediones [##REF##15039476##37##–##REF##17848652##40##].</p> <p>Interestingly, similar mitochondrial impairment by PPAR agonists was observed by Nadanaciva et al. [##REF##17658574##41##], who used a phosphorescent oxygen-sensitive probe and an immunocapture technique to evaluate the mitochondrial respiration and the activity of individual oxidative phosphorylation complexes on isolated rat liver mitochondria. By this dual approach, the authors were also able to obtain a rank order of the mitochondrial toxicity of thiazolidinediones, fibrates, and statins. These results could be important as they suggest the possibility of a screening strategy to evaluate potential mitochondrial toxicity, reducing in such a way the incidence of clinical side effects.</p> <p>Moreover, a novel mitochondrial target
protein has recently been identified for the thiazolidinediones [##REF##14570702##42##]. This
protein, called mitoNEET, is an iron-containing outer mitochondrial membrane
protein that seems to play a role in regulating mitochondrial oxidative
processes. Recently, Wiley et al. [##REF##17376863##43##, ##REF##17766440##44##] showed that cardiac mitochondria isolated
from mitoNEET-null mice demonstrated a reduced oxidative capacity, confirming
that mitoNEET is a protein involved in the control of maximal mitochondrial
respiratory rates.</p> <p>These results underline the importance of carefully defining
the direct interrelationships between pharmacological PPAR ligands and
mitochondria, also to better clarify the physiology and pathophysiology of
PPARs.</p> <p>In
conclusion, many PPAR ligands possess extrareceptor biological activities that
can complicate interpretation of the results of experiments investigating the
pathophysiology of PPARs. This caveat has serious consequences, not only for
our ability to correctly understand the metabolic roles of PPARs, but also for
our ability to determine their roles in the (de)differentiation of cancer
cells.</p> <title>2.2. Metabolic and genetic studies of PPARs in rodents</title> <p>Most of the
available data on the pathophysiology of PPARs has been obtained from metabolic
studies in rodents. However, species-specific differences in metabolism and
diet can be an obstacle when applying the results of animal studies to human
patients [##REF##15545227##35##, ##UREF##10##45##].</p> <p>Another
source of data that may be misleading is genetic studies on knock-out rodents
for PPARs or their transcriptional coactivators (i.e., PGC-1<italic>α</italic>). Conclusions
drawn from these studies about the metabolic roles of the different PPARs
neglect the interaction between PPARs and their coactivators in mitochondrial
biogenesis in general, and in mitochondrial lipid metabolism in particular [##UREF##1##5##, ##REF##10412986##46##–##REF##15760270##49##].</p> <title>3. PPAR AND CANCER: WHAT IS THE ROLE
FOR MITOCHONDRIA?</title> <p>Various
molecular links between PPARs and cancer have been considered in other
articles. Here, we highlight some intriguing observations related to PPARs and
cancer that seem to indicate a role for mitochondria in this disease of cell
proliferation and differentiation. Since the aim of this review is to discuss
the potential molecular link between PPARs and cancer from a mitochondrial
point of view, we focus on the particular “extrareceptor” interrelationships
between PPARs and fibrates or their thiazolidinedione derivatives.</p> <title>3.1. PPARs, mitochondria, and carcinogenesis</title> <p>PPARs and their pharmacological ligands were
originally considered to be “oncopromoters.” Specifically, PPAR ligands were
considered to be nongenotoxic carcinogens. It is well known that these
molecules can induce hepatocarcinoma in rodents; however, their administration
first provokes hepatomegalia and induces expression of a series of antioxidant
enzymes, such as catalase, superoxide dismutase, and glutathione peroxidase. In
this way, these molecules may create an imbalance in oxidative metabolism in
general and oxidative stress in particular [##REF##7562957##50##, ##REF##11248139##51##]. The ability of PPAR
ligands to induce oxidative stress has since been confirmed for
thiazolidinediones as well [##REF##11328671##52##–##REF##14623026##54##]. Interestingly, reactive oxygen species
(ROS) and cellular oxidative stress have long been implicated in
carcinogenesis, despite the fact that the precise pathogenic molecular
mechanisms are complex, debated, and at times paradoxical [##REF##16959608##55##]. More specifically,
the following hold.</p> <p>
<list list-type="roman-lower"><list-item><p>In
normal cells, mutations in nuclear or mitochondrial genes encoding
components of the mitochondrial electron transport chain (ETC) or xenobiotics
capable of disrupting the mitochondrial electron flux can lead to an increase
in the generation of ROS, particularly superoxide. This radical is rapidly
dismuted by superoxide dismutase to yield hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>),
which can diffuse to the nucleus and attack DNA before cellular antioxidant
defenses adjust to the new level of oxidative stress. This oxidative damage may
contribute to genetic instability in congenital and/or acquired predisposed
subjects [##REF##18032788##56##, ##REF##17307400##57##].</p></list-item><list-item><p>Cancer cells generally generate more ROS than normal cells. This
difference may relate to the greater number of metabolic and proliferative
