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Abstract. The DNA methyltransferase (DNMT) inhibitors
azacytidine and decitabine are the most successful epigenetic
drugs to date and are still the most widely used as epigenetic
modulators, even though their application for oncological
diseases is restricted by their relative toxicity and poor
chemical stability. Zebularine (1-(β-D-ribofuranosyl)-1,2-
dihydropyrimidin-2-one), a more stable and less toxic
cytidine analog, is another inhibitor of DNMT with
concomitant inhibitory activity towards cytidine deaminase.
Unfortunately, there is no new information related to the
possible clinical applications of zebularine. Although many
new inhibitors of DNMT have been identified, none of them
can so far replace azacytidine, decitabine and, to a lesser
degree, zebularine. This review summarizes the current data
and knowledge about azacytidine, decitabine and zebularine,
and their role in present and possible future epigenetic
cancer therapy. We also discuss the molecular modes of
action of these agents with consideration of their different
toxicities and demethylation profiles, reflecting their complex
and partially overlapping biological effects.
The term ‘epigenetic’ was coined by C. H. Waddington in
1942 and from that time, the definition of ‘epigenetic’ has
evolved (1). Finally, in 2008 a consensus definition of
‘epigenetics’ was established as “stably-heritable phenotype
resulting from changes in a chromosome without alterations
in the DNA sequence”, which may lead to disease e.g.
Beckwith-Wiedemann and Prader Willi/Angelman syndromes,
as well as cellular aging and cancer (2). Although for many
years scientists have been reporting that epigenetic changes
may influence cancer development, the methodology to prove
this had some limitations. Nowadays, it is known that
epigenetic alterations are as equally responsible for
carcinogenesis as are genetic mutations. In cancer, gene
silencing through methylation occurs at least as frequently as
mutations or deletions, and leads to aberrant silencing of
normal tumor-suppressor function (3). It has also been proven
that epigenetic changes can be detected in carcinogenesis
earlier than the well-known genetic origins of cancer (4).
Epigenetic modifications rely on re-building of chromatin
structure resulting in an open or closed configuration and
thereby expressing or repressing genes which control such
basic cellular processes such as differentiation, proliferation
and apoptosis, and as a consequence, cell functions. Two
covalent modifications are responsible for these processes:
DNA methylation and nucleosomal histone tail acetylation,
which influence the epigenetic regulation of the gene
expression pattern.
DNA methylation is the most characterized epigenetic
phenomenon described as a stable epigenetic marker (5). This
process involves enzymes belonging to the DNA
methyltransferase family (DNMTs). In humans, DNMTs bind
a methyl group (-CH
3
) at the carbon 5-position of the
pyrimidine ring of cytosine in CpG dinucleotides (Figure 1A).
In somatic cells, most CpG dinucleotides are methylated,
except those located in CpG islands (6-8). These islands are
mainly located near or in the promoter regions (nearly 60% of
mammalian gene promoters) in repeated sequences as long
interspersed nuclear elements (LINEs) and short interspersed
nuclear elements (SINEs) (9), as well as in CpG island shores,
where the methylation status depends on the tissue origin (10).
The regulation of histone acetylation is controlled by two
enzyme families: histone acetyltransferases (HATs) and
histone deacetylases (HDACs). The latter promotes a higher-
order chromatin structure, which is equated with
transcriptional gene repression.
Histone proteins are not the only target of HATs and
HDACs. It is important to note that HDACs might also
2989
*These Authors contributed equally to this work.
Correspondence to: Sylwia Flis, Ph.D., Department of
Pharmacology, National Medicines Institute, Chelmska Street 30/34,
00-725 Warsaw, Poland. E-mail: sylwia.flis@yahoo.pl
Key Words: Epigenetics, DNA methylation, DNMT inhibitors,
cancer therapy, review.
ANTICANCER RESEARCH 33: 2989-2996 (2013)
Review
DNA Methyltransferase Inhibitors and Their Emerging
Role in Epigenetic Therapy of Cancer
AGNIESZKA GNYSZKA*, ZENON JASTRZĘBSKI and SYLWIA FLIS*
Department of Pharmacology, National Medicines Institute, Warsaw, Poland
0250-7005/2013 $2.00+.40
directly modulate acetylation of the non-histone proteins such
as p53, signal transducer and activator of transcription (STAT),
transcription factor E2F and others. Hence, HDACs do not
only act in an epigenetic manner. Nevertheless, DNA
methylation and histone modifications are closely related.
