Michael Frezza

Novel Metals and Metal Complexes as Platforms for Cancer Therapy

Metals are essential cellular components selected by nature to function in several indispensable biochemical processes for living organisms. Metals are endowed with unique characteristics that include redox activity, variable coordination modes, and reactivity towards organic substrates. Due to their reactivity, metals are tightly regulated under normal conditions and aberrant metal ion concentrations are associated with various pathological disorders, including cancer. For these reasons, coordination complexes, either as drugs or prodrugs, become very attractive probes as potential anticancer agents. The use of metals and their salts for medicinal purposes, from iatrochemistry to modern day, has been present throughout human history. The discovery of cisplatin, cis-[Pt(II) (NH(3))(2)Cl(2)], was a defining moment which triggered the interest in platinum(II)- and other metal-containing complexes as potential novel anticancer drugs. Other interests in this field address concerns for uptake, toxicity, and resistance to metallodrugs. This review article highlights selected metals that have gained considerable interest in both the development and the treatment of cancer. For example, copper is enriched in various human cancer tissues and is a co-factor essential for tumor angiogenesis processes. However the use of copper-binding ligands to target tumor copper could provide a novel strategy for cancer selective treatment. The use of nonessential metals as probes to target molecular pathways as anticancer agents is also emphasized. Finally, based on the interface between molecular biology and bioinorganic chemistry the design of coordination complexes for cancer treatment is reviewed and design strategies and mechanisms of action are discussed.

Metal complexes, their cellular targets and potential for cancer therapy

The development of metal complexes with anticancer activity has had an enormous impact on cancer chemotherapy. The discovery of cisplatin in the 1960s represented a landmark achievement and ushered in a new era in cancer treatment. Despite the fact that cisplatin has achieved significant clinical benefit for several types of solid tumors, its effectiveness has been hampered by toxic side effects and tumor resistance that often leads to the occurrence of secondary malignancies. However, discovery and use of cisplatin have encouraged investigators to search for and develop novel non platinum-containing metal species with superior anti-cancer activity and low side effects. As examples, gallium salts and gold complexes have been evaluated in phase I and phase II trials. Copper-chelating compounds have also shown promising results in both preclinical and clinical studies. This review provides a comprehensive overview of various non platinum metal complexes and metal-chelating compounds and discusses their potential molecular targets in tumor cells and their applications in cancer therapy.

Inhibition of tumor proteasome activity by gold dithiocarbamato complexes via both redox-dependent and -independent processes

We have previously reported on a gold(III) complex, namely [AuBr2(DMDT)] (N,N‐dimethyldithiocarbamate) showing potent in vitro and in vivo growth inhibitory activities toward human cancer cells and identifying the cellular proteasome as one of the major targets. However, the importance of the oxidation state of the gold center and the involved mechanism of action has yet to be established. Here we show that both gold(III)− and gold(I)–dithiocarbamato species, namely [AuBr2(ESDT)] (AUL12) and [Au(ESDT)]2 (AUL15), could inhibit the chymotrypsin‐like activity of purified 20S proteasome and 26S proteasome in human breast cancer MDA‐MB‐231 cells, resulting in accumulation of ubiquitinated proteins and proteasome target proteins, and induction of cell death, but at significantly different levels. Gold(I)‐ and gold(III)‐compound‐mediated proteasome inhibition and cell death induction were completely reversed by the addition of a reducing agent, dithiothreitol or N‐acetyl‐L‐cysteine, suggesting the involvement of redox processes. Furthermore, treatment of MDA‐MB‐231 cells with gold(III) compound (AUL12), but not the gold(I) analog (AUL15), resulted in the production of significant levels of reactive oxygen species. Our study provides strong evidence that the cellular proteasome is an important target of both gold(I) and gold(III)–dithiocarbamates, but distinct cellular mechanisms of action are responsible for their different overall effect. J. Cell. Biochem. 109: 162–172, 2010.

Metals in Anticancer Therapy: Copper(II) Complexes as Inhibitors of the 20S Proteasome

Selective 20S proteasomal inhibition and apoptosis induction were observed when several lines of cancer cells were treated with a series of copper complexes described as [Cu(L(I))Cl] (1), [Cu(L(I))OAc] (2), and [Cu(HL(I))(L(I))]OAc (3), where HL(I) is the ligand 2,4-diiodo-6-((pyridine-2-ylmethylamino)methyl)phenol. These complexes were synthesized, characterized by means of ESI spectrometry, infrared, UV-visible and EPR spectroscopies, and X-ray diffraction when possible. After full characterization species 1-3 were evaluated for their ability to function as proteasome inhibitors and apoptosis inducers in C4-2B and PC-3 human prostate cancer cells and MCF-10A normal cells. With distinct stoichiometries and protonation states, this series suggests the assignment of species [CuL(I)](+) as the minimal pharmacophore needed for proteasomal chymotryspin-like activity inhibition and permits some initial inference of mechanistic information.

