Iron-targeting antitumor activity of gallium compounds and novel insights into triapine(®)-metal complexes

Abstract

Significance: Despite advances made in the treatment of cancer, a significant number of patients succumb to this disease every year. Hence, there is a great need to develop new anticancer agents.

Recent advances: Emerging data show that malignant cells have a greater requirement for iron than normal cells do and that proteins involved in iron import, export, and storage may be altered in cancer cells. Therefore, strategies to perturb these iron-dependent steps in malignant cells hold promise for the treatment of cancer. Recent studies show that gallium compounds and metal-thiosemicarbazone complexes inhibit tumor cell growth by targeting iron homeostasis, including iron-dependent ribonucleotide reductase. Chemical similarities of gallium(III) with iron(III) enable the former to mimic the latter and interpose itself in critical iron-dependent steps in cellular proliferation. Newer gallium compounds have emerged with additional mechanisms of action. In clinical trials, the first-generation-compound gallium nitrate has exhibited activity against bladder cancer and non-Hodgkin's lymphoma, while the thiosemicarbazone Triapine(®) has demonstrated activity against other tumors.

Critical issues: Novel gallium compounds with greater cytotoxicity and a broader spectrum of antineoplastic activity than gallium nitrate should continue to be developed.

Future directions: The antineoplastic activity and toxicity of the existing novel gallium compounds and thiosemicarbazone-metal complexes should be tested in animal tumor models and advanced to Phase I and II clinical trials. Future research should identify biologic markers that predict tumor sensitivity to gallium compounds. This will help direct gallium-based therapy to cancer patients who are most likely to benefit from it.

FIG. 1.

Iron proteins in breast cancer cells. Under physiologic conditions, iron is bound to transferrin (Tf) in the circulation and is incorporated into cells by transferrin receptor1 (TfR1)-mediated endocytosis of Tf-Fe complexes. The binding site of the wild-type hemochromatosis protein (wt HFE) partially overlaps with the Tf-binding site on TfR1 and can, thus, competitively inhibit Tf binding to its receptor. This regulatory effect of HFE on Tf-Fe-TfR binding is lost with the HFE C282Y mutation, as the latter is degraded within the cell and no longer associates with the TfR to interfere with its binding to Tf-Fe. The Tf-Fe-TfR complex translocates from the cell surface to an intracellular acidic endosome, where Fe(III) dissociates from Tf and is reduced to Fe(II) by STEAP3 (six-membrane epithelial antigen of the prostate 3) (not shown). Fe(II) exits the endosome through divalent metal transporter1 (DMT1, not shown) to a labile iron “pool.” From here, iron trafficks to different compartments (mitochondria, ribonucleotide reductase [RR], and others). Excess iron is stored in ferritin. Iron exits from the cell through cell membrane-based ferroportin. Ferroportin levels can be lowered by hepcidin, which binds to it and translocates it to the lysosome for degradation. Cytoplasmic iron regulatory proteins (IRPs) function as sensors of cellular iron status and regulate the synthesis of Tf receptors, ferritin, and ferroportin at the mRNA translational level by interactions with iron response elements (IREs) present in the untranslated regions of their respective mRNAs. Iron proteins known to be altered in breast cancer are marked with an asterisk (*) and include an increase in TfR and ferritin as well as a reduction in ferroportin levels. In addition, the C282Y HFE mutation may be associated with an increased risk of breast cancer development. (To see this illustration in color, the reader is referred to the web version of this article at www.liebertpub.com/ars.)

FIG. 2.

Interaction of gallium with cellular iron metabolism. The potential sites of gallium's interaction with cellular iron metabolism are identified in the bordered boxes. Membrane transport: TfR mediated. In the circulation, gallium(III) is transported and bound to Tf as Tf-Ga and is incorporated into TfR1-expressing cells by TfR1-mediated endocytosis of Tf-Ga complexes. Tf-Ga inhibits cellular iron incorporation by interfering with TfR-mediated uptake of Tf-Fe and the release of Fe from Tf within the endosome. Non-TfR mediated. Low-molecular-weight gallium and iron chelates in the circulation may also be taken up by certain cells through a shared Tf-independent pathway. In this pathway, gallium may enhance cellular iron uptake and vice versa. In cells, gallium can be detected in a low-molecular-weight pool. TfR1 and ferritin mRNA translation: Consistent with the induction of cellular iron deprivation, cells exposed to gallium display an increase in TfR1 mRNA and protein. This results from an increase in IRP-IRE mRNA interaction that leads to increased TfR mRNA translation. RR: Gallium blocks iron incorporation into the R2 subunit and may itself be incorporated into this subunit, rendering it inactive. Gallium also blocks RR enzyme activity more directly through competitive inhibition of substrate binding. This could result from the formation of Ga-ADP/CDP complexes that compete with (Fe)-ADP/CDP binding to the enzyme. The inhibition of RR activity contributes to gallium-induced cell death. Iron-dependent mitochondrial function: Gallium may perturb mitochondrial function by action on the numerous iron-containing proteins present in the citric acid cycle and the electron transport chain (shown in Fig. 4). Upstream from the mitochondrion (mito), gallium may activate proapoptotic Bax, which translocates to the mitochondria, produces a loss of mitochondrial membrane potential, and the release of cytochrome c and apoptogenic factors to the cytoplasm. This triggers the activation of effector caspases-3 and -7, leading to apoptotic cell death. ADP, adenosine diphosphate; CDP, cytidine diphosphate.

FIG. 3.

Ribonucleotide reductase (RR). RR is responsible for the reduction of ribonucleoside diphosphates (NDPs) to deoxyribonucleoside diphosphates (dNDPs). The enzyme is a heterodimer that consists of an R1 and R2 subunit. The R2 subunit contains a dinuclear iron center and a tyrosyl-free radical (*Tyr) that is detected by EPR spectroscopy to produce the signal shown in the figure. The amplitude of the EPR signal is closely correlated with enzyme activity and is dependent on the presence of iron in the R2 subunit. EPR, electron paramagnetic resonance.

FIG. 4.

Potential sites of action for gallium in the mitochondria. The iron-sulfur cluster (Fe-S) proteins in the citric acid cycle and mitochondrial complexes are potential targets for the cytotoxic action of gallium compounds.

FIG. 5.

Cytoprotection and cell death with gallium compounds. Events that confer cytoprotection or cell death by gallium compounds are summarized. The exposure of cells to gallium nitrate produces an early increase in cellular oxidative stress that leads to an increase in metallothionein-2A (MT2A) and hemoxygenase-1 (HO-1) as a cytoprotective reaction. The negative effect of gallium on cellular iron balance, RR activity, mitochondrial function, and other processes tilts the balance toward cytotoxicity. Cell death ensues when the cytoprotective responses are overcome. It is possible that cytoprotective responses to gallium may vary in different cells and may, thus, contribute to differences in cell sensitivity to gallium compounds.

FIG. 6.

Chemical structures of gallium compounds currently in use in the clinic. The chemical structures of (A) gallium nitrate, (B) gallium maltolate, and (C) Tris(8quinolato)gallium(III) (KP46) are shown.

FIG. 7.

Chemical structures of various thiosemicarbazones.

FIG. 8.

Chemical structures for cupric complexes of bis-thiosemicarbazones.