Methylated CpG sites in gene promoter regions are easily
recognized by specific methyl CpG binding proteins (MBPs)
which act as adapters between methylated DNA and chromatin
modifying factors. MBPs can recruit co-repressors such as
HDAC, methyltransferase and chromatin remodeling factors,
creating the protein complex which regulates gene expression
(11). If the promoter region is methylated, the corresponding
gene is repressed due to its poor recognition by transcription
factors (12). Indeed, DNMTs affect protein DNA interactions
by chromatin remodeling, determine the accessibility of DNA
to transcription factors, and are associated with under- or
overexpression of certain proteins, ultimately leading to
diverse pathologies, among which cancer (13).
Distinct changes of DNA methylation are termed
‘epimutations’ and appear to play an important role in
carcinogenesis. Consequences of epimutation are similar to
those of classic genetic mutations because the affected genes
are silenced and functional gene products cannot be
generated (14). In many types of tumors, the genomic DNA
methylation pattern is changed either through
hypermethylation (increased methylation) or
hypomethylation (decreased methylation). In cancer cells,
hypomethylation of CpG dinucleotides, especially in the
pericentromeric regions of the chromosome, may lead to
genomic instability. On the other hand, dense methylation of
CpG islands, particularly in the promoter region of tumor
suppressor genes, is associated with aberrant silencing of
transcription (15). An elevated level of methylation in cancer
cells probably results from increased activity of DNMTs,
often as a consequence of overexpression. De-regulation of
the DNMTs has been shown in many types of cancer
including of the lung, breast, stomach and colon, and as well
as in leukemia (16). Fortunately, epigenetic alterations are
potentially reversible, unlike genetic mutations. Therefore,
such alterations have become an attractive target for cancer
therapy. Since hypermethylation of tumor suppressor genes
and overexpression of DNMTs have been established as the
major key players in carcinogenesis, demethylating agents
seem to be especially promising as anticancer drugs. Re-
expression of aberrantly silenced genes and restoration of
their normal function can be achieved through the use of
DNMT inhibitors which are incorporated into the growing
DNA strand and covalently bind DNMTs.
ANTICANCER RESEARCH 33: 2989-2996 (2013)
2990
Figure 1. Scheme of DNA methylation (A) and DNA methyltansferase inhibition (B). A: DNA methylation at the 5-position of the pyrimidine ring of
cytosine is catalyzed by DNA methyltransferase. The methyl group (-CH
3
) is transferred from the cofactor S-adenosyl-L methionine (SAM), resulting
in creation of 5-methylcytosine, then the enzyme is released by β elimination. B: Trapping reaction relies on prevention of β elimination due to the
presence of the nitrogen atom at the 5-position of azacytidine, resulting in a covalent irreversible complex (6).
Currently, two DNMT inhibitors (azanucleosides) have
been approved by the US Food and Drug Administration
(FDA): azacytidine (Vidaza; Celgene) and decitabine (5 aza
2’ deoxycytidine) (Dacogen; SuperGen). These two types of
drugs are the first molecules that have been characterized as
the archetypal DNMT inhibitors and the only epidrugs that
have been approved for the treatment of patients with acute
myeloid leukemia (AML) and myelodyplastic syndrome
(MDS). Azacytidine has also been approved by the FDA and
the European Medicines Agency (EMA) for use against
chronic myelomonocytic leukemia (CMML) (10).
Therapeutic use of DNMT inhibitors can provide new and
effective solutions for patients not only with hematological
malignancies but also with other tumor types (especially
since azacytidine and decitabine are currently in phase I
clinical trials in patients with solid tumors) (17). The third
novel member of the nucleoside DNMT inhibitor family is
zebularine, a cytidine analog, which has been characterized
as a potent and promising agent because of good results
achieved in in vitro experiments, encouraging zebularine use
for future clinical trials.
In the present article, we discuss the action mechanism of
DNMT inhibitors (azacytidine, decitabine and zebularine)
with consideration of their different activities towards cancer
cells which may impact future clinical trials.