activities that often occur in a transformed cell, or to a qualitative or
quantitative imbalance between cellular antioxidant defenses and the oxidative
environment [##REF##16959608##55##, ##REF##17079457##58##–##REF##18388260##60##].</p></list-item><list-item><p>In cancer cells, the levels of
expression of some components of the antioxidant system are amplified
independently by drug treatment (e.g., thioredoxin, DJ-1 protein,
peptidyl-prolyl cis-trans isomerase A, cyclophilin A, protein disulfide
isomerase A3, ERP 60/GRP58) [##REF##16959608##55##, ##REF##21136693##61##]. Interestingly, the increase in
thioredoxin activity in cells with elevated oxidative stress may relate to its
essential role in facilitating transcription in an environment where increased
oxidative stress signaling in the cytosol is required for stimulating cell
proliferation. Furthermore, drug-induced cancer cell differentiation typically
reduces the expression of these antioxidant proteins [##REF##21136827##62##].</p></list-item><list-item><p>Numerous studies implicate
increased oxidative stress in the cell death induced by diverse
chemotherapeutic agents. Anthracycline derivatives, newer redox cycling agents,
and, more recently, histone deacetylase inhibitors and proteasome inhibitors
all appear to increase oxidative stress in cells. Although the mechanism
responsible for the increase has not been established, mitochondria are fundamental
in ROS generation and seem to be involved, either directly or indirectly [##REF##16959608##55##, ##REF##17593552##63##–##REF##16146732##65##].</p></list-item></list> This
puzzling picture suggests that PPAR-related rodent hepatocarcinogenesis depends
on strong stimulation of ROS generation, mainly in mitochondria dysregulated by
the PPAR ligand in question. Moreover, this oxidative stress may be reinforced
by the specific membrane composition and the abundant H<sub>2</sub>O<sub>2</sub> production from peroxisomal lipid metabolism in rodents, as already discussed.</p> <p>Could
a similar pathogenic mechanism have a role in human carcinogenesis?</p> <p>A
direct role for PPARs in carcinogenesis is hardly credible, considering their
fundamental physiological role in cell metabolism. The altered expression of
different PPAR isoforms observed in some neoplasias may be the result of
secondary metabolic changes in transformed cells relative to normal cells [##REF##15192441##6##, ##REF##17443682##66##].</p> <p>On
the other hand, the question of whether the synthetic PPAR ligands play a role
in human carcinogenesis is still open and intriguing. In fact, one of the first
large clinical studies on gemfibrozil, a classic PPAR-<italic>α</italic> ligand, [##REF##3313041##67##] showed a small but significant increase (<italic>P</italic> = .032 by the Fisher
exact test) in the incidence of basal cell carcinoma in patients taking
gemfibrozil; this finding, unfortunately has been largely ignored by
investigators. Moreover, in an
intermediate follow-up study
[##REF##8283157##68##], cancer occurred at equal rates in both the
untreated group and the group treated with gemfibrozil, but the cancer in the
latter led more often to mortality, primarily during the last 1.5 years of
follow-up. To be sure, results recently obtained from an 18-year mortality
follow-up of this study do not seem to confirm this increase in cancer
mortality, but the follow-up design failed to address certain possible
interpretations and also in the Autors opinion of this cited study some of the
follow-up data can be misleading [##REF##16606810##69##].</p> <p>Nevertheless, the data obtained in the
original study are intriguing considering the peculiar molecular epidemiology
and pathogenesis of basal cell carcinoma and the relatively short (5-year)
period of drug exposure used in the study. These findings, together with the
demonstrated ability of fibrates and thiazolidinediones to alter mitochondrial
oxidative metabolism and induce ROS generation, indicate that care should be
taken when this class of drugs is used in the treatment of nutrition-sensitive
tumors [##REF##11121836##70##, ##REF##12489852##71##].</p> <title>3.2. PPARs, mitochondria, and inhibition of tumor
growth</title> <p>It is well established that activation of PPARs (<italic>α</italic>,
<italic>β</italic>/<italic>δ</italic>, and <italic>γ</italic>) by natural or synthetic agonists can inhibit growth and induce
differentiation or death of tumor cells. The original observation was of PPAR-<italic>γ</italic>
ligands and liposarcoma, consistent with the physiological function of PPAR-<italic>γ</italic>
[##REF##10839530##3##, ##REF##14708026##72##, ##REF##10097144##73##]. Subsequently, PPAR-<italic>γ</italic> and PPAR-<italic>α</italic> ligands were shown to promote the
differentiation of various tumor cell lines, including breast, lung, prostate, leukemia, colon, melanoma, and liver. This
differentiation was often independent of the relative expression levels of the
different PPAR isoforms [##REF##15192441##6##, ##REF##10467352##15##, ##REF##11507074##16##, ##REF##17443682##66##, ##REF##11121836##70##, ##REF##14708026##72##]. These studies also suggested
extrareceptor activities of fibrates and thiazolidinediones as the basis of
their ability to induce cancer cell differentiation [##REF##10467352##15##–##REF##14615970##18##, ##REF##14708026##72##]. Moreover, a
recent study by Panigrahy et al.
