Chiara Nardon

Cellular and computational studies of proteasome inhibition and apoptosis induction in human cancer cells by amino acid Schiff base–copper complexes

Proliferation and apoptosis pathways are tightly regulated in a cell by the ubiquitin-proteasome system (UPS) and alterations in the UPS may result in cellular transformation or other pathological conditions. Indeed, the proteasome is often found to be overactive in cancer cells. It has also been found that cancer cells are more sensitive to proteasome inhibition than normal cells, and therefore proteasome inhibitors are pursued as antitumor drugs. The use of the proteasome inhibitor Bortezomib for treatment of multiple myeloma and mantle cell lymphoma has proved this principle. Recent studies have suggested that copper complexes can inhibit proteasome activity and induce apoptosis in some human cancer cells. However, the involved molecular mechanism is unknown. In this study, we investigated the biological activities of four amino acid Schiff base-copper(II) complexes by using human breast (MDA-MB-231 and MCF-7) and prostate (PC-3) cancer cells. The complexes C1 and C3, but not their counterparts C2 and C4, inhibit the chymotrypsin-like activity of purified 20S proteasome and human cancer cellular 26S proteasome, cause accumulation of proteasome target proteins Bax and IκB-α, and induce growth inhibition and apoptosis in concentration- and time-dependent manners. Docking analysis shows that C1, but not C2 has hydrophobic, pi-pi, pi-cation and hydrogen bond interactions with the proteasomal chymotrypsin-like pocket and could stably fit into the S3 region, leading to specific inhibition. Our study has identified the mechanism of action of these copper complexes on inhibiting tumor cell proteasome and suggested their great potential as novel anticancer agents.

Toward the selective delivery of chemotherapeutics into tumor cells by targeting peptide transporters: tailored gold-based anticancer peptidomimetics.

Complexes [Au(III)X(2)(dtc-Sar-AA-O(t-Bu))] (AA = Gly, X = Br (1)/Cl (2); AA = Aib, X = Br (3)/Cl (4); AA = l-Phe, X = Br (5)/Cl (6)) were designed on purpose in order to obtain gold(III)-based anticancer peptidomimetics that might specifically target two peptide transporters (namely, PEPT1 and PEPT2) upregulated in several tumor cells. All the compounds were characterized by means of FT-IR and mono- and multidimensional NMR spectroscopy, and the crystal structure of [Au(III)Br(2)(dtc-Sar-Aib-O(t-Bu))] (3) was solved and refined. According to in vitro cytotoxicity studies, the Aib-containing complexes 3 and 4 turned out to be the most effective toward all the human tumor cell lines evaluated (PC3, DU145, 2008, C13, and L540), reporting IC(50) values much lower than that of cisplatin. Remarkably, they showed no cross-resistance with cisplatin itself and were proved to inhibit tumor cell proliferation by inducing either apoptosis or late apoptosis/necrosis depending on the cell lines. Biological results are here reported and discussed in terms of the structure-activity relationship.

Noble metal-dithiocarbamates precious allies in the fight against cancer.

To date, cisplatin and its analogs are among the most effective chemotherapeutic agents for cancer treatment. However, high systemic toxicity and the propensity for patients to develop tumor resistance remain the main challenges in the clinical application. Therefore, the discovery and development of novel active chemotherapeutic agents are largely needed and the research of new metal-based anticancer drugs continues to be a very active international field. In this review paper we aim to give a detailed overview on our research work devoted to the design of novel dithiocarbamato complexes with different noble metals (such as palladium, platinum, copper, ruthenium and gold), which have gained considerable interest in both the development and the treatment of cancer. In particular, we summarize the results of the metal complexes achieved so far, focusing on the gold(III) compounds, that show outstanding in vitro and in vivo antitumor properties and reduced, or even no, systemic and renal toxicity, compared to the reference drug cisplatin.

Targeting the ubiquitin–proteasome pathway with inorganic compounds to fight cancer: a challenge for the future

Proteasomes are large multicatalytic complexes endowed with proteinase activity, located both in the cytosol and in the nucleus of eukaryotic cells. The ubiquitin-proteasome system is responsible for selective degradation of most intracellular proteins and therefore plays an essential regulatory role in many critical cellular processes. The proteasomal activity can also contribute to the pathological states of many diseases, including inflammation, neurodegeneration and cancer, through a disregulation in the level of regulatory proteins. These diseases may be targeted by modulating components of the ubiquitin-proteasome pathway, using small molecules as inhibitors. Bortezomib (Velcade(®)), used for the treatment of relapsed multiple myeloma, is the first and, up to now, the only proteasome inhibitor approved by the US FDA. Nowadays, the discovery that some metal-based complexes exert their antiproliferative action by affecting proteasomal activities provides the possibility of developing new opportunities in cancer therapy.

Gold(III)-Dithiocarbamato Peptidomimetics in the Forefront of the Targeted Anticancer Therapy: Preclinical Studies against Human Breast Neoplasia

Since the serendipitous discovery of cisplatin, platinum-based drugs have become well-established antitumor agents, despite the fact that their clinical use is limited by many severe side-effects. In order to both improve the chemotherapeutic index and broaden the therapeutic spectrum of current drugs, our most recent anti-neoplastic agents, Au(III) complexes, were designed as carrier-mediated delivery systems exploiting peptide transporters, which are up-regulated in some cancers. Among all, we focused on two compounds and tested them on human MDA-MB-231 (resistant to cisplatin) breast cancer cell cultures and xenografts, discovering the proteasome as a major target both in vitro and in vivo. 53% inhibition of breast tumor growth in mice was observed after 27 days of treatment at 1.0 mg kg−1 d−1, compared to control. Remarkably, if only the most responsive mice are taken into account, 85% growth inhibition, with some animals showing tumor shrinkage, was observed after 13 days. These results led us to file an international patent, recognizing this class of gold(III) peptidomimetics as suitable candidates for entering phase I clinical trials.

