|Recent efforts to develop novel strategies for the treatment of
cancer have focused on targeting the essential element, iron, which
is crucial for DNA synthesis, cell cycle progression and metabolism
. This strategy led to the development of a potent and selective
new class of anti-cancer agents known as the thiosemicarbazone iron
chelators . One of the most well characterized thiosemicarbazones
is 3-AP or Triapine®, which has entered over 20 Phase I and Phase II
clinical trials [2-4]. Although 3-AP has shown some promise [2,5],
its further development has been hampered by a host of serious side
effects including hypoxia and methemoglobinemia [3,4]. To overcome
these problems, a new series of dipyridyl thiosemicarbazone (DpT)
iron chelators have been developed, with the lead agents being di-2-
pyridylketone 4, 4-dimethyl-3-thiosemicarbazone (Dp44mT) and di-
2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC)
. These novel compounds were demonstrated to selectively inhibit
tumor growth in a wide range of cancers both in vitro and in vivo,
being well tolerated and having no marked toxicity at the optimal
doses required for tumor inhibition [6,7]. These agents are also able to
target a number of crucial molecules that control cancer progression
including ribonucleotide reductase , p53 , cyclin D1 , p21
 etc. Furthermore, DpC does not induce metHb in vivo in animal
models, and thus, this agent shows considerable advantages over both
3-AP and an earlier DpT analog, Dp44mT .
|Although these studies present a significant step forward towards
developing an effective treatment strategy for cancer, an important
question still must be answered: can these agents inhibit cancer
metastasis? To address this, a number of recent studies have examined
one of the key down-stream targets of iron chelators, namely the
metastasis suppressor, N-myc down-stream regulated gene 1 (NDRG1)
[6,7,11]. NDRG1 has been demonstrated to potently inhibit metastasis
in a number of tumors including prostate, breast, lung, colon and
pancreatic cancer, with its expression being correlated with less
aggressive neoplasms and better patient prognosis [11-13]. Although
the molecular mechanisms behind these effects remain to be completely
established, there is increasing evidence that NDRG1 can inhibit the
key pathways that are involved in metastasis, namely the transforming
growth factor-β (TGF-β), phosphatidylinositol 3-kinase (PI3K), Ras
and WNT signaling pathways [13,14].
|TGF-β normally activates a signaling cascade that regulates cell
proliferation . However, in many cancer cells, this pathway is
de-regulated due to the loss of a crucial down-stream target, namely
the tumor suppressor, SMAD4 . As a result, TGF-β becomes
oncogenic, leading to the activation of Ras and PI3K pathways that
promote cancer progression and metastasis (Figure 1; [15-17]). In
fact, TGF-β can initiate epithelial to mesenchymal transition (EMT) in
advanced cancer cells, leading to reduced membrane expression of the
adhesion molecules E-cadherin and β-catenin, as well as increased cell
migration and invasion . Recent studies have revealed that NDRG1
can inhibit the oncogenic effects of TGF-β in cancer cells [14,16]. In
fact, over-expression of NDRG1 increased the levels of E-cadherin and
β-catenin at the membrane and inhibited both migration and invasion
of colon and prostate cancer cells upon TGF-β treatment (Figure 1).
|Moreover, NDRG1 also prevented the oncogenic down-stream
effects of TGF-β in pancreatic cancer cells and this was mediated by an
increase in the key tumor suppressor molecules phosphatase and tensin
homologue deleted on chromosome 10 (PTEN) and SMAD4 (Figure 1;
). As a result, NDRG1 over-expression led to the inactivation of the
oncogenic PI3K and Ras [14,16].
|Further studies examining the mechanisms behind the effect
of NDRG1 on β-catenin expression revealed that this metastasis
suppressor is also able to inhibit the WNT signaling pathway [13,16].
WNT signaling plays a crucial role in EMT and metastasis, leading to
down-regulation of E-cadherin and an increase in oncogenic molecules
such as cyclin D1 and Myc (Figure 1; ). The down-stream effects
of WNT signaling are primarily mediated by β-catenin, which
translocates from the membrane to the nucleus where it can function as
part of a transcriptional complex with Lef-1 to mediate expression of its
target genes (Figure 1; [18,19]). Recent studies have demonstrated that
NDRG1 can inhibit WNT signaling by directly binding to the WNT
co-receptor LRP6 . In addition, NDRG1 also activates molecules
such as GSK3β, that are involved in degrading free β-catenin that is not
part of the adherens junction (Figure 1; ). Hence, the combined
effects of NDRG1 on the TGF-β, PI3K, Ras and WNT signaling
pathways leads to increased levels of E-cadherin and β-catenin at the
cell membrane, where they promote cell adhesion and inhibit cell
invasion and metastasis.
|Considering the potent ability of NDRG1 to inhibit metastatic
progression, targeting this latter molecule may be a promising
strategy for cancer treatment. In fact, recent studies examining novel
thiosemicarbazone iron chelators that markedly up-regulate NDRG1
expression in cancer cells have shown these agents to be highly
effective at inhibiting cancer progression [6,7,13]. The mechanism
involved in the up-regulation of NDRG1 involves hypoxia-inducible
factor-1α (HIF-1α)-dependent and independent pathways . The
thiosemicarbazones, Dp44mT and DpC, were found to markedly
up-regulate NDRG1 expression in pancreatic cancer cells, and this
probably played a role in significantly reducing tumor growth in vivo
. Indeed, these latter agents also inhibited important down-stream
targets of the Ras pathway, namely pERK and pSMAD2L in vivo, both
of which are also down-regulated by NDRG1 .
|In addition, Dp44mT and the “gold standard” iron chelator,
desferrioxamine, also markedly up-regulated the cell adhesion
markers, E-cadherin and β-catenin, in prostate and colon cancer cells,
leading to reduced migration and invasion . This was accompanied
by a marked decrease in the mesenchymal marker, vimentin, further
indicating that these agents are able to reverse the EMT . Finally,
Dp44mT was also demonstrated to inhibit breast cancer metastasis
in vivo, with this effect being dependent on its ability to up-regulate
|In summary, the ability of novel thiosemicarbazone iron chelators
to markedly inhibit cancer progression extends to more than just an
inhibition of primary tumour growth. Indeed, these agents are also able
to markedly reduce cancer metastasis and this is likely to occur, at least
in part, through the up-regulation of the metastasis suppressor, NDRG1
(Figure 1). The ability of thiosemicarbazones to target NDRG1 and
other key molecular players (e.g., cyclin D1, ribonucleotide reductase,
p21, etc.) via their ability to bind iron demonstrates the potential of this
exciting new group of anti-cancer agents.
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