Azacytidine and decitabine are cytidine analogs modified
in position 5 of the pyrimidine ring (Figure 2). Both
compounds were synthesized by Sorm and co-workers in
1964 (18). The compounds were initially used as
antimetabolite agents in leukemia chemotherapy until their
hypomethylating properties were discovered. These cytidine
analogs are transported into cells by human concentrative
nucleoside transporter-1 and converted to active triphosphate
forms, i.e. azacytidine by uridine cytidine kinase to 5-
azacytidine 5’-triphosphate and decitabine by deoxycytidine
kinase to 5-aza-2’-deoxycytidine-5’-triphosphate and then
degraded by cytidine deaminase (CDA). Azacytidine, being a
ribonucleoside, is incorporated into RNA and, to a lesser
extent, into DNA, whereas decitabine, as a deoxyribose
analog, is incorporated only into DNA strands (Figure 3A).
To be active, these compounds need to be integrated into the
genome of rapidly proliferating cells during the S phase of
the cell cycle (19). The incorporated 5-azanucleoside disrupts
the interaction between DNA and DNMTs through nitrogen
instead of carbon, in the 5-position of the modified
pyrimidine, which precludes the resolution of the complex
and finally promotes its proteosomal degradation (Figure
1B). Thus, the enzyme remains covalently bound to DNA
and its DNMT and the function is blocked. In addition, the
covalent protein adduction also compromises the
functionality of DNA and triggers DNA damage signaling,
resulting in the degradation of the trapped DNMTs (20, 21).
Therefore, further methylation of cytosine residues is
inhibited, causing the passive loss of cytosine methylation in
the daughter cells after replication.
Another demethylating agent in the family of nucleoside
analogs is zebularine, which is characterized by chemical
stability, apparent bioavailability and low cytotoxicity (22).
Such properties distinguish zebularine among nucleoside
inhibitors (Figure 2). Zebularine was originally synthesized
and evaluated as an inhibitor of CDA to prevent de-amination
of nucleoside analogs. It acts, however, primarily as a DNMT
inhibitor by trapping the DNMT protein and forming tight
covalent complexes between the DNMT protein and
zebularine-substituted DNA (23). Zebularine is also activated
after incorporation into DNA and metabolized presumably in
a similar way to azacytidine. The initial phosphorylation is
most likely mediated by uridine-cytidine kinase, followed by
conversion to the 2’-deoxyzebularine-5’-diphosphate by
ribonucleotide reductase. Furthermore, 2’ deoxyzebularine-
5’-diphosphate is converted to 2’-deoxyzebularine-5’
triphosphate, which appears to be crucial for its incorporation
into DNA, but this step of metabolic activation still needs to
be clarified. In brief, inactivation of the DNMT by zebularine
may be related to the absence of the 4-amino group in the
6-position of its pyrimidine ring (Figure 3B). In this way,
zebularine does not allow the activation of cytosine C5
Gnyszka et al: DNMT Inhibitors in Cancer Therapy (Review)
2991
Figure 2. Nucleoside analog inhibitors of DNA methyltranferase.
position and methyl group transfer. The lack of methylation,
even if it is reversible, can clarify the stabilization of the
zebularine containing DNA binding complex and slowing its
dissociation (24).
Even though azacytidine and decitabine are used in the
clinic, they are characterized by poor chemical stability,
which depends on temperature and pH. In alkaline solutions
both agents undergo irreversible decomposition. In vitro
studies have shown that at 37˚C, in neutral aqueous
solutions, the half-lives were 7 hours for azacytidine and 21
hours for decitabine, whereas the corresponding times in vivo
are 41 minutes for azacytidine and from 15 to 25 minutes for
decitabine. At 4˚C, both agents have considerable chemical
stability, but with elevation of the temperature, they undergo
more rapid degradation (21, 25, 26).
In humans, azacytidine and decitabine are metabolized by
CDA, the enzyme which renders these drugs inactive by
converting them into 5-azauridine compounds. The high level
of CDA in the human liver and spleen is largely responsible
for shorter half-lives of both agents in vivo than in vitro.
Therefore, an increase in CDA activity may reduce efficacy
by lowering drug levels and shortening the half-life times.
Unlike azacytidine and decitabine, zebularine is highly
stable at acid and neutral pH. Zebularine has a half-life of
~44 hours at 37˚C in phosphate buffered saline (PBS) at pH
1.0 and ~508 hours at pH 7.0; such properties allow for its
oral administration (27). Moreover, taking into consideration
the half-life times of zebularine after intravenous and oral
dosing in mice, rats and rhesus monkeys, it is likely that
relatively frequent dosing or continuous intravenous infusion
of zebularine will be necessary to maintain prolonged
DNMT inhibition, which seems to be possible because of the
low toxicity of zebularine (28).