Comparative Activities of Nickel(II) and Zinc(II) Complexes of Asymmetric [NN'O] Ligands as 26S Proteasome Inhibitors

In this study, we compare the proteasome inhibition capabilities of two anticancer candidates, [Ni(L(IA))(2)] (1) and [Zn(L(IA))(2)] (2), where L(IA-) is the deprotonated form of the ligand 2,4-diiodo-6-(((2-pyridinylmethyl)amino)methyl)phenol. Species 1 contains nickel(II), a considerably inert ion that favors covalency, whereas 2 contains zinc(II), a labile transition metal ion that favors predominantly ionic bonds. We report on the synthesis and characterization of 1 and 2 using various spectroscopic, spectrometric, and structural methods. Furthermore, the pharmacological effects of 1 and 2, along with those of the salts NiCl(2) and ZnCl(2), were evaluated in vitro and in cultured human cancer cells in terms of their proteasome-inhibitory and apoptotic cell-death-inducing capabilities. It is shown that neither NiCl(2) nor 1 have the ability to inhibit the proteasome activity at any sustained levels. However, ZnCl(2) and 2 showed superior inhibitory activity versus the chymotrypsin-like activity of both the 26S proteasome (IC(50) = 5.7 and 4.4 micromol/L, respectively) and the purified 20S proteasome (IC(50) = 16.6 and 11.7 micromol/L, respectively) under cell-free conditions. Additionally, inhibition of proteasomal activity in cultured prostate cancer cells by 2 was associated with higher levels of ubiquitinated proteins and apoptosis. Treatment with either the metal complex or the salt was relatively nontoxic toward human normal cells. These results strengthen the current working hypothesis that fast ligand dissociation is required to generate an [ML(IA)](+) pharmacophore, capable of interaction with the proteasome. This interaction, possibly via N-terminal threonine amino acids present in the active sites, renders the proteasome inactive. Our results present a compelling rationale for 2 along with its gallium(III) and copper(II) congeners to be further investigated as potential anticancer drugs that act as proteasome inhibitiors.

Anti-tumor activity of N-thiolated β-lactam antibiotics

An ongoing strategy for cancer treatment is selective induction of apoptosis in cancer over normal cells. N-thiolated beta-lactams were found to induce DNA damage, growth arrest and apoptosis in cultured human cancer cells. However, whether these compounds have a similar effect in vivo has not been studied. We report here that treatment with the beta-lactam L-1 caused a significant inhibition of tumor growth in a breast cancer xenograft mouse model, associated with induction of DNA damage and apoptosis in vivo. These results suggest that the synthetic antibiotic N-thiolated beta-lactams hold great potential to be developed as novel anti-cancer drugs.

Modulation of the tumor cell death pathway by androgen receptor in response to cytotoxic stimuli

Despite an initial response from androgen deprivation therapy, most prostate cancer patients relapse to a hormone‐refractory state where tumors still remain dependent on androgen receptor (AR) function. We have previously shown that AR breakdown correlates with the induction of cancer cell apoptosis by proteasome inhibition. However, the involvement of AR in modulating the cell death pathway has remained elusive. To investigate this, we used an experimental model consisting of parental PC‐3 prostate cancer cells that lack AR expression and PC‐3 cells stably overexpressing wild type AR gene. Here, we report that both chemotherapeutic drugs (cisplatin) and proteasome inhibitors induced caspase‐3‐associated cell death in parental PC‐3 cells whereas non‐caspase‐3 associated cell death in PC3‐AR cells. The involvement of AR in modulating tumor cell death was further confirmed in PC‐3 cells transiently expressing AR. Consistently, treatment with the clinically used proteasome inhibitor Bortezomib (Velcade/PS‐341) of (AR+) LNCaP prostate cancer cells caused AR cleavage and cell death with low levels of caspase activation. However, co‐treatment with Bortezomib and the AR antagonist Bicalutamide (Casodex) caused significant decrease in AR expression associated with an increase in caspase‐3 activity in both LNCaP and PC3‐AR cells. Thus our results provide compelling evidence for involvement of AR in deciding types of tumor cell death upon cytotoxic stimuli, and specifically, blockade of AR activities could change necrosis to apoptosis in tumor cells. Our findings may help guide clinicians based on AR status in the design of favorable treatment strategies for prostate cancer patients. J. Cell. Physiol. 226: 2731–2739, 2011.

Chapter 98 – Synthetic Analogs of (−)-Epigallocatechin-Gallate: Bioavailability and Molecular Mechanisms of Action

(−)-Epigallocatechin-gallate [(−)-EGCG], a major constituent of green tea extract, is thought to have chemopreventive and anti-cancer properties. The challenge in using it for cancer prevention and therapy is its low bioavailability, in part due to its instability under neutral or alkaline conditions and due to biologically inactivating processes such as methylation. In addition, the mechanisms of action of (−)-EGCG still remain elusive. In this chapter we discuss the various synthetic analogs of EGCG, designed to enhance its bioavailability, and we explore the contributions of the various rings and hydroxyl groups of (−)-EGCG to its molecular mechanisms of action.

Synthetic Analogs of (-)-Epigallocatechin-Gallate: Bioavailability and Molecular Mechanisms of Action

(−)-Epigallocatechin-gallate [(−)-EGCG], a major constituent of green tea extract, is thought to have chemopreventive and anti-cancer properties. The challenge in using it for cancer prevention and therapy is its low bioavailability, in part due to its instability under neutral or alkaline conditions and due to biologically inactivating processes such as methylation. In addition, the mechanisms of action of (−)-EGCG still remain elusive. In this chapter we discuss the various synthetic analogs of EGCG, designed to enhance its bioavailability, and we explore the contributions of the various rings and hydroxyl groups of (−)-EGCG to its molecular mechanisms of action.

Synthetic Analogs of (−)-Epigallocatechin-Gallate

(−)-Epigallocatechin-gallate [(−)-EGCG], a major constituent of green tea extract, is thought to have chemopreventive and anti-cancer properties. The challenge in using it for cancer prevention and therapy is its low bioavailability, in part due to its instability under neutral or alkaline conditions and due to biologically inactivating processes such as methylation. In addition, the mechanisms of action of (−)-EGCG still remain elusive. In this chapter we discuss the various synthetic analogs of EGCG, designed to enhance its bioavailability, and we explore the contributions of the various rings and hydroxyl groups of (−)-EGCG to its molecular mechanisms of action.