The Anticancer Agent, Di-2-Pyridylketone 4,4-Dimethyl-3-Thiosemicarbazone (Dp44mT), Up-Regulates the AMPK-Dependent Energy Homeostasis Pathway in Cancer Cells

Abstract

Adenosine monophosphate-activated protein kinase (AMPK) is a cellular energy sensor that monitors ATP levels. There is also evidence that AMPK has onco-suppressive properties. Iron plays a crucial role in cellular energy transducing pathways and tumor cell proliferation. Therefore, metals (e.g., iron) could play an important role in the regulation of AMPK-dependent pathways. Hence, this investigation examined the effect of the iron and copper chelator and potent anti-cancer agent, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), on the AMPK-mediated pathway. These studies demonstrated that Dp44mT, which forms intracellular redox-active complexes with iron and copper, significantly activated AMPK (i.e., p-AMPK/AMPK ratio) in 5 different tumor cell-types. Furthermore, examination of the Dp44mT-metal complexes demonstrated that the effect of Dp44mT on AMPK was due to a dual mechanism: (1) its ability to chelate metal ions; and (2) the generation of reactive oxygen species (ROS). The activation of the AMPK-pathway by Dp44mT was mediated by the upstream kinase, liver kinase B1 (LKB1) that is a known tumor suppressor. Moreover, using AMPKα1-selective silencing, we demonstrated that Dp44mT activated AMPK, resulting in inhibition of acetyl CoA carboxylase 1 (ACC1) and raptor, and activation of Unc-51 like kinase (ULK1). These effects are vital for inhibition of fatty acid synthesis, suppression of protein synthesis and autophagic activation, respectively. Together, this AMPK-mediated repair response aims to rescue the loss of metal ions via chelation and the induction of cytotoxic damage mediated by redox cycling of the Dp44mT-metal ion complex. In conclusion, this study demonstrates for the first time that chelators target the AMPK-dependent pathway.

Increased intracellular iron in mouse primary hepatocytes in vitro causes activation of the Akt pathway but decreases its response to insulin

“…It has been shown that di-2-pyridylketone 4,4-dimethyl-3-thiose-micarbazone (Dp44mT), a novel iron and copper chelator that has been shown to have anti-cancer activity, increases AMPK activity in various cancer cell lines [52]. This is in contrast to the results of the present study where we have shown activation of AMPK in response to an increase in intracellular iron (Fig.…”

“…Chelation of iron and copper by Dp44mT can therefore inhibit oxidative phosphorylation resulting in accumulation of AMP, which is a potent activator of AMPK [21]. Dp44mT is also known to generate ROS and induce oxidative stress in cells [52]; oxidative stress is known to activate AMPK [21].…”

Synergy of Iron Chelators and Therapeutic Peptide Sequences Delivered via a Magnetic Nanocarrier

“…D [KLAKLAK] 2 is capable of disrupting mitochondrial membranes, thus leading to apoptosis of cancer cells [ 6 ]. We have chosen a derivative of the iron-chelator Dp44mT [ 9 , 10 , 11 , 12 , 13 ] as a small molecule drug, because it is known to target several pathways in tumors. PLFAERL is cleaved by calpains, Ca 2+ -dependent cytosolic proteases that are overexpressed in virtually all solid tumors.…”

“…Iron is an important element for multiple cellular processes, as for instance proliferation, and DNA synthesis [ 20 , 21 ]. Dp44mT (Di-2-pyridylketone-4, 4, -dimethyl-3-thiosemicarbazone) is known to effectively chelate iron(II) and iron(III), generate reactive radicals[ 10 ], and interact with several pathways: (1) Dp44mT regulates the AMPK-dependent energy homeostasis pathway in cancer cells [ 11 ]; (2) Dp44mT induces endoplasmic reticulum (ER) stress and activates PERK/eIF3α, IRE1α, ATF6, and calmodulin kinase, resulting in ER-stress-associated pro-apoptotic signaling [ 12 ]; (3) Dp44mT decreases the efficacy of mitochondrial peroxidases, such as peroxiredoxin-3 (Prx3), which are important for cell survival in multiple tumors; (4) Dp44mT can selectively inhibit topoisomerase II α in (breast) cancer cells, leading to G 1 cell cycle arrest at nanomolar concentrations [ 9 ]. The dopamine-tethered Dp44mT-derivative is released from the surface of Fe/Fe 3 O 4 nanocarriers, because the dopamine-anchors are replaced by glutathione and other thiol-containing peptides and proteins in the cytoplasm.…”

“…We will then activate several signaling cascades triggering apoptosis: The D [KLAKLAK] 2 sequence will disrupt the mitochondrial cell walls [ 6 ]. Dp44mT will regulate the AMPK-dependent pathway [ 11 ], create ER-stress [ 12 ], reduce mitochondrial peroxidase activity and cause G 1 cell cycle arrest [ 13 ]. AERL D [KLAKLAK] 2 CGKRK and the Dp44mT-derivative will be released from the nanocarrier after dopamine-coated Fe 3 O 4 -nanoparticles have escaped from early endosomes into the cytoplasm, as previously demonstrated [ 15 ].…”