Azanucleosides in Cancer Therapy –
Mechanism of Action
The importance of DNA hypermethylation of CpG islands as
a key player during cancer development and progress is not
questioned. Especially since this phenomenon occurs
infrequently in normal cells, the abnormal transcriptional
silencing of tumor suppressor genes by hypermethylation of
CpG islands has become an attractive and selective tumor
specific therapeutic cancer target (3, 27). During gene
methylation, the DNA sequence, as well as protein product,
remains unaltered. Therefore, pharmacological intervention
in the form of chemical inhibitors seems to be one of the
possible ways for de-repression of inappropriately silenced
genes and restoration of their normal functions.
The ability of azacytidine and decitabine to deplete the
DNA methylating activity of DNMT can be achieved at low
doses of both agents. Stresemann et al. have shown that
azacytidine and decitabine are able to induce demethylation
of epigenetically silenced genes at least at concentrations that
ANTICANCER RESEARCH 33: 2989-2996 (2013)
2992
Figure 3. Metabolic activation of 5 azanucleosides (A) and zabularine (B). After cellular uptake 5-azanucleosides and zebularine are modified by
different metabolic pathways as described in the text. CDA: Cytidine deaminase; UCK: uridine cytidine kinase; dCK: deoxycytidine kinase; CMP
kinase: cytidylate kinases; dCMP: deoxycytidylate kinase; 5-Aza-U: 5-aza-uridine; 5-Aza-CTP: 5 azacytidine 5’-triphosphate; 5-Aza-dCTP: 5-aza
2’-deoxycytidine-5’-triphosphate; ZTP: zebularine-5’-triphosphate; dZTP: 2’-deoxyzebularine-5’-triphosphate (19, 22).
exceed 20% inhibition of cell growth (IC
20
). In vitro
experiments on whole-genome methylation, as well as local
methylation at a defined genomic locus revealed that
azacytidine and decitabine at 5-fold the IC
20
concentration
strongly reduces the genomic DNA methylation level in
lymphoid cancer cells as compared to controls, i.e. by 60%
and 40%, respectively. A similarly strong concentration-
dependent demethylation was observed after incubation of
colon carcinoma cells with both agents at concentrations
centered on the respective drug specific IC
20
concentration
(29). Independent studies have shown that these two epidrugs
may significantly reactivate silenced genes [tissue inhibitor
of metalloproteinase 3 (TIMP3), p15, p16, cyclin-dependent
kinase inhibitor 1C (CDKN1C), RAS-association domain
family 1 (RASSF1)] responsible for basic cellular mechanisms
such as apoptosis, cell cycle, and DNA repair (29), and that
the concentrations required of both agents to achieve this
effect are not high. Azacytidine and decitabine should be used
at low inhibitory concentrations because at higher
concentrations these agents exert strong cytotoxicity, interfere
with DNA synthesis and cause DNA damage (30, 31). Apart
from the use of low concentrations, for their demethylation
function, the S phase of the cell cycle is needed. The DNA
replication phase of the cell cycle enables selective and
effective incorporation of these substances into the DNA of
rapidly dividing cancer cells, reducing, thereby,
hypomethylation in normal cycling cells (32). However, the
hypomethylation activity of these two agents is not equal. It
has been shown that azacytidine has only about 10% the
potency of decitabine at inhibiting DNA methylation (33).
Decitabine as a deoxyribonucleotide is directly incorporated
into the DNA after phosphorylation to inhibit DNA
methylation, whereas azacytidine is additionally incorporated
into RNA. The overall incorporation of azacitydine into RNA
can account for about 80-90% (34). Decrease of tRNA
acceptor activity, polyribosome breakdown and incorporation
into mRNA causing the subsequent inhibition of protein
synthesis and enzyme induction are the functional
consequences of RNA synthesis inhibition by azacytidine
(35). All these in effect may influence both cancer and normal
cells, resulting in greater in vitro and in vivo side-effects.