[##REF##18199835##74##, ##REF##17327920##75##] on endothelial and mesenchymal tumor cells and mice showed that PPAR-<italic>α</italic>
ligands such as fenofibrate directly suppress tumor growth through
receptor-dependent and -independent pathways, and that they indirectly suppress
tumor growth by inhibiting angiogenesis and the inflammatory response in the
microenvironment of the tumor. Therefore, the noncancerous host tissue could be
an important target for cancer treatment with pharmacological PPAR ligands.</p> <p>These data illustrate the extreme complexity of the
interrelationships among PPARs, mitochondria, and cancer. Nevertheless, the
most important aspects of these interrelationships are the activities of the
synthetic PPAR ligands, particularly their extrareceptor activities.
Mitochondria are becoming increasingly important as targets for these
drug-induced extrareceptor activities, as discussed in recent reviews [##REF##15545227##35##, ##REF##15925327##76##, ##REF##16728570##77##]. To better understand the interactions among PPAR ligands, mitochondria,
and cancer, it may be useful to describe our work, which parallels that of
other groups. Curiously, the differentiating activity of fibrates was
originally hypothesized in binding studies of fibrates and hemoglobin. The
physicochemical properties of fibrates and their toxicological profile allow
them to interact with some hydrophobic components of the mitochondrial electron
respiratory chain. The resulting oxidative metabolic stress may induce
differentiation of cancer cells, similar to the effects of heat shock [##REF##10467352##15##].
Importantly, this effect does not depend on PPAR agonism, but it is related to
the physicochemical properties (pKa, log P, log D, water solubility, and pH
profile) of the molecules. These properties should favor permeation,
accumulation, and interaction with components of the internal mitochondrial
membrane [##REF##17475665##10##].</p> <p>For
example, therapeutic doses of bezafibrate inhibited proliferation of human
leukemia cell lines HL-60, U-937, and K-562 in a dose-dependent manner. In
HL-60 cells, growth inhibition was associated with an increased number of cells
in the G0/G1 phase and a significant decrease in the number of cells in the
G2/M phase. Analysis of cell differentiation markers (CD) showed a
dose-dependent increase in expression of CD11b and CD14 in HL-60 cells and of
CD14 in U-937 cells. Functional assays confirmed that the phenotypes of these
cells were more mature. Both HL-60 and U-937 cells showed a dose-dependent
restoration of the respiratory burst stimulated by PMA and zymosan. K-562
erythroleukemia cells showed a dose-dependent increase in hemoglobin synthesis.
