Mechanism of the induction of endoplasmic reticulum stress by the anti-cancer agent, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT): Activation of PERK/eIF2α, IRE1α, ATF6 and calmodulin kinase.

The endoplasmic reticulum (ER) plays a major role in the synthesis, maturation and folding of proteins and is a critical calcium (Ca(2+)) reservoir. Cellular stresses lead to an overwhelming accumulation of misfolded proteins in the ER, leading to ER stress and the activation of the unfolded protein response (UPR). In the stressful tumor microenvironment, the UPR maintains ER homeostasis and enables tumor survival. Thus, a novel strategy for cancer therapeutics is to overcome chronically activated ER stress by triggering pro-apoptotic pathways of the UPR. Considering this, the mechanisms by which the novel anti-cancer agent, Dp44mT, can target the ER stress response pathways were investigated in multiple cell-types. Our results demonstrate that the cytotoxic chelator, Dp44mT, which forms redox-active metal complexes, significantly: (1) increased ER stress-associated pro-apoptotic signaling molecules (i.e., p-eIF2α, ATF4, CHOP); (2) increased IRE1α phosphorylation (p-IRE1α) and XBP1 mRNA splicing; (3) reduced expression of ER stress-associated cell survival signaling molecules (e.g., XBP1s and p58(IPK)); (4) increased cleavage of the transcription factor, ATF6, which enhances expression of its downstream targets (i.e., CHOP and BiP); and (5) increased phosphorylation of CaMKII that induces apoptosis. In contrast to Dp44mT, the iron chelator, DFO, which forms redox-inactive iron complexes, did not affect BiP, p-IRE1α, XBP1 or p58(IPK) levels. This study highlights the ability of a novel cancer therapeutic (i.e., Dp44mT) to target the pro-apoptotic functions of the UPR via cellular metal sequestration and redox stress. Assessment of ER stress-mediated apoptosis is fundamental to the understanding of the pharmacology of chelation for cancer treatment.

Expanding horizons in iron chelation and the treatment of cancer: role of iron in the regulation of ER stress and the epithelial-mesenchymal transition.

Cancer is a major public health issue and, despite recent advances, effective clinical management remains elusive due to intra-tumoural heterogeneity and therapeutic resistance. Iron is a trace element integral to a multitude of metabolic processes, including DNA synthesis and energy transduction. Due to their generally heightened proliferative potential, cancer cells have a greater metabolic demand for iron than normal cells. As such, iron metabolism represents an important "Achilles' heel" for cancer that can be targeted by ligands that bind and sequester intracellular iron. Indeed, novel thiosemicarbazone chelators that act by a "double punch" mechanism to both bind intracellular iron and promote redox cycling reactions demonstrate marked potency and selectivity in vitro and in vivo against a range of tumours. The general mechanisms by which iron chelators selectively target tumour cells through the sequestration of intracellular iron fall into the following categories: (1) inhibition of cellular iron uptake/promotion of iron mobilisation; (2) inhibition of ribonucleotide reductase, the rate-limiting, iron-containing enzyme for DNA synthesis; (3) induction of cell cycle arrest; (4) promotion of localised and cytotoxic reactive oxygen species production by copper and iron complexes of thiosemicarbazones (e.g., Triapine(®) and Dp44mT); and (5) induction of metastasis and tumour suppressors (e.g., NDRG1 and p53, respectively). Emerging evidence indicates that chelators can further undermine the cancer phenotype via inhibiting the epithelial-mesenchymal transition that is critical for metastasis and by modulating ER stress. This review explores the "expanding horizons" for iron chelators in selectively targeting cancer cells.