1 INTRODUCTION
Cancer is one of the leading causes of human death worldwide,
characterized by uncontrolled proliferation and locally invasive
infiltration.1-3 Because of the evading immune
recognition through genetic mutations, the balance between cell death
and proliferation is broken, leading to the unrestricted cell
proliferation and ultimately cancer.4,5 While
advanced techniques have been applied to basic and clinical cancer
research, most of them are arduous and unsatisfactory in performance, as
existing approaches can barely kill cancer cells while sparing
surrounding normal cells.6,7 Selectively inducing
programmed cell death (PCD) in cancer cells is a promising option for
cancer therapy, including apoptosis, autophagy, necrosis. Among them,
apoptosis is considered the preferred alternative, but the therapeutic
effects are still far from satisfactory due to the intrinsic resistance
induced by tumor heterogeneity.8,9 Furthermore,
acquired resistance results in high doses of medication, which brings
severe side effects.10,11 Thus, development of novel
cell death mode with more efficiency and less side effect is an urgent
concern in the field of cancer therapy. Fortunately, recent studies have
proposed several unidentified cell death forms with unique regulatory
pathways, including ferroptosis, pyroptosis and cuproptosis, which can
circumvent the limitations of classical cell death methods and open up
new opportunities for the cancer treatment.12-15
Among these non-apoptotic forms of PCDs, cuproptosis has received much
attention as the most emerging regulatory pathways of cell death.
Interestingly, the connection between copper homeostasis and physical
health was explored long before the term cuproptosis was established.
The disequilibrium of copper homeostasis was repeatedly found to be
associated with development of various diseases, such as Menkes disease,
Wilson’s disease, neurodegenerative diseases, cardiovascular diseases,
and cancer.16-20 Despite the apparent importance of
copper for physiology and pathology, the underlying cellular mechanisms
are still largely unknown, which prompts extensive exploration of copper
in the treatment of various diseases. For example, the same
morphological and molecular changes were observed after treating cancer
cells with disulfiram (copper ionophore), pyrazole-pyridine copper
complexes and inorganic copper, indicating that copper overload was the
cause of cell death.21 Moreover, the killing effect of
elesclomol (copper ionophore) was totally lost on coadministration of
MDA-MB435 melanoma cells in the absence of serum (the source of copper),
while it can be restored after adding copper instead of iron, manganese
and zinc to the serum-free medium, suggesting a potential copper
ion-regulated cell death mode.22 Afterwards, the
anti-cancer effect of elesclomol was further corroborated through
inducing a variety of cells, including melanoma cells, lung cancer
cells, glioblastoma stem cells (GSCs) and gynecological tumor cells, to
produce reactive oxygen species (ROS).23-26 However,
the oxidative stress alone does not fully explain the mechanism of cell
death, because the use of ROS scavenger N-acetylcysteine (NAC) can only
partly reverse the elesclomol induced cancer cell
death.27,28 Therefore, apart from oxidative stress,
elesclomol should have additional mechanisms to regulate cancer cell
death.
Gratifyingly, the “cuproptosis” proposed by Tsvetkov et al .
provides a definitive explanation for the anti-cancer mechanism of
elesclomol (Figure 1 ).29 Firstly, different
metal ions including copper were carried by elesclomol to verify that
only copper ions can mediate cancer cell death, which could be reversed
by the copper ion chelating agents glutathione (GSH) and
tetrathiomolybdate (TTM) but not by known inhibitors of various cell
death pathways (ferrostatin-1, necrostatin-1, NAC), confirming that cell
death induced by copper ions, namely cuproptosis, may be a new type of
cell death mode different from traditional cell death such as apoptosis,
ferroptosis, necrosis and autophagy. Then, mitochondrial
respiration-dependent cells showed higher sensitivity to copper ions
compared with glycolysis-dependent cells, suggesting that cuproptosis
was related to mitochondrial metabolism. Further investigation revealed
that the respiratory reserve capacity was significantly reduced after
copper ion treatment, while basic respiration or ATP-related respiration
remained stable, indicating that copper acted on the components of the
tricarboxylic acid (TCA) cycle rather than electron transport chain
(ETC). After that, seven TCA cycle genes were identified as relevant for
cuproptosis mechanism, including FDX1 (a reductase reducing
Cu2+ to Cu+), LIPT1, LIAS, DLD
(three key enzymes of the lipoic acid pathway), DLAT, PDHA1, and PDHB
(three components of the pyruvate dehydrogenase complex). Moreover, FDX1
and LIAS knockdown alleviated copper ionophore-triggered cytotoxicity,
emphasizing the inherent association between TCA cycle and cuproptosis.
Furthermore, the copper ions reduced by FDX1 bound to the lipoacyl group
of DLAT to promote its lipoacylation and aggregation, thereby exerting
cytotoxicity. In addition, FDX1 also leaded to the instability of Fe–S
cluster protein, which exacerbated cell death. Notably, in vivoexperiments confirmed that cell death caused by copper homeostasis
imbalance and cell death induced by copper ionophore belonged to the
same cuproptosis mechanism. Altogether, cuproptosis is a
copper-dependent form of novel cell death. The establishment of the
concept not only illuminates the cytotoxic mechanism in copper
ionophore, but also provides new insights for the treatment of various
diseases including cancer.