Neuroblastoma (NB) is the most common and aggressive pediatric cancer, characterized by a remarkable phenotypic diversity and high malignancy. The heterogeneous clinical behavior, ranging from spontaneous remission to fatal metastatic disease, is attributable to NB biology and genetics. Despite major advances in therapies, NB is still associated with a high morbidity and mortality. Thus, novel diagnostic, prognostic, and therapeutic approaches are required, mainly to improve treatment outcomes of high-risk NB patients. Among neuroepithelial cancers, NB is the most studied tumor as far as PPAR ligands are concerned. PPAR ligands are endowed with antitumoral effects, mainly acting on cancer stem cells, and constitute a possible add-on therapy to antiblastic drugs, in particular for NB with unfavourable prognosis. While discussing clinical background, this review will provide a synopsis of the major studies about PPAR expression in NB, focusing on the potential beneficial effects of hypoglycemic drugs, thiazolidinediones and metformin, to reduce the occurrence of relapses as well as tumor regrowth in NB patients.
NB represents the second most common extracranial malignancy of childhood, accounting for 8 to 10% of all childhood cancers (NB prevalence is about one case in 7,000–10,000 live births) and for approximately 15% of the pediatric deaths for malignant conditions [
The clinical presentation of NB ranges from asymptomatic masses to primary tumors that cause critical illness due to local invasion and/or widely disseminated disease. Most primary NB (65%) usually present in the abdominal region, often in the adrenal medulla. Other common sites of disease include the neck and head (5%), chest (20%), and pelvis (5%) [
NB is a disease of the sympaticoadrenal lineage of the neural crest and originates from neuroblasts in the developing peripheral nervous system [
In recent years, it has been suggested that NB tumorigenesis is dependent on the presence of cancer stem cells (CSCs), which have been also isolated from NB cell lines [
Cellular heterogeneity is a hallmark of NB nodules and the prognosis of these tumors depends on their differentiation levels [
Biedler et al. described three cell subtypes, often discernible also in NB cell line cultures, based on cell morphology, biochemical features, and growth patterns [
Several studies have shown that these cell types derive from a common precursor and are able to bidirectionally differentiate. This bidirectional conversion between well-defined differentiation lineages of the neural crest has been termed “transdifferentiation” [
Because the transdifferentiation process is able to also allow the differentiating of malignant CSCs into benign phenotype, a novel concept in cancer biology was introduced: “induction of differentiation” as possible treatment (e.g., using retinoids to treat NB and acute promyelocytic leukaemia [
The cause of NB development is still unclear occurring mostly as sporadic disease but also rare (about 1% of all cases) familial cases were reported [
Traditional NB treatments include surgery, chemotherapy, radiotherapy, and biotherapy [
Unfortunately, in many cases, by the time of diagnosis, the disease has usually spread already. In these cases, the mainstay treatment is frequently intensive regimens including combinations of high doses of chemotherapeutics [
PPARs are activated by fatty acids, eicosanoids, other dietary lipids, and their metabolites, or synthetic ligands [
There are three PPAR isoforms (
Through the regulation of the expression of multiple genes [
Although all PPAR isoforms display a partially overlapping spectrum of activity, essentially as far as the control of lipid and energy metabolism is concerned, they differ in tissue expression pattern and functional roles [
PPAR-
PPAR-
PPAR-
PPAR-
PPAR-
It regulates energy storage and has a key role in fatty acid metabolism and glucose homeostasis [
TZDs are the best-characterized pharmacological PPAR-
In addition, another oral hypoglycemic drug, metformin, which directly improves insulin action, modulating AMPK activity (a key energy regulator), increases PPAR-
Interestingly, it has been suggested to use PPARs as target for cancer treatment, and several PPAR agonists, in particular acting on PPAR-
All three PPARs isoforms have been identified in NB, although human NB cell lines express PPAR-
In addition, it has been documented that embryonic rat brain and neural stem cells have higher concentration of PPAR-
Interestingly, PPAR-
Several studies have assessed the activity of PPAR-
Preclinical and experimental studies on PPAR agonists in neuroblastoma.