Similar cellular differentiation was observed following treatment with two
other fibrate derivatives, clofibric acid and gemfibrozil. Interestingly,
fibrate-induced differentiation was partially inhibited by antioxidants
including acetylcysteine (NAC), and electron microscopy revealed that
fibrate-treated cells had mitochondrial damage [##REF##10467352##15##]. Functional evaluation of
this drug-induced mitochondriopathy showed that fibrates and ciglitazone
specifically inhibited NADH cytochrome c reductase activity in a dose-dependent
manner in HL-60, TE-671 human rabdomyosarcoma, and Hep-G2 human hepatocarcinoma
cell lines, whereas the activity of other mitochondrial respiratory chain
enzymes remained unchanged [##REF##14615970##18##, ##REF##15184076##33##, ##REF##17499714##36##]. The impairment of NADH oxidation induced a cellular metabolic
shift towards anaerobic glycolysis and/or <italic>β</italic>-oxidation, as shown by the dose-dependent
increases of certain metabolites (lactate, alanine, glycolytic, and
nonglycolytic derived acetate) [##REF##14615970##18##].</p> <p>A fundamental
observation from this research was the correlation of mitochondrial
dysfunction, metabolic shift, and differentiation activity in tumor cells
treated with increasing concentrations of PPAR ligands. Furthermore, quantitative comparison
on a molar ratio basis between these PPAR ligands (bezafibrate, clofibric acid,
gemfibrozil, and ciglitazone) for inhibition of NADH cytochrome c reductase
activity, metabolic adaptations, differentiation potency, and antiproliferative
index confirmed a strict correlation between these parameters [##REF##14615970##18##]. These results suggested that
inhibition of mitochondrial NADH dehydrogenase could contribute to both the
pharmacological and toxicological profiles of fibrate derivatives (strong
hypolipidemic/weak hypoglycemic effect, liver and muscle toxicity) and
thiazolidinediones (hypoglycemic/insulin sensitizer effect, liver and heart
toxicity) [##REF##12475986##17##, ##REF##14615970##18##, ##REF##9493760##25##, ##REF##10330687##26##, ##REF##15184076##33##–##REF##17848652##40##].</p> <p>In terms of
mitochondrial oncology, these data suggest a possible molecular mechanism for
the peroxisome proliferator activity and carcinogenicity of fibrates typically
observed in rodents. These data also indicate a strict correlation among
fibrate- and thiazolidinedione-induced cellular respiration dysfunction,
stimulation of glycolysis, and cancer cell differentiation that strongly
implicates mitochondria and oxidative metabolism in the pathophysiology of
cancer.</p> <p>Importantly,
these results confirm and extend the results of other studies focusing on
nongenomic activities of fibrates and thiazolidinediones [##REF##11507074##16##, ##REF##12475986##17##, ##UREF##13##78##–##REF##17015690##81##].
Furthermore, these observations explain some contradictory data related to the
role of PPARs in cancer cell differentiation [##REF##10839530##3##, ##REF##15192441##6##, ##REF##14708026##72##, ##REF##16728570##77##]. Above all, the
intriguing data concerning the induction of differentiation associated with a
shift towards aerobic glycolysis (a paradoxical Warburg
effect) confirms the need to reconsider cancer cell metabolism in general
and the Warburg effect in particular [##REF##16728625##11##, ##REF##16982728##13##, ##REF##16766262##82##]. To that end, our
understanding of the role of PPARs in cancer should assume a new level of
complexity that takes into account their fundamental functions in lipid
metabolism, in inflammation and, directly or indirectly, in angiogenesis [##REF##18199835##74##, ##REF##17327920##75##].</p> <p>The
molecular link among the synthetic PPAR ligands, mitochondria, and cancer
indicates the need for a careful evaluation of some aspects of cancer cell
pathophysiology, such as the following.</p> <p>
<list list-type="roman-lower"><list-item><p>The possible existence of a
transduction pathway master signal as the basis of the complex cellular
differentiation program related to PPAR. ROS, nitric oxide (NO), and reactive
nitric oxide species (RNS) should form an important branch of this program. In
addition, there should be a role for the NADH/NAD+ ratio.</p></list-item><list-item><p>The role of some
oncogenes/oncosuppressors in cancer pathogenesis, given that mitochondrial
respiratory chain dysfunction can induce a more differentiated phenotype in
tumor cells and thereby influence their activity [##REF##10467352##15##–##REF##14615970##18##].</p></list-item><list-item><p>The significance of the modulation
of the expression of proteins with oncogenic and antioxidant functions
(stathmin 1, DJ-1 protein, peroxiredoxin 2, nucleoside diphosphate kinase A, etc.) in
PPAR-related cancer cell differentiation. This is an important topic given the
potential pathophysiological role of PPAR in cancer [##REF##21136827##62##, ##REF##15963265##83##].</p></list-item></list>At last, an
understanding of the molecular mechanisms involved in the interrelationships
between mitochondrial respiration and PPAR-related cancer regression may have
important clinical implications for cancer diagnosis, prognosis, and therapy.</p>
<fig id="fig1" position="float"><label>Figure 1</label><caption><p>Structure of some drugs acting as PPAR ligands.</p></caption></fig> <fig id="fig2" position="float"><label>Figure 2</label><caption><p>Flow chart of possible molecular mechanisms caused by a mitochondrial
dysfunction induced by fibrates (PPAR-<italic>α</italic> ligands) and thiazolidinediones (PPAR-<italic>γ</italic>
ligands).</p></caption></fig>
<graphic xlink:href="PPAR2008-256251.001"/> <graphic xlink:href="PPAR2008-256251.002"/>
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PPAR Res. 2008 Jul 14; 2008:256251