It has been proven that the mechanism of action of both
these agents is dose-dependent. Therefore, the concept of a
dual mechanism of action of these two agents has arisen. For
decitabine, the ‘dual mechanism’ refers to the inhibition of
cell proliferation at high doses and to the DNA
hypomethylation-mediated effect on gene re-expression at
low doses affecting the processes of cell differentiation and
tumor suppression, whereas for azacytidine the ‘dual
mechanism’ refers to the cytotoxicity at high doses, via
incorporation into RNA and DNA, and to the DNA
hypomethylation effect at lower doses (34). Their mechanism
of action at the highest doses is related to the formation of
covalent DNMT-DNA adducts in aza-containing DNA,
leading to DNA damage and cytotoxicity. Experiments
conducted on human tumor cell lines have shown that
treatment with these agents causes growth inhibition by cell
cycle arrest (specific to G
2
/M phase for decitabine and cell
cycle non-specific for azacytidine) and reduction in
clonogenic survival. Moreover, decitabine and azacytidine
can induce apoptosis in p53-dependent or p53-independent
manners, respectively (36-38).
Unfortunately, these agents also have some limitations. In
spite of their clinical efficacy, azacytidine and decitabine are
characterized by poor bioavailability, instability in
physiological media and high toxicity, restricting their use.
For this reason, one of the most promising molecules appears
to be zebularine. It has been characterized as a potential
antitumor agent, based on its stability (half-life of >500
hours at pH 7.4) and minimal toxicity both in vitro and in
vivo (39-41). Once incorporated into DNA, zebularine is
involved in the reactivation of the silenced genes, such as
cyclin-dependent kinase inhibitors (CKI) p15 in AML cells,
p16 in bladder, colon and pancreatic cells, and p57 in
myeloid leukemia cells (42, 43). Zebularine also leads to the
re-expression of the cell cycle and apoptosis modulator Ras
association domain family 1 isoform A (RASSF1A) as a
result of induced demethylation of its promoter region in
ovarian cancer cells. Moreover, microarray analysis
demonstrated that decitabine and zebularine may
demethylate 78 genes, with 32 exclusive to decitabine and
only eight specific for zebularine (44). This might suggest
that both agents exert their demethylating effects by different
mechanisms. Cheng et al. observed that zebularine caused
complete depletion of DNMT1 and partial depletion of
DNMT3A and DNMT3B2/3 in cancer cells (45). This
suggests zebularine preference for DNMT1 over DNMT3A
and DNMT3B. Moreover, DNMT1 may be an important
indicator of the demethylating ability of zebularine. It has
also been reported that DNMT3A/DNMT3B double-null
embryonic stem cells are more resistant to decitabine than
are DNMT1 null cells, suggesting that decitabine may be
more effective for selected types of cancer cells, in which
DNMT3 expression is up-regulated (46). Furthermore,
DNMTs have a higher affinity for zebularine-containing
DNA than for the unmodified DNA (24). Such results
indicate that the differential specificity and affinity of these
drugs for DNMT isoforms could lead to divergent cellular
responses. It has been demonstrated that higher doses of
zebularine are required to obtain demethylation and gene re-
expression levels comparable to those that are induced by
azacytidine and decitabine (29). This might be a result of
lower binding affinity of uridine-cytidine kinase for
zebularine and its slow conversion to 2’-deoxyzebularine-5’-
diphosphate (40). However, owing to its minimal toxicity
profile, zebularine, unlike azacytidine and decitabine, can be
Gnyszka et al: DNMT Inhibitors in Cancer Therapy (Review)
2993
used at high micromolar concentrations in prolonged
treatment periods (41, 47). The lower toxicity of zebularine,
as a ribonucleoside, may result from its different
incorporation into RNA and DNA in normal and
neoplastically-transformed cells, due to the overexpression
of uridine-cytidine kinase in cancer cells, facilitating
zebularine insertion into nucleic acids (44). Moreover,
Champion et al. have shown that the absence of the 4-amino
group in zebularine does not allow for the activation of the
cytosine C5 position after covalent intermediate formation
and methyl group transfer (24). Such lack of zebularine
methylation seems to prevent dissociation of the DNA-
enzyme complex, indicating the stabilization of the
zebularine-containing DNA-binding complex.
Future medical use of zebularine could involve
combinations of this drug with other therapeutic modalities,
such as chemotherapy, immunotherapy or radiotherapy.