Drug/s | Reference/s | Year | Target | Study types | Cell lines/animal model | Effects |
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15-deoxy-PGJ2 | [ | 2001, 2003, 2004 | PPAR- | | NB cell lines and primary cultures of cortical neurons | Inhibition of growth and apoptosis induction, through PPAR- |
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GW1929 | [ | 2001 | PPAR- | | LA-N-5 | Prodifferentiating effect and inhibition of proliferation. |
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Rosiglitazone and 15-deoxy-PGJ2 | [ | 2004 | PPAR- | | SH-SY5Y, SH-EP1, SK-N-AS, SK-N-FI, LA-N-5, SMS-KCNR, SK-N-DZ, and LA-N-1 | Inhibition of cell growth with different sensitivity related to the cell phenotype. |
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Ciglitazone and 15-deoxy-PGJ2 | [ | 2004 | PPAR- | | SK-N-AS, IMR-32, SK-N-SH, and ND-7 | Overexpression of Rb protein and inhibition of PPAR- |
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Ciglitazone, pioglitazone, troglitazone, and rosiglitazone | [ | 2005 | PPAR- | | Kelly, LA-N-1, LA-N-5, LS, IMR-32, SK-N-SH, and SH-SY5Y | Inhibition of cell proliferation and viability in a dose-dependent manner. |
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Rosiglitazone | [ | 2010 | PPAR- | | SK-N-SH xenograft NB mouse model | Inhibition of tumor growth. |
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Troglitazone | [ | 2002 | PPAR- | | NB-1 cell line | Increase of PPAR- |
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Troglitazone | [ | 2006 | PPAR- | | SHEP NB cell line | Increase of PPAR- |
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Rosiglitazone | [ | 2006, 2007 | PPAR- | | SH-SY5Y cell line | Antiapoptotic effects of rosiglitazone which protected NB cells subjected to MPP+-induced mitochondrial injury reducing ROS production. |
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Rosiglitazone | [ | 2006 | PPAR- | | SK-N-AS and SH-SY5Y cell lines | Inhibition of cell adhesion, invasiveness, and proapoptotic effects. |
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Rosiglitazone | [ | 2010 | PPAR- | | SK-N-AS xenograft NB mouse model | Significant decrease of tumor growth (−70%) as compared to control mice. |
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Rosiglitazone | [ | 2008 | PPAR- | | Rat primary cortical neurons | Induction of cell differentiation, increasing dendritic spine density. |
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Pioglitazone and rosiglitazone | [ | 2011 | PPAR- | Both | Adult male Wistar rats | Induction of proliferation, differentiation, and migration of neural stem cells |
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Pioglitazone | [ | 2009 | PPAR- | | SH-SY5Y cell line | Induction of differentiation and neurite outgrowth, promoting differentiation and outgrowth of cell processes. |
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Rosiglitazone | [ | 2014 | PPAR- | | Mouse NB Neuro 2a (N2A) cell line | Stimulation of neurite outgrowth and significant increase of the population of neurite-bearing cells, via PPAR- |
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Oleic acid or GW0742 | [ | 2007 | PPAR- | | SH-NH-5YSY cell line | Induction of G1 cell cycle arrest, reduction of cell migration and invasiveness, and an increase of neuronal differentiation. |
15-deoxy-Δ12,14-prostaglandin J2 (15-deoxy-PGJ2), a high-affinity natural ligand of PPAR-
In addition, Rodway et al. have found that the inhibition of NB growth induced by 15-deoxy-PGJ2 can be reduced by the presence of serum lysolipids in the culture medium [
Synthetic PPAR-
Han and coworkers firstly evaluated the effect of the synthetic ligand, GW1929, in the NB cell line LA-N-5, and found that this compound induces cell differentiation and inhibits proliferation [
GW1929 prodifferentiating effect was shown to be dependent on PPAR-
In 2005, Valentiner et al. tested the effects of four TZDs (ciglitazone, pioglitazone, troglitazone, and rosiglitazone) in seven NB cell lines (i.e., Kelly, LAN-1, LAN-5, LS, IMR-32, SK-N-SH, and SH-SY5Y) [
The antiproliferative effect of rosiglitazone was confirmed by the same group
Ciglitazone was also used in association with 15-deoxy-PGJ2 to overexpress Rb protein and inhibit PPAR-
Servidei et al. tested 15-deoxy-PGJ2 and rosiglitazone on 8 NB cell lines, with different phenotypes, including N- and S-types [
Many studies have documented that the inhibitory effects of TZDs on neuroblastoma cell growth are partially due to an increase of apoptosis. Indeed, troglitazone induced PPAR-
Proapoptotic effects of rosiglitazone were also reported [
In addition, rosiglitazone induces differentiation, increasing density of dendritic spines in rat primary cortical neurons [
Moreover, in neural stem cells (NSC) from adult mammalian brain, pioglitazone and rosiglitazone directly regulate proliferation, differentiation, and migration [
Accordingly, Miglio et al. described the effects of pioglitazone on SH-SY5Y NB cells, in which this agonist promotes differentiation and outgrowth of cell processes, in a dose-dependent manner [
All these observations are in agreement with previous findings indicating that PPAR-
While PPAR-
In summary, all these results suggest the possible use of PPAR agonists as novel therapy for NB, but to date clinical trials are not yet underway (
Beyond TZDs, metformin is another hypoglycemic drug able to modulate PPAR expression or activity, although these effects are rather cell specific and mainly indirectly mediated by the activation of AMPK.