Gold(III)–pyrrolidinedithiocarbamato Derivatives as Antineoplastic Agents

Transition metals offer many possibilities in developing potent chemotherapeutic agents. They are endowed with a variety of oxidation states, allowing for the selection of their coordination numbers and geometries via the choice of proper ligands, leading to the tuning of their final biological properties. We report here on the synthesis, physico-chemical characterization, and solution behavior of two gold(III) pyrrolidinedithiocarbamates (PDT), namely [AuIIIBr2(PDT)] and [AuIIICl2(PDT)]. We found that the bromide derivative was more effective than the chloride one in inducing cell death for several cancer cell lines. [AuIIIBr2(PDT)] elicited oxidative stress with effects on the permeability transition pore, a mitochondrial channel whose opening leads to cell death. More efficient antineoplastic strategies are required for the widespread burden that is cancer. In line with this, our results indicate that [AuIIIBr2(PDT)] is a promising antineoplastic agent that targets cellular components with crucial functions for the survival of tumor cells.

Target selective micelles for bombesin receptors incorporating Au(III)-dithiocarbamato complexes

Pure sterically stabilized micelles (SSM) of DSPE-PEG2000, and sterically stabilized mixed micelles (SSMM) containing PC or DOPC phospholipids (5, 10 or 20% mol/mol with respect to DSPE-PEG2000) are developed as delivery systems for the gold based cytotoxic drug Au(III)-dithiocarbamato complex AuL12. In particular, SSMM containing 5% of PC at 5mM of lipid concentration encapsulates 61.0 μg of AuL12 with a DL% of 1.13. The gold complex remains stable up to 72 h when incorporated in the aggregate, as indicated by UV-vis measurements. Incorporation in micelle composition of a low amount of the peptide derivative MonY-BN-AA1, containing a bombesin peptide analogue does not influence structural parameters of the micelles (diameter around 20 nm) neither the AuL12 loading parameters. Target selective properties of the peptide containing full aggregate on PC-3 cells overexpressing the GRP/bombesin receptors are observed by in vitro cytotoxic studies: a decrease of cell viability, ∼ 50%, is obtained in cells treated with AuL12-targeted micelles at 10 μM drug concentration for 48 h with respect to untargeted micelles.

Gold(III) Complexes in the Oncological Preclinical Arena: From Aminoderivatives to Peptidomimetics.

In the last decade, we have been developing some gold(III) derivatives showing interesting antitumor properties and reduced systemic and renal toxicity, compared to the clinically-established reference drug cisplatin. Starting from the rationale at the base of our investigations, this review has been divided into two sections, with respect to our patented first- (aminoderivatives) and secondgeneration (peptidomimetics) potential drugs. Every section describes the in vitro and in vivo anticancer activity of the compounds, chosen as models, towards different types of tumor. In particular, we summarize the results achieved so far, in particular taking into account the latest in-depth studies related to their activity, mechanism of action and toxicological profile. Taken together, our data could open up new prospects for further advanced preclinical pharmacological testing.

Gold Complexes for Therapeutic Purposes: an Updated Patent Review (2010-2015).

Gold has always aroused great interest in the history of mankind. It has been used for thousands of years for jewelry, religious cult valuables, durable goods and in the art world. However, few know that such a precious and noble metal was exploited in the past by the ancients also for its therapeutic properties. More recently, in the twentieth century some complexes containing gold centers in the oxidation state +1 were studied for the treatment of the rheumatoid arthritis and the orally-administered drug Auranofin was approved by the FDA in 1985. From the chemical point of view, gold derivatives deserve special attention due to the unique position of this metal within the periodic table, which results in unconventional relativistic effects and, ultimately, in the highest electronegativity, electron affinity and redox potential among all metals. In this review, after an introduction concerning the use of gold complexes in medicine, we have examined all the patents internationally or nationally published in the years 2010-2015 (until December 31, 2015) and describing new inorganic compounds containing gold(I) and gold(III) with proved therapeutic properties. These patents were filed to mainly protect compounds with promising anticancer and anti-inflammatory activities (total 18 and 4, respectively). In particular, this work explores both coordination compounds containing ligands with various donor atoms (e.g., N-, O-, S- and -P) and organo-gold derivatives with at least one Au-C bond. The toxicological profile and the intracellular targets reported for some among the patented gold derivatives are discussed.

CCK8 peptide-labeled Pluronic® F127 micelles as a targeted vehicle of gold-based anticancer chemotherapeutics

The bioavailability and target selectivity of chemotherapeutics are significant issues in drug development. Here, we report the loading of the antiproliferative gold(III) complex, dibromo[ethyl-N-(dithiocarboxy-kS,kS′)-N-methylglycinato] gold(III) (AuL12), into the lipophilic core of micelles produced from the surfactant Pluronic® F127 (PF127). When AuL12 is encapsulated in PF127-based micelles it remains stable in saline solution up to 72 h with the gold center in the +3 oxidation state. PF127-based aggregates are efficient carriers as they enhance the water solubility of the gold complex. In vitro studies indicate that after micelle encapsulation, AuL12 gold complex preserves its antiproliferative efficacy. Moreover, by labeling the hydrophilic shell of micelles with the bioactive CCK8 peptide, the aggregates act as target-selective vehicles. In fact, cytotoxic activity towards the A431 cells overexpressing the CCK2 receptors is 10-fold higher than that towards the control cells.