Clinical trials with azacytidine or decitabine administered as
single agents or in co-administration with other
chemotherapeutics resulted in significant toxicity (48, 49).
Therefore, zebularine and other demethylation agents with
similar properties could be less detrimental and more
promising drug candidates.
Conclusion and Future Perspectives
Azacytidine, decitabine and zebularine have differential
activity, complex and partially overlapping mechanisms of
action. Studies have indicated that the methylation patterns
of tumor suppressor genes might differ depending on the
drug being used. Azacytidine and decitabine induce a strong
demethylating effect, leading at lower doses to re expression
of aberrantly silenced genes associated with reduced
proliferation, cell differentiation, apoptosis, and senescence,
while at higher doses, their main effect consists of DNA
damage after incorporation into genomic DNA. The impact
of zebularine on the DNA methylation level is more
moderate. However, this cytidine analog is less toxic and
can, therefore, be given continuously at high doses.
Additionally, zebularine seems to target tumor cells
preferentially.
The use of demethylating agents azacytidine and decitabine
in the treatment of myelodysplastic syndromes is well-
documented, in spite of their high toxicity. But the status of
knowledge for using demethylating agents for solid tumors is
still insufficient and needs to be further evaluated. It appears
to be reasonable to use demethylating agents in combination
with chemotherapeutic agents. In vitro experiments indicate
that decitabine can potentiate the cytotoxic effect of classical
chemotherapeutics, such as doxorubicin, 5-fluorouracil and
oxaliplatin, and induce highly synergistic effects (50, 51).
Interestingly, encouraging results were obtained with
combination of decitabine and carboplatin in patients with
solid tumors (52). The authors concluded that decitabine
combines safely with carboplatin and that the specific
regimen causes epigenetic changes. In another phase I study,
a combination of cisplatin with decitabine resulted in one
partial response in a patient with cervical cancer, and two
minor responses: one in a patient with non-small cell lung
cancer and another in a patient with cervical cancer (17).
Generally, combinations of azacytidine or decitabine with
standard chemotherapy clearly have clinical activity, but it is
difficult to distinguish the effect of the epigenetic therapy
from the cytotoxic therapy (17). On the other hand, the
present demethylating agents used in clinics are cytotoxic,
mutagenic and exhibit lack of specificity towards genes,
which may limit their clinical application. Thus, the next
generation of DNMT inhibitors with lower toxicity might be
more fruitful for future research. Zebularine is an alternative
derivative of cytidine and promises to be a better drug than
azacytidine and decitabine for epigenetic cancer therapy.
Based on its stability, it was the first DNMT inhibitor
showing in vivo antitumor activity against T-cell lymphoma
after oral administration (53). Attention has also been
attributed on the identification of small non-nucleoside
DNMT inhibitors, such as epigallocatechin-3-gallate,
hydralazine, procainamide, procaine and RG108, which bind
directly to the catalytic region of DNMTs without
incorporation into DNA. However, the results of in vitro
studies showed that non-nucleoside compounds induce
limited epigenetic changes in living cells (54). Among the
novel agents of demethylation, the most intensively studied
are DNMT1 antisense and siRNA. Down-regulation of
DNMT1 by antisense or siRNA is sufficient to restore the
expression of aberrantly hypermethylated genes (55, 56), but
such therapy is still controversial mainly because of the
problems associated with the administration to humans,
which include instability, toxicity, risk of non-specific effects,
and complexity in developing a suitable delivery system.
Even though many DNMT inhibitors other then cytidine
analogues have been developed, their effects are not
satisfactory and they do not seem to be able to replace
azacytidine and decitabine, as yet. Therefore, azacytidine and
decitabine, in spite of their limitations, are still used for
combination epigenetic therapy. However, attempts to
discover and to develop novel compounds targeting DNMTs
should be continued. It is important to find more selective and
less toxic DNMT inhibitors which will be effective especially
in patients with solid tumors. It would be a challenge to
design inhibitors whose mechanism of action will rely only
on reactivation of abnormally silenced suppressor genes.
Acknowledgements
This work was supported by National Science Centre, Poland (grant
N405 139139).
ANTICANCER RESEARCH 33: 2989-2996 (2013)
2994
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Received May 10, 2013
Revised May 24, 2013
Accepted May 27, 2013
ANTICANCER RESEARCH 33: 2989-2996 (2013)
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