Metformin is biguanide with a well-known safety profile, mainly used as oral antidiabetic drug [
In particular, metformin seems to selectively affect cancer stem cell survival, inhibiting cancer metastases and thus represents a good potential adjuvant agent for chemotherapy (as reviewed by [
However, the molecular mechanisms of action of metformin are still not completely defined, although it seems that the antiproliferative mechanisms induced by this drug are at least partially diverging from those regulating glucose homeostasis. While the latter is mainly dependent on the AMPK activation, the antitumor activity of metformin is mediated by inhibition of AKT/mTOR (also involving AMPK), the inhibition of TK activity, or the regulation of chloride channels [
As far as the effects of metformin on PPAR activity are concerned, several studies were performed but the results are extremely dependent on the receptor subtype and the cells analyzed. For example, metformin increased PPAR-
However, although demonstrated in several models, the role of the modulation of PPAR expression and/or activity in the antiproliferative effects of metformin in neuroblastoma has not been addressed yet.
The effect of metformin on NB was firstly demonstrated by our groups [
We reported that the effects of metformin treatment in human SKNBE2 and SH-SY5Y NB cell lines are a significant reduction in the proliferation rate and cell viability, due to inhibition of AKT phosphorylation and an increased cell death, via apoptosis-independent pathways. These effects were more pronounced in SKNBE2, which are less differentiated, highly proliferative cells than SH-SY5Y cells. Notably, metformin effects were different depending on the differentiating stimuli, being abolished by retinoic acid, but were potentiated by overexpression of NDM29, a noncoding RNA affecting NB malignancy, although both conditions were characterized by a neuron-like differentiated phenotype [
These
In the same year, Vujic et al. successfully used metformin to inhibit cell proliferation and induce apoptosis in NRAS mutant NB cell lines (SK-N-AS and CHP-212), in which NRAS signaling is constitutively active through the PI3K/AKT/mTOR pathway [
In 2015, Mouhieddine and colleagues found that metformin reduces proliferation rate, viability, and invasive potential of NB cell lines SH-SY5Y [
Interestingly, focusing on metformin effect on stem cell population within a 3D culture model, these authors reported that this drug is able to decrease, but not abolish, cell sphere-forming ability, significantly targeting and reducing cancer stem/progenitor cell population and thus possibly preventing recurrence.
Notably, metformin also reduces MMP-2, a protein involved in extracellular matrix degradation, favoring metastasis and cancer progression [
Thanks to the highly selective and cytotoxic effects of metformin on NB cells and on stem cell population in particular (Table
Studies on metformin treatment to date in neuroblastoma.
Reference/s | Year | Study types | Cell lines/animal model | Effects |
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[ | 2014 | | SKNBE2 and SH-SY5Y cell lines | Significant reduction in the proliferation rate and cell viability, due to inhibition of AKT phosphorylation, and an increased cell death, via apoptosis-independent pathways. |
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[ | 2014 | | SH-SY5Y and SK-N-BE xenograft NB mouse models | Significant inhibition of tumor growth and NB cell viability, interfering with spheroid formation in 3D cultures. Modulation of Rho-GTPases and AMPK activation mediate metformin effects on NB cell survival. |
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[ | 2015 | | SK-N-AS and CHP-212 cell lines | Inhibition of cell proliferation and induction of apoptosis. |
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[ | 2015 | | SH-SY5Y cell line | Reduction of proliferation rate, viability, and invasive potential. |
Despite advances in standard therapeutic protocols, the prognosis of NB has not gained significant progress, especially concerning the rates of metastasis, the incidence of recurrences, and the long-term survival, which are all correlated with the presence of CSCs.
Recently, many groups focused their attention on PPARs, suggesting the use of several PPAR agonists, currently used as hypoglycemic drugs, for NB treatment. Interestingly, many
Although further studies
The authors declare no competing interests.
Tullio Florio and Aldo Pagano equally contributed to the study and should be considered as senior authors.
The financial support of Compagnia di San Paolo is gratefully acknowledged. Aldo Pagano was supported by the IRCCS-AOU San Martino-IST, Genoa, Italy (Progetto 5 × 1000), by the University of Genoa, Genoa, Italy (Progetti di Ricerca di Ateneo, 2013), and by the Associazione Italiana per la Lotta al Neuroblastoma/Fondazione Neuroblastoma (Genoa, Italy).